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Exenatide and Pioglitazone Regulate Fatty Acid-Induced Gene Expression in Normal and Diabetic Human Islets | OMICS International
ISSN: 2153-0769
Metabolomics:Open Access

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Exenatide and Pioglitazone Regulate Fatty Acid-Induced Gene Expression in Normal and Diabetic Human Islets

Hamedeh Ghanaat-Pour and Åke Sjöholm*

Karolinska Institutet, Department of Internal Medicine, Stockholm South Hospital, SE-118 83 Stockholm, Sweden

*Corresponding Author:
Dr. Åke Sjöholm
Karolinska Institutet
Department of Internal Medicine, Stockholm South Hospital
SE-118 83 Stockholm, Sweden
Tel: +4686163219
Fax: +4686163146
E-mail: [email protected]

Received date: August 25, 2011; Accepted date: October 27, 2011; Published date: October 29, 2011

Citation: Ghanaat-Pour H, Sjöholm Å (2011) Exenatide and Pioglitazone Regulate Fatty Acid-Induced Gene Expression in Normal and Diabetic Human Islets. Metabolomics 1:102. doi:10.4172/2153-0769.1000102

Copyright: © 2011 Ghanaat-Pour H, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Abstract

Chronically elevated free fatty acids (FFAs) are believed to have a detrimental effect on glucose-stimulated insulin secretion (GSIS), ?-cell function, and ?-cell survival in type 2 diabetes, a phenomenon often referred to as lipotoxicity. The objective of the present study was to investigate the global alterations in gene expression induced by long term exposure to palmitate with or without the PPAR-? agonist pioglitazone (a thiazolidinedione) or the GLP-1 receptor agonist exenatide (synthetic exendin-4, an incretin mimetic) in human diabetic and non-diabetic pancreatic islets, using microarray technology and confirmed by qRT-PCR. Gene expression profiling revealed alterations in several functional categories, such as epigenetic regulation of gene expression, cell proliferation and differentiation, metabolism, response to stimulus, transport, and signal transduction. Moreover, PPY, TCF4/TCF7L2, REG3A, GADD45B, TXNIP, ARNT, EGFR, CEL, ACACA, and GAPDH are some of the interesting genes that were differently regulated in our study. The apoptotic pathway was not substantially influenced by either pioglitazone or exenatide in the presence of palmitate. The function and survival of the human ?-cell seems to be controlled directly through the epigenetic control of gene expression rather than a direct effect on the apoptotic pathway. Considering that the nutritional state directly induces the epigenetic modifications, pioglitazone and exenatide appear to normalize these epigenetic misregulations and may protect the ?-cell from lipotoxic insult. The epigenetic modifications of the genome provide new promising targets for clinical diagnostics and also therapeutic purposes in the treatment of human type 2 diabetes by conferring ?-cell protection from lipotoxicity.

Keywords

Apoptosis; Exenatide; GK rat; Lipotoxicity; Microarray, Pancreatic islet, Pioglitazone, Type 2 diabetes, ARNT, TCF7L2/TCF4, GLP-1, PPAR

Abbreviations

ACACA, Acetyl-Coenzyme A carboxylase alpha; ARNT, Aryl hydrocarbon receptor nuclear translocator; CEL, Carboxyl ester lipase; EGFR, Epidermal growth factor receptor; FFA, free fatty acids; GADD45B, Growth arrest and DNA-damage-inducible beta; GAPDH, Glyceraldehyde-3-phosphate dehydrogenase; GLP- 1, glucagon-like peptide 1; PPY, Pancreatic polypeptide; PPAR-γ, peroxisome proliferator-activated receptor γ; REG3A, Regenerating islet-derived 3 alpha; TZDs, thiazolidinediones; TXNIP, Thioredoxin interacting protein; TCF7L2/TCF4, Transcription factor 7-like 2

Introduction

Elevated plasma free fatty acids (FFAs), caused by the excessive release of fatty acids from an expanded adipose tissue mass, are often linked to β-cell dysfunction in diabetes [1,2]. β-cell mass appears to be reduced in many cases of type 2 diabetic patients [3-5] and considering that type 2 diabetic patients usually are obese, it is unclear whether this defect arises due to genetic and/or environmental factors. The respective role of genetic and environmental factors is controversial, since most of the previous studies have been performed in animal models with a genetic predisposition to diabetes/obesity. Considering the fact that non-diabetic obese subjects show an increased β-cell volume [6], a genetic predisposition may be essential to β-cell failure and thereby β-cell mass reduction.

β-cell failure in type 2 diabetes has been previously suggested to be induced by increased FFAs alone [7,8], or in synergy with glucotoxicity [9-12]. Whether FFAs alter β-cell function and apoptosis in synergy with glucose, or by its own right, thus remains controversial.

Pioglitazone, a PPAR-γ agonist (member of the TZD family), seems to attenuate FFA-induced apoptosis in β-cells [13]. In contrast, troglitazone (another member of the TZD family) showed no protective effect against FFA-induced toxicity in β-cells [14].

Exenatide (synthetic exendin-4), an incretin mimetic and GLP-1 receptor agonist, has been shown to increase β-cell mass by enhancing proliferation/neogenesis, and its function [15-19]. Moreover, it stimulates β-cell fat metabolism [20,21] and protects against β-cell apoptosis [22,23]. The mechanism and the protective effect of these drugs against lipotoxicity in the β-cell remain unclear.

The aim of this study was to determine the long-term in vitro effect of palmitate, with or without pioglitazone or exenatide, on global gene expression in human pancreatic islets from healthy non-obese non-diabetic and non-obese diabetic subjects by using microarray technology. We thus wanted to identify genes influenced by palmitate, pioglitazone and/or exenatide, and also to compare the expression of genes in between the diabetic and non-diabetic groups of conditions.

Materials and Methods

Materials

The chemicals were obtained from the following sources: Collagenase type CLS (EC 3.4.24.3) from Roche (Mannheim, Germany). Culture medium RPMI-1640, fetal calf serum, L-glutamine, benzylpenicillin and streptomycin from Flow Laboratories (Irvine, U.K.). Sodium palmitate from Sigma (St. Louis, MO). All other chemicals of analytical grade were obtained from E. Merck (Darmstadt, Germany). Pioglitazone (Actos®) was generously provided by Takeda Pharmaceuticals North America (Deerfield, IL) and exenatide (Byetta®) was graciously donated by Amylin Inc. (San Diego, CA).

Human islet preparation and culture

Diabetic or non-diabetic human pancreatic islets were kindly provided by the Uppsala University Hospital facility for isolation of human islets from Scandinavian brain-dead donors. The type 2 diabetic donor was a 63-year-old male with BMI 22.6 and the nondiabetic donor was a 57-year-old female with BMI 20.7. The islets were maintained in RPMI-1640 tissue culture medium in the presence of 6 mM glucose, supplemented with 1 % (v/v) FBS, 2 mM L-glutamine, 100 IU/ml penicillin, and 100 μg/ml streptomycin, and were used for experiments ~ a week after isolation. A total of 36 (18 non-diabetic and 18 diabetic) islet samples were used for gene chip analyses after 48 hrs of culture: 3 non-diabetic samples of islets cultured in 6 mM glucose, 3 non-diabetic samples of islets cultured in 6 mM glucose and 0.125 mM palmitate, 3 non-diabetic samples of islets cultured in 6 mM glucose and 10 mM pioglitazone, 3 non-diabetic samples of islets cultured in 6 mM glucose, 0.125 mM palmitate and 10 mM pioglitazone, 3 nondiabetic samples of islets cultured in 6 mM glucose and 1 nM exenatide, 3 non-diabetic samples of islets cultured in 6 mM glucose, 0.125 mM palmitate and 1 nM exenatide, 3 diabetic samples of islets cultured in 6 mM glucose, 3 diabetic samples of islets cultured in 6 mM glucose and 0.125 mM palmitate, 3 diabetic samples of islets cultured in 6 mM glucose and 10 mM pioglitazone, 3 diabetic samples of islets cultured in 6 mM glucose, 0.125 mM palmitate and 10 mM pioglitazone, 3 diabetic samples of islets cultured in 6 mM glucose and 1 nM exenatide, and 3 diabetic samples of islets cultured in 6 mM glucose, 0.125 mM palmitate and 1 nM exenatide. At the end of the culture period, batches of ~ 500 islets were washed in PBS and transferred to Eppendorf tubes with 0.5 ml Trizol (Invitrogen, Carlsbad, CA), in which the islets were homogenized and then snap-frozen in liquid nitrogen and maintained in –70°C pending analysis. All experiments were approved by the Karolinska Institute Ethics Committee.

RNA extraction and sample preparation for Affymetrix gene analysis

Total RNA was extracted using the Qiagen RNAesy kit (WVR, Stockholm, Sweden) according to the manufacturer’s instructions. The integrity of the extracted RNA was confirmed by Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). Double-stranded cDNA was synthesized with 50 ng of total RNA using the SuperScript Choice system (Invitrogen, Carlsbad, CA). Synthesis of single-stranded complementary DNA (cDNA) and purification of double-stranded cDNA, synthesis and isolation of biotin-labeled complementary RNA (cRNA), quantification and fragmentation of cRNA, and target hybridization of fragmented cRNA, washing and staining of GeneChip expression probe arrays were all performed according to the manufacturer’s instructions (Technical manual of Affymetrix GeneChip products). Samples were analyzed by the Affymetrix Human Genome U133 Plus 2.0 Array interrogating over 47,000 transcripts. The level of expression of each gene was evaluated by the intensity of hybridization of labeled mRNA.

The probe arrays were scanned using the Agilent Gene Array Scanner (Agilent, Santa Clara, CA) and the scanned images were inspected and analyzed using established quality control criteria according to manufacturer’s instructions.

Data analysis

Data normalization and statistical analysis were performed using GeneSpring 7.3 software (Agilent, Santa Clara, CA). The data were normalized by utilizing the 50th percentile (per chip normalization) and the normalization to median (per gene normalization). To determine genes differentially expressed across conditions, the ANOVA test (considering False Discovery Rate, FDR) at P values <0.05, and a fold change of >1.5 were performed. Moreover, these genes were assigned to functional groups as described in GeneSpring (Agilent, Santa Clara, CA), NetAffx (Affymetrix) and other databases. Detailed descriptions of a selection of the differentially expressed transcripts, classified according to function and average fold change, are listed in Tables 1-4 and supplementary Tables 6-13.

cDNA synthesis

RNA was denatured for 10 min at 65°C and immediately chilled on ice. First strand cDNA synthesis was performed in a 20 μl reaction mixture containing 2 μg total RNA in a solution of 10 μl, 4 μl 5 × reverse transcriptase buffer (Invitrogen, Carlsbad, CA), 10 mM deoxynucleoside triphosphate, 1 μl random hexamer primers (100 pmol/μl), 1.5 μl dithiothreitol (Amersham Pharmacia Biotech, Piscataway, NJ) (100 mmol/l) and 1 μl RT1 reverse transcriptase (200 U/ μl; Invitrogen, Carlsbad, CA). The reagents were mixed and incubated at 37°C for 45 min. cDNA solutions were incubated for 5 min at 95°C to inactivate reverse transcriptase and then stored at –20°C.

