Cytokine-mediated induction of anti-apoptotic genes that are lined to nuclear factor-κB (NF-κB) signalling in human islets and in a mouse β cell line

Barbara Davis Center for Childhood Diabetes, University of Colorado Denver, Aurora, CO 80045, USA.
Diabetologia (Impact Factor: 6.67). 05/2009; 52(6):1092-101. DOI: 10.1007/s00125-009-1331-x
Source: PubMed

ABSTRACT The destruction of pancreatic beta cells leading to type 1 diabetes in humans is thought to occur mainly through apoptosis and necrosis induced by activated macrophages and T cells, and in which secreted cytokines play a significant role. The transcription factor nuclear factor kappa-B (NF-kappaB) plays an important role in mediating the apoptotic action of cytokines in beta cells. We therefore sought to determine the changes in expression of genes modulated by NF-kappaB in human islets exposed to a combination of IL1beta, TNF-alpha and IFN-gamma.
Microarray and gene set enrichment analysis were performed to investigate the global response of gene expression and pathways modulated in cultured human islets exposed to cytokines. Validation of a panel of NF-kappaB-regulated genes was performed by quantitative RT-PCR. The mechanism of induction of BIRC3 by cytokines was examined by transient transfection of BIRC3 promoter constructs linked to a luciferase gene in MIN6 cells, a mouse beta cell line.
Enrichment of several metabolic and signalling pathways was observed in cytokine-treated human islets. In addition to the upregulation of known pro-apoptotic genes, a number of anti-apoptotic genes including BIRC3, BCL2A1, TNFAIP3, CFLAR and TRAF1 were induced by cytokines through NF-kappaB. Significant synergy between the cytokines was observed in NF-kappaB-mediated induction of the promoter of BIRC3 in MIN6 cells.
These findings suggest that, via NF-kappaB activation, cytokines induce a concurrent anti-apoptotic pathway that may be critical for preserving islet integrity and viability during the progression of insulitis in type 1 diabetes.

