DOCSLIB.ORG

Nardilysin Is Required for Maintaining Pancreatic Β-Cell Function

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Nardilysin Is Required for Maintaining Pancreatic Β-Cell Function Diabetes Volume 65, October 2016 3015 Kiyoto Nishi,1 Yuichi Sato,2 Mikiko Ohno,1 Yoshinori Hiraoka,1,3 Sayaka Saijo,1 Jiro Sakamoto,1 Po-Min Chen,1 Yusuke Morita,1 Shintaro Matsuda,1 Kanako Iwasaki,2 Kazu Sugizaki,2 Norio Harada,2 Yoshiko Mukumoto,4 Hiroshi Kiyonari,4,5 Kenichiro Furuyama,6 Yoshiya Kawaguchi,6,7 Shinji Uemoto,6 Toru Kita,8 Nobuya Inagaki,2 Takeshi Kimura,1 and Eiichiro Nishi1 Nardilysin Is Required for Maintaining Pancreatic b-Cell Function Diabetes 2016;65:3015–3027 | DOI: 10.2337/db16-0178 Type 2 diabetes (T2D) is associated with pancreatic factors play important roles in the pathogenesis of the b-cell dysfunction, manifested by reduced glucose- disease, b-cell dysfunction, mainly manifested by deteriorated stimulated insulin secretion (GSIS). Several transcription glucose-stimulated insulin secretion (GSIS), seems to be pre- factors enriched in b-cells, such as MafA, control b-cell dominant for the transition from simple obesity to T2D (2). function by organizing genes involved in GSIS. Here we The observation that impaired GSIS in islets from patients demonstrate that nardilysin (N-arginine dibasic conver- with T2D cannot be accounted for by reduced insulin content tase; Nrd1 and NRDc) critically regulates b-cell function also supports this concept (1). Several biological processes, ISLET STUDIES Nrd12/2 through MafA. mice showed glucose intoler- including endoplasmic reticulum stress, inflammation, and ance and severely decreased GSIS. Islets isolated from oxidative stress, are suggested to impair GSIS in T2D (3,4). Nrd12/2 mice exhibited reduced insulin content and The pancreas comprises an exocrine compartment, impaired GSIS in vitro. Moreover, b-cell-specificNRDc- b consisting of acinar and ductal cells, and an endocrine com- deficient (Nrd1del ) mice showed a diabetic phenotype partment, consisting of a, b, d, e, and pancreatic polypep- with markedly reduced GSIS. MafA was specifically downregulated in islets from Nrd1delb mice, whereas tide cells. These different types of endocrine cells express overexpression of NRDc upregulated MafA and insulin and secrete the hormones glucagon, insulin, somatostatin, expression in INS832/13 cells. Chromatin immunopre- ghrelin, and pancreatic polypeptide, respectively. Sequen- cipitation assay revealed that NRDc is associated with tial expression of pancreatic transcription factors deter- Islet-1 in the enhancer region of MafA, where NRDc mines cell fate and pancreatic development. Among them, controls the recruitment of Islet-1 and MafA transcrip- several transcriptional factors enriched in b-cells, such as tion. Our findings demonstrate that NRDc controls b-cell Pdx-1, Nkx6.1, and MafA, play essential roles in the main- function via regulation of the Islet-1–MafA pathway. tenance of b-cell identity and function. b-Cell-specific ablation of Pdx-1 results in hyperglycemia with fewer Type 2 diabetes (T2D) is a common metabolic disorder insulin-positive cells and more glucagon-positive cells (5,6), that afflicts more than 300 million people globally (1). It whereas conditional Nkx6.1 inactivation in adult b-cells is characterized by impaired insulin secretion from pan- causes reduced insulin production (7). In contrast to the creatic b-cells and insulin resistance of the target tissues, earlier expression of Pdx-1 and Nkx6.1 in endocrine pro- such as liver, adipose tissue, and muscle. Although both genitor cells, MafA is first produced at embryonic day 13.5 1Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto 7Department of Clinical Application, Center for iPS Cell Research and Application, University, Sakyo-ku, Kyoto, Japan Kyoto University, Sakyo-ku, Kyoto, Japan 2Department of Diabetes, Endocrinology and Nutrition, Graduate School of Med- 8Kobe City Medical Center General Hospital, Chuo-ku, Kobe, Japan icine, Kyoto University, Sakyo-ku, Kyoto, Japan Corresponding author: Eiichiro Nishi, [email protected]. 