豬隻淋巴球對第二型豬環狀病毒感受性之探討

The Susceptibility of Swine Lymphocytes to Porcine Circovirus Type Ⅱ

林俊明(Chun-Ming Lin)

指導教授 ： 龐飛

共同指導教授 ： 鄭謙仁(Chian-Ren Jeng)

國立臺灣大學/生物資源暨農學院/獸醫學研究所/碩士(2006年)

https://doi.org/10.6342/NTU.2006.00406

摘要

豬隻離乳後多系統消耗性症候群（PMWS）是豬隻新興的疾病，其主要病原為豬第二型環狀病毒（PCV2），並且以免疫系統內的淋巴球流失以及組織球浸潤為其主要病理特徵。截至目前為止，PCV2是否會於免疫系統內複製以及PMWS的致病機制仍不甚清楚。在本實驗中，我們建立了以致裂原刺激豬隻周邊血液淋巴球（PBL）的PCV2感染模式。首先，在第一階段實驗中針對健康無PCV2帶原的豬隻進行測試，當同時對其PBL以致裂原刺激與PCV2攻毒處理後，其PBL中僅0.1％的細胞呈現PCV2抗原陽性；藉由MTT與blastogenesis分析，均顯示PCV2可以降低PBL對於致裂原刺激的反應，然而PCV2對於PBL的存活率或細胞凋亡率則無顯著影響。在第二階段的實驗中，針對臨床上健康的PCV2帶原豬隻進行測試，結果顯示經由免疫螢光染色或原位雜交的方式，PCV2的抗原或核酸可以顯著地在致裂原刺激後的PBL中觀測得到;在致裂原刺激後第4天，PCV2抗原陽性率於PBL中最高可達20％，其中以表面抗原呈現IgM+的細胞最具感受性，病毒力價亦有顯著上升的情況;此外，PCV2 ORF1蛋白的表現通常位於細胞核的周邊或同時出現於核内，而ORF2的蛋白則主要侷限於細胞質內；相較於ORF2蛋白的表現，ORF1蛋白於細胞內的表現與TUNEL陽性訊號呈現密切的相關性。本實驗的結果顯示PBL對於PCV2是具有感受性的，並且PCV2傾向於在分裂中的淋巴球內複製。根據目前所得的實驗結果，我們更進一步推測PCV2導致淋巴球凋亡與PCV2在其內複製可能具某種相關性，這些證據可以對PMWS的致病機制提供一部份的解釋。

關鍵字

淋巴球 ； 豬隻第二型環狀病毒

並列摘要

The porcine circovirus type Ⅱ (PCV2)-associated newly emerged disease, postweaning multisystemic wasting syndrome (PMWS), is characterized by lymphoid depletion in lymphoid organs and replacement by macrophages. Until now, whether PCV2 replicates in immune system and the pathogenesis of PMWS are still not well understood. In this study, we established a model of PCV2 infection in mitogen-treated peripheral blood lymphocytes (PBL). In the first part of the study, it was demonstrated that there was only 0.1% of PCV2 antigen-containing rate in PBL isolated from PCV2-free pigs and treated simultaneously with mitogen and PCV2. The results of both MTT assay and blastogenesis indicated that PCV2 could reduce the responsiveness of PBL to mitogen stimulation, but the results of survival and apoptotic rates indicated that PCV2 had no effects on the viability of PBL. In the second part of study, it was found PCV2 nucleic acid and antigens could be more easily detected in mitogen-stimulated PBL from clinically healthy PCV2-carrier pigs by in situ hybridization and immunofluorescence analysis, respectively. After 4 days of mitogen treatment, the PCV2 antigen-containing rate in PBL could reach 20% with IgM-positive cells predominant. The mitogen-stimulated PBL had higher PCV2 titer. Additionally, PCV2 ORF1-associated protein was usually present perinuclearly or within the nuclei; ORF2-associated protein was chiefly restricted in the cytoplasm; the expression of ORF1-associated protein, but not ORF2, as well matched with TUNEL signal. Results of this study indicate that PBL are indeed susceptible to PCV2 infection and PCV2 is capable to replicate in dividing lymphocytes. This replication may play a role on PCV2-induced apoptosis in PBL. The result may explain in part the pathogenesis of PMWS.

並列關鍵字

lymphocyte ； porcine circovirus type 2

參考文獻

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