| Objective: To investigate the roles of mitochondrial damage in hepatocye apoptosis in nonalcoholic fatty liver (NAFL) rats and the pathogenesis.Methods: (1) A total of 30 Wistar rats were randomly divided into basic diet-control group (Group C) and high-fat diet group (Group F). Each of the two groups was subdivided into 3 subgroups (4, 8, and 12 weeks) (n = 5 in each group). (2) The following parameters in each group were observed dynamically: ①changes in animal weight, ② pathological changes in the liver tissue observed by HE staining and light microscopy, ③ ultrastructural changes in the hepatocyte observed by electron microscopy, ④ respiratory chain function of mitochondria detected by Clark oxygen electrode, and mitochondrial membrane potential by fluorescence spectrophotometer, ⑤ levels of malondialdehyde (MDA) in liver tissue detected by using thio-barbituric acid, ⑥ hepatocyte apoptosis index detected by flow cytometry, ⑦ expressions of Cyt C, Bcl-2, Bax, and Caspase-3 detected by Western blotting and immunohistochemistry.Results: (1) Animal weight in Group F group was significantly higher than that in Group C (P<0.01). (2) HE staining revealed that fatty degeneration at 4 w, mild fatty liver at 8 w, moderate to severe fatty liver at 12 w in Group F, and electron microscopy showed ultrastructural changes in hepatocytes Group F (12 w), including swelling of hepatic mitochondria, shortened or even disappearance of crests, deeply stained chromatin concentrated at the edge of nuclear membrane, ③ Respiratory control rates (RCR) in Group F at 8 w (3.87±0.44) and 12 w (3.51±0.18) were significantly lower than those in Group C at 8 w (4.93±0.79) and 12 w (4.85±0.47) (P<0.05, P<0.01). P/O in Group F at 4 w (3.61±0.39), 8 w (3.56±0.34), and 12 w (3.37±0.25) were significantly lower than those in Group C at 4 w (4.22±0.63), 8 w (4.11±0.29), and 12 w (4.13±0.39) (P<0.05 or P<0.01). ④ Mitochondrial membrane potentials in Group F at 4 w (833.1±53.5), 8 w (772.2±73.4), and 12 w (587.2±63.1) were significantly lower than those in Group C at 4 w (951.2±64.6), 8 w (977.2±78.3), and 12 w (958.6±70.4) (P<0.05or P<0.01). ⑤ Liver tissue MDA levels in Group F were significantly higher than those in Group C (P<0.05). ⑥ Apoptosis indexes in Group F at 8 w (9.2±0.4) and 12 w (11.6±1.1) were lower than those in Group C at 8 w (4.3±0.3) and 12 w (4.3±0.2) (P<0.01). ⑦ Immunohistochemistry showed that Cyt C was in the form of lightly stained brown-yellow granules distributed in spot-like form, and in Group F, the staining deepened and the stained area enlarged in a time-dependent manner, Caspase-3 was distributed diffusedly in normal liver tissue with weak positive staining of inner cytoplasm, but in Group F at 4 w, it was more deeply stained, and in Group F at 8 w and 12 w, the staining was gradually deepened, and the stained area enlarged gradually and distributed diffusedly, diffusedly distributed weak positive expressions of Bcl-2 and Bax were found in the normal group, but in Group F at 4, 8, and 12 w, the stained area enlarged, and deep staining was found at the sites with obvious fatty degeneration. ⑧ Protein expressions of Bcl-2, Bax, and Caspase-3 in Group F, particularly at 8 and12 w increased in a time-dependent manner with significant difference as compared with those in Group C group at 12 w (P < 0.05 or P<0.01), Bcl-2/ Bax in Group F decreased with the progression of fatty liver, Cyt C expressions in Group F at 4, 8, and 12 w increased significantly as compared with those in the control group (P<0.05), but no difference was found between the subgroups in Group F.Conclusions: (1) Structural and functional damages in hepatic mitochondria occur in the genesis and progression of NAFL. (2) The increase of MDA in liver tissue and lipid peroxidation due to the functional damage of mitochondria may lead to decline of mitochondrial membrane potential, release of cytochrome C, activation of Caspase-3, resulting in hepatocyte apoptosis in NAFL. (3) Bcl-2 family may participate in the regu...
|
PDF Full Text Request