本文選題：骨髓間充質干細胞 + 心肌細胞　； 參考：《蘇州大學》2009年碩士論文

【摘要】： 目的:通過體外實驗,觀察化學誘導劑和模擬的生物微環(huán)境誘導骨髓間充質干細胞向心肌細胞分化。 方法:取6~8w齡SD大鼠股骨骨髓,培養(yǎng)至第二代,經流式細胞儀檢測骨髓間充質干細胞純度97%以上,然后分成五組:A組,單純DMEM細胞培養(yǎng)組(對照組);B組,5-氮雜胞苷誘導組;C組,心肌細胞裂解液組;D組,心肌細胞裂解液組+5-氮雜胞苷組;E組,細胞間接接觸組。根據各自實驗條件誘導至第2w和第4w時,收獲細胞。應用免疫組化法檢測誘導后骨髓間充質干細胞表達心肌特異性肌鈣蛋白T(cTnT)、連接蛋白43(CX-43),血管內皮細胞表面抗原標志CD31及肌性標志α-橫紋肌肌動蛋白(α-SCA)的表達情況。應用RT-PCR法檢測各組早期心肌細胞特異性轉錄因子GATA4、NKX2.5、β-MHC及成熟心肌細胞特異性轉錄因子α-MHC基因的表達情況,并進行簡單量的比較。 結果:免疫組化檢測:實驗組誘導2w后,cTnT、CX-43表達陰性或低表達,誘導4w后呈陽性表達,實驗組較對照組有統(tǒng)計學差異(P0.05),實驗組之間無顯著差異(P0.05);對照組及B組無CD31表達,C, D, E組表達呈弱陽性,組間無統(tǒng)計學差異(P0.05);α-SCA誘導前骨髓間充質細胞中既有低度表達,誘導后濃度明顯增高,實驗組間無差異(P0.05)。 RT-PCR表達情況:實驗組誘導2w后條帶不明顯;誘導4w后均表達GATA4, NKX2.5,β-MHC,無α-MHC表達,從條帶密度和亮度分析,實驗組較對照組有顯著差異(P0.05),實驗組間無顯著差異(P0.05)。 結論: 1. 5-氮雜胞苷可以誘導骨髓間充質干細胞向心肌細胞分化。 2.骨髓間充質干細胞因其“環(huán)境依賴性分化”特性,具有分化為心肌細胞的潛能。心肌細胞裂解液和細胞間接接觸模擬的心肌微環(huán)境也可以誘導骨髓間充質干細胞分化成心肌樣細胞,且誘導的效果與誘導的時間及誘導劑濃度相關,分化的細胞介于成熟的心肌細胞和心肌祖細胞之間的心肌細胞前體。
[Abstract]:Aim: to observe the differentiation of bone marrow mesenchymal stem cells (BMSCs) into cardiomyocytes induced by chemical inducers and simulated microenvironment in vitro. Methods: bone marrow of SD rats aged 6 to 8 weeks was cultured to the second generation. The purity of bone marrow mesenchymal stem cells was detected by flow cytometry. The bone marrow mesenchymal stem cells were divided into five groups: group A, group B (control group) and group C (group C) induced by 5-azacytidine. Group D was treated with cardiomyocyte lysate, group E with 5-azacytidine, group with indirect cell contact. According to the experimental conditions, the cells were harvested at the 2nd and 4th week. The expression of myocardial specific troponin T (cTnT), connexin 43 (CX-43), CD31 and 偽 -rhabdomyactin (偽 -SCA) in bone marrow mesenchymal stem cells (BMSCs) were detected by immunohistochemistry. RT-PCR was used to detect the expression of NKX2.5, 尾 -MHC and 偽 -MHC genes in the early cardiac myocyte specific transcription factor GATA4, NKX2.5, and mature cardiomyocyte specific transcription factor 偽 -MHC, and a simple comparison was made. Results: the expression of cTnTnTnTnTnTnCX-43 in the experimental group was negative or low after 2 weeks of induction, and was positive after 4 weeks of induction. There was statistical difference between the experimental group and the control group (P0.05), there was no significant difference between the experimental group and the experimental group (P0.05); in the control group and group B, the expression of CD31 was weakly positive, but there was no statistical difference between the two groups (P0.05); there was a low expression in the bone marrow mesenchymal cells before 偽 -SCA induction. After induction, the concentration of GATA4, NKX2.5, 尾 -MHC and 偽 -MHC were significantly increased (P0.05). The expression of RT-PCR: the bands were not obvious after 2 weeks of induction, but GATA4, NKX2.5, 尾 -MHCand 偽 -MHC were all expressed at 4 weeks after induction. There was significant difference between the experimental group and the control group (P0.05), but there was no significant difference between the experimental group (P0.05). Conclusion: 1. 5-azacytidine could induce the differentiation of bone marrow mesenchymal stem cells into cardiomyocytes. 2. Bone marrow mesenchymal stem cells (BMSCs) have the potential to differentiate into cardiomyocytes due to their environmental dependent differentiation. Myocardial cell lysate and cell contact with the simulated myocardial microenvironment could also induce bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells, and the induction effect was related to the time of induction and the concentration of inducer. Differentiated cells are precursors of cardiac myocytes between mature cardiomyocytes and cardiac progenitor cells.
【學位授予單位】：蘇州大學
【學位級別】：碩士
【學位授予年份】：2009
【分類號】：R329

【引證文獻】

相關碩士學位論文 前1條

1 閆穎穎;兔骨髓間充質干細胞向心肌樣細胞的體外誘導分化研究[D];西北農林科技大學;2010年

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本文編號：2080066