本文選題：腎型傳染性支氣管炎病毒　切入點：雞腎細胞　出處：《江西農(nóng)業(yè)大學》2016年碩士論文　論文類型：學位論文

【摘要】：為研究NIBV對雞腎細胞活性,抗氧化功能以及與細胞凋亡相關基因表達量等的影響,探討NIBV的致病機理。本實驗通過已建立的雞腎細胞體外培養(yǎng)模型,用劑量為半數(shù)組織感染量的NIBV病毒感染雞腎細胞,檢測NIBV病毒在腎細胞內(nèi)的動態(tài)復制變化;通過MTT四唑鹽法和乳酸脫氫酶測定試劑盒檢測NIBV病毒對腎細胞活性和細胞膜的損傷情況,用試劑盒測定細胞和細胞培養(yǎng)液中超氧化物歧化酶和黃嘌呤氧化酶活性、丙二醛和尿酸含量;SYBR-PCR檢測原代雞腎細胞內(nèi)XOD,Bak1,Bcl2,P53,Caspase-3,DDIT3的m RNA相對基因表達量變化。結果表明:感染NIBV后,雞腎細胞的形態(tài)和結構發(fā)生明顯變化,病毒組細胞活力極顯著下降(P0.01),細胞內(nèi)的病毒粒子數(shù)量先升高后下降,在36 hpi達到最高;與對照組相比,病毒組細胞LDH含量、XOD活性、UA濃度、上清液SOD活性和胞內(nèi)MDA濃度均極顯著升高(P0.01);胞內(nèi)SOD活性在72 hpi極顯著下降(P0.01);熒光定量PCR結果顯示病毒組細胞XOD,Bak1,Bcl2,P53,Caspase-3 mRNA基因表達量較對照組都極顯著升高(P0.01),病毒組Bcl2上游調(diào)控基因DDIT3的mRNA基因表達量較對照組極顯著下降(P0.01)。說明,腎型傳染性支氣管炎病毒感染抑制原代雞腎細胞的增殖,同時引起細胞黃嘌呤氧化酶活性上升和抗氧化功能下降,NIBV病毒感染影響雞腎細胞凋亡相關基因的表達。
[Abstract]:In order to study the effects of NIBV on the activity, antioxidant function and expression of apoptosis-related genes of chicken kidney cells, and to explore the pathogenesis of NIBV, we established a model of chicken renal cell culture in vitro. Chicken renal cells were infected with NIBV virus with a dose of 50% tissue infection. The dynamic replication of NIBV virus in renal cells was detected. MTT tetrazolium salt assay and lactate dehydrogenase assay were used to detect the damage of NIBV virus to renal cells and cell membrane. The activities of superoxide dismutase (SOD) and xanthine oxidase (xanthine oxidase) in cell and cell culture medium were measured by the kit. Malondialdehyde (MDA) and uric acid (uric acid) were detected by SYBR-PCR in primary chicken renal cells. The expression of m RNA in primary chicken kidney cells was detected by Bcl2P53Caspase-3 caspase-3 DDIT3. The results showed that the morphology and structure of chicken renal cells changed significantly after NIBV infection. The activity of virus group cells decreased significantly (P 0.01), and the number of virus particles increased first, then decreased, and reached the highest level at 36 hpi. Compared with the control group, the concentration of LDH activity and UA in virus group cells were increased. The activity of SOD and the concentration of intracellular MDA in supernatant were significantly higher than those in control group, and the intracellular SOD activity decreased significantly at 72 hpi. The results of fluorescence quantitative PCR showed that the expression of Caspase-3 mRNA gene in XODN Bak1 Bcl2P5Caspase-3 cells was significantly higher than that in control group, and the expression of Caspase-3 mRNA in virus group Bcl2 was significantly higher than that in control group. The expression of mRNA gene in DDIT3 was significantly lower than that in the control group (P 0.01). The infection of renal infectious bronchitis virus inhibited the proliferation of primary chicken renal cells and caused the increase of xanthine oxidase activity and the decrease of antioxidant function. The infection of NIBV virus affected the expression of apoptosis-related genes in chicken kidney cells.
【學位授予單位】：江西農(nóng)業(yè)大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：S858.31

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