中期保存液和甘油保存的血清HBsAg阳性供体角膜中HBsAg的表达和定位  

Expression and localization of HBsAg in corneas from HBsAg-seropositive donors preserved in medium-term preservation solution and glycerol

作　　者：仁虹 蔡子怡 杜佳琪 李高阳 胡舒钰 欧阳莎 刘寒涵 邓志宏[1] REN Hong;CAI Ziyi;DU Jiaqi;LI Gaoyang;HU Shuyu;OUYANG Sha;LIU Hanhan;DENG Zhihong(Department of Ophthalmology,Third Xiangya Hospital,Central South University,Changsha 410013,China)

机构地区：[1]中南大学湘雅三医院眼科,长沙410013

出　　处：《中南大学学报(医学版)》2024年第10期1642-1648,共7页Journal of Central South University :Medical Science

基　　金：国家自然科学基金(82271057)。

摘　　要：目的:中国角膜移植供体缺口巨大,可通过角膜成分移植来扩大供体角膜来源。本研究旨在通过观察采用不同储存介质保存的血清乙型肝炎表面抗原(hepatitis B surface antigen,HBsAg)阳性捐献者角膜中HBsAg的表达和分布,以评估板层角膜移植的可行性和安全性。方法:本研究的研究对象为2019年12月至2021年3月间临床捐献的保存在中南大学湘雅三医院眼库的10枚角膜(来自6例捐献者)。采用酶联免疫吸附分析的方法检测血清HBsAg的表达,聚合酶链反应(polymerase chain reaction,PCR)法检测血清HBsAg阳性捐献者血中乙型肝炎病毒(hepatitis B virus,HBV)DNA滴度,免疫荧光染色检测角膜中的HBsAg的表达及其在角膜各层的定位,对血中HBV DNA滴度和角膜中HBsAg荧光强度进行相关性分析。结果:4枚来自2例血清HBsAg阳性捐献者的角膜、1枚来自1例血清HBsAg阴性捐献者的角膜均短期(<2周)保存在中期保存液中,2枚来自1例血清HBsAg阳性捐献者的角膜和2枚来自1例血清HBsAg阳性捐献者的角膜分别在甘油中保存1个月和1年,1枚来自1例血清HBsAg阴性捐献者的角膜在甘油中保存1年。免疫荧光染色结果显示:采用中期保存液短期(<2周)保存的2枚角膜上皮层和1枚角膜内皮层、甘油保存1年的1枚角膜上皮细胞均可见HBsAg阳性信号。免疫荧光染色半定量分析结果显示:角膜上皮层和内皮层中HBsAg荧光强度差异有统计学意义(P=0.012),不同捐献者及保存方法的角膜上皮中HBsAg荧光强度的差异无统计学意义(P>0.05)。血中HBV DNA滴度与角膜上皮中的HBsAg阳性表达呈正相关(R^(2)=0.707,P=0.036)。结论:HBsAg血清阳性捐献者的供体角膜存放于中期保存液或在甘油中长期保存后,上皮层或内皮层仍有HBsAg的阳性表达,但角膜基质层中均无HBsAg的阳性表达。血中HBV DNA滴度高表达的捐献者,其角膜中HBsAg表达阳性的可能性越高。本研究为进一步研究HBsObjective:Due to the severe shortage of donor corneas for transplantation in China,corneal component transplantation may expand the available donor pool.This study aims to evaluate the safety and feasibility of corneal component transplantation by examining the distribution of hepatitis B surface antigen(HBsAg)in corneas from HBsAg-seropositive donors under different storage media.Methods:Ten corneas(from 6 donors)donated between December 2019 and March 2021 and stored at the Eye Bank of Xiangya Third Hospital,Central South University,were analyzed.Serum HBsAg levels were quantified using enzyme-linked immunosorbent assay(ELISA),while HBV DNA titers in donor blood were measured via polymerase chain reaction(PCR).Immunofluorescence staining was employed to detect HBsAg expression and its localization within corneal layers.Correlations between blood HBV DNA titers and HBsAg fluorescence intensity in corneal tissues were statistically evaluated.Results:Four corneas from 2 HBsAg-seropositive donors and 1 cornea from 1 HBsAg-seronegative donor were preserved in medium-term preservation solution for<2 weeks.Two corneas from 1 HBsAg-seropositive donor and 2 corneas from another HBsAg-seropositive donor were preserved in glycerol for 1 month and 1 year,respectively.One HBsAg-seronegative donor cornea was stored in glycerol for 1 year.Immunofluorescence staining revealed HBsAg-positive signals in the epithelium of 2 corneas and the endothelium of 1 cornea preserved in medium-term preservation solution(<2 weeks),as well as in the epithelium of 1 cornea stored in glycerol long-term(1 year).Semi-quantitative analysis demonstrated statistically significant differences in HBsAg fluorescence intensity between the corneal epithelium and endothelium(P=0.012),while no significant variations were observed across preservation methods or donors(P>0.05).A positive correlation was identified between blood HBV DNA titers and HBsAg expression in the corneal epithelium(R^(2)=0.707,P=0.036).Conclusion:HBsAg-positive expression persists in

关 键 词：角膜 板层角膜移植 乙型肝炎病毒 乙型肝炎病毒表面抗原 乙型肝炎病毒表面抗体 上皮层 内皮层 基质层 

分 类 号：R51[医药卫生—内科学]