CAJ | 학술논문

目的运用CCl4肝纤维化模型研究血管紧张素转换酶(ACE)2在肝纤维化形成中的表达及其与纤维化的关系. 方法将64只雄性SD大鼠随机分为2组.模型组按3m/kg皮下注射40％CC14,对照组按3ml/kg皮下注射橄榄油,每周2次.分别在注射前(0周)及注射后第2、4、6周取肝组织及血清.实时荧光定量PCR检测ACE2和ACE的mRNA相对表达量,Western blot检测ACE2蛋白相对表达量,酶联免疫吸附法检测血清中ACE2浓度.两组间差异比较用t检验,多组样本的比较用单因素方差分析或Kruskal Wallis检验,等级相关分析用Spearman等级相关检验.结果造模0、2、4、6周组的ACE2 mRNA相对表达量分别为0.991±0.079、3.055±1.034、3.545±1.947、6.448±1.836,差异有统计学意义(H=23.224,P＜0.01); ACE2 mRNA的表达在造模2、4、6周逐步升高,且均明显高于对照组(t值分别为-5.760、-3.678和-8.453,P值均＜0.01).造模0、2、4、6周组肝组织ACE mRNA相对表达量分别为1.005±0.193、3.956±0.934、4.974±2.004、6.265±2.718,差异有统计学意义F=12.982,P＜0.01); ACE mRNA的表达在造模2、4、6周逐步升高,且均明显高于对照组(t值分别为-8.790、-5.544和-5.431,P值均＜0.01).ACE2蛋白在正常组低表达,随着造模时间的延长,其表达逐渐升高,造模0、2、4、6周组的ACE2蛋白相对表达量分别为0.034、0.097、0.356、0.512,造模第6周表达最高.ACE2 mRNA的表达与肝纤维化等级评分及ALT、AST变化具有良好的相关性(r值分别为0.850、0.669和0.815,P值均＜0.01),外周血中游离ACE2的含量与肝纤维化等级评分具有良好相关性(r=0.730,P＜0.01).结论 ACE2参与肝纤维化的形成,在肝纤维化形成过程中的表达明显上调,且其表达量与肝纤维化程度明显相关.
목적 운용CCl4간섬유화모형연구혈관긴장소전환매(ACE)2재간섬유화형성중적표체급기여섬유화적관계. 방법 장64지웅성SD대서수궤분위2조.모형조안3m/kg피하주사40％CC14,대조조안3ml/kg피하주사감람유,매주2차.분별재주사전(0주)급주사후제2、4、6주취간조직급혈청.실시형광정량PCR검측ACE2화ACE적mRNA상대표체량,Western blot검측ACE2단백상대표체량,매련면역흡부법검측혈청중ACE2농도.량조간차이비교용t검험,다조양본적비교용단인소방차분석혹Kruskal Wallis검험,등급상관분석용Spearman등급상관검험.결과 조모0、2、4、6주조적ACE2 mRNA상대표체량분별위0.991±0.079、3.055±1.034、3.545±1.947、6.448±1.836,차이유통계학의의(H=23.224,P＜0.01); ACE2 mRNA적표체재조모2、4、6주축보승고,차균명현고우대조조(t치분별위-5.760、-3.678화-8.453,P치균＜0.01).조모0、2、4、6주조간조직ACE mRNA상대표체량분별위1.005±0.193、3.956±0.934、4.974±2.004、6.265±2.718,차이유통계학의의F=12.982,P＜0.01); ACE mRNA적표체재조모2、4、6주축보승고,차균명현고우대조조(t치분별위-8.790、-5.544화-5.431,P치균＜0.01).ACE2단백재정상조저표체,수착조모시간적연장,기표체축점승고,조모0、2、4、6주조적ACE2단백상대표체량분별위0.034、0.097、0.356、0.512,조모제6주표체최고.ACE2 mRNA적표체여간섬유화등급평분급ALT、AST변화구유량호적상관성(r치분별위0.850、0.669화0.815,P치균＜0.01),외주혈중유리ACE2적함량여간섬유화등급평분구유량호상관성(r=0.730,P＜0.01).결론 ACE2삼여간섬유화적형성,재간섬유화형성과정중적표체명현상조,차기표체량여간섬유화정도명현상관.
Objective To investigate the expression and pathogenic relevance of angiotensinconverting enzyme 2 (ACE2) in liver fibrosis by using the rat model of CCl4-induced liver fibrosis.Methods The liver fibrosis model was generated by delivering subcutaneous injections of CCl4 (dissolved in olive oil at a 2:3 ratio; injection dose:3 ml/kg) every three days for six weeks into male Sprague-Dawley rats.Another group of rats that received simultaneous injections of olive oil alone (3 ml/kg) were used as controls.At week 0,2,4,or 6 after the first injection,a subset of rats from each group was sacrificed to obtain liver tissues and serum samples.Pathological analyses were carried out to detect the presence and extent of liver cell degeneration,necrosis,inflammatory cell infiltration,and collagen deposition.ACE2 and ACE gene and protein expressions were measured by real-time PCR and Western blotting,respectively.The significance of differential expression between groups and time points was assessed by t-test and one-way ANOVA or Kruskal-Wallis tests,and correlation with fibrosis was assessed by Spearman's rank correlation coefficient.Results CCl4 administration led to significantly up-regulated ACE2 mRNA levels at week 2 (3.055 ± 1.034),4 (3.545± 1.947),and 6 (6.448± 1.836) (vs.controls; H=23.224,P＜0.001).Similarly,hepatic ACE mRNA was significantly increased after the CCl4 injections (week 2:3.055 ± 1.034,week 4:3.545 ± 1.947,week 6:6.448 ± 1.836; vs.controls:F=12.982,P＜ 0.001).There was a significant correlation between the ACE and ACE2 gene expression levels (r=0.750,P＜ 0.001).Protein levels of ACE2 also showed an increasing trend following CCl4 administration (week 0:0.034,week 2:0.097,week 4:0.356,week 6:0.512).The hepatic ACE2 gene expression strongly correlated with levels of alanine aminotransferase (r=0.669,P＜ 0.0001)and aspartate aminotransferase (r=0.815,P＜0.0001),and with the Ishak fibrosis score (r=0.850,P＜0.001).Finally,there was a significant correlation between circulating ACE2 and the Ishak fibrosis score (r =0.730,P＜0.001).Conclusion A significant relationship exists between ACE2 gene expression and extent of liver fibrosis.ACE2 may play a crucial role in liver fibrogenesis.