Quantitative real-time PCR

Expression of selected genes (Table 5) was performed from total RNA using an ABI Prism 7700 polymerase chain reaction machine (PE Applied Biosystems, Foster City, CA). These genes were selected based on differential expression grounded on statistical analysis and fold change threshold, a select range of intensities, biological interest and potential roles in diabetes, and primer availability. Results are expressed relative to with and the ratio of different groups (Table 5).

cDNA samples were used as templates whereas an internal control (18S rRNA), was quantified as a positive control and used for normalization of the different template values. 18S rRNA was chosen among 34 candidate control genes previously checked using the TaqMan Human Endogenous Control Plate. Data were normalized using 18S rRNA because of the consistency of its level from sample to sample. To amplify the cDNA, 100 ng of the reversed transcribed cDNA from each sample were subjected to 40 cycles of real-time quantitative PCR in 20 μl of total reaction volume. Moreover, GADD45B, GAPDH, PPY, REG3A, TXNIP, CEL, ARNT, ISL1, TCF7L2/TCF4, and DAD1 probes and primers were used to flank the target DNA sequence. Each realtime TaqMan PCR reaction (20 μl) consisted of the cDNA template (except for the negative controls) and H2O to final volume of 9 μl, 1μl of TaqMan® Gene Expression Assay in 20 X format (containing the target primers and TaqMan probe) and 10 μl of TaqMan Fast Universal PCR Master Mix (2X), No AmpErase UNG (According to manufacturer’s instruction “TaqMan Gene Expression Assays”, Applied Biosystems). Aliquots were then amplified by an initial period of 2 min at 50°C and 10 min at 95°C followed by 40 concurrent cycles involving denaturation at 95°C for 15 s and annealing/extension at 60°C for 1 min The samples were placed in 96-well optical PCR plate (N 801- 0560, Perkin Elmer) and amplified in an automated fluorometer (ABI PRISM 7700 Sequence Detection System, Applied Biosystems). For relative quantification of different mRNAs, the following arithmetic formula was used according to the Perkin-Elmer Instruction Manual of 1997: Relative alteration = 2-ΔΔCT, where Ct is the point (cycle) at which the amplification plot crosses the threshold, which is defined as ΔCt = (Ct_sample2 – Ct_18S rRNA) - (Ct_sample1 Ct_18S rRNA). Therefore, the final value indicates an increase or decrease in mRNA for selected genes after normalization against 18S rRNA (Table 5).

Statistical analysis

Means ± S.E.M. were calculated and groups of data compared using Student’s t-test for paired or unpaired data. Differences were deemed statistically significant when P<0.05.

Results

Microarray analysis of global gene expression changes regulated by palmitate, pioglitazone and exenatide in normal and diabetic human islets

To gain detailed insight into the molecular mechanisms of palmitate, pioglitazone and exenatide action in isolated islets, we performed global gene expression profiling studies using Affymetrix GeneChip technology. Among the ~ 47,000 transcripts interrogated on the array, there were over 2,700 genes with known or inferred function that were affected by both pioglitazone and exenatide treatment, during palmitate exposure (Figure 1). A large number of differentially expressed genes belonged to several functional categories, such as epigenetic regulation of gene expression, cell proliferation and differentiation, metabolism, response to stimulus, transport, and signal transduction (supplementary Figure 2). Interestingly, epigenetic regulation of gene expression turned out to be the largest and most influenced transcript group in this study. The expressional profiles for these genes or ESTs are listed in Table 1 to 4 and supplementary Tables 6-13. In none of the analyses, the apoptotic pathway was noticeably influenced either by palmitate, pioglitazone or exenatide.

metabolomics-Hierarchical-clustering

Figure 1: Hierarchical clustering. Each gene is represented by a single row of colored bars. The red color indicates down-regulation, black color shows unchanged and the green color denotes up-regulation of gene expression compared to controls.

