中华肝脏病杂志
中華肝髒病雜誌
중화간장병잡지
CHINESE JOURNAL OF HEPATOLOGY
2013年
12期
914-919
,共6页
谢聪%李龙%许雅苹%朱月永%江家骥
謝聰%李龍%許雅蘋%硃月永%江傢驥
사총%리룡%허아평%주월영%강가기
肝硬化%炎症因子%超氧化物歧化酶%非诺贝特%过氧化酶增殖物激活受体
肝硬化%炎癥因子%超氧化物歧化酶%非諾貝特%過氧化酶增殖物激活受體
간경화%염증인자%초양화물기화매%비낙패특%과양화매증식물격활수체
Liver cirrhosis%Inflammatory cytokines%Superoxide dismutase%Fenofibrate%PPARα receptor
目的 观察过氧化酶增殖物激活受体(PPAR α)配体非诺贝特对四氯化碳(CCl4)诱导的小鼠肝纤维化的作用,并探讨其可能的机制.方法 26只C57BL雄性小鼠随机分为3组:正常对照组(n=6)、模型对照组(n=10)、非诺贝特治疗组(n=10).以CCl4腹腔注射制备小鼠肝纤维化模型;造模的同时,以非诺贝特进行干预治疗;以橄榄油腹腔注射作为空白对照组.小鼠均在第5周末处死,取其血清及肝脏组织.采用比色法检测血清中ALT、AST含量;放射免疫分析法测定纤维化血清标志物透明质酸(HA)、Ⅲ型前胶原氨基端肽(PⅢNP)含量的变化;紫外分光光度法测肝组织中羟脯氨酸(HYP)、丙二醛(MDA)、超氧化物歧化酶(SOD)的含量;Real-tiime PCR技术测定肝脏中纤维化相关因子α-平滑肌肌动蛋白(α-SMA)、转化生长因子β1(TGF β l)、Ⅰ型胶原,炎症因子肿瘤坏死因子(TNF)α、白细胞介素6(IL-6)及PPAR-αmRNA表达量.另取肝组织作冰冻切片,行苏木素-伊红(HE)、天狼星红染色观察肝脏组织的病理变化.数据以均数±标准差(-X±s)表示,多组间比较方法采用单因素方差分析(one-way ANOVA)检验.结果 5周后,非诺贝特治疗组与模型对照组相比,小鼠血清中ALT、HA、PⅢNP的含量有明显降低[(38.72±1.25) U/L对比(55.72±1.20)U/L,P＜0.01; (152.9±13.06) μg/L对比(236.2±17.57) μg/L,P＜0.01;(34.32±1.53)μg/L对比(41.66±1.89) μg/L,P＜0.05];而小鼠肝组织中SOD水平明显升高[(101.1±5.32)U/mg对比(67.0±4.65)U/mg,P＜0.01],MDA含量下降[(10.17±0.60) nmol/mg对比(14.67±0.93) nmol/mg,P＜0.01).Real-time PCR结果显示,非诺贝特治疗组与模型对照组相比,肝脏中PPARαmRNA表达上调(相对表达量为0.30±0.03对比0.18±0.03,P＜0.01),而α-SMA、TGFβ 1、Ⅰ型胶原mRNA表达均下降(相对表达量为6.83±0.88对比11.57±1.31,P＜ 0.05;67.83±4.65对比112.30±4.81,P＜0.01 ; 164.0±14.74对比232.5±12.37,P＜0.01);同时非诺贝特治疗组肝组织中TNFα、IL-6 mRNA的相对表达量也减少(17.43±2.32对比37.83±4.69,P＜ 0.01 ; 4.0±0.49对比5.62±0.54,P＜0.05).病理切片显示非诺贝特治疗组较模型对照组肝脏炎症及纤维化程度有明显改善,天狼猩红染色肝纤维半定量评分为3.4±0.3对比9.6±0.8,P＜0.01;同时肝组织中HYP含量分别为(0.41±0.5)mg/g对比(0.67±0.8) mg/g,P＜ 0.01.结论 非诺贝特可抑制CCI4诱导的C57BL小鼠肝纤维化程度,其机制可能与上调肝脏中PPARα表达,抑制炎症反应,升高SOD发挥抗氧化作用有关.
