本文選題：雷公藤內(nèi)醇酯　切入點(diǎn)：多發(fā)性骨髓瘤　出處：《中國(guó)實(shí)驗(yàn)血液學(xué)雜志》2017年04期 　論文類型：期刊論文

【摘要】：目的:探討雷公藤內(nèi)酯醇(triptolide,TPL)抑制組蛋白甲基化酶SMYD3對(duì)多發(fā)性骨髓瘤RPMI8226細(xì)胞增殖和凋亡的影響及其作用機(jī)制。方法:采用MTT法檢測(cè)不同濃度(10、20、40、80、160 nmol/L)TPL作用不同時(shí)間(24、48、72 h)對(duì)RPMI8226細(xì)胞增殖的影響;應(yīng)用流式細(xì)胞術(shù)檢測(cè)不同濃度(40、80、160 nmol/L)TPL作用48 h后RPMI8226細(xì)胞的凋亡率;實(shí)時(shí)熒光定量PCR檢測(cè)SMYD3和MMP-9基因的mRNA表達(dá)水平;Western blot法檢測(cè)組蛋白H3K4me2,H3K4me3的甲基化水平。結(jié)果:TPL能夠明顯抑制RPMI8226細(xì)胞的增殖活性,且隨作用時(shí)間延長(zhǎng)及藥物濃度增高,細(xì)胞增殖抑制率明顯升高(P0.05);不同濃度TPL作用RPMI8226細(xì)胞48 h后,隨著藥物濃度的增高,細(xì)胞凋亡比例逐漸增加,與空白對(duì)照組比較,差異均有統(tǒng)計(jì)學(xué)意義(r=0.974,P0.05);實(shí)時(shí)熒光定量PCR結(jié)果顯示,不同濃度TPL作用RPMI8226細(xì)胞48 h后,隨著藥物濃度的增高,SMYD3的mRNA相對(duì)表達(dá)水平呈濃度依賴性下降,與空白對(duì)照組比較,差異均有統(tǒng)計(jì)學(xué)意義(r=-0.882,P0.05);siRNA-SMYD3轉(zhuǎn)染RPM I8226細(xì)胞48 h后,siRNA-SM YD3組與空白對(duì)照組比較MMP-9的mRNA相對(duì)表達(dá)水平明顯下降,與80nmol/L的TPL作用一致;Western blot結(jié)果顯示,不同濃度的TPL作用RPM I8226細(xì)胞48 h后,隨著藥物濃度的增高,組蛋白H3K4me2和H3K4me3的甲基化水平呈濃度依賴性下降,分別與空白對(duì)照組比較,差異均有統(tǒng)計(jì)學(xué)意義(r=-0.971,r=-0.985,P0.05);siRNA-SMYD3轉(zhuǎn)染RPMI8226細(xì)胞48 h,組蛋白H3K4me2和H3K4me3的甲基化水平也明顯下降,分別與siRNA陰性對(duì)照組、空白對(duì)照組比較,差異均有統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論:TPL對(duì)多發(fā)性骨髓瘤RPMI8226細(xì)胞增殖具有明顯的抑制作用,并可誘導(dǎo)細(xì)胞凋亡,其機(jī)制與抑制SMYD3,調(diào)控組蛋白H3K4甲基化和MMP-9基因轉(zhuǎn)錄激活有關(guān)。
[Abstract]:Objective: to investigate the effect of triptolide triptolide triptolide (TPL) on the proliferation and apoptosis of multiple myeloma RPMI8226 cells induced by histone methylase SMYD3. Methods: MTT assay was used to detect the effect of triptolide triptolide (TPL) on the proliferation and apoptosis of multiple myeloma RPMI8226 cells. Effects on proliferation of RPMI8226 cells; Flow cytometry was used to detect the apoptotic rate of RPMI8226 cells after 48 h treatment with different concentrations of 40,80,160 nmol/L)TPL. Real-time fluorescence quantitative PCR was used to detect the mRNA expression of SMYD3 and MMP-9 genes. Western blot assay was used to detect the methylation level of histone H3K4ME2H3K4me3.Results: TPL could significantly inhibit the proliferative activity of RPMI8226 cells and increase the drug concentration with the prolongation of time and the increase of drug concentration. The inhibition rate of cell proliferation was significantly increased (P 0.05), and the ratio of apoptosis of RPMI8226 cells increased with the increase of drug concentration after 48 h treatment with different concentrations of TPL, compared with the control group. The results of real-time fluorescence quantitative PCR showed that the relative mRNA expression of RPMI8226 cells decreased in a concentration-dependent manner with the increase of drug concentration after 48 h treatment with different concentrations of TPL, compared with the control group. The difference was statistically significant. The relative expression of MMP-9 mRNA in the siRNA-SMYD3 transfected RPM I8226 cells 48 h after transfection was significantly lower than that in the control group, which was consistent with the effect of 80 nmol / L TPL. The results showed that different concentrations of TPL treated RPM I8226 cells for 48 h. The methylation level of histone H3K4me2 and H3K4me3 decreased in a concentration-dependent manner with the increase of drug concentration, and the methylation levels of histone H3K4me2 and H3K4me3 were significantly decreased after transfection of histone H3K4me2 and H3K4me3 into RPMI8226 cells for 48 h after transfection of histone H3K4me2 and H3K4me3. Compared with siRNA negative control group and blank control group, the difference was statistically significant (P 0.05). Conclusion: TPL can significantly inhibit the proliferation of multiple myeloma RPMI8226 cells and induce apoptosis. The mechanism is related to inhibition of SMYD3, regulation of histone H3K4 methylation and transcriptional activation of MMP-9 gene.
【作者單位】： 福建省人民醫(yī)院福建中醫(yī)藥大學(xué)附屬人民醫(yī)院血液科;福建醫(yī)科大學(xué)附屬協(xié)和醫(yī)院血液科福建省血液病研究所;
【基金】：福建省自然科學(xué)基金項(xiàng)目(2015J01327) 福建省衛(wèi)計(jì)委青年科研基金(2016-1-77)
【分類號(hào)】：R733.3

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