JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 JPET FastThis articleForward. has not Published been copyedited on and October formatted. 31,The final2005 version as DOI:10.1124/jpet.105.093831 may differ from this version. JPET# 93831

Blood glucose lowering nuclear receptor agonists only

partially normalize hepatic gene expression in db/db

mice Downloaded from

jpet.aspetjournals.org Michael Loffler, Martin Bilban, Mark Reimers, Werner Waldhäusl and

Thomas M. Stulnig

at ASPET Journals on September 25, 2021 Clinical Division of Endocrinology and Metabolism, Department of Internal Medicine

III, Medical University Vienna, Vienna, Austria (M.L., W.W., T.M.S.); CeMM - Center

of Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria (M.L.,

W.W., T.M.S.); Clinical Institute of Medical and Chemical Laboratory Diagnostics,

Medical University Vienna and Ludwig Boltzmann Institute for clinical and

experimental Oncology, Vienna, Austria (M.B.); Laboratory of Molecular

Pharmacology, National Cancer Institute, Bethesda, Maryland (M.R.)

1

Copyright 2005 by the American Society for Pharmacology and Experimental Therapeutics. JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

Running Title

Blood Glucose Lowering Drugs and Hepatic Gene Expression

Corresponding author Downloaded from Thomas M. Stulnig, Clinical Division of Endocrinology and Metabolism, Department of

Internal Medicine III, Medical University Vienna, Währinger Gürtel 18-20, 1090 Vienna,

Austria; e-mail : [email protected] jpet.aspetjournals.org

Number of Text pages 23 at ASPET Journals on September 25, 2021 Tables 2 Figures 6 + 3 Supplementary Figures References 40

Number of words Abstract 220 Introduction 333 Discussion 402

ABBREVIATIONS PPAR, peroxisome proliferator-activated receptor; QPCR, quantitative real-time reverse-transcriptase PCR; SREBP, sterol regulatory element-binding protein; UPR, unfolded protein response.

Section Endocrine and Diabetes

2 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

ABSTRACT

Agonists of the nuclear receptors peroxisome proliferator-activated receptor (PPAR)

γ, PPARα, and liver X receptors (LXR) reduce blood glucose in type 2 diabetic patients and comparable mouse models. Since the capacity of these drugs to normalize hepatic gene expression is not known we compared groups of obese diabetic db/db mice treated with agonists for PPARγ (rosiglitazone [Rosi]; 10

mg/kg/day), PPARα (Wy 14643 [Wy]; 30 mg/kg/day), and LXR (T0901317 [T09]; 40 Downloaded from mg/kg/day) and from untreated non-diabetic littermates (db/+) by oligonucleotide microarrays and quantitative reverse transcriptase polymerase chain reaction. The jpet.aspetjournals.org 10-day treatment period of db/db mice with Rosi, Wy, and T09 altered expression of

300, 620, and 735 genes including agonist-specific target genes, respectively.

However, from the 337 genes differentially regulated in untreated db/+ vs db/db at ASPET Journals on September 25, 2021 animals, only 34 (10%), 51 (15%) and 82 (24%) were regulated in the direction of the db/+ group by Rosi, Wy and T09, respectively. Gene expression normalization by drug treatment involved glucose homeostasis, lipid homeostasis and local glucocorticoid activation. In addition, our data pointed to hitherto unknown interference of these nuclear receptors with growth hormone receptor gene expression and endoplasmic reticulum stress. However, many diabetes-associated gene alterations remained unaffected or were even aggravated by nuclear receptor agonist treatment. These results suggest that diabetes-induced gene expression is minimally reversed by potent blood glucose lowering nuclear receptor agonists.

3 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

INTRODUCTION

Modern anti-diabetic agents targeting nuclear receptors are designed to modulate blood glucose levels and gene expression on a molecular level. Transcriptional regulation by agonists of peroxisome proliferator-activated receptor (PPAR)γ, PPARα and LXR comprise genes involved in gluconeogenesis, fatty acid metabolism and ketogenesis (Willson et al.,

2000; Venkateswaran et al., 2000). PPARγ is a critical transcription factor involved in energy Downloaded from balance and activated by well-established anti-diabetic drugs, the thiazolidinediones

(Lehmann et al., 1995). Nuclear receptor agonist or fatty acid dependent activation of PPARα promotes peroxisomal proliferation, hepatic fatty acid oxidation and the generation of ketone jpet.aspetjournals.org bodies thereby providing substrates for energy metabolism in peripheral tissues (Issemann and

Green, 1990). LXRα, the predominant LXR paralogue in liver (Repa et al., 2000) regulates at ASPET Journals on September 25, 2021 intracellular cholesterol and bile acid metabolism as well as expression of sterol regulatory binding protein (SREBP)-1c, the major lipogenic transcription factor (Schultz et al., 2000;

Repa et al., 2000). Activation of LXR is associated with down-regulation of key genes involved in hepatic gluconeogenesis (Stulnig et al., 2002a; Cao et al., 2003; Laffitte et al.,

2003). Moreover, nuclear receptors can modulate each other’s gene expression as shown for

PPARγ and LXRα (Ide et al., 2003; Seo et al., 2004) pointing to a close relation of their transcriptional regulations and metabolic function. Agonists of PPARγ, PPARα and LXR all decrease blood glucose concentrations in type 2 diabetes patients and/or comparable animal models (Lehmann et al., 1995; Guerre-Millo et al., 2000; Laffitte et al., 2003) by regulating gene expression. In order to address the question whether normalization of blood glucose levels by these nuclear receptor agonists is accompanied by normalized gene expression, we analyzed genome-wide hepatic gene expression profiles of diabetic db/db mice treated with nuclear receptor agonists and their untreated non-diabetic littermates and compared them with

4 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 those of untreated db/db mice. By providing a comprehensive overview of drug-induced changes in gene expression in obese diabetic mice our data revealed that reversal of diabetes- associated alterations in hepatic gene expression occurs only to a very limited extent. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021

5 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

MATERIAL AND METHODS

Animals. Male C57BL/KsJ-leprdb/leprdb diabetic (db/db) mice and their nondiabetic littermates (db/+) were purchased from Charles River Laboratories Inc. (Germany) at seven weeks of age and maintained under standard light (12h light/dark) and temperature conditions

(23°C). During one week of acclimatization mice were provided with a low-fat standard

rodent diet (<3% fat; N1324, Altromin, Germany) and water ad libitum. Downloaded from

Treatment. For the experiment, the low-fat standard diet was either mixed with vehicle alone

(ethanol; untreated) or supplemented with 0.005% (w/w) PPARγ agonist Rosiglitazone (Rosi; jpet.aspetjournals.org

Avandia, Smithkline Beecham, PLC, Middlesex, U.K.; corresponding to approximately 10 mg/kg/day; Hori et al., 2002), 0.02% PPARα agonist Wy-14.643 ([4-chloro-6-(2,3-xylidino)-

2-pyrimidinylthioacetic acid; Wy; Sigma-Aldrich, St Louis, MO, U.S.A.; corresponding to at ASPET Journals on September 25, 2021 approximately 30 mg/kg/day; Edvardsson et al., 1999) or 0.025% of the synthetic LXR agonist T0901317 (N-(2,2,2-trifluoroethyl)-N-[4-[2,2,2-trifluoro-1-hydroxy-

1(trifluoromethyl)-ethyl]phenyl]-benzenesulfonamide; T09; generously provided by Amgen

Inc., formerly Tularik Inc.; corresponding to approximately 40 mg/kg/day; Stulnig et al. ,

2002a) followed by extensive evaporation of ethanol. Four groups of db/db (untreated, Rosi,

Wy, T09; n = 5) and one group of db/+ mice (untreated; n = 8) were treated for 10 days.

Tissue and blood analyses. Mice were anesthetized with Isoflurane and sacrificed by neck dislocation after cardiac puncture. The liver was immediately cut into small homogenous regions, snap frozen in liquid nitrogen and kept at -80°C until isolation of total RNA. Blood samples were drawn from the tail vein before starting the experimental diets. EDTA plasma separated from cardiac blood was stored in aliquots at -20°C until further analyses. Blood glucose was measured by an automated analyzer (ALCYON 300i, Abbott Laboratories,

6 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

Illinois, U.S.A.) at the beginning and the end of the experiment. Plasma cholesterol and triglycerides were determined by the Alcyon 300i analyzer (Abbott Laboratories). Serum concentrations of non-esterified fatty acids were measured with the Wako FFA-kit (Wako chemicals, Richmond, VA, U.S.A.), insulin by ELISA (Linco Research, St. Charles, MO,

U.S.A.). Liver triglycerides were determined following lipid extraction as described

(Haemmerle, 2002) but by using a commercially available enzymatic reagent (Rolf Greiner

Biochemica, Flacht, Germany). The study protocol was approved by the local ethics Downloaded from committee for animal experiments and the Guidelines for the Care and Use of Laboratory

Animals as adopted and promulgated by the U.S. National Institutes of Health and the jpet.aspetjournals.org European Convention for the Protection of Vertebrate Animals Used for Experimental and

Other Scientific Purposes were followed.

RNA preparation. Total RNA was prepared by disrupting equivalent regions of at ASPET Journals on September 25, 2021 approximately 50 mg of liver tissue from each animal per group in TRIzol reagent

(Invitrogen) with a tissue homogenizer followed by RNA isolation according the manufacturer’s instructions. Total RNA samples were checked for integrity by agarose gel electrophoresis and the 2100 bioanalyzer (Agilent, Palo Alto, CA). For microarray cDNA synthesis total RNA samples were repurified with RNeasy MinElute kit (Qiagen).

Microarray hybridization and data analysis. Ten µg of total RNA from each animal (n=3 per group) was transcribed into first strand cDNA by Superscript II (Invitrogen, Carlsbad,

CA, U.S.A.) using T7-Oligo(dT) primers followed by second strand synthesis and repurification according to the manufacturer’s protocols (all Affymetrix, Santa Clara, CA,

U.S.A.). Following in vitro transcription, 15µg of labeled and fragmented cRNA from each individual sample were hybridized to U74Av2 GeneChips (12k) which were scanned using the GeneArray scanner and further analyzed with Microarray Suite version 5.0 software

7 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 according to the manufacturer’s protocols (all Affymetrix, Santa Clara, CA, U.S.A.). Array quality criteria for all chips included control of expression report files for background, background noise, scale factor <2, internal control gene 3' to 5' ratio, and hybridization control ratios, respectively. Normalization was performed by global scaling to an average intensity of 100 arbitrary units. Gene abundances were estimated by robust multi-array analysis (Irizarry, 2003) using the probe-level modeling (affyPLM) package from

Bioconductor (www.bioconductor.org). This algorithm provides for calculation of means and Downloaded from standard errors of logarithmically transformed estimates, reflecting the inconsistency among the different probes for the same gene. We used a strategy similar to analysis of variance and jpet.aspetjournals.org computed a consensus estimate for the variability among all groups as a basis for t-test statistics. The multiple comparisons problem was addressed by estimating the false discovery rate (FDR) in a simple manner as the ratio of the expected number of false positives (resulting at ASPET Journals on September 25, 2021 from permutation analyses with samples being randomly assigned to different groups) at a given p-value threshold to the number of positives actually found (Irizarry, 2003; Storey and

Tibshirani, 2003). Using this statistical approach comparisons of gene expression with absolute fold changes of at least 1.5-fold (increase or decrease) were selected at p<0.01 as a default and p<0.001 where indicated. Abbreviation, annotation and analyses of genes meeting the selection criteria were done by combining available information from Affymetrix, Applied

Biosystems (PANTHER), Jax, Genelynx, Kegg, Ensembl, Swiss-Prot and PubMed in a

Filemaker Pro database.

Hierarchical clustering analysis. Genes altered by treatment with all nuclear receptor agonists by at least 1.5-fold (in either direction) and at p < 0.01 between mean gene expression and untreated db/db animals were selected for a cluster analysis. The genes were clustered using a distance measure defined as 1 minus the correlation between gene pair

8 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 measures, and using complete-linkage hierarchical clustering. The results are shown in

Supplementary Figure 2.

Quantitative Real-Time Polymerase Chain Reaction. One µg of total RNA from each animal (n=5 per group) was treated with DNase I and reverse transcribed into cDNA by

Superscript II using random hexamer priming (all Invitrogen, Carlsbad, CA, U.S.A.).

Quantitative real-time polymerase chain reaction (QPCR) was performed for all samples per Downloaded from group using gene-specific FAM-TAMRA-labeled commercial Assays-on-Demand (Assay ID:

Mm00839363_m1, G6pc; Mm00484574_m1, G6pt1; Mm00440636_m1, Pck1;

Mm00662319_m1, Fasn; Mm00772290_m1, Scd1; Mm00833328_m1, Gpam; jpet.aspetjournals.org

Mm00483985_m1, Crat; Mm00451571_m1, Slc25a20; Mm00443579_m1, Acox1;

Mm00470091_s1, Ehhadh; Mm00478137_m1, Pex11a; Mm00550050_m1, Hmgcs2;

Mm00476182_m1, Hsd11b1; Mm00439093_m1, Ghr; Mm00439561_m1, Igf1; Applied at ASPET Journals on September 25, 2021

Biosystems, Foster City, CA, U.S.A.) or self-designed primer/probe combinations (SREBP-

1c; ref. Stulnig et al., 2002b) normalized to 18S VIC-TAMRA labeled endogenous control

(Applied Biosystems). Expression of specific mRNAs was quantitated in duplicates with a tolerated variance of ≤10% in an ABI PRISM 7000 Cycler (Applied Biosystems).

Statistics and calculations. Data are given in means ± S.E.M. unless indicated otherwise.

Study groups were compared to untreated db/db mice by univariate ANOVA using Dunnett’s t-test for post-hoc analysis except for evaluation of gene expression profiles (see above). Data from quantitative PCR and microarrays as well as other data exhibiting unequality of variances between groups according to Levene’s test were log-transformed before ANOVA.

The effect of diabetes was evaluated by comparing db/+ to db/db mice resulting in its reciprocal (diabetes-1) in order to facilitate direct comparisons with the normalizing effect of the compounds.

9 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

RESULTS

Metabolic data. This study was performed to elucidate to which extent blood glucose- lowering nuclear receptor agonists alter hepatic gene expression profiles of obese diabetic mice in convergence to non-diabetic animals. Gene expression profiles of db/db mice treated with agonists of PPARγ (Rosi), PPARα (Wy) and LXR (T09) and untreated db/+ mice were compared to untreated diabetic mice. Treatment with each of the compounds resulted in

reductions in blood glucose concentrations near to those of non-diabetic mice (Table 1). Downloaded from

Plasma insulin concentrations were significantly increased only in the T09-treated group indicating that stimulation of insulin secretion as shown for cultured pancreatic islets and β- jpet.aspetjournals.org cells (Efanov, 2004) could also occur in vivo. Body weight remained constant during the treatment except a borderline (p = 0.088) increase in the Rosi group. The Wy and particularly

T09-treated animals showed significant hepatomegaly with increased liver to body weight at ASPET Journals on September 25, 2021 ratio by 1.8 and 2.5-fold, respectively, due to the known development of hepatic steatosis by

T09 as revealed by highly elevated hepatic triglyceride contents in T09-treated db/db mice

(Table 1).

Gene expression profiling. Gene expression profiles were evaluated using 12k oligonucleotide microarrays for 3 individual mice of each group and robust multiarray analysis. A p-value of < 0.01 was predefined together with a fold change of ≥ 1.5 / ≤ -1.5-fold as selection criteria for profile comparisons including detection of functional groups. 337 genes were altered in untreated diabetic vs. non-diabetic mice (Table 2). Drug treatment of db/db mice elicited significant changes in the expression of a comparable number of genes, namely 300, 620, and 735 genes for Rosi, Wy, and T09, respectively. Estimated false discovery rates (FDR) were between 6-11% for p < 0.01 but about 80% of genes were altered at a p < 0.001 with FDR of 1-3%, respectively, indicating excellent confidence of data.

Moreover, standardized logarithmic expression estimates from microarrays highly correlated

10 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 with those from quantitative PCR (adjusted r2 = 0.854, p < 0.0005) indicating valid relative quantitation by microarray hybridization (data not shown).

We used the reciprocal form of the diabetes effect (diabetes-1), i.e. untreated non- diabetic vs. diabetic mice, in order to facilitate direct comparison with treatment effects.

Treatment-induced normalization of gene expression was defined as genes that were significantly changed by a compound in the same direction as diabetes-1, i.e. in convergence to db/+ animals, without testing whether the gene actually reached the level of non-diabetic Downloaded from mice. From the genes (microarray probe sets) altered in db/db vs db/+ mice, only 34 (10% of all genes altered in diabetes-1), 51 (15%) and 82 (24%) were normalized by Rosi, Wy and

T09, respectively (Fig. 1A). In total, only a set of 19 genes (6%) were normalized by all blood jpet.aspetjournals.org glucose lowering drugs (listed in Supplementary Figure 1A) indicating that these genes could be particularly important in mediating the glucose lowering effects. These genes included

some with clear implication in glucose metabolism and diabetes such as those involved in at ASPET Journals on September 25, 2021 gluconeogenesis (e.g., glucose-6-phosphatase, fructose-1,6-bisphosphatase) and the glucocorticoid-activating enzyme 11β-hydroxysteroid dehydrogenase type 1, whereas the functional implication of others for glucose lowering have to be assessed in detail. However,

56 (17%), 85 (25%) and 59 (18%) genes altered in db/db vs db/+ mice were regulated by

Rosi, Wy and T09, respectively, in the direction opposite to diabetes-1 (Fig. 1B). The 30 genes

(probe sets) regulated by all compounds in the direction opposite to diabetes-1 (listed in

Supplementary Figure 1B) thus possibly giving insight into augmented adaptive processes, e.g., comprised genes involved in mitochondrial and peroxisomal β-oxidation (Acaa1, Dci,

Ehhadh), including the important peroxisomal biogenesis factor (Pex11a) as discussed in detail below. Treatment with nuclear receptor agonists regulated many genes not altered by diabetes-1 itself and resulted in considerably overlaps in gene expression profiles in diabetic mice (Fig. 1C) similar to that shown for PPARα, LXR, and retinoid X receptor in non- diabetic mice (Anderson et al., 2004). The overlap of PPARγ agonist treatment with other

11 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 compounds is noteworthy, since PPARγ is only weakly expressed in liver and thus direct crosstalk between nuclear receptors cannot be accounted for. Supplementary Figure 2 provides a tree obtained by hierarchical clustering of genes altered in parallel by all nuclear receptors agonists to highlight the correlation of their regulation. A complete list of all genes altered in db/db vs db/+ mice (shown as diabetes-1) or by treatment with any compound is given in Supplementary Figure 3.

Gluconeogenesis. Hepatic gluconeogenesis and glucose output is significantly enhanced Downloaded from in type 2 diabetes patients and comparable mouse models and contributes to high blood glucose levels. Treatment with each of the blood glucose lowering nuclear receptor agonists

resulted in normalization of gluconeogenetic key enzyme gene expression as shown for jpet.aspetjournals.org phosphoenol pyruvate carboxykinase 1 (PEPCK; gene Pck1; 1.2 fold; p < 0.05), fructose-1,6- bisphosphatase (Fbp1) and the catalytic and transport units of glucose-6-phosphatase (G-6-

Pase; G6pc and G6pt1) (Fig. 2). Interestingly, the PPARα agonist treatment led to an at ASPET Journals on September 25, 2021 increased expression of fructose-2,6-bisphosphatase (Fbp2) which could imply an additional regulation of Fbp1 through competitive inhibition by fructose-2,6-bisphosphate. Expression of aldolase B (Aldo2) that catalyzes cleavage of fructose-1,6-bisphosphate and functions as a feed forward activator of pyruvate kinase was normalized by both Wy (-1.8 fold; p < 0.002) and Rosi (-1.4 fold; p = 0.042). Glucokinase (Gck) whose expression was increased by 1.4 fold in diabetes-1, was normalized in the Rosi group (1.4 fold; p = 0.003) and T09 group (1.8 fold; p < 0.001). Additional transfer of glucose-6-phosphate through the pentose phosphate shunt in the LXR-treated group is indicated by elevated expression of Gck, X-linked glucose-

6-phosphate dehydrogenase (G6pdx), glucose-6-phosphate dehydrogenase 2 (G6pd2), ribose

5-phosphate isomerase A (Ripa) and transketolase (Tkt; Supplementary Figure 3). In general these data indicate that treatment of db/db mice with different blood glucose lowering nuclear receptor agonist resulted in normalized expression of key genes involved in glucose homeostasis even though some genes were regulated in an agonist-specific manner.

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Lipogenesis. Hepatic lipogenesis and fatty acid desaturation has been implicated in various pathological conditions including obesity and diabetes (Ntambi et al., 2002). Hepatic gene expression of lipogenic enzymes, e.g., stearoyl-Coenzyme A desaturase 1 (Scd1), was generally increased in db/db vs db/+ animals (Fig. 3A). Treatment of db/db mice with any blood glucose lowering compound led to a further strong increase of Scd1 gene expression

(Fig. 3A). Scd1 expression is regulated by sterol regulatory element-binding protein-

(SREBP)-1c-dependent and independent mechanisms (Miyazaki et al., 2004). Since induction Downloaded from of Scd1 by drugs correlated with elevated SREBP-1c expression levels only in the T09 group

(Fig. 3A), Scd1 upregulation by PPAR agonists appears to occur predominantly by SREPB-

1c-independent mechanisms. Parallel to Scd1 expression, fatty acid synthase (Fasn) and jpet.aspetjournals.org glycerol-3-phosphate acyltransferase (Gpam) was strongly induced by treatment with any agonist whereas expression of hepatic lipase was decreased (Fig. 3A). Thereby, the nuclear

receptor agonists aggravated diabetes-associated alterations in lipogenesis in parallel with at ASPET Journals on September 25, 2021 their glucose lowering effect (Way et al., 2001). Such regulation opposite to diabetes-1 could on the one hand indicate enhancement of adaptive processes that were already induced by diabetes itself but also worsening of detrimental diabetes-associated alterations. Recent data suggest that upregulation of lipogenesis provides an effective means to inhibit diabetes development by shifting the lipogenic burden from adipose tissue to the liver (Nadler and

Attie, 2001). Altogether alterations in hepatic lipid homeostasis provoked hepatic triglyceride accumulation particularly in the T09-treated and – to a lesser extent – in Rosi-treated animals.

Moreover, the LXR agonist increased blood plasma triglyceride concentrations, whereas Rosi and Wy lowered plasma triglyceride concentrations (Table 1; ref. Chisholm et al., 2003;

Oakes et al., 1994). In contrast to lipogenic enzymes, genes involved in cholesterogenesis such as 3-hydroxy-3-methylglutaryl-Coenzyme A reductase (Hmgr) and synthase 1

(Hmgcs1), isopentenyl-diphosphate delta isomerase (Idi1), farnesyl diphosphate farnesyl transferase 1 (Fdft1) and NAD(P)-dependent steroid dehydrogenase-like (Nsdhl) were not

13 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 altered by diabetes and were differently changed by nuclear receptor agonists (Supplementary

Figure 3. and data not shown).

Mitochondrial and peroxisomal β-oxidation. β-oxidation of free fatty acids provides energy and substrates for ketogenesis. Expression of genes implicated in mitochondrial fatty acid import including the carnitine acyltransferase Crat and the translocase Slc25a20 (Ramsay et al., 2001; Sekoguchi et al., 2003). Expression of both genes was significantly increased in diabetic vs. non-diabetic mice and further elevated by treatment with blood glucose-lowering Downloaded from nuclear receptor agonists indicating augmented mitochondrial fatty acid import (Fig. 3B).

