组织蛋白酶B抑制剂CA-074Me对多发性肌炎豚鼠模型肌纤维保护机制的研究
조직단백매B억제제CA-074Me대다발성기염돈서모형기섬유보호궤제적연구
Protective mechanisms of a cathepsin B inhibitor, CA-074 Me, on muscle fibers in coxsackievirus B1-induced polymyositis in guinea pigs: an experimental study
  • 万方数据
    中华皮肤科杂志 중화피부과잡지
    Chinese Journal of Dermatology
    2013年 2期 100-104 ,共5页

    倪立燕%王强

    예립연%왕강

    多发性肌炎%组织蛋白酶B%细胞凋亡%CD8阳性T淋巴细胞 多髮性肌炎%組織蛋白酶B%細胞凋亡%CD8暘性T淋巴細胞
    다발성기염%조직단백매B%세포조망%CD8양성T림파세포
    Polymyositis%Cathepsin B%Apoptosis%CD8-positive T-lymphocytes
    目的 探讨组织蛋白酶B抑制剂CA-074Me对组织蛋白酶B的影响,以阐明其对肌纤维的保护机制.方法 40只健康雌性短毛英国种豚鼠,随机分为5组,每组8只,分别为:阳性对照γ干扰素组、PM模型组、健康组、假干预组(即生理氯化钠溶液对照组)和阴性对照CA-074Me组.相应时间点处死动物,留取血液检测肌酸激酶(CK)及其同工酶(CK-MM)、门冬氨酸氨基转移酶(AST)、乳酸脱氢酶(LDH)变化;取肌肉组织进行组织病理学观察(HE染色);用免疫组化法(Envision二步法)分别检测CD8+细胞、组织蛋白酶B的表达水平;原位末端标记法(TUNEL)检测各组豚鼠骨骼肌细胞的凋亡变化.肌酶、肌肉炎症程度评分、肌细胞凋亡的比较均采用单因素方差分析(ANOVA);CD8+T细胞、组织蛋白酶B表达的结果采用Pearsonx2检验.结果 PM模型组、假干预组、γ干扰素组、CA-074 Me组肌酶谱(CK、CK-MM、AST和LDH)均异常升高,以CK升高最为明显,4个组的CK分别为(3537.3±2141.6) U/L、(2222.0±226.9) U/L、(973.8±423.2) U/L、(814.0±268.4) U/L,均高于健康组(410.7±167.9) U/L (P＜ 0.05);γ干扰素组、CA-074 Me组的肌酶谱呈低水平升高,与假干预组比较,差异均有统计学意义(P＜0.05).PM模型组、假干预组、γ干扰素组、CA-074Me组的骨骼肌炎症程度评分分别为1.75±0.50、1.40±0.55、2.38±0.74和1.20±0.45,均较健康组明显增加,尤其以γ干扰素组炎症细胞浸润明显(P＜0.05).与健康组相比,PM模型组、假干预组、γ干扰素组、CA-074Me组的CD8+细胞、组织蛋白酶B表达增加,凋亡指数升高,差异均有统计学意义(P＜0.05);CA-074 Me组的CD8+细胞(42.3±27.4)个、组织蛋白酶B表达(31.3±6.7)个均低于假干预组[分别为(68.0±13.2)个和(37.5±9.2)个],P值均＜0.05;干扰素组组织蛋白酶B表达(49.3±17.0)个、凋亡指数(40.1±6.7)个均较假干预组[分别为(37.5±9.2)和(25.4±5.0)个]升高(P＜0.05).结论 组织蛋白酶B在PM豚鼠模型中存在着高表达,CA-074 Me通过抑制组织蛋白酶B表达减少其介导的炎症和细胞凋亡,起到对PM模型豚鼠肌组织的保护作用.
    목적 탐토조직단백매B억제제CA-074Me대조직단백매B적영향,이천명기대기섬유적보호궤제.방법 40지건강자성단모영국충돈서,수궤분위5조,매조8지,분별위:양성대조γ간우소조、PM모형조、건강조、가간예조(즉생리록화납용액대조조)화음성대조CA-074Me조.상응시간점처사동물,류취혈액검측기산격매(CK)급기동공매(CK-MM)、문동안산안기전이매(AST)、유산탈경매(LDH)변화;취기육조직진행조직병이학관찰(HE염색);용면역조화법(Envision이보법)분별검측CD8+세포、조직단백매B적표체수평;원위말단표기법(TUNEL)검측각조돈서골격기세포적조망변화.기매、기육염증정도평분、기세포조망적비교균채용단인소방차분석(ANOVA);CD8+T세포、조직단백매B표체적결과채용Pearsonx2검험.결과 PM모형조、가간예조、γ간우소조、CA-074 Me조기매보(CK、CK-MM、AST화LDH)균이상승고,이CK승고최위명현,4개조적CK분별위(3537.3±2141.6) U/L、(2222.0±226.9) U/L、(973.8±423.2) U/L、(814.0±268.4) U/L,균고우건강조(410.7±167.9) U/L (P＜ 0.05);γ간우소조、CA-074 Me조적기매보정저수평승고,여가간예조비교,차이균유통계학의의(P＜0.05).PM모형조、가간예조、γ간우소조、CA-074Me조적골격기염증정도평분분별위1.75±0.50、1.40±0.55、2.38±0.74화1.20±0.45,균교건강조명현증가,우기이γ간우소조염증세포침윤명현(P＜0.05).여건강조상비,PM모형조、가간예조、γ간우소조、CA-074Me조적CD8+세포、조직단백매B표체증가,조망지수승고,차이균유통계학의의(P＜0.05);CA-074 Me조적CD8+세포(42.3±27.4)개、조직단백매B표체(31.3±6.7)개균저우가간예조[분별위(68.0±13.2)개화(37.5±9.2)개],P치균＜0.05;간우소조조직단백매B표체(49.3±17.0)개、조망지수(40.1±6.7)개균교가간예조[분별위(37.5±9.2)화(25.4±5.0)개]승고(P＜0.05).결론 조직단백매B재PM돈서모형중존재착고표체,CA-074 Me통과억제조직단백매B표체감소기개도적염증화세포조망,기도대PM모형돈서기조직적보호작용.
    Objective To evaluate the effect of a specific cathepsin B inhibitor,CA-074 Me,on the expression of cathepsin B in coxsackievirus B1-induced polymyositis in guinea pigs,and to elucidate the protective mechanisms of CA-074 Me on muscle fibers.Methods Polymyositis model was established in 32 guinea pigs by infection with coxsackievirus B1,which were then equally divided into 4 groups: γ-interferon group treated with intraperitoneal γ-interferon (150 000 IU per kilogram per day) from week 5 to week 8,polymyositis model group receiving no treatment,pseudo-intervention group treated with