目的 觀察過氧化酶增殖物激活受體(PPAR α)配體非諾貝特對四氯化碳(CCl4)誘導的小鼠肝纖維化的作用,併探討其可能的機製.方法 26隻C57BL雄性小鼠隨機分為3組:正常對照組(n=6)、模型對照組(n=10)、非諾貝特治療組(n=10).以CCl4腹腔註射製備小鼠肝纖維化模型;造模的同時,以非諾貝特進行榦預治療;以橄欖油腹腔註射作為空白對照組.小鼠均在第5週末處死,取其血清及肝髒組織.採用比色法檢測血清中ALT、AST含量;放射免疫分析法測定纖維化血清標誌物透明質痠(HA)、Ⅲ型前膠原氨基耑肽(PⅢNP)含量的變化;紫外分光光度法測肝組織中羥脯氨痠(HYP)、丙二醛(MDA)、超氧化物歧化酶(SOD)的含量;Real-tiime PCR技術測定肝髒中纖維化相關因子α-平滑肌肌動蛋白(α-SMA)、轉化生長因子β1(TGF β l)、Ⅰ型膠原,炎癥因子腫瘤壞死因子(TNF)α、白細胞介素6(IL-6)及PPAR-αmRNA錶達量.另取肝組織作冰凍切片,行囌木素-伊紅(HE)、天狼星紅染色觀察肝髒組織的病理變化.數據以均數±標準差(-X±s)錶示,多組間比較方法採用單因素方差分析(one-way ANOVA)檢驗.結果 5週後,非諾貝特治療組與模型對照組相比,小鼠血清中ALT、HA、PⅢNP的含量有明顯降低[(38.72±1.25) U/L對比(55.72±1.20)U/L,P＜0.01; (152.9±13.06) μg/L對比(236.2±17.57) μg/L,P＜0.01;(34.32±1.53)μg/L對比(41.66±1.89) μg/L,P＜0.05];而小鼠肝組織中SOD水平明顯升高[(101.1±5.32)U/mg對比(67.0±4.65)U/mg,P＜0.01],MDA含量下降[(10.17±0.60) nmol/mg對比(14.67±0.93) nmol/mg,P＜0.01).Real-time PCR結果顯示,非諾貝特治療組與模型對照組相比,肝髒中PPARαmRNA錶達上調(相對錶達量為0.30±0.03對比0.18±0.03,P＜0.01),而α-SMA、TGFβ 1、Ⅰ型膠原mRNA錶達均下降(相對錶達量為6.83±0.88對比11.57±1.31,P＜ 0.05;67.83±4.65對比112.30±4.81,P＜0.01 ; 164.0±14.74對比232.5±12.37,P＜0.01);同時非諾貝特治療組肝組織中TNFα、IL-6 mRNA的相對錶達量也減少(17.43±2.32對比37.83±4.69,P＜ 0.01 ; 4.0±0.49對比5.62±0.54,P＜0.05).病理切片顯示非諾貝特治療組較模型對照組肝髒炎癥及纖維化程度有明顯改善,天狼猩紅染色肝纖維半定量評分為3.4±0.3對比9.6±0.8,P＜0.01;同時肝組織中HYP含量分彆為(0.41±0.5)mg/g對比(0.67±0.8) mg/g,P＜ 0.01.結論 非諾貝特可抑製CCI4誘導的C57BL小鼠肝纖維化程度,其機製可能與上調肝髒中PPARα錶達,抑製炎癥反應,升高SOD髮揮抗氧化作用有關.
목적 관찰과양화매증식물격활수체(PPAR α)배체비낙패특대사록화탄(CCl4)유도적소서간섬유화적작용,병탐토기가능적궤제.방법 26지C57BL웅성소서수궤분위3조:정상대조조(n=6)、모형대조조(n=10)、비낙패특치료조(n=10).이CCl4복강주사제비소서간섬유화모형;조모적동시,이비낙패특진행간예치료;이감람유복강주사작위공백대조조.소서균재제5주말처사,취기혈청급간장조직.채용비색법검측혈청중ALT、AST함량;방사면역분석법측정섬유화혈청표지물투명질산(HA)、Ⅲ형전효원안기단태(PⅢNP)함량적변화;자외분광광도법측간조직중간포안산(HYP)、병이철(MDA)、초양화물기화매(SOD)적함량;Real-tiime PCR기술측정간장중섬유화상관인자α-평활기기동단백(α-SMA)、전화생장인자β1(TGF β l)、Ⅰ형효원,염증인자종류배사인자(TNF)α、백세포개소6(IL-6)급PPAR-αmRNA표체량.령취간조직작빙동절편,행소목소-이홍(HE)、천랑성홍염색관찰간장조직적병리변화.수거이균수±표준차(-X±s)표시,다조간비교방법채용단인소방차분석(one-way ANOVA)검험.결과 5주후,비낙패특치료조여모형대조조상비,소서혈청중ALT、HA、PⅢNP적함량유명현강저[(38.72±1.25) U/L대비(55.72±1.20)U/L,P＜0.01; (152.9±13.06) μg/L대비(236.2±17.57) μg/L,P＜0.01;(34.32±1.53)μg/L대비(41.66±1.89) μg/L,P＜0.05];이소서간조직중SOD수평명현승고[(101.1±5.32)U/mg대비(67.0±4.65)U/mg,P＜0.01],MDA함량하강[(10.17±0.60) nmol/mg대비(14.67±0.93) nmol/mg,P＜0.01).Real-time PCR결과현시,비낙패특치료조여모형대조조상비,간장중PPARαmRNA표체상조(상대표체량위0.30±0.03대비0.18±0.03,P＜0.01),이α-SMA、TGFβ 1、Ⅰ형효원mRNA표체균하강(상대표체량위6.83±0.88대비11.57±1.31,P＜ 0.05;67.83±4.65대비112.30±4.81,P＜0.01 ; 164.0±14.74대비232.5±12.37,P＜0.01);동시비낙패특치료조간조직중TNFα、IL-6 mRNA적상대표체량야감소(17.43±2.32대비37.83±4.69,P＜ 0.01 ; 4.0±0.49대비5.62±0.54,P＜0.05).병리절편현시비낙패특치료조교모형대조조간장염증급섬유화정도유명현개선,천랑성홍염색간섬유반정량평분위3.4±0.3대비9.6±0.8,P＜0.01;동시간조직중HYP함량분별위(0.41±0.5)mg/g대비(0.67±0.8) mg/g,P＜ 0.01.결론 비낙패특가억제CCI4유도적C57BL소서간섬유화정도,기궤제가능여상조간장중PPARα표체,억제염증반응,승고SOD발휘항양화작용유관.
