| To explore whether the immunity enhancing effect of Arg is attributable to promotion of IGF-1 secretion by liver. Serum-free culture, MTT , FCM, Hoechst 33258 fluorescence label, ELISA, RIA, RT-PCR, Western blotting etc were adopted. Three aspects were studied including: 1. Establishment of primary cultured rat hepatocytes in serum-free medium to explore the regulation of Arg on IGF-1 production; 2. Effects of Arg on IGF-1 mRNA expression, IGF-1 secretion and cell immunity of primary cultured rat hepatocytes in serum-free medium; 3. Effects of Arg on IGF-1 secretion, expression and immunity of injured rats.No significant difference was observed in survival rates, apoptotic rates and DNA contents of the hepatocytes treated by different concentrations of Arg (0 or 37,500umol/L). The above results provided a preliminary idea for further studies about the regulation of Arg on IGF-1 secretion by hepatocytes.Treatment with Arg (0-7,500umol/L) for 48h led to succeeding increases in IGF-1 secretion by hepatocytes; The changes in IGF-1 secretion showed the same tendency as that of IGF-1 mRNA, but no IGF-1R mRNA expression was detected by RT-PCR in hepatocytes , which probably explained why Arg did not exert the effect in liver. The IGF-1 secreted by hepatocyte under Arg treatment did not act on hepatocyte per se, but other types of cells.Arg (0-7,500umol/L)-treated hepatocyte supernatant promoted T cell proliferation in dose-dependent manner; Lymphocytes co-cultured with hepatocyte supernatant (750umol/L, 48h) led to the induction of IL-2 and the instant opening of lymphocytes membrane Ca2+ channel in the early phase of lymphocyte activation; Hepatocyte supernatant treatment also increased the ability of NK cells to kill K562 cells. Hepatic IGF-1 mRNA levels, IGF-1 expression and serum IGF-1 levels were all decreased in injured rats(IC), but these indices significantly increased after Arg-supplement (IA). Spleen index of IA rats was significantly higher than that of normal control (NC) group and activities of thymus T cells were significantly increased. The abundance of BMNC IGF-1R mRNA in IA group was higher than that of IC. Scrum Arg and ornithinc levels of IA rats were significantly higher than those of NC rats while serum NO levels of injured rats (IC and IA) were significantly lower thanthose of NC rats, serum GH and cortisol levels showed no difference among the groups. Arg regulated IGF-1 production in a dose-dependent and GH-independent manner; and both in cultured hepatocytes and in injured rats, the regulation of Arg on IGF-1 was observed at mRNA and protein expression levels. Arg may exert the immune-enhancing effect from stimulating IGF-1 production by the liver. |
