Vascular grafts are used to repair, replace, or bypass diseased arteries, and there is a growing need for tissue-engineered blood vessels (TEBVs) as replacement grafts. Three-dimensional, self-assembled smooth muscle cell (SMC) rings can be fabricated and fused to create SMC tissue tubes with a structure similar to native vessels; however, this approach is limited by the underdeveloped mechanical integrity of the tissue. Thus, the goal of this research is to design, manufacture, and validate a cyclic circumferential stretch bioreactor to mechanically stimulate SMC tissue rings, with the goal of developing rings that can withstand the physiological forces of the in vivo environment. The bioreactor consists of a closed cam-syringe-tubing system that forces fluid into the tubing with each rotation of the cam, thereby distending and relaxing the tubing. Various sized cams were implemented to modify the distension of the tubing (5%, 7.5%, 10%, and 15% stretch magnitudes). Tissue rings are placed on the tubing, which is housed in a custom culture chamber. The tubing was validated using DVT® imaging technology to distend approximately 5, 7.5, 10, and 15% under static conditions. High density mapping was used to analyze the dynamic distension of the tubing and tissue rings. During bioreactor operation, the tubing distends 1-2% less than expected for the fabricated cams (5, 7.5, 10, 15%), and the tissue ring distends 31-56% less than the tubing on which it is located. To assess the effects of cyclic distension, 7-day-old SMC rings were cultured dynamically for 7 days and exposed to 0%, 5%, 7.5%, 10%, or 15% cyclic stretch (1 Hz, 100% duty cycle). Histology and immunohistochemistry indicate that both stretched and non-stretched rings synthesized collagen and glycosaminoglycans, but the contractile proteins á-smooth muscle actin and calponin were not synthesized. A decrease in cell density was observed as the magnitude of stretch increased, and the 5-15% stretched samples demonstrated more cellular alignment than the 0% stretch control samples. Mechanical testing analysis concluded that the stretched rings exhibited a reduction in ultimate tensile strength, maximum tangent modulus, maximum strain, and maximum load compared to unstretched control samples. It is anticipated that future work, including modifications of the culture medium and mechanical stimulation parameters (eg. reduced duty cycle, reduced frequency), has the potential to achieve the expected outcome of this research - a strong, aligned, contractile vascular smooth muscle cell tissue ring through dynamic culture using a cyclic circumferential stretch bioreactor.

Congenital heart defects are the number one cause of birth defect-related deaths. Cardiovascular diseases are the most common cause of death worldwide. Layered cellular sheet constructs offer one very valuable option for cardiac patch implantation during surgical treatment of both pediatric and adult patients with cardiac defects or damage. A very exciting, relatively unexplored, autologous, available cell source for making patches are placenta-derived mesenchymal stem cells (pMSCs). In this study, pMSCs were assessed as a potential cell source for cardiac repair and regeneration by evaluating their differentiation capacity into cardiomyocytes, their effects on cardiac cell migration and proliferation, and their ability to be grown into cell sheets. It was found that pMSC cardiac protein content was enhanced by differentiation media treatment, but no beating cells were produced. Undifferentiated pMSCs improved migration and proliferation of a cardiac cell population and formed intact, aligned cell sheets. However, like many new cell sources for cardiac repair, pMSCs should still be functionally characterized to understand how compatible they will be with resident heart tissue. Implanting non-autologous, potentially pluripotent, non-myocyte (non-beating) cells presents concerns regarding electromechanical mismatch and implant rejection. The characterization of non-traditional cell sources such as pMSCs motivated the design of a bio-MEMS device that assesses contractile force and conduction velocity in response to electrical and mechanical stimulation of a cell source as it is grown and once it forms a cellular sheet. This ideally creates the ability for patient specific cell sheets to be cultured, characterized, and conditioned to be compatible with the patient’s cardiac environment in vitro, prior to implantation. In this work, the device was designed to achieve the following: cellular alignment, electrical stimulation, mechanical stimulation, conduction velocity readout, contraction force readout, and upon characterization, cell sheet release. The platform is based on a set of comb electrical contacts which are three dimensional wall contacts made of polydimethylsiloxane and coated with electrically conductive metals. Device fabrication and initial validation experiments were completed as part of this study; ultimately the device will allow for the complete functional characterization and conditioning of variable cell source cell sheet implants for myocardial implantation. / 2019-07-02T00:00:00Z

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