Function Gene Title Public ID Fold change  
      Palmitate Palmitate + pioglitazone Palmitate + exenatide
Regulation of gene expression, epigenetic Thyroid hormone receptor associated protein 1 (THRAP1) BC024236 11.2    
  Phosphoinositide-3-kinase, class 3 (PIK3C3) BC010388 9.1    
  Cardiac-MyBP-C associated Ca/CaM kinase (MLCK) BC039103 7.1    
  Acetyl-Coenzyme A carboxylase alpha (ACACA) BC007115 5.2    
  GLI-Kruppel family member GLI3 (Greig cephalopolysyndactyly syndrome) (GLI3) BC032660 5.6    
  Deoxynucleotidyltransferase, terminal (DNTT) AA585152 5.4    
  RAD51 homolog (RAD51) AL833420 4.7    
  Pleckstrin homology, Sec7 and coiled-coil domains 1(cytohesin 1)( PSCD1) CA442689 3.4    
  Sterol regulatory element binding transcription factor 1 (SREBF1) S66168 3.3    
  Rho-guanine nucleotide exchange factor (RGNEF) AB082529 3.2    
  Leukemia inhibitory factor receptor (LIFR) BC038371 3.0    
  Insulin-like growth factor binding protein 2, 36kDa (IGFBP2) NM_000597 2.9    
  Glutamate receptor, metabotropic 3 (GRM3) AI733361 2.8    
  Glutathione peroxidase 3 (GPX3) AW149846 2.5    
  Pre-B-cell leukemia transcription factor 1 (PBX1) AL832146 2.4    
  Transforming growth factor, beta receptor II (TGFBR2) AI809493 2.4    
  CAMP responsive element binding protein 1 (CREB1) AW945589 2.2    
  Nuclear receptor subfamily 1, group H, member 2 (NR1H2) NM_007121 1.8    
  Arginase (ARG1) AV649309 1.7    
  Fibroblast growth factor 10 (FGF10) NM_004465 1.7    
  Nodal homolog (NODAL) AI670948 1.7    
  X (inactive)-specific transcript (XIST) BE644917 1.6    
  Aryl hydrocarbon receptor nuclear translocator (ARNT) AL137290 1.6    
  Sp3 transcription factor (SP3) AW470841 1.5    
  Ribosomal protein L28 (RPL28) NM_000991   21.2 5.6
  Chromatin modifying protein 5 (CHMP5) NM_015961   18.4 15.6
  Hypothetical protein LOC285463 (CTBP1) BF984434   14.0 14.9
  Glutathione S-transferase A1(GSTA1) NM_000846   11.7 4.9
  Protease, serine, 3 (PRSS3) NM_002771   9.4 3.2
  Aldolase B, fructose-bisphosphate (ALDOB) BC005314   8.0 8.0
  ISL1 transcription factor, LIM/homeodomain, (islet-1)( ISL1) NM_002202   7.3 8.6
  Trypsinogen C (TRY6) U66061   7.3  
  Calreticulin (CALR) AA910371   7.2 9.8
  Ectonucleotide pyrophosphatase/phosphodiesterase 2 (autotaxin)( ENPP2) L35594   7.2 3.2
  Cytochrome P450, family 3, subfamily A, polypeptide 5 (CYP3A5) X90579   7.1 1.9
  Cysteine-rich, angiogenic inducer, 61 (CYR61) NM_001554   7.0 6.6
  Protease, serine, 2 (trypsin 2)( PRSS2) NM_002770   6.9 3.3
  Fibrinogen beta chain (FGB) NM_005141   6.9 4.3
  Human T-cell leukemia virus enhancer factor (HTLF) BF590117   5.7 6.4
  Regulator of G-protein signalling 4 (RGS4) AL514445   5.3 4.4
  Pancreatic polypeptide (PPY) M15788   5.2  
  CD53 antigen (CD53) NM_000560   4.8 3.6
  Leukocyte-derived arginine aminopeptidase (LRAP) BE889628   4.7 4.3
  Colipase, pancreatic (CLPS) NM_001832   4.6 1.9
  Potassium large conductance calcium-activated channel, subfamily M, beta member 2 (KCNMB2) AF209747   4.6 4.7
  Regenerating islet-derived 3 alpha (REG3A) NM_002580   4.2 2.0
  Phospholipase C, epsilon 1 (PLCE1) NM_016341   4.0 1.8
  Transcription factor 4 (TCF4) BF592782   3.9 5.3
  cAMP responsive element binding protein 1 (CREB1) M34356   3.7 2.5
  Transforming growth factor, beta 2 (TGFB2) BF061658   3.4 3.7
  lymphoid-restricted membrane protein (LRMP) NM_006152   3.3 2.3
  Cytochrome P450, family 2, subfamily A, polypeptide 6 (CYP2A6) M33318   3.3 3.2
  Bombesin-like receptor 3 (BRS3) Z97632   3.2 2.3
  Cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) AF182273   3.2 1.8
  Eukaryotic translation initiation factor 2-alpha kinase 1 (EIF2AK1) NM_014413   3.1 2.0
  Glutathione peroxidase 2 (gastrointestinal)(GPX2) NM_002083   3.3 3.3
  Ferritin, heavy polypeptide 1(FTH1) AA083483   3.1 3.7
  Meningioma expressed antigen 5 (hyaluronidase)( MGEA5) AK002091   3.0 1.7
  ADAM metallopeptidase domain 28 (ADAM28) NM_021778   3.0  
  Nuclear receptor subfamily 4, group A, member 1 (NR4A1) D85245   2.9 3.5
  High-mobility group box 1 (HMGB1) N92507   2.8 2.5
  Peripherin (PRPH) NM_006262   2.8 4.1
  Activating transcription factor 6 (ATF6) NM_007348   2.8 1.9
  GLIS family zinc finger 3 (GLIS3) AW025602   2.7 2.3
  DNA-damage-inducible transcript 3 (DDIT3) BC003637   2.7  
  Calcium channel, voltage-dependent, P/Q type, alpha 1A subunit (CACNA1A) AA769818   2.6 2.5
  Paired box gene 6 ( PAX6) NM_000280   2.6  
  Interleukin 7 receptor (IL7R) NM_002185   2.5 2.2
  Villin 2 (ezrin)( VIL2) BF663141   2.5 2.0
  Xanthine dehydrogenase (XDH) U06117   2.4 3.2
  Glutamate receptor, metabotropic 2 (GRM2) NM_000839   2.4 2.4
  Heparin-binding EGF-like growth factor (HBEGF) NM_001945   2.3 2.3
  Tumor necrosis factor (ligand) superfamily, member 13b (TNFSF13B) AW151360   2.3 2.5
  Major histocompatibility complex, class II, DR alpha (HLA-DRA) M60333   2.3  
  S100 calcium binding protein A8 (calgranulin A)( S100A8) NM_002964   2.2 2.0
  Major histocompatibility complex, class I, A (HLA-A) AI923492   2.2 2.1
  Major histocompatibility complex, class II, DR beta 1(HLA-DRB1) NM_002125   2.1  
  Apolipoprotein C-I (APOC1) W79394   2.1 1.9
  Pantothenate kinase 2 (PANK2) AV703394   2.0 2.0
  Neuropeptide Y (NPY) NM_000905   2.0 2.1
  Rho guanine nucleotide exchange factor (GEF) 12 (ARHGEF12) AI807672   2.0 1.8
  keratin 20 (KRT20) AI732381   2.0 1.9
  Nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 3 (NFATC3) U85430   2.0  
  Fms-related tyrosine kinase 1 (FLT1) AA058828   1.9 2.3
  Sp4 transcription factor (SP4) BF438799   1.9 2.1
  Synaptotagmin I (SYT1) AV731490   1.8  
  Major histocompatibility complex, class II, DQ beta 1(HLA-DQB1) AI583173   1.8  
  Interleukin 10 receptor, beta (IL10RB) BC001903   1.8  
  Phosphofructokinase, muscle (PFKM) U24183   1.7  
  Rap guanine nucleotide exchange factor (GEF) 3 (RAPGEF3) U78168   1.7 2.3
  Epidermal growth factor receptor (EGFR) BE878463   1.7  
  Chemokine (C-C motif) receptor 1 (CCR1) NM_001295   1.6 2.1
  Carboxypeptidase M (CPM) AW663908   1.6  
  FXYD domain containing ion transport regulator 3 (FXYD3) NM_005971   3.1 3.6
  PR domain containing 1, with ZNF domain (PRDM1) AI692659   1.9 3.1
  Myosin binding protein C (MYBPC3) NM_000256   2.7 2.4
  Bombesin-like receptor 3 (BRS3) Z97632   3.2 2.3
  Arachidonate 5-lipoxygenase (ALOX5) NM_000698     2.1
  Regenerating islet-derived 3 alpha (REG3A) NM_002580   4.2 2.0
  Bone morphogenetic protein 2 (BMP2) NM_001200     1.9
  Cytochrome P450, family 3, subfamily A, polypeptide 5 (CYP3A5) X90579   7.1 1.9
  Major histocompatibility complex, class II, DR beta 4 (HLA-DRB4) BC005312   1.6 1.9
  Colipase, pancreatic (CLPS) NM_001832   4.6 1.9
  Alpha-2-HS-glycoprotein (AHSG) AF130057     1.6
  WNT1 inducible signaling pathway protein 3 (WISP3) AF143679     1.5
Metabolism Potassium channel, subfamily K, member 1 (KCNK1) AL833343 4.3    
  Choline dehydrogenase (CHDH) AA609488 3.7    
  Nephronophthisis 3 (adolescent)(NPHP3) NM_152530 2.9    
  Hydroxysteroid (17-beta) dehydrogenase 12 (HSD17B12) BC012536 2.9    
  NADPH oxidase 1 (NOX1) AI826277 2.9    
  Tyrosinase (TYR) BC027179 2.9    
  Eukaryotic translation initiation factor 3, subunit 9 eta (EIF3S9) AA628539 2.6    
  Proteasome 26S subunit, non-ATPase, 7 (Mov34 homolog)( PSMD7) AW361702 2.3    
  Solute carrier family 18, member 2 (SLC18A2) NM_003054 2.3    
  Mevalonate kinase (MVK) AU150926 1.9    
  Mitochondrial ribosomal protein S12 (MRPS12) NM_021107 1.9    
  Transferrin receptor (TFRC) N76327 1.9    
  Nitric oxide synthase 1 (NOS1) BE299830 1.8    
  Transient receptor potential cation channel, subfamily M, member 6 (TRPM6) AK026281 1.8    
  Sorbin and SH3 domain containing 1 (SORBS1) AI377221 1.7    
  Solute carrier family 22, member 5 (SLC22A5) NM_003060 2.0 1.8 2.0
  Cystathionine-beta-synthase (CBS) BE613178 1.7    
  Sucrase-isomaltase (alpha-glucosidase)( SI) NM_001041 1.6    
  Secretin (SCT) NM_021920 1.5    
  Glutamate decarboxylase 2 (GAD2) NM_000818   10.7 8.2
  Glutaredoxin (thioltransferase) (GLRX) AF162769   5.2 2.3
  Voltage-dependent anion channel 3 (VDAC3) U90943   4.9 2.1
  Lectin, mannose-binding, 1 (LMAN1) U09716   4.7 4.7
  Collagen, type I, alpha 2 (COL1A2) NM_000089   4.0 3.8
  NADH dehydrogenase 1 beta subcomplex, 8 (NDUFB8) NM_005004   3.9  
  Ferritin, light polypeptide (FTL) BG538564   3.8  
  Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) AFFX-HUMGAPDH/M33197_5   3.6  
  Potassium inwardly-rectifying channel, subfamily J, member 15  (KCNJ15) D87291   3.6 2.0
  NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 2 (NDUFA2) BC003674   3.3 2.4
  Thioredoxin interacting protein (TXNIP) AI439556   3.2 1.8
  Potassium inwardly-rectifying channel, subfamily J, member 8 (KCNJ8) NM_004982   3.2 1.8
  Glycogenin (GYG) NM_004130   3.1  
  kynurenine 3-monooxygenase (kynurenine 3-hydroxylase) (KMO) AI074145   2.7 2.2
  Ceruloplasmin (ferroxidase) (CP) NM_000096   2.7  
  Biotinidase (BTD) AI767414   2.5 1.6
  Sterol O-acyltransferase 2 (SOAT2) AF059203   2.4 2.6
  Carbohydrate (chondroitin 4) sulfotransferase 11 (CHST11) AI806905   2.3 2.7
  SH3-domain GRB2-like 2 (SH3GL2) NM_003026   2.3 1.8
  Syntaxin 7 (STX7) NM_003569   2.2 1.9
  Matrix metallopeptidase 3 (stromelysin 1, progelatinase) (MMP3) NM_002422   2.2 2.1
  Carbonic anhydrase I (CA1) NM_001738   2.1 2.2
  Pyruvate dehydrogenase kinase, isoenzyme 3 (PDK3) NM_005391   2.1 1.6
  Catalase (CAT) AY028632   2.0  
  Synaptotagmin V(SYT5) NM_003180   2.0 1.8
  Phosphogluconate dehydrogenase (PGD) NM_002631   1.9  
  Steroid sulfatase, arylsulfatase C, isozyme S (STS) NM_000351   1.8 1.7
  ATP-binding cassette, sub-family C (CFTR/MRP), member 1 (ABCC1) NM_004996   1.8 2.3
  Glutathione S-transferase A3 (GSTA3) NM_000847   1.8  
  Sorbitol dehydrogenase (SORDa) L29008   1.7 2.1
  Carnitine palmitoyltransferase 1A (CPT1A) BC000185   1.7  
  Holocarboxylase synthetase (biotin-(proprionyl-Coenzyme A-carboxylase (ATP-hydrolysing)) ligase) (HLCS) D87328   1.6  
  Argininosuccinate synthetase (ASS) NM_000050   1.6  
  Monoglyceride lipase (MGLL) BC006230   1.6  
  NADH dehydrogenase 1 alpha subcomplex, 9 (NDUFA9) AF050641   1.5  
  NADH dehydrogenase 1 alpha subcomplex, 10 (NDUFA10) NM_004544   1.5  
  DEAD (Asp-Glu-Ala-Asp) box polypeptide 18 (DDX18) NM_006773   9.2 9.8
  Proprotein convertase subtilisin/kexin type 2 (PCSK2) AL031664   7.2 6.5
  Cyclic nucleotide gated channel alpha 1 (CNGA1) NM_000087   3.7 2.8
  Cytochrome b-245, alpha polypeptide (CYBA) NM_000101   2.6 2.1
  Cyclic nucleotide gated channel alpha 3 (CNGA3) NM_001298   1.8 2.1
  Ceruloplasmin (ferroxidase)(HPS3, CP) AI922198   2.4 2.0
  Phospholipase A2, group IIA (PLA2G2A) NM_000300     1.6
  Interleukin 27 receptor, alpha (IL27RA) NM_004843     1.5
Apoptosis Mitogen-activated protein kinase 1 (MAPK1) AL833111 4.7    
  BCL2/adenovirus E1B interacting protein like (BNIPL) NM_138279 3.2    
  Serine/threonine kinase 17b ( STK17B) AI221707 2.3    
  Ras homolog gene family, member B (RHOB) AI263909   9.5 13.3
  Defender against cell death 1 (DAD1) NM_001344   8.4 3.4
  Lectin, galactoside-binding, soluble, 1 (galectin 1)( LGALS1) NM_002305   6.6 3.0
  Engulfment and cell motility 1 (ELMO1) NM_014800   5.5 4.8
  Synuclein, alpha ( SNCA) L36675   3.6 2.7
  Ring finger and FYVE-like domain containing 1(RFFL) AI760772   3.3 3.5
  Growth arrest and DNA-damage-inducible, beta (GADD45B) AF078077   2.9 1.7
  Serine/threonine kinase 4 (STK4) BE222274   2.8 3.0
  Caspase 1, apoptosis-related cysteine peptidase (interleukin 1, beta, convertase)(CASP1) U13699   2.7  
  Mitogen-activated protein kinase kinase kinase 10 (MAP3K10) NM_002446   2.7 2.7
  Protein phosphatase 2 (formerly 2A), regulatory subunit A (PR 65), beta isoform (PPP2R1B) AF163473   2.0 1.9
  Peroxiredoxin 2 (PRDX2) L19185   1.8  
  NADH dehydrogenase 1 alpha subcomplex, 13 (NDUFA13) NM_015965   1.7  
  Programmed cell death 8 (PDCD8 ) NM_004208   1.7  
  TNF receptor-associated factor 2 (TRAF2) NM_021138   1.6  
  BCL2-associated athanogene (BAG1) AA394039   1.5  
  Apolipoprotein E (APOE) N33009   1.5 1.8
  Caspase 10, apoptosis-related cysteine peptidase (CASP10) NM_001230   1.5  
  Synuclein, alpha (non A4 component of amyloid precursor)( SNCA) L36675   3.6 2.7
  Secreted phosphoprotein 1 (osteopontin)( SPP1) AB019562     1.8
  S100 calcium binding protein, beta (S100B) BC001766     1.5
Response to stimulus Lymphotoxin beta (TNF superfamily, member 3)( LTB) BC018898 2.8    
  Calcium-sensing receptor (CASR) BF061333 2.1    
  Decay accelerating factor for complement (CD55)( DAF) CA448665 2.1    
  DnaJ (Hsp40) homolog, subfamily A, member 1(DNAJA1) NM_001539   8.0 5.6
  Chemokine (C-X-C motif) ligand 2 (CXCL2) M57731   5.0 5.1
  CD59 antigen p18-20 (CD59) BF983379   4.5 1.6
  CD58 antigen, (lymphocyte function-associated antigen 3) (CD58) D28586   3.6 2.3
  Decay accelerating factor for complement (DAF) BC001288   2.1  
  Coagulation factor XII (Hageman factor)( F12) BF530348   1.7 1.8
  Chemokine (C-C motif) ligand 5 (CCL5) NM_002985     1.8
  Interleukin 1 receptor antagonist (IL1RN) U65590     1.8
Signal transduction Leucine-rich repeat-containing G protein-coupled receptor 7 (LGR7) NM_021634 3.0    
  SH3 multiple domains 2 (SH3MD2) AB062480 3.0    
  GTPase, IMAP family member 1 (GIMAP1) NM_130759 2.1    
  Glutamate receptor, metabotropic 5 (GRM5) S64316 1.9    
  Growth differentiation factor 15 (GDF15) BC000529 1.7    
  Thromboxane A2 receptor (TBXA2R) U27325   1.7 2.1
  Protein tyrosine phosphatase, non-receptor type 1 (PTPN1) NM_002827   1.5  
  SH3-domain binding protein 2 (SH3BP2) AB000462   1.5 1.9
  TBC1 domain family, member 4 (TBC1D4) NM_014832   2.7 2.0

Table 1: Transcripts down-regulated by palmitate, with or without pioglitazone or exenatide, in diabetic human islets.

