Introduction: Articular cartilage defects are common, particularly in the young and active population.

Background: Most treatments available for articular cartilage defects are not completely satisfactory as they tend to result in the production of fibrocartilage instead of hyaline cartilage.

Aim: We aimed at evaluating the chondrogenic differentiation of marrow-isolated adult multilineage inducible (MIAMI) cells in contact with pharmacologically active microcarriers (PAMs) releasing TGF-β3, microcartilage, or a combination thereof. We hypothesize that combination of TGF-β3-releasing PAMs and microcartilage will enhance and guide the chondrogenic differentiation of MIAMI cells toward a hyaline phenotype.

Materials and Methods: MIAMI cells were isolated from swine bone marrow. PAMs are biocompatible and biodegradable 60 µm PLGA microspheres coated with a biomimetic surface and delivering TGF-β3 in a controlled fashion. Microcartilage is made of micronized and dried cadaveric human articular cartilage. The attachment, survival and chondrogenic differentiation of swine MIAMI (swMIAMI) cells in contact with PAMs and/or microcartilage was assessed by microscopy, histological staining, RT-qPCR and immunohistochemistry of the cartilage pellets formed after 21 days of chondrogenic differentiation in vitro.

Results: SwMIAMI completely attached to PAMs and microcartilage in 24 hours in vitro, and were then pelleted to undergo in vitro chondrogenesis. Microcartilage and PAMs TGF-β3 contributed to the overall size of the neo-cartilage, with PAMs having a strong effect on swMIAMI cell survival and glycosaminoglycan production. Using RT-qPCR, we observed the strongest effect in terms of hyaline-specific cartilage gene expression, i.e. high aggrecan and low type X collagen, with the combination of both PAMs TGF-β3 and microcartilage. We confirmed by immunohistochemistry the overexpression of aggrecan and down-regulation of type X collagen in swMIAMI cells exposed to both microcartilage and PAMs delivering TGF-β3.

Discussion: Our main finding was that the exclusive combination of the natural microcartilage and synthetic PAMs delivering TGF-β3 provide a unique environment with adequate support and signaling molecules to direct the differentiation of swMIAMI cells in vitro towards a phenotype closely resembling that of hyaline cartilage instead of a more fibrocartilaginous phenotype.

Conclusions: In the future, we hope this study will help designing a minimally invasive combinatorial strategy to provide a faster healing of the damaged articular cartilage with improved microarchitecture and mechanical properties.