Although mesenchymal stem/stromal cells (MSCs) chondrogenic differentiation has been thoroughly investigated, the rudiments for enhancing chondrogenesis have remained largely dependent on external cues. Since aggregation of MSCs, a prerequisite for chondrogenesis, generates tension within the cell agglomerate, we theorized that the initial number of the cells within the aggregate could function as an intrinsic activator of a mechanobiology paradigm and alter the outcomes. We discovered that reducing aggregate cell number (ACN) from 500k to 70k leads to activation and acceleration of the chondrogenic differentiation, independent of soluble chondro-inductive factors, via β-catenin dependent TCF/LEF transcriptional activity and expression of anti-apoptotic protein survivin. Our state-of-the-art mechanical testing revealed a correlation between progression of chondrogenesis and emergence of stiffer cell phenotype. In-depth Affymetrix gene array analysis proposed that the down-regulation of genes associated with lipid synthesis and regulation could account for observed outcomes. Furthermore, we illustrate that implanting aggregates within collagenous matrix not only decreases the necessity for high quantity of cells but also leads to drastic improvement in quality of the deposited tissue. In summary, our study presents a simple and donor-independent strategy to enhance the efficiency of MSCs chondrogenic differentiation and demonstrates a correlation between MSCs chondrogenesis and mechanical properties with potential translational applications.

 

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