3Division of Clinical Pharmacy, Faculty of Pharmaceutical Sciences, Kobe Gakuin Received 5 February 2016 and accepted 15 June 2016. University, Chuo-ku, Kobe, Japan 4Genetic Engineering Team, RIKEN Center for Life Science Technologies, Chuo-ku, This article contains Supplementary Data online at http://diabetes Kobe, Japan .diabetesjournals.org/lookup/suppl/doi:10.2337/db16-0178/-/DC1. 5Animal Resource Development Unit, RIKEN Center for Life Science Technologies, © 2016 by the American Diabetes Association. Readers may use this article as Chuo-ku, Kobe, Japan long as the work is properly cited, the use is educational and not for profit, and the 6Department of Surgery, Graduate School of Medicine, Kyoto University, Sakyo-ku, work is not altered. More information is available at http://www.diabetesjournals Kyoto, Japan .org/content/license. 3016 Nardilysin in Pancreatic b-Cell Function Diabetes Volume 65, October 2016 fl fl and is observed only in insulin-positive mature b-cells. ICR background (.98%). Nrd1 ox/ ox mice (accession no. MafA binds to an insulin promoter, like Pdx-1, and regu- CDB1019K; http://www.clst.riken.jp/arg/mutant%20mice% lates b-cell-selective insulin transcription (8). Mafa-deficient 20list.html) were generated by gene targeting in TT2 mice consistently develop diabetes because of impaired (23) embryonic stem (ES) cells (http://www.clst.riken.jp/ GSIS without a defect of islet cell development (9). More- arg/Methods.html). The targeting vector was designed to over, b-cell-specific ablation of pan-endocrine transcription insert loxP sites upstream and downstream of exon 1 of factors such as NeuroD1 and Islet-1 demonstrated their Nrd1 (Supplementary Fig. 1A). Successful homologous critical roles in b-cell maturation and function (10,11). In recombination in TT2 ES cells was confirmed by PCR particular, postnatal ablation of Islet-1 in b-cells resulted in (Supplementary Fig. 1A) and Southern blotting. Result- impaired GSIS without significantly reducing b-cell mass ingmutantmiceweregenotypedwithtailDNAbyPCR (11). Collectively, these findings demonstrated that tran- (Supplementary Fig. 1B) and crossed with CAG-FLPe trans- scription factors involved in the determination of endocrine genic mice to remove the neomycin selection cassette sur- or b-cell identity also play essential roles in b-cell functions. rounded by Flp recombinase target sites, then backcrossed A regulatory network of b-cell-enriched transcriptional to the C57BL/6J background (.98%). b-Cell-specificNRDc- fl fl factors (e.g., Pdx-1, NeuroD1, NKx6.1, and MafA) critically deficient mice were generated by crossing Nrd1 ox/ ox mice controls GSIS (5–10). These transcriptional factors also con- with rat insulin II promoter-Cre transgenic (RIP-Cre) nect oxidative stress with impaired GSIS in diabetic mouse mice (24). Male mice were used unless otherwise indi- models and human T2D (4). Among them, several lines of cated.Allanimalexperimentswereperformedaccording evidence have suggested that the Islet-1–MafA pathway to procedures approved by the Institute of Laboratory critically controls GSIS in adults. First, ablation of these Animals, Kyoto University. Mice were maintained on a genes in b-cells results in impaired GSIS without a severe diet of standard rodent chow in environmentally con- defect in b-cell development, as described above (9,11). trolled rooms. Second, MafA is a direct target gene of Islet-1 (11,12). Third, MafA directly regulates several genes critical for Glucose and Insulin Tolerance Tests GSIS, such as Ins1, Ins2,andGlut2 (also known as Slc2a2). For glucose tolerance tests, mice were fasted for 16 h and In this study, we demonstrate a novel regulatory intraperitoneally injected with 2 g/kg body weight of mechanism of GSIS by nardilysin (N-arginine dibasic D-glucose. For insulin tolerance tests, mice were fasted for convertase; Nrd1 and NRDc) through the Islet-1–MafA 5 h and intraperitoneally injected with insulin (Humulin pathway. NRDc is a zinc peptidase of the peptidase M16 R; Eli Lilly). To