Survival of rat pancreatic islets is partly controlled by a TCF7L2-p53-p53INP1 dependent pathway

• Background and aims: TCF7L2 is both an activator and an inhibitor of transcription and the most highly associated type 2 diabetes gene known to date. It influences beta cell survival and function, i.e. incretin hormonal effects, insulin processing and secretion. However, its target genes in pancreatic islets are not fully described and the molecular mechanism whereby it propagates its effects on islet function is not known. The aim of this study is to identify the molecular mechanisms through which TCF7L2 influence beta cell survival and function. Materials and methods: Wister rat primary islets and INS-1 (832/13) cells were incubated with siRNA against Tcf7l2, both Tcf7l2 and TP53INP1 or both TCF7L2 and TP53 in 5.5 mM and 14.3 mM glucose. TCF7L2 activity, p53 activity and target gene expression (using qPCR) were measured after siRNA treatment. INS-1 cell apoptosis was measured by DNA degradation levels, caspase-3/7 levels and by using antibodies against Annexin V, and 7-AAD, visualized using confocal microscopy. Rat islet viability was estimated measuring metabolic rate. Rat islet apoptosis was estimated by measuring Caspase-3/7 level. Results: The type 2 diabetes associated genes TP53INP1, FTO, GIPR and ADAMTS9 were identified as TCF7L2 potential target gene using chromatin immunoprecipitation on microarrays. In INS-1 cells, siRNA mediated Tcf7l2 knock down (69.5 %) resulted in decreased TCF7L2 activity (91%) and differential expression of the target genes: Tp53 (14.5% increase), TP53INP1 (65.9% increase) and ADAMTS9 (82.8% decrease). TCF7L2 knockdown also lead to reduced cell viability (65%) and increased apoptosis (113%). The TCF7L2 induced cell death was replicated in rat primary islets. When restoring (decreasing) the Tp53inp1 expression level in TCF7L2 depleted islets, the decrease in cell viability and increase in apoptosis were prevented, suggesting that the Tcf7l2 effect is mediated via Tp53inp1. Furthermore, p53 depletion prohibited TCF7L2 down regulation induced cell death and elevation of Tp53inp1 expression in both INS-1 cells and rat primary islets. Conclusion: The type 2 diabetes associated genes TP53INP1 and ADAMTS9 are target genes of TCF7L2 in pancreatic islets. TCF7L2 induced apoptosis and decreased cell viability are mediated through activation of p53 and increased p53INP1 expression.

Subject headings

MEDICIN OCH HÄLSOVETENSKAP  -- Klinisk medicin -- Endokrinologi och diabetes (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Clinical Medicine -- Endocrinology and Diabetes (hsv//eng)

Keyword

protein p53

small interfering RNA

caspase

DNA

antibody

lipocortin 5

incretin

insulin

glucose

diabetes mellitus

rat

pancreas islet

survival

gene

apoptosis

cell viability

cell survival

cell death

DNA degradation

processing

confocal microscopy

metabolic rate

chromatin immunoprecipitation

secretion (process)

gene expression

down regulation

Publication and Content Type

art (subject category)

ref (subject category)