缺血后处理通过抑制细胞焦亡减轻大鼠肺缺血/再灌注引发的肝损伤

Ischemic postconditioning attenuates lung ischemia/reperfusion-induced liver damage in a rat model by inhibiting pyroptosis

目的:探讨细胞焦亡在大鼠肺缺血/再灌注(I/R)引发的肝脏损伤中的作用及缺血后处理(I-post-C对其干预的保护机制。方法:将32只SPF级SD雄性大鼠随机分为对照(control)组、I/R组、I/R+I-post-C组及I/R+INF39(细胞焦亡抑制剂)组,每组8只。普通光镜下观察肝脏组织形态;用相关化学试剂盒测定丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)和caspase-1活性;采用ELISA的方法检测血清中白细胞介素1β(IL-1β)和IL-18水平;使用Western blot及免疫荧光法检测肝组织细胞焦亡相关蛋白的表达情况。结果:与control组相比,I/R组肝小叶结构紊乱,部分肝细胞出现水肿、空泡样变性和坏死,炎症细胞浸润,血清ALT和AST水平显著升高(P<0.01);细胞焦亡相关指标核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、胱天蛋白酶1(caspase-1)和gasdermin D N端片段(GSDMD-N)蛋白表达增加(P<0.05);NLRP3的免疫荧光表达显著增强(P<0.01);caspase-1活性显著上升(P<0.01);血清中IL-1β和IL-18水平显著升高(P<0.01)。与I/R组相比,I/R+I-post-C组和I/R+INF39组的肝细胞水肿减轻、坏死减少,中央静脉、肝血窦扩张及淤血减轻;大鼠血清ALT和AST水平显著降低(P<0.05);NLRP3免疫荧光表达减弱(P<0.05);细胞焦亡相关指标NLRP3、caspase-1和GSDMD-N的蛋白水平显著降低(P<0.01);血清中IL-1β和IL-18含量显著降低(P<0.01)。结论:I-post-C可通过抑制细胞焦亡减轻大鼠肺I/R引发的肝损伤。

AIM To explore the role of pyroptosis in liver damage induced by lung ischemia/reperfusion(I/R),as well as the protective mechanism of ischemic postconditioning(I-post-C)in rats.METHODS Thirty-two SPF SD male rats were randomly divided into control group,I/R group,I/R+I-post-C group and I/R+INF39(pyroptosis inhibitor)group.The morphological changes of liver tissues were subsequently observed under light microscope.The activity of alanine aminotransferase(ALT),aspartate transaminase(AST)and caspase-1 was tested by chemical test kit.Interleukin-1β(IL-1β)and IL-18 levels were detected by ELISA.Western blot and immunofluorescence were used to detect the expression of pyroptosis-related proteins in extracted liver tissues.RESULTS Compared with control group,the hepatic lobular structure in I/R group was more disordered.Furthermore,serum ALT and AST levels were elevated(P<0.01)along with liver MDA levels(P<0.01)and pyroptosis-related protein levels.More specifically,nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),caspase-1 and gasdermin D N-terminal fragment(GSDMD-N)were elevated(P<0.05).Immunofluorescence studies showed significant increases in NLRP3 expression level(P<0.01),caspase-1 activity(P<0.01),as well as serum IL-1βand IL-18 levels(P<0.01).Compared with I/R group,the level of edema in hepatic cells in I/R+I-post-C and I/R+INF39 groups was significantly reduced.Furthermore,central vein and hepatic blood sinus dilation was decreased along with a decrease in MDA level(P<0.05).Serum ALT and AST levels were significantly reduced(P<0.05).The expression of NLRP3 was significantly reduced(P<0.05),as well as the expression levels of caspase-1 and GSDMD-N(P<0.01).Serum IL-1βand IL-18 levels were also significantly reduced(P<0.01).CONCLUSION I-post-C attenuates liver damage caused by lung I/R in rats through inhibition of pyroptosis.