Interplay of Substrate Conductivity, Cellular Microenvironment, and Pulsatile Electrical Stimulation toward Osteogenesis of Human Mesenchymal Stem Cells in Vitro

Abstract

The influences of physical stimuli such as surface elasticity, topography, and chemistry over mesenchymal stem cell proliferation and differentiation are well investigated. In this context, a fundamentally different approach was adopted, and we have demonstrated the interplay of inherent substrate conductivity, defined chemical composition of cellular microenvironment, and intermittent delivery of electric pulses to drive mesenchymal stem cell differentiation toward osteogenesis. For this, conducting polyaniline (PANI) substrates were coated with collagen type 1 (Coll) alone or in association with sulfated hyaluronan (sHya) to form artificial extracellular matrix (aECM), which mimics the native microenvironment of bone tissue. Further, bone marrow derived human mesenchymal stem cells (hMSCs) were cultured on these moderately conductive (10(-4)-10(-3) S/cm) aECM coated PANI substrates and exposed intermittently to pulsed electric field (PEF) generated through transformer-like coupling (TLC) approach over 28 days. On the basis of critical analysis over an array of end points, it was inferred that Coll/sHya coated PANI (PANI/Coll/sHya) substrates had enhanced proliferative capacity of hMSCs up to 28 days in culture, even in the absence of PEF stimulation. On the contrary, the adopted PEF stimulation protocol (7 ms rectangular pulses, 3.6 mV/cm, 10 Hz) is shown to enhance osteogenic differentiation potential of hMSCs. Additionally, PEF stimulated hMSCs had also displayed different morphological characteristics as their nonstimulated counterparts. Concomitantly, earlier onset of ALP activity was also observed on PANI/Coll/sHya substrates and resulted in more calcium deposition. Moreover, real-time polymerase chain reaction results indicated higher mRNA levels of alkaline phosphatase and osteocalcin, whereas the expression of other osteogenic markers such as Runt-related transcription factor 2, Col1A, and osteopontin exhibited a dynamic pattern similar to control cells that are cultured in osteogenic medium. Taken together, our experimental results illustrate the interplay of multiple parameters such as substrate conductivity, electric field stimulation, and aECM coating on the modulation of hMSC proliferation and differentiation in vitro.

Time‐dependent effect of electrical stimulation on osteogenic differentiation of bone mesenchymal stromal cells cultured on conductive nanofibers

“…While it is still not quite clear how the ES would affect the differentiation if it is applied during different periods of the osteogenic differentiation. It is interesting to investigate this issue because varied parameters were applied in ES studies on bone regeneration from different studies …”

“…It is interesting to investigate this issue because varied parameters were applied in ES studies on bone regeneration from different studies. 31,32,49 To achieve the goal, conductive PLLA/MWCNT composite nanofibers were prepared for cell culture under ES in this study, and the contents of MWCNTs were changed from 0 wt % to 5 wt % to adjust the conductivity of resulting composite nanofibers. Most of the MWCNTs were aligned along the fiber direction that fiber morphology of PLLA/MWCNT composite nanofibers did not show much difference at different MWCNTs loading contents.…”

“…39,43,50 There was a big difference in cell tolerance when ES was conducted to cells, even cell types were the same but from different individuals. 32,49,51 For the present study, the voltage (1.5 V) and the duration (1.5 h per day) were optimized via pretests (Supporting Information Figs. S2 and S3).…”

Electrical stimulation in bone tissue engineering treatments

“…Previous in vitro experiments that exposed cells and/or scaffolds to EStim, demonstrated its ability to influence cell functions associated with enhanced bone healing [7][8][9][10][11][12][13][14]. These experiments were conducted on a variety of different cell types; bone marrow-derived mesenchymal stem cells (BM-MSC), from human and animal origin [10,11,[15][16][17][18][19][20][21][22][23][24][25]; adipose-derived mesenchymal stem cells (AT-MSC) [11,20,[26][27][28][29][30], mouse osteoblast-like cells [31][32][33] and more recently, human dental pulp stem cells (DPSC) [34]. The above cell types are commonly studied for use in BTE applications.…”

Synthesis of Conductive Carbon Aerogels Decorated with β-Tricalcium Phosphate Nanocrystallites

“…The phenomenon that the electrical current can have a detrimental effect on differentiation and proliferation capacity of the cells has triggered recent research interest towards quest for biocompatible and conductive substrates [1][2][3][4] . In particular, carbon-based composite materials have emerged as potentially promising targets for growth of electroactive tissues such as cardiac muscle, nerve and bone since the stimulation of osteogenesis due to inherent piezoelectric features of bone tissues attainable on conductive scaffolds [5][6][7][8][9] . Among these, blended composites between nanostructured carbon materials (carbon nanofibers/nanotubes and graphene) and biocompatible polymers (poly(vinyl alcohol), alginate, poly(lactide-co-glycolide and collagen) have been widely investigated as future materials for tissue engineering [10][11][12][13][14][15][16][17] .…”