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Effects of Classical Swine Fever Virus Infection on Phenotypic and Inflammatory Cytokine Expression of Porcine Alveolar Macrophages
|關鍵字:||classical swine fever;alveolar macrophages;cytokine||出版社:||獸醫病理學研究所||摘要:||
肺泡巨噬細胞(alveolar macrophages; AMs)為下呼吸道重要之防禦細胞，並參與免疫反應進行。當AMs受到感染時，除了表面分子表現量改變外，亦會誘發多種炎症性細胞激素之分泌，並媒介肺炎病變形成。AMs為豬瘟病毒主要標的細胞之一，因此在豬瘟病毒導致之肺炎中，可能佔有決定性角色。為瞭解豬瘟病毒感染AMs後對細胞表面分子及炎症性細胞激素表現之影響，本實驗以in vitro的方式進行AMs之培養，於豬瘟病毒(CSFV-YL strain)攻毒後各時間點收取細胞，以流式細胞儀分析表面分子(MHC class I, class II及CD80/86)之表現程度，並分別利用RT-PCR及ELISA技術評估炎症性細胞激素IL-1alpha, -6, -12 p40, TNF-alpha基因及IL-1 beta, TNF-alpha蛋白質層面表現之情形，另以吞噬作用作為功能分析。實驗結果顯示，豬瘟病毒感染AMs後，可見MHC及CD80/86表面分子表現量之減少，即使在IFN-gamma刺激下，病毒感染細胞其MHC class I及class II分子被誘發表現的程度亦不如對照組細胞明顯。在功能分析方面，豬瘟病毒感染s對其吞噬功能之影響並無顯著差異(p>0.05)。以炎症性細胞激素表現角度來看，遭受豬瘟病毒感染的AMs雖然會誘發炎症性細胞激素的產生，但不論是在IL-1alpha, -6, -12 p40, TNF-alpha基因及IL-1beta, TNF-alpha蛋白質之表現上，皆無顯著之差異(p>0.05)；若於豬瘟病毒感染48小時後以LPS刺激，則會誘發上述細胞激素更大量之表現，尤其是IL-6及IL-12，在刺激後12-24小時其基因有顯著表現(p<0.05)。而蛋白質表現層面，豬瘟病毒感染的AMs同樣在LPS刺激後，會誘發IL-1更大量之產生；然而卻導致TNF-alpha的降低。綜合以上實驗結果，顯示肺泡巨噬細胞在豬瘟病毒感染後，會降低MHC分子之呈現，並誘發炎症性細胞激素表現，亦可能促進其他病原對肺臟組織之傷害，尤其在細菌混合感染之下，更會藉由促進IL-6及IL-12等炎症性媒介物質的產生，而加重肺臟之炎症反應。
Alveolar macrophages (AMs) are important defensive cells of lower respiratory tract, which play a prominent role in pulmonary immune response. When AMs are infected, there are not only alteration of cell surface molecules, but also induction of inflammatory cytokine, which contribute to the pathogenesis of pneumonia. AMs are one of main target cells for classical swine fever virus (CSFV), therefore they might play a determinative role in CSFV induced pneumonia. To investigate the effects of CSFV infection, AMs were cultured in vitro, challenged with CSFV (YL strain), and their phagocytic activity, phenotypic expression, and inflammatory cytokine expression were evaluated. By flow cytometry analysis, we found that the expression of MHC class I, class II and CD80/86 decreased after CSFV infection. Even in IFN-gamma stimulated AMs, the MHC class I and class II expression in virus-infected cells were not as high as control cells. In addition, there was no significant difference (p>0.05) in phagocytic activity in AMs infected with CSFV. To study the inflammatory cytokine profile, expression of IL-1 alpha, -6, -12 p40 and TNF-alpha were assessed by RT-PCR, and protein level of IL-1 beta and TNF-alpha were determined by ELISA assay. We found that expression of IL-1 alpha, -6, -12 p40, IL-1 beta, TNF-alpha in CSFV-infected AMs were elevated at mRNA and/or protein level, but these elevations were statistically insignificant (p>0.05). Furthermore, expression of IL-6 and IL-12 were greatly increased (p<0.05) in AMs infected with CSFV for 48 hours, followed by stimulation with LPS for 12-24 hours, with the exception that expression of TNF-alpha was reduced. In summary, CSFV infected AMs showed decreased level of MHC molecule expression and induction of inflammatory cytokine. These responses might promote the injury of pulmonary tissue by other pathogens, especially while in complication with bacterial infection which might enhance pulmonary inflammatory by inducing production of inflammatory mediators such as IL-6 and IL-12.
|Appears in Collections:||獸醫病理生物學所|
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