Function Gene Title Public ID Fold change  
      Palmitate Palmitate + pioglitazone Palmitate + exenatide
Regulation of gene expression, epigenetic Chromatin modifying protein 5 (CHMP5) NM_015961 17.5    
  Aldolase B, fructose-bisphosphate (ALDOB) BC005314 17.4    
  PHD finger protein 20 (PHF20) AL109965 12.7    
  ISL1 transcription factor, LIM/homeodomain, (ISL1) NM_002202 12.4    
  Potassium large conductance calcium-activated channel, subfamily M, beta member 2 (KCNMB2) AF209747 12.1    
  Nuclear receptor subfamily 2, group F, member 2 (NR2F2) AL037401 11.9    
  Ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2 )(autotaxin) L35594 10.5    
  Forkhead box N2 (FOXN2) BF590117 9.7    
  Transcription factor 4 (TCF4) BF592782 8.6    
  Regulator of G-protein signalling 4 (RGS4) AL514445 8.6    
  Calreticulin (CALR) AA910371 8.5    
  Fibrinogen beta chain (FGB) NM_005141 8.4    
  Glutathione S-transferase A1 (GSTA1) NM_000846 7.4    
  KH domain containing, RNA binding, signal transduction associated 1 (KHDRBS1) NM_006559 6.1    
  Leukocyte-derived arginine aminopeptidase (LRAP) BE889628 5.5    
  Ribosomal protein S19 (RPS19) BC000023 5.5    
  Cysteine-rich, angiogenic inducer, 61 (CYR61) NM_001554 5.5    
  GLIS family zinc finger 3 (GLIS3) AW025602 5.0    
  High-mobility group box 1(HMGB1) N92507 4.2    
  Regenerating islet-derived 3 alpha (REG3A ) NM_002580 4.1    
  Adducin 1 (alpha) (ADD1) NM_001119 3.9    
  Protease, serine, 2 (trypsin 2)( PRSS2) NM_002770 3.8    
  MAX-like protein X (MLX) N40555 3.7    
  Eukaryotic translation initiation factor 2-alpha kinase 1 (EIF2AK1) NM_014413 3.6    
  Sp4 transcription factor (SP4) BF438799 3.5    
  Aldolase B, fructose-bisphosphate (ALDOB) AV652403 3.5    
  Meningioma expressed antigen 5 (MGEA5) AK002091 3.4    
  Major histocompatibility complex, class I, A (HLA-A) AI923492 3.4    
  Runt-related transcription factor 1 (RUNX1) D43968 3.3    
  CD53 molecule (CD53) NM_000560 3.3    
  Corticotropin releasing hormone (CRH) NM_000756 3.2    
  Lymphoid-restricted membrane protein (LRMP) NM_006152 3.2    
  Platelet/endothelial cell adhesion molecule (PECAM1 )(CD31 antigen) AW574504 3.2    
  Cytochrome P450, family 3, subfamily A, polypeptide 5 (CYP3A5) X90579 3.0    
  Major histocompatibility complex, class II, DQ beta 1 (HLA-DQB1) AI583173 3.0    
  Retinoic acid receptor responder 2 (RARRES2) BC000069 3.0    
  Major histocompatibility complex, class II, DR beta 1 (HLA-DRB1) NM_002125 2.9    
  Calcium channel, voltage-dependent, P/Q type, alpha 1A subunit (CACNA1A) AA769818 2.9    
  Major histocompatibility complex, class II, DR beta 4 (HLA-DRB4) BC005312 2.8    
  Glutathione peroxidase 1(GPX1) NM_000581 2.8    
  Bombesin-like receptor 3 (BRS3) Z97632 2.8    
  Aldolase A, fructose-bisphosphate (ALDOA) AK026577 2.7    
  Neuropeptide Y (NPY) NM_000905 2.7    
  Major histocompatibility complex, class II, DR alpha (HLA-DRA) M60333 2.7    
  Fibroblast growth factor receptor 2 (FGFR2) NM_022975 2.7    
  ADAM metallopeptidase domain 28 (ADAM28) NM_021778 2.7    
  PHD finger protein 13 (PHF13) AL039384 2.7    
  Heparin-binding EGF-like growth factor (HBEGF) NM_001945 2.7    
  Amylase, alpha 2A (AMY2A) NM_004038 2.7    
  Signal transducer and activator of transcription 6, interleukin-4 induced (STAT6) BC004973 2.6    
  Pantothenate kinase 2 (PANK2) AV703394 2.6    
  Pancreatic polypeptide (PPY) M15788 2.5    
  Phosphofructokinase, muscle (PFKM) U24183 2.5    
  Major histocompatibility complex, class II, DR beta 5 (HLA-DRB5) NM_021983 2.5    
  PHD finger protein 15 (PHF15) AI735639 2.5    
  Guanine nucleotide binding protein (G protein), beta 5 (GNB5) NM_016194 2.5    
  Chemokine (C-C motif) receptor 1 (CCR1) NM_001295 2.4    
  Colipase, pancreatic (CLPS) NM_001832 2.4    
  Nuclear receptor subfamily 4, group A, member 1 (NR4A1) D85245 2.3    
  Acetyl-Coenzyme A carboxylase beta (ACACB) AI970898 2.3    
  Fms-related tyrosine kinase 1 (FLT1) AA058828 2.3    
  Rho guanine nucleotide exchange factor (GEF) 12 (ARHGEF12) AI807672 2.3    
  Vav 3 oncogene (VAV3) AF118887 2.2    
  Cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) AF182273 2.2    
  Islet cell autoantigen 1 (ICA1) L21181 2.2    
  Tumor necrosis factor receptor superfamily, member 13B (TNFRSF13B) NM_012452 2.1    
  S100 calcium binding protein A8 (S100A8) NM_002964 2.1    
  keratin 20 (KRT20) AI732381 2.1    
  Mitogen-activated protein kinase kinase 3 (MAP2K3) AA780381 2.1 2.4 2.0
  V-set domain containing T cell activation inhibitor 1 (VTCN1) NM_024626 2.0    
  Apolipoprotein C-I (APOC1) W79394 2.0    
  CDK5 regulatory subunit associated protein 1 (CDK5RAP1) NM_016408 2.0    
  Paired box gene 6 (PAX6) NM_000280 1.9    
  Phospholamban (PLN) NM_002667 1.9    
  DNA-damage-inducible transcript 3 (DDIT3) BC003637 1.9    
  Major histocompatibility complex, class II, DM alpha (HLA-DMA) X76775 1.8    
  Interleukin 7 receptor (IL7R) NM_002185 1.8    
  Solute carrier family 3, member 2 (SLC3A2) NM_002394 1.8    
  Epidermal growth factor receptor (EGFR) BE878463 1.8    
  Bone morphogenetic protein 2 (BMP2) NM_001200 1.8    
  Transforming growth factor, beta 2 (TGFB2) BF061658 1.7    
  Kruppel-like factor 5 (KLF5) AB030824 1.7    
  Glutamate-ammonia ligase (GLUL)(glutamine synthetase) U08626 1.7    
  Melanocortin 4 receptor (MC4R) NM_005912 1.7    
  Nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 3 (NFATC3) U85430 1.7    
  YY1 transcription factor (YY1) AA748649 1.7    
  EPH receptor B2 (EPHB2) AL530874 1.7    
  Mitogen-activated protein kinase kinase kinase 3 (MAP3K3) BG231756 1.6 2.1  
  Activating transcription factor 6 (ATF6) NM_007348 1.6    
  Xanthine dehydrogenase (XDH) U06117 1.6    
  Acetyl-Coenzyme A carboxylase beta (ACACB) AI057637 1.6    
  Forkhead box A3 (FOXA3) R99562 1.5 2.6 2.6
  Potassium voltage-gated channel, Isk-related family, member 3 (KCNE3) AI692703 1.5    
  Potassium voltage-gated channel, Shab-related subfamily, member 2 (KCNB2) AF338730 1.5    
  Thyroid hormone receptor associated protein 1 (THRAP1) BC024236   14.5 15.1
  LATS, large tumor suppressor, homolog 1 (LATS1) BC015665   11.6  
  Phosphoinositide-3-kinase, class 3 (PIK3C3) BC010388   11.4 11.0
  ATG10 autophagy related 10 homolog (ATG10) AF318326   11.1 12.6
  Protein tyrosine phosphatase, non-receptor type 18 (PTPN18) BC031076   9.9 11.2
  Cardiac-MyBP-C associated Ca/CaM kinase (MLCK) BC039103   8.5 9.4
  Acetyl-Coenzyme A carboxylase alpha (ACACA) BC007115   6.5 6.4
  Deoxynucleotidyltransferase, terminal (DNTT) AA585152   6.4 6.1
  GLI-Kruppel family member GLI3 (Greig cephalopolysyndactyly syndrome)(GLI3) BC032660   6.2 6.4
  Insulin-like growth factor binding protein 2 (IGFBP2) NM_000597   5.7 6.1
  Rho-guanine nucleotide exchange factor (RGNEF) AB082529   5.1 4.7
  RAD51 homolog (RAD51) AL833420   4.0 3.8
  Pre-B-cell leukemia transcription factor 1 (PBX1) AL832146   4.0 4.6
  Leukemia inhibitory factor receptor (LIFR) BC038371   3.8 4.1
  Glutamate receptor, metabotropic 3 (GRM3) AI733361   3.4 3.6
  Pleckstrin homology, Sec7 and coiled-coil domains 1(cytohesin 1)( PSCD1) CA442689   3.3  
  Cystatin C (CST3) NM_000099   3.1 5.3
  CAMP responsive element binding protein 1 (CREB1) AW945589   3.0 3.0
  Transforming growth factor, beta receptor II (TGFBR2) AI809493   2.9 2.9
  Apoptosis antagonizing transcription factor (AATF) AW471180   2.7 2.6
  Sterol regulatory element binding transcription factor 1 (SREBF1) S66168   2.5 3.0
  Glutathione peroxidase 3 (GPX3) AW149846   2.4 4.7
  Paired box gene 5 (B-cell lineage specific activator)( PAX5) AW405428   2.3 2.2
  LIM homeobox 4 (LHX4) AB055704   2.2 2.3
  Arginase (ARG1) AV649309   2.1 2.0
  Aryl hydrocarbon receptor nuclear translocator (ARNT) AL137290   2.1 2.4
  Fibroblast growth factor 12 (FGF12) D60438   2.1 2.3
  RAR-related orphan receptor B (RORB) R18374   2.1 2.1
  POU domain, class 1, transcription factor 1 (Pit1, growth hormone factor 1)( POU1F1) NM_000306   1.9 1.9
  Serpin peptidase inhibitor, clade H (heat shock protein 47), member 1 (SERPINH1) NM_004353   1.9 2.9
  Nodal homolog (NODAL) AI670948   1.8 1.6
  Fibroblast growth factor 14 (FGF14) BE221273   1.8  
  Forkhead box K2 (FOXK2) AV763408   1.8 1.7
  Pantothenate kinase 2 (PANK2) AI870137   1.8 1.7
  Fibroblast growth factor 10 (FGF10) NM_004465   1.8 1.6
  Major histocompatibility complex, class I, B (HLA-B) L07950   1.8 3.3
  Hepatocyte nuclear factor 4, gamma (HNF4G) NM_004133   1.7 1.7
  Guanidinoacetate N-methyltransferase (GAMT) NM_138924   1.7 2.0
  X (inactive)-specific transcript (XIST) BE644917   2.0  
  Nuclear receptor subfamily 1, group H, member 2 (NR1H2) NM_007121   1.6 1.6
  Pantothenate kinase 4 (PANK4) NM_018216   1.6 2.1
  RAR-related orphan receptor C (RORC) AI218580   1.6  
  Retinoid X receptor, beta (RXRB) BF337038   1.5 1.6
  5'-3' exoribonuclease 1 (XRN1) BC039314     12.4
  Homeodomain interacting protein kinase 1 (HIPK1) BC036361     10.1
  LATS, large tumor suppressor, homolog 1 (LATS1) BC015665     10.0
  CDC14 cell division cycle 14 homolog A (CDC14A) U29111     9.4
  High mobility group AT-hook 2 (HMGA2) U29113     8.1
  Mediator complex subunit 16 (MED16) AA314406     7.1
  Thyroid hormone receptor associated protein 1 (THRAP1) BC024236     4.0
  Insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) H93038     2.6
  HLA-G histocompatibility antigen, class I, G (HLA-G) M90684     2.4
  NAD(P)H dehydrogenase, quinone 1 (NQO1) BC000906     2.3
  Tubby like protein 4 (TULP4) AL157491     2.3
  Major histocompatibility complex, class II, DR beta 1 (HLA-DRB1) AJ297586     2.2
  X (inactive)-specific transcript (XIST) BF223193   2.0 1.9
