葫芦素B调节Hippo-Yes相关蛋白通路对宫颈癌中调节性T细胞介导免疫逃逸的有关机制研究  

Mechanism research of cucurbitacin B regulating Hippo-Yes-associated protein pathway on Treg-mediated immune escape in cervical cancer

作　　者：陈国斌[1] 祝文峰 史文娟[1] 董倩 CHEN Guobin;ZHU Wenfeng;SHI Wenjuan;DONG Qian(Department of Obstetrics and Gynecology,Shenzhen Maternal and Child Health Hospital,Guangdong Province,Shenzhen518000,China)

机构地区：[1]深圳市妇幼保健院妇产科,广东深圳518000

出　　处：《中国当代医药》2023年第13期15-19,24,F0003,共7页China Modern Medicine

基　　金：广东省医学科学技术研究基金项目(A2022374)。

摘　　要：目的探讨葫芦素B调节Hippo-Yes相关蛋白(YAP)通路对宫颈癌中调节性T细胞(Treg)介导的免疫逃逸的影响。方法选取BALB/c裸鼠32只作为研究对象,皮下注射人宫颈癌Hela细胞建立裸鼠移植瘤模型,将造模成功的裸鼠按照随机数字表法分为CON组、葫芦素B组、LPA组和葫芦素B-LPA组,每组8只。CON组采用腹腔注射生理盐水,葫芦素B组和LPA组采用腹腔注射剂量0.5 mg/kg的葫芦素B和剂量为3 mg/kg的JAP激活剂LPA,葫芦素B-LPA组采用腹腔同时注射0.5 mg/kg的葫芦素B和3 mg/kg的LPA,每2天给药一次,共处理21 d后取材;采用ELISA试剂盒法比较各组裸鼠血清白介素-2(IL-2)、IL-10和转化生长因子-β_(1)(TGF-β_(1))水平;采用流式细胞术比较各组裸鼠淋巴细胞CD4^(+)、CD4^(+)CD25^(+)叉状头状螺旋转录因子3(Foxp3^(+))Treg水平;采用Western blot法比较各组瘤组织Foxp3、TGF-β、YAP、大肿瘤抑制基因1(LATS1)、哺乳动物不育系20样激酶1(MST1)蛋白表达。结果21 d后,葫芦素B组裸鼠的肿瘤质量、血清IL-2、IL-10、TGF-β_(1)表达、CD4^(+)CD25^(+)Foxp3^(+)Treg/CD4^(+)比例、Foxp3、TGF-β、YAP蛋白表达低于CON组,LATS1、MST1蛋白表达高于CON组,差异有统计学意义(P<0.05);LPA组裸鼠的肿瘤质量、血清IL-2、IL-10、TGF-β_(1)表达、CD4^(+)CD25^(+)Foxp3^(+)Treg/CD4^(+)比例、Foxp3、TGF-β、YAP蛋白表达高于CON组,差异有统计学意义(P<0.05);LPA组和CON组的LATS1、MST1蛋白表达比较,差异无统计学意义(P>0.05);葫芦素B-LPA组裸鼠的肿瘤质量、血清IL-2、IL-10、TGF-β_(1)表达、CD4^(+)CD25^(+)Foxp3^(+)Treg/CD4^(+)比例、Foxp3、TGF-β、YAP蛋白表达高于葫芦素B组,低于LPA组,差异有统计学意义(P<0.05);葫芦素B-LPA组和葫芦素B组的LATS1、MST1蛋白表达比较,差异无统计学意义(P>0.05);葫芦素B-LPA组的LATS1、MST1蛋白表达高于LPA组,差异有统计学意义(P<0.05)。结论葫芦素B可通过调节Hippo-YAP通路抑制宫颈癌中Treg介导的免�Objective To investigate the influences of cucurbitacin B on regulatory T cells(Treg)-mediated immune escape in cervical cancer by regulating Hippo-Yes-associated protein(YAP)pathway.Methods 32 BALB/c nude mice were selected as the research objects,and the nude mice xenograft model was established by subcutaneous injection of human cervical cancer Hela cells.The successfully modeled nude mice were grouped into CON group,cucurbitacin B group,LPA group and cucurbitacin B-LPA group according to the random number table method,with 8 mice in each group;The CON group received intraperitoneal injection of normal saline.The cucurbitacin B group and the LPA group were respectively administered the cucurbitacin B with a dose of 0.5 mg/kg or the JAP activator LPA with a dose of 3 mg/kg by intraperitoneal injection.The cucurbitacin B-LPA group was injected intraperitoneally with 0.5 mg/kg cucurbitacin B and 3 mg/kg LPA at the same time.The drug was administered once every 2 days,and the materials were collected after a total of 21 days of treatment.ELISA kits were used to detect serum levels of interleukin-2(IL-2),IL-10 and transforming growth factor-β_(1)(TGF-β_(1))in nude mice;flow cytometry was used to detect the levels of CD4^(+),CD4^(+)CD25^(+)fork-head helix transcription factor 3(Foxp3^(+))Treg.Western blot was used to detect the protein expression of Foxp3,TGF-β,YAP,large tumor suppressor gene 1(LATS1)and mammalian sterile line 20-like kinase 1(MST1)in tumor tissues.Results After 21 days,the tumor mass,serum IL-2,IL-10,TGF-β_(1) expression,CD4^(+)CD25^(+)Foxp3^(+)Treg/CD4^(+)ratio,Foxp3,TGF-β,and YAP protein expression in nude mice in cucurbitacin B group were lower than those in CON group,LATS1,MST1 protein expression were higher than those in the CON group,the differences were statistically significant(P<0.05).The tumor mass,serum IL-2,IL-10,TGF-β_(1) expression,CD4^(+)CD25^(+)Foxp3^(+)Treg/CD4^(+)ratio,Foxp3,TGF-β,and YAP protein expression in nude mice of the LPA group were higher than those of the CON gro

关 键 词：葫芦素B Hippo-Yes相关蛋白通路 宫颈癌 调节性T细胞 免疫逃逸 

分 类 号：R737.33[医药卫生—肿瘤]