Expression of the long chain-specific carnitine acyltransferases (Cpt1a, Cpt2) was increased

by diabetes and further elevated in response to Wy treatment. Expression of enzymes jpet.aspetjournals.org involved in mitochondrial fatty acyl oxidation such as dodecenoyl-Coenzyme A delta isomerase (Dci) and t acetyl-CoA dehydrogenases (Acads, Acadm, Acadl) was somewhat

increased in diabetic animals and further elevated by nuclear receptor agonists (Fig. 3C and at ASPET Journals on September 25, 2021

Supplementary Figure 3). Peroxisomes oxidize fatty acids by different pathways with preference for long chain and very long chain fatty acids (Van Veldhoven and Mannaerts,

1999). Expression of enzymes of the classical pathway, namely acyl-Coenzyme A oxidase 1

(Acox1), enoyl-Coenzyme A hydratase/3-hydroxyacyl Coenzyme A dehydrogenase (Ehhadh) and acetyl-Coenzyme A acyltransferase 1 (Acaa1) was more pronounced in db/db vs. db/+ animals (Lan et al., 2003) and further increased by nuclear receptor agonist treatment, particularly Wy (Fig. 3C) in parallel with peroxisomal biogenesis factor 11a (Pex11a; Fig.

3C). Notably, hepatic triglyceride accumulation particularly in T09-treated animals revealed that increased β-oxidation did not sufficiantly counteract elevated lipogenesis to prevent hepatic steatosis. Increased ketogenesis in diabetes was indicated by elevated expression of key genes 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 2 (Hmgcs2) and 3-hydroxy-3- methylglutaryl-Coenzyme A lyase (Hmgcl; Fig. 3D). Particularly PPAR agonists further increased Hmgcl and Wy also increased Hmgcs2 expression suggesting increased

14 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 ketogenesis. In conclusion, treatment with different blood glucose-lowering nuclear receptor agonists resulted in increased expression of most genes contributing to mitochondrial and peroxisomal fatty acid oxidation and ketogenesis thereby aggravating the diabetes-associated shift to fatty acid metabolism.

Glucocorticoid activation. Type 2 diabetes has been shown to be associated with alterations in local activation of inactive glucocorticoid precursors (cortisone, 11-dehydro corticosterone) to active glucocorticoids (corticosterone, cortisol) by 11β-hydroxysteroid Downloaded from dehydrogenase type 1 (Hsd11b1). Hsd11b1 is particularly expressed in human and rodent liver and adipose tissue (Bujalska et al., 2002) and increased expression has been linked to

development of diabetes and the metabolic syndrome (Masuzaki et al., 2001; Aoki et al., jpet.aspetjournals.org

2001). Treatment with agonists of the PPAR and LXR family has been shown recently to affect expression of Hsd11b1 in adipose tissue and liver (Stulnig et al., 2002b; Cao et al.,

2003). Treatment of either nuclear receptor agonists diminished Hsd11b1 expression even at ASPET Journals on September 25, 2021 below the level of non-diabetic mice (Fig. 4). Since oxo-reductase activity of Hsd11b1 depends on NADPH, mutations in hexose-6-phosphate dehydrogenase (H6pd) synergize with weak Hsd11b1 mutations in glucocorticoid activation (Draper et al., 2003). However, H6pd gene expression was not changed in this study (Fig. 4). Notably, Hsd11b1 was one of the only

19 transcripts that were normalized by all blood glucose lowering nuclear receptor agonists

(Supplementary Fig. 1A). These data emphasize the importance of 11β-hydroxysteroid dehydrogenase 1 for the development of insulin resistance and suggest the use of specific inhibitors of this enzyme in diabetes treatment.

Growth hormone signaling. Growth hormone (GH) antagonizes insulin signaling and elicits insulin resistance resulting in increased hepatic glucose production. In contrast to unchanged expression in diabetic vs. non-diabetic mice, treatment with blood nuclear receptor agonists downregulated growth hormone receptor gene (Ghr) expression (Fig. 5), whose abundance is related to GH tissue sensitivity (Dominici and Turyn, 2002; Iida et al., 2004), as

15 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 well as expression of insulin-like growth factor-1 (Igf-1), a prototype GH-responsive gene in the liver. These data indicate interference with GH action by nuclear receptor agonist treatment. Since inhibition of GH action improves insulin sensitivity (Yakar et al., 2004), this could be an additional mechanism how PPAR and LXR agonists ameliorate glucose homeostasis in diabetes.

Endoplasmic reticulum stress. Endoplasmic reticulum stress is a major contributor for

the development of obesity and insulin resistance as shown very recently (Özcan et al., 2004). Downloaded from

Obesity triggers an unfolded protein response (UPR) in liver that is controlled by X-box- binding protein-1 (XBP-1) (Yoshida et al., 2001). Notably, XBP-1+/- mice are prone to jpet.aspetjournals.org insulin resistance due to impaired insulin signal transduction during endoplasmic reticulum stress (Özcan et al., 2004). Glucose-regulated/binding immunoglobulin protein Grp78

(Hspa5), an endoplasmic reticulum chaperone, and the protein kinase inhibitor p58ipk at ASPET Journals on September 25, 2021 (Dnajc3) are upregulated during UPR and the latter has been shown to be a XBP-1 target gene as is thioredoxin domain containing protein-7 (Txndc7) (Lee et al., 2003). XBP-1 expression was significantly lowered by all blood glucose-lowering nuclear receptor agonists (Fig. 6) in parallel with Grp78, DNajc3 and Txndc7 pointing to reduced XBP-1 activity and amelioration of endoplasmic reticulum stress. Reduced expression of Grp78 in untreated db/db animals compared to lean mice could be secondary to the nearly twofold reduced expression of XBP-1 in untreated db/db vs. db/+ mice. It is intriguing to speculate that the reduced expression of

XBP-1 in db/db animals indicates a reduced capacity to deal with endoplasmic reticulum stress similar to that seen in XBP-1+/- mice whereas further reduction of the expression of

UPR genes by nuclear receptor agonist treatment of diabetic mice reflects a decline of endoplasmic reticulum stress. Thus, reduction of endoplasmic reticulum stress could be a novel mechanism how PPAR and LXR agonists lower blood glucose concentrations in diabetic mice prone to UPR by lowered XBP-1 expression.

16 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

DISCUSSION

Insulin-sensitizing compounds such as agonists for PPARγ and PPARα are widely used in clinical practice to ameliorate diabetes-induced alterations in glucose and lipid metabolism, respectively. In this study we show that although blood glucose lowering agonists for the nuclear receptors PPARγ, PPARα and LXR regulate expression of a large number of genes, they improve only some diabetes-associated alterations mainly by normalization of Downloaded from gluconeogenetic gene expression. A large number of diabetes-associated alterations in gene expression were not reversed towards levels found in non-diabetic mice but even deteriorated jpet.aspetjournals.org by drug treatment including genes implicated in lipogenesis, peroxisomal and mitochondrial function. Some of these regulations, e.g., the elevated expression of genes involved in lipogenesis, indicate that nuclear receptor agonists enhanced adaptive processes protecting at ASPET Journals on September 25, 2021 obese mice from further metabolic derangements, e.g. by shifting the lipogenic burden to the liver. A gene expression study by itself cannot discriminate between causal and adaptive alterations. However, irrespective of whether diabetes-associated alterations by nuclear receptor agonist treatment were primarily of causal or adaptive nature, these changes indicate that a cure in a molecular sense, i.e., correction of causal diabetes-associated molecular alterations, has been achieved by the compounds only to a very limited extent. In addition, our study disclosed hints on possible novel modes of action how nuclear receptor agonists ameliorate blood glucose concentrations. Particularly interference with growth hormone signaling and reduction of endoplasmic reticulum stress warrant investigations in future focused studies. Moreover, the significantly increased insulin plasma concentration by T09 treatment revealed that LXR agonism or selective LXR modulation could provide a novel pharmacological approach to stimulate insulin secretion. However, issues of preventing hepatic steatosis by these compounds and possible β-cell lipotoxicity by increased SREBP-1c

17 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 expression (Efanov, 2004) have to be addressed first. Notably, the model for type 2 diabetes used here cannot discriminate between alterations induced by diabetes or obesity alone, respectively. Hence changes provoked by obesity but not by metabolic derangements cannot be overcome by the action of the compounds that did not induce weight loss. However, alterations induced by obesity and metabolic derangements are usually combined also in type

2 diabetes patients. Downloaded from In conclusion, this study revealed that apart from pointing to novel modes of action, currently available blood glucose-lowering nuclear receptor agonists by far do not normalize diabetes-associated molecular alterations. Despite our advances during recent years, there is jpet.aspetjournals.org still a need for developing novel drugs for effective treatment of type 2 diabetes at the molecular level. at ASPET Journals on September 25, 2021 Acknowledgments. We thank Sylvia Molzer for technical assistance and Jelena Todoric for liver triglyceride determination.

18 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

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FOOTNOTES

Title footnote: This work was supported by CeMM – Center of Molecular Medicine, a basic research institute within the companies of the Austrian Academy of Sciences (to T.M.S. and

W.W.), and the Joseph Skoda Award of the Austrian Society for Internal Medicine (to

T.M.S.).

Downloaded from

Address reprint requests to Thomas M. Stulnig, Clinical Division of Endocrinology and

Metabolism, Department of Internal Medicine III, Medical University Vienna, Währinger

Gürtel 18-20, 1090 Vienna, Austria; e-mail : [email protected] jpet.aspetjournals.org

at ASPET Journals on September 25, 2021

24 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

FIGURE LEGENDS

Fig. 1. Overlap of diabetes and drug effects. Out of the 337 genes (microarray probe sets) altered by diabetes figures in overlapping wheels of panels (A) and (B) indicate the number of genes that were also regulated by PPARγ, PPARα and LXR agonists, respectively, in direction to normalize (A) or deteriorate (B) diabetes-associated changes (see Supplementary

Figure 1 and 2). Panel (C) gives total numbers of genes regulated by the compounds

illustrating considerable overlap of drug effects. Numbers with connecting lines to the wheels Downloaded from indicate the total number of genes regulated by the respective nuclear receptor agonist. Note that only a minority of diabetes-associated gene regulations were normalized by the treatments, and that many genes not associated with diabetes-associated alterations were jpet.aspetjournals.org regulated by the compounds.

Fig. 2. Expression of genes involved in gluconeogenesis. Diabetic db/db mice were treated for at ASPET Journals on September 25, 2021

10 days with Rosi, Wy, or T09, respectively, or left untreated as were non-diabetic db/+ mice.

Gene expression was evaluated by QPCR (default; n=5) or by microarrays (indicated by black dot; n=3) and is given in percent of that in untreated db/db mice. G6pc, glucose-6- phosphatase, catalytic subunit; G6pt, glucose-6-phosphatase, transport subunit; PEPCK

(Pck1), phosphoenolpyruvate carboxykinase 1, cytosolic; Fbp1, fructose bisphosphatase 1;

Aldo2, aldolase 2, B isoform; ut, untreated. Significant differences compared to untreated db/db are indicated as follows: *, p < 0.05; †, p < 0.01; ‡, p < 0.001.

Fig. 3. Lipid homeostasis and ketogenesis. A) Expression of genes involved in lipogenesis.

Gene expression from mice is given as detailed in legend to Fig. 2. Gene expression was evaluated by QPCR (default; n=5) or by microarrays (indicated by black dot; n=3) and is given in percent of that in untreated db/db mice. Fasn, fatty acid synthase; Scd1, stearoyl-

25 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

Coenzyme A desaturase 1; Gpam, glycerol-3-phosphate acyltransferase, mitochondrial;

Dgat1, diacylglycerol O-acyltransferase 1; Lipc, hepatic lipase; Srebp-1c, sterol regulatory element binding factor 1; ut, untreated; geometric mean is given for columns exceeding scale of Y-axis. Significant differences compared to untreated db/db are indicated as follows: *, p <

0.05; †, p < 0.01; ‡, p < 0.001. B) Expression of genes involved in mitochondrial fatty acid import. Crat, carnitine acetyltransferase; Slc25a20, solute carrier family 25 (mitochondrial carnitine/acylcarnitine translocase), member 20; Cpt1a, carnitine palmitoyl- transferase 1, Downloaded from liver; Cpt2, carnitine palmitoyl- transferase 2. C) Expression of genes involved in mitochondrial and peroxisomal β-oxidation. Acox1, acyl-Coenzyme A oxidase 1, palmitoyl;

Dci, dodecenoyl-Coenzyme A delta isomerase (3,2 trans-enoyl-Coenyme A isomerase); jpet.aspetjournals.org

Ehhadh, enoyl-Coenzyme A, hydratase/3-hydroxyacyl Coenzyme A dehydrogenase; Acaa1,

3-ketoacyl-CoA thiolase B, acetyl-Coenzyme A acyltransferase 1; Pex11a, peroxisomal

biogenesis factor 11a. D) Expression of genes involved in ketogenesis. Hmgcs2, 3-hydroxy-3- at ASPET Journals on September 25, 2021 methylglutaryl-Coenzyme A synthase 2; Hmgcl, 3-hydroxy-3-methylglutaryl-Coenzyme A lyase.

Fig. 4. Glucocorticoid metabolism. Gene expression from mice is given as detailed in legend to Fig. 2. Gene expression was evaluated by QPCR (default; n=5) or by microarrays

(indicated by black dot; n=3) and is given in percent of that in untreated db/db mice.

Hsd11b1, hydroxysteroid 11-beta dehydrogenase 1; H6pd, hexose-6-phosphate dehydrogenase (glucose 1-dehydrogenase); ut, untreated. Significant differences compared to untreated db/db are indicated as follows: ‡, p < 0.001.

Fig. 5. Growth hormone signaling. Gene expression from mice is given as detailed in legend to Fig. 2. Gene expression was evaluated by QPCR (default; n=5) or by microarrays

(indicated by black dot; n=3) and is given in percent of that in untreated db/db mice. Ghr,

26 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831 growth hormone receptor; Igf-1, insulin-like growth factor 1; ut, untreated. Significant differences compared to untreated db/db are indicated as follows: †, p < 0.01; ‡, p < 0.001.

Fig. 6. Expression of genes involved in endoplasmic reticulum stress. Gene expression from mice is given as detailed in legend to Fig. 2. Gene expression was evaluated by QPCR

(default; n=5) or by microarrays (indicated by black dot; n=3) and is given in percent of that in untreated db/db mice. Xbp1, X-box binding protein 1; Grp78 (Hspa5), heat shock 70kD

protein 5 (glucose-regulated protein); Dnajc3, DnaJ (Hsp40) homolog, subfamily C, member Downloaded from

3; Txndc7, thioredoxin domain containing protein-7; ut, untreated; *, p < 0.05; †, p < 0.01; ‡, p < 0.001.

jpet.aspetjournals.org

Supplementary Figure 1. Genes regulated by all nuclear receptor agonists in the direction to normalize (A) or aggravate (B) diabetes-induced alterations. Gene Ontology (GO) identifiers

were added as a functional annotation. Microarray probe sets representing identical genes at ASPET Journals on September 25, 2021 show up repeatedly.

Supplementary Figure 2. Clustering of genes regulated by all nuclear receptor agonists. The profiles cluster according to the pattern across all conditions including alterations between untreated diabetic and non-diabetic animals. Genes downregulated by the compounds are represented in the upper part, upregulated genes in the lower part of the figure. Several different patterns of response are observed; for example the upmost group of the lower half corresponds to genes that are up-regulated in all conditions including the diabetes-1 effect (i.e., db/+ vs. db/db animals). On the other hand, the downmost genes of the upper half correspond to genes with increased expression in db/+ vs. db/db animals but downregulated by all compounds.

27 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

Supplementary Figure 3. Hepatic gene expression profiles of drug-treated db/db and untreated db/+ compared to untreated db/db. Diabetic db/db mice were treated for 10 days with Rosi (R), Wy (W), or T09 (T), respectively, or left untreated as were non-diabetic db/+ mice. Probe sets numbered with affy ID# represent genes identified by genbank entry numbers (genbank ID#), gene symbols and names. The fold change difference to untreated db/db mice is given as a color code and in numbers. The mean estimate of expression in untreated db/db mice is related to the mean of all genes on each microarray that was set to Downloaded from 100.

jpet.aspetjournals.org

at ASPET Journals on September 25, 2021

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TABLE 1. Animal characteristics and blood analyses

Parameter (units) db/db db/+

untreated Rosi Wy T09 untreated

Body weight (g)

before treatment 42.7±1.8a 42.6±1.6 42.0±1.7 42.5±1.3 27.3±1.4***

end of treatment 43.7±1.7 46.7±1.9 42.3±2.7 43.4±2.4 27.9±1.4*** Downloaded from

Liver weight (g) 2.30±0.30 2.68±0.35 3.83±0.36*** 5.67±0.29*** 1.43±0.13***

Liver/body weight (%) 5.2±0.5 5.7±0.6 9.1±0.9*** 13.1±0.2*** 5.1±0.3 jpet.aspetjournals.org

Blood glucoseb (mmol/l)

before treatment 23.4±3.2 19.5±5.1 24.5±3.7 25.0±4.8 8.1±0.9*** at ASPET Journals on September 25, 2021

end of treatment 28.7±2.0 10.4±1.7*** 14.7±4.5*** 11.9±4.0*** 7.9±0.7***

Insulinc (nmol/l) 0.52±0.26 0.53±0.26 0.41±0.22 1.75±0.58*** 0.37±0.19

Triglyceridesc (mmol/l) 2.3±0.5 0.7±0.3*** 1.1±0.8** 4.4±1.7 1.7±0.5

Cholesterolc (mmol/l) 4.2±0.9 3.5±0.6 6.7±1.0*** 14.8±2.2*** 3.4±0.4

NEFAc (mmol/l) 0.25±0.04 0.14±0.02** 0.18±0.01 0.29±0.05 0.21±0.07

Liver triglycerides 36.9±9.3 63.1±11.1** 33.8±5.1 174.6±40.1** 5.3±2.2**

(nmol/mg liver)

a Data are given in means ± S.D.

b Blood parameters were analyzed from plasma unless stated otherwise.

c log-transformed for ANOVA.

d All groups were compared to untreated db/db; * p < 0.05; ** p < 0,01; *** p < 0.001. 29 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. JPET# 93831

TABLE 2. Overall statistics of changes in gene expression profiles

Effect Comparison vs Number of genesa

untreated db/db Increased Decreased Total

Diabetes-1b db/+ 155 182 337

PPARγ Rosi 158 142 300

PPARα Wy 362 258 620 Downloaded from

LXR T09 402 333 735

jpet.aspetjournals.org a Number of genes changed at least ±1.5-fold and a p<0.01. b The diabetes effect was given as reciprocal (db/+ vs db/db; diabetes-1) in order to facilitate direct comparisons with normalizing effects of drugs. at ASPET Journals on September 25, 2021

30 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 JPET Fast Forward. Published on October 31, 2005 as DOI: 10.1124/jpet.105.093831 This article has not been copyedited and formatted. The final version may differ from this version. Downloaded from jpet.aspetjournals.org at ASPET Journals on September 25, 2021 Supplementary Figure 1A List of genes regulated by all agonists in the same direction to db/+ fold changes estimate

affy ID# genbank ID# symbol gene name B W T + B W T + db/db

104617_at AI042964 0610005 RIKEN cDNA 0610005C13 gene -1,8 -2,5 -2,9 -1,5 795 C13Rik 104566_at AI845009 Asl argininosuccinate lyase -2,3 -4,7 -3,1 -2,1 226 102373_at M29961 Enpep glutamyl aminopeptidase -2,7 -3,5 -2,5 -2,0 260 95660_at AI851815 Ethe1 RIKEN cDNA 0610025L15 gene -1,7 -2,6 -1,9 -1,6 634 96918_at AI790931 Fbp1 fructose bisphosphatase 1 -1,7 -2,2 -2,5 -1,6 3975 103333_at U00445 G6pc glucose-6-phosphatase, catalytic -2,7 -2,9 -3,4 -2,9 2329 97430_at AF080469 G6pt1 glucose-6-phosphatase, transport protein 1 -1,9 -3,8 -2,7 -1,6 901 96828_at D89664 Gnmt glycine N-methyltransferase -1,5 -3,7 -3,3 -1,5 5650 97867_at X83202 Hsd11b1 hydroxysteroid 11-beta dehydrogenase 1 -1,8 -5,9 -4,0 -1,6 1624 95611_at AA726364 Lpl lipoprotein lipase 3,8 17,111,2 2,3 31 160083_at M63335 Lpl lipoprotein lipase 2,6 11,1 7,2 1,9 18 101473_at U86108 Nnmt nicotinamide N-methyltransferase -2,3 -7,0 -4,7 -1,7 1084 104343_f_at AI845798 Pla2g12a phospholipase A2, group XII 1,8 1,6 2,4 1,7 90 93926_at M22957 Prlr prolactin receptor -2,0 -2,2 -2,7 -2,6 236 102816_at X69832 Serpina3 serine (or cysteine) proteinase inhibitor, clade A, member -2,8 -3,6 -2,7 -1,8 696 m 3M 100341_g_at U95132 Slc10a1 solute carrier family 10 (sodium/bile acid cotransporter -1,8 -2,5 -4,2 -1,7 1414 family), member 1 104539_at AF022894 Sult1b1 sulfotransferase family 1B, member 1 -1,6 -2,3 -2,4 -1,6 169 93662_s_at AI386093 Zap70 zeta-chain (TCR) associated protein kinase -2,5 -3,2 -3,6 -1,8 173 93661_at U04379 Zap70 zeta-chain (TCR) associated protein kinase -1,7 -2,3 -2,2 -1,8 78

B ... BRL vs untreated db/db ; W ... Wy vs untreated db/db ; T... T09 vs untreated db/db ; +... db/+ vs db/db Fold change <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure 1B List of genes regulated by all agonists in the opposite direction to db/+ fold changes estimate

affy ID# genbank ID# symbol gene name B W T + B W T + db/db

99571_at AW012588 Acaa1 3-ketoacyl-CoA thiolase B 2,5 2,8 2,0 -2,5 4993 102004_at AI530403 Acaa1 3-ketoacyl-CoA thiolase B, acetyl-Coenzyme A 4,2 4,0 2,6 -2,7 836 acyltransferase 1 98589_at M93275 Adrp adipose differentiation related protein 2,3 4,1 2,8 -1,6 812 99559_at U14390 Aldh3a2 aldehyde dehydrogenase family 3, subfamily A2 2,0 4,1 2,1 -3,9 857 103703_f_at AI647632 C730048 RIKEN cDNA C730048C13 gene -1,8 -2,7 -2,7 1,7 312 C13Rik 103702_i_at AI647632 C730048 RIKEN cDNA C730048C13 gene -2,3 -3,6 -4,4 2,1 251 C13Rik 104424_at X05475 C9 complement component 9 -2,4 -5,4 -2,0 1,8 1044 103646_at X85983 Crat carnitine acetyltransferase 3,0 6,6 2,8 -1,6 106 103581_at Y14004 Cte1 cytosolic acyl-CoA thioesterase 1 2,2 20,0 4,6 -4,1 145 161345_f_at AV141027 Cyp7b1 cytochrome P450, family 7, subfamily b, polypeptide 1 -2,1 -2,0 -2,6 12,8 65 92898_at U36993 Cyp7b1 cytochrome P450, family 7, subfamily b, polypeptide 1 -2,6 -2,7 -3,6 12,8 108 98527_at Z14050 Dci dodecenoyl-Coenzyme A delta isomerase (3,2 1,6 3,7 1,6 -1,9 449 trans-enoyl-Coenyme A isomerase) 101842_g_at AW049716 Egfr epidermal growth factor receptor -3,1 -3,4 -3,1 2,5 219 101841_at AW049716 Egfr epidermal growth factor receptor -4,2 -5,2 -4,7 2,5 136 101840_at L06864 Egfr epidermal growth factor receptor -1,7 -1,8 -1,8 1,7 64 97316_at AJ011864 Ehhadh RIKEN cDNA 1300002P22 gene 4,0 22,1 6,9 -1,7 76 104325_at AI461631 Falp Fat cell-specific low molecular weight protein 2,1 2,1 1,7 -1,6 198 97820_at AB027012 Galk1 galactokinase 1 2,5 1,6 2,0 -1,6 99 101659_at M75886 Hsd3b2 hydroxysteroid dehydrogenase-2, delta<5>-3-beta -5,5 -5,6 -7,3 5,4 115 99669_at X15986 Lgals1 lectin, galactose binding, soluble 1 3,9 2,9 4,5 -2,0 107 104658_at D17444 Lifr leukemia inhibitory factor receptor -3,5 -3,6 -4,0 3,4 330 104659_g_at D17444 Lifr leukemia inhibitory factor receptor -3,5 -3,9 -4,1 3,2 197 160553_at X63782 Ly6d lymphocyte antigen 6 complex, locus D 2,2 4,0 7,7 -3,2 273 96258_at AI843448 Mgst3 microsomal glutathione S-transferase 3 6,8 3,4 2,6 -2,1 98 101082_at J02652 Mod1 malic enzyme, supernatant 3,7 5,8 3,7 -2,0 239 103660_at AF093669 Pex11a peroxisomal biogenesis factor 11a 2,7 5,5 2,2 -1,7 386 100927_at U28960 Pltp phospholipid transfer protein 8,2 7,6 4,6 -1,7 128 102013_at AF030513 Rdh6 retinol dehydrogenase 6 2,7 2,1 1,9 -3,1 154 96038_at AI840339 Rnase4 ribonuclease, RNase A family 4 -2,3 -1,6 -1,8 2,0 458 92242_at U65403 Saa4 serum amyloid A 4 -2,1 -2,6 -3,3 1,6 227

