Department of Bioengineering, University of California, Berkeley, B108A Stanley Hall, Berkeley, CA 94720-1762, USA.
Biomaterials. 2011 Jun;32(16):3921-30. doi: 10.1016/j.biomaterials.2011.02.019.
Bone marrow mesenchymal stem cells (MSCs) are a valuable cell source for tissue engineering and regenerative medicine. Transforming growth factor β (TGF-β) can promote MSC differentiation into either smooth muscle cells (SMCs) or chondrogenic cells. Here we showed that the stiffness of cell adhesion substrates modulated these differential effects. MSCs on soft substrates had less spreading, fewer stress fibers and lower proliferation rate than MSCs on stiff substrates. MSCs on stiff substrates had higher expression of SMC markers α-actin and calponin-1; in contrast, MSCs on soft substrates had a higher expression of chondrogenic marker collagen-II and adipogenic marker lipoprotein lipase (LPL). TGF-β increased SMC marker expression on stiff substrates. However, TGF-β increased chondrogenic marker expression and suppressed adipogenic marker expression on soft substrates, while adipogenic medium and soft substrates induced adipogenic differentiation effectively. Rho GTPase was involved in the expression of all aforementioned lineage markers, but did not account for the differential effects of substrate stiffness. In addition, soft substrates did not significantly affect Rho activity, but inhibited Rho-induced stress fiber formation and α-actin assembly. Further analysis showed that MSCs on soft substrates had weaker cell adhesion, and that the suppression of cell adhesion strength mimicked the effects of soft substrates on the lineage marker expression. These results provide insights of how substrate stiffness differentially regulates stem cell differentiation, and have significant implications for the design of biomaterials with appropriate mechanical property for tissue regeneration.
骨髓间充质干细胞（MSCs）是组织工程和再生医学中非常有价值的细胞来源。转化生长因子β（TGF-β）可以促进 MSC 分化为平滑肌细胞（SMCs）或软骨细胞。在这里，我们发现细胞黏附底物的刚度调节了这些差异效应。与硬基底上的 MSC 相比，软基底上的 MSC 铺展面积更小，应力纤维更少，增殖速度更慢。硬基底上的 MSC 表达更高的 SMC 标志物 α-肌动蛋白和钙调蛋白-1；相比之下，软基底上的 MSC 表达更高的软骨标志物胶原蛋白-II 和脂肪生成标志物脂蛋白脂肪酶（LPL）。TGF-β增加了硬基底上的 SMC 标志物表达。然而，TGF-β增加了软基底上的软骨标志物表达并抑制了脂肪生成标志物表达，而脂肪生成培养基和软基底有效地诱导了脂肪生成分化。Rho GTPase 参与了所有上述谱系标志物的表达，但不能解释底物刚度的差异效应。此外，软基底并没有显著影响 Rho 活性，但抑制了 Rho 诱导的应力纤维形成和α-肌动蛋白组装。进一步分析表明，软基底上的 MSC 黏附力较弱，细胞黏附力的抑制作用模拟了软基底对谱系标志物表达的影响。这些结果提供了关于底物刚度如何差异调节干细胞分化的深入了解，对设计具有适当机械性能的生物材料以促进组织再生具有重要意义。
