【摘要】： [目的]研究腎上腺髓質(zhì)素(ADM)基因轉(zhuǎn)染對骨髓間充干細(xì)胞(MSCs)在缺氧及無血清培養(yǎng)條件下的增殖、凋亡、分泌血管內(nèi)皮生長因子(VEGF)的影響。 [方法]貼壁法體外分離、擴(kuò)增培養(yǎng)大鼠骨髓間充質(zhì)干細(xì)胞。用x-gal染色測含腎上腺髓質(zhì)素基因的重組腺病毒(Ad-ADM)對骨髓間充質(zhì)干細(xì)胞感染率。骨髓間充質(zhì)干細(xì)胞分為未轉(zhuǎn)染組,空載體組,Ad-ADM轉(zhuǎn)染組。在缺氧、無血清條件下進(jìn)行骨髓間充質(zhì)干細(xì)胞的培養(yǎng),在缺氧0、3、6、9、12、16、20、24h CCK-8檢測細(xì)胞增殖、ELISA檢測檢測細(xì)胞上清液腎上腺髓質(zhì)素及血管內(nèi)皮生長因子的含量、Tunel法檢測細(xì)胞凋亡。 [結(jié)果]重組腺病毒對骨髓間充質(zhì)干細(xì)胞的轉(zhuǎn)染率與病毒感染復(fù)數(shù)(MOI)具有量效關(guān)系,MOI為150時細(xì)胞的感染率達(dá)95.4%。缺氧、無血清培養(yǎng)條件下,3組骨髓間充質(zhì)干細(xì)胞均出現(xiàn)生長抑制、凋亡增加,但是腎上腺髓質(zhì)素基因轉(zhuǎn)染組于0,3,6,9、12、16h細(xì)胞凋亡率顯著低于其它2組(P0.05)。缺氧9,12h,骨髓間充質(zhì)干細(xì)胞分泌腎上腺髓質(zhì)素及血管內(nèi)皮生長因子達(dá)到高峰,且Ad-ADM轉(zhuǎn)染組顯著增高于其它2組(P0.05)。 [結(jié)論]在缺氧、無血清培養(yǎng)條件下(20h),腎上腺髓質(zhì)素基因轉(zhuǎn)染增強(qiáng)骨髓間充質(zhì)干細(xì)胞抗凋亡能力,這可能與腎上腺髓質(zhì)素、血管內(nèi)皮生長因子表達(dá)增加有關(guān)。
[Abstract]:[objective] to study the effect of adrenomedullin (ADM) gene transfection on proliferation, apoptosis and secretion of vascular endothelial growth factor (VEGF) in bone marrow mesenchymal stem cells (MSCs) in hypoxic and serum-free culture. [methods] Rat bone marrow mesenchymal stem cells were isolated and cultured by adherent method in vitro. The infection rate of recombinant adenovirus containing adrenomedullin gene (Ad-ADM) on bone marrow mesenchymal stem cells (BMSCs) was detected by x-gal staining. Bone marrow mesenchymal stem cells were divided into untransfected group, empty vector group and Ad-ADM transfection group. Bone marrow mesenchymal stem cells (BMSCs) were cultured in anoxic and serum-free condition. Cell proliferation was detected by CCK-8 at 24 h after hypoxia, and the content of adrenomedullin and vascular endothelial growth factor (VEGF) in supernatant was detected by ELISA. Apoptosis was detected by Tunel. [results] the transfection efficiency of recombinant adenovirus on bone marrow mesenchymal stem cells was dose-dependent with the infection of complex (MOI), and the infection rate reached 95.4when MOI was 150. Under the condition of anoxia and serum-free culture, the growth inhibition and apoptosis of bone marrow mesenchymal stem cells in all three groups were inhibited, but the apoptotic rate in the adrenomedullin gene transfection group was significantly lower than that in the other two groups (P0.05). The secretion of adrenomedullin and vascular endothelial growth factor (VEGF) from bone marrow mesenchymal stem cells reached the peak at 9h after hypoxia, and the levels of adrenomedullin and vascular endothelial growth factor (VEGF) in the Ad-ADM transfected group were significantly higher than those in the other two groups (P0.05). [conclusion] under hypoxic and serum-free culture conditions (20 h), adrenomedullin gene transfection enhances the anti-apoptosis ability of bone marrow mesenchymal stem cells, which may be related to the increased expression of adrenomedullin and vascular endothelial growth factor.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2010
【分類號】：R329

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