Effects Of Baoganning On Hepatic Fibrosis And Its Regulation On Nuclear Transcriptional Factor-κB

To investigate the effects of Baoganning on proliferation and apoptosis of HSC-T6; to elucidate the regulation of Baoganning on the expression of IL-6 and Bcl-2; to study the action of Baoganning on the activity of NF-B and B, try to interpret the protective effects of Baoganning on hepatic fibrosis. Methods:1. in vitro study: Rats were divided into three groups that were fed with low dosage Baoganning (1.35g/ml), high dosage Baoganning (2.7g/ml) and saline for seven days. The serum with or without Baoganning ingredients was prepared two hours after the last feeding. HSC cultured and passaged were divided into three groups that were treated with serum of low dosage Baoganning, serum of high dosage Baoganning and normal rat serum respectively. Cell proliferation was detected by the SRB colorimetric assay. The apoptosis of cells were observed by phase-contrast fluorescent micrography and agarose gel electrophoresis. The expression of Bcl-2, monomer of NF-KB (p50 ) and IicBa was detected by the Western Blot. Secret of IL-6 was detected by Enzyme-linked immunoadsorden assay (ELISA).2. in vivo study: The liver fibrosis model of rats was made by hypodermic injection of carbon tetrachloride. Model rats were divided into four groups that were treated with low dosage Baoganning (low dosage baoganning group), high dosage Baoganning (high dosage baoganning group), saline (pathology group) and colchicines (colchicines group); moreover, rats without carbon tetrachloride that fed with saline were set as control. Histopathology of liver was observed with HE stain. Iinmunohistochemical examination of NF-KB was performed onformalin-fixed, and paraffin-embedded sections of rat liver. Ixfiaphosphorylation of liver was detected by the Western Blot.Results:1. Baoganning low dosage and high dosage could significantly inhibit HSC proliferation compared with normal group (P<0.01). However, no significant difference was observed between low dosage and high dosage baogannig groups (P0.05). It demonstrated that the inhibition effect of baoganning didn't increase pro rata.2. After HSC have been cultured for three hours in different culture medium, the phosphorylation level of Ixfla, which demonstrated translocation of NF-KB to nuclear, decreased significantly in baoganning group compared with normal group (P<0.01). The phosphorylation level of IicBa has no significant difference 48 hours after treatment(P>0.05).3. Compared with the control group, Baoganning could significantly increase the apoptosis rates of HSC (P <0.01 ).4. Baoganning could significantly discrease the level of NF-KB, IicBa, Bcl-2 and IL-6 in which the expression of Bcl-2 and IL-6 were regulated by NF-KB (PO.01).5. After the rats were treated with Baoganning, Western blot analysis suggested that iKBa phosphorylation level in model rats increased compared with control (P<0.01). Baoganning high dosage, low dosage and colchicines lowered the IicBa phosphorylation level compared with pathology group (PO.01). Compared with colchicines group, baoganning decreased IicBa phosphorylation level significantly (P<0.01).6. Immunohistochemical examination of NF-KB showed that there were a small quantity of NF-KB hi cell cytoplasm of normal liver. But in pathology groupliver NF-KB expressde strong positive. In Baoganning group and colchicines group NF-icB expressed secondarily and NF-KB main lies in interstitial cell nuclear. Contrast to pathology group, the level of NF-xfi expression hi Baoganning group and colchicines group were lower. Thedifference had significant means (P<0.01). No difference was betweenBaoganning group and colchicines group (P>0.05). Between Baoganninggroups there was not different (P>0.05).Conclusion:1. Baoganning could inhibit the activation and proliferation of HSC, and acceleration apoptosis of HSC;2. The inhibition of activation and proliferation of HSC may be mediated by Baoganning inhibiting the expres...