Growth Of Human Chondrocytes Research Articles

Evaluation of the effect of mesenchymal stromal cells from different sources on human chondrocyte proliferation

Objective: to study the effect of a conditioned medium of mesenchymal stromal cells (MSCs) from different sources on human chondrocyte proliferation.Materials and methods. To confirm functional activity, chondrocytes were cultured in a cartilage cell-engineered construct (CEC), including 5 × 105 cells and 5 mg of tissue-specific matrix from decellularized cartilage. The conditioned medium was obtained after culturing MSCs derived from human adipose tissue (AT), MSCs derived from the pulp of primary teeth and MSCs isolated from umbilical cord-derived Wharton’s jelly in a complete cell growth medium (CCGM). To evaluate the effect of MSC-derived secretome on chondrocyte proliferation, the conditioned medium, diluted 1 : 1 with CCGM, was added to wells containing chondrocytes. The effect of MSCs on human chondrocyte proliferation was studied by indirectly coculturing cells in CCGM using Transwell inserts. 5 × 104 MSCs were applied to the bottom of the lower chamber, and 5 × 104 human chondrocytes and 5 mg of matrix were placed in the upper chamber. Chondrocyte proliferation was assessed at days 7 and 14 by DNA quantification. Interleukin-6 content was determined as a marker of secretory activity of MSCs in the conditioned medium. The morphology of the samples was studied using histological staining methods.Results. The ability of chondrocytes to produce cartilage-specific extracellular matrix was confirmed when forming cartilage CEC with tissue-specific matrix in a chondrogenic differentiation medium. When comparing the effect of the conditioned medium of MSCs obtained from different sources on the growth of human chondrocytes in vitro, increased proliferation was observed in all samples compared to controls. Indirect co-culture of MSCs with chondrocytes as part of CEC showed increased DNA amount in all samples at day 14, with the amount of DNA in the sample with MSC conditioned medium significantly higher than the control.Conclusion. Studies on the effect of MSC conditioned medium on chondrocyte proliferation in 2D culture indicate a possible regenerative potential of MSCs for cartilage tissue repair. Within the scope of this work, we did not identify significant differences in the effect of secretome derived from MSCs that were obtained from different sources on chondrocyte proliferation. However, additional in vivo studies are warranted in the future.

PLLA scaffolds produced by thermally induced phase separation (TIPS) allow human chondrocyte growth and extracellular matrix formation dependent on pore size

Damage of hyaline cartilage species such as nasoseptal or joint cartilage requires proper reconstruction, which remains challenging due to the low intrinsic repair capacity of this tissue. Implantation of autologous chondrocytes in combination with a biomimetic biomaterial represents a promising strategy to support cartilage repair.The aim of this work was to assess the viability, attachment, morphology, extracellular matrix (ECM) production of human articular and nasoseptal chondrocytes cultured in vitro in porous poly(l-lactic) (PLLA) scaffolds of two selected pore sizes (100 and 200μm).The PLLA scaffolds with 100 and 200μm pore sizes were prepared via ternary thermally induced phase separation (TIPS) technique and analyzed using scanning electron microscopy (SEM). Articular and nasoseptal chondrocytes were seeded on the scaffold and cultures maintained for 7 and 14days. Live/dead staining, (immuno-)histology and gene expression analysis of type II, type I collagen, aggrecan and SOX9 were performed to assess scaffold cytocompatibility and chondrocyte phenotype.The majority of both chondrocyte types survived on both scaffolds for the whole culture period. Hematoxylin-eosin (HE), alcian blue (visualizing glycosaminoglycans) stainings, immunoreactivity and gene expression of ECM proteins and cartilage marker (type II, I collagen, aggrecan, SOX9) of the chondrocyte scaffold constructs indicated that the smaller pore dimensions promoted the differentiation of the chondrocytes compared with the larger pore size.The present work revealed that the scaffold pore size is an important factor influencing chondrocyte differentiation and indicated that the scaffolds with 100μm pores serve as a cytocompatible basis for further future modifications.

Fluoroquinoloneʼs Effect on Growth of Human Chondrocytes and Chondrosarcomas

Clinical and in vitro studies have demonstrated that fluoroquinolones are toxic to chondrocytes; however, the exact mechanism of fluoroquinolone arthropathy is unknown. We investigated the toxicity of ciprofloxacin on normal cartilage and on cartilaginous tumors. Normal human cartilage, enchondroma, and chondrosarcoma explants were cultured either alone or with the addition of ciprofloxacin at 1, 10, or 20 mg/L of medium. Samples were collected up to twenty-one days after treatment and were processed for electron microscopy and conventional light microscopy. The specimens were characterized morphologically with use of conventional light microscopy, electron microscopy, and immunohistochemistry to identify extracellular matrix, cell proliferation, and apoptosis. Cultures of normal chondrocytes expressed type-II collagen. Electron microscopy revealed a large amount of glycogen in the cells; the presence of fat droplets, rough endoplasmic reticulum, and prominent Golgi apparatus; and a proteoglycan layer surrounding the cells. With prolonged ciprofloxacin treatment and with increased doses, there was an increase in dilated rough endoplasmic reticulum, the appearance of phagosomes, and disintegrated bundles of vimentin filaments. The treated chondrocytes showed a decrease in cell proliferation, but there was no induction of apoptosis or effect on the expression of extracellular matrix proteins. Ciprofloxacin-treated chondrosarcoma cultures and tissue samples showed changes in cartilage matrix composition. Ultrastructural analysis demonstrated clumped glycogen, dilation of endoplasmic reticulum, numerous abnormal lysosomes containing degeneration products, and a decreased proteoglycan deposit surrounding the tumor cells. Treated chondrosarcoma cells and tissue specimens did not proliferate, and apoptosis was induced. In contrast, the in vitro growth of other noncartilaginous malignant tumors like osteosarcoma and liposarcoma was unaffected by ciprofloxacin. Our results indicate that ciprofloxacin is toxic to chondrocytes. In vitro and in vivo treated chondrosarcomas are the most affected.