{"id":3685,"date":"2011-07-11T14:37:24","date_gmt":"2011-07-11T14:37:24","guid":{"rendered":"https:\/\/mtlsites.mit.edu\/annual_reports\/2011\/?p=3685"},"modified":"2011-07-19T20:54:52","modified_gmt":"2011-07-19T20:54:52","slug":"microfluidic-perfusion-for-modulating-stem-cell-diffusible-signaling-2","status":"publish","type":"post","link":"https:\/\/mtlsites.mit.edu\/annual_reports\/2011\/microfluidic-perfusion-for-modulating-stem-cell-diffusible-signaling-2\/","title":{"rendered":"Microfluidic Perfusion for Modulating Stem Cell Diffusible Signaling"},"content":{"rendered":"

Stem cell phenotype and function are influenced by microenvironmental cues that include cell-cell, cell-extracellular matrix (ECM), and cell-media interactions, as well as mechanical forces. Our research focuses on developing microscale systems for controlling the cellular microenvironment of mouse embryonic stem cells (mESCs), in particular mechanical forces (i.e.,<\/em> shear stress) and cell-media interactions (i.e., diffusible signaling).<\/p>\n

Many emerging technologies used for ESC expansion or differentiation require perfusion culture, an example being pluripotent stem cell expansion in bioreactors for clinical applications [1<\/a>] <\/sup>. We employ a multiplex microfluidic perfusion array to study the effects of shear stress on mESCs across a wide range of flow rates in a graded, quantitative manner. Using this device, we are able to show that perfusion elicits phenotypic changes and that the specific shear-responsive phenotype is due to mechanosensing by heparan sulfate proteoglycans (HSPGs, Figure 1A-C). This is the first study describing the ESC machinery capable of responding to shear stress, thus providing a foundation for further shear mechanotransduction studies [2<\/a>] <\/sup>.<\/p>\n

Cells are constantly secreting and responding to soluble signals, the removal of which can be mediated by modulating flow properties at the microscale. To assess the contribution of cell-secreted factors to mESC differentiation and self-renewal, we utilized a two-layer microfluidic perfusion device allowing for parallel comparison of different cell culture conditions (Figure 2A) [3<\/a>] <\/sup>. Our results demonstrate that mESCs do not grow in differentiation conditions with minimal autocrine signaling, even with supplementation by Fgf4, a signal that has been shown to be a crucial factor in differentiation toward a neuronal stem cell fate (Figure 2B). Conversely, under self-renewal conditions, mESCs proliferate but lose self-renewal markers and upregulate differentiation markers (Figure 2C). These results, together with signaling and downstream differentiation assays, indicate that a differentiation towards an epiblast-like early differentiation state under conditions that had previously been shown as sufficient for self-renewal. Together, these results indicate the importance of cell-secreted signals for mESC growth and self-renewal.<\/p>\n\n\t\t