Objective To investigate the anti-fibrosis effects and mechanisms of fenofibrate on hepatic fibrosis using a mouse model of fibrosis induced by carbon tetrachloride (CCI4).Methods Twenty-six male C57BL mice were divided into the following three groups:CCL4-induced untreated model control (n =10),CCl4-induced fenofibrate-treated model (n =10),and uninduced/untreated normal control (n =6).All animals were sacrificed after the 5 weeks of induction and treatment.Serum levels of alanine aminotransferase (ALT),aspartate aminotransferase (AST),hyaluronic acid (HA) and procollagen Ⅲ amino-terminal peptide (PIIINP) were determined by routine biochemistry assays.Liver content of hydroxyproline (HYP) was measured by spectrophotometry.Liver content of malonic aldehyde (MDA) and superoxide dismutase (SOD) was measured by enzymatic assays.mRNA expression levels of liver fibrosis-associated factors were determined by PCR,and included alpha-smooth muscle actin (a-SMA),transforming growth factor-β1 (TGFβ1),type Ⅰ collagen-alpha (Collagenlα),peroxisome proliferatoractivated receptor-alpha (PPARα),and the inflammatory cytokines tumor necrosis factor alpha (TNFα) and interleukin-6 (IL-6).Finally,the degree of inflammation and fibrosis were assessed by histological analysis using hematoxylin-eosin and Sirius red staining.Results Compared to the untreated model group,the fenofibrate-treated model group showed significantly lower levels of serum ALT (55.72 ± 1.20 vs.38.72 ± 1.25IU/L),HA (236.20 ± 17.57 vs.152.9 ± 13.06 μg/L) and PmNP (41.66 ± 1.89 vs.34.32 ± 1.53 μg/L) (all P ＜ 0.05).The fenofibrate-treated group also showed a significantly higher level of hepatic SOD content (untreated model:67.00 ± 4.65 vs.101.1 ± 5.32) but significantly lower level of hepatic MDA content (14.67 ± 0.93 vs.10.17 ± 0.60nmol/mg) and lower level of hepatic HYP content (0.67 ± 0.80 vs.0.41 ± 0.50 mg/g) (all,P ＜ 0.05).In addition,the fenofibrate-treated group showed significantly reduced mRNA expression levels of α-SMA (6.83 ± 0.88 vs.untreated model:11.57 ± 1.31),TGFβ1 (67.83 ± 4.65 vs.112.30 ± 4.81),Collagen1α (67.83 ± 4.65 vs.112.30 ±4.81),TNFα (17.43 ± 2.32 vs.37.83 ± 4.69),and IL-6 (4.00 ± 0.49 vs.5.62 ± 0.54),but significantly increased PPARα (0.30 ± 0.03 vs.0.18 ± 0.03) (all,P ＜ 0.05).Finally,the degree of CCL4-induced hepatic fibrosis was attenuated by the fenofibrate treatment.Conclusion Fenofibrate can reduce the degree of liver fibrosis in mice induced by CCi4.The mechanism may involve up-regulation of PPARα,inhibition of the inflammatory response,and enhancement of SOD antioxidant activity.