B ... BRL vs untreated db/db ; W ... Wy vs untreated db/db ; T... T09 vs untreated db/db ; +... db/+ vs db/db Fold change <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure 3. Hepatic gene expression profiles of drug-treated db/db and untreated db/+ compared to untreated db/db

fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

145 Cellular Processes 4 chemotaxis 16 apoptosis 102938_atAB009687 Lect2 leukocyte cell-derived chemotaxin 2 -1,7-2,1 -1,5 -1,0 374 102016_atM61737 Fsp27 fat specific gene 27 5,43,1 3,6 -1,3 20 103448_atM83218 S100a8 S100 calcium binding protein A8 (calgranulin 1,1-1,0 1,8 -1,1 17 103029_atD86344 Pdcd4 programmed cell death 4 1,2-1,1 -1,4 1,6 174 97360_atAI841689 Cklfsf3 chemokine-like factor super family 3 1,11,1 1,6 1,0 11 93968_atAA870917 Pdcd5 programmed cell death 5 -1,01,5 1,0 -1,1 241 103240_f_atAF017258 Ear2 eosinophil-associated, ribonuclease, RNase A 1,2-1,0 1,9 1,0 35 93766_atU32170 Rgn regucalcin -1,2-1,3 -1,6 -1,0 1356 38 cytoskeleton 93869_s_atU23781 Bcl2a1d Bcl-2-related protein A1 (BFL-1 protein) 1,01,1 1,6 1,0 24 160065_s_atAI837625 Csrp1 cysteine and glycine-rich protein 1 1,21,2 1,7 1,0 21 161500_i_atAV102186 Bad Bcl-associated death promoter -1,1-1,1 -1,1 1,5 50 96298_f_atAF020185 Dnclc1 dynein, cytoplasmic, light chain 1 -1,6-1,1 1,2 -1,1 315 161980_f_atAV373612 Bag3 Bcl2-associated athanogene 3 1,1-1,3 1,7 1,2 30 99441_atAW123852 Gphn gephyrin 1,01,0 -1,5 1,5 85 93536_atL22472 Bax Bcl2-associated X protein 1,21,3 2,0 -1,1 64 93571_atM74773 Spnb2 spectrin beta 2 1,11,2 1,5 1,1 148 93697_atU63387 Cbx4 chromobox homolog 4 (Drosophila Pc class) -1,5-1,7 -1,9 -1,2 62 102043_atAF047704 Tuft1 tuftelin 1 -1,01,1 1,9 1,7 54 93093_atU35623 Mcl1 myeloid cell leukemia sequence 1 1,2-1,0 1,6 1,4 66 162379_r_atAV245272 Vim vimentin 1,1-1,0 1,0 1,6 20 97825_atAI854029 Perp-pen p53 apoptosis effector related to Pmp22 1,21,7 1,3 -1,3 196 160150_f_atAW125626 Cnn3 calponin 3, acidic -1,1-1,6 -1,2 1,2 261 99994_atAF041376 Cidea cell death-inducing DNA fragmentation factor, 5,49,5 2,0 1,0 28 101578_f_atM12481 Actb actin, beta, cytoplasmic 2,21,0 1,6 1,5 119 102114_f_atAI326963 Angptl4 angiopoietin-like 4 -1,11,5 -1,0 -1,3 108 96573_atM21495 Actg actin, gamma, cytoplasmic -1,21,1 1,6 -1,1 573 96119_s_atAA797604 Angptl4 angiopoietin-like 4 1,11,6 1,1 -1,5 156 103574_atAI841606 AbLIM1 actin-binding LIM protein 1 1,31,5 1,5 1,2 27 97890_atAW046181 Sgk serum/glucocorticoid regulated kinase -1,11,2 1,5 1,0 34 104578_f_atAI195392 Actn1 actinin, alpha 1 1,11,4 1,6 1,7 66 101521_atAB013819 Birc5 baculoviral IAP repeat-containing 5 -1,11,2 1,5 -1,1 16 93364_atX59990 Catna1 catenin alpha 1 1,11,1 1,6 1,1 102 40 cell adhesion 98107_atAW123801 Coro1c coronin, actin binding protein 1C 1,21,3 1,7 1,1 71 161996_f_atAV328452 Bcan brevican -1,6-1,2 -1,8 -1,6 25 96865_atM60474 Marcks myristoylated alanine rich protein kinase C 1,31,5 1,8 1,1 27 104083_atAI853217 Cdh5 cadherin 5 1,31,2 1,6 1,2 33 94991_atAW046661 Synpo synaptopodin 1,42,1 1,6 -1,0 16 92835_atAI840501 Cml1 camello-like 1 -2,4-3,1 -3,1 1,1 588 96426_atU38967 Tmsb4x thymosin, beta 4, X chromosome 1,31,0 2,7 1,3 109 95016_atD50086 Nrp neuropilin 1,01,7 1,1 2,0 74 93100_atX13297 Acta2 actin, alpha 2, smooth muscle, aorta 1,11,0 1,8 1,3 24 100120_atL17324 Nid1 nidogen 1 1,41,3 1,7 1,1 11 92608_atD88793 Csrp1 cysteine and glycine-rich protein 1 1,01,1 1,5 1,0 29 93318_atU91513 Ninj1 ninjurin 1 -1,2-1,5 -1,3 -1,2 203 103330_atAI838709 Strbp spermatid perinuclear RNA binding protein 1,11,6 1,4 1,1 30 96774_atAW047139 Plekhc1 pleckstrin homology domain containing, family -1,2-1,3 -1,5 1,8 78 100446_r_atX91825 Sprr1b small proline-rich protein 1B -1,11,1 1,6 -1,1 1204 92558_atM84487 Vcam1 vascular cell adhesion molecule 1 1,2-1,0 1,9 1,2 18 100554_atAF053367 Pdlim1 PDZ and LIM domain 1 (elfin) 1,01,0 1,8 1,2 109 161188_f_atAV238648 Lamb3 laminin, beta 3 1,12,2 -1,0 -1,1 51 160417_atU86090 Kif5b kinesin family member 5B -1,21,0 -1,0 1,6 25 92759_atU43298 Lamb3 laminin, beta 3 1,57,2 1,1 -1,3 39 98462_s_atAI322968 Klc8 kinesin-like protein 8 -1,3-1,3 -1,2 -1,6 41 102805_atM77196 Ceacam1 CEA-related cell adhesion molecule 1 -1,6-1,8 -2,1 -1,5 278 99458_i_atX70764 Mark2 MAP/microtubule affinity-regulating kinase 2 -1,21,1 1,5 1,1 104 102806_g_atM77196 Ceacam1 CEA-related cell adhesion molecule 1 -1,7-1,8 -2,1 -1,5 130 97447_atAI836718 Map1lc3a microtubule-associated protein 1 light chain 3 1,91,5 1,4 -1,1 106 101908_s_atAF101164 Ceacam2 CEA-related cell adhesion molecule 2 -1,4-1,5 -1,7 -1,5 135 100915_atAW125698 Myh9 myosin heavy chain IX 1,11,1 1,7 -1,0 155 93063_atU82624 App amyloid beta (A4) precursor protein 1,21,3 1,8 1,1 155 98409_atL00923 Myo1b myosin IB -1,4-2,9 -2,0 1,2 220 92593_atD13664 Osf2-pen osteoblast specific factor 2 (fasciclin I-like) 1,71,2 2,0 -1,2 20 92254_atM55253 Myo5b myosin Vb 1,31,6 1,5 -1,3 77 93353_atAF013262 Lum lumican -1,1-1,1 1,7 1,4 22 160532_atM22479 Tpm1 tropomyosin 1, alpha 1,31,4 2,5 1,1 75 160583_atAA880988 Xlkd1 extra cellular link domain-containing 1 1,41,3 1,9 -1,2 18 94270_atM22832 Krt1-18 keratin complex 1, acidic, gene 18 -1,01,3 2,7 1,0 209 104337_f_atAW208938 Pkp2 plakophilin 2 1,71,4 1,4 -1,1 116 101009_atX15662 Krt2-8 keratin complex 2, basic, gene 8 1,21,5 3,1 -1,1 377 102852_atM31131 Cdh2 cadherin 2 -1,2-1,1 -1,3 2,1 64 100342_i_atM28729 Tuba1 tubulin, alpha 1 1,21,0 1,6 1,2 82 98549_atM77123 Vtn vitronectin -1,5-2,0 -1,6 1,1 2239 100343_f_atM28729 Tuba1 tubulin, alpha 1 1,1-1,0 1,6 1,0 301 100990_g_atAJ001373 Itgb1bp1 integrin beta 1 binding protein 1 -1,2-1,1 1,6 -1,1 38 98759_f_atM28727 Tuba2 tubulin, alpha 2 1,11,1 1,7 1,1 540 94963_atAI462105 Vcl vinculin 1,11,6 1,6 1,3 63 94835_f_atM28739 Tubb2 tubulin, beta 2 4,22,8 6,4 -1,8 395 104257_g_atAI120844 Pscdbp pleckstrin homology, Sec7 and coiled-coil 1,21,1 1,3 1,5 24 160462_f_atAW050256 Tubb3 tubulin, beta 3 2,42,0 3,4 -1,3 150 92880_atM38337 Mfge8 milk fat globule-EGF factor 8 protein 1,11,6 1,7 1,1 170 94788_f_atX04663 Tubb5 tubulin, beta 5 1,71,8 2,5 1,0 256 95706_atX16834 Lgals3 lectin, galactose binding, soluble 3 1,11,2 5,3 1,0 22 160461_f_atAW215736 Tubb6 tubulin, beta 6 1,31,6 2,4 -1,2 67 160099_atAF026799 Lgals4 lectin, galactose binding, soluble 4 1,11,7 -1,1 -1,3 54 5 peroxisome organization and biogenesis 99669_atX15986 Lgals1 lectin, galactose binding, soluble 1 3,92,9 4,5 -2,0 107 98965_atAC002397 Grcc3f, gene rich cluster, C3f gene 1,33,0 1,9 -1,4 141 95566_atAF050666 Gpld1 glycosylphosphatidylinositol specific -1,4-4,6 -2,5 1,3 288 95074_atAW125309 Pxf peroxisomal farnesylated protein 1,51,5 1,2 -1,4 96 160179_atAW124122 Col12a1 collagen, type XII, alpha1 -1,11,0 -1,1 1,6 54 104098_atL28835 Pxmp2 peroxisomal membrane protein 2 -1,0-1,3 -1,7 -1,1 1281 103709_atAA763466 Col1a1 procollagen, type I, alpha 1 1,11,2 1,8 1,1 58 94491_atAW123194 Pex13 peroxisomal biogenesis factor 13 1,21,6 -1,1 1,0 35 94305_atU03419 Col1a1 procollagen, type I, alpha 1 1,41,8 6,1 1,1 39 99469_atAW121865 Pex6 peroxisomal biogenesis factor 6 1,51,1 -1,1 1,2 127 101130_atX58251 Col1a2 procollagen, type I, alpha 2 1,11,2 2,4 1,1 45 299 Environmental Information Processing 161984_f_atAV234303 Col3a1 procollagen, type III, alpha 1 1,01,1 1,5 -1,0 10 3 angiogenesis 98331_atX52046 Col3a1 procollagen, type III, alpha 1 -1,01,3 3,0 -1,1 34 94392_f_atU22516 Ang angiogenin -1,8-1,4 -1,4 2,6 348 101093_atM15832 Col4a1 procollagen, type IV, alpha 1 1,61,5 2,6 1,2 47 101735_f_atU22519 Ang2 angiogenin, ribonuclease A family, member 2 -1,3-1,2 -1,1 1,8 50 101039_atX04647 Col4a2 procollagen, type IV, alpha 2 1,21,6 1,8 1,1 93 160484_atAI840118 Rtn4 reticulon 4 1,52,3 1,3 -2,0 185 92567_atL02918 Col5a2 procollagen, type V, alpha 2 1,21,2 2,3 1,1 17 17 blood coagulation 101110_atAF064749 Col6a3 procollagen, type VI, alpha 3 1,11,2 1,8 1,2 57 99638_atD17546 Col18a1 procollagen, type XVIII, alpha 1 -1,2-1,3 -1,6 -1,1 878 102291_atM23109 F9 coagulation factor IX -1,2-1,4 -1,8 1,1 335 93934_atAF072128 Cldn2 claudin 2 -1,2-1,3 -1,1 -1,6 195 97435_atD10071 F13b coagulation factor XIII, beta subunit -1,4-1,7 -1,7 -1,1 197 96868_atAI196896 Fgb fibrinogen, B beta polypeptide -1,3-1,4 -1,5 1,3 672 25 cell cycle 93096_atAA986050 FGG fibrinogen, gamma polypeptide -1,3-1,8 -1,6 1,4 1362 96146_atD83745 Btg3 B-cell translocation gene 3 -1,11,2 1,6 1,6 16 161458_atAV298640 F7 coagulation factor VII -1,4-1,5 -1,4 1,2 534 101906_atAA032310 Smc4l1 SMC4 structural maintenance of 1,01,8 1,6 1,0 23 92918_atU66079 F7 coagulation factor VII , serum prothrombin -1,5-2,6 -1,9 1,2 201 160623_atAI847045 Cdkl2 cyclin-dependent kinase-like 2 (CDC2-related 1,21,5 1,5 1,1 21 102748_atU52925 F5 coagulation factor V -1,7-2,0 -1,7 1,1 317 96319_atAW061324 Cdc20 cell division cycle 20 homolog (S. cerevisiae) 1,01,6 2,0 1,0 68 104344_atM58588 Klkb1 kallikrein B, plasma 1 -1,3-1,7 -2,3 1,4 141 94048_atAW120683 Cdc34 cell division cycle 34 homolog (S. cerevisiae) 1,51,2 1,6 -1,2 178 95585_atAF034569 Proc protein C -1,3-1,3 -1,6 1,1 976 103821_atAJ223087 Cdc06 cell division cycle 6 homolog (S. cerevisiae) 1,11,6 2,1 1,4 16 104519_atX70392 Itih2 inter-alpha trypsin inhibitor, heavy chain 2 -1,7-1,2 -1,4 -1,0 1777 94294_atX66032 Ccnb2 cyclin B2 1,01,5 2,0 1,3 48 160611_atAA212964 Klkb1 cytochrome P450, family 4, subfamily v, -1,3-1,2 -1,9 1,1 261 94232_atAI849928 Ccnd1 cyclin D1 1,12,9 4,7 1,7 30 100002_atX70393 Itih3 inter-alpha trypsin inhibitor, heavy chain 3 -2,4-1,3 -1,9 1,2 670 98478_atU95826 Ccng2 cyclin G2 1,21,5 -1,0 1,1 44 94383_atD50586 Tfpi2 tissue factor pathway inhibitor 2 1,31,8 -1,3 -1,0 186 102781_atU37351 Ccnl2 cyclin L2 1,0-1,0 -1,1 1,5 115 98014_atX70391 Itih1 inter-alpha trypsin inhibitor, heavy chain 1 -1,3-2,0 -1,9 1,2 385 101638_s_atD26137 Cdk8 cyclin-dependent kinase 8, cytochrome P450, 1,2-1,7 1,3 -1,8 3872 92858_atAF002719 Slpi secretory leukocyte protease inhibitor 1,21,1 3,3 -1,1 50 101639_r_atD26137 Cdk8 cyclin-dependent kinase 8, cytochrome P450, -1,0-1,4 1,2 -1,6 48 98467_atAF023919 Itih4 inter alpha-trypsin inhibitor, heavy chain 4 -1,7-5,6 -1,6 1,1 843 94881_atAW048937 Cdkn1a cyclin-dependent kinase inhibitor 1A (P21) 1,21,7 2,0 -1,0 58 103977_atAF087644 F10 coagulation factor X -1,3-1,7 -1,6 1,1 380 97411_atL11316 Ect2 ect2 oncogene -1,11,5 1,6 1,1 12 1 blood pressure regulation 103309_atAI853977 Frap1 FK506 binding protein 12-rapamycin 1,22,0 1,3 -1,2 92 94971_atAI317217 Cdkn3 Mus musculus cDNA clone MGC:58593 1,11,3 2,0 1,0 26 101887_atAF045887 Agt angiotensinogen -1,4-1,8 -2,1 -1,8 1987 160846_atAI846534 Nek6 NIMA (never in mitosis gene a)-related 1,41,3 1,7 1,2 54 10 immune defense 94024_atAA671429 Ris2 retroviral integration site 2 -1,01,4 1,6 1,0 78 101054_atX00496 Ii Ia-associated invariant chain 1,2-1,1 2,0 1,4 139 92770_atX66449 S100a6 S100 calcium binding protein A6 (calcyclin) 1,11,1 3,8 -1,1 19 99109_atM59821 Ier2 immediate early response 2 -1,1-1,2 -1,1 1,5 49 97531_atX95280 G0s2 G0/G1 switch gene 2 1,93,6 1,2 -2,9 352 92773_atAF079528 Ier5 immediate early response 5 1,11,4 1,1 1,8 17 94813_atX65128 Gas1 growth arrest specific 1 -1,2-1,4 -1,7 -1,3 93 98822_atX56602 G1p2 interferon, alpha-inducible protein -1,1-1,5 1,3 1,5 74 94338_g_atM21828 Gas2 growth arrest specific 2 -1,81,2 1,2 -1,8 129 160253_atAW125390 Ifitm3 interferon induced transmembrane protein 3 ; -2,1-4,4 -1,1 1,4 1105 99067_atX59846 Gas6 growth arrest specific 6 1,31,4 3,9 -1,3 64 97409_atU19119 Ifi1 interferon inducible protein 1 -1,3-1,2 -1,1 1,6 39 99632_atU83902 Mad2l1 MAD2 (mitotic arrest deficient, homolog)-like 1,11,8 1,9 -1,0 22 100981_atU43084 Ifit1 interferon-induced protein with -1,3-1,4 -1,0 1,8 17 98603_s_atU20857 Rangap1 RAN GTPase activating protein 1 -1,6-1,3 -1,4 -1,4 117 103639_atU43085 Ifit2 interferon-induced protein with 1,01,0 1,6 -1,0 16 10 cell growth 103963_f_atAA914345 Iigp-pendi interferon-inducible GTPase -1,5-1,2 -1,5 1,8 173 97426_atX98471 Emp1 epithelial membrane protein 1 1,01,2 1,8 1,0 21 96764_atAJ007971 Iigp-pendi interferon-inducible GTPase -1,61,0 -1,3 2,4 330 104646_atAJ011967 Gdf15 growth differentiation factor 15 1,21,8 2,0 -1,1 82 99452_atU49507 Lisch7-pe liver-specific bHLH-Zip transcription factor 1,1-1,1 1,5 1,1 334 100327_atM17015 Lta lymphotoxin alpha (TNF superfamily, member 1,21,0 1,1 1,6 45 160553_atX63782 Ly6d lymphocyte antigen 6 complex, locus D 2,24,0 7,7 -3,2 273 96632_atAB025049 Morf4l2 mortality factor 4 like 2 -1,0-1,0 1,5 1,1 241 94425_atAB007599 Ly86 lymphocyte antigen 86 1,01,0 1,6 1,0 24 94932_atM29464 Pdgfa platelet derived growth factor, alpha -1,01,1 1,6 1,0 93 99071_atL20315 Mpeg1 macrophage expressed gene 1 1,2-1,2 3,6 1,7 83 104533_atAA764261 Pim1 proviral integration site 1 1,2-1,1 1,0 1,5 193 95723_r_atAI841464 2610009E Parathymosin (zinc binding 11.5 kDA protein -1,2-1,6 -1,3 -1,1 818 100494_atM30641 Fgf1 fibroblast growth factor 1 -1,4-1,3 -1,9 -1,4 94 96038_atAI840339 Rnase4 ribonuclease, RNase A family 4 -2,3-1,6 -1,8 2,0 458 103001_atU43836 Vegfb vascular endothelial growth factor B 1,51,5 1,5 -1,0 81 103794_i_atAA986114 Timd2 T-cell immunoglobulin and mucin domain -1,7-1,5 -1,6 1,0 31 160292_atAW123987 Ecgf1 endothelial cell growth factor 1 1,11,1 -1,7 -1,2 490 97335_atAA795198 Timd2 T-cell immunoglobulin and mucin domain -1,5-2,0 -1,7 -1,2 224 161666_f_atAV138783 Gadd45b growth arrest and DNA-damage-inducible 45 1,43,0 2,2 -1,1 17 100397_atAF024637 Tyrobp TYRO protein tyrosine kinase binding protein 1,21,0 2,4 1,2 44 93661_atU04379 Zap70 zeta-chain (TCR) associated protein kinase -1,7-2,3 -2,2 -1,8 78 5 cell proliferation 93662_s_atAI386093 Zap70 zeta-chain (TCR) associated protein kinase -2,5-3,2 -3,6 -1,8 173 92471_i_atAF099973 Slfn2 schlafen 2 1,11,2 1,6 -1,0 53 103422_atM63695 Cd1d1 CD1d1 antigen 1,01,2 -1,6 -1,3 455 162014_i_atAV248951 Tax1bp3 Tax1 (human T-cell leukemia virus type I) -1,4-2,0 -1,9 -1,4 194 101897_g_atM63697 Cd1d2 CD1d2 antigen -1,01,4 -1,5 -1,7 945 93327_atAI842665 Tax1bp3 Tax1 (human T-cell leukemia virus type I) -1,1-1,1 1,6 -1,3 62 93445_atAF011428 Cd5l CD5 antigen-like 1,31,1 5,1 1,6 35 99191_atAI844939 Cri1 CREBBP/EP300 inhibitory protein 1 1,31,5 1,5 1,4 19 95661_atL08115 Cd9 CD9 antigen -1,11,9 2,2 -1,1 39 97228_atAI844417 Lzf leucine zipper domain protein 1,51,6 1,3 -1,1 137 98088_atX13333 Cd14 CD14 antigen 1,01,1 1,5 1,2 88 2 cell shape 100600_atM58661 Cd24a CD24a antigen 1,02,7 1,4 1,1 23 97160_atX04017 Sparc secreted acidic cysteine rich glycoprotein 1,71,7 4,0 1,1 27 93332_atL23108 Cd36 CD36 antigen 1,32,1 1,3 -1,4 33 98059_s_atD49733 Lmna lamin A 1,21,3 2,1 -1,1 142 94939_atX97227 Cd53 CD53 antigen 1,21,0 2,2 1,2 12 160493_atD16432 Cd63 Cd63 antigen 1,11,6 5,5 1,1 26

