Introduction. Microbial infections are suspected to take part in the pathogenesis of diabetes mellitus type 1 (T1DM). Glucose stimulation of cells is accompained by release of free arachidonic acid (AA) mainly by the action of cytosolic and calcium independent phospholipases A2 (cPLA2 and iPLA2). E. coli infection also activates AA release: the exact relationship between E. coli infection and insulin secretion has been a matter of speculation. Materials and Methods. Insulinoma cell line (INS-1E) was infected with enterohemorrhagic E. coli (EHEC). Invasion assay, Scanning Electron Microscopy and Transmission Electron Microscopy were used to demonstrate the capacity of EHEC to enter cells. Glucose-induced insulin secretion was evaluated after acute (6h) and chronic (72h) infection and after transfection of INS-1E with specific cPLA2- or iPLA2-siRNAs. PLA2s activities in presence of specific inhibitors and Western blot analysis were performed. Results. After acute infection, the insulin release was very similar to control cells but after chronic infection the insulin release significantly decreased. PLA2s activities were significantly activated and the presence of EDTA (cPLA2 inhibitor) or BEL (iPLA2 inhibitor) demonstrated that cPLA2 activity is mainly responsible for the AA production. Western blot analysis showed an increase of cPLA2 , iPLA2, phospho-cPLA2, and COX-2 expressions after acute and, even more, after chronic E. coli infection. After acute infection, the silencing of the two isoforms of PLA2s, with specific cPLA2- or iPLA2-siRNAs, reduced the insulin secretion. After chronic infection, the silencing of the PLA2s determined a rise in the insulin release. Discussion and conclusions. The results obtained demonstrated that cPLA2 and iPLA2 play a key role in the process of insulin secretion after EHEC infection, probably because a high enzymatic activation leads to the release of AA. This polyunsaturated fatty acid per se or its metabolites, since it is the substrate of cyclooxygenase, which could produce a high amount of prostaglandins, could be responsible for the reduced secretion of insulin. The understanding of the molecular mechanisms activated by the bacterial infection is a necessary requirement in order to develop new therapeutic approaches in patients suffering from an imbalance in the secretion of insulin in response to bacterial infection.
|Titolo:||E. coli infection impairs insulin secretion through a PLA2-dependent mechanism|
|Data di pubblicazione:||2015|
|Appare nelle tipologie:||4.2 Abstract in Atti di convegno|