Effects Of L-carnitine On Proliferation, Apoptosis And Fibronectin Secretion Of Human Peritoneal Mesothelial Cells On The Condition Of High Glucose

IntroductionPeritoneal dialysis is an essential component of renal replacement therapies. The structural and functional integrity of the mesothelium is the base of permeability and molecule transport of the peritoneal membrane. Recent research showed the continuous exposure to the nonphysiologic high glucose could inhibit proliferation, induce apoptosis and increase FN secretion of human peritoneal mesothelial cells ( HPMCs) , was one of the reasons of PD related peritoneal sclerosis, and would compromise peritoneal membrane performance, possibly led in turn to PD failure. Carnitine presents in various tissues and cells. It plays a role in carrying long chain fatty acids across mitochondial membrane to accelerate tricarboxylic acid cycle and fulfill the normal physiological function and energy metabolism in cell. Carnitine deficiency has been associated with synthesis decrease and low nutritional intake in uremic patients. In our study we try to investigate the role of L - carnitine on proliferation, apoptosis and fibrone-ctin secretion of HPMCs on the condition of high glucose in order to know if we use carnitine in conventional peritoneal dialysis fluids (PDF) the clinical symptoms that have been associated with this disorder, inhibition of HPMCs proliferation and apoptosis caused by high glucose in PDFmight be improved.Materials and Methods1. Materials1. 1 Human greater omenta of nonuremic patients.1. 2 Reagents for cell culture and identify.1. 3 L - carnitine.1. 4 Reagents for MTT colorimetric assay.1. 5 Reagents for Annexin V -FITC/PI double labeled assay.1. 6 Reagents for ELISA.2. MethodsEnzymatic disaggregation was used for primary culture and passage. HPMCs were cultured in vitro and different concentrations of glucose and L -carnitine were added in experimental groups. Cell proliferation was evaluated by MTT colorimetric assay, apoptosis was analyzed by Annexin V - FITC/PI double labeled assay, concentrations of fibronectin in culture medium were assessed by ELISA . All data appear as x ± s. SPSS12. 0 software was employed to analyze the variance with oneway ANOVA between the different groups, the difference is significant when P <0. 05.ResultsIncubation with glucose in different concentrations for 24 hours the proliferation effect was observed in 1.5% glucose group (P <0. 05) , 2. 5% glucose group and 4. 25% glucose group( P <0. 01) . HPMC proliferation was increased in 1.5% and 2.5% glucose group(P <0. 05) ,and inhibited in A. 25% glucose group( P <0. 01) at 72 hours. Incubation with L - carnitine in different concentrations for 24 hours the proliferation effect was observed in each L - carnitine group ( P < 0. 01) . HPMC proliferation was inhibited at 72 hours in 0. 5% ( P < 0. 05 ) and 1. 0% L - carnitine group( P <0. 01) . HPMC apoptosis was inhibited in 0. 05% and 0. 1% L - carnitine group at 72 hours. The concentration of Fibronectin in 0. 05% L - camitine group was increased at 24 hours, in each L -carnitine group the concentration of FN was decreased at 72 hours( P <0. 01).DiscussionRecent research showed the continuous exposure to the nonphysiologic high glucose could induce HPMC to express cell cycle inhibitate protein and lead to proliferation stasis in Gj period, finally consenescence and apoptosis. A study by Weibin S et al approved that high glucose promoted HPMC decease mainly by inducing cell apoptosis. High glucose can increase FN secretion of HPMC found in some research, suggesting increasing in extracellular matrix, might be associated with peritoneal sclerosis and decline in dialysis efficacy. In humans, carni-tine is synthesized in liver, kidney and brain from the essential amino acids ly-sine and methionine. Only liver and kidney release newly synthesized carnitine into systemic circulation. The main physiological function of carnitine is the transport of long — chain fatty acids to the sites of p - oxidation in the mitochon-drial matrix involving several enzymes. L - carnitine deficiency leads to the accumulation of free fatty acids in cell cytoplasm and of acylCoA in the mitochondria, producing a toxic effect on the cell. E. GAGGIOTTI et al compared in vitro and in the rabbit a new peritoneal dialysis solution containing carnitine and found the solution seems to be more biocompatible than standard glucose solutions . They also found phospholipid secretion of mesothelial cells was much higher in test group . This is an important index of cell function, as phospholip-ids are essential for peritoneal physiology. Guangxu Z et al found L - carnitine decreased the ratios of DNA fragments. A study by Yong Y et al suggested L -carnitine prevented apoptosis through its action on interfering with oxygen free radical formation, promoting antioxidant properties of cells and inhibiting apoptosis mediated by oxygen free radical. Our results indicated that high glucose could promote proliferation of HPMC in short period, and inhibit it after long period exposure;L - carnitine on certain concentration could promote proliferation and FN secretion, inhibit apoptosis of HPMC on the condition of high glucose;L - carnitine on high concentration and in long — term exposure might be inhibit proliferation of HPMC.ConclusionHigh glucose can promote proliferation of HPMC in short period, and inhibit it after long period exposure. L - carnitine on certain concentration can promote proliferation and FN secretion, inhibit apoptosis of HPMC on the condition of high glucose. L - carnitine on high concentration and in long - term exposure maybe inhibit proliferation of HPMC.