Ⅰ型和Ⅲ型前胶原基因核酶抑制增生性瘢痕的实验  

Inhibition of hyperplastic scar with ribozymes target in typeⅠand Ⅲ pre-collagen

作　　者：朱斌[1] 朱家源[1] 张涛[2] 唐冰[1] 李新强[2] 陈东[1] 张伟[1] 李爽[1] 

机构地区：[1]中山大学附属第一医院烧伤科,广东省广州市510080 [2]中山大学附属第一医院

出　　处：《中国临床康复》2006年第28期85-87,F0003,共4页Chinese Journal of Clinical Rehabilitation

基　　金：国家自然科学基金(39770842)~~

摘　　要：目的:在细胞及动物体内证实前期已合成,并证实其活性的、针对I型和Ⅲ型前胶原基因的核酶的有效性。方法:实验于2004-01/08在中山大学附属第一医院烧伤科、中山大学附属第一医院外科实验室、广州市红十字会医院创伤研究所、中山大学医学院动物实验中心完成。选取80只SPF级4～6周龄裸鼠,体质量15～25g。随机分为注射Ⅰ型前胶原核酶A组、注射Ⅰ型前胶原核酶B组、注射Ⅲ型前胶原核酶C组、注射Ⅲ型前胶原核酶D组、注射Ⅲ型前胶原核酶E组、注射Ⅲ型前胶原核酶F组、对照组G组和空白对照组H组,每组10只。注射Ⅰ型及Ⅲ型前胶原核酶A-F组,7d。对照组G组注射生理盐水7d。应用苦味酸-天狼猩红染色偏振光法,观察分析各组胶原相对含量及分布的改变。同期选取增生性瘢痕来自烧伤愈后、瘢痕增生半年之患者手术切除标本。分为甲、乙、丙、丁、戊、己六种浓度组及空白对照组庚组。脂质体包裹核酶转染实验组培养细胞,测定培养上清羟脯氨酸含量、反转录多聚链反应法测量细胞内Ⅰ型和Ⅲ型胶原mRNA含量。结果:①细胞上清羟脯氨酸含量:甲-已组明显低于庚组(2.33±0.04,2.32±0.04,2.42±0.04,2.41±0.05,2.20±0.03,2.12±0.04,2.85±0.07)μg/L,P<0.01。②I型胶原mRNA表达甲-已组明显低于庚组:(0.796±0.034,0.834±0.017,0.860±0.026,0.838±0.023,0.842±0.031,0.858±0.037,0.940±0.037)μg/L,P<0.05。③Ⅲ型胶原蛋白mRNA表达:甲-已组明显低于庚组:(1.496±0.039,1.668±0.052,1.154±0.093,1.078±0.093,1.270±0.060,1.182±0.111,2.001±0.099)μg/L。④应用核酶前后瘢痕体积变化:A组、G组、H组实验前明显高于实验后[(28.31±2.10,25.60±1.20)(33.65±1.76,32.71±3.92)(29.53±2.50,28.80±2.71)mm3]。结论:所合成及扩增、转染的核酶可抑制人瘢痕成纤维细胞合成胶原,并能有效减少人增生性瘢痕裸鼠动物模型中瘢痕的胶原含量。AIM： To confirm the efficiency of Ribozymes built in cell and animal models target on type I and m pre-collagen. METHODS： The experiment was conducted in the Department of Bum and laboratory of Department of Surgery, First Hospital affiliated to the Sun Yat-sen University, Institute of Wound, Guangzhou Red Cross Hospital, Animal Experimental Center, Medical College of Sun Yat-sen University from January to August 2004. A total of 80 SPF nude mice aged 4-6 weeks, body mass 15-25 g were selected and randomly divided into type Ⅰ pre-collagen ribozymes A （rgⅠA） group, rgⅠB, rgⅢC, rgⅢD, rgⅢE, rgⅢF, control G, blank control H groups with 10 mice in each group. RgⅠ and rg Ⅲ A-F groups were injected with different concentrations of ribozymes, respectively for 7 days, and the G group with normal saline for 7 days. The total collagen synthesis and changes of distribution were assessed by picrosirius-pelarization method. Meanwhile, the excised samples of the hypertrophic scar from bum healing and hypertrophic lasted for half a year were selected and divided into group A1, B1, C1, D1, E1 and F1 and blank control group G1. Supematant bydroxyproline （HYP） content of cultured fibroblast cells was determined, and content of type Ⅰ and Ⅲ pre-collogen mRNA were measured by RT-PCR. RESULTS： （1）HYP content： The content of the group A1 to F1 were obviously lower than the group G1 [（2.33±0.04）, （2.32±0.04）, （2.42±0.0405）, （2.41±0.05）, （2.20±0.03）, （2.12±0.04）, （2.85±0.07） μg/L, P 〈 0.01]. （2） Expressions of collagen type Ⅰ mRNA of the A1 to F1 groups were significantly lower than the group G1 [（0.796±0.034）, （0.834±0.017）, （0.860±0.026）, （0.838±0.023）, （0.842±0.031）, （0.858 ±0.037）, （0.940 ±0.037）μg/L, P 〈 0.05]. （3）Expression of collagen type Ⅲ： The group At to F1 were lower than the group G1 [（1.496±0.039）, （1.668±0.052）, （1.154 ±0.093）, （1.078±0.093）, （1.270±0.060）, （1.18

关 键 词：瘢痕 成纤维细胞 胶原 核酶 动物模型 

分 类 号：R619.6[医药卫生—外科学]