Background: Stable cartilage regeneration in immunocompetent large animals remains a bottleneck problem that restricts clinical application. The inflammation elicited by degradation products of scaffolds has a decisive influence on cartilage formation. Although prolonged preculture in vitro could form mature engineered cartilage and allow sufficient degradation of scaffolds, the inflammatory reaction was still observed. This study explored the feasibility of using chondrocyte sheet technology to regenerate stable cartilage in the subcutaneous environment with a pig model. Methods: Passage 1 chondrocytes were used to form cell sheets by high-density culture. As a control, chondrocytes were seeded onto polyglycolic acid/polylactic acid scaffolds for 6 and 12 weeks’ in vitro preculture, respectively. Then, they were autologously implanted subcutaneously into pigs for 2, 8, and 24 weeks. Gross view, histologic staining, and biochemical and biomechanical characteristics were evaluated. Results: With prolonged culture in vitro, relatively homogeneous engineered cartilages were formed with less scaffold residue. However, the chondrocyte–polyglycolic acid/polylactic acid group still encountered severe inflammation and inferior cartilage formation at 2 and 8 weeks in vivo. The engineered cartilage with cell sheet technique exhibited a relatively more stable and mature tissue structure without obvious inflammatory response at 24 weeks in vivo, which was similar to the native auricular cartilage. Conclusions: The chondrocyte sheet technique could successfully regenerate mature and stable engineered cartilages in pig models. It is possibly an effective method of repairing cartilage defects in the clinic that uses regenerated substitutes derived from autologous cell sheets.
Tissue engineering cartilage is a promising strategy to reconstruct the craniofacial cartilaginous defects. It demands plenty of chondrocytes to generate human-sized craniofacial frameworks. Partly replacement of chondrocytes by adipose-derived stem cells (ADSCs) can be an alternative strategy. The study aimed at evaluating the chondrogenic outcome of ADSCs and chondrocytes in direct co-culture with transforming growth factor-beta (TGF-β3). Porcine ADSCs and chondrocytes were obtained from abdominal wall and external ears. Four groups: ADSCs or chondrocytes monocultured in medium added with TGF-β3; ADSCs and ACs co-cultured with or without TGF-β3. Cell growth rate was performed to evaluate the cell proliferation. Morphological, histologic and real-time polymerase chain reaction analysis were performed to characterize the chondrogenic outcome of pellets. ADSCs had favorable multi-lineage differentiation potential. Further, when ADSCs were co-cultured with chondrocytes in medium added with TGF-β3, the cell proliferation was promoted and the chondrogenic differentiation of ADSCs was enhanced. We demonstrate that pellet co-culture of ADSCs and chondrocyte with TGF-β3 could construct high quantity cartilages. It suggests that this strategy might be useful in future cartilage repair.
Paenibacillus polymyxa is a well-known Gram-positive biocontrol bacterium. It has been reported that many P. polymyxa strains can inhibit bacteria, fungi and other plant pathogens. Paenibacillus polymyxa employs a variety of mechanisms to promote plant growth, so it is necessary to understand the biocontrol ability of bacteria at the genome level. In the present study, thanks to the widespread availability of Paenibacillus genome data and the development of bioinformatics tools, we were able to analyze and mine the genomes of 43 P. polymyxa strains. The strain NCTC4744 was determined not to be P. polymyxa according to digital DNA-DNA hybridization and average nucleotide identity. By analysis of the pangenome and the core genome, we found that the pan-genome of P. polymyxa was open and that there were 3,192 core genes. In a gene cluster analysis of secondary metabolites, 797 secondary metabolite gene clusters were found, of which 343 are not similar to known clusters and are expected to reveal a large number of new secondary metabolites. We also analyzed the plant growth-promoting genes that were mined and found, surpisingly, that these genes are highly conserved. The results of the present study not only reveal a large number of unknown potential secondary metabolite gene clusters in P. polymyxa, but also suggest that plant growth promotion characteristics are evolutionary adaptations of P. polymyxa to plant-related habitats.
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