本文關(guān)鍵詞： LPS 小鼠β防御素-2 小鼠schlafen 小鼠成纖維細(xì)胞 信號轉(zhuǎn)導(dǎo)　出處：《蘭州大學(xué)》2011年碩士論文　論文類型：學(xué)位論文

【摘要】：目的：通過研究LPS對于小鼠成纖維細(xì)胞(L929)β-防御素(defb)和schlafen (slfn)基因表達(dá)的影響,以及P38MAPK和NF-κB信號轉(zhuǎn)導(dǎo)通路對defb2和slfn2基因表達(dá)的調(diào)控,探討成纖維細(xì)胞防御病原微生物作用的機(jī)制,為進(jìn)一步研究機(jī)體抗感染機(jī)理提供理論基礎(chǔ)和實(shí)驗(yàn)依據(jù)。 方法：以低、中、高劑量組LPS(50ng/ml,200ng/ml,500ng/ml)分別處理L929細(xì)胞2h,4h,6h,以PBS處理L929細(xì)胞相同時間段作為對照組,提取細(xì)胞總RNA,用RT-PCR和熒光定量PCR的方法檢測defb1,2和slfn1,2,3基因表達(dá)；應(yīng)用Western Blotting的方法檢測defb2蛋白的表達(dá)。BMS345541和PD169316兩種信號轉(zhuǎn)導(dǎo)通路的化學(xué)抑制劑預(yù)處理細(xì)胞,再用LPS刺激細(xì)胞后觀察defb2和slfn2基因、蛋白表達(dá)的變化,探討defb2和slfn2基因表達(dá)的信號轉(zhuǎn)導(dǎo)通路。 結(jié)果： 1.LPS誘導(dǎo)小鼠成纖維細(xì)胞β-防御素-2(defb2)的表達(dá),其在中劑量組、2h時基因、蛋白表達(dá)最強(qiáng)；p-防御素-1(defb1)則未被誘導(dǎo)表達(dá)。 2. schlafen2 (slfn2)在中劑量組、2h時其相對mRNA表達(dá)最多；在不同濃度和不同時間下,與對照組相比,slfnl,3的相對mRNA表達(dá)均無明顯差異。 3.IL-6相對mRNA表達(dá)在2h組基因表達(dá)最強(qiáng),4h和6h組相對mRNA表達(dá)逐漸下降；IL-1相對mRNA表達(dá)在2h、4h、6h組與對照組相比均無明顯差異。 4.信號轉(zhuǎn)導(dǎo)效應(yīng)分子TLR4、MYD88、NF-κB的相對]nRNA表達(dá)在2h、4h、6h組與對照組相比均有統(tǒng)計學(xué)意義。 5.與LPS組相比,BMS345541和PD169316組defb2相對mRNA與蛋白的表達(dá)均明顯降低,slfn2相對mRNA表達(dá)也明顯降低。 結(jié)論：LPS誘導(dǎo)小鼠成纖維細(xì)胞β-防御素-2基因的表達(dá),上調(diào)schlafen-2和白細(xì)胞介素-6基因的表達(dá),對p-防御素-1、schlafen-1,-3和白細(xì)胞介素-1基因的表達(dá)無影響；小鼠成纖維細(xì)胞defb2和slfn2基因表達(dá)都受到P38MAPK及NF-κB信號轉(zhuǎn)導(dǎo)通路調(diào)控。提示成纖維細(xì)胞可能通過誘導(dǎo)defb2、上調(diào)schlafen-2和白細(xì)胞介素-6的表達(dá)發(fā)揮機(jī)體抗感染的作用。
[Abstract]:Objective: to investigate the effect of LPS on the expression of 尾 -defb and schlafen slfn in mouse fibroblasts. And the regulation of P38 MAPK and NF- 魏 B signal transduction pathway on the expression of defb2 and slfn2 genes, to explore the mechanism of fibroblast defense against pathogenic microorganisms. It provides theoretical and experimental basis for further study on the mechanism of anti-infection. Methods: L929 cells were treated with L929 cells for 2 h and 4 h for 6 h with low, medium and high doses of LPSN 50ng / ml, 200ng / ml and 500ng / ml, respectively. L929 cells were treated with PBS at the same time as control group. Total RNAs were extracted from L929 cells. Defb1m-2 and slfn1m-2 were detected by RT-PCR and fluorescence quantitative PCR. 3Gene expression; Application of Western. The expression of defb2 protein. BMS345541 and PD169316 signal transduction pathway were detected by Blotting. The changes of defb2 and slfn2 gene and protein expression were observed after stimulation with LPS, and the signal transduction pathway of defb2 and slfn2 gene expression was discussed. Results: 1. LPS-induced expression of 尾 -defin -2tdefb _ 2 (尾 -defb _ 2) in mouse fibroblasts was the strongest at 2 h in the middle dose group. The expression of P-defin-1 was not induced. 2.The relative mRNA expression of schlafen2 / slfn2 was the highest at 2 h in the middle dose group. There was no significant difference in the expression of mRNA between the two groups at different concentrations and at different time. 3. The relative mRNA expression of IL-6 decreased gradually in the groups of 2h and 6h, respectively. There was no significant difference in the expression of IL-1 relative mRNA between the two groups at 2 h and 4 h after 6 h compared with the control group. 4. The expression of nRNA in the signal transduction effector molecule TLR4, MYD8, NF- 魏 B was significantly higher than that in the control group. 5. Compared with LPS group, the expression of defb2 and protein in BMS345541 and PD169316 group were significantly lower than those in LPS group. The expression of slfn2 was also significantly decreased relative to mRNA. Conclusion the expression of 尾-defensin-2 gene in mouse fibroblasts was induced by 1: 1 LPS, and the expression of schlafen-2 and IL-6 genes was up-regulated, and the expression of 尾-defensin-1 was up-regulated. The expression of Schlafen-1- 3 and interleukin-1 gene was not affected. The expression of defb2 and slfn2 genes in mouse fibroblasts were regulated by P38 MAPK and NF- 魏 B signal transduction pathway, suggesting that fibroblasts might induce defb2. Upregulation of schlafen-2 and IL-6 expression may play an anti-infection role.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R392.12

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