determine fasting plasma glucagon con- domain (M16) family that selectively cleaves dibasic sites centrations, mice were fasted for 16 h. To determine pan- (13,14). We rediscovered NRDc as a specificreceptorfor creatic insulin content and the proinsulin-to-insulin ratio, heparin-binding epidermal growth factor–like growth fac- whole pancreata were homogenized in 4 mL of a 2% tor (HB-EGF) (15), and our subsequent studies showed HCl/75% ethanol solution. After neutralization, samples multiple functions of NRDc, which depend on its cellular were diluted 1:1,000 in PBS before measuring insulin and localization. In the extracellular space, NRDc enhances proinsulin. Glucose was measured using a glucometer. Insulin ectodomain shedding of HB-EGF and other membrane pro- (MS303; Morinaga Institute of Biological Science, Yokohama, teins, such as tumor necrosis factor-a, amyloid precursor pro- Japan) and proinsulin (AKMPI-111; Shibayagi, Shibukawa- tein, and neuregulin-1 (16–20). We also revealed that NRDc City,Gunma,Japan)weremeasuredbyELISA.Glucagon in the nucleus works as a transcriptional coregulator was measured by enzyme immunosorbant assay (YK090; through the modulation of NCoR/SMRT corepressor or Yanaihara Institute, Fujinomiya-City, Shizuoka, Japan). PGC-1a coactivator function (21,22). We show here that Cell Culture fi 2/2 NRDc-de cient (Nrd1 ) mice and pancreatic b-cell- MIN6 and INS832/13 cells were gifts from Dr. Miyazaki fi fi delb 2 2 speci c NRDc-de cient (Nrd1 ) mice show glucose in- and Dr. Newgard, respectively. Nrd1 / mouse embryonic tolerance and severely impaired GSIS. Islets isolated from fi 2 2 broblasts (MEFs) were prepared as described previously / delb 2 2 Nrd1 and Nrd1 mice also show lower insulin pro- (19). MIN6 cells and Nrd1 / MEFs were grown in 2 duction and GSIS than wild-type islets. Notably, NRDc and DMEM (4.5 g L 1 glucose) supplemented with 10% FBS Islet-1 colocalize in the MafA enhancer, where NRDc reg- and antibiotics. INS832/13 cells were cultured as previ- ulates the recruitment of Islet-1 and MafA transcription, ously described (25). GM6001 (26) was purchased from indicating the essential role of NRDc in GSIS through Islet- Calbiochem (CA). For overexpression or gene knockdown – 1 MafA regulation.
Load more

Recommended publications
  • Nardilysin Is Involved in Autoimmune Arthritis Via the Regulation of Tumour Necrosis Factor Alpha Secretion
    Animal models RMD Open: first published as 10.1136/rmdopen-2017-000436 on 13 July 2017. Downloaded from ORIGINAL articLE Nardilysin is involved in autoimmune arthritis via the regulation of tumour necrosis factor alpha secretion Takayuki Fujii,1 Eiichiro Nishi,2,3 Hiromu Ito,1 Hiroyuki Yoshitomi,4 Moritoshi Furu,5 Namiko Okabe,6 Mikiko Ohno,2 Kiyoto Nishi,2 Yusuke Morita,2 Yugo Morita,1 Masayuki Azukizawa,1 Akinori Okahata,1 Takuya Tomizawa,1 Takeshi Kimura,2 Shuichi Matsuda1 To cite: Fujii T, Nishi E, ABSTRACT Key messages Ito H, et al. Nardilysin is Objective Tumour necrosis factor alpha (TNF-α) plays involved in autoimmune an important role in rheumatoid arthritis (RA). TNF-α is arthritis via the regulation synthesised as a membrane-anchored precursor and is What is already known about this subject? of tumour necrosis factor fully activated by a disintegrin and metalloproteinase 17 ► Tumour necrosis factor alpha (TNF-α), which alpha secretion. RMD Open (ADAM17)-mediated ectodomain shedding. Nardilysin is activated by ectodomain shedding, plays an 2017;3:e000436. doi:10.1136/ important role in rheumatoid arthritis (RA). rmdopen-2017-000436 (NRDC) facilitates ectodomain shedding via activation of ADAM17. This study was undertaken to elucidate the role ► Nardilysin facilitates TNF-α shedding via enhancing of NRDC in RA. a disintegrin and metalloproteinase 17 activity. Additional material is –/– ► Methods NRDC-deficient Nrdc( ) mice and macrophage- published online only. To view What does this study add? specific NRDC-deficientNrdc ( delM) mice were examined please visit the journal online ► Deletion or inhibition of nardilysin prevents (http:// dx. doi. org/ 10.
