Choi C, Hwang K-K, Chae C
Department of Veterinary Pathology, College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul National University, Seoul, Republic of Korea.
Arch Virol. 2004 May;149(5):875-89. doi: 10.1007/s00705-003-0275-6. Epub 2004 Jan 29.
The expression of tumor necrosis factor (TNF)-alpha and apoptosis was studied in lymph nodes from pigs infected with Classical swine fever virus (CSFV). Pigs were inoculated with CSFV and euthanized at 3, 5, 7, and 10 days postinoculation. An increase in TNF-alpha expression was detected in CSFV-infected lymph nodes using a reverse transcription-polymerase chain reaction, and TNF-alpha protein was detected in lymph nodes by immunohistochemistry. The majority of TNF-alpha-positive cells also expressed the SWC3a antigen, a specific marker for porcine leukocytes. By combined use of in situ hybridization and immunohistochemistry, CSFV infection was detected in lymph nodes macrophage. Lymphocytes death occurred by apoptosis that was characterized by condensed shrunken cells and the formation of apoptotic bodies, some of them contained pyknotic nuclear remnants. Apoptosis was detected in situ by TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling) reaction. A double-labeling experiment using immunohistochemistry and TUNEL reaction for the detection of CSFV and apoptosis demonstrated that the majority of labeled cells were positive for CSFV or apoptosis. This suggests that CSFV can induce apoptosis directly and indirectly. Apoptotic cells induced by viral infection were more abundant than CSFV-infected cells in all lymph nodes tested. A double-labeling experiment using immunohistochemistry and TUNEL reaction for the detection of TNF-alpha and apoptosis demonstrated that labeled cells were positive for either TNF-alpha or apoptosis, and both. The present study addressed two important issues regarding CSFV-induced apoptosis: (i) viral infection and apoptosis colocalize at the cell level; and (ii) one or more factors (e.g., TNF-alpha) released from macrophages may induce apoptosis in uninfected bystander cells.
对感染经典猪瘟病毒（CSFV）的猪的淋巴结中肿瘤坏死因子（TNF）-α的表达及细胞凋亡情况进行了研究。给猪接种CSFV，并在接种后第3、5、7和10天实施安乐死。采用逆转录-聚合酶链反应检测发现，CSFV感染的淋巴结中TNF-α表达增加，通过免疫组织化学在淋巴结中检测到了TNF-α蛋白。大多数TNF-α阳性细胞也表达SWC3a抗原，这是猪白细胞的一种特异性标志物。通过原位杂交和免疫组织化学联合使用，在淋巴结巨噬细胞中检测到了CSFV感染。淋巴细胞通过凋亡发生死亡，其特征为细胞浓缩皱缩以及凋亡小体形成，其中一些含有固缩的核残余物。通过TUNEL（末端脱氧核苷酸转移酶介导的dUTP-生物素缺口末端标记）反应原位检测到了细胞凋亡。使用免疫组织化学和TUNEL反应进行双标记实验以检测CSFV和细胞凋亡，结果表明大多数标记细胞CSFV或凋亡呈阳性。这表明CSFV可直接和间接诱导细胞凋亡。在所有检测的淋巴结中，病毒感染诱导的凋亡细胞比CSFV感染的细胞更为丰富。使用免疫组织化学和TUNEL反应进行双标记实验以检测TNF-α和细胞凋亡，结果表明标记细胞TNF-α或凋亡呈阳性，或两者均呈阳性。本研究探讨了关于CSFV诱导细胞凋亡的两个重要问题：（i）病毒感染和细胞凋亡在细胞水平上共定位；（ii）巨噬细胞释放的一种或多种因子（如TNF-α）可能诱导未感染的旁观者细胞发生凋亡。