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 1 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

103016_s_atX68273 Cd68 CD68 antigen -1,0-1,0 1,8 1,2 34 99596_f_atM13963 Gnai2 guanine nucleotide binding protein, alpha 1,11,0 1,6 1,2 411 98129_atAI852553 Tmsb10 thymosin, beta 10 1,31,4 4,0 -1,2 68 99598_g_atAI841629 Gnai2 guanine nucleotide binding protein, alpha 1,11,1 1,6 1,1 290 99862_atAF025821 Ahsg alpha-2-HS-glycoprotein -1,6-2,3 -1,6 1,3 4606 101441_i_atAF031127 Itpr5 inositol 1,4,5-triphosphate receptor 5 -1,01,1 -1,2 1,9 32 161827_f_atAV105307 Hpxn hemopexin 1,2-1,5 2,0 1,6 1078 160530_atAW120976 Ghitm growth hormone inducible transmembrane 1,01,5 1,0 1,0 102 98116_atU89889 Hpxn hemopexin 1,2-1,5 1,9 1,4 1823 99107_atM31680 Ghr growth hormone receptor -1,7-1,7 -2,1 -1,2 214 102575_atS38219 Orm3 orosomucoid 3 1,5-1,8 1,7 -1,8 112 99108_s_atU15012 Ghr growth hormone receptor -1,7-2,3 -2,5 -1,0 496 96787_atAA880891 Serpina10 serine (or cysteine) proteinase inhibitor, clade -1,3-1,5 -1,1 1,4 441 104658_atD17444 Lifr leukemia inhibitory factor receptor -3,5-3,6 -4,0 3,4 330 160951_atAI645694 Serpina12 serine (or cysteine) proteinase inhibitor, clade -1,3-1,5 -1,4 8,2 47 104659_g_atD17444 Lifr leukemia inhibitory factor receptor -3,5-3,9 -4,1 3,2 197 101565_f_atM75720 Serpina1a serine (or cysteine) proteinase inhibitor, clade -1,3-1,7 -1,8 1,1 ? 99142_atAI834895 2310021 membrane progestin receptor alpha 1,91,9 1,2 1,0 96 101572_f_atM75721 Serpina1a serine (or cysteine) proteinase inhibitor, clade -1,4-2,0 -2,0 1,3 7060 104354_atX06368 Csf1r colony stimulating factor 1 receptor 1,2-1,0 1,5 1,2 50 101576_f_atM25529 Serpina1b serine (or cysteine) proteinase inhibitor, clade -1,2-1,7 -1,7 1,1 8856 160674_atAI596094 Trfr2 transferrin receptor 2 -1,2-1,4 -1,6 -1,3 547 100329_atX00945 Serpina1d serine (or cysteine) proteinase inhibitor, clade -1,4-2,2 -2,2 1,2 7106 92847_s_atX56831 M6pr mannose-6-phosphate receptor, cation 1,21,6 1,3 1,1 53 93109_f_atM75718 Serpina1d serine (or cysteine) proteinase inhibitor, clade -1,2-1,7 -1,8 1,1 8718 94828_atAF004927 Oprs1 opioid receptor, sigma 1 2,01,8 1,7 -1,3 428 101574_f_atM75717 Serpina1e serine (or cysteine) proteinase inhibitor, clade -1,3-1,9 -2,0 1,3 6961 97507_atX67809 Lgals3bp lectin, galactose binding, soluble, 3 binding 1,21,3 1,8 -1,0 84 102707_f_atX61597 Serpina3c serine (or cysteine) proteinase inhibitor, clade -1,7-3,0 -2,2 -1,1 700 94801_atAI607332 Pgrmc2 progesterone receptor membrane component 1,61,6 1,1 -1,1 35 92583_atD00725 Serpina3k serine (or cysteine) proteinase inhibitor, clade -1,9-3,0 -3,3 -1,1 64 96104_atAI047107 3732413I RIKEN cDNA 3732413I11 gene 1,31,6 3,2 1,2 30 102816_atX69832 Serpina3 serine (or cysteine) proteinase inhibitor, clade -2,8-3,6 -2,7 -1,8 696 96534_atAA408956 Vldlr very low density lipoprotein receptor 1,32,0 1,1 -1,0 11 104374_atM64086 Serpina3n serine (or cysteine) proteinase inhibitor, clade -1,6-1,5 -1,5 1,0 806 55 signal transduction 96227_atX70533 Serpina6 serine (or cysteine) proteinase inhibitor, clade -1,31,1 -1,8 -1,6 859 99106_atAF071315 Cops6 COP9 (constitutive photomorphogenic) 1,21,8 1,2 -1,3 448 96060_atU25844 Serpinb6a serine (or cysteine) proteinase inhibitor, clade 1,31,3 4,3 1,7 20 96592_atU50413 Pik3r1 phosphatidylinositol 3-kinase, regulatory -1,11,5 1,2 -1,4 54 100966_atU07425 Serpind1 serine (or cysteine) proteinase inhibitor, clade -1,6-1,5 -1,8 1,1 1025 102338_atAF031147 Pde9a phosphodiesterase 9A -1,2-1,6 -1,3 1,1 34 101928_atZ36774 Serpinf2 serine (or cysteine) proteinase inhibitor, clade -1,5-2,3 -1,9 1,3 590 100555_atAI846152 Dscr1 Down syndrome critical region homolog 1 -1,3-1,7 -1,1 -2,4 131 94817_atX60676 Serpinh1 serine (or cysteine) proteinase inhibitor, clade -1,11,1 1,7 -1,1 32 161610_atAV349686 Ndr2 N-myc downstream regulated 2 -1,4-1,8 -2,7 1,5 419 97444_atAI844520 Ifi30 interferon gamma inducible protein 30 1,21,2 2,3 1,0 39 96088_atAB033921 Ndr2 N-myc downstream regulated 2 -1,3-1,9 -2,4 1,1 1165 104407_atL25274 Alcam activated leukocyte cell adhesion molecule -1,1-1,0 1,2 1,8 17 104268_atX51975 Il6ra interleukin 6 receptor, alpha -1,5-1,9 -1,9 -1,0 92 93454_atAF081789 C1qr1 complement component 1, q subcomponent, 1,01,0 1,6 -1,1 31 102321_atM93422 Adcy6 adenylate cyclase 6 1,31,5 1,2 -1,0 54 102961_atAI118358 Hrg histidine-rich glycoprotein -1,7-2,7 -3,1 -1,0 1043 104412_atAI153412 Gnai1 guanine nucleotide binding protein, alpha 1,41,8 1,1 -1,1 27 100882_atAF003525 Defb1 defensin beta 1 1,22,2 1,2 -1,1 19 161609_atAV349152 Rgs16 regulator of G-protein signaling 16 1,0-2,8 -2,4 -2,1 83 92852_atM18194 Fn1 fibronectin 1 -1,5-1,3 -1,4 -1,1 408 93783_atM27347 Ela1 elastase 1, pancreatic 1,21,0 1,1 2,4 120 104388_atU49513 Syca9 chemokine (C-C motif) ligand 9 1,81,2 2,6 1,5 127 97227_atM63659 Gna12 guanine nucleotide binding protein, alpha 12 -1,11,0 -1,3 -1,8 43 99671_atX04673 Adn adipsin 13,5-1,0 -1,1 1,1 34 92647_atU35141 Rbbp4 retinoblastoma binding protein 4 -1,1-1,1 -1,1 1,8 52 104424_atX05475 C9 complement component 9 -2,4-5,4 -2,0 1,8 1044 93559_atD90374 Apex1 apurinic/apyrimidinic endonuclease 1 1,11,5 1,4 1,1 105 101853_f_atM12660 Cfh complement component factor h -1,3-1,5 -1,7 1,2 195 97458_atAI845935 Gnb1 guanine nucleotide binding protein, beta 1 1,51,0 1,4 1,2 114 94743_f_atM29009 Cfh complement component factor h -1,6-2,1 -2,1 1,2 300 93771_atX84215 Gjb1 gap junction membrane channel protein beta -1,4-1,3 -1,7 -1,1 1025 94994_atM29007 Cfh complement component factor h -1,8-1,8 -1,9 -1,2 62 102366_atAA718169 Retn resistin 3,7-1,2 -1,1 1,0 47 103458_atM35525 Hc hemolytic complement -1,5-1,9 -1,4 2,2 535 93212_atZ97207 AW74231 butyrate-induced transcript 1 1,02,1 1,0 -1,8 81 101468_atX12905 Pfc properdin factor, complement 1,21,0 1,8 -1,0 23 160320_atU58883 Sorbs1 sorbin and SH3 domain containing 1 1,61,8 1,4 -1,3 114 98562_atX58861 C1qa complement component 1, q subcomponent, 1,2-1,1 2,1 1,2 80 97803_atU38196 Mpp1 membrane protein, palmitoylated 1,22,1 1,9 1,0 116 96020_atM22531 C1qb complement component 1, q subcomponent, 1,5-1,0 3,1 1,2 61 93747_atAW122599 Hspc121- butyrate-induced transcript 1 1,12,2 1,2 -1,0 644 92223_atX66295 C1qg complement component 1, q subcomponent, 1,1-1,0 1,9 1,0 140 97909_atAI838080 Stmn1 stathmin 1/oncoprotein 18 1,11,9 2,7 -1,2 54 103673_atM57891 C2 complement component 2 (within H-2S) -1,1-1,4 -1,1 1,7 109 103020_s_atAI317205 Map3k1 mitogen activated protein kinase kinase 1,21,1 1,4 1,6 26 102799_atM17122 C4bp complement component 4 binding protein -1,5-2,8 -1,7 -1,2 927 102195_atU88984 Map4k4 mitogen-activated protein kinase kinase 1,31,4 1,6 -1,0 17 103033_atX06454 C4 complement component 4 (within H-2S) 1,1-1,8 -1,1 1,5 1675 93285_atAI845584 Dusp6 dual specificity phosphatase 6 1,21,2 2,0 -1,4 57 99927_atU47810 Cfi complement component factor i -1,3-1,5 -1,3 1,2 1132 161257_r_atAV321519 Snx17 sorting nexin 17 -1,2-1,3 -1,5 2,4 61 101474_atU09010 Mbl1 mannose binding lectin, liver (A) -1,3-1,6 -1,7 -1,2 235 96199_atAF073996 Mtm1 X-linked myotubular myopathy gene 1 1,11,8 1,2 1,0 23 97427_atU09016 Mbl2 mannose binding lectin, serum (C) -2,0-3,0 -2,1 1,0 306 98927_atAI851048 Rab6 RAB6, member RAS oncogene family -1,6-1,1 -1,1 1,1 142 99081_atAF010254 Serping1 serine (or cysteine) proteinase inhibitor, clade -1,7-1,6 -1,8 -1,0 2005 101254_atL32751 Ran RAN, member RAS oncogene family 1,01,5 1,4 -1,1 850 104726_atAA670737 C8G Complement component C8 gamma chain 1,0-1,6 -1,6 1,2 1187 162011_f_atAV246963 Arhu ras homolog gene family, member U 1,31,5 1,4 -1,1 115 92291_f_atM29008 Cfhl1 complement component factor h-like 1 -1,2-1,7 -1,8 1,3 188 96747_atAW121294 Arhu ras homolog gene family, member U 1,81,9 1,8 1,1 106 102736_atM19681 Ccl2 chemokine (C-C motif) ligand 2 = MCP-1 1,11,0 3,7 -1,0 12 97991_atX02452 Kras2 Kirsten rat sarcoma oncogene 2, expressed 1,11,3 1,6 1,0 19 95348_atJ04596 Cxcl1 chemokine (C-X-C motif) ligand 1 1,21,1 1,3 2,7 40 94394_atM21019 Rras Harvey rat sarcoma oncogene, subgroup R 1,31,2 1,9 -1,1 83 95349_g_atJ04596 Cxcl1 chemokine (C-X-C motif) ligand 1 1,21,0 1,2 2,6 41 103062_atAB004664 Rab33b RAB33B, member of RAS oncogene family -1,1-1,0 -1,0 1,6 38 100112_atL12030 Cxcl12 chemokine (C-X-C motif) ligand 12 -1,5-1,5 -1,8 -1,1 212 95516_atAB027290 Rab9 RAB9, member RAS oncogene family 1,12,3 1,3 -1,1 190 160511_atL12029 Cxcl12 chemokine (C-X-C motif) ligand 12 -1,1-1,3 -1,5 1,1 64 95547_atD89821 Rhod ras homolog gene family, member D 1,31,3 1,6 -1,1 98 162234_f_atAV139913 Cxcl12 chemokine (C-X-C motif) ligand 12 -1,2-1,4 -1,6 -1,1 60 94362_atAI843682 Nras neuroblastoma ras oncogene 1,11,5 1,6 1,6 54 97519_atX13986 Spp1 secreted phosphoprotein 1 -1,11,6 3,6 1,2 129 101030_atX99963 Arhb ras homolog gene family, member AB 1,01,4 1,8 1,2 58 101487_f_atU47737 Ly6e lymphocyte antigen 6 complex, locus E -1,2-1,9 1,8 2,1 1113 96056_atX80638 Arhc ras homolog gene family, member C 1,11,2 1,6 1,0 56 97312_atAB014464 Cd164 CD164 antigen -1,2-1,2 -1,1 1,5 262 93687_r_atU39904 Cit citron -1,3-1,4 -1,7 1,2 32 99624_atAW125517 Igk-V4 immunoglobulin kappa chain variable 4 (V4) 1,21,4 1,6 1,1 160 160840_atAI853706 Arhgef3 Rho guanine nucleotide exchange factor 1,21,8 1,2 1,0 24 104750_atM63630 Ifi47 interferon gamma inducible protein -1,1-1,2 1,0 2,4 44 160977_atAA726063 Arhgef5 Rho guanine nucleotide exchange factor 1,21,3 1,6 1,1 18 93775_atAI841894 D12Ertd6 DNA segment, Chr 12, ERATO Doi 647, -1,3-1,3 1,3 1,7 330 96360_atAW125498 Arhgdia Rho GDP dissociation inhibitor (GDI) alpha 1,41,5 2,0 1,2 204 100611_atM21050 Lyzs lysozyme 1,01,1 3,2 1,5 26 98485_atAI850288 Rhpn2 rhophilin, Rho GTPase binding protein 2 1,21,6 2,3 1,2 12 101078_atY16258 Bsg basigin 1,71,7 1,5 -1,7 428 98545_atX78683 Bcap37 B-cell receptor-associated protein 37 1,31,8 1,0 -1,2 270 92866_atX52643 H2-Aa histocompatibility 2, class II antigen A, alpha 1,1-1,1 1,9 1,2 95 96033_atZ22532 Sdc1 syndecan 1 -1,3-1,6 -1,3 -1,7 202 102904_atV00833 H2-Ea histocompatibility 2, class II antigen E alpha 1,21,1 2,6 1,3 30 100561_atAW209098 Iqgap1 IQ motif containing GTPase activating protein 1,11,2 1,6 1,1 17 97541_f_atX00246 H2-D1 histocompatibility 2, D region locus 1 1,2-1,2 1,2 1,5 434 104432_atAF016482 Arhn ras homolog N (RhoN) 1,1-1,5 -1,0 -1,8 92 92837_f_atM65736 Mug1 murinoglobulin 1 -2,8-4,0 -4,5 1,2 1406 98084_atAI849834 Arl2bp ADP-ribosylation factor-like 2 binding protein 1,41,5 1,6 1,0 129 AFFX-MurIL4M25892 Il4 interleukin 4 1,81,1 -1,3 -1,4 565 99475_atU88327 Socs2 suppressor of cytokine signaling 2 -1,01,1 1,0 1,7 21 100325_atM65027 Gp49a glycoprotein 49A 1,01,1 1,5 1,1 9 160758_atAW124855 Sec13l-pe RIKEN cDNA 2610007A16 gene 1,21,6 1,2 1,3 27 94448_atAJ006289 Bcl10 B-cell leukemia/lymphoma 10 1,31,4 1,5 1,1 47 162206_f_atAV374868 Socs3 suppressor of cytokine signaling 3 -1,1-1,1 -1,0 1,6 26 100436_atM27008 Orm1 orosomucoid 1 -1,2-1,8 -1,0 1,2 995 96935_atAW011791 Map17-pe RIKEN cDNA 2700030M23 gene 1,51,6 1,9 1,2 72 100437_g_atM27008 Orm1 orosomucoid 1 -1,1-2,0 1,3 1,1 4717 93896_atD13903 Ptprd protein tyrosine phosphatase, receptor type, -1,3-1,4 -1,6 -1,1 74 94734_atM12566 Orm2 orosomucoid 2 -1,1-2,4 2,5 -3,7 507 162231_r_atAV309750 Mesdc2 mesoderm development candiate 2 -1,3-1,5 -1,3 -1,5 19 104606_atM55561 Cd52 CD52 antigen 1,0-1,0 1,8 1,0 35 11 stress response 96867_atD44593 Azgp1 alpha-2-glycoprotein 1, zinc -1,2-1,4 -1,7 -1,1 1070 96048_atU50631 Hrsp12 heat-responsive protein 12 -1,0-1,0 -1,7 -1,2 459 101516_atU60473 Cd59a CD59a antigen -1,2-2,4 -2,0 -1,5 418 95057_atAI846938 Herpud1 homocysteine-inducible, endoplasmic -1,4-1,7 -2,0 -1,1 475 95415_f_atAI132585 C1r complement component 1, r subcomponent -1,4-2,1 -1,3 1,7 183 96831_atAA755004 Pdir-pendi protein disulfide isomerase-related -1,5-2,0 -1,8 1,1 104 7 insulin-like growth factor 162220_r_atAV111954 Ucp2 uncoupling protein 2 (mitochondrial, proton -1,5-1,2 -1,7 -1,7 26 95117_atU04710 Igf2r insulin-like growth factor 2 receptor 1,21,2 1,6 1,2 64 94343_atAI604013 Dnajc3 DnaJ (Hsp40) homolog, subfamily C, member -1,2-1,4 -1,3 1,5 80 98627_atX81580 Igfbp2 insulin-like growth factor binding protein 2 -1,32,3 1,7 5,7 173 101955_atAJ002387 Hspa5 heat shock 70kD protein 5 (glucose-regulated -1,3-1,8 -1,6 1,5 841 101571_g_atX76066 Igfbp4 insulin-like growth factor binding protein 4 -1,4-2,4 -2,4 -1,1 1209 95282_atJ04633 Hspca heat shock protein 1, alpha 1,11,7 1,7 1,2 181 100566_atL12447 Igfbp5 insulin-like growth factor binding protein 5 -2,4-2,4 -1,3 -2,3 75 93277_atX53584 Hspd1 heat shock protein 1 (chaperonin) 1,21,7 1,2 1,1 458 97987_atU66900 Igfals insulin-like growth factor binding protein, acid -1,8-1,8 -2,5 1,1 294 95654_atAF109905 Hspa1l heat shock protein 1-like 1,21,1 2,2 1,1 32 95545_atX04480 Igf1 insulin-like growth factor 1 -1,5-1,8 -2,0 -1,1 803 93875_atM12571 Hspa1a heat shock protein 1A -1,11,0 1,9 -1,1 30 95546_g_atX04480 Igf1 insulin-like growth factor 1 -2,0-2,9 -3,1 -1,3 801 95505_atAW060509 Tor1b torsin family 1, member B 1,11,6 1,5 1,2 36 1 protein amino acid phosphorylation 2 thyroid hormone 97429_atAW048113 Snrk SNF related kinase 1,72,0 1,3 1,0 57 95350_atD00073 Ttr transthyretin -1,2-2,3 -1,8 1,2 854 38 receptor 95552_atU49861 Dio1 deiodinase, iodothyronine, type I 1,7-1,5 -2,2 3,2 74 93433_s_atD13517 Asgr1 asialoglycoprotein receptor 1 -1,2-1,8 -1,6 -1,6 1360 40 transporter 96674_atAW123553 D6Ertd31 DNA segment, Chr 6, ERATO Doi 313, 1,21,2 1,2 1,5 48 94563_atAJ011080 Afm afamin -1,3-1,4 -1,8 1,4 189 92937_atX59927 Fgfr4 fibroblast growth factor receptor 4 -1,4-1,3 -1,5 -1,2 77 92531_atAI844545 Tloc1 translocation protein 1 -1,01,0 -1,1 1,6 21 93872_atAF014117 Gfra1 glial cell line derived neurotrophic factor family -2,1-2,5 -2,0 -1,3 77 160590_r_atAI853323 Vdac2 voltage-dependent anion channel 2 1,31,1 1,6 -1,4 164 99926_atAB001489 Pigr polymeric immunoglobulin receptor -1,4-4,6 -4,3 1,6 1838 102703_s_atU48398 Aqp4 aquaporin 4 1,2-1,2 -1,9 -2,6 49 93926_atM22957 Prlr prolactin receptor -2,0-2,2 -2,7 -2,6 236 102704_atU88623 Aqp4 aquaporin 4 1,3-1,2 -1,9 -3,0 73 95030_atD10214 Prlr prolactin receptor -1,2-1,2 -1,2 -1,6 94 102200_atAF018952 Aqp8 aquaporin 8 1,71,2 2,6 1,0 189 95586_atAF089751 P2rx4 purinergic receptor P2X, ligand-gated ion 1,21,2 1,6 1,6 99 103899_atAA690863 Atp11a ATPase, class VI, type 11A 1,21,1 1,4 1,6 11 93871_atL32838 Il1rn interleukin 1 receptor antagonist 1,22,7 1,4 -1,1 26 94255_g_atAI845237 Clic4 chloride intracellular channel 4 (mitochondrial) 1,21,3 1,5 1,4 56 93914_atM20658 Il1r1 interleukin 1 receptor, type I -1,11,0 -1,0 1,9 30 97288_atAW260404 Pdzk1 PDZ domain containing 1 1,1-1,3 1,2 1,9 49 99491_atU53696 Il10rb interleukin 10 receptor, beta 1,31,2 1,7 1,3 45 94464_atAF029347 Clcn3 chloride channel 3 1,51,6 1,2 1,4 40 101840_atL06864 Egfr epidermal growth factor receptor -1,7-1,8 -1,8 1,7 64 93573_atV00835 Mt1 metallothionein 1 -1,2-1,2 -1,1 -10,7 1422 101841_atAW049716 Egfr epidermal growth factor receptor -4,2-5,2 -4,7 2,5 136 101561_atK02236 Mt2 metallothionein 2 -1,4-1,6 -1,3 -11,1 1527 101842_g_atAW049716 Egfr epidermal growth factor receptor -3,1-3,4 -3,1 2,5 219 97473_atAW124470 Tm4sf7 transmembrane 4 superfamily member 7 -1,6-1,8 -1,2 -1,3 245 160172_atY13832 Lphn1 latrophilin 1 -1,7-1,9 -1,7 -1,4 188 97680_atM65237 Mug-ps1 murinoglobulin, pseudogene 1 -2,2-3,4 -3,4 1,2 4406 160173_atAI852838 Lphn1 latrophilin 1 -2,1-2,2 -2,4 -1,3 52 102973_f_atM65238 Mug2 murinoglobulin 2 -1,8-2,8 -3,1 1,3 448 95474_atAW123850 F2r coagulation factor II (thrombin) receptor 1,0-1,2 1,3 1,7 137 101566_f_atM17818 Mup1 major urinary protein 1 1,1-1,2 -2,1 1,6 3921 104135_atAW045474 Arl3 ADP-ribosylation factor-like 3 -1,3-1,7 -1,4 -1,1 43 101910_f_atM16359 Mup1 major urinary protein 1 -1,7-3,4 -5,5 1,4 6040 92340_atY12738 Adra1b adrenergic receptor, alpha 1b -1,1-1,4 -1,4 -1,6 105 102096_f_atAI255271 Mup2 major urinary protein 2 -1,2-1,8 -4,2 2,0 1318 161189_r_atAV240013 Avpr1a arginine vasopressin receptor 1A 1,0-1,0 -1,1 2,2 12 161815_f_atAV355798 Mup2 major urinary protein 2 1,0-1,3 -2,7 1,8 1799 92532_atD49730 Avpr1a arginine vasopressin receptor 1A -1,2-1,0 -1,3 1,6 33 101909_f_atM16357 Mup3 major urinary protein 3 -1,8-3,3 -4,6 1,4 7048

