结直肠癌患者血清外泌体整合素的表达及其临床意义  

Expression of serum exosomes integrins and their clinical significance in patients with colorectal cancer

作　　者：焦蕉[1,2] 唐继云 刘佳琴 唐麒 李东芳 胡广生[2] 蒋益兰 Jiao Jiao;Tang Ji-Yun;Liu Jia-Qin;Tang Qi;Li Dong-Fang;Hu Guang-Sheng;Jiang Yi-Lan(Department of Integrated Chinese and Western Medicine,Hunan Cancer Hospital,Changsha 410013,China;Graduate School,Hunan University of Traditional Chinese Medicine,Changsha 410208,China;Department of Oncology,Hengyang Hospital of Traditional Chinese Medicine,Hengyang,Hunan 421000,China;Hunan Key Laboratory of Chinese Medicine Oncology,Hunan Academy of Traditional Chinese Medicine Affiliated Hospital,Changsha 410006,China)

机构地区：[1]湖南省肿瘤医院中西医结合科,长沙410013 [2]湖南中医药大学研究生院,长沙410208 [3]衡阳市中医医院肿瘤科,湖南衡阳421000 [4]湖南省中医药研究院附属医院中医肿瘤学湖南省重点实验室,长沙410006

出　　处：《解放军医学杂志》2022年第6期586-592,共7页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金(81774287);湖南省中医药科研计划项目(2021193);长沙市科技计划项目(kq1706040);湖南创新型省份建设专项经费资助项目(2019SK4013)。

摘　　要：目的探讨不同分期结直肠癌患者血清外泌体中整合素α_(5)、β_(5)、β_(6)的表达水平及其临床意义。方法收集2016年1月-2020年10月就诊于湖南省中医药研究院附属医院和湖南省肿瘤医院的50例结直肠癌患者的血清标本,按照TNM分期分为结直肠癌Ⅰ期组(n=5)、Ⅱ期组(n=19)、Ⅲ期组(n=14)与Ⅳ期组(仅肝转移,n=12)。另收集2018年8月-2020年5月于湖南省中医药研究院附属医院健康管理中心体检的10名健康者的血清标本作为健康对照组。对血清标本进行外泌体提纯,运用纳米微粒追踪分析(NTA)及Western blotting鉴定外泌体;采用Western blotting检测血清外泌体中整合素α_(5)、β_(5)、β_(6)的表达水平。随访追踪50例结直肠癌患者的生存期,采用Kaplan-Meier法绘制生存曲线,采用Cox比例风险回归模型进行生存分析。结果50例结直肠癌患者中,男26例(52.0%),女24例(48.0%),以中老年群体为主,平均年龄58岁;TNM分期Ⅰ期5例(10.0%),Ⅱ期19例(38.0%),Ⅲ期14例(28.0%),Ⅳ期(仅肝转移)12例(24.0%)。随访截至2021年2月,随访时间5~60个月,中位随访时间44个月,无病例脱落或失访。NTA分析结果显示,各组血清外泌体的粒径分布集中在50~150 nm。Western blotting检测结果显示,各组血清外泌体中ALIX、HSP70等标志性蛋白均有表达,证实提取物为外泌体;各组血清外泌体中整合素α_(5)、β_(5)、β_(6)均有不同程度表达,其中整合素α_(5)、β_(5)的表达水平随着TNM分期增加而升高,肝转移Ⅳ期患者α_(5)、β_(5)表达水平明显高于非Ⅳ期患者(P<0.05),而整合素β_(6)表达与TNM分期无明显关系(P>0.05)。Kaplan-Meier生存曲线分析显示,整合素α_(5)、β_(5)及TNM分期与结直肠癌患者的生存期有关(P<0.05);Cox比例风险回归模型分析结果显示,整合素α_(5)及病理分化程度是结直肠癌患者生存期的独立影响因素(P<0.05)。结论外泌体整合素α_(5)、β_(5)与结肠癌肝转移及TNObjective To investigate the expression levels of serum exosomes integrinα_(5),β_(5)andβ_(6)and their clinical significance in patients with different stages of colorectal cancer(CRC).Methods The serum samples were collected from 50 patients with CRC admitted in the Affiliated Hospital of Hunan Academy of Traditional Chinese Medicine and Hunan Cancer Hospital from January 2016 to October 2020,and were divided into CRC stageⅠgroup(n=5),CRC stageⅡgroup(n=19),CRC stageⅢgroup(n=14),and CRC stageⅣgroup(only liver metastasis,n=12)according the TNM stage.The serum samples were collected from 10 healthy people who underwent physical examination in the Health Management Center of the Affiliated Hospital of Hunan Academy of Traditional Chinese Medicine from August 2018 to May 2020,and set as healthy people group.The exosomes were purified,and then identified by nanoparticle tracking assay(NTA)and Western blotting.The expression levels of integrinα_(5),β_(5)andβ_(6)were detected by Western blotting,the survival time of 50 patients with CRC were followed up.Kaplan-Meier method was used to draw the survival curve,and Cox proportional hazards model was used for survival analysis.Results The 50 patients with CRC[26 males(52.0%)and 24 females(48.0%)]are mainly middle-and elderly-aged with average age of 58 years.Five cases(10.0%)in TNM stageⅠ,19 cases(38.0%)in stageⅡ,14 cases(28.0%)in stageⅢand 12 cases(24.0%)in stageⅣ(only liver metastasis).Up to February 2021,the follow-up time was 5-60 months,and the median follow-up time was 44 months,no cases fell off or lost follow-up.NTA analysis showed that the particle diameter of serum exosomes in all serum samples were concentrated in 50-150 nm.Western blotting results showed that the marker proteins of serum exosomes ALIX and HSP70 were expressed in all serum samples,which confirmed that these extracts were exosomes.Integrinα_(5),β_(5)andβ_(6)were expressed in different degrees,the expression levels of integrinα_(5)andβ_(5)increased with the increase of TNM

关 键 词：结直肠癌 整合素 外泌体 生存期 

分 类 号：R735[医药卫生—肿瘤]