Matrix compliance and RhoA direct the differentiation of mammary progenitor cells

The regenerative capacity of the mammary gland following post-lactational involution depends on the presence of multipotent stem or progenitor cells. Mammary progenitor cells exist as a quiescent and self-renewing population capable of differentiating into luminal epithelial and myoepithelial cells and generating ductal and alveolar structures. The fate choices of these cells are regulated by several soluble signals as well as their surrounding extracellular matrix. Whereas matrix stiffness has been implicated in organ-specific differentiation of embryonic and mesenchymal stem cells, the effects of substratum compliance on the more limited fate switches typical of tissue-specific progenitor cells are unknown. Here, we examined how the mechanical properties of the microenvironment affect the differentiation of mammary progenitor cells. Immortalized human mammary progenitor cells were cultured on synthetic hydrogels of varying stiffness, and their self-renewal and fate decisions were quantified. We found that cells cultured on soft substrata differentiated preferentially into luminal epithelial cells, whereas those cultured on stiff substrata differentiated preferentially into myoepithelial cells. Furthermore, pharmacological manipulations of cytoskeletal tension in conjunction with analysis of gene expression revealed that mechanical properties of the microenvironment signal through the small GTPase RhoA and cytoskeletal contractility to modulate the differentiation of mammary progenitor cells. These data suggest that subtle variations in the mechanical compliance of a tissue can direct the fate decisions of its resident progenitor cells.