CAJ | 학술논문

目的：探讨三七皂甙( panax notoginseng saponins,PnS) Rg1、PnS Rb1干预下,肝纤维化大鼠线粒体DNA三磷酸腺苷( adenine triphosphate,ATP)酶6亚基、ATP酶8亚基的基因突变和ATP含量变化。方法96只Wistar大鼠分为对照组、四氯化碳( carbon tetrachloride,CCl4)肝纤维化大鼠模型组、PnS Rg1组、PnS Rb1组各24只。除对照组外,其余3组用5％ CCl4橄榄油按5 ml/kg灌胃制作肝纤维化大鼠模型。 PnS Rg1组在每次CCl4灌胃同时腹腔注射PnSRg1(5 mg/kg)。 PnS Rb1组在每次CCl4灌胃同时腹腔注射Rb1(5 mg/kg)。用生物发光法测定各组大鼠实验前及实验开始2、4、6周末时线粒体内ATP含量并比较。提取肝细胞线粒体总RNA,逆转录为互补脱氧核糖核酸( complementary DNA,cDNA),用基因重组、测序,通过GenBank与线粒体 DNA已知序列进行比较,研究PnS Rg1、PnS Rb1干预6周后线粒体 DNA的ATP酶6亚基7904-8584区域和ATP酶8亚基7743-7946区域基因突变情况。结果(1)与对照组相比,模型组大鼠线粒体DNA ATP酶6亚基7904-8584区域有3处突变：C8294G、T8505G、G8584A。用Fisher确切概率法分析,突变增加有显著性意义( P<0.01)。 PnS Rg1组、PnS Rb1组与对照组相比突变增加没有显著性意义( P>0.05)。(2)与对照组相比,ATP酶8亚基7743-7946区域有2处突变：A7797non、T7863C。用Fisher确切概率法分析,突变增加有显著性意义(P<0.01)。 PnS Rg1、Rb1组与对照组相比突变增加没有显著性意义( P>0.05)。(3)各组肝纤维化大鼠线粒体 ATP 含量与 CCl4处理时间的相关性分析,用pearson＇s 检验r=-0.935,呈负相关。(4)肝纤维化大鼠线粒体内ATP含量与ATP酶6亚基突变的相关性分析,用pearson＇s 检验r=-0.943,呈负相关；肝纤维化大鼠线粒体内ATP含量与ATP酶8亚基突变的相关性分析,用pearson＇s检验r=-0.961,呈负相关。结论 PnS Rg1、Rb1可以显著抑制线粒体ATP含量下降,可以显著抑制线粒体DNA ATP酶6亚基、ATP酶8亚基突变,从而推断PnS Rg1、Rb1可能通过减少线粒体DNA ATP酶6、8亚基突变而抑制线粒体ATP含量下降,从而延缓肝纤维化发生发展。目的：探討三七皂甙( panax notoginseng saponins,PnS) Rg1、PnS Rb1榦預下,肝纖維化大鼠線粒體DNA三燐痠腺苷( adenine triphosphate,ATP)酶6亞基、ATP酶8亞基的基因突變和ATP含量變化。方法96隻Wistar大鼠分為對照組、四氯化碳( carbon tetrachloride,CCl4)肝纖維化大鼠模型組、PnS Rg1組、PnS Rb1組各24隻。除對照組外,其餘3組用5％ CCl4橄欖油按5 ml/kg灌胃製作肝纖維化大鼠模型。 PnS Rg1組在每次CCl4灌胃同時腹腔註射PnSRg1(5 mg/kg)。 PnS Rb1組在每次CCl4灌胃同時腹腔註射Rb1(5 mg/kg)。用生物髮光法測定各組大鼠實驗前及實驗開始2、4、6週末時線粒體內ATP含量併比較。提取肝細胞線粒體總RNA,逆轉錄為互補脫氧覈糖覈痠( complementary DNA,cDNA),用基因重組、測序,通過GenBank與線粒體 DNA已知序列進行比較,研究PnS Rg1、PnS Rb1榦預6週後線粒體 DNA的ATP酶6亞基7904-8584區域和ATP酶8亞基7743-7946區域基因突變情況。結果(1)與對照組相比,模型組大鼠線粒體DNA ATP酶6亞基7904-8584區域有3處突變：C8294G、T8505G、G8584A。用Fisher確切概率法分析,突變增加有顯著性意義( P<0.01)。 PnS Rg1組、PnS Rb1組與對照組相比突變增加沒有顯著性意義( P>0.05)。(2)與對照組相比,ATP酶8亞基7743-7946區域有2處突變：A7797non、T7863C。用Fisher確切概率法分析,突變增加有顯著性意義(P<0.01)。 PnS Rg1、Rb1組與對照組相比突變增加沒有顯著性意義( P>0.05)。(3)各組肝纖維化大鼠線粒體 ATP 含量與 CCl4處理時間的相關性分析,用pearson＇s 檢驗r=-0.935,呈負相關。(4)肝纖維化大鼠線粒體內ATP含量與ATP酶6亞基突變的相關性分析,用pearson＇s 檢驗r=-0.943,呈負相關；肝纖維化大鼠線粒體內ATP含量與ATP酶8亞基突變的相關性分析,用pearson＇s檢驗r=-0.961,呈負相關。結論 PnS Rg1、Rb1可以顯著抑製線粒體ATP含量下降,可以顯著抑製線粒體DNA ATP酶6亞基、ATP酶8亞基突變,從而推斷PnS Rg1、Rb1可能通過減少線粒體DNA ATP酶6、8亞基突變而抑製線粒體ATP含量下降,從而延緩肝纖維化髮生髮展。
목적：탐토삼칠조대( panax notoginseng saponins,PnS) Rg1、PnS Rb1간예하,간섬유화대서선립체DNA삼린산선감( adenine triphosphate,ATP)매6아기、ATP매8아기적기인돌변화ATP함량변화。방법96지Wistar대서분위대조조、사록화탄( carbon tetrachloride,CCl4)간섬유화대서모형조、PnS Rg1조、PnS Rb1조각24지。제대조조외,기여3조용5％ CCl4감람유안5 ml/kg관위제작간섬유화대서모형。 PnS Rg1조재매차CCl4관위동시복강주사PnSRg1(5 mg/kg)。 PnS Rb1조재매차CCl4관위동시복강주사Rb1(5 mg/kg)。용생물발광법측정각조대서실험전급실험개시2、4、6주말시선립체내ATP함량병비교。제취간세포선립체총RNA,역전록위호보탈양핵당핵산( complementary DNA,cDNA),용기인중조、측서,통과GenBank여선립체 DNA이지서렬진행비교,연구PnS Rg1、PnS Rb1간예6주후선립체 DNA적ATP매6아기7904-8584구역화ATP매8아기7743-7946구역기인돌변정황。결과(1)여대조조상비,모형조대서선립체DNA ATP매6아기7904-8584구역유3처돌변：C8294G、T8505G、G8584A。용Fisher학절개솔법분석,돌변증가유현저성의의( P<0.01)。 PnS Rg1조、PnS Rb1조여대조조상비돌변증가몰유현저성의의( P>0.05)。(2)여대조조상비,ATP매8아기7743-7946구역유2처돌변：A7797non、T7863C。용Fisher학절개솔법분석,돌변증가유현저성의의(P<0.01)。 PnS Rg1、Rb1조여대조조상비돌변증가몰유현저성의의( P>0.05)。(3)각조간섬유화대서선립체 ATP 함량여 CCl4처리시간적상관성분석,용pearson＇s 검험r=-0.935,정부상관。(4)간섬유화대서선립체내ATP함량여ATP매6아기돌변적상관성분석,용pearson＇s 검험r=-0.943,정부상관；간섬유화대서선립체내ATP함량여ATP매8아기돌변적상관성분석,용pearson＇s검험r=-0.961,정부상관。결론 PnS Rg1、Rb1가이현저억제선립체ATP함량하강,가이현저억제선립체DNA ATP매6아기、ATP매8아기돌변,종이추단PnS Rg1、Rb1가능통과감소선립체DNA ATP매6、8아기돌변이억제선립체ATP함량하강,종이연완간섬유화발생발전。
