Neurogenesis-on-Chip: Electric field modulated transdifferentiation of human mesenchymal stem cell and mouse muscle precursor cell coculture

Abstract

A number of bioengineering strategies, using biophysical stimulation, are being explored to guide the human mesenchymal stem cells (hMScs) into different lineages. In this context, we have limited understanding on the transdifferentiation of matured cells to another functional-cell type, when grown with stem cells, in a constrained cellular microenvironment under biophysical stimulation. While addressing such aspects, the present work reports the influence of the electric field (EF) stimulation on the phenotypic and functionality modulation of the coculture of murine myoblasts (C2C12) with hMScs [hMSc:C2C12=1:10] in a custom designed polymethylmethacrylate (PMMA) based microfluidic device with in-built metal electrodes. The quantitative and qualitative analysis of the immunofluorescence study confirms that the cocultured cells in the conditioned medium with astrocytic feed, exhibit differentiation towards neural-committed cells under biophysical stimulation in the range of the endogenous physiological electric field strength (8 ± 0.06 mV/mm). The control experiments using similar culture protocols revealed that while C2C12 monoculture exhibited myotube-like fused structures, the hMScs exhibited the neurosphere-like clusters with SOX2, nestin, βIII-tubulin expression. The electrophysiological study indicates the significant role of intercellular calcium signalling among the differentiated cells towards transdifferentiation. Furthermore, the depolarization induced calcium influx strongly supports neural-like behaviour for the electric field stimulated cells in coculture. The intriguing results are explained in terms of the paracrine signalling among the transdifferentiated cells in the electric field stimulated cellular microenvironment. In summary, the present study establishes the potential for neurogenesis on-chip for the coculture of hMSc and C2C12 cells under tailored electric field stimulation, in vitro.

The secretome of human dental pulp stem cells protects myoblasts from hypoxia‑induced injury via the Wnt/β‑catenin pathway

“…The mouse cells mirror the biology of human cells well in various aspects (33). Previous studies (34)(35)(36) have also demonstrated that human cells and/or their cM have protective effects in mouse and rat cells. Therefore, c2c12 cells were used in the present study.…”

Electrical Stimulation Promotes Stem Cell Neural Differentiation in Tissue Engineering

“…Dong et al treat NSCs with ES for 3 days at 150 mV/mm, resulting in increased achaete-scute homolog (Ascl1) expression that was further proven to regulate phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway in NSCs [ 42 ]. In addition, hMSCs also showed increased levels of SOX2, nestin, β III-tubulin expression, and Ca 2+ oscillation after nine days of continuous exposure to 8 mV/mm ES for 20 h/day [ 43 ]. Taken together, these studies confirm the ES can induce cell orientation and migration, and enhance the differentiation of stem cells into neural cell lineages.…”

Anodic TiO2 Nanotubes: Tailoring Osteoinduction via Drug Delivery

“…Further, despite a long history of clinical trials, the mechanism linking electric stimuli and its sensing on the cell surface is still elusive. Therefore, during recent decades, the mechanism of EF (e.g., how cells sense and react to EFs) affecting cell adhesion, migration, proliferation, and differentiation has been extensively investigated in various primary cell/cell lines [ 206 , 207 , 208 , 209 , 210 , 211 ] including epithelial, mesenchymal, and neural cells [ 212 , 213 , 214 , 215 ]. Exposure of the cells to an electric current, if a certain current value threshold is exceeded, leads to cell death, while currents lower than 11 As/m 2 have shown no decrease in cell viability [ 216 ].…”