Abstract
BackgroundAdult mesenchymal stem cells (MSCs) are an important resource for tissue growth, repair, and regeneration. To utilize MSCs more effectively, a clear understanding of how they react to environmental cues is essential. Currently, relatively little is known about how the composition of the plasma membranes affects stem cell phenotype and properties. The presence of lipid molecules, including cholesterol in particular, in the plasma membrane plays a crucial role in regulating a variety of physiological processes in cells. In this study, we examined the effects of perturbations in cholesterol/caveolin-1 (CAV-1)/caveolae homeostasis on the membrane properties and adhesive characteristics of MSCs. Findings from this study will contribute to the understanding of how cholesterol/CAV-1/caveolae regulates aspects of the cell membrane important to cell adhesion, substrate sensing, and microenvironment interaction.MethodsWe generated five experimental MSC groups: 1) untreated MSCs; 2) cholesterol-depleted MSCs; 3) cholesterol-supplemented MSCs; 4) MSCs transfected with control, nonspecific small interfering (si)RNA; and 5) MSCs transfected with CAV-1 siRNA. Each cell group was analyzed for perturbation of cholesterol status and CAV-1 expression by performing Amplex Red cholesterol assay, filipin fluorescence staining, and real-time polymerase chain reaction (PCR). The membrane fluidity in the five experimental cell groups were measured using pyrene fluorescence probe staining followed by FACS analysis. Cell adhesion to collagen and fibronectin as well as cell surface integrin expression were examined.ResultsCholesterol supplementation to MSCs increased membrane cholesterol, and resulted in decreased membrane fluidity and localization of elevated numbers of caveolae and CAV-1 to the cell membrane. These cells showed increased expression of α1, α4, and β1 integrins, and exhibited higher adhesion rates to fibronectin and collagen. Conversely, knockdown of CAV-1 expression or cholesterol depletion on MSCs caused a parallel decrease in caveolae content and an increase in membrane fluidity due to decreased delivery of cholesterol to the cell membrane. Cells with depleted CAV-1 expression showed decreased cell surface integrin expression and slower adhesion to different substrates.ConclusionsOur results demonstrate that perturbations in cholesterol/CAV-1 levels significantly affect the membrane properties of MSCs. These findings suggest that modification of membrane cholesterol and/or CAV-1 and caveolae may be used to manipulate the biological activities of MSCs.
Highlights
Adult mesenchymal stem cells (MSCs) are an important resource for tissue growth, repair, and regeneration
Our results demonstrate that perturbations in cholesterol/CAV-1 levels significantly affect the membrane properties of MSCs
These findings suggest that modification of membrane cholesterol and/or CAV-1 and caveolae may be used to manipulate the biological activities of MSCs
Summary
Adult mesenchymal stem cells (MSCs) are an important resource for tissue growth, repair, and regeneration. A landmark discovery in stem cell-environment interactions was made by Engler et al [6] who reported that the stiffness of two-dimensional (2D) adhesion substrates can determine the differentiation of MSCs in vitro. These investigators showed that MSCs differentiate into neurogenic, myogenic, and osteogenic phenotypes on substrates that have elasticity values similar to those of brain, muscle, and bone tissue, respectively [6]. This mechanosensory behavior is a potentially critical parameter in the application of MSCs for tissue engineering. While many studies have focused on derivation and molecular regulation of stem cells, relatively little is known about the composition of the stem cell membrane, the organization of which can critically affect cell responses to external stimuli
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
