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  本文選題：COX-2　切入點：AA　出處：《南華大學》2014年碩士論文　論文類型：學位論文

【摘要】：目的 本實驗擬從COX-2代謝底物花生四烯酸這一視角，，闡明COX-2調(diào)控HSC細胞增殖及ACSL家族基因表達的機制。 實驗一 方法 1、利用MTT比色法檢測大鼠HSC-T6細胞在花生四烯酸20μM、40μM、60μM24h、48h的增殖情況。 2、利用RT-PCR檢測AA20μM、AA40μM、AA60μM組時HSC的COX-2、ACSL家族基因的表達情況。 結(jié)果 1、花生四烯酸20μM、40μM、60μM時HSC細胞生存率增加（p0.05），在40μM濃度下作用48h時增加顯著（p0.05）。 2、COX-2基因在AA20μM、AA40μM表達增高(p0.05)，ACSL1基因在AA60μM表達相對增高(p0.05)，ACSL3、ACSL6基因在AA40μM、AA60μM表達增高(p0.05)，ACSL4、ACSL5基因在各組表達無顯著變化(p0.05）。 實驗二 方法 1、向HSC-T6轉(zhuǎn)染構(gòu)建的pYr-1.1-hU6-EGFP-COX-2shRNA1，熒光顯微鏡下評估其轉(zhuǎn)染效率。分組為①HSC組；②AA40μM組；③pYr-1.1HK+AA組；④COX-2shRNA1+AA組。AA組濃度為MTT篩選的最佳反應濃度40μM。 2、利用MTT比色法檢測轉(zhuǎn)染后各組HSC細胞增殖情況。 3、利用WB技術(shù)檢測轉(zhuǎn)染48h后各組COX-2、ACSL家族基因的表達情況。 結(jié)果 1、AA40μM組較HSC組OD值增加（p0.05），COX-2shRNA1+AA組與pYr-1.1HK+AA組比較OD值降低（p0.05），COX-2shRNA1+AA組較HSC組OD值無顯著差異。 2、 AA組COX-2、 ACSL1、 ACSL6基因表達較HSC組增加(p0.05），AA+COX-2shRNA轉(zhuǎn)染組較pYr-1.1HK+AA空質(zhì)粒組COX-2、ACSL1、ACSL6蛋白表達降低(p0.05），而對照組COX-2、ACSL1、ACSL6表達無顯著差異(p0.05）。 結(jié)論 1、COX-2可以通過代謝底物花生四烯酸促進肝星狀細胞的增殖作用。 2、COX-2可以通過代謝底物花生四烯酸調(diào)控HSC細胞ACSL1、ACSL6基因表達。
[Abstract]:objective
This study aims to elucidate the mechanism of COX-2 regulation of HSC cell proliferation and ACSL gene expression from the perspective of COX-2 metabolite groundnut four enoic acid.
Experiment 1
Method
1, MTT colorimetric assay was used to detect the proliferation of HSC-T6 cells in peanut four enoic acid 20 mu M, 40 mu M, 60 mu M24h, and 48h.
2, the expression of the COX-2 and ACSL family of HSC in the group of AA20 M, AA40, M and AA60 mu M was detected by RT-PCR.
Result
1, the survival rate of HSC cells increased (P0.05) when the peanut four enoic acid 20 mu M, 40 mu M, 60 micron M, and increased significantly at the concentration of 40 mu M (P0.05).
2, the expression of COX-2 gene increased at AA20, M, AA40 and M (P0.05), ACSL1 gene expression was increased at AA60 (M), P0.05 (ACSL3), and the expression of M gene increased in the two groups.
Experiment two
Method
1, transfection of pYr-1.1-hU6-EGFP-COX-2shRNA1 constructed into HSC-T6, and the transfection efficiency under fluorescent microscope. It was grouped as follows: HSC group; AA40 AA40 M group; pYr-1.1HK+AA group; COX-2shRNA1+AA group.AA group concentration was MTT screening for the best reaction concentration of M..
2, the proliferation of HSC cells after transfection was detected by MTT colorimetric assay.
3, WB technique was used to detect the expression of COX-2 and ACSL family genes in each group after transfection of 48h.
Result
1, the AA40 M group than in HSC group was increased (P0.05), COX-2shRNA1+AA group and pYr-1.1HK+AA group was lower than that in group HSC (P0.05), OD group COX-2shRNA1+AA had no significant difference.
2, the expression of COX-2, ACSL1 and ACSL6 genes in group AA increased compared with that in HSC group (P0.05), and the expression of COX-2, ACSL1 and ACSL6 protein in AA+COX-2shRNA transfection group was lower than that in pYr-1.1HK+AA empty plasmid group.
conclusion
1, COX-2 can promote the proliferation of hepatic stellate cells by metabolic substrates, arachidic acid (arachidic acid).
2, COX-2 can regulate the expression of ACSL1 and ACSL6 gene in HSC cells by metabolizing the substrate, arachidic acid.

【學位授予單位】：南華大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R575.5

【參考文獻】

相關期刊論文 前1條

1 李惠俠,楊公社,盧建雄;花生四烯酸對大鼠前體脂肪細胞生長與分化的影響(英文)[J];中國生物化學與分子生物學報;2005年06期

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