Free Fatty Acid On People Proximal Convoluted Tubules Epithelial Cells Of The Influence And Function Mechanism

Objective:For the study of kidney damage lipid toxic effects, we use free fatty acid (Free Fatty Acids,FFAs) intervention people proximal convoluted tubules epithelial cells (HK - 2), explore free fatty acid to proximal convoluted tubules epithelial cells of the influence and function mechanism.Methods:People proximal convoluted tubules epithelial cells(HK - 2), were cultured with DMEM-F12 of different concentrations of palmitic acid (0,125,250,500μmol/L) (using defat bovine serum albumin as vector), normal medium as control grop. on the 0 hour,24 hour,48 hour respectively:①Morphological observation;②Oil red O dyeing detection cell lipid;③The MTT method to detect different environment for HK - 2 FFAs cell proliferation influence;④Flow cytometric FFAs intraoperative detection of apoptosis and cell cycle impact;⑤Immune cell chemical method before and after treatment were detected HK - 2 cells iNOS of nitric oxide synthase (iNOS), apoptosis related proteins caspase8 protein, and interleukin (IL-1β), interleukin (IL - 8) expression changes.Results:①HK - 2 cells form Normal HK - 2 cells assumes the circular posted parietal cells, strewn with cell fusion after a pebble, swelling. The FFAs working, HK - 2 cells by circular into spun spindle cells, the connections between looser.②Oil red O dyeing detection cell lipid deposition Palmitic acid intervention group, 48h, HK - 2 cell cytoplasm appeared in very clear red dye lipid drops particles. The control HK - 2 cells not seen red dye lipid drops particles.③The MTT detection HK - 2 cell proliferation OD value with the increase of palmitic acid concentration and role of prolonged decrease gradually. Three different dose group of binary comparison between are statistically significant (P < 0.05).The same concentration, 24 h 48h between groups compared with high, middle dose group of statistically significant (P < 0.05), low dose group was not statistically significant (P > 0.05) explain palmitic acid inhibit HK - 2 cell proliferation, and has the dose and time dependence trend. Each palmitic acid group compared with controls have significant difference (P < 0.01), d - BSA group and normal nutrient-containing medium group compared was not statistically significant (P > 0.05). ④Flow cytometric intraoperative determination result FFAs role 48h adopted when double marking method to detect cell apoptosis,HK - 2 apoptosis rate in order 5.86±1.39,9.61±0.82,12.3±1.19,With normal control group 2.23±0.92, d - BSA group 2.64±1.02 compared with significant difference (P < 0.05).PI ChanBiao detection cell cycle changes, compared with the control group cell group G0 / G1 ratio increases, S period with the control group rate by significant difference (P < 0.05).⑤Cell immunochemical test results In normal controls,d-BSA control 48h, training, iNOS, caspase8, IL-1β, IL-8 hardly express, between two control group was not statistically significant (P > 0.05). Different concentrations of FFAs processing HK - 2 cell, iNOS, caspase8, IL-1β, IL-8 expression, and has a significantly higher concentration dependence trend, and normal controls and d - BSA rcpmaj, iNOS, IL - 1β, IL-8, in the high-dosage groups was statistically significant difference were, caspase8 each dose group of differences are statistically significant (P < 0.05) Conclusion:FFAs can cause renal tubular epithelial cells of lipid deposition, inhibit renal tubular epithelial cell proliferation, possibly by blocking the cell cycle and promote its apoptosis role, and with time concentration dependence. This effect may be with FFAs through activating HK - 2 within the iNOS, cause excessive NO relevant, caspase8 participated in its occurrence and development process, simultaneously the inflammatory cytokines IL - 1β, IL - 8 express increased, and jointly promote the renal tubular epithelial cell apoptosis.