  Rho guanine nucleotide exchange factor (GEF)10 (ARHGEF10) BC036809   2.2 1.9
  Major histocompatibility complex, class II, DQ beta 1 (HLA-DQB1) M17565     1.6
  Acetyl-Coenzyme A carboxylase beta (ACACB) AI057637     1.6
  Interleukin 10 receptor, beta (IL10RB) BC001903     1.5
  Angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (ACE2) NM_021804     1.5
Metabolism Proprotein convertase subtilisin/kexin type 2 (PCSK2) AL031664 19.4    
  DEAD (Asp-Glu-Ala-Asp) box polypeptide 3, Y-linked (DDX3Y) NM_004660 14.0    
  Thioredoxin domain containing 13 (TXNDC13) BF572868 13.0    
  Collagen, type I, alpha 2 (COL1A2) NM_000089 12.0    
  SEC24 related gene family, member A (SEC24A) AJ131244 11.3    
  Aftiphilin (AFTPH) BF055271 10.4    
  Fumarylacetoacetate hydrolase domain containing 1 (FAHD1) AW070436 9.7    
  Glutamate decarboxylase 2 (GAD2) NM_000818 8.8    
  SH3-domain GRB2-like 2 (SH3GL2) NM_003026 6.0    
  Glutathione reductase (GSR) AI888037 4.7    
  Matrix metallopeptidase 3 (MMP3) NM_002422 4.4    
  Solute carrier family 22, member 5 (SLC22A5) NM_003060 3.9 2.2 2.2
  Triosephosphate isomerase 1 (TPI1) BF116254 3.8    
  Potassium inwardly-rectifying channel, subfamily J, member 8 (KCNJ8) NM_004982 3.3    
  CDP-diacylglycerol synthase (phosphatidate cytidylyltransferase) 2 (CDS2) Y16521 3.3    
  Syntaxin 7 (STX7) NM_003569 3.3    
  Sorbitol dehydrogenase (SORD) L29008 3.1    
  Ras homolog gene family, member Q (RHOQ) BF978689 3.0    
  Voltage-dependent anion channel 3 (VDAC3) BC002456 2.8    
  Argininosuccinate synthetase 1(ASS1) NM_000050 2.7    
  ATP-binding cassette, sub-family C, member 1 (ABCC1) NM_004996 2.7    
  Prosaposin (PSAP) NM_002778 2.7    
  Sodium channel, nonvoltage-gated 1 alpha (SCNN1A) NM_001038 2.7    
  Potassium channel tetramerisation domain containing 10 (KCTD10) AW073741 2.5    
  Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) AFFX-HUMGAPDH/M33197_3 2.5    
  Pancreatic lipase-related protein 1 (PNLIPRP1) BC005233 2.5    
  Pyruvate dehydrogenase kinase, isozyme 3 (PDK3) NM_005391 2.4    
  Glutaredoxin (GLRX) AF162769 2.4    
  Triosephosphate isomerase 1 (TPI1) NM_000365 2.2    
  Ceruloplasmin (ferroxidase) (CP) AI922198 2.2    
  Chloride intracellular channel 5 (CLIC5) AL049313 2.1    
  Cytochrome b-245, alpha polypeptide (CYBA) NM_000101 2.1    
  Steroid-5-alpha-reductase, alpha polypeptide 1 (3-oxo-5 alpha-steroid delta 4-dehydrogenase alpha 1)(SRD5A1) BC006373 2.0    
  Carnitine palmitoyltransferase 1A (CPT1A) BC000185 2.0    
  Sterol O-acyltransferase 2 (SOAT2) AF059203 1.9    
  A kinase (PRKA) anchor protein 10 (AKAP10) AA456929 1.9    
  Thioredoxin interacting protein (TXNIP) AI439556 1.9    
  Acid phosphatase 1, soluble (ACP1) BE872974 1.8    
  Glutathione S-transferase M3 (GSTM3) AL527430 1.8    
  Glutathione S-transferase theta 1 (GSTT1) NM_000853 1.8    
  Lectin, galactoside-binding, soluble, 2 (LGALS2) NM_006498 1.8    
  Potassium inwardly-rectifying channel, subfamily J, member 15 (KCNJ15) D87291 1.8    
  Amine oxidase, copper containing 3 (AOC3) NM_003734 1.7    
  Chromosome 11 open reading frame 24 (C11orf24) NM_022338 1.6    
  Phospholipase A2, group IIA (PLA2G2A) NM_000300 1.5    
  Vacuolar protein sorting 13B (COH1) BC016375   15.5 13.3
  Carboxyl ester lipase (CEL) BC042510   12.8 10.9
  Potassium channel, subfamily K, member 1 (KCNK1) AL833343   6.7 6.4
  Choline dehydrogenase (CHDH) AA609488   6.0 6.2
  Tyrosinase (TYR) BC027179   4.0 4.2
  Hydroxysteroid (17-beta) dehydrogenase 12 (HSD17B12) BC012536   3.5 3.7
  NADPH oxidase 1 (NOX1) AI826277   3.2 2.7
  Lipocalin 2 (LCN2) NM_005564   3.1 3.1
  Sucrase-isomaltase (alpha-glucosidase)( SI) NM_001041   2.1 2.1
  Sorbin and SH3 domain containing 1 (SORBS1) AI377221   2.1 1.8
  Proprotein convertase subtilisin/kexin type 2 (PCSK2) AU158871   2.0 1.7
  Transferrin receptor (TFRC) N76327   2.0  
  Solute carrier family 18, member 2 (SLC18A2) NM_003054   1.9 2.1
  Paraoxonase 1 (PON1) U53784   1.9  
  Mevalonate kinase (MVK) AU150926   1.8 2.1
  Cystathionine-beta-synthase (CBS) BE613178   1.8 1.9
  HLA-B associated transcript 3 (BAT3) BC003133   1.6 1.5
  Alcohol dehydrogenase IB (class I), beta polypeptide (ADH1B) M21692   1.5  
  Solute carrier family 6, member 15 BC039443     12.7
  DEAD box polypeptide 3, Y-linked (DDX3Y) BC011022     12.1
  Protein phosphatase 2 (formerly 2A), regulatory subunit B, beta isoform (PPP2R2B) AF086066     5.0
  Recombination activating gene 1 (RAG1) BC037344     4.8
  Lipid storage droplet protein 5 (LSDP5) BC033570   4.6 4.6
  Transferrin receptor (TFRC) N76327     2.0
  Hexosaminidase (glycosyl hydrolase family 20, catalytic domain) containing (HEXDC) BM980844   2.4 2.4
  Paraoxonase 1 (PON1) U53784     1.8
  Solute carrier family 14, member 2 (SLC14A2) AL049328     1.7
  Farnesyl diphosphate synthase (FDPS) AL022163     1.6
  Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) BF689355     1.5
Apoptosis Defender against cell death 1 (DAD1) NM_001344 9.0    
  Ras homolog gene family, member B (RHOB) AI263909 9.4    
  Engulfment and cell motility 1 (ELMO1) NM_014800 5.9    
  Pleckstrin homology-like domain, family A, member 1 (PHLDA1) AI795908 5.2    
  Lectin, galactoside-binding, soluble, 1 (galectin 1)( LGALS1) NM_002305 5.2    
  Serine/threonine kinase 4 (STK4) BE222274 4.3    
  HIV-1 Tat interactive protein 2 (HTATIP2) NM_006410 3.8    
  Caspase 10, apoptosis-related cysteine peptidase (CASP10) NM_001230 2.9    
  NADH dehydrogenase 1 alpha subcomplex, 13 (NDUFA13) NM_015965 2.8    
  Caspase recruitment domain family, member 10 (CARD10) AY028896 2.5    
  Secreted phosphoprotein 1 (SPP1) AB019562 2.4    
  Inhibitor of growth family, member 4 (ING4) AA887083 2.3    
  Brain and reproductive organ-expressed (BRE) AL566299 2.2    
  Cytoplasmic FMR1 interacting protein 2 (CYFIP2) NM_030778 2.1    
  Nucleolar protein 3 (apoptosis repressor with CARD domain) AI912351 2.0    
  Death-associated protein (DAP) NM_004394 2.0    
  DNA fragmentation factor, beta polypeptide (DFFB) NM_004402 2.0    
  Tumor necrosis factor superfamily, member 13 (TNFSF13) AF114013 1.9    
  Mitogen-activated protein kinase kinase kinase 10 (MAP3K10) NM_002446 1.8    
  TNF receptor-associated factor 2 (TRAF2) NM_021138 1.7    
  Testis-specific kinase 2 (TESK2) NM_007170 1.6    
  CD14 molecule (CD14) NM_000591 1.6    
  Clusterin (CLU) M25915 1.6    
  Growth arrest and DNA-damage-inducible, beta (GADD45B) AF087853 1.6    
  Tumor necrosis factor receptor superfamily, member 12A (TNFRSF12A) NM_016639 1.6 1.9 2.0
  TNF receptor-associated factor 5 (TRAF5) BC032830   9.4 9.1
  Mitogen-activated protein kinase 1 (MAPK1) AL833111   6.1 6.3
  Dedicator of cytokinesis 1 (DOCK1) AI377320   4.0 3.6
  Serine/threonine kinase 17b ( STK17B) AI221707   3.7 3.9
  Programmed cell death 6 (PDCD6) BC02055   3.6 3.0
  Adrenergic, alpha-1A-, receptor (ADRA1A) BF221864   3.6 3.6
  BCL2/adenovirus E1B interacting protein like (BNIPL) NM_138279   3.0 2.7
  Sphingosine kinase 1 (SPHK1) NM_021972   2.3 2.4
  Protein phosphatase 1, regulatory (inhibitor) subunit 13B (PPP1R13B) AK023188   1.7 1.5
  Tumor necrosis factor receptor superfamily, member 10a (TNFRSF10A) W65310   1.6  
  Sterile alpha motif and leucine zipper containing kinase AZK (ZAK) AI475164   1.5 2.0
  Clusterin (CLU) M25915     4.1
  Apoptosis-inducing factor, mitochondrion-associated, 1(AIFM1) NM_004208     1.7
  NLR family, pyrin domain containing 1 (NLRP1) NM_021730     1.7
  Von Hippel-Lindau tumor suppressor (VHL) BF972755     1.6
Response to stimulus DnaJ (Hsp40) homolog, subfamily A, member 1 (DNAJA1) NM_001539 12.6    
  Interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) AI075407 7.5    
  Fusion (FUS)( CHOP) NM_004960 3.6    
  Major histocompatibility complex, class I, E (HLA-E) NM_005516 2.8    
  DnaJ (Hsp40) homolog, subfamily B, member 2 (DNAJB2) NM_006736 2.6    
  CD55 molecule, decay accelerating factor for complement (CD55) BC001288 2.5    
  CD58 molecule (CD58) D28586 1.9    
  Interleukin 1 receptor antagonist (IL1RN) U65590 1.9    
  Major histocompatibility complex, class I-related (MR1) NM_001531 1.7    
  Paraoxonase 3 (PON3) L48516 1.5    
  Activated leukocyte cell adhesion molecule (ALCAM) BC041127   8.9 8.5
  Decay accelerating factor for complement (DAF )(CD55) CA448665   3.5 8.8
  T cell receptor alpha locus ([email protected]) AE000659   1.8 1.9
  Lymphotoxin beta (TNF superfamily, member 3)( LTB) BC018898   1.7 1.8
  Olfactory receptor, family 2, subfamily H, member 1 (OR2H1) AJ459850     8.2
  Recombination activating gene 1 (RAG1) BC037344     4.8
  Tyrosinase (TYR) BC027179     4.2
  Lysozyme (LYZ) U25677     3.0
  REV1 homolog (REV1) W87784     2.7
  Paraoxonase 1 (PON1) U53784     1.8
Signal transduction Chemokine (C-X-C motif) ligand 2 (CXCL2) M57731 10.7    
  TRAF family member-associated NFKB activator (TANK) NM_004180 4.9    
  CD59 molecule (CD59) BF983379 2.9    
  TBC1 domain family, member 4 (TBC1D4) NM_014832 2.7    
  Urocortin (UCN) NM_003353 1.7    
  c-mer proto-oncogene tyrosine kinase (MERTK) NM_006343 1.7    
  Growth differentiation factor 15 (GDF15) BC000529   4.3 1.9
  SH3 multiple domains 2 (SH3MD2) AB062480   3.0 4.3
  Relaxin/insulin-like family peptide receptor 1(RXFP1) NM_021634   3.7 3.3
  SH3-domain GRB2-like 3 (SH3GL3) U79301   1.9 1.9
  Gastrin-releasing peptide (GRP) BF110750   1.8 1.7
  Hypothetical LOC554175 BC006530     8.3
  Ras and Rab interactor 3 (RIN3) BC034153     4.8
  GTPase, IMAP family member 1 (GIMAP1) NM_130759     3.4
  Mitogen-activated protein kinase associated protein 1 (MAPKAP1) NM_024117     1.8