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 2 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

101682_f_atM16358 Mup4 major urinary protein 4 -1,4-2,6 -3,7 1,8 2450 98956_atAI844979 Tram1 translocating chain-associating membrane -1,3-1,9 -1,2 1,4 260 101635_f_atM16360 Mup5 major urinary protein 5 -1,1-1,9 -2,5 1,6 2607 22 proteolysis & peptidolysis 161799_r_atAV350205 Kcnj9 potassium inwardly-rectifying channel, -1,7-1,2 -1,7 -1,9 89 100610_atAF058298 Capns1 calpain, small subunit 1 1,21,1 1,5 -1,1 194 96708_atAW120643 Tmed3 RIKEN cDNA 1200002G13 gene 1,0-1,2 1,1 1,5 48 104109_atAI853773 Fbxo21 F-box only protein 21 1,31,8 -1,4 1,5 75 97535_atD87661 Ywhah tyrosine 3-monooxygenase/tryptophan 1,21,4 1,9 -1,1 58 104182_atAI042655 Hgfac hepatocyte growth factor activator -1,1-1,8 -1,6 1,0 260 93070_atAI847564 Kpnb3 karyopherin (importin) beta 3 1,31,6 1,4 1,2 32 93261_atAJ000990 Lgmn legumain 1,1-1,1 2,3 1,1 39 160564_atX81627 Lcn2 lipocalin 2 1,2-1,3 4,4 -2,2 47 93025_atAI835257 Ndfip1 Nedd4 family interacting protein 1 -1,4-1,4 -1,5 1,0 550 101588_atAF058055 Slc16a1 solute carrier family 16 (monocarboxylic acid 1,31,7 1,3 -1,2 16 101985_atJ04766 Plg plasminogen -1,2-1,6 -1,9 1,1 2544 103065_atM73696 Slc20a1 solute carrier family 20, member 1 -1,11,3 2,1 1,2 54 92230_atAI642033 4930472 RIKEN cDNA 4930472G13 gene 1,52,1 1,4 -1,1 16 161149_r_atAV222871 Slc23a3 solute carrier family 23 (nucleobase -1,7-1,1 -1,5 -1,6 52 101040_atD38117 Capn2 calpain 2 1,52,0 1,6 1,1 68 102991_s_atAF100956 Slc39a7 solute carrier family 39 (zinc transporter), 1,51,1 -1,1 -1,1 147 103882_atAI182588 Cpn1 carboxypeptidase N, polypeptide 1 1,41,5 1,1 -1,1 323 103510_atM97632 Slc6a12 solute carrier family 6 (neurotransmitter 1,1-1,4 -1,7 -1,0 112 98481_atAB009459 Masp2 mannan-binding lectin serine protease 2 1,11,0 -1,5 -1,0 159 98322_atAB015800 Slc22a5 solute carrier family 22 (organic cation 1,34,8 1,3 -1,2 39 95338_s_atM82831 Mmp12 matrix metalloproteinase 12 -1,11,1 1,6 -1,0 17 95733_atAI838274 Slc29a1 solute carrier family 29 (nucleoside -1,3-1,9 -1,7 -1,0 488 95339_r_atM82831 Mmp12 matrix metalloproteinase 12 1,1-1,1 1,8 1,1 27 101877_atAI854432 Slc31a1 solute carrier family 31, member 1 -1,3-1,5 -1,8 -1,1 352 94831_atM65270 Ctsb cathepsin B 1,1-1,1 1,5 -1,2 1405 103845_atAI839005 Slc31a1 solute carrier family 31, member 1 -1,2-1,3 -1,5 -1,0 211 95608_atAI851255 Ctsb cathepsin B 1,11,3 1,9 -1,1 32 97320_atAI842734 Slc39a4 solute carrier family 39 (zinc transporter), 1,21,3 1,7 1,3 163 93810_atX68378 Ctsd cathepsin D 1,71,5 1,9 -1,2 535 104267_atAI844736 Slc23a2 solute carrier family 23 (nucleobase 1,21,7 1,1 1,1 51 104696_atAJ009840 Ctse cathepsin E 1,22,6 1,1 -1,1 16 98457_atAF020195 Slc4a4 solute carrier family 4 (anion exchanger), 1,31,8 1,2 -1,2 38 101963_atX06086 Ctsl cathepsin L -1,5-1,5 1,1 -1,0 606 96077_atX77241 Slc17a1 solute carrier family 17 (sodium phosphate), -1,3-1,7 -1,2 -1,4 21 98543_atAJ223208 Ctss cathepsin S 1,4-1,0 3,2 1,6 56 17 vesicle-mediated transport 92633_atAJ242663 Ctsz cathepsin Z 1,2-1,3 1,8 1,3 162 92220_s_atU60884 Bin1 bridging integrator 1 1,01,2 1,5 1,1 42 104036_atAI845447 Dpp7 dipeptidylpeptidase 7 1,31,6 1,8 1,3 76 95593_atAW125446 Golph2 golgi phosphoprotein 2 -1,01,9 1,2 1,0 58 161626_f_atAV028204 Plg plasminogen -1,2-1,3 -1,6 1,1 2925 99143_atAA614914 Tgoln1 trans-golgi network protein 1,41,7 1,2 -1,1 245 102284_atD16492 Masp1 mannan-binding lectin serine protease 1 -1,2-1,8 -1,7 1,4 80 99144_s_atD50031 Tgoln1 trans-golgi network protein 1,11,9 1,0 -1,2 164 59 regulation of transcription 93881_i_atD50032 Tgoln2 trans-golgi network protein 2 1,21,7 1,1 -1,3 32 98455_atU46026 Atf2 activating transcription factor 2 -1,2-1,2 -1,4 1,6 63 93882_f_atD50032 Tgoln2 trans-golgi network protein 2 1,62,2 1,3 -1,3 39 104155_f_atU19118 Atf3 activating transcription factor 3 1,11,4 1,9 1,2 23 160734_atAF084575 Ap3s1 adaptor-related protein complex 3, sigma 1 1,31,6 1,4 -1,2 74 160495_atM94623 Ahr aryl-hydrocarbon receptor -1,9-1,2 -1,2 1,3 35 93711_atD12713 Sec23a Sec23A (S. cerevisiae) -1,21,2 -1,1 -1,5 51 94466_f_atU19891 Cebpa-rs CCAAT/enhancer binding protein alpha 1,11,2 1,1 1,5 69 99847_atX73523 Siat4a sialyltransferase 4A (beta-galactoside -1,4-1,6 -1,3 -1,2 132 160502_atAF084524 Creg cellular repressor of E1A-stimulated genes -1,21,5 1,0 -1,9 352 104642_atD88577 Clecsf13 C-type (calcium dependent, carbohydrate 1,2-1,1 1,8 1,4 92 93546_s_atD14572 Cbfb core binding factor beta 1,01,3 1,4 1,6 106 98944_atAI848343 Sec23b Sec23B (S. cerevisiae) -1,1-1,5 -1,2 1,3 172 93547_atL03279 Cbfb core binding factor beta 1,11,2 1,4 1,5 258 95531_atAI854771 Amot angiomotin -1,21,3 1,5 -1,0 24 98083_atAW049031 Copeb core promoter element binding protein 1,11,5 1,8 -1,0 36 161406_atAV280754 Rnp24 Cop-coated vesicle membrane protein p24 -1,4-1,2 -1,6 -1,1 27 96023_atAI852472 D8Wsu15 DNA segment, Chr 8, Wayne State University 1,71,6 1,4 -1,3 21 160252_atAI840975 Cltc clathrin, heavy polypeptide (Hc) 1,01,4 1,5 1,0 333 94246_atJ04103 Ets2 E26 avian leukemia oncogene 2, 3' domain 1,11,3 1,8 -1,1 54 102208_atAI153959 Siat10 sialyltransferase 10 1,32,0 1,0 -1,0 17 98579_atM28845 Egr1 early growth response 1 1,31,9 1,5 2,3 40 99087_atAW060179 ZWINT ZW10 interactor (ZW interacting protein-1) 1,21,7 1,3 -1,1 273 103891_i_atAI197161 Ell2 elongation factor RNA polymerase II 2 -1,4-1,0 -1,6 1,7 101 96667_atAI851542 Vps41 vacuolar protein sorting 41 (yeast) 1,31,5 1,2 -1,0 87 98324_atX74938 Hnf-3g forkhead box A3 -1,2-1,2 1,1 2,0 56 155 Genetic Information Processing 92713_atD29015 Hnf4 hepatic nuclear factor 4 -1,4-1,4 -1,5 -1,3 233 19 DNA 100130_atX12761 Jun Jun oncogene 1,41,7 1,6 1,0 53 103460_atAI849939 Ddit4 DNA-damage-inducible transcript 4 -1,4-1,5 -1,4 -1,7 64 93753_atAI852632 Litaf LPS-induced TN factor 1,1-1,0 1,6 -1,1 80 97896_r_atAW125218 Hat1 histidine aminotransferase 1 1,11,5 1,6 1,2 16 100307_atAA002843 Lyl1 lymphoblastomic leukemia -1,3-1,5 -1,5 1,5 43 96806_atAI843802 Lpin2 lipin 2 1,82,1 1,7 -1,1 163 94289_r_atAI851574 Maged2 melanoma antigen, family D, 2 1,31,3 1,6 -1,0 49 100144_atX07699 Ncl nucleolin 1,11,6 1,5 1,0 306 96495_atU86338 Myt1l myelin transcription factor 1-like -1,3-1,2 -1,5 -1,1 140 98587_atX61449 Nap1l1 nucleosome assembly protein 1-like 1 1,42,0 1,4 -1,2 55 104712_atL00039 Myc myelocytomatosis oncogene 1,01,3 1,6 1,1 9 93987_f_atAW121568 Orc5l origin recognition complex, subunit 5-like 1,41,8 1,8 2,5 233 103654_atAB018374 Nsbp1 nucleosome binding protein 1 -1,5-1,3 -1,6 1,0 138 101065_atX57800 Pcna proliferating cell nuclear antigen -1,11,3 1,8 -1,0 80 102344_s_atAI132239 Tcea3 transcription elongation factor A (SII), 3 -1,8-1,3 -1,4 -1,3 98 103944_atU92068 Rad51l1 RAD51-like 1 (S. cerevisiae) 4,510,7 4,8 1,0 23 94821_atAW123880 Xbp1 X-box binding protein 1 -1,3-1,7 -1,4 1,9 272 99621_s_atAA690583 Sfpq splicing factor proline/glutamine rich 1,11,5 1,6 1,4 20 160617_atAW125783 Klf13 Kruppel-like factor 13 -1,6-1,6 -1,9 -1,1 113 98463_atAW124186 Smarca4 SWI/SNF related, matrix associated, actin 2,31,0 1,6 -1,2 111 160869_atAI849490 Sirt3 sirtuin 3 (silent mating type information -1,1-1,2 -1,9 -1,6 211 161077_f_atAV252495 Smarcd2 SWI/SNF related, matrix associated, actin -1,1-1,6 -1,4 -1,1 288 98038_atAF022465 Hmgb3 high mobility group box 3 1,42,5 1,9 1,3 64 99457_atX82786 Mki67 antigen identified by monoclonal antibody Ki 1,11,2 1,9 1,1 11 99158_atU58888 Sh3d3 SH3 domain protein 3 1,21,4 1,6 -1,3 110 162169_r_atAV047268 Hist1h2bp histone 1, H2bp -1,5-1,2 -1,5 -1,2 77 103553_atAA867646 Mcm10 minichromosome maintenance deficient 10 -1,1-1,3 -1,8 2,1 85 93068_r_atU62674 Hist2h2aa histone 2, H2aa1 -1,1-1,1 -1,5 -1,2 248 100062_atX62154 Mcm3 minichromosome maintenance deficient 3 (S. 1,01,5 1,7 -1,3 18 93019_atZ35401 H2afx H2A histone family, member X 1,11,2 1,7 1,1 74 160496_s_atAA929330 Mcm3 minichromosome maintenance deficient 3 (S. 1,01,7 1,8 -1,1 37 104230_atAI225445 Dclre1a DNA cross-link repair 1A, PSO2 homolog (S. -1,7-1,5 -2,0 -2,6 91 93041_atD26089 Mcm4 minichromosome maintenance deficient 4 -1,11,9 1,6 -1,1 31 93994_atAW212131 Sycp3 synaptonemal complex protein 3 1,41,2 -1,7 -2,4 343 100156_atD26090 Mcm5 minichromosome maintenance deficient 5, -1,11,8 1,7 -1,2 47 94897_atD87896 Gpx4 glutathione peroxidase 4 1,6-1,0 1,4 -1,5 247 96672_atAW123564 Hod homeobox only domain -1,01,0 -1,5 1,6 221 97421_atU42385 Smc2l1 SMC2 structural maintenance of 1,01,4 1,6 1,0 10 103671_atAF061972 Htatip2 HIV-1 tat interactive protein 2, homolog 1,31,6 1,7 -1,2 59 160182_atAI846595 Sfrs6 splicing factor, arginine/serine-rich 6 1,21,2 1,4 1,6 67 1 protein amino acid dephosphorylation 100050_atM31885 Idb1 inhibitor of DNA binding 1 1,42,6 1,9 -1,0 43 160862_atAF035645 Ptp4a3 protein tyrosine phosphatase 4a3 -1,01,5 1,2 -1,3 44 97334_atAW048812 Hes6 hairy and enhancer of split 6 (Drosophila) -1,0-1,8 -1,4 1,4 107 2 protein amino acid phosphorylation 96703_atAB029448 Maged1 melanoma antigen, family D, 1 1,21,3 1,5 1,2 187 93431_atZ38015 Dm15 dystrophia myotonica kinase, B15; -1,01,5 1,5 1,0 84 98465_f_atM31419 Ifi204 interferon activated gene 204 1,31,1 2,2 1,2 18 92310_atM96163 Snk serum-inducible kinase 1,21,5 1,6 1,0 81 94224_s_atM74123 Ifi205 interferon activated gene 205 1,11,1 2,3 1,4 17 11 protein biosynthesis 160215_atL12140 Aes amino-terminal enhancer of split -1,0-1,5 -1,4 -1,7 727 98149_s_atAW046496 1110033J RIKEN cDNA 1110033J19 gene -1,6-1,0 -1,4 -1,4 84 92440_atU73029 Irf6 interferon regulatory factor 6 -1,1-1,0 -1,3 1,6 44 95743_atAI846078 Paip2-pen RIKEN cDNA 2310050K10 gene -1,2-1,1 -1,5 1,1 72 95785_s_atY13361 Rab7 RAB7, member RAS oncogene family -1,2-1,1 1,0 -1,7 139 96016_atAW045665 Lamr1 Mus musculus cDNA clone MGC:67366 1,01,2 1,5 -1,1 164 92697_atU44752 Foxa1 forkhead box A1 -1,11,2 -1,1 1,6 28 95449_atAW049793 2310075 RIKEN cDNA 2310075G12 gene -1,3-1,4 -1,5 -1,2 343 92658_atAF010405 Foxq1 forkhead box Q1 1,0-1,2 -1,1 1,8 34 101007_atAI845732 Mknk2 MAP kinase-interacting serine/threonine 1,31,7 1,3 -1,3 209 160535_atX78709 Nfe2l1 nuclear factor, erythroid derived 2,-like 1 1,41,6 1,2 -1,1 201 102126_atX15962 Rps12 ribosomal protein S12 1,11,2 1,7 1,2 994 92562_atU70475 Nfe2l2 nuclear, factor, erythroid derived 2, like 2 1,01,7 2,1 -1,5 143 93270_atAI839918 Gars glycyl-tRNA synthetase 1,01,5 1,6 1,1 51 160859_s_atY07685 Nfib nuclear factor I/B -1,5-1,3 -1,7 -1,1 71 104144_atAW124369 Gtpbp2 GTP binding protein 2 1,11,3 1,5 1,2 97 99440_atY07686 Nfib nuclear factor I/B -1,6-1,2 -2,0 -1,2 109 93089_atX12507 Eif4a2 eukaryotic translation initiation factor 4A2 1,21,5 -1,1 1,3 197 101930_atY07688 Nfix nuclear factor I/X -1,5-1,7 -1,7 1,4 123 93859_atAI875598 Mtif2 mitochondrial translational initiation factor 2 -1,21,1 -1,7 1,1 79 102955_atU83148 Nfil3 nuclear factor, interleukin 3, regulated -1,2-1,6 -1,6 -1,1 27 94826_atY11460 Itgb4bp integrin beta 4 binding protein 1,1-1,0 1,8 -1,3 145 97123_atL76567 Nr0b2 nuclear receptor subfamily 0, group B, -1,2-1,4 -1,6 -1,2 173 16 protein catabolism 102171_r_atAI132241 Nr1i3 nuclear receptor subfamily 1, group I, -1,31,2 -2,0 -2,0 406 96890_atAW060971 1300002A RIKEN cDNA 1300002A08 gene 1,31,9 1,0 1,1 385 104507_g_atAF009328 Nr1i3 nuclear receptor subfamily 1, group I, -1,11,3 -1,9 -1,7 350 102052_atAA871791 1300003 RIKEN cDNA 1300003D03 gene 1,71,6 1,7 1,0 59 102668_atX57638 Ppara peroxisome proliferator activated receptor 1,11,5 -1,4 -1,3 300 97485_atAI850953 Pcl1-pend prenylcysteine oxidase 1 -1,01,6 1,5 -1,2 394 97926_s_atU10374 Pparg peroxisome proliferator activated receptor 1,41,0 1,5 -1,9 19 94833_atM91380 Fstl follistatin-like -1,01,1 1,6 -1,1 18 93264_atAI843895 Srebf1, sterol regulatory element binding factor 1 1,01,0 2,1 -1,1 733 100581_atU59807 Cstb cystatin B 1,11,4 2,2 -1,2 313 96192_atAF062567 Sp3 trans-acting transcription factor 3 1,42,0 1,4 1,5 31 94302_atAF013099 Psmd4 proteasome (prosome, macropain) 26S 1,31,9 1,3 -1,3 436 161595_atAV292586 Zfhx1a zinc finger homeobox 1a -1,4-1,6 -1,5 -1,1 23 98522_atAI839158 Psmd8 proteasome (prosome, macropain) 26S 1,21,5 2,1 -1,1 151 1 regulation of transcritpion 101510_atAB007136 Psme1 proteasome (prosome, macropain) 28 1,51,5 1,3 -1,1 227 101959_r_atX72310 Tfdp1 transcription factor Dp 1 1,11,7 1,5 1,0 29 94841_atAW048997 Psma5 proteasome (prosome, macropain) subunit, 1,21,5 1,1 -1,0 232 1 ribosome biogenesis 101558_s_atAB003304 Psmb5 proteasome (prosome, macropain) subunit, 1,41,5 1,4 -1,2 279 160531_atAC002397 Grcc2f gene rich cluster, C2f gene -1,3-1,7 -1,5 1,2 107 102791_atU22033 Psmb8 proteosome (prosome, macropain) subunit, -1,5-2,3 -1,3 1,3 154 8 RNA 92715_atAL078630 Ubd ubiquitin D 1,51,0 1,5 -1,0 19 97255_atAW061318 Cugbp2 CUG triplet repeat,RNA binding protein 2 1,41,5 1,2 1,1 15 161870_atAV359471 Usp15 ubiquitin specific protease 15 -1,0-1,3 -1,4 1,9 79 101634_atM33212 Npm1 nucleophosmin 1 1,01,5 1,2 1,3 227 95024_atAW047653 Usp18 ubiquitin specific protease 18 1,21,0 1,7 1,7 36 103888_atAA636171 Rbpms RNA binding protein gene with multiple 1,42,2 2,2 1,0 111 102776_atAI155095 Usp38 ubiquitin specific protease 38 1,21,5 -1,0 -1,0 63 98923_atAI852608 Rcl1 RNA cyclase homolog -1,2-1,3 -1,8 1,1 196 160247_atAW060527 Ube2v2 ubiquitin-conjugating enzyme E2 variant 2 1,21,3 1,5 -1,0 17 96041_atAB016424 Rbm3 RNA binding motif protein 3 1,61,2 1,9 -1,0 51 11 protein metabolism 93999_atAW060597 Snrpg small nuclear ribonucleoprotein polypeptide G 1,21,5 1,4 -1,0 115 97964_atAW122851 Fkbp11 FK506 binding protein 11 -1,1-1,1 1,6 1,9 67 98574_atAI835685 Prpf8 pre-mRNA processing factor 8 -1,4-1,1 -1,6 -1,2 201 92809_r_atX17069 Fkbp4 FK506 binding protein 4 -1,4-1,1 -1,6 1,0 97 103251_atAW125822 Cdadc1 RIKEN cDNA 2310010M10 gene -1,3-1,5 -1,5 1,0 58 102761_atAF041060 Grpel2 GrpE-like 2, mitochondrial 1,72,8 1,7 -1,2 50 1 RNP 94915_atX58990 Ppib peptidylprolyl isomerase B -1,2-1,5 -1,1 1,5 315 103793_atAI874858 Mvp major vault protein 1,31,4 1,7 -1,1 73 161400_f_atAV276050 Rpn1 ribophorin I -1,0-1,2 -1,1 1,6 61 97696_r_atM29015 Rpl7 ribosomal protein L7 1,31,1 1,6 1,6 1156 429 Metabolism 101966_s_atAF037206 Rnf13 ring finger protein 13 -1,3-1,1 -1,3 -1,5 353 100634_atAF056194 Rdh7 retinol dehydrogenase 7 -1,2-1,2 -1,6 1,1 1314 160257_atX60203 Fkbp1a FK506 binding protein 1a 1,31,7 2,2 1,3 114 93688_atD21826 Cmah cytidine monophospho-N-acetylneuraminic -1,4-1,7 -1,5 -1,2 39 94297_atU16959 Fkbp5 FK506 binding protein 5 -1,2-1,2 -1,3 -1,7 148 93542_atU28016 Pter phosphotriesterase related -1,5-1,3 -1,0 -1,1 123 101838_r_atAB007798 Ppm1b protein phosphatase 1B, magnesium 1,11,1 -1,2 -1,7 86 1 acetyl-CoA biosynthesis 92850_atAI836446 Rrbp1 Mus musculus, clone IMAGE:4209489, 1,21,1 1,1 1,6 116 102049_atAJ001418 Pdk4 pyruvate dehydrogenase kinase, isoenzyme 4 1,812,5 2,3 1,1 25 3 protein targeting 2 acyl CoA metabolism 160291_atAI838624 Sec61a Sec61 alpha 1 subunit (S. cerevisiae) -1,3-1,6 -1,1 1,2 289 94850_atAJ238894 Acate3-pe acyl-Coenzyme A thioesterase 3, 1,11,4 2,0 -1,2 18 160275_atAI840996 Sepr selenoprotein R -1,5-1,1 -1,6 -1,1 1831 160171_f_atAI854821 Acate2-pe RIKEN cDNA 0610041P13 gene 1,21,7 2,1 -1,1 27

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 3 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

