發(fā)布時間：2018-06-09 21:08

  本文選題：干擾素 + 樂復能　； 參考：《中南大學》2014年碩士論文

【摘要】：目的：檢驗樂復能是否具有種間交叉生物活性；測定樂復能在VSV感染的小鼠成纖維細胞系L929中的抗病毒活性；探討其進一步進行臨床前的人類疾病小鼠模型研究的可行性。 方法：培養(yǎng)小鼠成纖維細胞系L929,將其制備成細胞懸液,接種于96孔板中置溫箱中過夜培養(yǎng)。以不同的濃度起始,連續(xù)對倍稀釋樂復能和重組小鼠干擾素α1(MuIFN-α1),并依次加入96孔板的培養(yǎng)細胞中,設置正常細胞對照孔,繼續(xù)培養(yǎng)24小時。棄培養(yǎng)液后,在樂復能組、MuIFN-α1組添加含有VSV病毒的完全培養(yǎng)基(MOI:0.1),并設置正常細胞對照組和病毒對照組,連續(xù)培養(yǎng)36小時。吸出細胞培養(yǎng)液,0.85%NaCl漂洗后拍干,添加細胞染色液室溫下染色2小時。自來水漂凈染色液,室溫下完全干燥后,每孔加入2-甲氧基乙醇(2-methoxyethanol),靜置45分鐘脫色,酶標儀檢測各細胞孔的吸光值。實驗重復三次,結果取平均值,并計算保護率。以藥物濃度的對數(shù)值為橫軸,保護率為縱軸分別繪制樂復能和MuIFN-α1在L929-VSV系統(tǒng)中的回歸分析圖,并通過線性回歸方法計算兩種藥物的50%最大效應濃度(EC50)。 結果：①樂復能組和MuIFN-α1組細胞的OD550值均明顯升高。計算兩組藥物對小鼠L929細胞的保護率,均隨著藥物濃度的增高而增大,兩者呈正相關(樂復能：r=0.972, p0.001; MuIFN-α=0.950,p0.001)。通過藥物濃度和細胞保護率相關曲線發(fā)現(xiàn),相同濃度下,MuIFN-α1在小鼠L929細胞中的對細胞的保護率較樂復能高,即抗病毒活性更明顯。②通過線性回歸的方法計算出樂復能和MuIFN-α1的EC50值分別為：170.752pg和38.810pg。由此推算樂復能在L929-VSV系統(tǒng)中抗病毒活性為6.32×106IU/mg。證實樂復能在VSV病毒感染的小鼠L929細胞中具有良好的抗病毒活性,提示其具有較強的種間交叉作用。 結論：①樂復能和MuIFN-α1均對感染VSV病毒的小鼠L929細胞具有保護作用,且保護率與藥物濃度呈正相關；②相同藥物濃度下,MuIFN-α1對小鼠L929細胞的保護率較樂復能高,MuIFN-α1的抗病毒活性高于樂復能；③樂復能在VSV病毒感染的小鼠L929細胞中具有較強的種間交叉作用,可用于樂復能小鼠動物實驗。
[Abstract]:Objective: to test whether lofenin has interspecific cross biological activity and to determine its antiviral activity in VSV infected mouse fibroblast cell line L929. Methods: the mouse fibroblast cell line L929 was cultured and the suspension was prepared and cultured overnight in a 96-well plate. At the beginning of different concentrations, the cells were continuously diluted with Lefergic and recombinant mouse interferon 偽 1, MuIFN- 偽 1, and added in turn to 96 well plate culture cells. The normal control cells were set up and cultured for 24 hours. After the culture medium was abandoned, the complete medium containing VSV virus was added to the MuIFN- 偽 1 group, and the normal cell control group and the virus control group were set up for 36 hours. The cell culture medium (0.85 NaCl) was bleached and dried, and the cells were stained at room temperature for 2 hours. After completely drying at room temperature, 2-methoxyethanolol was added to each pore, and the dye was decolorized for 45 minutes. The absorptivity of each cell was measured by enzyme labeling instrument. The experiment was repeated three times, the results were averaged and the protection rate was calculated. The logarithmic values of drug concentration were taken as horizontal axis and the protection rate was used as longitudinal axis to draw the regression analysis of L929-VSV system for L929-VSV system with L929-VSV system and MuIFN- 偽 1, respectively. The 50% maximum effective concentration (MEC) of the two drugs was calculated by linear regression method. Results the OD550 values of cells in the two groups were significantly higher than those in the control group and MuIFN- 偽 1 group. The protective rate of the two drugs on L929 cells increased with the increase of drug concentration, and there was a positive correlation between the two groups (Le Fu Neng 0.972, p 0.001; MuIFN- 偽 0.950 p 0.001). It was found that the protective rate of MuIFN- 偽 1 in mouse L929 cells at the same concentration was higher than that in L929 cells. The EC50 values of Lefeng and MuIFN- 偽 1 were calculated by linear regression method, and the EC50 values were: 1: 170.752pg and 38.810pg. respectively. The antiviral activity of L929-VSV was estimated to be 6.32 脳 10 ~ (6) IU / mg. It was confirmed that L929 cells infected with VSV virus had good antiviral activity, which indicated that L929 cells infected with VSV virus had strong interspecific cross effect. Conclusion both of them have protective effect on L929 cells infected with VSV virus. The protective rate of MuIFN- 偽 1 on L929 cells was higher than that of L929 cells at the same drug concentration, and the antiviral activity of MuIFN- 偽 1 was higher than that of L929 cells. 3 lofenac has strong interspecific cross effect in L929 cells infected with VSV virus, and it can be used in the animal experiment of L929 cells infected with VSV virus.
【學位授予單位】：中南大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R965

【參考文獻】

相關期刊論文 前1條

1 李富軍;薛梅;盧放根;鄒益友;;樂復能對LPS介導的健康人外周血單核細胞分泌TNF-α的影響及其機制[J];中南大學學報(醫(yī)學版);2013年01期

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本文編號：2000836