Table 2: Transcripts up-regulated by palmitate, with or without pioglitazone or exenatide, in diabetic human islets.

Function Gene Title Public ID Fold change  
      Palmitate Palmitate + pioglitazone Palmitate + exenatide
Regulation of gene expression, epigenetic Fibroblast growth factor receptor 2 (FGFR2) NM_022975 1.7 3.4 2.4
  Calreticulin (CALR) AA910371 1.6    
  Aldolase B, fructose-bisphosphate (ALDOB) BC005314   6.5 5.4
  Tropomyosin 3, 4 (TPM3, TPM4) AF362887   4.9 7.4
  Insulinoma-associated 1 (INSM1) BF108585   2.5 2.1
  YY1 transcription factor (YY1) AA748649   2.5  
  Mitogen-activated protein kinase kinase 3 (MAP2K3) AA780381   2.4 1.7
  Acetyl-Coenzyme A carboxylase beta (ACACB) AI970898   1.9 1.8
  GLIS family zinc finger 3 (GLIS3) AW025602   1.7  
  Caveolin 2 (CAV2) AA150110   1.6 1.7
  Calcium channel, voltage-dependent, P/Q type, alpha 1A subunit (CACNA1A) AA769818   1.6  
  Aryl hydrocarbon receptor nuclear translocator (ARNT) AL137290     2.0
  Sp1 transcription factor (SP1) AU121035     1.9
  Cysteine-rich, angiogenic inducer, 61 (CYR61) NM_001554     1.8
  ATPase, Cu++ transporting, alpha polypeptide (ATP7A) NM_000052     1.6
  Transcription factor 4 (TCF4) BF592782     1.6
Metabolism Voltage-dependent anion channel 3 (VDAC3) U90943   2.3 2.4
  Ras homolog gene family, member Q (RHOQ) BF978689   2.0 2.3
  Solute carrier family 22, member 5 (SLC22A5) NM_003060   1.8 1.7
  ATP-binding cassette, sub-family C (CFTR/MRP), member 1 (ABCC1) NM_004996   1.6  
  ATPase, H+ transporting, lysosomal 9kDa, V0 subunit e (ATP6V0E) NM_003945   5.1 4.2
  Transferrin receptor (TFRC) N76327     1.6
  Transient receptor potential cation channel, subfamily C, member 1(TRPC1) U31110     1.6
  Alcohol dehydrogenase IB (class I), beta polypeptide (ADH1B) M21692     1.5
  Nuclear factor related to kappaB binding protein (NFRKB) AI869532     1.5
  Syntaxin 7 (STX7) NM_003569     1.5
Apoptosis Lectin, galactoside-binding, soluble, 1 (galectin 1)( LGALS1) NM_002305 1.9    
  Caspase recruitment domain family, member 10 (CARD10) AY028896 1.6    
  Serine/threonine kinase 4 (STK4) BE222274 1.5    
  Pleckstrin homology-like domain, family A, member 1 (PHLDA1) AI795908 1.7    
  Apolipoprotein E (APOE) N33009   2.5 1.5
  Secreted phosphoprotein 1 (SPP1) AB019562   4.8 3.0
  Tumor necrosis factor receptor superfamily, member 10a (TNFRSF10A) W65310   2.5  
  Ras homolog gene family, member B (RHOB) AI263909   2.4 2.1
  Neuronal thread protein AD7c-NTP (AD7C-NTP) AF010144   1.6 1.6
  Sema domain, immunoglobulin domain (Ig), transmembrane domain (TM) and short cytoplasmic domain, (semaphorin) 4D (SEMA4D) NM_006378   1.5  
  DNA fragmentation factor, 40kDa, beta polypeptide (caspase-activated DNase) (DFFB) NM_004402   1.5  
  Protein phosphatase 1, regulatory (inhibitor) subunit 13B (PPP1R13B) AK023188     1.8
  Programmed cell death 6 (PDCD6) AI692169     1.6
Signal transduction Growth differentiation factor 15 (GDF15) BC000529 1.9 7.4 7.5
  Striatin, calmodulin binding protein 4 (STRN4) NM_013403 1.6    
  Protein tyrosine kinase 9 (PTK9) N25562   3.1 1.7
  Chemokine (C-X-C motif) ligand 2 (CXCL2) M57731   2.3 2.0

Table 3: Transcripts down-regulated by palmitate, with or without pioglitazone or exenatide, in non-diabetic human islets.

Function Gene Title Public ID Fold change  
      Palmitate Palmitate + pioglitazone Palmitate + exenatide
Regulation of gene expression, epigenetic Hydroxyprostaglandin dehydrogenase 15-(NAD)(HPGD) NM_000860 2.3    
  Vav 3 oncogene (VAV3) AF118887 1.8    
  Major histocompatibility complex, class II, DR alpha (HLA-DRA) M60333 1.8 2.7 3.8
  Alpha-2-glycoprotein 1, zinc-binding (AZGP1) D90427 1.8 1.8 2.5
  Nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 3 (NFATC3) U85430 1.7 1.7 2.3
  Glutathione peroxidase 3 (GPX3) AW149846 1.7    
  Glutamate-ammonia ligase (GLUL)(glutamine synthetase) U08626 1.7    
  Cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) AF182273 1.6    
  Angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (ACE2) AK026461 1.6 2.4  
  Paired box gene 8 (PAX8) BF056746 2.5 2.5 2.2
  Ribosomal protein S5 (RPS5) NM_001009   13.2 13.8
  Ribosomal protein L29 (RPL29) NM_000992   11.4 14.8
  Pancreatic polypeptide (PPY) M15788   9.9 7.7
  Ribosomal protein L28 (RPL28) NM_000991   8.6 10.1
  Heat shock 27kDa protein 1 (HSPB1) NM_001540   8.2 12.7
  Glutathione peroxidase 1(GPX1) NM_000581   7.8 10.4
  Colipase, pancreatic (CLPS) NM_001832   6.2 8.7
  Major histocompatibility complex, class II, DR beta 1 (HLA-DRB1) NM_002125   5.8 8.5
  Major histocompatibility complex, class II, DQ beta 1 (HLA-DQB1) AI583173   5.4 6.0
  Glutathione peroxidase 2 (gastrointestinal) NM_002083   4.7 5.9
  Glutathione S-transferase A1 (GSTA1) NM_000846   4.6 3.5
  Aldolase A, fructose-bisphosphate (ALDOA) AK026577   4.2 4.9
  Y box binding protein 1 (YBX1) BE966374   4.1 3.5
  Epidermal growth factor receptor (EGFR) BE878463   4.0 2.2
  Protease, serine, 2 (trypsin 2)( PRSS2) NM_002770   3.7 4.7
  Cytochrome P450, family 3, subfamily A, polypeptide 5 (CYP3A5) X90579   3.6 4.5
  Amylase, alpha 2A (AMY2A) NM_004038   3.4 2.7
  Regenerating islet-derived 3 alpha (REG3A ) NM_002580   3.3 3.6
  Major histocompatibility complex, class II, DR beta 5 (HLA-DRB5) NM_021983   3.1 3.9
  Eukaryotic translation initiation factor 4E family member 2 (EIF4E2) AF047695   3.2 4.0
  Eukaryotic translation initiation factor 4B (EIF4B) NM_001417   3.0 3.0
  ADAM metallopeptidase domain 28 (ADAM28) NM_021778   3.0 2.7
  Major histocompatibility complex, class II, DM alpha (HLA-DMA) X76775   2.9 3.0
  Interleukin 32 (IL32) NM_004221   2.8 3.0
  Cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) AF182273   2.7 2.4
  Bone morphogenetic protein 2 (BMP2) NM_001200   2.7 2.3
  Solute carrier family 3, member 2 (SLC3A2) NM_002394   2.7 4.2
  Vav 3 oncogene (VAV3) AF118887   2.6 2.2
  Eukaryotic translation initiation factor 2-alpha kinase 1 (EIF2AK1) NM_014413   2.6 2.7
  DNA-damage-inducible transcript 3 (DDIT3) BC003637   2.6 3.5
  Glutamate-ammonia ligase (GLUL)(glutamine synthetase) U08626   2.6 3.0
  Aldolase B, fructose-bisphosphate (ALDOB) BC005314   2.5 2.3
  S100 calcium binding protein A8 (S100A8) NM_002964   2.5 2.3
  Interleukin 10 receptor, beta (IL10RB) BC001903   2.4 3.7
  V-set domain containing T cell activation inhibitor 1 (VTCN1) NM_024626   2.3 3.0
  Serpin peptidase inhibitor, clade H (heat shock protein 47), member 1 (SERPINH1) NM_004353   2.3 4.0
  X (inactive)-specific transcript (XIST) AV646597   2.3 3.3
  Major histocompatibility complex, class I, B (HLA-B) L07950   2.3 4.8
  MAX-like protein X (MLX) N40555   2.3 2.1
  Phosphofructokinase, muscle (PFKM) U24183   2.3 2.2
  Carboxypeptidase M (CPM) AW663908   2.2 2.8
  Islet cell autoantigen 1(ICA1) L21181   2.1 3.9
  Pantothenate kinase 4 (PANK4) NM_018216   2.1 2.8
  Fibrinogen beta chain (FGB) NM_005141   2.1  
  Activating transcription factor 6 (ATF6) NM_007348   2.0  
  Ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2 )(autotaxin) L35594   2.0 2.3
  Retinoic acid receptor responder 2 (RARRES2) BC000069   2.0 3.6
  CD53 molecule (CD53) NM_000560   1.9 2.0
  Ghrelin/obestatin preprohormone (GHRL) AB035700   1.9 1.9
  Serpin peptidase inhibitor, clade F, member 1 (SERPINF1) NM_002615   1.9 2.0
  Regulator of G-protein signalling 4 (RGS4) AL514445   1.9 1.9
  HLA-G histocompatibility antigen, class I, G (HLA-G) M90685   1.8 3.0
  Major histocompatibility complex, class I, C (HLA-C) U62824   1.7 2.6
  keratin 20 (KRT20) AI732381   1.7 1.6
  NAD(P)H dehydrogenase, quinone 1 (NQO1) BC000906   1.7 3.4
  HLA-G histocompatibility antigen, class I, G (HLA-G) M90684   1.7 3.0
  Nuclear receptor subfamily 1, group H, member 3 (NR1H3) NM_005693   1.6  
  Kruppel-like factor 5 (KLF5) AB030824   1.6 1.8
  PHD finger protein 15 (PHF15) AI735639   1.5  
  Rho guanine nucleotide exchange factor (GEF) 12 (ARHGEF12) AI807672   1.5 1.8
  Cystatin C (CST3) NM_000099   1.8 3.9
  Major histocompatibility complex, class II, DR beta 1 (HLA-DRB1) NM_002125   3.1 3.7
  Zinc finger protein 148 (ZNF148) L04282   2.6 3.2
  Major histocompatibility complex, class II, DM alpha (HLA-DMA) X76775   2.9 3.0
  CDK5 regulatory subunit associated protein 1(CDK5RAP1) NM_016408     2.7
  Cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) AF182273   2.7 2.4
  Forkhead box A3 (FOXA3) R99562     2.2
  Spleen tyrosine kinase (SYK) NM_003177     2.0
  Serpin peptidase inhibitor, clade H (heat shock protein 47), member 1 (SERPINH1) NM_004353   2.3 2.0
  cAMP responsive element binding protein 1 (CREB1) M34356     2.0
  General transcription factor IIH, polypeptide 1, 62kDa (GTF2H1) NM_005316     1.9
  Major histocompatibility complex, class I, A (HLA-A) AI923492     1.8
  Lamin B2 (LMNB2) M94363     1.7
  Guanine nucleotide binding protein (G protein), beta 5 (GNB5) NM_016194     1.6
  Eukaryotic translation initiation factor 4E family member 2 (EIF4E2) AF047695   3.2 1.6
  MAX-like protein X (MLX) N40555   1.7 1.6
  Lymphoid-restricted membrane protein (LRMP) NM_006152     1.6
  Growth arrest-specific 6 (GAS6) L13720     1.5
Metabolism Butyrobetaine (gamma), 2-oxoglutarate dioxygenase (gamma-butyrobetaine hydroxylase) 1 (BBOX1) NM_003986 2.6    
  Phosphogluconate dehydrogenase (PGD) NM_002631 1.5 2.0 3.0
  Pancreatic lipase-related protein 1 (PNLIPRP1) BC005233 1.5 4.4 4.9
  Thioredoxin interacting protein (TXNIP) AI439556 1.5 2.7 3.2
  Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) AFFX-HUMGAPDH/M33197_3   8.5 12.8
  Voltage-dependent anion channel 3 (VDAC3) BC002456   5.6 5.9
  Esterase D/formylglutathione hydrolase (ESD) BC001169   4.4 4.5
  Glutathione S-transferase M3 (GSTM3) AL527430   3.8  
  Argininosuccinate synthetase 1(ASS1) NM_000050   3.7 3.5
  Coenzyme Q9 homolog (COQ9) AC004382   3.3 4.2
  Acid phosphatase 1, soluble (ACP1) BE872974   3.0 3.3
  Potassium inwardly-rectifying channel, subfamily J, member 8 (KCNJ8) NM_004982   2.5 2.2
  Aminocarboxymuconate semialdehyde decarboxylase (ACMSD) AW025340   2.4 2.4
  Potassium inwardly-rectifying channel, subfamily J, member 15 (KCNJ15) D87291   2.4 2.5
  Glutathione S-transferase theta 1 (GSTT1) NM_000853   2.2 2.2
  Lipocalin 2 (LCN2) NM_005564   2.1 2.5
  Triosephosphate isomerase 1 (TPI1) BF116254   2.1 3.0
  Ceruloplasmin (ferroxidase) (CP) AI922198   2.0 2.1
  Syntaxin 17 (STX17) AW014619   2.0 3.1
  Cytochrome b-245, alpha polypeptide (CYBA) NM_000101   2.0 2.1
  kynurenine 3-monooxygenase (kynurenine 3-hydroxylase)( KMO) AI074145   1.9  
  Glutaredoxin (GLRX) AF162769   1.9 1.8
  Triosephosphate isomerase 1 (TPI1) NM_000365   1.9  
  Carboxyl ester lipase (bile salt-stimulated lipase)(CEL) BC042510   1.9 1.6
  Lectin, galactoside-binding, soluble, 2 (LGALS2) NM_006498   1.8  
  Matrix metallopeptidase 3 (MMP3) NM_002422   1.8 1.8
  Glutathione S-transferase A3 8 GSTA3) NM_000847   1.7 1.7
  Ceruloplasmin (ferroxidase) (CP) AI922198   1.7  
  SCO cytochrome oxidase deficient homolog 1 (SCO1) AF183424   1.6 3.0
  A kinase (PRKA) anchor protein 10 (AKAP10) AA456929   1.5 1.9
  Proprotein convertase subtilisin/kexin type 2 (PCSK2) AL031664   1.5 1.5
  Ferritin, light polypeptide (FTL) BG538564   8.4 18.1
  Ribosomal protein, large, P0 (RPLP0) AA555113   8.1 12.7
  Esterase D/formylglutathione hydrolase (ESD) BC001169   4.4 4.5
  Glutathione S-transferase M3 (GSTM3) AL527430   3.8 3.8
  Vesicle-associated membrane protein 5 (VAMP5) NM_006634   1.9 3.2
  Triosephosphate isomerase 1 (TPI1) BF116254   2.1 3.0
  Sulfotransferase family, cytosolic, 1A, phenol-preferring, member 1 (SULT1A1) NM_001055     2.2
  Potassium inwardly-rectifying channel, subfamily J, member 8 (KCNJ8) NM_004982   2.5 2.2
  HLA-B associated transcript 3 (BAT3) BC003133     1.8
  Chromosome 11 open reading frame 24 (C11orf24) NM_022338     1.7
  Glutamate dehydrogenase 2 (GLUD2) AC006144     1.6
  Prosaposin (PSAP) NM_002778     1.6
  Sorbin and SH3 domain containing 1 (SORBS1) AI377221     1.5
Apoptosis Apoptosis-inducing factor, mitochondrion-associated, 1 (AIFM1) NM_004208 1.8 1.8 2.2
  Apolipoprotein E (APOE) N33009 1.7   1.7
  Macrophage migration inhibitory factor (glycosylation-inhibiting factor)(MIF) NM_002415   9.3 10.3
  Lectin, galactoside-binding, soluble, 1 (galectin 1)( LGALS1) NM_002305   8.2 12.3
  Defender against cell death 1(DAD1) NM_001344   7.3 6.4
  Clusterin (CLU) M25915   3.9 6.3
  NADH dehydrogenase 1 alpha subcomplex, 13 (NDUFA13) NM_015965   3.3 4.1
  CD14 molecule (CD14) NM_000591   2.7 2.4
  Growth arrest and DNA-damage-inducible, beta (GADD45B) AF087853   2.6 4.3
  Tumor necrosis factor receptor superfamily, member 12A (TNFRSF12A) NM_016639   2.2 2.7
  Death-associated protein (DAP) NM_004394   1.9 3.9
  HIV-1 Tat interactive protein 2 (HTATIP2) NM_006410   1.9 2.7
  Inhibitor of growth family, member 4 (ING4) AA887083   1.8  
  NLR family, pyrin domain containing 1 (NLRP1) NM_021730   1.6 1.9
  Brain and reproductive organ-expressed (BRE) AL566299   1.6  
  Sphingosine kinase 1(SPHK1) NM_021972   1.5 1.9
  Engulfment and cell motility 1 (ELMO1) NM_014800     2.3
  Tumor necrosis factor (ligand) superfamily, member 13 (TNFSF13) AF114013     1.7
  caspase 1, apoptosis-related cysteine peptidase (interleukin 1, beta, convertase)( CASP1) U13699     1.7
  Cell division cycle 2-like 2 (CDC2L2) AF067524     1.5
  Programmed cell death 5 (PDCD5) NM_004708     1.5
Response to stimulus Interferon induced transmembrane protein 2 (1-8D)(IFITM2) NM_006435   6.4 7.8
  Interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) AI075407   4.2 5.3
  CD59 molecule, complement regulatory protein (CD59) BF983379   3.3 2.6
  CD55 molecule, decay accelerating factor for complement (CD55) BC001288   2.6 3.6
  C-mer proto-oncogene tyrosine kinase (MERTK) NM_006343   2.4 2.2
  Paraoxonase 3 (PON3) L48516   2.3 2.6
  Major histocompatibility complex, class I-related (MR1) NM_001531   2.2 2.0
  Major histocompatibility complex, class I, E (HLA-E) NM_005516   2.0 2.3
  DnaJ (Hsp40) homolog, subfamily A, member 1 (DNAJA1) NM_001539   1.5 2.3
  Calcium binding atopy-related autoantigen 1 (CBARA1) AK022697     1.8
Signal transduction Mitogen-activated protein kinase associated protein 1 (MAPKAP1) NM_024117   2.5 3.1
  TRAF family member-associated NFKB activator (TANK) NM_004180   2.2 4.7
  Docking protein 2, 56kDa (DOK2) AI828929     1.6

