Leptin对猪原代脂肪细胞的影响及脂滴包被蛋白介导的机制研究

V cells were separated from subcutaneous adipose tissue of weaned piglet. Cells were cultured to 80% confluence followed by differentiation for 3 days. The cells were treated with 10-8 M and 10-7 M leptin respectively for 4h (acute treatment) or 48h (chronic treatment). We used oil-red O and immunofluorescence histochemistry to identify adipocytes, lipid droplets and perilipin. Cultured media were collected for quantitation of glycerol content. Perilipin, HSL and ATGL mRNA levels were determined by Real-time RT-PCR. Activity of lipases (HSL and ATGL) was determined. Perilipin and phosphorylated perilipin protein levesls were quantitated by Western blot analysis. After Leptin acute treatment for 4h,the viability of cells was increased significantly in 10-7 M leptin treatment group; cells in both experimental groups released much more glycerol than in the control group; Perilipin, HSL and ATGL mRNA expression were significantly increased by 10-7 M leptin treatment, whereas10-8 M Leptin treatment increased the mRNA expression of ATGL only; there was no alteration of lipase activity and perilipin content, and the phosphorylation of perilipin wasn't detected. After Leptin chronic treatment for 48h, Perilipin mRNA was down-regulated by 10-8 M leptin;10-7 M leptin treatment could significantly down-regulate the expression of perilipin mRNA whereas up-regulate ATGL mRNA expression;importantly the phosphorylated perilipin were higher than control group in both treatement groups. The results indicate that acute treatment leptin may influence mRNA expression of related genes, and chronic leptin treatment activated phosphorylation of perilipin and increased lipolytic activity of mature adipocytes.