Collagen is a key element of basal lamina in physiological systems that participates in cell and tissue culture. Its function is for cell maintenance and growth, angiogenesis, disease progression, and immunology. The goal of our present study was to integrate a micrometer resolution membrane that is synthesized out of rat-tail type I collagen in a microfluidic device with apical and basolateral chambers. The collagen membrane was generated by lyophilization. In order to evaluate the compatibility of the resulting membrane with organs-on-chips technology, it was sandwiched between layers of polydimethylsiloxane (PDMS) that had been prepared by replica molding, and the device was used to culture human colon caco 2 cells on the top of the membrane. Membrane microstructure, transport, and cell viability in the organs-on-chips were observed to confirm the suitability of our resulting membrane. Through transport studies, we compared diffusion of two different membranes: Transwell and our resulting collagen membrane. We found that mass transport of 40 kDa dextran was an order of magnitude higher through the collagen membrane than that through the Transwell membrane. Human colon caco 2 cells were cultured in devices with no, Transwell, or ECM membrane to evaluate the compatibility of cells on the ECM membrane compared to the other two membranes. We found that caco 2 cells cultured on the collagen membrane had excellent viability and function for extended periods of time compared to the other two devices. Our results indicate a substantial improvement in establishing a physiological microenvironment for in vitro organs-on-chips.
An alginate/halloysite nanotube (HNT) nanocomposite was developed with sustained release of bone morphogenetic proteins (BMPs) at picogram low levels. BMP-2, 4, and 6 and osteoblasts were chosen as our model "growth factor" and "cell type" as the interaction of BMPs with osteoblasts is well known and thoroughly investigated. Alginate hydrogels with HNTs doped with BMP-2, 4, or 6 only or BMP-4 and 6 in combination. Osteoblasts were seeded within the hydrogels and studied for changes in cell proliferation, phenotypic expression, and mineralization over a 28-day experimental period. Osteoblast behavior was enhanced in BMP doped hydrogel/HNTs nanocomposites as compared with control groups. Release profiles showed that BMP-2 was released in a sustained fashion over a 7-day period and at picogram levels. Mineralization, as showed by Von Kossa staining, and protein synthesis peaked at 28 days, for all three growth factor combinations. BMP-4 provided a marked stimulus for osteoblast functionality base and was comparable to BMP-6 in terms of osteoblast differentiation and mineralization. BMP-4 and 6, in combination, showed a marked enhancement in osteoblast differentiation and functionality; however, the response seemed to be delayed when compared with BMP-4 and 6 release. Hydrogel surfaces had a complex surface topography and greater structural integrity with increased halloysite addition. The data suggest that these nanocomposites may provide a mechanism to enhance repair and regeneration in damaged or diseased tissues, reducing the need for more invasive treatment modalities.
Successful fracture healing requires the simultaneous regeneration of both the bone and vasculature; mesenchymal stem cells (MSCs) are directed to replace the bone tissue, while endothelial progenitor cells (EPCs) form the new vasculature that supplies blood to the fracture site. In the elderly, the healing process is slowed, partly due to decreased regenerative function of these stem and progenitor cells. MSCs from older individuals are impaired with regard to cell number, proliferative capacity, ability to migrate, and osteochondrogenic differentiation potential. The proliferation, migration and function of EPCs are also compromised with advanced age. Although the reasons for cellular dysfunction with age are complex and multidimensional, reduced expression of growth factors, accumulation of oxidative damage from reactive oxygen species, and altered signaling of the Sirtuin-1 pathway are contributing factors to aging at the cellular level of both MSCs and EPCs. Because of these geriatric-specific issues, effective treatment for fracture repair may require new therapeutic techniques to restore cellular function. Some suggested directions for potential treatments include cellular therapies, pharmacological agents, treatments targeting age-related molecular mechanisms, and physical therapeutics. Advanced age is the primary risk factor for a fracture, due to the low bone mass and inferior bone quality associated with aging; a better understanding of the dysfunctional behavior of the aging cell will provide a foundation for new treatments to decrease healing time and reduce the development of complications during the extended recovery from fracture healing in the elderly.
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