摘要
目的 探讨α干扰素-1b调控蛋白激酶Cε（protein kinase Cε,PKCε）蛋白激酶Cε、蛋白激酶Cα（protein kinase Cα,PKCα）抑制肝星状细胞（hepatic stellate cell,HSC）纤维化及机制.方法 采用不同浓度α干扰素-1b（100、200、400、800、1000 U/mL）处理HSC-T6细胞株,同步设置无α干扰素-1b处理的对照组;四甲基偶氮唑盐比色法分析HSC-T6细胞增殖;分析羟脯氨酸含量变化;免疫荧光染色检测PKCε和PKCα的表达,RT-PCR检测PKCε、PKCα、β-链蛋白（β-catenin）、生存素（Survivin）mRNA的水平.蛋白质印迹法检测PKCε、PKCα、β-catenin、Survivin蛋白水平.采用One-Way ANOVA分析.结果 给药24 h后,100、200、400、800、1000 U/mLα干扰素-1b组抑制率分别为（15.85±1.05）％ 、（36.59±1.03）％ 、（45.12±1.05）％ 、（50.00±1.01）％ 和（62.20±1.02）％,组间比较差异有统计学意义（F=27.478,P〈0.01）;48 h抑制率分别为（20.87±1.09）％ 、（43.96±1.08）％ 、（53.85±1.08）％ 、（64.84±1.06）％ 和（74.72±1.07）％,组间比较差异有统计学意义（F=25.321,P〈0.01）.48 h时半抑制浓度为343.47 U/mL.100、200、400 U/mLα干扰素-1b组的羟脯氨酸水平分别为（7.48±0.28）、（6.26±0.17）、（3.86±0.20）μg/mL,均低于对照组的（8.47±0.32）μg/mL,差异均有统计学意义（t值分别为4.033、10.564和21.160,均P〈0.05）.100、200、400 U/mLα干扰素-1b组的PKCε荧光强度均低于对照组,比较差异均有统计学意义（t值分别为1.984、2.457、7.771,均P〈0.05）;PKCα的荧光强度分别也均低于对照组,比较差异均有统计学意义（t值分别为9.232、15.921、22.222,均P〈0.01）.随着α干扰素-1b水平的增加,HSC-T6细胞PKCε、PKCα、β-catenin、Survivin mRNA水平较对照组明显下降,比较差异均有统计学意义（t值分别为7.020、24.562、45.701和14.241,均P〈0.01）.随着α干扰素-1b水平的增加,HSC-T6细胞PKCε、PKCα、β-catenin和Survivin�
Objective To investigate the regulation of interferon α-1b （IFNα-1b） on protein kinase Cε（PKCε） and protein kinase Cα（PKCα） which inhibit the fibrosis of hepatic stellate cells （HSC） ,and to explore its mechanism .Methods HSC-T6 cells were treated with different levels of IFNα-1b （100 , 200 ,400 ,800 and 1000 U/mL） and the proliferation of HSC-T6 cells was analyzed by methyl thiazol tetrazolium （MTT） assay .Changes of hydroxyproline level were analyzed .The expressions of PKCεand PKCαwere detected by immunofluorescence staining . PKCε, PKCα,β-catenin and Survivin mRNA levels were detected by RT-PCR . PKCε, PKCα,β-catenin and Survivin protein levels were detected by Western blot . Variance analysis was conducted by using one-way ANOVA approach . Results The inhibition rates of 100 , 200 , 400 , 800 and 1000 U/mL IFNα-1b treatment after 24 hours of administration were （15 .85 ± 1 .05）% ,（36 .59 ± 1 .03）% ,（45 .12 ± 1 .05）% ,（50 .00 ± 1 .01）% and （62 .20 ± 1 .02）% ,respectively ,with statistically significant differences among groups （F=27 .478 , P〈0 .01） .The 48h inhibition rates were （20 .87 ± 1 .09）% ,（43 .96 ± 1 .08）% ,（53 .85 ± 1 .08）% ,（64 .84 ± 1 .06）% and （74 .72 ± 1 .07）% ,respectively ,with statistically significant differences among groups （F=25 .321 , P〈 0 .01 ） . half maximal inhibitory concentration at 48 h was 343 .47 U/mL . The levels of hydroxyproline in 100 ,200 and 400 U/mL IFNα-1b groups were （7 .48 ± 0 .28） ,（6 .26 ± 0 .17） and （3 .86 ± 0 .20） μg/mL ,respectively ,which were lower than that in control group （8 .47 ± 0 .32） μg/mL .The differences were all statistically significant （t=4 .033 ,10 .564 and 21 .160 ,respective ,all P〈0 .05） .The fluorescence intensities of PKCεin 100 ,200 and 400 U/mL IFNα-1b groups were all lower than that of control group .The differences were statistically significant （t=1 .984 ,2 .457 and 7 .771 ,respectively ,all P〈0 .05） .Th
作者
秦文燕
李国军
林琪
宋凌云
沈萍萍
宋章章
许春芳
Qin Wenyan;Li Guojun;Lin Qi;Song Lingyun;Shen Pinpin;Song Zhangzhang;Xu Chunfang(Department of Gastroenterology, the First Affiliated Hospital of Soochow University, Suzhou 215006, China)
出处
《中华传染病杂志》
CAS
CSCD
2018年第3期145-149,共5页
Chinese Journal of Infectious Diseases
基金
2016年浙江省医药卫生一般研究项目（2016KYA162）
2016年宁波市科技重大专项肝病基金（2016C51008）
关键词
IFNα—1b
肝星状细胞
增殖
分子机制
IFNα-1b
hepatic stellate cells
proliferation
molecular mechanism
