Enterostatin alters protein trafficking to inhibit insulin secretion in Beta-TC6 cells

Abstract

Enterostatin is a peptide that regulates dietary fat intake in rodents and inhibits insulin secretion from pancreatic beta cells. Microarray studies of the genomic response of both a human hepatoma cell line (HepG2 cells) and a mouse hypothalamic cell line (GT1-7 cells) to enterostatin suggested that it might regulate protein trafficking. Using semi quantitative real-time PCR and western blot analysis, we confirmed that enterostatin upregulated Scamp2 and down regulated Dynamin2 in these cell lines. The receptor for enterostatin is the F 1 -ATPase beta subunit. We transfected HepG2 cells with either a Green Fluorescent protein (GFP) tagged F 1 -ATPase beta subunit or a Red Fluorescent protein (RFP) tagged F 1 -ATPase alpha subunit to study the effects of enterostatin on translocation of its own receptor protein. Enterostatin induced movement of GFP-beta subunit to the cell periphery area but did not have any effect on the localization of RFP-alpha subunit protein in HepG2. As Scamp2 is involved in glucose uptake in mouse Beta-TC6 insulinoma cells we tested enterostatin's effect in Beta-TC6 cells. Glucose stimulated insulin release was inhibited by enterostatin as reported previously. Using siRNA to Scamp2 did not change glucose stimulated insulin release but siRNA to Dynamin2 and dominant negative Dynamin2 (Dyn K44A) inhibited glucose stimulated insulin release and abolished the response to enterostatin. This suggests enterostatin inhibits glucose stimulated insulin release in pancreatic beta cells through down regulation of Dynamin2. This study also suggests that enterostatin might have a more generalized effect on protein trafficking in various cells.

Salivary extracellular RNA biomarkers for insulin resistance detection in hispanics

“…For example, IL1R2 was found to be positively correlated with reduced insulin secretion and higher HbA1c levels[38]; VPS4B was up-regulated in response to the inhibition of glucose stimulated insulin secretion[39]; and CAP1 expression level was associated with insulin sensitivity[40]. These studies corroborate our findings and strengthen the premise that our panel could be used in the clinical detection of IR.…”

Anthocyanin inhibits high glucose-induced hepatic mtGPAT1 activation and prevents fatty acid synthesis through PKCζ

“…HepG2 cells were stimulated with high glucose levels (25 mM, HG) in the absence or presence of 1, 10, or 100 M Cy-3-g for 24 h. Normal glucose (5.5 mM) was used as a control (Ctr). A: Cellular subfractions containing the endoplasmic reticulum (ER) and outer mitochondrial membrane (OMM) were isolated and GPAT1 expression was measured by by guest, on www.jlr.org An F1-ATPase ␤ -subunit clone was generated by PCR using an F1-ATPase ␤ -subunit forward primer (5 ′ -GAGAGGAGCT CC-ACTATTGCTATGGATGGC-3 ′ , SacI recognition site underlined) and an F1-ATPase ␤ -subunit reverse primer (5 ′ -GAGAGA AG-CTTCACGACCCATGCTC-3 ′ , Hin dIII recognition site underlined) as described ( 16 ). The PCR product was cloned into the pCMV5 vector (Clontech, Palo Alto, CA) between the SacI and Hin dIII sites and then transformed into DH5 ␣ cells.…”

Hypothalamic KLF4 mediates leptin's effects on food intake via AgRP

“…The transcription factor KLF4 is one of the most recently discovered activators of AgRP. In vitro studies involving over-expression or silencing of KLF4 have shown that KLF4 is required for activation of AgRP [8,39] . In vivo , a compound named PMI-5011 has also been found to activate both KLF4 and AgRP expression; however, PMI-5011 also activated hypothalamic orexin and melanin-concentrating hormone, stimulating food intake by that route [8] .…”