Al‑Sulaiti et al. J Transl Med (2018) 16:175 https://doi.org/10.1186/s12967-018-1548-x Journal of Translational Medicine RESEARCH Open Access Triglyceride profling in adipose tissues from obese insulin sensitive, insulin resistant and type 2 diabetes mellitus individuals Haya Al‑Sulaiti1, Ilhame Diboun2, Sameem Banu1, Mohamed Al‑Emadi3, Parvaneh Amani3, Thomas M. Harvey1, Alex S. Dömling4, Aishah Latif1 and Mohamed A. Elrayess1,5* Abstract Background: Lipid intermediates produced during triacylglycerols (TAGs) synthesis and lipolysis in adipocytes inter‑ fere with the intracellular insulin signaling pathway and development of insulin resistance. This study aims to compare TAG species and their fatty acid composition in adipose tissues from insulin sensitive (IS), insulin resistant (IR) and type 2 diabetes mellitus (T2DM) obese individuals. Methods: Human subcutaneous and omental adipose tissue biopsies were obtained from 64 clinically characterized obese individuals during weight reduction surgery. TAGs were extracted from the adipose tissues using the Bligh and Dyer method, then were subjected to non-aqueous reverse phase ultra-high performance liquid chromatography and full scan mass spectrometry acquisition and data dependent MS/MS on LTQ dual cell linear ion trap. TAGs and their fatty acid contents were identifed and compared between IS, IR and T2DM individuals and their levels were cor‑ related with metabolic traits of participants and the adipogenic potential of preadipocyte cultures established from their adipose tissues. Results: Data revealed 76 unique TAG species in adipose tissues identifed based on their exact mass. Analysis of TAG levels revealed a number of TAGs that were signifcantly altered with disease progression including C46:4, C48:5, C48:4, C38:1, C50:3, C40:2, C56:3, C56:4, C56:7 and C58:7. Enrichment analysis revealed C12:0 fatty acid to be associated with TAGs least abundant in T2DM whereas C18:3 was found in both depleted and enriched TAGs in T2DM. Signifcant cor‑ relations of various adipose tissue-derived TAG species and metabolic traits were observed, including age and body mass index, systemic total cholesterol, TAGs, and interleukin-6 in addition to adipogenic potential of preadipocytes derived from the same adipose tissues. Conclusion: Pilot data suggest that adipose tissues from obese IR and T2DM individuals exhibit TAG-specifc signa‑ tures that may contribute to their increased risk compared to their IS counterparts. Future experiments are warranted to investigate the functional relevance of these specifc lipidomic profles. Keywords: Lipidomics, Adipose tissue, Triaclyglycerols, Insulin sensitivity, Insulin resistance, Type 2 diabetes mellitus Background of the adipose tissue, leading to progression of insu- Adipose tissue is the main site for storing and mobiliz- lin resistance and type 2 diabetes mellitus (T2DM) [1]. ing energy in response to metabolic demand. Obesity is However, a subset of obese individuals, known as the associated with changes in the structure and function insulin sensitive (IS) obese, maintain insulin sensitivity and exhibit better adipose tissue functions compared to equally obese insulin resistant (IR) counterparts [2]. Obe- *Correspondence: [email protected] 1 Toxicology and Multipurpose Lab, Anti Doping Laboratory Qatar, Sports sity triggers hypertrophy of adipocytes within the subcu- City, Doha, Qatar taneous (SC) adipose tissues to enable accumulation of Full list of author information is available at the end of the article excess triacylglycerols (TAGs). Additional energy intake © The Author(s) 2018. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creat​iveco​mmons​.org/licen​ses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creat​iveco​mmons​.org/ publi​cdoma​in/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Al‑Sulaiti et al. J Transl Med (2018) 16:175 Page 2 of 13 causes further fat accumulation within the omental (OM) Other chemicals and reagents were from Sigma (Munich, depot, which is associated with ectopic fat deposition in Germany). the liver, skeletal muscle and heart tissues [3]. Te subse- quent hyperinsulinemia inhibits hormone sensitive lipase Cohort and triggers the lipoprotein lipase causing additional glu- Participants’ recruitment criteria were described previ- cose intolerance, hyperinsulinemia, hypertriglyceridemia ously [12]. Briefy, 64 consented obese individuals under- and higher risk of insulin resistance in these tissues [4]. going bariatric surgery at AlEmadi hosptial (Doha, Qatar) Analysis of complex biological systems has become