Download full-text


Available from: Subbiah Pugazhenthi, Mar 12, 2014
68 Reads
  • Source
    • "Baculoviral inhibitor of apoptosis repeat-containing 2 (BIRC2), another well-documented downstream regulator of the TNF-α signaling network, has also been reported to moderate the activity of NF-κB, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase, thus providing feed-forward regulation of the TNF-α signaling pathway (Tan et al., 2013). Extensive studies have also shown that BIRC2 has roles in many processes, including caspase regulation in apoptosis, inflammatory and mitogen kinase signaling , immunity, cell proliferation, cell invasion and metastasis (Gyrd-Hansen and Meier, 2010; Sarkar et al., 2009). In the shrimp Penaeus monodon, the inhibitor of apoptosis proteins (IAPs), homologs of BIRC2, was demonstrated to block apoptosis induced by the Drosophila Reaper protein (Leu et al., 2008). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The TNF-α signaling cascade is involved in the regulation of a variety of biological processes, including cell proliferation, differentiation, apoptosis and the immune response in vertebrates. Here, two regulatory genes, lipopolysaccharide-induced tumor necrosis factor α factor (LITAF) and baculoviral inhibitor of apoptosis repeat-containing 2 (BIRC2), were identified in coelomocytes from the sea cucumber Apostichopus japonicus by RNA-seq and RACE (denoted as AjLITAF and AjBIRC2, respectively). The full-length cDNA of AjLITAF was 1417 bp, with a 5' untranslated region (UTR) of 189 bp, a 3' UTR of 637 bp with one cytokine RNA instability motif (ATTTA) and an open reading frame (ORF) of 591 bp encoding a polypeptide of 196 amino acid residues and a predicted molecular weight of 22.1 kDa. The partial AjBIRC2 cDNA was 2324 bp with a 5' UTR of 145 bp, a 3' UTR of 469 bp and a complete ORF of 1710 bp encoding a polypeptide of 569 amino acid residues. Analysis of the deduced amino acid sequences revealed that both genes shared a remarkably high degree of structural conservation with their mammalian orthologs, including a highly conserved LITAF domain in AjLITAF and three types of BIR domains in AjBIRC2. Spatial expression analysis revealed that AjLITAF and AjBIRC2 were expressed at a slightly lower level in the intestine and tentacle tissues compared with the other four tissues examined. After challenging the sea cucumbers with Vibrio splendidus, the expression levels of AjLITAF and AjBIRC2 in coelomocytes were increased by 2.65-fold at 6 h and 1.76-fold at 24 h compared with the control group. In primary cultured coelomocytes, a significant increase in the expression of AjLITAF and AjBIRC2 was detected after 6 h of exposure to 1 µg mL(-1) LPS. Together, these results suggest that AjLITAF and AjBIRC2 might be involved in the sea cucumber immune response during the course of a pathogenic infection or exposure to pathogen-associated molecular pattern (PAMP) molecules.
    Developmental & Comparative Immunology 10/2014; 48(1). DOI:10.1016/j.dci.2014.10.001 · 2.82 Impact Factor
  • Source
    • "diabetes (Kharroubi et al. 2004; Sarkar et al. 2009) and its relationship to LT-a, we examined the association of LT-a ?249A/G with diabetes among Tunisian subjects. Our data show that the LT-a ?249A/G "
    [Show abstract] [Hide abstract]
    ABSTRACT: We investigated the association of the lymphotoxin (LT)-α gene polymorphism +249A/G with type 1 diabetes. The distribution of genotypes of the LT-α +249A/G single nucleotide polymorphism (SNP) was assessed in 115 diabetic patients and 123 normoglycemic control subjects, using PCR-restriction fragment length polymorphism analysis. Among unselected patients, the SNP was significantly associated with increased risk of diabetes (χ(2) = 8.44, p = 0.014) and was found to be more pronounced among female (χ(2) = 8.37, p = 0.02) than male (χ(2) = 6.11, p = 0.047) patients. A significant association was detected between LT-α +249A/G and increased risk of diabetes, in particular for young-onset patients (χ(2) = 6.92, p = 0.031). Moreover, we reported significant differences in levels of HbA1c, triglycerides, alanine transaminase, and anti-glutamic acid decarboxylase-65 among alleles. Additional studies with extended patient age groups and different ethnicities are needed to confirm our findings.
    Biochemical Genetics 11/2013; 52(1). DOI:10.1007/s10528-013-9629-2 · 0.87 Impact Factor
  • Source
    • "Among various inflammatory cytokines, IL-1β is the major inflammatory cytokine whose expression is upregulated during islet isolation procedure, culture and upon exposure to hyperglycemic conditions [24]–[26]. Human islets exposed to IL-1β undergo NFkB activation, Fas upregulation and DNA fragmentation, eventually leading to β-cell death [27], [28]. To prevent IL-1β induced β-cell apoptosis, human islets have been genetically modified to overexpress hIL-1Ra locally [5]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The objective of this study was to determine the potential of human bone marrow derived mesenchymal stem cells (hBMSCs) as gene carriers for improving the outcome of human islet transplantation. hBMSCs were characterized for the expression of phenotypic markers and transduced with Adv-hVEGF-hIL-1Ra to overexpress human vascular endothelial growth factor (hVEGF) and human interleukin-1 receptor antagonist (hIL-1Ra). Human islets were co-cultured with hBMSCs overexpressing hVEGF and hIL-1Ra. Islet viability was determined by membrane fluorescent method and glucose stimulation test. Transduced hBMSCs and human islets were co-transplanted under the kidney capsule of NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl) /SzJ (NSG) diabetic mice and blood glucose levels were measured over time to demonstrate the efficacy of genetically modified hBMSCs. At the end of study, immunofluorescent staining of kidney section bearing islets was performed for insulin and von Willebrand Factor (vWF). hBMSCs were positive for the expression of CD73, CD90, CD105, CD146 and Stro-1 surface markers as determined by flow cytometry. Transduction of hBMSCs with adenovirus did not affect their stemness and differentiation potential as confirmed by mRNA levels of stem cell markers and adipogenic differentiation of transduced hBMSCs. hBMSCs were efficiently transduced with Adv-hVEGF-hIL-1Ra to overexpress hVEGF and hIL-1Ra. Live dead cell staining and glucose stimulation test have shown that transduced hBMSCs improved the viability of islets against cytokine cocktail. Co-transplantation of human islets with genetically modified hBMSCs improved the glycemic control of diabetic NSG mice as determined by mean blood glucose levels and intraperitoneal glucose tolerance test. Immunofluorescent staining of kidney sections was positive for human insulin and vWF. In conclusion, our results have demonstrated that hBMSCs may be used as gene carriers and nursing cells to improve the outcome of islet transplantation.
    PLoS ONE 10/2013; 8(10):e77591. DOI:10.1371/journal.pone.0077591 · 3.23 Impact Factor
Show more