Objective To explore the changes of the mitochondrial DNA ( mt DNA ) adenine triphospholibase(ATPase)6 gene , ATPase 8 gene mutation under the intervention of panax notoginseng saponins(PnS)Rg1, PnS Rb1 for hepatic fibrosis rats which may have theoretical sense in the prevention and cure hepatic fibrosis. Methods 96 Wistar rats divided into contrast group, carbon tetrachloride (CCl4) group, liver fibrosis rats model group, PnS Rg1 group and PnS Rb1group, with 24 rats in each group. Except contrast group, other three groups were reproduced by repeated injection 5% CCl4 solution in rats into liver fibrosis rats model. PnS Rg1 group were also injected 5 ml/kg, PnS Rb1 group were injected 5 ml/kg, and at the same time given 5% CCl4 solution. Extract RNA of hepatic mitochondrial and reverse transcription complementary DNA (cDNA). Gene-recommended and sequencing have been researched. Then further study the PnS Rg1 group, PnS Rb1 group ATPase 6 gene 7904-8584 region and ATPase 8 gene 7743-7946 region gene mutation after 6 weeks. Biological fluorescent method detect ATP content. Re-sults (1)Compare with the contrast group, gene analysis of mtDNA ATPase 6 in hepatic fibrosis of model group rats showed that there were three mutation sites:C8294G, T8505G, C8584A. Using Fisher's exact test to analyze, mutations have increased significantly(P<0. 01). Compare with the contrast group, PnS Rg1 group、PnS Rb1 group mutation increasing does not have marked sense ( P >0. 05 ) . ( 2 ) Compare with the contrast group , the gene analysis of mtDNA ATPase 8 were two mutation sites: A7797non, T7863C. Using Fisher's exact test to analyze, mutations have increased significantly(P<0. 01). (3) Liv-er fibrosis rats mitochondrial DNA ATP of each group with CCl4 manage time correlation analysis, undergo-ing pearson's test r= -0. 935 negative correlation. (4) Liver fibrosis rats mitochondrial DNA ATP of each group with ATPase6 gene mutation has correlation analysis, undergoing pearson's test r= -0. 943 negative correlation. Liver fibrosis rats mitochondrial DNA ATP of each group with ATPase8 gene mutation has cor-relation analysis, undergoing pearson's test r= -0. 961 negative correlation. Conclusion The mutation of mtDNA ATPase 6 gene and ATPase 8 gene may be responsible for hepatic fibrosis development. PnS Rg1, PnS Rb1 were protective function remarkably for the mitochondrial of hepatic fibrosis rats through inhibition of mtDNA ATPase 6 gene, ATPase 8 gene mutation rate. Therefore delay hepatic fibrosis development.