【作者】 黄生祥；

【导师】 胡懿郃；

【作者基本信息】 中南大学 ， 外科学， 2009， 硕士

【摘要】 目的:研究体外实验中榄香烯对人骨肉瘤细胞系（MG-63）多药耐药的逆转效果,探讨与GST-π的关系。方法:采用MTT法测定阿霉素对人骨肉瘤细胞系（MG-63）的半数抑制浓度(IC₅₀)以及榄香烯对人骨肉瘤细胞系（MG-63）的低细胞毒浓度(IC₁₅)和非细胞毒浓度(IC₀₅)。将细胞分为A、B、C三组,A组为对照组,加入半数抑制浓度的阿霉素;B、C组在A组基础上还加入非细胞毒浓度、低细胞毒浓度的榄香烯,细胞传代培养至A组中MG-63细胞对半数抑制浓度的阿霉素完全耐药后停止干预,检测以下指标:①阿霉素对干预后各组MG-63细胞的半数抑制浓度,计算耐药倍数（RF）;②采用荧光分光光度计测定各组OD值,计算细胞内阿霉素药物浓度;③免疫组化法测定原始MG-63细胞和A、B、C组MG-63细胞谷胱甘肽S-转移酶-π（GST-π）的表达,并对以上数据进行分析比较各处理组间的差异。结果:①阿霉素对干预后A、B、C组MG-63细胞的半数抑制浓度分别为3.37μg/ml、2.21μg/ml、1.19μg/ml,A、B、C组MG-63细胞对阿霉素的耐药倍数（RF）分别为11.62、7.62和4.10,A组＞B组＞C组,差异有统计学意义。②干预后A、B、C组MG-63细胞内阿霉素药物浓度分别为1.21μg/10⁶细胞、1.96μg/10⁶细胞、2.71μg/10⁶细胞,A组＜B组＜C组,差异有统计学意义。③细胞GSTs的表达率:原始MG-63细胞为10.25%、A组为35.75%、B组为25.25%、C组为20.5%,A组＞B组＞C组＞原始MG-63细胞,差异有统计学意义。结论:①证实在体外运用非细胞毒浓度或低细胞毒浓度的榄香烯乳能有效抑制阿霉素所诱导的人骨肉瘤细胞MG-63的多药耐药性（MDR）。②榄香烯体外能下调人骨肉瘤MG-63细胞GST-π的表达。③榄香烯乳逆转人骨肉瘤MG-63细胞产生多药耐药性（MDR）机制可能是通过抑制GST-π的过度表达,提高细胞内抗癌药的药物浓度而实现的。更多还原

【Abstract】 Objective:To study the Reversal Effect of Elemene on Multidrug-resistant of human osteosarcoma cell line（MG-63/ADM） in vitro and investigated the relationship between GST-πand the effect.Methods:To investigate the IC₅₀ of adriamycin（ADM）,IC₀₅ and IC₁₅ of elemene（ELE） to MG-63 human osteosarcoma cell by MTT assay. The MG-63 human osteosarcoma cell divide into three groups randomly （A、B、C Group）.Group A is the control group,to establish a multi-drug resistant of human MG-63 cells by short-time pulse exposure of the cell to the IC₅₀ of adriamycin.The cell of Group B、C are exposed both in the same dosage of ADM with the control group and IC₀₅（group B） or IC₁₅ （group C）of elemene.The exposure continued until the cell of Group A can be cultured stably in the nutritive medium which contain IC₅₀ of ADM,then drug exposure are stopped in all experiment Groups.we repeat to investigate the IC₅₀ of adriamycin to all post-drug intervention MG-63 human osteosarcoma cells by MTT assay.And intracellular accumulation of ADM were determined by fluorospectrophotometery. Immunohistochemistry was applied to study the GST-πexpression of initial and post-drug intervention MG-63 cells.At the end,we analyse the experimental result by statistics. Results:①The IC₅₀ of adriamycin（ADM） to MG-63 cells were 3.37μg/ml（GroupA）、2.21μg/ml（GroupB）、1.19μg/ml（GroupC）,and the RF values were 11.62（GroupA）、7.62（GroupB） and 4.10（GroupC）,the comparisons among Group A,GroupB and Group C have statistical significance（P＜0.05）.②The intracellular accumulation of ADM all experiment groups were 1.21μg/10⁶ cells（Group A）、1.96μg/10⁶ cells（GroupB）、2.71μg/10⁶ cells（Group C）,Group A＜GroupB＜Group C,the comparisons among Group A,GroupB and Group C have statistical significance（P＜0.05）.③The GST-πexpression rates of initial and post-drug intervention MG-63 cells were 10.25%（initial cells）, 35.75%（Group A）,25.25%（Group B） and 20.50%（Group C）,the comparisons among Group A,GroupB and Group C have statistical significance（P＜0.05）Conclusion:Firstly,we conclude that the multi-drug resistance to ADM of human osteosacoma cell（MG-63/ADM） could be reversed by Elemene with non-toxic or low-toxic dosage;Secondly the expression of GST-πcould be regulated down by Elemene;The last,the increased intracellular accumulation of ADM and downregulation of GST-πwould contribute to the reversal mechanism of Elemene.更多还原