    [Show full text]
  • A High-Resolution Luminescent Assay for Rapid and Continuous Monitoring of Protein Translocation Across Biological Membranes
    De Castro Pereira, G., Allen, W., Watkins, D., Buddrus, L., Noone, D., Liu, X., Richardson, A., Collinson, I., & Chacinska, A. (2019). A high- resolution luminescent assay for rapid and continuous monitoring of protein translocation across biological membranes. Journal of Molecular Biology, 431(8), 1689-1699. https://doi.org/10.1016/j.jmb.2019.03.007 Publisher's PDF, also known as Version of record License (if available): CC BY Link to published version (if available): 10.1016/j.jmb.2019.03.007 Link to publication record in Explore Bristol Research PDF-document This is the final published version of the article (version of record). It first appeared online via Elsevier at https://doi.org/10.1016/j.jmb.2019.03.007 . Please refer to any applicable terms of use of the publisher. University of Bristol - Explore Bristol Research General rights This document is made available in accordance with publisher policies. Please cite only the published version using the reference above. Full terms of use are available: http://www.bristol.ac.uk/red/research-policy/pure/user-guides/ebr-terms/ Methods Notes A High-Resolution Luminescent Assay for Rapid and Continuous Monitoring of Protein Translocation across Biological Membranes Gonçalo C. Pereira 1, William J. Allen 1, Daniel W. Watkins 1, Lisa Buddrus 1,2, Dylan Noone 1, Xia Liu 1, Andrew P. Richardson 1, Agnieszka Chacinska 3 and Ian Collinson 1,2 1 - School of Biochemistry, University of Bristol, Bristol, UK 2 - BrisSynBio, University of Bristol, Bristol, UK 3 - Centre of New Technologies, University of Warsaw, S. Banacha 2c, 02-097, Warsaw, Poland Correspondence to Ian Collinson: School of Biochemistry, University Walk, University of Bristol, Bristol BS8 1TD, UK.
    [Show full text]
  • Functional Coverage of the Human Genome by Existing Structures, Structural Genomics Targets, and Homology Models
    Functional Coverage of the Human Genome by Existing Structures, Structural Genomics Targets, and Homology Models Lei Xie, Philip E. Bourne* San Diego Supercomputer Center and Department of Pharmacology, University of California, San Diego, California, United States of America The bias in protein structure and function space resulting from experimental limitations and targeting of particular functional classes of proteins by structural biologists has long been recognized, but never continuously quantified. Using the Enzyme Commission and the Gene Ontology classifications as a reference frame, and integrating structure data from the Protein Data Bank (PDB), target sequences from the structural genomics projects, structure homology derived from the SUPERFAMILY database, and genome annotations from Ensembl and NCBI, we provide a quantified view, both at the domain and whole-protein levels, of the current and projected coverage of protein structure and function space relative to the human genome. Protein structures currently provide at least one domain that covers 37% of the functional classes identified in the genome; whole structure coverage exists for 25% of the genome. If all the structural genomics targets were solved (twice the current number of structures in the PDB), it is estimated that structures of one domain would cover 69% of the functional classes identified and complete structure coverage would be 44%. Homology models from existing experimental structures extend the 37% coverage to 56% of the genome as single domains and 25% to 31% for complete structures. Coverage from homology models is not evenly distributed by protein family, reflecting differing degrees of sequence and structure divergence within families. While these data provide coverage, conversely, they also systematically highlight functional classes of proteins for which structures should be determined.
