Mechanical conditioning is often used to enhance collagen synthesis, remodeling and maturation and, hence, the structural and mechanical properties of engineered cardiovascular tissues. Intermittent straining, i.e., alternating periods of cyclic and static strain, has previously been shown to result in more mature tissue compared with continuous cyclic straining. Nevertheless, the underlying mechanism is unknown. We have determined the short-term effects of continuous cyclic strain and of cyclic strain followed by static strain at the gene expression level to improve insight into the mechano-regulatory mechanism of intermittent conditioning on collagen synthesis, remodeling and maturation. Tissue-engineered constructs, consisting of human vascular-derived cells seeded onto rapidly degrading PGA/P4HB scaffolds, were conditioned with 4% strain at 1 Hz for 3 h in order to study the immediate effects of cyclic strain (n=18). Next, the constructs were either subjected to ongoing cyclic strain (4% at 1 Hz; n=9) or to static strain (n=9). Expression levels of genes involved in collagen synthesis, remodeling and maturation were studied at various time points up to 24 h within each straining protocol. The results indicate that a period of static strain following cyclic strain favors collagen synthesis and remodeling, whereas ongoing cyclic strain shifts this balance toward collagen remodeling and maturation. The data suggest that, with prolonged culture, the conditioning protocol should be changed from intermittent straining to continuous cyclic straining to improve collagen maturation after its synthesis and, hence, the tissue (mechanical) properties.
Synthetic polymers are widely used to fabricate porous scaffolds for the regeneration of cardiovascular tissues. To ensure mechanical integrity, a balance between the rate of scaffold absorption and tissue formation is of high importance. A higher rate of tissue formation is expected in fast-degrading materials than in slow-degrading materials. This could be a result of synthetic cells, which aim to compensate for the fast loss of mechanical integrity of the scaffold by deposition of collagen fibers. Here, we studied the effect of fast-degrading polyglycolic acid scaffolds coated with poly-4-hydroxybutyrate (PGA-P4HB) and slow-degrading poly-ɛ-caprolactone (PCL) scaffolds on amount of tissue, composition, and mechanical characteristics in time, and compared these engineered values with values for native human heart valves. Electrospun PGA-P4HB and PCL scaffolds were either kept unseeded in culture or were seeded with human vascular-derived cells. Tissue formation, extracellular matrix (ECM) composition, remaining scaffold weight, tissue-to-scaffold weight ratio, and mechanical properties were analyzed every week up to 6 weeks. Mass of unseeded PCL scaffolds remained stable during culture, whereas PGA-P4HB scaffolds degraded rapidly. When seeded with cells, both scaffold types demonstrated increasing amounts of tissue with time, which was more pronounced for PGA-P4HB-based tissues during the first 2 weeks; however, PCL-based tissues resulted in the highest amount of tissue after 6 weeks. This study is the first to provide insight into the tissue-to-scaffold weight ratio, therewith allowing for a fair comparison between engineered tissues cultured on scaffolds as well as between native heart valve tissues. Although the absolute amount of ECM components differed between the engineered tissues, the ratio between ECM components was similar after 6 weeks. PCL-based tissues maintained their shape, whereas the PGA-P4HB-based tissues deformed during culture. After 6 weeks, PCL-based engineered tissues showed amounts of cells and ECM that were comparable to the number of human native heart valve leaflets, whereas values were lower in the PGA-P4HB-based tissues. Although increasing in time, the number of collagen crosslinks were below native values in all engineered tissues. In conclusion, this study indicates that slow-degrading scaffold materials are favored over fast-degrading materials to create organized ECM-rich tissues in vitro, which keep their three-dimensional structure before implantation.
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