Research Of The Regulatory Effect Of Leptin On Apoptosis Of Thymocytes Induced By LPS And Mechanism Of Anti-apoptosis

Objective: To observe the effect of leptin on apoptosis of thymocytes. To explore weather phosphatidylinositol 3-kinase/AKT (PI3-K/AKT) signal pathway and cPLA2 signal pathway are involved in anti-apoptotic action, and mechanism of anti-apoptosis of leptin.Methods: 1. Apoptosis was induced by lipopolysaccharide (LPS) in mice. The toxicity of LPS on thymocytes was evaluated using MTT colorimetric assay. Thymocytes apoptosis was detected using Annexin V-F ITC /PI double staining flow cytometry.2. Caspase3 mRNA was measured by RT-PCR.3. The effects of a specific inhibitor an inhibitor (AACOCF3) of cPLA2 and (LY294002) of PI3-K on apoptosis of thymocytes was detected using Annexin V-FITC /PI double staining flow cytometry.4. cPLA2 mRNA were measured by RT-PCR. cPLA2 activity was detected by cPLA2 Assay Kit.Results: 1. Leptin decreases the toxicity of LPS on thymocytes in a dose-dependent manner; Leptin restores the percentage of LPS-inhibited apoptotic cells determined by FACS after annexin V binding and PI staining.2. The mRNA expressions of caspase3 were inhibited by leptin in a dose-dependent manner. These demonstrate that the leptin directly inhibits cytotoxic effect and apoptosis in thymocytes, which is very likely derived from a caspase-dependent apoptosis.3. LPS increased the percentage of apoptotic cells from 12.58 % ( control) to 42.20%. But AACOCF3 (10μM) restores LPS-inhibited thymic apoptosis, from 42.20% to 37.89%. At the same time, the expressions of cPLA2mRNA were inhibited by leptin,and cPLA2 activity was measured by cPLA2 assay kit, which suggested that AACOCF3 depressed cPLA2 activity. These data clearly demonstrate that cPLA2 signaling pathways participates partly LPS-induced apoptosis in thymus.4. leptin protected apoptosis of thymocytes induced by LPS in mice, thymocytes viability was elevated from 57.80% to 64.09 % (P<0.05). LY294002 (10μmol/L) reversed the protection of leptin, thymocytes viability decreased respectively from 64.09% to 60.33%, which leptin activates the PI3-K/AKT signal transduction pathway in anti-apoptotic action.Conclusions:1. leptin is able to decrease the apoptosis of thymocytes induced by LPS.2. leptin inhibits apoptosis in thymus through cPLA2 and PI3-K/AKT signal pathway.3. caspase3 is involved in mechanism of anti-apoptosis of leptin.