Dual role of electrical stimulation and a biomimetic matrix in neural differentiation within a microfluidic platform†

Abstract

Neurodegenerative diseases mostly stem from oxidative stress and/or misfolded proteins in the central and peripheral nervous systems, posing clinical and economic burdens globally. Despite the advances in this field, biomimetic models recapitulating the neural microphysiological environment of both patients and healthy individuals are needed to accelerate drug development. Herein, a biomimetic microfluidic platform was developed to promote neural differentiation of stem cells by recapitulating physicochemical and physicomechanical factors in the neural microenvironment. In order to address this, the supportive role of electrical stimulation (ES) was assessed under various conditions by using immunofluorescence staining of mesenchymal stromal cell markers (CD45, CD90), the neuroepithelial stem cell protein marker (Nestin) and the microtubule-associated protein 2 marker (MAP2). Moreover, the combinational effect of ES and a cell-derived matrix (CDM), or a three-dimensional tissue-derived matrix (TDM), was explored. The matrices were obtained and characterized by scanning electron microscopy, contact angle analysis, DNA analysis, agarose gel electrophoresis, and in terms of extracellular matrix proteins. Neural differentiation was further validated by analysis of changes in gene expressions. ES applied in a rectangular manner with a 10 ms frequency at an intensity of 200 mV cm⁻¹ for 1 h per day for 7 days, followed by an additional 7 day recovery phase, revealed optimum neural differentiation for the combinational approach with brain TDM in both 2D and 3D. In conclusion, this work highlights the critical role of both physicochemical and physicomechanical factors in neural differentiation, offering valuable insights for advancing biomimetic models and stem cell research.