禽网状内皮组织增生病病毒感染对SPF雏鸡法氏囊细胞Bcl-2和C-myc基因表达及凋亡的影响

doi:10.16431/j.cnki.1671-7236.2022.04.031

摘要/Abstract

摘要： 【目的】探究禽网状内皮组织增生病病毒(Reticuloendotheliosis virus,REV)感染禽类后导致免疫器官发生细胞凋亡的机理。【方法】以1日龄SPF雏鸡为试验对象,将100只SPF雏鸡随机均分为REV感染组和未感染病毒的对照组,REV感染组雏鸡经腹腔感染500 μL REV稀释液,对照组雏鸡经相同途径注射等量灭菌生理盐水,于病毒感染后第1、7、14、21、28和42天,2组雏鸡随机各抽取5只,心脏采血处死雏鸡后快速摘取法氏囊。分别应用HE染色和病理切片成像系统测定分析法氏囊细胞核浆比,TUNEL细胞凋亡原位检测试剂盒测定凋亡细胞数,免疫组化法测定Bcl-2和C-myc阳性细胞数量,实时荧光定量PCR和ELISA法分别检测法氏囊Bcl-2和C-myc 基因mRNA表达和蛋白含量。【结果】 ①REV感染1日龄SPF雏鸡后21~42 d,其法氏囊淋巴细胞凋亡百分比显著或极显著高于对照组雏鸡(P<0.05;P<0.01);②SPF雏鸡感染REV后21和28 d,其法氏囊细胞核浆比显著低于对照组雏鸡(P<0.05);③REV感染SPF雏鸡法氏囊中Bcl-2和C-myc阳性细胞数在病毒感染后21和28 d显著高于对照组雏鸡(P<0.05);④REV感染SPF雏鸡后21 d,其法氏囊Bcl-2和C-myc 基因mRNA表达极显著高于对照组雏鸡(P<0.01)。⑤SPF雏鸡感染REV后,其法氏囊中Bcl-2蛋白含量较对照组雏鸡有不同程度的增加,其中21和28 d分别差异极显著(P<0.01)和显著(P<0.05),病毒感染组雏鸡的C-myc蛋白含量也始终高于对照组雏鸡,且21和28 d极显著增高(P<0.01)。【结论】 REV感染所致SPF雏鸡法氏囊细胞Bcl-2和C-myc的mRNA表达以及蛋白含量异常均与病毒感染导致的法氏囊细胞凋亡密切相关,而法氏囊细胞凋亡数量增加与REV感染引发的机体免疫机能抑制密切相关。

关键词: 禽网状内皮组织增生病病毒(REV); SPF雏鸡; 法氏囊; 细胞凋亡

Abstract: 【Objective】 This experiment was aimed to investigate the mechanism of apoptosis of immune organs caused by avian Reticuloendotheliosis virus (REV).【Method】 SPF chickens were selected as experimental subjects in this study.100 1-day-old SPF chickens were randomly divided into REV-infected group and uninfected control group.Those REV-infected chickens were taken 500 μL of REV dilution by peritoneal injection,the control group chicks were injected with sterilized saline instead.On the 1st,7th,14th,21th,28th and 42th day after virus infection,5 chickens were randomly selected from these two groups of chickens,and their bursas were quickly removed after execution.HE staining and pathological imaging system were applied to observe the nuclear-cytoplasm ratio of bursa of Fabricius cells,TUNEL assay was used to determine the quantity of apoptotic cells,immunohistochemical method was used to count the number of Bcl-2 and C-myc positive cells,and Real-time PCR and ELISA methods were used to test the mRNA expression and protein content of Bcl-2 and C-myc genes in the bursa of Fabricius.【Result】 The results showed that:①21-42 days after SPF chickens infected with REV,the percentage of apoptosis of bursal lymphocytes was significantly or extremely significantly higher than that of the control chickens (P<0.05;P<0.01);②21-28 days after SPF chickens infected with REV,the ratio of lymphocyte to cytoplasm in the bursa of Fabricius was significantly lower than that of control chickens (P<0.05);③The number of Bcl-2 and C-myc positive cells in the bursa of Fabricius of SPF chickens infected with REV was significantly higher than that of control chickens at 21 and 28 d (P<0.05);④21 d after REV infection,the expression of Bcl-2 and C-myc genes mRNA in bursa of Fabricius was extremely significantly higher than that of control chickens (P<0.01);⑤After SPF chickens infected with REV,the Bcl-2 protein content in the bursa of Fabricius was increased to different degrees compared with control group chickens,and the difference between 21 and 28 d was extremely significant (P<0.01) and significant (P<0.05),respectively.The content of C-myc protein in the REV-infected group was always higher than that in the control group,and it was increased extremely significantly from 21 to 28 d (P<0.01).【Conclusion】 The above results indicated that the abnormal expression of Bcl-2 and C-myc in the bursa of Fabricius of SPF chickens caused by REV infection was related to the apoptosis of bursal of Fabricius cells caused by the virus,and the increased apoptosis in the number of bursal of Fabricius cell was related to the suppression of immune function caused by REV infection.

Key words: Reticuloendotheliosis virus (REV); SPF chickens; bursa of Fabricius; apoptosis

中图分类号:

S852.65^＋7

顾先哲, 侯宁, 吕晓萍, 高雪丽, 刘超男, 吴文杰, 郑世民. 禽网状内皮组织增生病病毒感染对SPF雏鸡法氏囊细胞Bcl-2和C-myc基因表达及凋亡的影响[J]. 中国畜牧兽医, 2022, 49(4): 1488-1496.

GU Xianzhe, HOU Ning, LYU Xiaoping, GAO Xueli, LIU Chaonan, WU Wenjie, ZHENG Shimin. Influence of Avian Reticuloendotheliosis Virus Infection on Expression of Bcl-2 and C-myc Genes and Apoptosis in Bursa of Fabricius Cells of SPF Chickens[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(4): 1488-1496.

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