    [Show full text]
  • The Ectodomain Shedding Database for Membrane-Bound Shed Markers Wei-Sheng Tien1,2, Jun-Hong Chen3 and Kun-Pin Wu1*
    The Author(s) BMC Bioinformatics 2017, 18(Suppl 3):42 DOI 10.1186/s12859-017-1465-7 RESEARCH Open Access SheddomeDB: the ectodomain shedding database for membrane-bound shed markers Wei-Sheng Tien1,2, Jun-Hong Chen3 and Kun-Pin Wu1* From The Fifteenth Asia Pacific Bioinformatics Conference Shenzhen, China. 16-18 January 2017 Abstract Background: A number of membrane-anchored proteins are known to be released from cell surface via ectodomain shedding. The cleavage and release of membrane proteins has been shown to modulate various cellular processes and disease pathologies. Numerous studies revealed that cell membrane molecules of diverse functional groups are subjected to proteolytic cleavage, and the released soluble form of proteins may modulate various signaling processes. Therefore, in addition to the secreted protein markers that undergo secretion through the secretory pathway, the shed membrane proteins may comprise an additional resource of noninvasive and accessible biomarkers. In this context, identifying the membrane-bound proteins that will be shed has become important in the discovery of clinically noninvasive biomarkers. Nevertheless, a data repository for biological and clinical researchers to review the shedding information, which is experimentally validated, for membrane-bound protein shed markers is still lacking. Results: In this study, the database SheddomeDB was developed to integrate publicly available data of the shed membrane proteins. A comprehensive literature survey was performed to collect the membrane proteins that were verified to be cleaved or released in the supernatant by immunological-based validation experiments. From 436 studies on shedding, 401 validated shed membrane proteins were included, among which 199 shed membrane proteins have not been annotated or validated yet by existing cleavage databases.
    [Show full text]
  • Wo2015188839a2
    Downloaded from orbit.dtu.dk on: Oct 08, 2021 General detection and isolation of specific cells by binding of labeled molecules Pedersen, Henrik; Jakobsen, Søren; Hadrup, Sine Reker; Bentzen, Amalie Kai; Johansen, Kristoffer Haurum Publication date: 2015 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Pedersen, H., Jakobsen, S., Hadrup, S. R., Bentzen, A. K., & Johansen, K. H. (2015). General detection and isolation of specific cells by binding of labeled molecules. (Patent No. WO2015188839). General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. Users may download and print one copy of any publication from the public portal for the purpose of private study or research. You may not further distribute the material or use it for any profit-making activity or commercial gain You may freely distribute the URL identifying the publication in the public portal If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2015/188839
    [Show full text]
  • Enzyme DHRS7
