ABSTRACT
ObjectiveTo investigate the effect of phytoestrogen biochanin A （BCA） on liver fibrosis induced by CCl4 in female mice with bilateral oophorectomy （ovariectomized） and its mechanism. MethodsA total of 50 ovariectomized Kunming mice were selected and given intraperitoneal injection of CCl4 to establish a model of liver fibrosis， and then according to body weight， they were randomly divided into model group， positive control group， and low-， middle-， and high-dose BCA groups， with 10 mice in each group. In addition， 10 female mice in the same litter were given resection of a small amount of adipose tissue near both ovaries to establish the sham-operation group. The mice in the positive control group were given estradiol 2 mg/kg by gavage， and those in the low-， middle-， and high-dose BCA groups were given BCA by gavage at a dose of 25， 50， and 100 mg/kg， respectively， once a day for 7 consecutive weeks； the mice in the sham-operation group and the model group were given an equal volume of 0.5% sodium carboxymethyl cellulose solution by gavage. The mice were anesthetized and sacrificed after administration to collect samples. Liver index and uterus index were measured； HE staining and Masson staining were used to observe liver histopathological changes； the biochemical analysis was used to measure the activity of aspartate aminotransferase （AST） and alanine aminotransferase （ALT）； ELISA was used to measure the levels of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α） in liver tissue， and Western blot was used to measure the relative protein expression levels of collagen Ⅰ， transforming growth factor-beta 1 （TGF-β1）， alpha-smooth muscle actin （α-SMA）， estrogen receptor beta （ERβ）， and p-NF-κBp65/NF-κBp65 in liver tissue. The t-test was used for comparison of continuous data between two groups； a one-way analysis of various was used for comparison between multiple groups， and the least significant difference t-test was used for further comparison between two groups. The Kruskal-Wallis H test was used for comparison of non-normally distributed continuous data between multiple groups and further comparison between two groups. ResultsCompared with the sham-operated group， the model group had a significant increase in liver index and a significant reduction in uterus index， as well as significant increases in the activities of serum AST and ALT， the levels of IL-6 and TNF-α in liver tissue， and the protein expression levels of collagen Ⅰ， TGF-β1， α-SMA， and p-NF-κBp65/NF-κBp65 in liver tissue （all P<0.05）， with no significant change in the expression of ERβ in liver tissue （P>0.05）， and the model group showed significant fibrosis lesions in the liver， such as hepatocyte edema， steatosis， and necrosis with inflammatory cell infiltration and hyperplasia， deposition， and staggered distribution of collagen fibers. Compared with the model group， the low-， middle-， and high-dose BCA groups had significant reductions in liver index， the activities of serum AST and ALT， the levels of IL-6 and TNF-α， and the protein expression levels of collagen Ⅰ， TGF-β1， α-SMA， and p-NF-κBp65/NF-κBp65 in liver tissue （all P<0.05）， with no significant change in uterine index （P>0.05）， as well as a significant increase in the protein expression level of ERβ in liver tissue （P<0.05） and varying degrees of improvement in liver fibrosis lesions. ConclusionBCA can effectively improve CCL4-induced liver fibrosis in ovariectomized female mice， possibly by upregulating ERβ to inhibit the NF-κB signaling pathway and then alleviating inflammatory response.