intraperitoneal sodium chloride physiological solution,CA-074 Me group treated with intraperitoneal CA-074 Me (4 mg per kilogram per day) for 7 days,after the infection with coxsackievirus B1.Eight guinea pigs receiving no infection or treatment served as the healthy control group.Blood samples and muscle tissue samples were obtained from the guinea pigs in the γ-interferon group on week 8 and in the other 4 groups on week 5.The serum level of muscle enzymes including creatine kinase (CK),CK-MM,aspartic transaminase (AST) and l-lactate dehydrogenase (LDH) were determined.Muscle tissue samples were studied by hematoxylin and eosin (HE) staining,and Envision two-step method was used to quantify the expression of cathepsin B and to numerate CD8+ T cells.The apoptosis in muscle cells was detected by terminal deoxynucleotide transferase-mediated dUTP-biotin nick end labeling (TUNEL).One-way analysis of variance (ANOVA) was conducted to compare the serum level of muscle enzymes,inflammation score of muscle and apoptosis index of muscular cells,and Pearson chi-square test to compare the count of CD8 + T cells and cathepsin B expression,among these groups.Results Polymyositis model was successfully established by infection with coxsackievirus B1 in the 32 guinea pigs with a marked increase in the serum level of the tested enzymes,especially in that of CK.In detail,the serum level of CK was (410.7 ±167.9) U/L in the healthy control group,significantly lower than that in the polymyositis model group ((3537.3 ± 2141.6) U/L,P ＜ 0.05),pseudo-intervention group ((2222.0 ± 226.9) U/L,P ＜ 0.05),γ-interferon group ((973.8 ± 423.2) U/L,P＜ 0.05) and CA-074 Me group ((814.0 ± 268.4) U/L,P＜ 0.05).Compared with the pseudo-intervention group,the γ-interferon group and CA-074 Me group showed a slight increase in the serum level of all the four enzymes (all P ＜ 0.05).There was a significant elevation in the inflammation score of skeletal muscles in the polymyositis model group,pseudo-intervention group,γ-interferon group and CA-074 Me group compared with the healthy control group (1.75 ± 0.50,1.40 ± 0.55,2.38 ± 0.74 and 1.20 ± 0.45 vs.0.00 ± 0.00,all P ＜ 0.05),with the most intense infiltration of inflammatory cells observed in the γ-interferon group.Moreover,the number of CD8 + T cells,cathepsin B expression and muscular cell apoptosis index were all significantly higher in the polymyositis model group,pseudo-intervention group,γ-interferon group and CA-074 Me group than in the healthy control group (all P ＜ 0.05).Compared with the pseudo-intervention group,the CA-074 Me group showed less CD8+ T cells (42.3 ± 27.4 vs.68.0 ± 13.2,P ＜ 0.05) and lower expression of cathepsin B (31.3 ± 6.7 vs.37.5 ± 9.2,P ＜ 0.05),whereas the γ-interferon group exhibited elevated cathepsin B expression (49.3 ± 17.0 vs.37.5 ± 9.2,P＜ 0.05) and apoptosis index (40.1 ± 6.7 vs.25.4 ± 5.0,P＜ 0.05).Conclusions Cathepsin B is highly expressed in the guinea pig model of polymyositis,while CA-074 Me may protect muscle tissue in this model by downregulating the expression of cathepsin B and attenuating the inflammation and apoptosis induced by cathepsin B.
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