摘要目的 观察靶向NF-κB基因的短发夹状RNA(shRNA)在治疗食蟹猴子宫内膜异位症的效果,分析NF-κB基因在食蟹猴子宫内膜异位症发生发展中的作用.方法 构建食蟹猴子宫内膜异位症模型,同时合成高特异性的靶向NF-κB基因的shRNA腺病毒及其阴性对照空载shRNA腺病毒.将建模成功的食蟹猴分成实验组、阴性对照组及单纯建模组,将NF-κBshRNA腺病毒在腹腔镜下直接注入实验组食蟹猴子宫内膜异位病灶内,阴性对照组在病灶内注射空载shRNA腺病毒,单纯建模组病灶内注射生理盐水.注射腺病毒后4周,腹腔镜下观察病灶变化,取各组子宫内膜异位病灶进行CD34免疫组化染色检测微血管密度,并采用蛋白印迹法(western blot)检测NF-κB及增殖细胞核抗原(PCNA)蛋白的表达.结果 成功构建了食蟹猴子宫内膜异位症模型.实验组食蟹猴异位病灶较前萎缩,其微血管密度(0.002 0±0.000 3)较阴性对照组(0.021 9±0.002 6)、单纯建模组(0.024 5±0.003 3)明显减小,分别比较,差异有统计学意义(P＜0.01).实验组食蟹猴NF-κB蛋白的表达水平(0.338±0.174)较阴性对照组(0.678±0.021)、单纯建模组(0.645±0.098)明显下降,分别比较,差异有统计学意义(P＜0.01).实验组食蟹猴PCNA蛋白的表达水平(0.37±0.17)低于阴性对照组(0.57±0.26)、单纯建模组(0.57±0.28),分别比较,差异有统计学意义(P＜0.01).结论 NF-κB shRNA通过靶向抑制NF-κB基因的表达后,能够抑制食蟹猴子宫内膜异位症的进展,抑制子宫内膜异位病灶的血管生成能力,减弱异位子宫内膜细胞的增殖能力.

abstractsObjective To observe the therapeutic effect of NF-κB gene short hairpin RNA (shRNA) on endometriosis and identify the function of NF-κB on the maintenance and development of endometriosis in Macaca fascicularis.Methods The Macaca fascicularis model of endometriosis was developed,which divided into experimental group,negative control group and simple model group.The high specificity adenovirus vector mediated shRNA targeting NF-κB gene and negative control shRNA adenovirus with no-load NF-κB gene were synthesised.The experimental group injected the adenovirus which carried the NF-κB shRNA into the endometriosis lesions under laparoscopy surgery,the negative control group with no-load shRNA adenovirus and the simple models group injected with normal saline.Four weeks later after the injection,an observed operation was performed through laparoscopy and some lesions were collected.The CD34 immunohistochemistry of these lesions were done to detect the microvessel density,then the variation of the microvessel density among each group were observed.The expression of the NF-κB and proliferating cell nuclear antigen (PCNA) were detected through western blot.Results First,the Macaca fascicularis model of endometriosis was successful developed,and the experimental group has an evident atrophy in ectopic lesions compared with the previous.The lesions' microvessel density in experimental group decreased evidently compared with the negative control group and simple model group (0.002 0±0.000 3 versus 0.021 9±0.002 6 versus 0.024 5±0.003 3),and the differences was statistically significant (P＜0.01).The expression of PCNA (0.37±0.17 versus 0.57±0.26 versus 0.57±0.28) and NF-κB (0.338 ± 0.174 versus 0.678 ± 0.021 versus 0.645 ±0.098) in experiment group was lower than the negative control group and simple model group,the differences were statistically significant (all P＜0.01).Conclusion Through targeting suppressed the NF-κB gene expression by NF-κB shRNA,we can inhibit the development of endometriosis through reducing the ability of angiogenesis and cell proliferation of ectopic endometrial cells.