Using different geometries to modulate the cardiac fibroblast phenotype and the biomechanical properties of engineered connective tissues.

Tissue engineering with human cardiac fibroblasts (CF) allows identifying novel mechanisms and anti-fibrotic drugs in the context of cardiac fibrosis. However, substantial knowledge on the influences of the used materials and tissue geometries on tissue properties and cell phenotypes is necessary to be able to choose an appropriate model for a specific research question. As there is a clear lack of information on how CF react to the mold architecture in engineered connective tissues (ECT), we first compared the effect of two mold geometries and materials with different hardnesses on the biomechanical properties of ECT. We could show that ECT, which formed around two distant poles (non-uniform model) were less stiff and more strain-resistant than ECT, which formed around a central rod (uniform model), independent of the materials used for poles and rods. Next, we investigated the cell state and could demonstrate that in the uniform versus non-uniform model, the embedded cells have a higher cell cycle activity and display a more pronounced myofibroblast phenotype. Differential gene expression analysis revealed that uniform ECT displayed a fibrosis-associated gene signature similar to the diseased heart. Furthermore, we were able to identify important relationships between cell and tissue characteristics, as well as between biomechanical tissue parameters by implementing cells from normal heart and end-stage heart failure explants from patients with ischemic or dilated cardiomyopathy. Finally, we show that the application of pro- and anti-fibrotic factors in the non-uniform and uniform model, respectively, is not sufficient to mimic the effect of the other geometry. Taken together, we demonstrate that modifying the mold geometry in tissue engineering with CF offers the possibility to compare different cellular phenotypes and biomechanical tissue properties.