Central of Clinical Laboratory, Shimane University Hospital, Izumo, Japan.
Eur J Haematol. 2010 Jun;84(6):538-46. doi: 10.1111/j.1600-0609.2010.01419.x. Epub 2010 Jan 5.
Effective ex vivo expansion of hematopoietic stem cells (HSCs) is a prerequisite for HSC transplantation. Growth and maintenance of HSC is dependent on cytokine and niche factors. We investigated whether mesenchymal stem cells (MSCs) or osteogenic cytokine-differentiated MSCs play a role in HSC expansion. We used the human HM3.B10 (B10) MSC cell line and the osteoblast-differentiated B10 (Ost-B10) as a feeder layer and examined ex vivo expansion of CD34(+)CD38(-) HSCs obtained from peripheral blood (PB) and cord blood (CB) with or without several growth cytokines. Both undifferentiated B10 and Ost-B10 cells exhibited similar effects on total HSC expansion; however, Ost-B10 demonstrated a higher potency in CD34(+)CD38(-) cell-specific proliferation in the presence of cytokines compared to undifferentiated B10 HSCs. Colony-forming cell assay and long-term culture initiating cell assay revealed that Ost-B10 displayed multipotent differentiation and enabled long-term ex vivo culture of HSCs. We next examined the relationship between HSC expansion and the presence of various chemokines. CXCL4 and CXCL12 expression were increased in Ost-B10 cells compared with the B10 cells. CD34(+)CD38(-) cells were significantly increased with CXCL12, but not CXCL4 treatment. siRNA inhibition of CXCL12 decreased CXCL12 secretion in both B10 and Ost-B10, whereas expansion of CD34(+)CD38(-) cells was decreased in Ost-B10 alone. These results demonstrated that ex vivo expansion of HSCs may be highly effective through osteoblast-differentiated MSCs acting as a feeder layer, and likely operates through the CXCL12 chemokines signaling pathway.
有效的造血干细胞（HSCs）体外扩增是 HSC 移植的前提。HSC 的生长和维持依赖于细胞因子和龛位因子。我们研究了间充质干细胞（MSCs）或成骨细胞分化的 MSC 是否在 HSC 扩增中发挥作用。我们使用人 HM3.B10（B10）MSC 细胞系和成骨细胞分化的 B10（Ost-B10）作为饲养层，并检查了来自外周血（PB）和脐带血（CB）的 CD34+CD38-HSCs 的体外扩增，有无几种生长细胞因子。未分化的 B10 和 Ost-B10 细胞均对总 HSC 扩增表现出相似的作用；然而，与未分化的 B10 HSCs 相比，Ost-B10 在存在细胞因子的情况下对 CD34+CD38-细胞的特异性增殖具有更高的效力。集落形成细胞测定和长期培养起始细胞测定显示，Ost-B10 具有多能分化能力，并能够实现 HSCs 的长期体外培养。我们接下来研究了 HSC 扩增与各种趋化因子之间的关系。与 B10 细胞相比，Ost-B10 细胞中 CXCL4 和 CXCL12 的表达增加。CXCL12 处理可显著增加 CD34+CD38-细胞，但 CXCL4 处理则不然。CXCL12 的 siRNA 抑制降低了 B10 和 Ost-B10 中 CXCL12 的分泌，而仅在 Ost-B10 中 CD34+CD38-细胞的扩增减少。这些结果表明，通过成骨细胞分化的 MSC 作为饲养层，HSC 的体外扩增可能非常有效，并且可能通过 CXCL12 趋化因子信号通路起作用。
