组蛋白去乙酰化酶9在骨髓间充质干细胞衰老过程中的表达

Expression of histone deacetylase 9 in bone marrow mesenchymal stem cells during senescence

背景:骨髓间充质干细胞在体外培养传代过程中存在细胞老化现象,但其机制尚不明确。组蛋白去乙酰化酶可以使组蛋白脱乙酰化而使染色质处于浓缩状态,进一步关闭基因。目的:探讨骨髓间充质干细胞衰老过程中组蛋白去乙酰化酶9的表达模式。方法:通过生物信息学挖掘技术分析组蛋白去乙酰化酶在骨髓间充质干细胞中的表达;通过体外传代培养骨髓间充质干细胞,β-半乳糖甘酶染色和qRT-PCR方法检测其衰老状态,qRT-PCR、Western blot及免疫荧光技术检测组蛋白去乙酰化酶9的表达模式。结果与结论:①数据挖掘、生物信息学分析显示在骨髓间充质干细胞衰老过程中,所有组蛋白去乙酰化酶亚型中,只有组蛋白去乙酰化酶9呈现高表达;②骨髓间充质干细胞在传至10代时可表现明显的衰老状态,如β-半乳糖甘酶染色阳性细胞最为明显,衰老相关基因表达上调;③组蛋白去乙酰化酶9的表达与数据挖掘结果一致,呈现上调;组蛋白去乙酰化酶9在年轻骨髓间充质干细胞的胞核与胞浆中均有表达,但其在胞核中的表达较低,衰老骨髓间充质干细胞胞浆中几乎不表达组蛋白去乙酰化酶9,而其在胞核中的表达增加;④结果表明,组蛋白去乙酰化酶9在骨髓间充质干细胞衰老过程中具有重要的作用,但其衰老表型与组蛋白去乙酰化酶9发挥作用的分子机制尚待进一步研究。

BACKGROUND:Bone marrow mesenchymal stem cells have aging phenomenon during in vitro culture,but the mechanism is still unclear.Histone deacetylases can deacetylate histones and make the chromatin condense,further shutting down genes.OBJECTIVE:To investigate the expression pattern of histone deacetylase 9 in bone marrow mesenchymal stem cells during senescence.METHODS:The expression of histone deacetylases in bone marrow mesenchymal stem cells was analyzed by bioinformatics.Bone marrow mesenchymal stem cells were cultured by in vitro subculture.The senescence status of bone marrow mesenchymal stem cells was detected byβ-galactosidase staining and qRT-PCR.The expression pattern of histone deacetylase 9 was detected by qRT-PCR,western blot assay,and immunofluorescence.RESULTS AND CONCLUSION:(1)Data mining and bioinformatics analysis showed that only histone deacetylase 9 expression was up-regulated among all histone deacetylase subtypes during bone marrow mesenchymal stem cell senescence.(2)Bone marrow mesenchymal stem cells presented obvious senescence at passage 10;for example,β-galactosidase-positive cells were the most obvious,and the expression of senescence-related genes was up-regulated.(3)The expression of histone deacetylase 9 was up-regulated and consistent with the data mining results.Histone deacetylase 9 was expressed in both the nucleus and cytoplasm of young bone marrow mesenchymal stem cells,but the expression in the nucleus was lower than that in the nucleus of senescence bone marrow mesenchymal stem cells.The expression of histone deacetylase 9 was almost not expressed in the cytoplasm of senescence bone marrow mesenchymal stem cells,but increased in the nucleus.(4)The results have shown that histone deacetylase 9 plays an important role in the senescence process of bone marrow mesenchymal stem cells,but the aging phenotype and molecular mechanism of histone deacetylase 9 need to be further studied.