    Toward the identification of a function of the “orphan” enzyme DHRS7 Inauguraldissertation zur Erlangung der Würde eines Doktors der Philosophie vorgelegt der Philosophisch-Naturwissenschaftlichen Fakultät der Universität Basel von Selene Araya, aus Lugano, Tessin Basel, 2018 Originaldokument gespeichert auf dem Dokumentenserver der Universität Basel edoc.unibas.ch Genehmigt von der Philosophisch-Naturwissenschaftlichen Fakultät auf Antrag von Prof. Dr. Alex Odermatt (Fakultätsverantwortlicher) und Prof. Dr. Michael Arand (Korreferent) Basel, den 26.6.2018 ________________________ Dekan Prof. Dr. Martin Spiess I. List of Abbreviations 3α/βAdiol 3α/β-Androstanediol (5α-Androstane-3α/β,17β-diol) 3α/βHSD 3α/β-hydroxysteroid dehydrogenase 17β-HSD 17β-Hydroxysteroid Dehydrogenase 17αOHProg 17α-Hydroxyprogesterone 20α/βOHProg 20α/β-Hydroxyprogesterone 17α,20α/βdiOHProg 20α/βdihydroxyprogesterone ADT Androgen deprivation therapy ANOVA Analysis of variance AR Androgen Receptor AKR Aldo-Keto Reductase ATCC American Type Culture Collection CAM Cell Adhesion Molecule CYP Cytochrome P450 CBR1 Carbonyl reductase 1 CRPC Castration resistant prostate cancer Ct-value Cycle threshold-value DHRS7 (B/C) Dehydrogenase/Reductase Short Chain Dehydrogenase Family Member 7 (B/C) DHEA Dehydroepiandrosterone DHP Dehydroprogesterone DHT 5α-Dihydrotestosterone DMEM Dulbecco's Modified Eagle's Medium DMSO Dimethyl Sulfoxide DTT Dithiothreitol E1 Estrone E2 Estradiol ECM Extracellular Membrane EDTA Ethylenediaminetetraacetic acid EMT Epithelial-mesenchymal transition ER Endoplasmic Reticulum ERα/β Estrogen Receptor α/β FBS Fetal Bovine Serum 3 FDR False discovery rate FGF Fibroblast growth factor HEPES 4-(2-Hydroxyethyl)-1-Piperazineethanesulfonic Acid HMDB Human Metabolome Database HPLC High Performance Liquid Chromatography HSD Hydroxysteroid Dehydrogenase IC50 Half-Maximal Inhibitory Concentration LNCaP Lymph node carcinoma of the prostate mRNA Messenger Ribonucleic Acid n.d.
    [Show full text]
  • Serine Proteases with Altered Sensitivity to Activity-Modulating
    (19) & (11) EP 2 045 321 A2 (12) EUROPEAN PATENT APPLICATION (43) Date of publication: (51) Int Cl.: 08.04.2009 Bulletin 2009/15 C12N 9/00 (2006.01) C12N 15/00 (2006.01) C12Q 1/37 (2006.01) (21) Application number: 09150549.5 (22) Date of filing: 26.05.2006 (84) Designated Contracting States: • Haupts, Ulrich AT BE BG CH CY CZ DE DK EE ES FI FR GB GR 51519 Odenthal (DE) HU IE IS IT LI LT LU LV MC NL PL PT RO SE SI • Coco, Wayne SK TR 50737 Köln (DE) •Tebbe, Jan (30) Priority: 27.05.2005 EP 05104543 50733 Köln (DE) • Votsmeier, Christian (62) Document number(s) of the earlier application(s) in 50259 Pulheim (DE) accordance with Art. 76 EPC: • Scheidig, Andreas 06763303.2 / 1 883 696 50823 Köln (DE) (71) Applicant: Direvo Biotech AG (74) Representative: von Kreisler Selting Werner 50829 Köln (DE) Patentanwälte P.O. Box 10 22 41 (72) Inventors: 50462 Köln (DE) • Koltermann, André 82057 Icking (DE) Remarks: • Kettling, Ulrich This application was filed on 14-01-2009 as a 81477 München (DE) divisional application to the application mentioned under INID code 62. (54) Serine proteases with altered sensitivity to activity-modulating substances (57) The present invention provides variants of ser- screening of the library in the presence of one or several ine proteases of the S1 class with altered sensitivity to activity-modulating substances, selection of variants with one or more activity-modulating substances. A method altered sensitivity to one or several activity-modulating for the generation of such proteases is disclosed, com- substances and isolation of those polynucleotide se- prising the provision of a protease library encoding poly- quences that encode for the selected variants.
