原核表达纯化的抗菌肽LL-37重组蛋白对小鼠外阴阴道假丝酵母菌病的治疗作用  

Effects of antimicrobial peptide LL-37 expressed and purified from prokaryotes in the murine model of vaginal candidiasis

作　　者：王芳[1] 霍彦[2] 尹利荣[1] 孙蓓[3] 张萍萍[1] 

机构地区：[1]天津医科大学第二医院妇科,300211 [2]天津医科大学第二医院计划生育科,300211 [3]天津医科大学代谢病医院内分泌研究所卫计委激素与发育重点实验室

出　　处：《中华妇产科杂志》2016年第7期530-534,共5页Chinese Journal of Obstetrics and Gynecology

基　　金：天津市卫生局科技基金（2013KZ103）

摘　　要：目的：探讨原核表达纯化的抗菌肽LL-37重组蛋白对小鼠外阴阴道假丝酵母菌病（VVC）的治疗作用。方法（1）雌性昆明小白鼠30只，经雌激素诱导并阴道内接种白假丝酵母菌悬液，构建VVC小鼠模型。建模成功后，按随机数字表法将小鼠随机分成实验组和对照组各15只，实验组小鼠阴道内灌注LL-37重组蛋白溶液30μl（100μg/ml，即3μg），每天1次，连续5 d；对照组小鼠阴道内灌注等量磷酸盐缓冲液（PBS）。（2）阴道用药结束后24 h，取两组小鼠的阴道灌洗液评估小鼠阴道内假丝酵母菌的载菌量[以菌落形成单位（CFU）表示]及假菌丝和芽生孢子的生成情况；摘取两组小鼠的阴道组织制备匀浆，ELISA法检测匀浆中免疫因子——白细胞介素10（IL-10）、γ干扰素（IFN-γ）的浓度。结果（1）本研究成功构建VVC小鼠模型，对照组小鼠阴道黏膜明显充血、水肿、阴道皱襞消失，实验组经LL-37重组蛋白治疗后阴道黏膜无明显水肿，阴道皱襞可见。（2）实验组小鼠阴道分泌物涂片中白假丝酵母菌的芽生孢子及假菌丝明显少于对照组。实验组小鼠阴道灌洗液中载菌量明显低于对照组[分别为（4.8±1.0）×10^4、（8.5±2.1）×10^4 CFU/ml，P=0.017]；阴道组织匀浆中IFN-γ浓度明显高于对照组[分别为（257±11）、（197±4）pg/ml，P=0.000]，而IL-10浓度明显低于对照组[分别为（287±15）、（379 ±17）pg/ml，P=0.000]，且IFN-γ/IL-10比值明显高于对照组（分别为0.892±0.008、0.496±0.013，P=0.000）。结论原核表达纯化的LL-37重组蛋白可以抑制VVC小鼠阴道内假丝酵母菌的生长，调节阴道内IFN-γ、IL-10的分泌水平，改善阴道局部免疫力，对小鼠VVC有明显的治疗作用。Objective To study the effects of antimicrobial peptide LL-37 expressed and purified from prokaryotes on candida albicans growth. Methods （1） Thirty female Kunming mice were treated with estrogen and white candida yeast suspension were poured into vagina to establish a vulvovaginal candidiasis （VVC） murine model. After successful establishing the VVC mouse model, mice were randomly sorted into test group （n=15） and control group （n=15） . Suspension（30μl, 100μg/ml）of recombinant peptide LL-37 expressed and purified in Prokaryotes was given by intravaginal administration to the test group for 5 days, while the same amount of phosphate buffered saline （PBS） was given to the control group. （2） Tweenty-four hours after treatment, the fungal burden and colony-forming unit （CFU） of vaginal fluids were evaluated. All mice were subsequently sacrificed and vaginal tissues were harvested for tissue homogenate preparation. ELISA was used to determine the levels of nterleukin-10（IL-10）and interferon-γ（IFN-γ）in the isolated vaginal tissues. Results （1） VVC mouse model was established successfully in this study. Vaginal mucosa congestion, edema, vaginal plica disappearing were obviously observed in the control group. After treatment＆amp;nbsp;with recombinant protein LL-37 vaginal mucosa has no obvious change in the test group. （2） Fungal burden and CFU of vaginal fluids were significantly lower in the test group [（4.8±1.0）×10^4 CFU/ml] than that in the control group [（8.5 ± 2.1） × 10^4 CFU/ml, P=0.017]. IFN-γlevel of the test group was increased [（257 ± 11） vs （197 ± 4） pg/ml, P=0.000], while the level of IL-10 was reduced [ （287 ± 15） vs （379 ± 17） pg/ml P=0.000] resulting in a the ratio of IFN-γ/IL-10 was in significantly higher in test group （0.892±0.008 vs 0.496±0.013, P=0.000）. Conclusion Recombinant protein LL-37 expressed and purified from prokaryotes inhibits the growth candida albicans and improves vaginal immunity by adjustin

关 键 词：念珠菌病 外阴阴道 内源性抗菌多肽类物质 干扰素Γ 白细胞介素10 INTERLEUKIN-10 

分 类 号：R711.3[医药卫生—妇产科学]