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Abstract：
OBJECTIVE: To investigate the protective effect of recombinant Schistosoma japonicum cystatin (rSj-Cys) against acute liver injury induced by lipopolysaccharide (LPS) and D-GalN in mice.
METHODS: Adult male C57BL/6J mice with or without LPS/D-GaIN-induced acute liver injury were given intraperitoneal injections of rSj-Cys or PBS 30 min after modeling (n=18), and serum and liver tissues samples were collected from 8 mice in each group 6 h after modeling. The survival of the remaining 10 mice in each group within 24 h was observed. Serum levels of ALT, AST, TNF-α and IL-6 of the mice were measured, and liver pathologies was observed with HE staining. The hepatic expressions of macrophage marker CD68, Bax, Bcl-2 and endoplasmic reticulum stress (ERS)-related proteins were detected using immunohistochemistry or immunoblotting, and TUNEL staining was used to detect hepatocyte apoptosis.
RESULTS: The survival rates of PBS- and rSj-Cys-treated mouse models of acute liver injury were 30% and 80% at 12 h and were 10% and 60% at 24 h after modeling, respectively; no death occurred in the two control groups within 24 h. The mouse models showed significantly increased serum levels of AST, ALT, IL-6 and TNF-α and serious liver pathologies with increased hepatic expressions of CD68 and Bax, lowered expression of Bcl-2, increased hepatocyte apoptosis, and up-regulated expressions of ERS-related signaling pathway proteins GRP78, CHOP and NF-κB p-p65. Treatment of the mouse models significantly lowered the levels of AST, ALT, IL-6 and TNF-α, alleviated liver pathologies, reduced hepatic expressions of CD68, Bax, GRP78, CHOP and NF-κB p-p65, and enhanced the expression of Bcl-2. In the normal control mice, rSj-Cys injection did not produce any significant changes in these parameters compared with PBS.
CONCLUSIONS: rSj-Cys alleviates LPS/D-GalN-induced acute liver injury in mice by suppressing ERS, attenuating inflammation and inhibiting hepatocyte apoptosis.
METHODS: Adult male C57BL/6J mice with or without LPS/D-GaIN-induced acute liver injury were given intraperitoneal injections of rSj-Cys or PBS 30 min after modeling (n=18), and serum and liver tissues samples were collected from 8 mice in each group 6 h after modeling. The survival of the remaining 10 mice in each group within 24 h was observed. Serum levels of ALT, AST, TNF-α and IL-6 of the mice were measured, and liver pathologies was observed with HE staining. The hepatic expressions of macrophage marker CD68, Bax, Bcl-2 and endoplasmic reticulum stress (ERS)-related proteins were detected using immunohistochemistry or immunoblotting, and TUNEL staining was used to detect hepatocyte apoptosis.
RESULTS: The survival rates of PBS- and rSj-Cys-treated mouse models of acute liver injury were 30% and 80% at 12 h and were 10% and 60% at 24 h after modeling, respectively; no death occurred in the two control groups within 24 h. The mouse models showed significantly increased serum levels of AST, ALT, IL-6 and TNF-α and serious liver pathologies with increased hepatic expressions of CD68 and Bax, lowered expression of Bcl-2, increased hepatocyte apoptosis, and up-regulated expressions of ERS-related signaling pathway proteins GRP78, CHOP and NF-κB p-p65. Treatment of the mouse models significantly lowered the levels of AST, ALT, IL-6 and TNF-α, alleviated liver pathologies, reduced hepatic expressions of CD68, Bax, GRP78, CHOP and NF-κB p-p65, and enhanced the expression of Bcl-2. In the normal control mice, rSj-Cys injection did not produce any significant changes in these parameters compared with PBS.
CONCLUSIONS: rSj-Cys alleviates LPS/D-GalN-induced acute liver injury in mice by suppressing ERS, attenuating inflammation and inhibiting hepatocyte apoptosis.
摘要：
目的：研究重组日本血吸虫胱抑素(rSj-Cys)对脂多糖(LPS)和D-GalN诱导的小鼠急性肝损伤的保护作用。
方法：成年雄性C57BL/6J小鼠有或没有LPS/D-GaIN诱导的急性肝损伤，在造模后30分钟腹腔注射rSj-Cys或PBS（n=18），并在造模后6h采集各组8只小鼠的血清和肝组织样本。观察各组剩余10只小鼠在24h内的存活情况。血清ALT水平，AST,检测小鼠的TNF-α和IL-6,HE染色观察肝脏病理。肝脏巨噬细胞标记物CD68、Bax的表达,采用免疫组织化学或免疫印迹法检测Bcl-2和内质网应激(ERS)相关蛋白,TUNEL染色检测肝细胞凋亡。
结果：经PBS和rSj-Cys处理的急性肝损伤小鼠模型在造模后12h生存率分别为30%和80%，24h生存率分别为10%和60%，两个对照组在24h内没有死亡。小鼠模型显示血清AST水平显着升高，ALT,IL-6和TNF-α与CD68和Bax表达增加的严重肝脏病变，Bcl-2表达降低，肝细胞凋亡增加，并上调ERS相关信号通路蛋白GRP78、CHOP和NF-κBp-p65的表达。小鼠模型的处理显著降低了AST的水平,ALT,IL-6和TNF-α,减轻肝脏病变，CD68、Bax、GRP78、CHOP和NF-κBp-p65增强Bcl-2的表达。在正常对照小鼠中，与PBS相比,rSj-Cys注射不产生这些参数的任何显著变化。
结论：rSj-Cys通过抑制ERS减轻LPS/D-GalN诱导的小鼠急性肝损伤,减轻炎症和抑制肝细胞凋亡。
方法：成年雄性C57BL/6J小鼠有或没有LPS/D-GaIN诱导的急性肝损伤，在造模后30分钟腹腔注射rSj-Cys或PBS（n=18），并在造模后6h采集各组8只小鼠的血清和肝组织样本。观察各组剩余10只小鼠在24h内的存活情况。血清ALT水平，AST,检测小鼠的TNF-α和IL-6,HE染色观察肝脏病理。肝脏巨噬细胞标记物CD68、Bax的表达,采用免疫组织化学或免疫印迹法检测Bcl-2和内质网应激(ERS)相关蛋白,TUNEL染色检测肝细胞凋亡。
结果：经PBS和rSj-Cys处理的急性肝损伤小鼠模型在造模后12h生存率分别为30%和80%，24h生存率分别为10%和60%，两个对照组在24h内没有死亡。小鼠模型显示血清AST水平显着升高，ALT,IL-6和TNF-α与CD68和Bax表达增加的严重肝脏病变，Bcl-2表达降低，肝细胞凋亡增加，并上调ERS相关信号通路蛋白GRP78、CHOP和NF-κBp-p65的表达。小鼠模型的处理显著降低了AST的水平,ALT,IL-6和TNF-α,减轻肝脏病变，CD68、Bax、GRP78、CHOP和NF-κBp-p65增强Bcl-2的表达。在正常对照小鼠中，与PBS相比,rSj-Cys注射不产生这些参数的任何显著变化。
结论：rSj-Cys通过抑制ERS减轻LPS/D-GalN诱导的小鼠急性肝损伤,减轻炎症和抑制肝细胞凋亡。
