Differentiation and Distribution of Marrow Stem Cells in Flex-Flow Environments Demonstrate Support of the Valvular Phenotype

Abstract

For treatment of critical heart valve diseases, prosthetic valves perform fairly well in most adults; however, for pediatric patients, there is the added requirement that the replacement valve grows with the child, thus extremely limiting current treatment options. Tissue engineered heart valves (TEHV), such as those derived from autologous bone marrow stem cells (BMSCs), have the potential to recapitulate native valve architecture and accommodate somatic growth. However, a fundamental pre-cursor in promoting directed integration with native tissues rather than random, uncontrolled growth requires an understanding of BMSC mechanobiological responses to valve-relevant mechanical environments. Here, we report on the responses of human BMSC-seeded polymer constructs to the valve-relevant stress states of: (i) steady flow alone, (ii) cyclic flexure alone, and (iii) the combination of cyclic flexure and steady flow (flex-flow). BMSCs were seeded onto a PGA: PLLA polymer scaffold and cultured in static culture for 8 days. Subsequently, the aforementioned mechanical conditions, (groups consisting of steady flow alone—850ml/min, cyclic flexure alone—1 Hz, and flex-flow—850ml/min and 1 Hz) were applied for an additional two weeks. We found samples from the flex-flow group exhibited a valve-like distribution of cells that expressed endothelial (preference to the surfaces) and myofibroblast (preference to the intermediate region) phenotypes. We interpret that this was likely due to the presence of both appreciable fluid-induced shear stress magnitudes and oscillatory shear stresses, which were concomitantly imparted onto the samples. These results indicate that flex-flow mechanical environments support directed in vitro differentiation of BMSCs uniquely towards a heart valve phenotype, as evident by cellular distribution and expression of specific gene markers. A priori guidance of BMSC-derived, engineered tissue growth under flex-flow conditions may serve to subsequently promote controlled, engineered to native tissue integration processes in vivo necessary for successful long-term valve remodeling.

Identifying Behavioral Phenotypes and Heterogeneity in Heart Valve Surface Endothelium

“…Valve cells are particularly challenging in this regard, since harvesting cells from a patient is considered to be extremely invasive if not impossible. Therefore, a suitable VEC facsimile would need to be derived from a different source, such as bone marrow stem cells (Rath et al 2015), peripheral blood endothelial progenitor cells (EPCs), umbilical cord blood EPCs, or possibly induced pluripotent stem cells (IPSCs).…”

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

“…Mechanical heart valves require patients to be on life-long anticoagulant medication; bioprosthetic valves are less durable and more prone to re-calcification. Scientists and engineers have been exploring alternative treatments such as tissue engineered heart valves using bioreactor systems to deliver the proper flow, flex, and stretch conditioning (116,117).…”

“…Some of the major in-house findings of tissue engineered heart valves show that when bone marrow stems cells seeded in scaffolds are exposed to a combination of flex and flow environments, the shear stresses from both flex and flow conditions generated a significantly higher amount of collagen compared flex only or flow only samples. In addition, under static (no flow) conditions, very little collagen is produced (116). This suggests that physiologically relevant flex-flow conditions promote early valvular tissue matrix development.…”

Developing a Clinically Relevant Tissue Engineered Heart Valve—A Review of Current Approaches

“…Since then, PGA, PGA/PLA hybrids, and their copolymer PLGA have been used in a variety of studies regarding the fabrication of functional, bioresorbable leaflet scaffolds . More recent studies have utilized these scaffolds to observe cellular behavior and mechanical properties . One group studied bone marrow stem cells seeded on PGA:PLLA scaffolds under different stress conditions, such as steady flow, cyclic flexure, and a combination of the two conditions, and found that the cells exhibited endothelial and myofibroblast phenotypes similar to valves under a combination of steady flow and cyclic flexure .…”

“…More recent studies have utilized these scaffolds to observe cellular behavior and mechanical properties . One group studied bone marrow stem cells seeded on PGA:PLLA scaffolds under different stress conditions, such as steady flow, cyclic flexure, and a combination of the two conditions, and found that the cells exhibited endothelial and myofibroblast phenotypes similar to valves under a combination of steady flow and cyclic flexure . Another group found that prolonged tissue culture reduced retraction of the tissue engineered leaflets that typically lose mechanical integrity due to rapid degradation of the PGA scaffold, and suggested that this resulted from an increased amount of GAGs in the tissue …”