Table 4: Transcripts up-regulated by palmitate, with or without pioglitazone or exenatide, in non-diabetic human islets.

Differential expression of a subset of genes confirmed with qrt-PCR

To confirm the gene regulation observed with microarray analysis, we performed qRT-PCR on samples for selected genes that were differently regulated on the microarrays. The results generated by qRTPCR showed a highly significant correlation between the levels of gene expression measured by both methods (Table 5).

Gene Name   Fold Change
Growth arrest and DNA-damage-inducible, beta (GADD45B) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 2.7 ± 0.001 * †
     
  Down-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets -1.8 ± 0.002 * †
     
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 8.4 ± 0.001 * †
  Up-regulated by pioglitazone & palmitate vs. palmitate in non-diabetic human islets 2.0 ± 0.001 * †
     
  Down-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets -7.1 ± 0.001 * †
     
Pancreatic polypeptide (PPY) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 4.1 ± 0.002 * †
     
  Up-regulated by pioglitazone & palmitate vs. palmitate in non-diabetic human islets 4.7 ± 0.001 * †
     
  Up-regulated by palmitate vs. control in diabetic human islets 6.6 ± 0.001 * †
     
Regenerating islet-derived 3 alpha (REG3A) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 2.1 ± 0.002* †
  Up-regulated by pioglitazone & palmitate vs. palmitate in non-diabetic human islets 5.2 ± 0.001* †
     
  Down-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets -8.4 ± 0.001* †
     
  Up-regulated by palmitate vs. control in diabetic human islets 10.3 ± 0.002* †
     
Thioredoxin interacting protein  (TXNIP) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 12.6 ± 0.002 * †
     
  Up-regulated by pioglitazone & palmitate vs. palmitate in non-diabetic human islets 16.0 ± 0.002* †
     
  Down-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets -16.6 ± 0.002* †
     
Carboxyl ester lipase (CEL) Up-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets 6.7 ± 0.001 * †
     
  Up-regulated by exenatide & palmitate vs. palmitate in diabetic human islets 7.2 ± 0.001 *†
     
Aryl hydrocarbon receptor nuclear translocator (ARNT) Down -regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets -14.9 ± 0.001 * †
     
  Up-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets 14.7 ± 0.001 * †
     
  Up-regulated by exenatide & palmitate vs. palmitate in diabetic human islets 3.2 ± 0.002 *†
     
ISL1 transcription factor, LIM/homeodomain (ISL1) Down-regulated by exenatide & palmitate vs. palmitate in diabetic human islets -3.1 ± 0.001 *†
     
  Up-regulated by palmitate vs. control in diabetic human islets 2.3 ± 0.001* †
     
Transcription factor 4 (TCF4/TCF7L2) Down-regulated by pioglitazone & palmitate vs. palmitate in diabetic human islets -1.9 ± 0.002 * †
     
  Down-regulated by exenatide & palmitate vs. palmitate in diabetic human islets -2.0 ± 0.002 *†
     
  Up-regulated by palmitate vs. control in diabetic human islets 5.1 ± 0.002* †
     
Defender against cell death 1 (DAD1) Up-regulated by exenatide & palmitate vs. palmitate in non-diabetic human islets 7.9 ± 0.002 * †
     
  Up-regulated by pioglitazone & palmitate vs. palmitate in non-diabetic human islets 4.1 ± 0.002 * †
     
  Up-regulated by palmitate vs. control in diabetic human islets 2.1 ± 0.001* †

Table 5: Validation by qRT-PCR of differently regulated human islet transcripts identified by microarray analysis.

Discussion

The interaction between the genetic predisposition to β-cell failure with environmental or lifestyle factors appears to lead to diabetes pathogenesis. In the present study, we identify a subset of genes that seems to be involved in maladaptive metabolic alterations underlying the lipotoxic state leading to β-cell dysfunction and diabetes.

Pancreatic polypeptide (PPY) belongs to the NPY family and is expressed in the pancreatic islets of Langerhans. Moreover, PPY is released from the pancreas in response to a meal, specially by fat [24,25]. PPY decreases food intake and increases energy expenditure [26,27]. In this study, PPY expression was up-regulated in non-diabetic islets by pioglitazone and exenatide in the presence of palmitate (Table 4), but not by palmitate alone (epigenetic gene regulation pathway). This induction of PPY gene expression could influence pancreatic and gastric hormone release, and modulate different physiological processes. Interestingly, palmitate had no effect alone on PPY gene expression, possibly due to the absence of obesity, considering that obese subjects have lower serum levels of PPY [28,29]. Also, the PPY gene could be silenced by palmitate in non-diabetic islets. On the other hand, PPY was up-regulated by palmitate (Table 2) and this was reduced/normalized by pioglitazone in diabetic islets (Table 1). The linkage of the region containing PPY [30] and also genetic association of this gene with type 2 diabetes [31-33], suggest that PPY may be pathogenetically and therapeutically relevant in diabetes.

