| Ojective:To investigate the effects of Ling Qi Juan Gan Capsule drug-containing serum at different concentrations on the proliferation and the expression of janus kinase-2 (JAK-2), signal transducer and activator of transcription-3 (STAT-3) and tissue inhibitor of metalloproteinase-1 (TIMP-1) proteins in hepatic stellate cell-T6 (HSC-T6) induced by platelet-derived growth factor (PDGF). Methods:1. Drug-containing serum preparation:25 SD rats were randomly divided into 5 groups with 5 rats in each group. The drug-containing serum was obtained by intragastric administration to the SD rats respectively with physiological saline (lOml/kg, group A), Fufang Biejia Ruangan Tablet solution (1.5g/kg, group B), low dose (2.125g/kg, group C1), middle dose (4.25g/kg, group C2) and high dose (8.5g/kg, group C3) of Ling Qi Juan Gan Capsule solution.2. HSC-T6 induced by 10ng/ml PDGF was incubated at 200ml/L of the each group drug-containing serum. After 24,48,72 hours, the proliferation of HSC-T6 was measured by Cell Counting Kit-8 (CCK-8) and the growth curve was drew. The HSC-T6 ultrastructure changes were observed by transmission electron microscopy. After 24 hours, the expression of JAK-2, STAT-3 and TIMP-1 of HSC-T6 in each group was measured by western blot.3. Statistic Analysis: Measurement data was expressed by x±s. Multiple groups comparisons were made by one-way ANOVA analysis with Student-Newman-Keuls post hoc analysis. Significance level:Statistical significance was accepted at P<0.05. Results:1. Inhibiting proliferation of HSC-T6:Compared with group A, groups B, C2, C3 had a significantly higher proliferation inhibition rate at 24 hours (all P<0.01), while group C1 had no proliferation inhibition rate (P>0.05). The HSC-T6 proliferation inhibition rate of groups B and C3 were higher than that of group C2 (P>0.05), but there was no difference between B and C3 (P>0.05). Compared with group A, the other four groups B, C1, C2, C3 had a higher proliferation inhibition rate at 48 hours (P<0.05). The HSC-T6 proliferation inhibition rate of groups B and C3 were higher than that of groups C1 and C2 (P>0.05), but neither groups B and C3 nor C1 and C2 had difference (P>0.05). Compared with group A, the other four groups B, C1, C2, C3 had a higher proliferation inhibition rate at 72 hours (P<0.05), but there was no difference between each two groups (P>0.05). With increasing concentration of the Ling Qi Juan Gan Capsule drug-containingl serum, proliferation inhibition rate for HSC-T6 increased.2. Transmission electron microscopy showed:After 24 hours, there were no significant abnormalities of HSC-T6 indicated in group A. The shape of the nucleus was normal with clear nuclear membrane. The chromatin was in normal distribution. Abundant organelles and the lipid droplet could be seen. While in groups B,C1,C2 and C3, the varying degrees of necrotic and apoptotic cells were there. The cell volume was smaller and the nuclear membrane disappeared, the vacuoles in cytoplasm increased, the organelles were less, the chromatin became pyknotic and irregular, karyorrhexis was visible. These changes were more obvious in groups B and C3 than others at the each same time point and aggravated as the time proceeded.3. The expression of JAK-2, STAT-3 and TIMP-1 was detected by western blot after 24 hours:Compared with group A, JAK-2, STAT-3 and TIMP-1 in the other four groups were significantly decreased (all P<0.01). The expression of JAK-2, STAT-3 and TIMP-1 in group C1 was increased (P<0.01), while that was decreased in group C2 compared with group B,but there was no significant difference (P>0.05). JAK-2, TIMP-1 and STAT-3 in group C3 were all decreased compared with group B or C2, but the former two had a significant difference (P<0.01), and the latter didn't have (P>0.05). Conclusion:1. Ling Qi Juan Gan Capsule can inhibit the proliferation of HSC-T6 with a dose-dependent manner.2. Ling Qi Juan Gan Capsule can decrease the expression of JAK-2 and STAT-3 in activated HSC, and thus block JAK/STAT signal transduction pathway.3. Ling Qi Juan Gan Capsule can decrease the expression of TIMP-1 in activated HSC, alleviate the inhibition to MMPs. So Ling Qi Juan Gan Capsule had therapeutic effect on hepatic fibrosis.
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