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姜黄素衍生物对肝纤维化小鼠核因子-κB表达的影响

The effect of curcumin derivatives on the nuclear factor-κB expression in mice with hepatic fibrosis

摘要:

目的观察姜黄素衍生物C66的抗小鼠肝纤维化效果及其相关机制。方法33只C57BL/6J小鼠分为正常对照组、模型对照组和C66治疗组。正常对照组小鼠9只,普通饲料和自来水喂饲,其余24只小鼠予腹腔注射40%四氯化碳(CCl4)混合液,首剂量为4 mL/kg ,以后按照剂量2 mL/kg腹腔注射,每周2次,共6周。于造模6周末随机选取6只小鼠,行病理学检测,判断造模是否成功。造模小鼠随机均分成模型对照组和C66治疗组,每组9只。第6周开始,给予治疗组C6610 mg/(kg · d)灌胃,其余组给予等量的0.5%羧甲基纤维素钠溶液灌胃。检测各组小鼠血清ALT、AST 和肝组织羟脯氨酸(Hyp)含量;肝组织行 HE染色及Masson染色,光学显微镜下观察病理学改变,并进行肝纤维化半定量评分;采用实时荧光定量PCR检测Ⅰ型胶原、α‐平滑肌肌动蛋白(α‐SMA) mRNA表达情况;Western印迹法检测各组Ⅰ型胶原、α‐SMA及核因子‐κB p65 p65、核因子‐κB抑制蛋白α(IκBα)蛋白表达情况。样本均数比较采用单因素方差分析。结果模型对照组ALT、AST水平分别为(202.71±19.66) U/L 和(233.42±23.97) U/L ,C66治疗组分别为(102.00±11.04)U/L和(120.87±13.83)U/L,正常对照组分别为(36.66±6.37)U/L和(43.33±8.08)U/L,模型对照组与正常对照组比较差异均有统计学意义(t值分别为23.96和22.39,均 P<0.05),C66治疗组与模型对照组比较差异均有统计学意义(t值分别为11.56和10.52,均 P<0.05)。模型对照组肝组织Hyp含量与正常对照组比较差异有统计学意义(t=17.50,P<0.05),C66治疗组肝组织Hyp含量与模型对照组比较差异有统计学意义(t=11.45,P<0.05)。C66治疗组Ⅰ型胶原和α‐SMA mRNA表达明显下降,与模型对照组比较差异有统计学意义(t值分别为7.23和7.95,均 P<0.05)。与模型对照组相比,C66治疗组Ⅰ型胶原、α‐SMA、核因子‐κB p65蛋白表达下调,同时IκBα蛋白表达上调,差异均有统计学意义(均 P<0.05)。结论 C66具有抗肝纤维化作用,其机制可能是调节核因子‐κB的表达。

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abstracts:

Objective To investigate the therapeutic effects and mechanisms of curcumin derivatives C66 treatment on hepatic fibrosis .Methods Thirty three C57BL/6J mice were randomly divided into 3 groups:normal control group ,model control group and curcumin derivatives C66 treatment group .Nine mice in normal control group were fed with water and food .Hepatic fibrosis model was induced in 24 mice by intraperitoneal injection of 40% carbon tetrachloride at a dose of 4 mL/kg for the first time ,followed by 2 mL/kg twice a week for 6 weeks . At week 6 ,6 mice were randomly selected to perform pathological examination to evaluate whether the hepatic fibrosis were successfully induced .Mice with hepatic fibrosis were randomized into model control group and curcumin derivatives C66 treatment group with 9 mice in each group .From week 6 on ,mice in the treatment group were lavaged with curcumin derivatives C66 at a dosage of 10 mg ·/(kg · d) .The rest mice were administered with equivalent dosage of 0 .5% carboxymethylcellulose sodium .Serum alanine aminotransferase (ALT) ,aspartate&nbsp;aminotransferase (AST ) and liver hydroxyproline ( Hyp ) contents were detected , and the semi‐quantitative analysis of liver fibrosis was performed by pathological examination in hepatic tissue by hematoxylin and eosin (HE) and Masson staining .The expressions of collagen Ⅰ ,α‐smooth muscle actin (α‐SMA) mRNA and collagen Ⅰ ,α‐SMA ,nuclear factor‐kappa B p65 (NF‐κB p65) ,inhibitor kappa B alpha (IκBα) protein in each group were detected by quantitative real‐time polymerase chain reaction (RT‐PCR) and Western blot .Data were analyzed with one‐way ANOVA analysis .Results The serum levels of ALT and AST in model control group ,C66 treatment group and normal control group were (202 .71 ± 19 .66 ) U/L , (233 .42 ± 23 .97 ) U/L ;(102 .00 ± 11 .04 ) U/L , (120 .87 ± 13 .83 ) U/L ;(36 .66 ± 6 .37) U/L and (43 .33 ± 8 .08)U/L ,respectively .The differences between model and normal control group were both significant (t=23 .96 and 22 .39 ,respectively ;both P<0 .05) .The C66 treatment group showed significantly lower levels of serum ALT and AST in contrast with model control group (t =11 .56 and 10 .52 ,respectively ;both P<0 .05) .Compared to the model control group ,hepatic Hyp contents in normal control group and C66 treatment group were significantly different (t= 17 .50 , P< 0 .05;t=11 .45 ,P<0 .05) .Collagen Ⅰand α‐SMA mRNA expressions in C66 treatment group were remarkably lower in contrast with that in model control group (t= 7 .23 and 7 .95 ,respectively ;both P< 0 .05) . Protein levels of Collagen Ⅰ ,α‐SMA and NF‐κB p65 decreased in C66 treatment group ,while IκBαincreased significantly (all P<0 .05) .Conclusion The application of C66 can contribute to the regression of liver fibrosis and the mechanism may rely on the regulation of NF‐κB expression .

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作者: 吴和 [1] 陈永平 [1] 黄思思 [1] 杜珊洁 [1] 陈瑞聪 [1] 董佳佳 [1]
期刊: 《中华传染病杂志》2016年34卷1期 27-31页 ISTICCSCD
栏目名称: 论著
DOI: 10.3760/cma.j.issn.1000-6680.2016.01.007
发布时间: 2016-05-17
基金项目:
国家“十二五”科技重大专项(2013ZX10005002-001) 浙江省自然科学基金(LY15H030017)@@@@National Twelfth Five-year Science and Technology Major Project of China (2013ZX10005002-001) Natural Science Foundation of Zhejiang Province of China
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