were recruited. Protocols were approved by Institutional possible by the newly emerging metabolomics tech- Review Board of ADLQ (X2017000224). Blood was taken niques where metabolites serve as direct indicators of prior to operation and 1–5 g of abdominal SC and OM biochemical activity of complex phenotypes such as insu- adipose tissues biopsies were collected during the surgery lin resistance and T2DM [5]. In this context, lipidomics and stored at − 80 °C until use. Plasma cholesterol, fast- studies were utilized to study diferences between SC and ing glucose and liver function enzymes were measured by OM depots. Tese studies have revealed depot-specifc COBAS INTEGRA (Roche Diagnostics, Basil). IL-6, leptin enrichment of specifc TAGs, glycerophospholipids, and and insulin were determined using commercially available sphingolipids and diferences in the association of lipid ELISA. Insulin resistance was computed by homeostatic species with body mass index, infammation and insulin model assessment (HOMA-IR) [13] using 30th percentile sensitivity [6, 7]. Although TAGs themselves are unlikely (HOMAIR = 2.4) as a threshold point. Accordingly, sub- to be signaling molecules, an increasing body of evidence jects were dichotomized into IS (HOMA-IR < 2.4, n = 18, suggests that lipid intermediates produced during TAG 3 males and 15 females) and IR (HOMA-IR > 2.4, n = 35, 9 synthesis or breakdown interfere with the intracellular males and 26 females). Eleven participants were clinically insulin signaling pathway and contribute to the develop- diagnosed with T2DM (8 males and 3 females). ment of insulin resistance, including free fatty acids, dia- cylglycerols and ceramides [8]. Indeed, elevated fatty acid Preadipocyte culture and diferentiation efux from the adipose tissue stimulates TAG synthesis Stromal vascular fraction (SVF) cells were obtained by col- in the liver and triggers stress of endoplasmic reticulum lagenase digestion of adipose tissues as described previously and stimulation of June kinase pathway in the adipose tis- [12]. Cell pellets were re-suspended in stromal medium sues [9, 10]. Tis leads to an overload of TAG’s synthetic containing Dulbecco’s modifed Eagle’s Medium-F12 capacity, causing an increase in both diacylglycerols (DMEM-F12) supplemented with 10% fetal bovine serum (DAGs) and ceramide levels and further development of (FBS) and Penicillin/Streptomycin, then maintained at insulin resistance in adipocytes [11]. 37 °C with 5% CO2 until confuence. To induce diferentia- Despite various studies investigating lipidomic difer- tion, early passaged stromal vascular fraction (SVF)-derived 4 2 ences in human serum and adipose tissues in relation to preadipocytes (passages 1–3) were grown at 2 × 10 /cm insulin sensitivity, no studies have compared diferences in stromal medium overnight, then incubated in diferen- in TAG signatures and their fatty acid composition in tiation medium (DMEM-F12, 3% FBS, 33 μM biotin, 17 μM adipose tissues from IS, IR and T2DM obese individu- d-pantothenate, 1 μM dexamethasone, 250 μM of methyl- als and their correlations with mediators of metabolic isobutylxanthine, 0.1 μM human insulin, 5 μM of Peroxi- disease. Identifcation of the fatty acids that are enriched some proliferator-activated receptor gamma PPARγ agonist, or depleted in tissues from insulin resistance and T2DM rosiglitazone) for 7 days, followed by 12 days in maintenance individuals could shed light on their functional role in medium containing the same components as the diferen- disease progression, thus providing potential novel tar- tiation medium but omitting methylisobutylxanthine and gets for therapeutic intervention. Te aims of this study rosiglitazone. Diferentiation potential (adipogenic capacity) were to profle TAG species and measure their levels in was determined as a percentage of lipidtox positive stained two fat depots and to compare their fatty acid composi- cells to total number of stained nuclei (DAPI). tion between IS, IR and T2DM individuals. Sample preparation Methods Human SC and OM adipose tissue specimens from IS, IR Materials and T2DM individuals were extracted using the Bligh and Interleukin 6 (IL-6) and leptin ELISAs were from R&D Dyer Method [14]. Homogenization of tissue was car- systems (Abingdon, UK). Insulin ELISA was from Mer- ried out in the gentle MACS Dissociator (Miltenyi Bio- codia Diagnostics (Uppsala, Sweden). 4′,6-Diamidino- tech, Germany) with one volume of PBS for every gram 2-phenylindole (DAPI), and LipidTOX Green Neutral of tissue. Following tissue homogenization, 1 mL of each Lipid were from