Adult Human Mesenchymal Stem Cell Differentiation at the Cell Population and Single-Cell Levels Under Alternating Electric Current

作者：Wechsler Marissa E; Hermann Brian P; Bizios Rena^*

来源：Tissue Engineering - Part C: Methods , 2016, 22(2): 155-164.

DOI：10.1089/ten.tec.2015.0324

Mesenchymal stem cells, precursors that can differentiate into osteoblasts, chondrocytes, and adipocytes, have tremendous potential for derivation of cells with specific (e.g., osteogenic) phenotypes for tissue engineering and tissue regeneration applications. To date, the predominant strategy to achieve directed differentiation of MSCs into osteoblasts was to recapitulate the normal developmental ontogeny of osteoblasts using growth factors (e.g., bone morphogenetic proteins). In contrast, the effects of biophysical stimuli alone on such outcomes remain, at best, partially understood. This in vitro study examined and optimized the effects of alternating electric current alone on the differentiation of adult human mesenchymal stem cells (hMSCs) at the cell population and single-cell levels. hMSCs, cultured on flat, indium-tin-oxide-coated glass in the absence of supplemented exogenous growth factors were exposed to alternating electric current (5-40 mu A, 5-10 Hz frequency, sinusoidal waveform), for 1-24 h daily for up to 21 consecutive days. Compared to results obtained from the respective controls, hMSC populations exposed to the alternating electric current alone (in the absence of exogenous growth factors) expressed genes at various stages of differentiation (specifically, TAZ, Runx-2, Osterix, Osteopontin, and Osteocalcin). Optimal osteogenic differentiation was achieved when hMSCs were exposed to a 10 mA, 10 Hz alternating electric current for 6 h daily for up to 21 days. Exclusive osteodifferentiation was observed since genes for the chondrocyte (Collagen Type II) and adipocyte (FABP-4) lineages were not expressed under all conditions of the biophysical stimulus tested. Single cell mRNAs for 45 genes (indicative of hMSC differentiation) were monitored using Fluidigm Systems. Homogeneous expression of the early osteodifferentiation genes (specifically, TAZ and Runx-2) was observed in hMSCs exposed to the alternating electric current at 7 and 21 days. Heterogeneity for all other genes monitored was observed in hMSCs exposed to alternating electric current and in their respective controls. These results provide the first glimpse of gene expression in differentiating hMSCs at the cell population and single-cell levels and represent novel approaches for stem cell differentiation pertinent to new tissue formation.

• 出版日期2016-2