Stem Cell Cytoskeletal Responses to Pulsatile Flow in Heart Valve Tissue Engineering Studies

Abstract

Heart valve replacement options remain exceedingly limited for pediatric patients because they cannot accommodate somatic growth. To overcome this shortcoming, heart valve tissue engineering using human bone marrow stem cells (HBMSCs) has been considered a potential solution to the treatment of critical congenital valvular defects. The mechanical environments during in vitro culture are key regulators of progenitor cell fate. Here, we report on alterations in HBMSCs, specifically in their actin cytoskeleton and their nucleus under fluid-induced shear stresses of relevance to heart valves. HBMSCs were seeded in microfluidic channels and were exposed to the following conditions: pulsatile shear stress (PSS), steady shear stress (SS), and no flow controls (n = 4/group). Changes to the actin filament structure were monitored and subsequent gene expression was evaluated. A significant increase (p < 0.05) in the number of actin filaments, filament density and angle (between 30° and 84°), and conversely a significant decrease (p < 0.05) in the length of the filaments were observed when the HBMSCs were exposed to PSS for 48 h compared to SS and no flow conditions. No significant differences in nuclear shape were observed among the groups (p > 0.05). Of particular relevance to valvulogenesis, klf2a, a critical gene in valve development, was significantly expressed only by the PSS group (p < 0.05). We conclude that HBMSCs respond to PSS by alterations to their actin filament structure that are distinct from SS and no flow conditions. These changes coupled with the subsequent gene expression findings suggest that at the cellular level, the immediate effect of PSS is to initiate a unique set of quantifiable cytoskeletal events (increased actin filament number, density and angle, but decrease in filament length) in stem cells, which could be useful in the fine-tuning of in vitro protocols in heart valve tissue engineering.

Physiologically Relevant Fluid-Induced Oscillatory Shear Stress Stimulation of Mesenchymal Stem Cells Enhances the Engineered Valve Matrix Phenotype

“…The combination of cyclic flexure and steady flow (flex-flow) induces pulsatile and/or oscillatory flow patterns on the surfaces of TEVCs. Our work, as well as others, have shown the importance of oscillatory flow conditions on developing valve tissues (6,(8)(9)(10). However, a physiologically relevant pulsatile flow waveform may be able to induce oscillations similar to the conditions experienced in the native valve, which will mechanically condition these TEVCs.…”

The Shape and Function of Solid Fascias Depend on the Presence of Liquid Fascias

“…The pressure of the fluids inside the kidneys influences the podocytes, involving cyclooxygenase-2 and prostaglandins type 2 in the mechanical response and other molecules (Akt-GSK3β-β-catenin) for correct adaptation (and shape) and kidney function [31]. The constant passage of fluids maintains the shape and function of the myocardium, stimulating the potassium channels (for correct electrical function), stimulating correct repair (through the intervention of stem cells) [32][33][34]. According to a model to understand the importance of mechanotransduction in the presence of fluids, the fluid mosaic model, highlights that the cell does not behave morphologically in unison during the passage of fluidic-mechanical information but reminds a lot of the fractal concept [35 ].…”

Oscillatory fluid-induced mechanobiology in heart valves with parallels to the vasculature

“…In addition to flex-flow conditions, our laboratory has also investigated the effects of pulsatile shear stress in comparison to non-pulsatile shear on tissue cell alignment. It is observed that bone marrow stem cells respond to pulsatile shear stress with an increased level of actin filaments and a decrease in filament length, suggesting transition from a mesenchymal to endothelial phenotype, which is a crucial process in the formation of the endothelium in tissue engineered heart valves ( 118 ).…”