Department of Anatomical sciences, Medical Sciences Faculty, Tarbiat Modares University, Jalale-Ale-Ahmad highway, P.O. Box 14115-175, Tehran, Iran.
In Vitro Cell Dev Biol Anim. 2011 Sep;47(8):521-5. doi: 10.1007/s11626-011-9429-0. Epub 2011 Jun 30.
Bone morphogenetic protein (BMP)-4 has a crucial role on primordial germ cells (PGCs) development in vivo which can promote stem cell differentiation to PG-like cells. In this study, we investigated the expression of Mvh as one of the specific genes in primordial germ cells after treatment with different doses of BMP4 on bone mesenchymal stem cells (BMSCs)-derived PGCs. Following isolation of BMSCs from male mouse femur and tibia, cells were cultured in medium for 72 h. Passage 4 murine BMSCs were characterized by CD90, CD105, CD34, and CD45 markers and osteo-adipogenic differentiation. Different doses of BMP4 (0, 0.01, 0.1, 1, 5, 25, 50, and 100 ng/ml) were added to BMSCs for PGCs differentiation during 4-days culture. Viability percent, proliferation rates, and expression of Mvh gene were analyzed by RT-qPCR. Data analysis was done with ANOVA test. CD90(+), CD105(+), CD34(-), and CD45(-) BMSCs were able to differentiate to osteo-adipogenic lineages. The results revealed that proliferation rate and viability percent were raised significantly (p ≤ 0.05) by adding 1, 5, 25 ng/ml of BMP4 and there were decreased to the lowest rate after adding 100 ng/ml BMP4 (p ≤ 0.05). There were significant up regulation (p ≤ 0.05) in Mvh expression between 25, 50, and 100 ng/ml BMP4 with other doses. So the selective dose of BMP-4 for treatment during 4-day culture was 25 ng/ml. The results suggest that addition of 25 ng/ml BMP4 had the best effects based on gene-specific marker expression.
骨形态发生蛋白 4（BMP-4）在体内原始生殖细胞（PGC）发育中起着关键作用，它可以促进干细胞向 PG 样细胞分化。在这项研究中，我们研究了不同剂量 BMP4 处理后，骨间充质干细胞（BMSC）来源的 PGC 中 Mvh 作为特定基因之一的表达。从小鼠股骨和胫骨中分离出 BMSC 后，将细胞在培养基中培养 72 小时。第 4 代鼠 BMSC 通过 CD90、CD105、CD34 和 CD45 标志物和成骨脂肪分化进行鉴定。在 4 天的培养过程中，向 BMSC 中添加不同剂量的 BMP4（0、0.01、0.1、1、5、25、50 和 100ng/ml）以诱导 PGC 分化。通过 RT-qPCR 分析细胞活力百分比、增殖率和 Mvh 基因的表达。数据采用方差分析进行分析。CD90(+)、CD105(+)、CD34(-)和 CD45(-)BMSC 能够分化为成骨脂肪谱系。结果表明，添加 1、5、25ng/ml 的 BMP4 可显著提高增殖率和活力百分比（p≤0.05），而添加 100ng/ml 的 BMP4 则降至最低（p≤0.05）。在 25、50 和 100ng/ml BMP4 之间，Mvh 表达显著上调（p≤0.05），而其他剂量组则下调。因此，在 4 天培养过程中，BMP-4 的治疗选择剂量为 25ng/ml。结果表明，基于基因特异性标志物的表达，添加 25ng/ml 的 BMP4 效果最佳。
