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Objective:To investigate the effects of ancient Chinese medical formula Xiayuxue Decoction(下瘀血汤,XYXD) on activation of hepatic stellate cells(HSCs) and defenestration of sinusoidal endothelial cells(SECs) in CCI_4-induced fibrotic liver of mice.Methods:High performance liquid chromatography was used to identify the main components of XYXD and control the quality of extraction.C57BL/6 mice were induced liver fibrosis by CCl_4 exposure and administered with XYXD for 6 weeks simultaneously.Liver tissue was investigated by hematoxylin-eosin and Sirius-red staining.Sinusoidal fenestrations were observed by scanning electronic microscopy and fluorescent immunohistochemistry of PECAM-1(CD31).Whole liver lysates were detected of α-smooth muscle actin(α-SMA) and type-1 collagen by Western blot.Primary rat HSCs-T6 cells were analyzed by detecting a-SMA,F-actin,DNA fragmentation through confocal microscopy,Western blot,terminal-deoxynucleoitidyl transferase mediated nick end labeling(TUNEL) assay and cellomics arrayscan,respectively.Results:Amygdalin and emodin in XYXD were identified.XYXD(993 mg/kg) inhibited Sinus red positive area up to 70.1%(P<0.01),as well as protein levels of α-SMA and type-1 collagen by42.0%and 18.5%(P<0.05) respectively.In vitro,XYXD(12.5 μg/mL,50 μg/mL) suppressed the activation of HSCs and reversed the myofibroblastic HSCs into quiescent,demonstrated as inhibition of fluorescent F-actin by 32.3%and 46.6%(P<0.05).Besides,XYXD induced the apoptosis of HSC-T6 cells by 20.0%(P<0.05)and 49.5%(P<0.01),evidenced by enhanced TUNEL positivity.Moreover,ultrastructural observation suggested XYXD inhibited defenestration of SECs,which was confirmed by 31.1%reduction of protein level of CD31(P<0.05).Conclusions:XYXD inhibited both HSCs activation and SECs defenestration which accompany chronic liver injuries.These data may help to understand the underlying mechanisms of XYXD for prevetion of chronic liver diseases.
Objective: To investigate the effects of ancient Chinese medical formula Xiayuxue Decoction on activation of hepatic stellate cells (HSCs) and defenestration of sinusoidal endothelial cells (SECs) in CCI_4-induced fibrotic liver of mice. Methods: High performance liquid chromatography was used to identify the main components of XYXD and control the quality of extraction. C57BL / 6 mice were induced liver fibrosis by CCl_4 exposure and administered with XYXD for 6 weeks simultaneously. Liver tissue was investigated by hematoxylin-eosin and Sirius -red staining. Inusoidal fenestrations were observed by scanning electronic microscopy and fluorescent immunohistochemistry of PECAM-1 (CD31). Whole liver lysates were detected of α-smooth muscle actin (α-SMA) and type-1 collagen by Western blot. Primary rat HSCs-T6 cells were analyzed by detecting a-SMA, F-actin, DNA fragmentation through confocal microscopy, Western blot, terminal-deoxynucleoitidyl transferase mediated nick end labeling (TU NEL) assay and cellomics arrayscan, respectively.Results: Amygdalin and emodin in XYXD were identified.XYXD (993 mg / kg) inhibited Sinus red positive area up to 70.1% (P <0.01) In addition, XYXD (12.5 μg / mL, 50 μg / mL) suppressed the activation of HSCs into reversed-myeloblastic HSCs into quiescent, demonstrated as inhibition of fluorescent F actin by 32.3% and 46.6% (P <0.05) .Besides, XYXD induced apoptosis of HSC-T6 cells by 20.0% (P <0.05) and 49.5% (P <0.01) respectively, evidenced by enhanced TUNEL positivity . Moreover, ultrastructural observation XYXD inhibited defection of SECs, which was confirmed by 31.1% reduction of protein level of CD31 (P <0.05) .Conclusions: XYXD inhibited both HSCs activation and SECs defenestration which accompany chronic liver lesions. to understand the underlying mechanisms of XYXD for prevetion of chronic liver diseases.