本文選題：骨髓間充質(zhì)干細(xì)胞 + 子宮內(nèi)膜間質(zhì)細(xì)胞��； 參考：《中南大學(xué)》2012年博士論文

【摘要】：目的：體外誘導(dǎo)骨髓間充質(zhì)干細(xì)胞(BMSCs)向子宮內(nèi)膜細(xì)胞分化,為干細(xì)胞移植治療薄型子宮內(nèi)膜提供實(shí)驗(yàn)基礎(chǔ)。 方法：分離、培養(yǎng)、鑒定大鼠BMSCs和人子宮內(nèi)膜間質(zhì)細(xì)胞(hECs)。將BMSCs和hECs在Transwell培養(yǎng)板中共培養(yǎng),添加雌激素(E2,1×10-7mol/L)和生長(zhǎng)因子(GF,10ng/ml TGF-β、10ng/ml EGF、 lOng/ml PDGF-BB),在培養(yǎng)的第7天免疫熒光法檢測(cè)BMSCs的波形蛋白、角蛋白表達(dá)情況。在普通培養(yǎng)板中培養(yǎng),將BMSCs和hECs按細(xì)胞組成分為3組：co-culture組、BMSCs組和hECs組(其中co-culture組為BMSCs和hECs直接共培養(yǎng),另外兩組僅培養(yǎng)一種細(xì)胞)。每組中采用3種不同的誘導(dǎo)方式,分別加入:E2+GF(1×10-7mol/L E2,10ng/ml TGF-β、10ng/ml EGF、10ng/ml PDGF-BB); cAMP (1mmol/ml8-Br-cAMP); E2+cAMP (1×10-7mol/L E2、1mmol/ml8-Br-cAMP)。在培養(yǎng)的第14天和第20天通過(guò)細(xì)胞免疫熒光法檢測(cè)各組細(xì)胞波形蛋白和角蛋白表達(dá)情況,第3天和第19天酶聯(lián)免疫吸附法(ELISA)檢測(cè)蛻膜化標(biāo)記泌乳素(PRL)和胰島素樣結(jié)合蛋白1(IGFBP1)濃度。 結(jié)果：通過(guò)細(xì)胞培養(yǎng)獲得了穩(wěn)定增殖的大鼠BMSCs和人子宮內(nèi)膜間質(zhì)細(xì)胞。在Transwell培養(yǎng)板中共培養(yǎng)的BMSCs第7天波形蛋白表達(dá)陽(yáng)性,角蛋白表達(dá)陰性。普通培養(yǎng)板中各組細(xì)胞在培養(yǎng)的第14天和第20天細(xì)胞波形蛋白表達(dá)均為陽(yáng)性,角蛋白表達(dá)均為陰性。第3天和第19天兩種蛋白在組間及組內(nèi)的表達(dá)均沒(méi)有顯著統(tǒng)計(jì)學(xué)差異(P0.05)；PRL和IGFBP1的表達(dá)第19天明顯高于第3天,結(jié)果有統(tǒng)計(jì)學(xué)差異(P0.001)。 結(jié)論：①骨髓間充質(zhì)干細(xì)胞可以向子宮內(nèi)膜間質(zhì)細(xì)胞分化；②雌激素、生長(zhǎng)因子、cAMP類似物在骨髓間充質(zhì)干細(xì)胞向子宮內(nèi)膜間質(zhì)細(xì)胞分化中起到誘導(dǎo)作用,且誘導(dǎo)能力相當(dāng)；③本實(shí)驗(yàn)條件下骨髓間充質(zhì)干細(xì)胞未能向子宮內(nèi)膜上皮細(xì)胞分化。
[Abstract]:Aim: to induce the differentiation of bone marrow mesenchymal stem cells (BMSCs) into endometrial cells in vitro. Methods: rat BMSCs and human endometrial stromal cells (hECs) were isolated, cultured and identified. BMSCs and hECs were co-cultured on Transwell culture plate. The expression of vimentin and keratin of BMSCs were detected by immunofluorescence on the 7th day after adding estrogen (E2N 1 脳 10-7mol / L) and growth factor (GFF10ngr / ml TGF- 尾 -10ng / ml EGF-, lOng / ml PDGF-BB). BMSCs and hECs were divided into three groups according to cell composition: BMSCs and hECs (co-culture group was directly co-cultured with hECs, and only one cell was cultured in the other two groups). In each group, three different induction methods were used, respectively, with 1 脳 10 ~ (-7) mol / L E _ (2) E _ (2) O _ (10) ng / ml TGF- 尾 (10 ~ (-10) ng / ml EGFN _ (10) ng / ml PDGF-BB); camp (1 mmol / ml 8-Br-cAMP); E _ (2) camp (1 脳 10 ~ (-7) mol / L E _ 2 路mol / ml 路ml ~ (-1) Br-cAMP). The expression of vimentin and keratin was detected by cell immunofluorescence on the 14th and 20th day of culture. The concentrations of decidualized prolactin (PRL) and insulin-like binding protein 1 (IGFBP1) were detected by enzyme-linked immunosorbent assay (Elisa) on day 3 and day 19. Results: rat BMSCs and human endometrial stromal cells were obtained by cell culture. BMSCs co-cultured on Transwell culture plate showed positive expression of vimentin and negative expression of keratin on day 7. The expression of vimentin was positive on the 14th and 20th day, and keratin expression was negative in the normal culture plate. There was no significant difference in the expression of PRL and IGFBP1 between groups on day 3 and day 19 (P0.05). The expression of PRL and IGFBP1 on day 19 was significantly higher than that on day 3 (P0.001). Conclusion Bone marrow mesenchymal stem cells (BMSCs) can differentiate into endometrial stromal cells. Growth factor camp analogues play an important role in inducing the differentiation of BMSCs into endometrial stromal cells. 3 in this experiment, bone marrow mesenchymal stem cells could not differentiate into endometrial epithelial cells.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2012
【分類號(hào)】：R329

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