    [Show full text]
  • The Positive Side of Proteolysis in Alzheimer's Disease
    Hindawi Publishing Corporation Biochemistry Research International Volume 2011, Article ID 721463, 13 pages doi:10.1155/2011/721463 Review Article Zinc Metalloproteinases and Amyloid Beta-Peptide Metabolism: The Positive Side of Proteolysis in Alzheimer’s Disease Mallory Gough, Catherine Parr-Sturgess, and Edward Parkin Division of Biomedical and Life Sciences, School of Health and Medicine, Lancaster University, Lancaster LA1 4YQ, UK Correspondence should be addressed to Edward Parkin, [email protected] Received 17 August 2010; Accepted 7 September 2010 Academic Editor: Simon J. Morley Copyright © 2011 Mallory Gough et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Alzheimer’s disease is a neurodegenerative condition characterized by an accumulation of toxic amyloid beta- (Aβ-)peptides in the brain causing progressive neuronal death. Aβ-peptides are produced by aspartyl proteinase-mediated cleavage of the larger amyloid precursor protein (APP). In contrast to this detrimental “amyloidogenic” form of proteolysis, a range of zinc metalloproteinases can process APP via an alternative “nonamyloidogenic” pathway in which the protein is cleaved within its Aβ region thereby precluding the formation of intact Aβ-peptides. In addition, other members of the zinc metalloproteinase family can degrade preformed Aβ-peptides. As such, the zinc metalloproteinases, collectively, are key to downregulating Aβ generation and enhancing its degradation. It is the role of zinc metalloproteinases in this “positive side of proteolysis in Alzheimer’s disease” that is discussed in the current paper. 1. Introduction of 38–43 amino acid peptides called amyloid beta (Aβ)- peptides.
    [Show full text]
  • Secretion of Matrix Metalloproteinases and Their
    [CANCER RESEARCH 53, 4493-4498, October 1, 1993] Secretion of Matrix Metalloproteinases and Their Inhibitors (Tissue Inhibitor of Metalloproteinases) by Human Prostate in Explant Cultures: Reduced Tissue Inhibitor of Metailoproteinase Secretion by Malignant Tissues 1 Balakrishna L. Lokeshwar, 2 Marie G. Seizer, Norman L. Block, and Zeenat Gunja-Smith Departments of Urology [B. L. L., M. G. S., N. L. B.] and Medicine [Z. G-S.], University of Miami School of Medicine, Miami, Florida 33101 ABSTRACT V, and IX, as well as the noncollagenous components of the extracel- lular matrix, including laminin and fibronectin (6). In addition, Unregulated secretion of matrix metalloproteinases (MMPs) or their MMP-3 is also known to activate procollagenases (7). Increased levels endogenous protein inhibitors (tissue inhibitor of metalloproteinases, of these proteinases have been implicated with the invasive potential TIMPs) has been implicated in tumor invasion and metastasis. Species of MMPs and TIMPs secreted by epithelial cultures of normal, benign, and of tumors (8-10). malignant prostate were identified and their levels were compared. Frag- Matrixins secreted by normal cells are proenzymes (zymogens) and ments of fresh tissue were cultured in a serum-free medium that supported are inactive (11). This is due to a highly conserved sequence of nine the outgrowth of prostatic epithelial cells. Biochemical analysis of the amino acid residues in the propeptide region containing a solitary conditioned media by gelatin zymography and enzyme assays showed that cysteine residue bound to the metal ion, Zn 2+, at the active center both normal and neoplastic tissues secreted latent and active forms of both (12).
    [Show full text]
  • High-Throughput Secretomic Analysis of Single Cells to Assess Functional Cellular Heterogeneity † ⊗ † ⊗ † ‡ † † § ∥ Yao Lu, , Jonathan J
    Article pubs.acs.org/ac High-Throughput Secretomic Analysis of Single Cells to Assess Functional Cellular Heterogeneity † ⊗ † ⊗ † ‡ † † § ∥ Yao Lu, , Jonathan J. Chen, , Luye Mu, , Qiong Xue, Yu Wu, Pei-Hsun Wu, Jie Li, ∥ † § † ⊥ Alexander O. Vortmeyer, Kathryn Miller-Jensen, Denis Wirtz, and Rong Fan*, , † Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, United States ‡ Department of Electrical Engineering, Yale University, New Haven, Connecticut 06520, United States § Department of Chemical and Biomolecular Engineering and the Johns Hopkins Physical Sciences-Oncology Center, Johns Hopkins University, Baltimore, Maryland 21218, United States ∥ Department of Pathology, Yale School of Medicine, New Haven, Connecticut 06520, United States ⊥ Yale Comprehensive Cancer Center, New Haven, Connecticut 06520, United States *S Supporting Information ABSTRACT: Secreted proteins dictate a range of cellular functions in human health and disease. Because of the high degree of cellular heterogeneity and, more importantly, polyfunctionality of individual cells, there is an unmet need to simultaneously measure an array of proteins from single cells and to rapidly assay a large number of single cells (more than 1000) in parallel. We describe a simple bioanalytical assay platform consisting of a large array of subnanoliter micro- chambers integrated with high-density antibody barcode microarrays for highly multiplexed protein detection from over a thousand single cells in parallel. This platform has been tested for both cell lines and complex biological samples such as primary cells from patients. We observed distinct heterogeneity among the single cell secretomic signatures that, for the first time, can be directly correlated to the cells’ physical behavior such as migration. Compared to the state-of-the-art protein secretion assay such as ELISpot and emerging microtechnology-enabled assays, our approach offers both high throughput and high multiplicity.
