Background and aim of study: The use of a biological, biodegradable scaffold remodeled by cells to resemble a valve leaflet is an attractive approach to tissue engineering. The study aim was to evaluate the suitability of a three-dimensional biodegradable collagen sponge for maintenance of cell viability, proliferation and phenotype of cultured human cardiac valve interstitial cells (ICs).
Methods: Pieces of valve leaflets were snap-frozen, sectioned and stained by immunoperoxidase. Interstitial cells were cultured from cardiac valves and plated onto glass coverslips or seeded in collagen sponge, then stained by immunofluorescence or immunoperoxidase. A panel of antibodies was used to determine cell phenotype. Cell viability was assessed using a dye-based cell proliferation assay, and cell death by lactate dehydrogenase measurement.
Results: ICs variably expressing the phenotypic markers were found throughout the native valve leaflet, but particularly on the ventricular side. Cultured ICs either on coverslips or in collagen sponge expressed vimentin, a fibroblast surface antigen and variable amounts of smooth muscle (SM) alpha-actin. Expression of the other phenotypic markers, SM myosin, desmin and prolyl 4-hydroxylase differed: interestingly, the ratio of cells in collagen sponge expressing these markers reflected that found in the native valve leaflet. Confocal microscopy of ICs in the collagen sponge revealed the presence of cells with long interconnecting extensions indicating cell communication. Cell proliferation and cell death assays established that cells were not only viable after four weeks in the sponge, but were also proliferating.
Conclusion: This study demonstrates that collagen sponge is a suitable biodegradable scaffold that can maintain viable valve ICs and appears to enhance the capacity of the cell to express its original phenotype.