1 amino acid activation 95066_atU67611 Taldo1 transaldolase 1 1,71,3 1,3 -1,4 327 100136_atM32017 Lamp2 lysosomal membrane glycoprotein 2 -1,3-1,3 -1,7 -1,1 315 101294_g_atZ84471 G6pd2 glucose-6-phosphate dehydrogenase 2 1,51,2 2,1 1,1 21 101964_atU05809 Tkt transketolase 1,51,3 1,5 -1,5 295 1 amino acid catabolism 95420_atAW120625 Pgd phosphogluconate dehydrogenase 1,31,4 2,0 -1,1 119 98966_atL42996 Dbt dihydrolipoamide branched chain -1,0-1,2 -1,6 1,1 75 97260_atAW121489 Arpp19-p cyclic AMP phosphoprotein -1,01,8 1,6 -1,1 36 53 amino acid metabolism 103982_s_atAJ245750 Adh4 alcohol dehydrogenase 4 (class II), pi -2,1-2,2 -2,1 -1,2 227 93625_atAA986718 Agxt alanine-glyoxylate aminotransferase -1,4-1,6 -3,7 1,3 274 103983_atAI046345 Adh4 alcohol dehydrogenase 4 (class II), pi -1,9-1,9 -2,0 -1,4 261 104286_atAA795541 Slc38a4 solute carrier family 38, member 4 -1,4-1,1 -1,5 -1,4 1991 161401_f_atAV276715 Aldh3a2 aldehyde dehydrogenase family 3, subfamily 1,52,5 1,4 -2,4 75 104007_atAA986782 Slc25a15 solute carrier family 25 (mitochondrial carrier -2,1-2,2 -2,3 -1,1 540 99559_atU14390 Aldh3a2 aldehyde dehydrogenase family 3, subfamily 2,04,1 2,1 -3,9 857 94433_atAW060684 Slc38a2 solute carrier family 38, member 2 -2,1-1,2 -2,0 1,1 51 97449_atAI835461 Aldh7a1 aldehyde dehydrogenase family 7, member -1,1-1,1 -1,5 -1,5 1293 92736_atL03290 Slc7a2 solute carrier family 7 (cationic amino acid -1,6-1,5 -1,7 -1,0 152 95084_f_atAI844995 Grhpr glyoxylate reductase/hydroxypyruvate 1,12,0 1,0 1,3 750 100323_atZ23077 Amd1 S-adenosylmethionine decarboxylase 1 -2,0-1,3 -1,1 1,0 82 100042_atAI837921 Hagh hydroxyacyl glutathione hydrolase -1,4-1,1 -1,7 -1,0 635 101489_atD12780 Amd1 S-adenosylmethionine decarboxylase 1 -1,5-1,2 1,2 1,6 98 101082_atJ02652 Mod1 malic enzyme, supernatant 3,75,8 3,7 -2,0 239 96657_atL10244 Sat spermidine/spermine N1-acetyl transferase 1 1,11,4 2,4 1,0 103 93308_s_atM97957 Pcx pyruvate carboxylase 1,91,0 -1,2 -1,5 253 96346_atAI854020 Cdo1 cysteine dioxygenase 1, cytosolic 1,1-2,6 -1,5 1,2 2018 100977_atAA691492 Pdk1 pyruvate dehydrogenase kinase, isoenzyme 1 1,31,3 -1,1 -1,7 163 103452_atAA675075 Prodh2 proline dehydrogenase (oxidase) 2 -1,3-1,1 -2,7 -1,4 453 101471_atD63764 Pklr pyruvate kinase liver and red blood cell -1,9-3,7 -1,4 -3,2 442 97385_atAJ242909 Nagk N-acetylglucosamine kinase 1,31,3 1,6 -1,0 60 101472_s_atAI195164 Pklr pyruvate kinase liver and red blood cell -1,5-2,3 -1,2 -1,9 233 99993_atU77083 Anpep alanyl (membrane) aminopeptidase -1,11,7 1,2 -1,0 98 95060_atAF058054 Slc16a7 solute carrier family 16 (monocarboxylic acid 1,51,6 1,0 -2,4 71 93015_atX65021 Gsta3 glutathione S-transferase, alpha 3 -1,4-1,4 -1,9 -1,2 900 100128_atM38724 Cdc2a cell division cycle 2 homolog A (S. pombe) -1,11,8 1,9 -1,3 39 96085_atL06047 Gsta4 glutathione S-transferase, alpha 4 -1,0-1,7 1,0 -1,3 154 100885_atAF013166 Nek2 NIMA (never in mitosis gene a)-related 1,01,4 1,5 -1,0 38 102094_f_atAI841270 Gstm1 glutathione S-transferase, mu 1 1,41,6 2,4 -1,0 782 101058_atJ00356 Amy1 amylase 1, salivary -1,3-1,9 -1,5 -1,4 307 93543_f_atJ03952 Gstm1 glutathione S-transferase, mu 1 1,41,7 3,1 -1,1 4522 103416_atAI844810 Mapk6 mitogen-activated protein kinase 6 1,51,5 2,0 1,5 76 93009_atJ04696 Gstm2 glutathione S-transferase, mu 2 1,2-1,4 4,8 -1,5 83 94481_atAI851321 Ugp2 UDP-glucose pyrophosphorylase 2 -1,7-2,1 -1,1 -1,3 167 97681_f_atJ03953 Gstm3 glutathione S-transferase, mu 3 1,52,4 7,9 -1,1 1062 4 catecholamine metabolism 97682_r_atJ03953 Gstm3 glutathione S-transferase, mu 3 1,42,3 15,6 -1,2 309 103597_atX63349 Dct dopachrome tautomerase -1,3-2,0 -2,5 -1,4 146 96258_atAI843448 Mgst3 microsomal glutathione S-transferase 3 6,83,4 2,6 -2,1 98 98920_g_atAA867773 Blp1-pend RIKEN cDNA 2410018G23 gene -1,3-1,2 -1,5 -1,3 90 95603_atAW123955 Gldc glycine decarboxylase -1,1-1,6 -1,7 -1,2 393 103421_atD50464 Sdfr2 stromal cell derived factor receptor 2 -1,21,3 1,6 1,4 32 104086_atAI787269 Dmgdh dimethylglycine dehydrogenase precursor -1,4-1,9 -2,2 -1,3 913 93749_atAI848045 Maoa monoamine oxidase A 1,01,3 1,7 -1,4 58 96828_atD89664 Gnmt glycine N-methyltransferase -1,5-3,7 -3,3 -1,5 5650 12 cofactors & vitamins 101844_atU94700 Pipox pipecolic acid oxidase -1,4-2,4 -2,4 -1,2 422 96763_atAI839995 Sardh sarcosine dehydrogenase -1,5-3,4 -3,6 -1,6 465 94241_atAI837229 Coasy-pe Coenzyme A synthase 1,91,6 1,3 -1,1 130 92833_atL07645 Hal histidine ammonia lyase -1,3-3,5 -6,9 -1,1 638 93736_atAF090686 Tcn2 transcobalamin 2 1,1-1,1 1,1 -1,7 316 97424_atAW049647 Arl6ip5 ADP-ribosylation factor-like 6 interacting 1,1-1,1 1,8 -1,2 95 102313_atL09737 Gch GTP cyclohydrolase 1 -1,4-1,7 -1,2 -1,0 174 96295_atAW122030 Psat1 phosphoserine aminotransferase 1 1,11,8 1,5 1,2 15 160850_atU32197 Fpgs folylpolyglutamyl synthetase -1,01,1 -1,8 -1,0 407 96243_f_atAW120804 Aldh9a1 aldehyde dehydrogenase 9, subfamily A1 1,11,9 -1,2 -1,1 333 99009_atZ49204 Nnt nicotinamide nucleotide transhydrogenase 1,01,7 -1,2 -1,3 163 103389_atAJ224761 Aass aminoadipate-semialdehyde synthase -1,2-1,5 -1,9 -1,3 478 101473_atU86108 Nnmt nicotinamide N-methyltransferase -2,3-7,0 -4,7 -1,7 1084 94049_atAF033381 Bhmt betaine-homocysteine methyltransferase -1,1-1,7 -1,9 -1,0 5563 102013_atAF030513 Rdh6 retinol dehydrogenase 6 2,72,1 1,9 -3,1 154 97418_atAA986608 Bhmt2 betaine-homocysteine methyltransferase 2 -1,3-1,6 -1,7 -1,0 507 96678_atAW260761 Dhrs4 dehydrogenase/reductase (SDR family) 1,12,3 1,0 -1,7 427 96024_atL32836 Ahcy S-adenosylhomocysteine hydrolase -1,2-1,9 -2,1 -1,3 4843 95620_atAW120882 Dhrs7 dehydrogenase/reductase (SDR family) 1,01,1 -1,2 -1,6 172 96025_g_atL32836 Ahcy S-adenosylhomocysteine hydrolase -1,2-1,6 -2,0 -1,1 3119 98320_atY12657 Cyp26a1 cytochrome P450, family 26, subfamily a, -1,8-3,6 -4,3 -3,2 168 96026_atAA987153 Ahcy S-adenosylhomocysteine hydrolase -1,4-2,8 -3,3 -1,4 700 102649_s_atD64162 Raet1c retinoic acid early transcript gamma 1,32,0 -2,1 -5,0 641 98459_atAA913994 Shmt1 serine hydroxymethyl transferase 1 (soluble) -1,3-1,4 -1,6 -1,3 517 104716_atX60367 Rbp1 retinol binding protein 1, cellular 1,41,3 2,4 2,2 38 100620_atD29987 Hpd 4-hydroxyphenylpyruvic acid dioxygenase -1,4-1,5 -1,6 -1,3 148 18 detoxification 160350_atAW060750 Gstz1 glutathione transferase zeta 1 1,0-1,0 -1,5 -1,1 2066 101991_atD16215 Fmo1 flavin containing monooxygenase 1 -1,1-1,1 -1,4 -1,9 549 95407_atX51942 Pah phenylalanine hydroxylase -2,1-2,6 -2,8 1,2 1345 103087_atL02331 Sult1a1 sulfotransferase family 1A, phenol-preferring, -1,3-3,2 -2,0 -1,6 182 96326_atAI255353 Tat tyrosine aminotransferase -1,4-2,1 -2,4 -1,0 2325 93434_atAF026074 Sultx1-pe sulfotransferase related gene X1 -1,6-1,5 -1,8 1,9 32 104539_atAF022894 Sult1b1 sulfotransferase family 1B, member 1 -1,6-2,3 -2,4 -1,6 169 99941_atL11333 Es31 esterase 31 -1,2-2,0 -4,1 2,0 1982 92492_atAB020203 Ak3l adenylate kinase 3 alpha-like -1,1-1,5 -2,3 -1,9 109 100630_f_atAA241764 Gstm5 glutathione S-transferase, mu 5 -1,1-1,1 -1,2 1,7 335 93557_atU43285 Sps2 selenophosphate synthetase 2 1,2-1,8 -1,6 1,2 372 99583_atX53451 Gstp2 glutathione S-transferase, pi 2 -1,4-1,0 1,4 4,0 1853 92589_atAI846545 Psph phosphoserine phosphatase 1,21,2 1,6 -1,0 107 95019_atX98055 Gstt1 glutathione S-transferase, theta 1 1,81,1 -1,3 -1,3 750 99184_atAW120896 Csad cysteine sulfinic acid decarboxylase 2,43,6 3,8 1,3 134 104603_atX98056 Gstt2 glutathione S-transferase, theta 2 2,03,3 1,7 -1,2 293 93827_atU24493 Tdo2 tryptophan 2,3-dioxygenase -1,71,1 -3,0 1,2 518 101872_atJ03958 Gsta2 glutathione S-transferase, alpha 2 (Yc2) -1,7-2,0 -1,3 -2,9 769 99269_g_atAI194855 Tdo2 tryptophan 2,3-dioxygenase -1,51,1 -2,9 -1,3 2720 96670_atAI841295 Gstk1 glutathione S-transferase kappa 1 1,21,4 -1,5 -1,5 642 98582_atU58988 Hgd homogentisate 1, 2-dioxygenase -1,3-2,1 -1,5 -1,5 1399 93780_atAW060827 Them2 thioesterase superfamily member 2 1,91,6 1,3 -1,4 130 104566_atAI845009 Asl argininosuccinate lyase -2,3-4,7 -3,1 -2,1 226 100596_atM32032 Selenbp1 selenium binding protein 1 1,2-1,1 -1,4 3,0 828 92848_atX64837 Oat ornithine aminotransferase -1,5-5,4 -2,6 -2,6 2268 162030_r_atAV366872 Selenbp1 selenium binding protein 1 -1,5-1,0 -1,5 -1,7 47 101408_atAF010499 Gamt guanidinoacetate methyltransferase -1,3-1,7 -1,4 -1,2 466 101587_atU89491 Ephx1 epoxide hydrolase 1, microsomal -1,71,5 1,3 -2,4 1430 94414_atX07092 Otc ornithine transcarbamylase -1,1-2,9 -2,4 1,2 1098 93051_atZ37107 Ephx2 epoxide hydrolase 2, cytoplasmic 1,92,6 1,0 -1,3 279 104153_atAW047743 Ivd isovaleryl coenzyme A dehydrogenase 1,31,1 -1,5 1,1 153 93021_atAA790008 Drpla dentatorubral pallidoluysian atrophy 1,1-1,5 -1,5 -2,7 53 73 carbohydrate metabolism 99854_atAF026075 Sult3a1 sulfotransferase related gene X2 1,9-7,4 -3,3 -6,7 141 102807_atAW048054 9230112 RIKEN cDNA 9230112O05 gene 1,11,7 1,5 -1,4 145 160088_atU90535 Fmo5 flavin containing monooxygenase 5 -1,4-1,1 1,1 -1,8 906 100565_atAW123396 Gnpda1 glucosamine-6-phosphate deaminase 1 -1,6-1,6 -1,4 -1,5 64 31 energy metabolism 97924_atAJ132236 Uae1 UDP-N-acetylglucosamine-2-epimerase/N-ac -1,2-1,3 -1,6 1,1 366 95660_atAI851815 Ethe1 ethylmalonic encephalopathy 1 -1,7-2,6 -1,9 -1,6 634 96115_atU28168 Dp1 deleted in polyposis 1 1,51,5 2,1 1,2 107 160637_atAW060325 Mocs2 molybdenum cofactor synthesis 2 -1,6-1,4 -1,8 -1,5 328 103781_atU76832 Stx4a syntaxin 4A (placental) 1,51,4 1,5 1,5 70 160537_atAF026073 Sultn N-sulfotransferase -1,6-1,7 -1,9 1,1 114 96870_atAI836740 Aco2 aconitase 2, mitochondrial 1,51,9 1,4 -1,2 758 94797_atAI120514 Slc26a1 solute carrier family 26 (sulfate transporter), -1,8-2,0 -1,5 -1,2 109 160207_atAW121639 Acly ATP citrate lyase 1,4-1,0 1,6 1,0 443 94540_atAI595772 Cyp2d26 cytochrome P450, family 2, subfamily d, -1,2-1,2 -2,2 -1,3 2863 99666_atAW125431 Cs citrate synthase 1,51,8 2,0 1,1 147 100069_atM77497 Cyp2f2 cytochrome P450, family 2, subfamily f, -1,3-1,2 -2,1 -1,1 1341 160289_s_atAI849904 Dlst dihydrolipoamide S-succinyltransferase (E2 1,31,5 -1,1 -1,7 249 92814_atAI114881 Cyp2j5 cytochrome P450, family 2, subfamily j, -1,31,1 -1,8 1,2 28 99148_atAI852862 Fh1 fumarate hydratase 1 -1,2-1,1 -1,4 -1,8 918 100892_atAI850186 Ndufaf1 NADH dehydrogenase (ubiquinone) 1 alpha -1,1-1,1 -1,6 -1,1 92 160571_atAF020039 Idh1 isocitrate dehydrogenase 1 (NADP+), soluble 1,11,5 -1,1 -1,0 375 92807_atX77585 Txn1 thioredoxin 1 (ATL-derived factor) (ADF) 1,21,3 1,6 1,2 898 95693_atU51167 Idh2 isocitrate dehydrogenase 2 (NADP+), 1,51,4 1,4 -1,6 199 94207_atAI842377 Txndc7 thioredoxin domain containing 7 -1,2-1,6 -1,1 1,8 79 93991_atX07295 Mor1 malate dehydrogenase, mitochondrial 1,31,6 1,3 -1,1 694 94209_g_atAW045202 Txndc7 thioredoxin domain containing 7 -1,3-1,8 -1,2 1,2 220 96268_atAI840979 Suclg1 succinate-CoA ligase, GDP-forming, alpha 1,41,7 1,0 -1,1 375 97869_atAI844043 Etfdh electron transferring flavoprotein, 1,12,0 -1,2 -1,7 216 93501_f_atAF058955 Sucla2 succinate-Coenzyme A ligase, ADP-forming, -1,1-1,1 -1,5 1,1 338 96909_atAI849803 Ndufab1 NADH dehydrogenase (ubiquinone) 1, 1,22,0 -1,0 -1,2 256 102092_s_atC86340 Ovgp1 oviductal glycoprotein 1 precursor -1,2-1,2 -1,3 -1,5 22 96878_atAW048566 Cyb5m RIKEN cDNA 1810044O22 gene 1,61,4 1,2 -1,0 130 93360_atAF007267 Pmm1 phosphomannomutase 1 1,73,3 1,3 -1,1 51 100059_atM31775 Cyba cytochrome b-245, alpha polypeptide 1,1-1,0 1,6 1,1 85 94375_atY11666 Hk2 hexokinase 2 1,51,1 1,4 1,1 36 93500_atM63245 Alas1 aminolevulinic acid synthase 1 -1,51,4 -1,1 -2,6 1844 100573_f_atM14220 Gpi1 glucose phosphate isomerase 1 1,51,1 1,9 -1,5 107 92642_atM25944 Car2 carbonic anhydrase 2 2,12,5 1,5 -1,4 56 161524_r_atAV295044 Gpi1 glucose phosphate isomerase 1 -1,0-1,1 -1,1 1,7 24 160375_atAJ006474 Car3 carbonic anhydrase 3 1,3-1,7 -3,1 1,1 2338 97820_atAB027012 Galk1 galactokinase 1 2,51,6 2,0 -1,6 99 98137_atX51971 Car5a carbonic anhydrase 5a, mitochondrial -1,2-2,8 -2,3 -1,3 303 96918_atAI790931 Fbp1 fructose bisphosphatase 1 -1,7-2,2 -2,5 -1,6 3975 102773_atX61397 Car8 carbonic anhydrase 8 -1,6-1,5 -1,8 1,1 51 97379_atD42083 Fbp2 fructose bisphosphatase 2 -1,22,4 1,0 -1,2 29 160135_atAW122914 D16Ertd5 DNA segment, Chr 16, ERATO Doi 502, -1,4-1,4 -1,9 -1,2 241 96789_i_atAW125222 Galm galactose mutarotase (aldose 1-epimerase) -1,3-1,3 -1,7 -1,7 282 104165_atAJ132098 Vnn1 vanin 1 3,16,0 1,5 -4,7 202 103333_atU00445 G6pc glucose-6-phosphatase, catalytic -2,7-2,9 -3,4 -2,9 2329 104181_atAJ132103 Vnn3 vanin 3 1,91,6 1,3 -1,9 128 97430_atAF080469 G6pt1 glucose-6-phosphatase, transport protein 1 -1,9-3,8 -2,7 -1,6 901 98098_atM32452 Car1 carbonic anhydrase 1 -1,5-1,4 -1,9 -1,2 66 160481_atAF009605 Pck1 phosphoenolpyruvate carboxykinase 1, -2,1-2,2 -7,8 -1,2 1009 98079_atAB005450 Car14 carbonic anhydrase 14 -1,4-2,7 -3,0 -1,6 233 100574_f_atL09104 Gpi1 glucose phosphate isomerase 1 1,21,0 1,6 -1,0 481 160124_r_atU13839 Atp6v1c1 ATPase, H+ transporting, V1 subunit C, 1,41,4 1,6 1,5 36 103357_atX15684 Slc2a2 solute carrier family 2 (facilitated glucose -1,3-1,9 -1,7 -1,3 850 96610_atAW046442 Atp6v1h ATPase, H+ transporting, lysosomal 1,51,2 1,4 1,0 160 98942_r_atAW125284 2310032 RIKEN cDNA 2310032D16 gene -1,11,1 -1,4 1,9 13 94301_atAI843269 Atp6v0e ATPase, H+ transporting, V0 subunit 1,01,0 1,5 -1,3 177 162262_f_atAV357306 Gyg1 glycogenin 1 1,11,6 1,4 -1,0 20 97402_atM88694 Temt thioether S-methyltransferase 1,11,0 1,5 -1,9 2389 103297_atX98848 Pfkfb1 6-phosphofructo-2-kinase/fructose-2,6-biphos 1,71,1 1,0 -1,2 89 93084_atU27315 Slc25a4 solute carrier family 25 (mitochondrial carrier 1,76,5 3,9 1,8 30 93560_atAI845882 Acyp1 acylphosphatase 1, erythrocyte (common) -1,3-1,2 -1,5 -1,2 127 99112_atAA683883 Slc25a10 solute carrier family 25 (mitochondrial carrier 2,11,6 1,2 -1,9 556 104258_atAA881576 Acyp2 acylphosphatase 2, muscle type 2,42,3 1,9 -1,1 24 1 glutathione metabolism 160090_f_atY00516 Aldo1 aldolase 1, A isoform 1,41,6 1,8 1,2 176 160646_atAI851983 Gsr glutathione reductase 1 1,21,3 1,7 -1,0 167 161889_f_atAV102160 Aldo1 aldolase 1, A isoform 1,31,3 1,6 1,3 77 1 heme biosynthesis 101532_g_atAI527354 Aldo2 aldolase 2, B isoform -1,4-1,8 -1,2 -1,6 2117 92768_s_atM15268 Alas2 aminolevulinic acid synthase 2, erythroid -1,6-2,0 -1,4 1,2 160 160546_atAW121134 Aldo3 aldolase 3, C isoform -1,2-1,1 -1,5 -1,2 77 2 heme metabolism 96746_atAW124813 Dlat dihydrolipoamide S-acetyltransferase (E2 1,61,6 1,9 -1,0 58 160568_atAI841389 Eno1 enolase 1, alpha non-neuron 1,0-1,3 1,0 -1,8 951 103085_atAB013095 Hebp1 heme binding protein 1 -1,1-1,8 -1,4 -1,3 604 102651_atL41631 Gck glucokinase 1,41,2 1,8 1,4 200 160573_atAA645537 Hccs holocytochrome c synthetase 1,11,5 1,2 1,3 37 101214_f_atM32599 Gapd glyceraldehyde-3-phosphate dehydrogenase -1,1-1,5 1,1 -1,7 3365 1 hemoglobin AFFX-GapdhM32599 Gapd glyceraldehyde-3-phosphate dehydrogenase 1,31,1 1,6 1,3 181 162457_f_atAV003378 Hba-a1 hemoglobin alpha, adult chain 1 1,11,2 2,2 1,4 241 96066_s_atX97047 Pkm2 pyruvate kinase, muscle 1,11,0 1,9 1,0 66 1 lipid catabolism 101753_s_atX51547 Lzp-s Lysozyme C, type P precursor (EC 3.2.1.17) 1,01,1 3,6 1,3 39 97965_atU88624 Pla2g6 phospholipase A2, group VI 1,72,8 1,5 1,0 114 94840_atU05837 Hexa hexosaminidase A 1,41,2 1,7 -1,3 131 20 lipid metabolism 102322_atAF061017 Ugdh UDP-glucose dehydrogenase 1,52,2 1,9 1,0 339 102381_atAA619207 Acsl4 acyl-CoA synthetase long-chain family 1,72,3 2,4 1,9 50 102826_atJ05663 Akr1b7 aldo-keto reductase family 1, member B7 -1,01,0 2,0 1,1 28 94354_atAI845514 Abca1 ATP-binding cassette, sub-family A (ABC1), 1,31,3 1,9 1,8 125 103322_atL35034 Rpia ribose 5-phosphate isomerase A 1,31,4 1,8 1,0 96 104630_atAA914114 BC02161 cDNA sequence BC021611 1,61,7 1,2 -1,2 65 94966_atZ11911 G6pdx glucose-6-phosphate dehydrogenase X-linked 1,31,1 1,9 1,0 27 95043_atAI047331 Cyp2c70 cytochrome P450, family 2, subfamily c, -1,3-2,5 1,0 4,3 316