    [Show full text]
  • Zinc Metalloproteinases and Amyloid Beta-Peptide Metabolism: the Positive Side of Proteolysis in Alzheimer’S Disease
    Hindawi Publishing Corporation Biochemistry Research International Volume 2011, Article ID 721463, 13 pages doi:10.1155/2011/721463 Review Article Zinc Metalloproteinases and Amyloid Beta-Peptide Metabolism: The Positive Side of Proteolysis in Alzheimer’s Disease Mallory Gough, Catherine Parr-Sturgess, and Edward Parkin Division of Biomedical and Life Sciences, School of Health and Medicine, Lancaster University, Lancaster LA1 4YQ, UK Correspondence should be addressed to Edward Parkin, [email protected] Received 17 August 2010; Accepted 7 September 2010 Academic Editor: Simon J. Morley Copyright © 2011 Mallory Gough et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Alzheimer’s disease is a neurodegenerative condition characterized by an accumulation of toxic amyloid beta- (Aβ-)peptides in the brain causing progressive neuronal death. Aβ-peptides are produced by aspartyl proteinase-mediated cleavage of the larger amyloid precursor protein (APP). In contrast to this detrimental “amyloidogenic” form of proteolysis, a range of zinc metalloproteinases can process APP via an alternative “nonamyloidogenic” pathway in which the protein is cleaved within its Aβ region thereby precluding the formation of intact Aβ-peptides. In addition, other members of the zinc metalloproteinase family can degrade preformed Aβ-peptides. As such, the zinc metalloproteinases, collectively, are key to downregulating Aβ generation and enhancing its degradation. It is the role of zinc metalloproteinases in this “positive side of proteolysis in Alzheimer’s disease” that is discussed in the current paper. 1. Introduction of 38–43 amino acid peptides called amyloid beta (Aβ)- peptides.
    [Show full text]
  • Polymorphisms of the Matrix Metalloproteinase Genes
    www.nature.com/scientificreports OPEN Polymorphisms of the matrix metalloproteinase genes are associated with essential hypertension in a Caucasian population of Central Russia Maria Moskalenko1, Irina Ponomarenko1, Evgeny Reshetnikov1*, Volodymyr Dvornyk2 & Mikhail Churnosov1 This study aimed to determine possible association of eight polymorphisms of seven MMP genes with essential hypertension (EH) in a Caucasian population of Central Russia. Eight SNPs of the MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, and MMP12 genes and their gene–gene (epistatic) interactions were analyzed for association with EH in a cohort of 939 patients and 466 controls using logistic regression and assuming additive, recessive, and dominant genetic models. The functional signifcance of the polymorphisms associated with EH and 114 variants linked to them (r2 ≥ 0.8) was analyzed in silico. Allele G of rs11568818 MMP7 was associated with EH according to all three genetic models (OR = 0.58–0.70, pperm = 0.01–0.03). The above eight SNPs were associated with the disorder within 12 most signifcant epistatic models (OR = 1.49–1.93, pperm < 0.02). Loci rs1320632 MMP8 and rs11568818 MMP7 contributed to the largest number of the models (12 and 10, respectively). The EH-associated loci and 114 SNPs linked to them had non-synonymous, regulatory, and eQTL signifcance for 15 genes, which contributed to the pathways related to metalloendopeptidase activity, collagen degradation, and extracellular matrix disassembly. In summary, eight studied SNPs of MMPs genes were associated with EH in the Caucasian population of Central Russia. Cardiovascular diseases are a global problem of modern healthcare and the second most common cause of total mortality1,2.
    [Show full text]