【摘要】：目的：研究正常腦組織、不同級別膠質瘤組織以及膠質瘤細胞系U87中H3乙�；M蛋白的表達，使用去乙�；敢种苿〢picidin干擾U87膠質瘤細胞系，研究組蛋白H3乙�；潭雀淖儗δz質瘤細胞中Nanog的影響，并探索組蛋白H3低乙�；瘜δz質瘤細胞體內及體外生物學活性的影響。為后續(xù)進一步研究Nanog的表觀遺傳學干預治療奠定基礎。 方法：①使用Western-Blot方法檢測53例不同級別腦膠質瘤組織及11例取自顱腦損傷行內減壓術患者的正常腦組織中H3乙�；M蛋白的表達情況，RealtimePCR檢測高級別和低級別膠質瘤組織內Nanog表達情況。②通過組蛋白去乙�；敢种苿〢picidin干預U87膠質瘤細胞，不同濃度和時間干預過后，，通過RT-PCR和Western-blot實驗分別檢測U87細胞株中H3乙酰化組蛋白和Nanog的mRNA和蛋白表達水平，ChIP檢測U87細胞系中Nanog啟動子區(qū)域H3組蛋白乙酰化水平。③通過MTT、細胞劃痕實驗、流式細胞術檢測細胞凋亡、細胞周期、透射電鏡檢測凋亡、裸鼠成瘤實驗、免疫組織化學染色等方法檢測組蛋白H3低乙酰化水平抑制Nanog后對膠質瘤細胞體內外生物學活性的影響。 結果：①與正常腦組織相比，乙�；M蛋白H3在不同級別膠質瘤中的表達均明顯上調（P＜0.05）。WHO Ⅲ～Ⅳ級膠質瘤組織中乙酰化組蛋白H3表達明顯高于WHOⅠ～Ⅱ級（P＜0.05）。與正常腦組織相比，在膠質瘤細胞系U87中的表達亦顯著升高（P＜0.05）。②利用組蛋白去乙酰化酶抑制劑Apicidin干預膠質瘤U87細胞系后，同時檢測乙酰化組蛋白H3及Nanog表達水平的變化，發(fā)現(xiàn)Nanog啟動子區(qū)域的組蛋白H3乙酰化水平降低，同時Nanog的表達亦明顯降低；③與空白對照組相比，U87-Apicidin膠質瘤細胞的增殖水平和遷移能力均減弱，細胞凋亡增加； U87-Apicidin皮下膠質瘤的體積與質量都明顯小于空白對照組，提示Apicidin處理組U87細胞裸鼠皮下成瘤能力亦明顯下降（P＜0.05）。 結論：①組蛋白H3乙�；赡芘c膠質瘤的發(fā)生發(fā)展關系密切。膠質瘤級別越高，乙酰化組蛋白H3的表達量越高；②Nanog啟動子區(qū)域的組蛋白H3乙酰化修飾水平可以影響Nanog的表達；③Apicidin可以特異性的降低Nanog啟動子區(qū)域乙酰化組蛋白H3水平，從而影響Nanog的表達，最終對膠質瘤的生物學活性產生抑制性作用。
[Abstract]:Objective: to study the expression of H3 acetylated histone in normal brain tissue, glioma tissue of different grades and glioma cell line U87, and to use deacetylase inhibitor Apicidin to interfere with U87 glioma cell line. The effect of acetylation of protein H3 on Nanog in glioma cells and the effect of low acetylation of histone H3 on the biological activity of glioma cells in vivo and in vitro were investigated. To lay a foundation for the further study of epigenetic intervention therapy of Nanog. Methods: 1the expression of H3 acetylated histone was detected by Western-Blot in 53 gliomas with different grades and 11 normal brain tissues from patients with craniocerebral injury undergoing internal decompression. RealtimePCR was used to detect the expression of Nanog in high grade and low grade gliomas. (2) U87 glioma cells were treated with histone deacetylase inhibitor (Apicidin). The mRNA and protein expressions of H3 acetylated histone and Nanog in U87 cell line were detected by RT-PCR and Western-blot assay respectively. The H3 histone acetylation level of Nanog promoter region in U87 cell line was detected by ChIP. 3 the H3 histone acetylation level was detected by MTT, cell scratch assay. Flow cytometry was used to detect cell apoptosis, cell cycle, transmission electron microscopy (TEM), and tumorigenesis in nude mice. The effects of low acetylation of histone H3 on the biological activity of glioma cells in vivo and in vitro were detected by immunohistochemical staining. Results: 1Compared with normal brain tissue, The expression of acetylated histone H3 was significantly up-regulated in different grade gliomas (P < 0. 05). The expression of acetylated histone H3 in gliomas of grade 鈪

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