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 4 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

101307_atY10221 Cyp4a12 cytochrome P450, family 4, subfamily a, -1,01,3 -1,2 -2,1 2749 160544_atAJ223066 Fabp5 fatty acid binding protein 5, epidermal 1,61,5 2,0 1,4 72 101103_atY11638 Cyp4a14 cytochrome P450, family 4, subfamily a, 1,52,9 1,3 -9,2 1418 98967_atU04827 Fabp7 fatty acid binding protein 7, brain 1,3-1,0 2,2 1,0 22 104129_atAI462001 Cyp4f13 cytochrome P450, family 4, subfamily f, -1,5-1,6 -1,9 -1,1 182 93486_atU15976 Slc27a1 solute carrier family 27 (fatty acid transporter), 1,22,9 1,1 -1,0 94 100996_atAF015811 AU04170 expressed sequence AU041707 1,51,2 -1,0 -1,3 32 100967_atAF072757 Slc27a2 solute carrier family 27 (fatty acid transporter), 1,41,6 -1,1 -1,0 659 104325_atAI461631 Falp Fat cell-specific low molecular weight protein 2,12,1 1,7 -1,6 198 97957_atAF072759 Slc27a4 solute carrier family 27 (fatty acid transporter), 1,72,0 1,4 -1,1 46 99592_f_atAB030505 Rdh11 retinol dehydrogenase 11 1,31,5 1,0 -1,0 402 96231_atAW123780 Bphl Valacyclovir hydrolase precursor (Biphenyl -1,3-1,0 -2,3 -1,4 366 103465_f_atU60438 Saa2 serum amyloid A 2 1,1-1,5 7,3 3,6 177 98984_f_atD50430 Gpd2 glycerol phosphate dehydrogenase 2, 1,62,4 1,9 -1,2 169 100333_atM13521 Saa2 serum amyloid A 2 1,11,0 2,6 2,1 87 92796_atJ02980 Akp2 alkaline phosphatase 2, liver 1,01,6 1,4 -1,5 101 102712_atX03505 Saa3 serum amyloid A 3 -1,0-1,1 1,7 2,0 32 102026_s_atAB011000 Chkl choline kinase-like 1,42,3 1,2 1,0 32 92242_atU65403 Saa4 serum amyloid A 4 -2,1-2,6 -3,3 1,6 227 102027_s_atAA204010 Chkl choline kinase-like 1,52,2 1,3 -1,0 311 104072_atM23552 Apcs serum amyloid P-component 2,71,2 1,7 1,8 172 104371_atAF078752 Dgat1 diacylglycerol O-acyltransferase 1 1,61,5 1,1 -1,1 82 103386_atAW046123 Pte1 peroxisomal acyl-CoA thioesterase 1 1,31,8 1,2 -1,1 76 97525_atU48403 Gyk glycerol kinase -1,11,9 1,1 1,2 243 103646_atX85983 Crat carnitine acetyltransferase 3,06,6 2,8 -1,6 106 161753_f_atAV290060 Gpd1 glycerol-3-phosphate dehydrogenase 1 1,71,1 -1,1 -1,1 40 161989_f_atAV238359 Crat carnitine acetyltransferase 1,73,5 1,7 -1,2 38 92592_atM25558 Gpd1 glycerol-3-phosphate dehydrogenase 1 2,11,3 -1,1 -1,3 541 95695_atAB017112 Slc25a20 solute carrier family 25 (mitochondrial 1,62,7 1,2 -1,8 418 97511_atAI846600 Mgll monoglyceride lipase 1,53,5 1,2 -1,2 155 100931_atX73230 Arsa arylsulfatase A 1,71,8 1,6 -1,3 110 101867_atU11680 Gpam glycerol-3-phosphate acyltransferase, 3,22,5 3,1 -1,3 701 101891_atY09517 Hsd17b2 hydroxysteroid (17-beta) dehydrogenase 2 1,1-1,3 -1,6 1,9 142 104343_f_atAI845798 Pla2g12a phospholipase A2, group XII 1,81,6 2,4 1,7 90 97515_atX89998 Hsd17b4 hydroxysteroid (17-beta) dehydrogenase 4 1,52,3 1,4 -1,6 936 98596_s_atY15003 Siat9 sialyltransferase 9 1,22,0 1,3 -1,4 404 102123_atZ31689 Lip1 lysosomal acid lipase 1 1,41,7 1,6 -1,3 95 161970_f_atAV371169 Hmgcl 3-hydroxy-3-methylglutaryl-Coenzyme A lyase 1,51,8 1,2 -1,4 123 104273_atU95215 Baat bile acid-Coenzyme A: amino acid -1,3-1,7 -1,7 -1,1 375 94324_f_atU49878 Hmgcl 3-hydroxy-3-methylglutaryl-Coenzyme A lyase 1,72,2 1,2 -1,6 484 95283_atAI173996 Abcc2 ATP-binding cassette, sub-family C 1,11,6 1,9 -1,1 157 92590_atU12791 Hmgcs2 3-hydroxy-3-methylglutaryl-Coenzyme A -1,31,2 -2,4 -2,4 395 103689_atAA833514 Abcc3 ATP-binding cassette, sub-family C 1,91,3 1,2 -2,0 388 93066_atD16195 Grn granulin -1,01,5 1,7 -1,6 300 93407_atAB028737 Abcc6 ATP-binding cassette, sub-family C -1,3-1,5 -1,6 1,1 70 102839_atD78354 Plscr1 phospholipid scramblase 1 1,41,6 2,0 1,2 41 99404_atL23754 Cyp7a1 cytochrome P450, family 7, subfamily a, -3,0-1,9 -1,3 -1,5 119 98589_atM93275 Adrp adipose differentiation related protein 2,34,1 2,8 -1,6 812 161345_f_atAV141027 Cyp7b1 cytochrome P450, family 7, subfamily b, -2,1-2,0 -2,6 12,8 65 102763_atAF064748 S3-12 plasma membrane associated protein, S3-12 3,91,4 1,4 -1,2 30 92898_atU36993 Cyp7b1 cytochrome P450, family 7, subfamily b, -2,6-2,7 -3,6 12,8 108 104448_atL47970 Mttp microsomal triglyceride transfer protein 1,11,6 1,1 -1,2 56 103284_atAF090317 Cyp8b1 cytochrome P450, family 8, subfamily b, -1,8-2,9 -5,4 -1,5 1063 101173_atZ50024 Pctp phosphatidylcholine transfer protein 1,92,8 1,2 -2,6 79 161668_f_atAV140072 Por P450 (cytochrome) oxidoreductase -1,11,1 1,0 -1,5 145 100927_atU28960 Pltp phospholipid transfer protein 8,27,6 4,6 -1,7 128 99019_atD17571 Por P450 (cytochrome) oxidoreductase -1,11,4 1,2 -2,1 485 102696_s_atAI747899 Pitpnb phosphotidylinositol transfer protein, beta -1,6-1,3 -1,6 -1,6 35 100339_atU95131 Slc10a1 solute carrier family 10 (sodium/bile acid -1,7-2,5 -4,0 -1,4 1965 92612_atAJ223959 Slc27a5 solute carrier family 27 (fatty acid transporter), -1,4-2,1 -2,2 -1,3 1444 100341_g_atU95132 Slc10a1 solute carrier family 10 (sodium/bile acid -1,8-2,5 -4,2 -1,7 1414 100095_atU37799 Scarb1 scavenger receptor class B, member 1 -1,4-1,8 -1,5 1,3 302 94325_atAW124932 Hmgcs1 3-hydroxy-3-methylglutaryl-Coenzyme A 1,33,0 -2,0 -1,1 230 96094_atU79573 Apoa1 apolipoprotein A-I 1,41,6 1,9 1,9 5607 102416_atM64863 Cyp17a1 cytochrome P450, family 17, subfamily a, -1,1-1,4 -1,7 -2,2 376 100078_atM64248 Apoa4 apolipoprotein A-IV -2,0-2,5 7,0 -3,8 583 101659_atM75886 Hsd3b2 hydroxysteroid dehydrogenase-2, -5,5-5,6 -7,3 5,4 115 95727_atAI785422 Apoa5 apolipoprotein A-V -2,0-2,1 -2,2 -1,4 2470 98401_atM77015 Hsd3b3 hydroxysteroid dehydrogenase-3, -1,3-1,4 -1,6 1,0 84 95728_g_atAI785422 Apoa5 apolipoprotein A-V -2,0-2,1 -1,9 -1,3 2444 94795_atL41519 Hsd3b5 hydroxysteroid dehydrogenase-5, -1,4-1,3 -1,7 44,5 23 95729_atAA674450 Apoa5 apolipoprotein A-V -1,8-2,2 -1,8 -1,4 964 160104_atAA824102 Hsd3b7 hydroxy-delta-5-steroid dehydrogenase, 3 -1,0-1,0 -1,4 1,5 654 97887_atZ22216 Apoc2 apolipoprotein C-II 1,5-1,3 1,9 -1,3 984 102729_f_atAF031170 Hsd3b6 hydroxysteroid dehydrogenase-6, -1,2-1,6 -1,5 1,3 102 96074_atAI528149 Apof apolipoprotein F -2,1-2,2 -1,6 1,1 956 95453_f_atAF087687 S100a1 S100 calcium binding protein A1 1,6-1,2 -1,0 -1,4 354 94318_atY11356 Apoh apolipoprotein H -1,3-1,6 -1,8 -1,3 4081 94284_atAW122731 Dia1 diaphorase 1 (NADH) 1,51,1 1,4 1,0 640 93840_atAA655303 Apom apolipoprotein M -1,5-1,7 -1,8 2,3 160 160344_atAB021289 Npc2 Niemann Pick type C2 1,31,2 1,7 1,0 264 100332_s_atAF093853 Prdx6 peroxiredoxin 6 -1,4-1,5 -1,6 -1,6 555 97132_atAA189890 Soat2 sterol O-acyltransferase 2 1,21,5 1,5 1,1 155 102820_atM60358 Cyp2b13 cytochrome P450, family 2, subfamily b, 2,93,0 -1,2 -2,1 78 94177_atY15733 Hsd17b7 hydroxysteroid (17-beta) dehydrogenase 7 -1,11,7 -1,5 -1,3 47 103023_atJ05154 Lcat lecithin cholesterol acyltransferase 1,0-1,3 -1,8 -1,4 741 96895_atU32684 Pon1 paraoxonase 1 -1,4-2,1 -2,9 -1,5 1739 161703_f_atAV003419 Anxa1 annexin A1 1,21,3 2,1 1,1 14 98600_atU41341 S100a11 S100 calcium binding protein A11 (calizzarin) 2,32,4 7,7 -1,2 29 93038_f_atM69260 Anxa1 annexin A1 1,31,1 2,3 -1,0 21 93042_atD21207 Bzrp benzodiazepine receptor, peripheral 1,71,4 1,5 1,1 591 100569_atM14044 Anxa2 annexin A2 4,44,4 9,1 -1,4 112 97248_atX61431 Dbi diazepam binding inhibitor 1,72,2 1,6 -1,3 1771 101393_atAJ001633 Anxa3 annexin A3 1,01,1 1,7 1,0 16 101923_atU34277 Pla2g7 phospholipase A2, group VII 1,611,3 1,9 1,1 20 93083_atD63423 Anxa5 annexin A5 1,91,4 2,9 -2,0 518 102370_atAA822174 Dhrs8 retinal short-chain dehydrogenase/reductase 1,96,5 1,9 -1,2 409 92539_atM16465 S100a10 S100 calcium binding protein A10 (calpactin) 1,31,1 2,0 -1,8 495 96035_atL47335 Bckdha branched chain ketoacid dehydrogenase E1, 1,51,1 -1,0 -1,2 311 160373_i_atAI839175 Sdpr serum deprivation response 2,61,6 1,5 -1,0 15 160083_atM63335 Lpl lipoprotein lipase 2,611,1 7,2 1,9 18 104627_atAW121668 Cds2 CDP-diacylglycerol synthase (phosphatidate -1,01,1 1,6 -1,2 55 95611_atAA726364 Lpl lipoprotein lipase 3,817,1 11,2 2,3 31 160577_atAF011336 Atp9a ATPase, class II, type 9A 1,41,6 1,3 -1,1 214 161779_r_atAV360058 Cryl1 crystallin, lamda 1 -1,51,0 -1,5 -1,4 56 101944_atU89352 Lypla1 lysophospholipase 1 -1,2-1,1 -1,5 -1,7 645 101006_atM35797 Acat2 acetyl-Coenzyme A acetyltransferase 2 1,51,8 1,1 -1,1 64 101945_g_atU89352 Lypla1 lysophospholipase 1 -1,3-1,1 -1,5 -1,6 47 95425_atU21489 Acadl acetyl-Coenzyme A dehydrogenase, 1,42,5 1,3 -1,2 57 101946_atAA840463 Lypla1 lysophospholipase 1 -1,5-1,2 -2,1 -1,8 229 92913_atZ48670 Abcd2 ATP-binding cassette, sub-family D (ALD), 1,62,6 1,1 -1,3 17 160807_atAW124201 Agpat3 1-acylglycerol-3-phosphate O-acyltransferase 1,21,6 1,2 -1,4 153 95646_atU01170 Cpt2 carnitine palmitoyltransferase 2 -1,12,0 -1,0 -1,3 849 96110_atU31966 Cbr1 carbonyl reductase 1 1,01,0 1,6 -1,8 145 102998_atX04283 Cyp1a2 cytochrome P450, family 1, subfamily a, -1,9-1,6 -1,2 2,5 1070 93585_atD17674 Cyp2c29 cytochrome P450, family 2, subfamily c, -1,5-1,7 1,1 -1,2 103 102847_s_atM19319 Cyp2a4 cytochrome P450, family 2, subfamily a, -2,0-1,6 1,6 -1,1 2082 99083_atAF047542 Cyp2c37 cytochrome P450, family 2. subfamily c, -2,9? -4,0 -1,4 3481 102701_atM21856 Cyp2b10 cytochrome P450, family 2, subfamily b, 1,3-1,1 1,1 -2,6 125 102084_f_atAF047725 Cyp2c38 cytochrome P450, family 2, subfamily c, -1,3-2,1 -1,5 -1,8 1226 101862_atM21855 Cyp2b9 cytochrome P450, family 2, subfamily b, 1,62,2 -1,7 -12,5 693 96334_f_atAF047727 Cyp2c40 cytochrome P450, family 2, subfamily c, -1,9-3,0 -2,3 -1,3 2046 101287_s_atM24264 Cyp2d10 cytochrome P450, family 2, subfamily d, -1,7-2,1 -2,4 -1,3 3909 93351_atU44389 Hpgd hydroxyprostaglandin dehydrogenase 15 -1,6-1,1 -2,1 -2,1 227 98612_atM24263 Cyp2d10 cytochrome P450, family 2, subfamily d, -1,8-2,5 -2,4 -1,0 2429 103660_atAF093669 Pex11a peroxisomal biogenesis factor 11a 2,75,5 2,2 -1,7 386 162174_atAV003830 Cyp2d9 cytochrome P450, family 2, subfamily d, -1,6-1,7 -1,8 1,1 249 97114_atU57999 Psap prosaposin 1,51,5 1,5 -1,1 454 94539_f_atM27168 Cyp2d9 cytochrome P450, family 2, subfamily d, -1,9-2,3 -2,8 -1,0 3846 100893_atU27455 Sptlc2 serine palmitoyltransferase, long chain base 1,41,5 2,0 1,1 64 99463_atX63023 Cyp3a13 cytochrome P450, family 3, subfamily a, -1,5-1,0 -1,6 -1,3 356 96623_atAI853172 Ugcg UDP-glucose ceramide glucosyltransferase 1,41,3 1,9 1,6 17 100539_atAI841279 Bach Cytosolic acyl CoA thioester hydrolase (Long 1,21,6 1,4 -1,1 58 92556_atD45850 Akr1c6 aldo-keto reductase family 1, member C6 -1,7-2,3 -1,7 1,4 3513 93754_atAF030343 Ech1 enoyl-Coenzyme A hydratase 1, peroxisomal 1,42,7 1,2 -1,8 1147 94276_atAF064635 Hsd17b12 hydroxysteroid (17-beta) dehydrogenase 12 2,42,8 1,4 1,2 262 94507_atU15977 Facl2 fatty acid Coenzyme A ligase, long chain 2 1,63,0 -1,3 -1,2 378 92437_atAW047445 Tm7sf2 transmembrane 7 superfamily member 2 1,0-1,0 -1,6 -1,3 311 104017_atAB033887 Facl4 fatty acid-Coenzyme A ligase, long chain 4 1,31,7 2,0 1,1 15 97867_atX83202 Hsd11b1 hydroxysteroid 11-beta dehydrogenase 1 -1,8-5,9 -4,0 -1,6 1624 103649_atAI648067 Hao3 hydroxyacid oxidase (glycolate oxidase) 3 -1,05,3 -1,2 -1,5 12 93316_atAB017026 Osbpl1a oxysterol binding protein-like 1A -1,11,0 -1,4 1,5 95 94485_atAI840013 Peci peroxisomal delta3, delta2-enoyl-Coenzyme A 1,33,1 1,1 -1,8 336 104484_atL27121 Sth2 sulfotransferase, hydroxysteroid preferring 2 -1,2-2,7 1,1 -8,6 188 96605_atAI787183 Keg2 RIKEN cDNA 0610011I04 gene 1,3-1,1 2,0 -1,1 290 94916_atAW122260 Cyp51 cytochrome P450, 51 -1,11,7 -1,7 1,2 66 160388_atAI848668 Sc4mol sterol-C4-methyl oxidase-like -1,61,0 -3,5 -1,2 393 97518_atD29016 Fdft1 farnesyl diphosphate farnesyl transferase 1 -1,11,6 -2,1 -1,2 332 160711_atAI844846 Decr1 2,4-dienoyl CoA reductase 1, mitochondrial 2,13,3 1,7 -1,4 169 96095_i_atAI648018 Hsdl2 hydroxysteroid dehydrogenase like 2 1,33,3 1,3 -1,8 53 99571_atAW012588 Acaa1 3-ketoacyl-CoA thiolase B 2,52,8 2,0 -2,5 4993 96096_f_atAI648018 Hsdl2 hydroxysteroid dehydrogenase like 2 1,42,9 1,5 -1,5 127 102004_atAI530403 Acaa1 3-ketoacyl-CoA thiolase B, acetyl-Coenzyme 4,24,0 2,6 -2,7 836 96269_atAA716963 Idi1 isopentenyl-diphosphate delta isomerase -1,02,4 -2,3 1,2 512 160482_atAI841705 Acaa1 3-ketoacyl-CoA thiolase B, acetyl-Coenzyme 2,42,2 1,5 -1,4 1663 160737_atAW060927 Lss lanosterol synthase 1,42,1 1,0 -1,1 95 92581_atU07159 Acadm acetyl-Coenzyme A dehydrogenase, medium 2,23,0 1,4 -1,5 837 95632_f_atAW122653 Mvk mevalonate kinase -1,31,0 -1,7 1,0 101 103401_atL11163 Acads acyl-Coenzyme A dehydrogenase, short chain 1,71,3 1,1 -1,2 163 93868_atAL021127 Nsdhl NAD(P) dependent steroid -1,21,6 -2,0 -1,1 250 101515_atAF006688 Acox1 acyl-Coenzyme A oxidase 1, palmitoyl 2,53,9 1,4 -1,8 888 98631_g_atAW106745 Nsdhl NAD(P) dependent steroid -1,41,4 -1,9 -1,2 487 95588_atU89906 Amacr alpha-methylacyl-CoA racemase -1,4-1,0 -1,6 -1,1 285 102768_i_atAB016248 Sc5d sterol-C5-desaturase (fungal ERG3, 1,41,5 1,0 1,2 189 93045_atL28836 Abcd3 ATP-binding cassette, sub-family D (ALD), 1,42,6 1,2 -1,2 1654 102769_f_atAB016248 Sc5d sterol-C5-desaturase (fungal ERG3, 1,31,7 -1,0 1,2 236 93320_atAF017175 Cpt1a carnitine palmitoyltransferase 1, liver -1,31,7 1,0 -1,3 897 98989_atAF057368 Dhcr7 7-dehydrocholesterol reductase 1,81,7 1,2 -1,3 86 92600_f_atAB018421 Cyp4a10 cytochrome P450, family 4, subfamily a, 1,52,8 -1,1 -9,5 656 160217_atAI842715 Apmap Adipocyte plasma membrane-associated 1,32,1 1,0 -1,5 56 98353_atAI647548 Cyp4a10 cytochrome P450, family 4, subfamily a, 1,22,4 -1,0 -2,3 231 160306_atX95279 Thrsp thyroid hormone responsive SPOT14 1,7-2,1 1,3 -1,8 1975 98527_atZ14050 Dci dodecenoyl-Coenzyme A delta isomerase 1,63,7 1,6 -1,9 449 93626_atAF103875 Abcg2 ATP-binding cassette, sub-family G (WHITE), 1,21,7 1,2 -1,1 142 97316_atAJ011864 Ehhadh enoyl-Coenzyme A hydratase/ 3-hydroxyacyl 4,022,1 6,9 -1,7 76 98962_atX58426 Lipc lipase, hepatic -2,1-2,1 -2,3 -1,2 330 97456_atAI838021 Facl5 fatty acid Coenzyme A ligase, long chain 5 1,93,4 2,1 -1,3 415 101539_f_atAW226939 Ces3 carboxylesterase 3 1,72,1 1,1 -2,0 1527 101045_atU96116 Hadh2 hydroxyacyl-Coenzyme A dehydrogenase 1,6-1,0 -2,1 -2,1 1042 101538_i_atAW226939 Ces3 carboxylesterase 3 1,52,0 -1,1 -2,8 1533 96913_atAW122615 Hadhb hydroxyacyl-Coenzyme A 1,43,5 1,7 -1,4 657 18 nucleotide metabolism 95485_atD29639 Hadhsc L-3-hydroxyacyl-Coenzyme A dehydrogenase, 1,31,6 -1,1 -1,7 642 102373_atM29961 Enpep glutamyl aminopeptidase -2,7-3,5 -2,5 -2,0 260 96608_atAF023463 Phyh phytanoyl-CoA hydroxylase -1,1-1,4 -1,9 -1,0 2062 102001_atM14223 Rrm2 ribonucleotide reductase M2 1,11,5 2,2 -1,0 16 103581_atY14004 Cte1 cytosolic acyl-CoA thioesterase 1 2,220,0 4,6 -4,1 145 104213_atAI266885 Upp2 uridine phosphorylase 2 -1,8-2,0 -2,0 1,3 1091 94393_r_atAI317360 Elovl2 elongation of very long chain fatty acids -1,41,5 1,2 2,0 43 95722_atAB013137 Glrx1 glutaredoxin 1 (thioltransferase) -1,3-1,1 -1,1 -1,6 200 93496_atAI852098 Elovl5, ELOVL family member 5, elongation of long 2,12,5 2,1 -1,0 1006 161161_r_atAV216468 Nme1 expressed in non-metastatic cells 1, protein -1,0-1,2 -1,4 1,8 44 103665_atAW122523 Elovl6 ELOVL family member 6, elongation of long 1,0-1,2 2,4 -2,7 271 97317_atAW122933 Enpp2 ectonucleotide 1,1-1,3 -1,5 1,7 132 94418_atAI839004 Elovl6 ELOVL family member 6, elongation of long -1,1-1,3 2,4 -2,7 205 101196_atD50060 Pace4 paired basic amino acid cleaving system 4 1,11,1 -1,1 -1,6 183 160391_atAI853364 Fads1 fatty acid desaturase 1 1,52,3 1,6 1,0 476 93202_atL12059 Nt5e 5' nucleotidase, ecto 1,21,7 1,4 -1,6 30 98575_atX13135 Fasn fatty acid synthase 2,31,4 2,8 -1,8 1192 99521_atAB020239 Ak4 adenylate kinase 4 -1,2-1,3 -1,6 -1,4 72 103622_atAW050255 Mlycd malonyl-CoA decarboxylase 1,51,6 1,2 1,0 240 99959_atAW061337 Ak4 adenylate kinase 4 -1,3-1,4 -2,0 -1,3 129 94056_atM21285 Scd1 stearoyl-Coenzyme A desaturase 1 2,22,0 2,2 -1,0 3441 99039_g_atL24554 Adss2 adenylosuccinate synthetase 2, non muscle 1,21,5 1,5 -1,0 109 94057_g_atM21285 Scd1 stearoyl-Coenzyme A desaturase 1 2,72,4 2,6 -1,1 1946 98071_f_atX77731 Dck deoxycytidine kinase 1,11,6 1,5 1,1 87 94058_r_atAW047819 Scd1 stearoyl-Coenzyme A desaturase 1 -1,6-1,3 -1,6 -1,8 42 92561_atAJ238636 Entpd5 ectonucleoside triphosphate 1,31,6 1,2 -1,7 227 95758_atM26270 Scd2, stearoyl-Coenzyme A desaturase 2 2,38,8 14,8 -1,1 41 160107_atK01515 Hprt hypoxanthine guanine phosphoribosyl 1,51,7 1,2 -1,5 318 162077_f_atAV327760 Scd2, stearoyl-Coenzyme A desaturase 2 1,72,9 5,1 1,2 39 160652_atU49385 Ctps2 cytidine 5'-triphosphate synthase 2 1,01,5 1,2 -1,3 26 94075_atY14660 Fabp1 fatty acid binding protein 1, liver 1,21,7 1,2 1,1 3706 99985_atAB027565 Txnrd1 thioredoxin reductase 1 1,11,2 1,6 1,1 191 97889_atM65034 Fabp2 fatty acid binding protein 2, intestinal 1,84,7 2,7 -1,2 151 94381_atL31783 Umpk uridine monophosphate kinase 1,41,3 1,0 -1,5 270 94214_atX14961 Fabp3 fatty acid binding protein 3, muscle and heart 1,435,4 2,5 1,4 17 103341_atU49350 Ctps cytidine 5'-triphosphate synthase 1,31,7 1,7 -1,1 108 100567_atM20497 Fabp4 fatty acid binding protein 4, adipocyte 2,12,6 1,9 1,0 25

