| Objective:To investigate the effects of Ling Qi Juan Gan Capsule medicated serum on the apoptosis and the expression of Bcl-2, Bax, tissue inhibitor of metalloproteinase-1(TIMP-1), matrix metallo-proteinase-2(MMP2), and collagen type I and III in hepatic stellate cell-T6(HSC-T6) activated by platelet-derived growth factor (PDGF). Methods:1. Preparation of drug-medicated serum.30SD rats were randomly divided into3groups with10rats in each group. The drug-medicated serum was obtained from the SD rats administrated intragastricly with physiological saline (10ml/kg, group A), Fufang Biejia Ruangan Tablet solution (1.5g/kg, group B)and Ling Qi Juan Gan Capsule solution respectively (4.25g/kg, group C).2. Detection of the apoptosis and the expression of Bcl-2/Bax in HSC-T6. HSC-T6activated by10ng/ml PDGF was incubated with drug-medicated serum at200ml/L The apoptosis of HSC-T6and the expression of BCL-2, Bax were measured by flow cytometry at0,12,24and48hours.3. The concentration of TIMP-1, MMP2, collagen type Ⅰ and Ⅲ in supernatant were measured by enzyme linked immunosorbent assay (ELISA).4. Statistic Analysis:Measurement data was expressed by x±s. Multiple group comparisons were made by one-way ANOVA analysis with Student-Newman-Keuls or Dunnett post hoc analysis. Significance level: Statistical significance was accepted at P<0.05. All the data was analysed by software SPSS13.0. Results:1. The apoptotic rate of HSC-T6in group B and group C was significantly induced compared with that of group A at12,24,48h respectively (P<0.05),while there were no significant difference of apoptotic rate between group B and group C (P>0.05). The significant differences of apoptotic rate have also been found at different time points both in group B and group C. With the increase of exposure time, the apoptotic rate increases correspond-ingly (P<0.05).2.The effects of drug-medicated serum on Bcl-2expression:The Bcl-2expression level of group B and C were lower than that of group A (P<0.05)at12,24,48h,there was no significant difference of its expression level between group B and group C (P>0.05). Further work shows that the expression level of Bcl-2decreased with the prolongation of exposure time (P>0.05) in both Group B and Group C.However, no such trend was observed in group A (P>0.05).3. The effects of drug-medicated serums on Bax expression:The Bax expression of group B and group C increased significantly compared with that of group A at12,24,48h (P<0.05). And there was no significant difference of its expression level between group B and group C (P>0.05). Further study shows that in both group B and group C, the expression level of Bax increased time-dependently compared with that of group A.4. The effects on TIMP-1, MMP2and type Ⅰ, Ⅲ collagen expression:the levels of TIMP-1, type Ⅰ, Ⅲ collagen of group B and group C decreased significantly compared with that of group A (P<0.05). And there was no significant difference of their expression level between group B and group C (P>0.05). The MMP2level of group B and C increased significantly compared with that of group A (P<0.01). The was no statistic difference of MMP2level between group C group B (P>0.05).Conclusions:1. LingQiJuanGan capsule can induce apoptosis of the activated HSC-T6with a time-dependent manner by means of influencing the expression of BCL-2/Bax.2. LingQiJuanGan capsule can downregulate the expression of TIMP-1and upregulate the expression of MMP2in the activated HSC-T6.3. LingQiJuanGan capsule can decrease the secretion of collagen type Ⅰ and Ⅲ in the activated HSC-T6. In conclusion, our study indicated that LingQiJuanGan capsules could mediate apoptosis of the activated rat hepatic stellate cell by influencing the expression of Bcl-2/Bax, ultimately decreased the secretion of extracellular matrix and enhanced the degradation of extracellular matrix to inhibit hepatic fibrosis. |
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