PCV-2感染仔猪淋巴结中的病毒定位与细胞凋亡

Virus Location and Apoptosis in Lymph Nodes of Piglets Infected With PCV-2

【目的】了解PCV-2感染仔猪后,病毒在腹股沟淋巴结中的位置、凋亡细胞的类型及其两者之间的关系。【方法】选用9头32日龄PCV-2抗体阴性的普通断奶仔猪,随机分为3组,即对照组、PCV-2攻毒组(PCV-2组)、PCV-2攻毒后再用钥匙孔血蓝蛋白(KLH)刺激组(PCV-2+KLH组),每组3头。感染后32d采集腹股沟浅淋巴结制作石蜡切片。采用免疫组织化学方法对病毒进行定位,用末端标记法(TUNEL)进行细胞凋亡的定位测定,并用流式细胞法对细胞周期和细胞凋亡进行定量检测。【结果】PCV-2组和PCV-2+KLH组仔猪淋巴结的主要病变为皮质和副皮质区淋巴细胞严重缺失,上皮样巨噬细胞浸润,巨噬细胞胞浆内见有病毒包涵体,病毒主要存在淋巴小结的上皮样巨噬细胞中;PCV-2组和PCV-2+KLH组仔猪淋巴结中的淋巴细胞凋亡严重,且主要是淋巴小结内的淋巴细胞发生凋亡;流式细胞仪检测PCV-2组和PCV-2+KLH组仔猪淋巴结的细胞凋亡率分别为3.34%、4.88%,明显高于对照组0.41%的细胞凋亡率(P<0.05和P<0.01);表明细胞增殖活性的增殖指数(PI)分别是:0.17±0.01、0.12±0.01和0.12±0.04,3组间虽无统计学差异,但对照组高于2个实验组。【结论】PCV-2可通过抑制细胞增殖和促进细胞凋亡导致淋巴结中淋巴细胞的严重缺失,这可能是PCV-2感染引起仔猪免疫抑制的重要机理之一。

[Objective] The objective of the present study was to know location of the virus, type of apoptotic cells and their relations after PCV-2 infected piglets. [Method] Nine 32-day-old conventional piglets not infected with PCV-2 were used, distributed in three groups： control group （n=3）, pigs inoculated with PCV-2 alone （PCV-2, n=3）, PCV-2 inoculated and KLH immunostimulated （PCV-2＋KLH, n=3）. From all pigs, superficial inguinal lymph nodes were collected for histological examination after 32 days postinoculation, immunohistochemistry for PCV-2 detection. Location of apoptotic cells was detected with TUNEL and cell cycle and apoptotic rate were measured by flow cytometry. [ Result ] The characteristic histopathological lesions of the lymph nodes of the pigs in PCV-2 and PCV-2＋KLH were lymphocyte depletion in cortex and paracortex, epithelioid-like macrophages infiltration, intracytoplasrulc inclusion bodies presented in epithelioid-like macrophages. PCV-2 was mainly found in epithelioid -like macrophages by immunohistochernistry. In the lymph nodes, lymphocytes presented higher apoptotic rates in the cortex by TUNEL, special B-cell areas, similar apoptotic cells were found in this compartment in control. Apoptotic rates of lymph nodes were 0.41%, 3.34% and 4.88% in control, PCV-2 and PCV-2＋KLH by flow cytometry, respectively. The apoptotic rates of lymph nodes for PCV-2 and PCV-2 ＋ KLH pigs were significantly higher than those for control group （P〈0.05 and P〈0.01）. Proliferation index （PI） was 0.17±-0.01, 0.12±0.01 and 0.12±0.04 in control, PCV-2 and PCV-2 ＋ KLH group, the date of control group was higher than the other groups, but without statistical difference. [ Conclusion ] The PCV-2 can induce lymphocyte depletion in lymph nodes of piglets by blocking cells proliferation and promoting apoptosis. This is one of the mechanism of immunosuppression on infected piglets by PCV-2.