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 5 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0 Supplementary Figure fold changes estimate fold changes estimate

affy ID#genbank ID# symbol gene name RWT + RWT + db/db affy ID#genbank ID# symbol gene name RWT + RWT + db/db

2 retinol metabolism 102863_atAI550530 9130423L RIKEN cDNA 9130423L19 gene -1,4-1,3 -1,5 1,1 45 101431_atAF033195 Rdh5 retinol dehydrogenase 5 -1,3-1,6 -1,4 -1,1 155 94995_atAI854331 A030007L RIKEN cDNA A030007L17 gene 1,92,1 1,9 1,0 24 102797_atX95281 Dhrs3 retinal short-chain dehydrogenase/reductase -1,0-1,5 -1,7 -1,2 238 103744_atAI852760 A930014 RIKEN cDNA A930014C21 gene 1,81,6 1,2 -1,1 27 99451_atAW060510 C230093 RIKEN cDNA C230093N12 gene 1,21,7 1,2 -1,4 129 178 unknown 95910_f_atAI844780 C330008I RIKEN cDNA C330008I15 gene 1,21,7 1,2 -1,3 78 17 unknown 97710_f_atAI425990 C530046 RIKEN cDNA C530046L02 gene 1,5-1,0 1,2 1,2 23 92718_at AI158810 1,11,1 1,6 1,0 42 103257_atAA690483 C730036 RIKEN cDNA C730036B01 gene 1,21,9 -1,0 -1,2 283 104761_atAA612450 Antxr2 anthrax toxin receptor 2 -1,5-1,6 -1,5 -1,4 62 103702_i_atAI647632 C730048 RIKEN cDNA C730048C13 gene -2,3-3,6 -4,4 2,1 251 95749_atAW122364 Armet arginine-rich, mutated in early stage tumors -1,2-2,3 -1,3 1,4 347 103703_f_atAI647632 C730048 RIKEN cDNA C730048C13 gene -1,8-2,7 -2,7 1,7 312 93984_atAF002718 Atpi ATPase inhibitor 1,01,5 1,4 -1,2 136 99366_atAI553536 E030024 RIKEN cDNA E030024M05 gene -1,11,4 1,8 -1,0 11 162349_i_atAV173028 C1qtnf1 C1q and tumor necrosis factor related protein -1,2-1,4 -1,6 -1,1 20 93627_atAI852287 E430019 RIKEN cDNA E430019N21 gene -1,2-1,2 -1,5 -1,1 112 160769_atAW120606 Flana-pen carcinoma related gene -1,1-1,6 -1,2 1,2 131 104621_atAW046060 F830029L RIKEN cDNA F830029L24 gene -1,1-1,1 -1,6 1,1 426 103817_atAJ006469 Crtap cartilage associated protein -1,11,5 1,6 -1,1 66 103675_atY17793 Robo1 roundabout homolog 1 (Drosophila) 1,11,1 1,1 -1,7 22 93908_f_atX16670 Ccrn4l CCR4 carbon catabolite repression 4-like (S. 1,0-1,1 -1,4 -1,5 53 96237_atAI118905 Smaf1 small adipocyte factor 1 1,91,0 1,1 1,1 15 103391_atAI988033 C430041 cDNA sequence BC037135 1,11,1 -1,8 -1,2 124 97817_atAW121136 Spec1-pe small protein effector 1 of Cdc42 1,21,2 1,6 -1,1 68 97928_atAW045278 Cln6 ceroid-lipofuscinosis, neuronal 6 1,21,2 1,7 1,0 47 99127_atX61506 Sca10 spinocerebellar ataxia 10 homolog (human) 1,22,0 1,2 -1,0 185 94061_atM13018 Crip1 cysteine-rich protein 1 (intestinal) 1,11,2 2,5 -1,0 32 94515_atAW208628 Sqrdl sulfide quinone reductase-like (yeast) -1,0-1,5 -1,5 -1,3 153 96134_atAA755260 Dp1l1 deleted in polyposis 1-like 1 1,21,4 -1,3 -1,8 2246 95104_atU00674 Sdc2 syndecan 2, PRECURSOR (FIBROGLYCAN) -1,5-1,9 -1,8 1,1 171 96882_atAI842936 Ga17 dendritic cell protein GA17 -1,11,5 -1,1 1,1 130 102315_atAW124570 Tex292 testis expressed gene 292 -1,6-1,1 -1,2 1,2 45 94967_atAI851365 D19Wsu1 DNA segment, Chr 19, Wayne State 2,41,9 1,6 -1,3 47 96773_atAW125408 Txndc4 thioredoxin domain containing 4 (endoplasmic -1,3-1,6 -1,5 1,1 113 104116_atAW124049 D5Ertd59 DNA segment, Chr 5, ERATO Doi 593, 1,11,7 1,4 1,2 137 104755_atAJ242778 Tnip1 TNFAIP3 interacting protein 1 1,31,3 1,5 -1,0 73 97770_s_atAA733372 D6Wsu17 DNA segment, Chr 6, Wayne State University -1,01,1 1,2 2,0 30 93538_atAW228036 Ttrap Traf and Tnf receptor associated protein 1,81,8 1,4 -1,3 37 95709_atAW012491 D7Wsu86 DNA segment, Chr 7, Wayne State University -1,2-1,5 -1,4 1,0 910 160162_atAI852545 Tagln2 transgelin 2 1,01,1 2,1 -1,0 131 98503_atU53586 Evi5 ecotropic viral integration site 5 -1,3-1,2 -1,6 1,0 116 95431_atAA623426 Tomm70a translocase of outer mitochondrial membrane -1,01,6 1,1 1,3 38 103531_f_atAI049144 endoplasmic oxidoreductase 1 beta -1,7-1,9 -1,6 1,3 96 95542_atAI835858 Tpm4 tropomyosin 4 1,01,3 2,1 1,1 53 95386_atAI847050 AA40811 EST AA408112 1,41,7 1,2 -1,1 81 104177_atAA204579 Vig1-pend viral hemorrhagic septicemia virus(VHSV) 1,1-1,0 1,5 1,7 29 98948_atAI785289 Ns-pendin EST C77032 -1,01,2 1,5 -1,1 41 100522_s_atU92454 Wbp5 WW domain binding protein 5 -1,4-1,0 2,2 2,1 50 97918_atAA623587 AA53674 expressed sequence AA536743 1,41,4 2,2 1,3 20 104583_atAI227492 Zdhhc6 zinc finger, DHHC domain containing 6 -1,3-1,3 -1,6 1,2 56 94359_atAI849556 AA96055 expressed sequence AA960558 1,21,3 1,7 -1,1 65 104588_atAI255961 Hamp RIKEN cDNA 1810073K19 gene -1,8-2,8 -4,0 -1,1 1183 103400_atD50523 D50523 expressed sequence AI316828 -1,01,1 1,0 1,7 27 96122_atAW049373 2310016A RIKEN cDNA 2310016A09 gene 1,21,5 -1,4 -1,6 1002 99849_atC85523 AI553587 expressed sequence AI553587 1,51,8 1,9 -1,0 1491 100516_atAB030621 Chk choline kinase 1,11,1 1,5 1,1 64 93963_atAA200748 AI661017 expressed sequence AI661017 1,41,5 1,6 -1,0 17 160909_atAF057156 Sprr1a small proline-rich protein 1A 1,11,8 10,6 1,0 54 103227_atAI788959 AI788959 expressed sequence AI788959 1,21,6 1,4 1,3 402 95701_atAW124069 4930415K RIKEN cDNA 4930415K17 gene -1,6-1,7 -2,0 1,0 75 104413_atAI834976 AI834976 expressed sequence AI834976 1,01,5 1,2 1,2 311 103714_atAI265638 0610009A RIKEN cDNA 0610009A07 gene -1,6-1,8 -1,7 -1,3 149 103429_i_atAW125330 AL024210 expressed sequence AL024210 -1,2-1,3 -1,5 -1,1 157 161122_f_atAV032952 NADH dehydrogenase (ubiquinone) 1, 1,12,1 -1,1 -1,0 217 104333_atU69488 G7e-pend G7e protein 1,12,1 3,8 1,0 18 160648_atAI047076 Fignl1 fidgetin-like 1 1,02,1 1,9 1,1 23 101444_atU83176 Gt(ROSA) gene trap ROSA 26 antisense, Philippe -1,11,5 1,2 1,0 68 101537_atAF034435 Es1 esterase 1 -2,0-3,8 -2,0 1,3 670 93028_atX58196 H19 H19 fetal liver mRNAH19, imprinted 1,11,2 1,6 1,4 21 93534_atX53929 Dcn decorin -1,5-1,5 1,0 -1,2 567 93276_atU90123 Hn1 hematological and neurological expressed 1,31,1 1,6 1,1 48 103704_atAW047625 2010305K RIKEN cDNA 2010305K11 gene 1,31,9 1,1 -1,3 53 96117_r_atAI843732 H13 histocompatibility 13 -1,0-1,6 -1,1 1,2 200 100586_i_atAI843959 5730403B RIKEN cDNA 5730403B10 gene -1,3-1,4 -1,6 1,1 47 93824_atAI182073 4732433 hypothetical protein 4732433M03 -1,4-2,1 -1,4 1,3 3295 95621_atAA606367 9030623 RIKEN cDNA 9030623C06 gene 1,11,8 1,5 1,0 24 161214_r_atAV265258 MGC4743 hypothetical protein MGC47434 -1,1-1,2 1,1 1,7 17 103407_atAA895838 1300017J RIKEN cDNA 1300017J02 gene -1,2-1,6 -2,1 1,3 878 94428_atAI842668 Ilvbl ilvB (bacterial acetolactate synthase)-like 1,11,6 1,2 -1,4 60 98915_atAI849082 RNF149 ring finger protein 149 -1,6-1,5 -1,6 -1,4 133 95622_atAW123907 Klhdc2 kelch domain containing 2 1,11,7 1,6 1,6 20 102035_atAF037044 Tpmt thiopurine S-methyltransferase 1,1-1,2 -1,5 -1,3 444 96938_atAB028071 Keg1 kidney expressed gene 1 -2,2-2,1 -4,0 1,4 158 94536_s_atAI843417 2900073 RIKEN cDNA 2900073G15 gene 1,11,0 1,7 -1,5 691 98123_atAW226961 Kat2 kynurenine aminotransferase II -1,2-1,0 -1,9 1,3 63 95417_atAI117848 Mgat2 mannoside acetylglucosaminyltransferase 2 -1,5-1,7 -1,6 1,2 39 98440_atAA596710 Ltb4dh leukotriene B4 12-hydroxydehydrogenase -1,2-1,1 2,1 -1,5 28 97351_i_atAW123567 cytochrome c oxidase subunit VIb, 1,03,7 1,3 -1,2 13 96012_f_atAI835367 Matr3 matrin 3 -1,21,1 -1,1 1,7 101 97352_f_atAW123567 cytochrome c oxidase subunit VIb, 1,28,2 1,6 1,0 28 160119_atAI845538 LOC2238 matrix gamma-carboxyglutamate (gla) protein 1,1-1,2 -1,0 1,6 58 103442_atAI843399 LOC2168 cDNA sequence BC003479 1,91,7 1,2 -1,0 123 93974_atAW212475 Mig6 mitogen-inducible gene 6 protein homolog -2,2-1,9 -2,6 1,0 768 96643_atAW121336 Wfdc2 WAP four-disulfide core domain protein 2 1,91,2 1,6 -1,2 39 93975_atAI853531 Mig6 mitogen-inducible gene 6 protein homolog -2,2-1,6 -2,6 1,3 1831 93251_atAJ001260 Nipsnap1 4-nitrophenylphosphatase domain and 1,1-1,3 -1,7 -1,2 502 104300_atAI117936 Mus musculus 11 days embryo head cDNA, 1,11,1 1,7 -1,0 34 160455_s_atAW123786 ZWINT ZW10 interactor (ZW interacting protein-1) 1,21,6 1,3 -1,1 237 104661_atAF031164 Mus musculus 12 days embryo spinal 1,31,8 1,2 -1,1 30 97013_f_atAW046124 cytochrome b-245, alpha polypeptide 1,2-1,0 1,9 1,1 36 160273_atAA960603 Mus musculus Brf2 gene, 3' UTR 1,21,6 1,2 -1,0 100 93541_atZ68618 Tagln transgelin 1,11,1 1,5 1,1 43 104032_atAI850087 Mus musculus cDNA clone MGC:37981 -1,4-1,5 -1,6 -1,2 87 102111_f_at C81361 -1,21,0 1,0 1,6 23 103100_atAI788543 Mus musculus, clone IMAGE:5372338, 1,51,6 1,2 -1,1 27 93909_f_at X04120 -1,1-1,1 -1,3 -1,8 129 100880_atAA816121 Mdnr1 Mus musculus diabetic nephropathy-related 1,1-1,1 1,0 1,6 22 161504_i_atAV127302 D10Ertd2 DNA segment, Chr 10, ERATO Doi 214, 1,11,1 1,1 1,5 16 103203_f_atAW259499 Mus musculus transcribed sequence with 1,11,7 1,5 1,2 11 94440_atAI173533 AI876593 expressed sequence AI876593 -1,1-1,0 -1,3 -1,6 1178 103204_r_atAW259499 Mus musculus transcribed sequence with 1,11,9 1,2 1,1 21 104617_atAI042964 0610005 RIKEN cDNA 0610005C13 gene -1,8-2,5 -2,9 -1,5 795 96556_atC81463 Mus musculus transcribed sequence with -1,4-1,3 -1,6 -1,2 28 95026_atAW047688 0610039 RIKEN cDNA 0610039N19 gene 1,86,3 1,5 -3,5 414 100944_atAA958903 Mus musculus transcribed sequences -1,0-1,9 -1,4 1,1 169 97514_atAW046438 1810063B RIKEN cDNA 1810063B05 gene 1,0-1,1 -1,2 -1,5 131 95901_f_atAI662099 Mus musculus transcribed sequences -1,5-2,1 -1,6 -1,1 36 103403_atAW060833 2310007 RIKEN cDNA 2310007H09 gene -1,0-1,2 -1,3 -1,5 77 102780_atZ31362 Npn3 neoplastic progression 3 1,62,2 4,1 -1,2 41 103529_atAW125505 2600017J RIKEN cDNA 2600017J23 gene -1,4-1,5 -1,5 1,6 47 161090_i_atAV374584 Npn3 neoplastic progression 3 -1,5-1,7 -1,8 1,5 98 162424_f_atAV291989 2610007K RIKEN cDNA 2610007K22 gene 1,21,2 1,0 1,8 46 92500_atAB025412 Odz3 odd Oz/ten-m homolog 3 (Drosophila) 1,21,8 1,2 -1,1 23 95134_atAW124483 3110038L RIKEN cDNA 3110038L01 gene -1,1-1,3 1,1 -1,5 111 93495_atU96746 Prdx4 peroxiredoxin 4 -1,7-1,3 -1,3 1,2 169 104445_atAW046694 4631408 RIKEN cDNA 4631408O11 gene -1,1-1,3 1,1 1,6 32 98092_atAA790307 Plac8 placenta-specific 8 -1,11,0 3,0 -1,0 53 96089_atAI255972 4931406 RIKEN cDNA 4931406C07 gene 1,61,5 1,0 -1,8 247 93389_atAF039663 Prom1 prominin 1 1,11,4 2,3 1,1 27 96091_atAW060190 4931406 RIKEN cDNA 4931406C07 gene 1,51,4 1,1 -1,9 131 93390_g_atAF039663 Prom1 prominin 1 1,11,1 1,7 1,1 12 160661_atAI840615 5730472 RIKEN cDNA 5730472N09 gene 1,21,2 1,0 -1,5 116 94208_atAW045202 P5-pendin protein disulfide isomerase-related protein -1,5-1,9 -1,3 1,1 335 94415_atAA710439 6230421P RIKEN cDNA 6230421P05 gene 1,11,2 1,2 1,8 38 94085_atM34603 Prg proteoglycan, secretory granule 1,21,2 1,5 -1,1 15 95395_atAI225869 9130022A RIKEN cDNA 9130022A11 gene -1,11,0 -1,4 1,6 20 97835_atAI842828 RIKEN cDNA 0610007L05 gene -1,1-1,7 -1,9 -1,1 1894 103697_atAW061234 A230075 RIKEN cDNA A230075M04 gene -1,5-1,4 -1,6 2,6 61 104206_atAA815845 0610012A RIKEN cDNA 0610012A05 gene -1,5-1,6 -1,6 1,3 41 96545_s_atAI447510 A730042J RIKEN cDNA A730042J05 gene -1,0-1,6 -1,3 1,5 22 96348_atAW121217 0610039 RIKEN cDNA 0610039C21 gene 1,52,0 1,2 -1,0 296 160682_at AI851770 1,41,6 1,1 -1,0 98 101912_atAI019679 1100001 RIKEN cDNA 1100001G20 gene 1,3-1,8 1,1 1,3 1838 160255_atAA657044 1110004P RIKEN cDNA 1110004P15 gene 1,0-1,0 1,8 1,2 46 97908_atAW120676 1110007A RIKEN cDNA 1110007A06 gene 1,1-1,2 -1,5 -1,3 433 102922_atAI851387 5830436L RIKEN cDNA 1110020B03 gene -1,3-1,8 -1,4 1,0 56 160376_atAW125508 1110029F RIKEN cDNA 1110029F20 gene -1,31,7 -1,1 1,3 242 95690_atAW047363 1110030L RIKEN cDNA 1110030L07 gene 1,61,5 1,4 -1,2 263 96086_atAW122530 1110031B RIKEN cDNA 1110031B06 gene -1,4-1,6 -1,5 -1,0 173 98594_atAW125453 1190002 RIKEN cDNA 1190002N15 gene 1,42,7 1,6 1,3 32 160112_atAI852865 1190006E RIKEN cDNA 1190006E07 gene 1,31,4 1,9 1,1 14 160195_atAI846961 1200013P RIKEN cDNA 1200013P24 gene -1,1-1,1 -1,5 -1,1 327 98633_atAI854863 1200015A RIKEN cDNA 1200015A22 gene -1,11,1 1,7 -1,4 81 103409_atAA656550 1300006 RIKEN cDNA 1300006M19 gene 1,31,8 1,2 -1,2 74 94537_atAI838859 1500001 RIKEN cDNA 1500001M02 gene 1,1-1,1 1,6 -1,1 343 97885_atAB031386 1810009 RIKEN cDNA 1810009M01 gene 1,1-1,2 1,9 -1,1 913 160916_atAI838150 2310005P RIKEN cDNA 2310005P05 gene 1,61,6 1,1 -1,1 26 160801_atAW061073 2310009 RIKEN cDNA 2310009N05 gene -1,2-1,5 -1,1 -1,0 421 95627_atAW046038 2310058J RIKEN cDNA 2310058J06 gene 1,71,8 1,4 -1,4 55 95501_atAW122970 C200rf43 RIKEN cDNA 2410001C21 gene 1,61,8 1,2 -1,4 92 93138_atAI853219 2410012 RIKEN cDNA 2410012H22 gene 1,31,5 1,2 -1,1 69 102056_f_atAA839379 2610002J RIKEN cDNA 2610002J02 gene 1,11,6 1,6 1,4 63 100902_atAI846549 2610019F RIKEN cDNA 2610019F03 gene 1,41,7 1,1 -1,3 67 104045_atAW122372 2610102 RIKEN cDNA 2610102M01 gene -1,4-1,3 -1,6 -1,2 22 162057_f_atAV269118 2610103J RIKEN cDNA 2610103J23 gene -1,01,4 1,1 -1,5 91 98490_atAA822412 2610313E RIKEN cDNA 2610313E07 gene 1,31,8 1,2 -1,2 21 104403_atAI854118 2810417 RIKEN cDNA 2810417M16 gene 1,01,1 1,6 1,3 87 103071_atAI843655 2810429 RIKEN cDNA 2810429C13 gene 1,21,8 1,6 -1,1 14 96135_atAA833425 Q8CHQ7 RIKEN cDNA 3110003A17 gene 1,31,3 1,5 1,2 43 102784_atAI843949 4632413K RIKEN cDNA 4632413K17 gene -1,0-1,6 -1,4 1,0 159 96090_g_atAI255972 4931406 RIKEN cDNA 4931406C07 gene 1,61,4 1,1 -1,4 730 104284_atAI846126 5031439 RIKEN cDNA 5031439G07 gene 1,51,7 1,3 -1,0 182 100587_f_atAI843959 5730403B RIKEN cDNA 5730403B10 gene -1,4-1,2 -1,6 -1,1 58 94247_atAA600542 5730453 RIKEN cDNA 5730453H04 gene -1,11,6 1,4 -1,0 77 96634_atAI850090 5730469 RIKEN cDNA 5730469M10 gene 2,14,0 1,4 -1,0 78 95655_atAA717740 5830411E RIKEN cDNA 5830411E10 gene 1,21,4 2,2 1,1 16 103889_atAI834833 C230089I RIKEN cDNA 8030499H02 gene 1,11,6 1,1 1,4 36 104327_atAA755234 9030612 RIKEN cDNA 9030612M13 gene -1,3-1,1 -1,5 1,4 48

R ... Rosi vs untreated db/db ; W ... Wy vs untreated db/db ; T ... T09 vs untreated db/db ; + ... db/+ vs db/db Fold change page 6 <-5.0 <-2.0 <-1.5 >1.5 >2.0 >5.0