Influence of Conditioned Media on the Re-Differentiation Capacity of Human Chondrocytes in 3D Spheroid Cultures

Abstract

A major challenge of cell-based therapy for cartilage lesions is the preservation of the chondrogenic phenotype during ex vivo cell cultivation. In this in vitro study, the chondro-inductive capacity of two different hyaline cartilage-conditioned cell culture media on human chondrocytes in 3D spheroids was determined. Media were conditioned by incubation of 200 mg/mL vital or devitalized cartilage matrix in growth media over 35 days. The media were analyzed for the content of soluble procollagen type (Col) II and glycosaminoglycans (GAGs) as well as released TGF-β1, IGF-1 and IGFBP3. Unconditioned medium served as a negative control while the positive medium control was supplemented with TGF-β1 and IGF-1. Spheroid cultures prepared from human chondrocytes were cultivated at 37 °C, 5% CO2 and 21% O2 in the respective media and controls. After 14 and 35 days, the deposition of ECM components was evaluated by histological analysis. Vital cartilage-conditioned medium contained significantly higher levels of Col II and active TGF-β1 compared to medium conditioned with the devitalized cartilage matrix. Despite these differences, the incubation with vital as well as devitalized cartilage conditioned medium led to similar results in terms of deposition of proteoglycans and collagen type II, which was used as an indicator of re-differentiation of human chondrocytes in spheroid cultures. However, high density 3D cell cultivation showed a positive influence on re-differentiation.

Osteogenic differentiation of mesenchymal stem cells cultured on allogenic trabecular bone grafts treated with high hydrostatic pressure

“…It is conceivable that these changes caused by HHP enable mediators to diffuse into the cell culture medium where they influence cell differentiation. Such an effect of HHP was already hypothesized by Klinder et al 32 Although the release of various matrix proteins into the surrounding medium over a prolonged period of time could be detected, no differences between control and HHP treatment were observed 32 . However, Klinder et al 32 focused on cartilage, which is characterized by a compact matrix 33 .…”

The Critical Role of Hypoxia in the Re-Differentiation of Human Articular Chondrocytes

“…Culturing chondrocytes in a 3D environment has been widely proposed as a promising approach as it can mimic native articular signaling by re-establishing a specific phenotype and cellular functions of the cartilage compared with other previously reported methods not related to the spheroid generation [ 29 , 30 , 31 ]. Significantly, the 3D model triggers a tissue-specific phenotype due to cell–cell and cell–ECM interactions.…”

Human Chondrocytes from Human Adipose Tissue-Derived Mesenchymal Stem Cells Seeded on a Dermal-Derived Collagen Matrix Sheet: Our Preliminary Results for a Ready to Go Biotechnological Cartilage Graft in Clinical Practice

“…In this translational study, we chose to use human dermal substrate as vector to transduce human AT-MSC-derived chondrocytes since the human skin/dermis remains one of the most efficient sites of type II collagen matrix, significant to accommodate chondrocytes in generating very close biosimilar cartilage-like tissue [ 19 , 20 ]. After a long-term follow-up (11 years) on 61 patients after autologous chondrocyte transplantation graft procedures, it was observed that all of the specimens stained positive for COMP and aggrecan.…”