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目的研究雌性大鼠骨髓干细胞(BMSCs)在含10%胎牛血清的高糖DMEM体外培养条件下能否分化为产生雌二醇(E2)及表达芳香化酶(aromatase,P450arom)的细胞,为卵巢早衰、老年人骨质疏松症等疾病的治疗提供新的可能途径。方法体外分离成年雌性SD大鼠的骨髓干细胞,用含10%胎牛血清的高糖DMEM培养增殖,加或不加全反式视黄酸(all-trans retinoic acid,ATRA)诱导分化,分为对照组(不加ATRA)和ATRA组(加ATRA),放射性免疫法(RIA)检测两组细胞培养液中E2的浓度;RT-PCR检测两组细胞中芳香化酶基因CYP19 mRNA的表达和FSHR mRNA的表达;免疫细胞化学检测两组细胞中芳香化酶的表达情况。离体培养第3代雌性大鼠骨髓干细胞,RT-PCR示对照组和ATRA组从培养第1天开始均表达CYP19 mRNA和FSHR mRNA,免疫细胞化学示两组细胞从培养第2~4天都有P450arom的阳性细胞表达,从培养24 h开始在两组细胞培养液中都检测到较高水平的E2。结果离体培养第3代雌性大鼠骨髓干细胞,RT-PCR示对照组和ATRA组从培养第1天开始均表达CYP19 mRNA和FSHR mRNA,免疫细胞化学示两组细胞从培养第2~4天都有P450arom的阳性细胞表达,从培养24 h开始在两组细胞培养液中都检测到较高水平的E2。结论雌性大鼠骨髓干细胞在离体培养条件下能从头合成雌二醇及表达芳香化酶。
Objective To study whether female rat bone marrow stem cells (BMSCs) can differentiate into estradiol (E2) and aromatase (P450arom) cells under high glucose DMEM containing 10% fetal bovine serum Ovarian premature aging, osteoporosis in the elderly and other diseases provide new possible ways. Methods Adult female Sprague-Dawley rat bone marrow stem cells were isolated and cultured in vitro with high glucose DMEM containing 10% fetal bovine serum. The cells were induced to differentiate with or without all-trans retinoic acid (ATRA) The concentrations of E2 in the two groups of cells were detected by radioimmunoassay (RIA) in the control group (without ATRA) and ATRA group (ATRA). The expression of CYP19 mRNA and FSHR mRNA expression of the two groups of cells detected by immunocytochemistry aromatase expression. The 3rd generation female rat bone marrow stem cells were cultured in vitro. RT-PCR showed that both CYP19 mRNA and FSHR mRNA were expressed in the control group and ATRA group from day 1 of culture. Immunocytochemistry showed that both groups were cultured from day 2 to day 4 Positive cells with P450arom expression, from 24 hours after culture in both cell cultures detected a high level of E2. Results The 3rd generation female rat bone marrow stem cells were cultured in vitro. Both CYP19 mRNA and FSHR mRNA were expressed in RT-PCR control group and ATRA group from the 1st day of culture. Immunocytochemistry showed that the two groups of cells from day 2 to day 4 Both of them had positive expression of P450arom, and higher levels of E2 were detected in both cell cultures from 24 h after culture. Conclusion Female rat bone marrow stem cells can de novo estradiol and express aromatase under in vitro culture conditions.