摘要
目的:观察将不同剂量的重组质粒pLXSN-hBD2转染大鼠子宫内膜异位症(EMS)模型后,其局部异位病灶组织中hBD-2蛋白表达的特点,以明确动物实验的疗效观察中重组质粒的剂量和时间。方法:构建重组质粒pLXSN-hBD2后,根据不同质粒的剂量,将大鼠EMS模型随机分为2μg组和0.2μg组,分别在大鼠异位病灶局部多点注射2μg和0.2μg pLXSN-hBD2,采用PCR、免疫印迹法检测异位病灶组织内基因组DNA和hBD-2蛋白表达,进行外源基因转染整合的鉴定。同时观察不同剂量pLXSN-hBD2转染后2周内(2、7、14 d)局部病灶组织中hBD-2的蛋白表达变化。结果:转染2 d后,大鼠EMS模型局部异位病灶组织中检测到明显的hBD-2蛋白的表达,其表达水平随着时间的延长而逐渐衰减,但在转染后2、7、14 d,2μg组的hBD-2蛋白表达均明显高于相应的0.2μg组。结论:通过局部多点注射的方法,可将携带hBD-2基因的重组质粒pLXSN-hBD2成功转染至大鼠EMS异位病灶组织中,并获得至少持续约2周的hBD-2的蛋白表达,2μg重组质粒可用于动物实验的疗效观察。
Objective ： To explore the character of protein expression of human defense-2 （hBD-2） on ectopic focus of endometriosis model in rats after the different dose of recombinant plasmid expressing hBD-2 was trans- fered. And to definite the appropriate dose and time of recombinant plasmid in animal experiment. Methods： After the recombinant plasmid pLXSN-hBD2 containing hBD-2 （295bp） was built, 2μg and 0. 2μg pLXSN-hBD2 were injected by multi-spots into every endometriosis focus of establishing endometriosis models in SD rats, respectively. And then, pLXSN-hBD2 DNA and protein were detected in ectopic endometrium by PCR and Western-blot meth- od. At last, the variation of hBD-2 protein was observed during two weeks after transfection. Results： The expres- sions of hBD-2 protein were detected in ectopic focus after rats were transfected for two days, but gradually de- creased over time. The expressions of hBD-2 protein of 2μg group were higher than those of 0.2μg group 2, 7, 14 days after transfection. Conclusions： The hBD-2 gene and protein could be transferred into endometriosis tissues of rat models successfully and expressed effectively by single local muti-spots injection method for two weeks at least. And 2μg pLXSN-hBD2 can be used in animal experiment for the curative effect observation.
出处
《新医学》
2013年第5期341-346,共6页
Journal of New Medicine
基金
国家自然科学基金(81070472)
关键词
防御素-2
重组质粒
基因转染
表达
剂量
Defensin-2
Recombinant plasmid
Gene transfection
Expression
Dose