Transcription factor 7-like 2 (TCF7L2/TCF4), identified as the most important type 2 diabetes susceptibility gene to date [34], is involved in Wnt signaling induced β-cell proliferation [35,36]. The expression of TCF7L2/TCF4 transcript was induced by palmitate in diabetic islets in this study (Table 2), whereas in non-diabetic islets it remained unchanged. In addition, exenatide reduced the gene expression of TCF7L2/TCF4 in both diabetic (Table 1) and non-diabetic (Table 3) islets incubated with palmitate. Moreover, pioglitazone downregulated TCF7L2/TCF4 expression in the presence of palmitate in diabetic islets (Table 1). The up-regulated level of TCF7L2/TCF4 gene expression by palmitate noticed in our diabetic islets is in agreement with previous result where TCF7L2/TCF4 was overexpressed in type 2 diabetic human islets in response to a variety of stimulus [37]. Moreover, according to Lyssenko et al. overexpression of TCF7L2/ TCF4 in the β-cell reduces insulin secretion, suggesting that its upregulation by palmitate might be involved in β-cell dysfunction evoked by the fatty acid and perhaps also relevant in diabetic lipotoxicity in patients in whom β-cell dysfunction occur. The down-regulation of the palmitate-induced TCF7L2/TCF4 elevation by exenatide and pioglitazone observed in our human islets is also very interesting as it might contribute to the beneficial long-term influence by these drugs on β-cell function and viability.

Regenerating islet-derived 3 alpha (REG3A) is a pancreatic secretory protein that seems to be involved in β-cell proliferation and differentiation [38] via a cell surface Reg receptor [39]. In the current study, REG3A expression was up-regulated by palmitate in diabetic islets (Table 2). Taking into account this and also another study, where similar effect has been seen for the other members of the REG family in the presence of hyperglycemia [40], it may indicate the β-cells attempt to ameliorate lipoapoptosis through stimulation of β-cell proliferation. Furthermore, elevated expression of REG3A that results in islet neogeneis, growth, and/or protection has also been reported by others [38,41]. Both pioglitazone and exenatide induced REG3A expression in non-diabetic islets exposed to palmitate (Table 4), and reduced REG3A expression in diabetic islets exposed to palmitate (Table 1). Possibly, pioglitazone and exenatide contribute to maintenance of β-cell mass through induction of REG3A when islets become exposed to lipotoxicity and so signaling through the REG3A pathway might be one mechanism by which these drugs exert their beneficial influences on the β-cell.

Growth arrest and DNA-damage-inducible beta (GADD45B) is a member of the GADD45 nuclear protein family that is involved in β-cell growth arrest, DNA repair, and apoptosis [42]. In this study, the expression of GADD45B was slightly up-regulated by palmitate in diabetic islets (Table 2). A similar effect in response to the cytotoxic IL- 1β in INS-1E cells has recently been reported [43]. Both pioglitazone and exenatide reduced GADD45B expression in diabetic islets exposed to palmitate (Table 1). This may indicate that pioglitazone and exenatide, by normalizing the overexpression of GADD45B by palmitate, attempt to restore the balance between apoptosis and proliferation/survival in these islets. Considering that GADD45 family proteins are coactivators for RXR or PPAR activation [44], the modulation of GADD45B expression could influence nuclear hormone receptor activity and their involvment in fatty acid metabolism, apoptosis/cell survival, and insulin production/secretion.

Thioredoxin interacting protein (TXNIP) regulates the cellular redox state and mediates oxidative stress [45-49]. Our results show that TXNIP was up-regulated by palmitate in both diabetic (Table 2) and non-diabetic (Table 4) islets. Previous reports have shown TXNIP upregulation by glucose [50,51], and it appears now that FFAs impose similar effects in the β-cell. Considering that TXNIP overexpression induces apoptosis in β-cells in response to glucose [50], and impairs glucose-induced insulin secretion [52], this could also be the ultimate scenario for β-cells exposed to FFAs. These results underline the importance of TXNIP for β-cell survival and insulin secretion. Our results reveal that TXNIP expression was reduced by both pioglitazone and exenatide in diabetic islets exposed to palmitate (Table 1), whereas it was induced by these drugs in the presence of palmitate in nondiabetic islets (Table 4). It seems that both pioglitazone and exenatide were able to reduce and normalize the overexpression of TXNIP induced by palmitate in diabetic islets and it is conceivable that this mechanism might in part explain the positive impact of these drugs in terms of β-cell funtion and insulin secretion possibly due to existence of diabetes (a certain genetic background) and also hyperglycemia in diabetic subjects. This effect of exenatide has been reported previously in pancreatic islets [53].

Aryl hydrocarbon receptor nuclear translocator (ARNT), a.k.a. hypoxia-inducible factor (HIF), form heterodimeric complexes that directly regulate the expression of genes involved in glucose transport and glucose metabolism [54]. Moreover, the ARNT gene is located in a chromosomal region that shows strong linkage to type 2 diabetes [55]. Our present results show that ARNT gene expression was reduced by palmitate in diabetic islets (Table 1). ARNT expression has previously been reported to be significantly decreased in islets from type 2 patients, ARNT gene silencing in isolated β-cells was found to grossly impair glucose-stimulated insulin secretion, and ARNT knock-out animals showed severe glucose intolerance [56]. Our findings suggest that palmitate, which is in excess in diabetic patients’ serum due to their diabetic dyslipidemia, might be one factor that contributes to this ARNT down-regulation in diabetic islets. More importantly, ARNT down-regulation by palmitate was normalized by both pioglitazone and exenatide in our diabetic islets (Table 2). This may indicate one important molecular target for pioglitazone and exenatide in islets, by which these drugs maintain optimal insulin secretion and β-cell function.

Epidermal growth factor receptor (EGFR) is involved in cellular differentiation, proliferation, and survival in many cell types of epithelial origin through interactions with EGF [57-59]. Also, EGFR transactivation in GLP-1–induced β-cell proliferation has been reported [60]. The gene expression of EGFR in our study was slightly up-regulated by palmitate in diabetic islets (Table 2), whereas in nondiabetic islets it remained unchanged. This may reflect the β-cell’s attempt to diminish the apoptotic effect of palmitate in diabetic situations by stimulating the proliferation and β-cell survival trough induction of EGFR gene expression. On the other hand, the unchanged level of EGFR expression in non-diabetic islets could be due to the silencing effect of palmitate on the gene expression of EGFR. The expression of EGFR was reduced by pioglitazone in diabetic islets (Table 1), whereas it was induced by both pioglitazone and exenatide in non-diabetic islets (Table 4) exposed to palmitate. The latter may reflect the protective effects conferred by pioglitazone and exenatide in β–cells through maintaining the balance between their apoptosis and proliferation.

Carboxyl ester lipase (CEL), a lipolytical enzyme, highly expressed and secreted from pancreas, contributes to the digestion of dietary fat [61,62]. As exocrine pancreatic dysfunction is common in diabetes, considerable interest for CEL in islet biology arose when it was discovered that CEL mutations occur in a hereditary syndrome of diabetes and exocrine pancreatic dysfunction [63]. CEL mutations are overrepresented in MODY patients with β–cell failure (63). In the current study, CEL expression was not altered by palmitate in diabetic or non-diabetic islets, possibly due to silencing effect of palmitate. However, the CEL gene expression was up-regulated by pioglitazone and exenatide in both diabetic (Table 2) and non-diabetic (Table 4) islets exposed to palmitate. As studies of the role of CEL in islet biology are still in their infancy, it seems warranted to pursue this track in future efforts to determine whether the CEL upregulation observed in islets exposed to pioglitazone and exenatide is causally linked to protection by these drugs against β–cell dysfunction in lipotoxicity.

Acetyl-Coenzyme A carboxylase alpha (ACACA) regulates fatty acid biosynthesis [64,65]. In the current study, ACACA gene expression was down-regulated by palmitate in diabetic islets (Table 1), thus confirming results in the INS-l β-cell line [66]. This indicates stimulation of fatty acid oxidation to produce energy at the expense of lipid synthesis and contributing to impaired glucose-sensitive insulin secretion [66]. Previous studies have shown similar results when polyunsaturated fatty acids reduced the transcriptional rates of lipogenic enzymes [67]. On the other hand, studies on the INS-l β-cell line [68], and also rats fed with high-carbohydrate diet [67], have indicated glucose as a potent inducer of ACACA gene expression. Interestingly, both pioglitazone and exenatide up-regulated ACACA gene expression suppressed by palmitate in diabetic islets (Table 2). This indicates the normalizing effect by these drugs on palmitateinduced imbalance between fatty acid breakdown and synthesis by increasing lipogenesis, thereby probably positively impacting long term β-cell function and survival by ameliorating lipotoxicity and restoring glucose-sensitivity. Commensurate with such a role, previous findings using ACAC gene silencing have revealed an essential role for ACAC in nutrient-induced insulin secretion [69,70].

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a classical glycolytic enzyme involved in β-cell fuel sensing and energy production that yields high-energy phosphate molecules [71]. Moreover, GAPDH is also involved in several other pathways, including apoptosis by its translocation into the nucleus, protein phosphotransferase/kinase reactions, translational control of gene expression, DNA replication and repair [72-74]. In this study, GAPDH gene expression was upregulated by palmitate in diabetic islets (Table 4), whereas it remained unchanged in non-diabetic islets. This may reflect an attempt of the diabetic β-cell to sustain ATP production to endure lipotoxicity, whereas in non-diabetic islets GAPDH transcription seems to be silenced. Furthermore, the influence of lipotoxicity on transcriptional regulation of GAPDH gene expression appear to be different from the translational regulation, considering that palmitoyl-CoA has been reported to inhibit GAPDH enzyme activity [75]. Both pioglitazone and exenatide enhanced GAPDH gene expression in non-diabetic islets exposed to palmitate (Table 4). Possibly the GAPDH gene that was silenced by palmitate in these normal islets became activated by the drugs to cope with lipotoxicity, thus promoting β-cell survival by stimulating energy production. On the other hand, exenatide upregulated (Table 2) while pioglitazone down-regulated (Table 1) GAPDH gene expression in diabetic islets exposed to palmitate. It is difficult to speculate why these drugs act differently in this scenario, but one conceivable explanation could be that they might act through different pathways (as mentioned above) to cope with lipotoxicity and to protect the β-cell.

We noticed small differences between the diabetic and non-diabetic samples when looking at the genes involved in the inflammatory process, as previously suggested [76]. This could perhaps suggest the presence of a mild inflammation in islets from T2D patients compared to normal samples. In agreement with a previous study [77], in our study Chemokine (C-X-C motif) ligand 2 (CXCL2) gene expression was induced (10.7 fold) by palmitate in diabetic islets (Table 2). Whether the induction of the inflammatory genes in islets from T2D patients could be the result of in vivo exposure to saturated NEFA and whether this inflammation causes loss of functional β-cell mass and thereby T2D remains to be elucidated and needs to be further investigated.

In this study, the expression of a number of key genes altered by palmitate, pioglitazone and/or exenatide were classified into epigenetic regulation of gene expression, which also belong to major pathways involved in lipid metabolism. We identified epigenetic targets with known significance in β-cell biology that are up-regulated, downregulated or silenced by palmitate, pioglitazone and/or exenatide. The function and survival of the human β-cell seem to be regulated directly through epigenetic control of expression of genes involved in growth, proliferation, and differentiation rather than a direct effect on the apoptotic pathway. Since regulation of gene expression seems to be tightly under the epigenetic control and individually programmed, it might be different in different individuals and not totally be comparable to each other/generalized, which needs to be addressed in future investigations. FFAs seem to contribute and play an important role in the development of β-cell failure in synergy with hyperglycemia, in obese individuals, or in subjects who are genetically more predisposed to type 2 diabetes. Modern antidiabetic drugs such as pioglitazone and exenatide appear to normalize these epigenetic misregulations induced by lipotoxicity in the human β-cell. These identified β-cell genes may be harnessed to advantage in the development of novel diagnostic biomarkers or targeted therapeutics for type 2 diabetes based upon promotion of human β-cell function, survival, and proliferation.

Acknowledgements

Financial support was provided by the Amylin-Lilly Alliance, Takeda Pharmaceuticals North America, Stiftelsen Olle Engkvist Byggmästare, the Janne Elgqvist Family Foundation, the Swedish Society of Medicine, the Sigurd and Elsa Golje Memorial Foundation, Svenska Försäkringsföreningen, Svenska Diabetesstiftelsen, Berth von Kantzow’s Foundation, Trygg-Hansa´s Research Foundation, and the Research Center at Södersjukhuset, Stockholm.

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