Effects Of Free Fatty Acids On Isletβcells Line NIT-1 Apoptosis And Secretion.

Objective: To explore the effects of free fatty acids on islet B cells lines (NIT-1 cells) apoptosis and secretion. Analyse the mechanism of different FFA concentration damaged to beta cells. To provide treatment of reference for diabetics merge with metabolic disturbances.Methods: The NIT-1 cells in this study were grown in RPMI 1640 medium with 10% calf bovineserum, 100u/ml penicillin,100mg/L streptomycin,which were put in CO2 incubator for static culture at 37℃. Cell morphology and growth pattern were observed under reverse microscope and cell proliferation was assayed using MTT method to draw the growth curve,divided into single-agent group and FFA group(0.25,0.5,1.0mmol/L). NIT cells were treated with different concentrations of FFA (0.25, 0.5, and 1.0 mmol/L) for 24,48, and 72 h. and cells proliferation was detected by MTT assay. the cells insulin level was evaluated by radioimmno assay, cells apoptosis was examined by Hoechst33342 assay, SPSS 13.0 software was adopted to carry out the data processing and analyses, results with analysis of variance,two-sample t test and paired- samples t test was used for statistical analysis and make 0. 05 the boundary of statistical significance.Results:①There were no difference between single-agent group and FFA 0.25 group, however cell proliferation was attenuated significantly in other group after 24hours later by MTT assay ( P <0.05). The cell proliferation of all FFA groups were attenuated significantly after 48hours and 72hours later and apoptotic rate was significantly increased.②The NIT-1 cells insulin secretion effect, the insulin levels in all group of supernatant was tested at 24hours 48hours and 72hours respectively after the treatment. With concentration and the role of the time increased, insulin levels was significantly reduced,even completely not exist.③Under the fluorescent microscope, the cell nucleolus by Hoechst33342 assay exhibited round or oval shape and the border was clear in single-agent group.The typical morphological changes of FFA group detected by fluorescence microscope included chromatin condensation and aggregation at the periphery of the nucleons and nuclear fragmentation.Conclusion:①The MTT assay indicated that the percentage of apoptic cells of beta NIT-1 expression were increased with the increasing of exposure time and concentration of FFA.Abnomal FFA concentration not conducive to the growth of beta NIT-1 cells.②The insulin levels were reduced with the increasing of exposure time and concentration of FFA.③Abnomal FFA concentration could increase cell apoptosis and reduced the NIT-1 cells insulin secretion. Consequently FFAs were observed to be related with the defects of pancreatic beta cell, lowered level of blood fats (cholesterol and triglycerides)may be useful in identifying outpatients at increased risk of diabetes mellitus.Significance: Recently the harmful of lipotoxicity for T2DM more and more respected.This study indicated that abnormal levels of FFA is not conducive to the survival of isletβNIT-1 cells ,which can induce NIT-1cells apoptosis and reduce insulin secretion. It is very important to maintain beta cells normal insulin secretion and plenty of beta cells, quantity and improving the insulin resistance for T2DM. Body has more ability to maintain a stable blood sugar levels to reduce the chances of complications in patients with diabetes mellitus. To solve the beta cells impairment offer a new rationale to treat diabetes mellitus for us. Facilitate cell regeneration and transplantation for the patients who almost lose function of Insulin secretion could be cure T2DM.