Bio- chemical and physical characterizations of mesenchymal stromal cells along the time course of directed differentiation.

Yin-Quan Chen,Yi-Chang Wu,Oscar K Lee,Yi-Shiuan Liu,Juan C Del Álamo,Yu-An Liu,Arthur Chiou

doi:10.1038/srep31547

Abstract

Cellular biophysical properties are novel biomarkers of cell phenotypes which may reflect the status of differentiating stem cells. Accurate characterizations of cellular biophysical properties, in conjunction with the corresponding biochemical properties could help to distinguish stem cells from primary cells, cancer cells, and differentiated cells. However, the correlated evolution of these properties in the course of directed stem cells differentiation has not been well characterized. In this study, we applied video particle tracking microrheology (VPTM) to measure intracellular viscoelasticity of differentiating human mesenchymal stromal/stem cells (hMSCs). Our results showed that osteogenesis not only increased both elastic and viscous moduli, but also converted the intracellular viscoelasticity of differentiating hMSCs from viscous-like to elastic-like. In contrast, adipogenesis decreased both elastic and viscous moduli while hMSCs remained viscous-like during the differentiation. In conjunction with bio- chemical and physical parameters, such as gene expression profiles, cell morphology, and cytoskeleton arrangement, we demonstrated that VPTM is a unique approach to quantify, with high data throughput, the maturation level of differentiating hMSCs and to anticipate their fate decisions. This approach is well suited for time-lapsed study of the mechanobiology of differentiating stem cells especially in three dimensional physico-chemical biomimetic environments including porous scaffolds.

Highlights

Optical tweezers[13,17], and video particle tracking microrheology (VPTM)[18]
The results of gene expressions demonstrated that differentiations by chemical inductions were incomplete on day 7 but gradually reached the maturations on day 28, which is consistent with our previous studies of human MSCs (hMSCs) cultured on Petri dishes[1]
We show that the alterations of viscoelastic properties of hMSCs are drastically different in osteogenic and adipogenic differentiation

Summary

Introduction

Optical tweezers[13,17], and video particle tracking microrheology (VPTM)[18]. AFM systems equipped with a sharp tip[19] have been shown to probe local cell stiffness caused by the interaction between cortex actin and cell membrane, whereas those equipped with colloidal force probe[20,21] have been demonstrated to analyze global cell stiffness. VPTM has two key merits compared to other techniques for measuring mechanical properties of living cells such as AFM, micropipette aspiration or optical tweezers It can be used in living cells embedded in 3-dimensional extracellular matrix (3-D ECM) as long as the probing particles are injected in the cells prior to 3D culture. We systematically measured biophysical parameters, including cell morphology, size of focal adhesion complex, actin arrangement, and intracellular viscoelasticity, during osteogenic and adipogenic differentiations of human MSCs (hMSCs) up to 28 days. We complemented these parameters with biochemical parameters along the time course of differentiation, including expression of differentiation genes, cytoskeleton related genes, and focal adhesion related genes. There is a strong negative correlation (PCC =−0.94) between |G*| and the gene expression of CCAAT-enhancer-binding proteins (C/EBP) during adipogenic differentiation

Methods

Results

Conclusion