Objective To explore the effect of rolling compression loading bioreactor on chondrogenesis of rabbit bone marrow mesenchymal stem cells (BMSCs) with different loading parameters. Methods BMSCs were isolated from New Zealand rabbits, aged 2.5 months. BMSCs at passage 3 were used to prepare BMSCs-agarose gels (4 mm in diameter and height, respectively). Samples were divided into 8 groups: 10% (group A1), 20% (group A2), and 30% (group A3) compression groups (0.4 Hz, 3 h/ d) and 20 minutes (group B1), 3 hours (group B2), and 12 hours (group B3) rolling time groups and static culture (control groups). The living cell rate, the collagen type II and Aggrecan gene expressions, and glycosaminoglycan (GAG) content were determined, and histological staining was done at 24 hours, 7 days, 14 days, and 21 days after culture. Results At 14 and 21 days, the living cell rates of groups A1 and A2 were significantly higher than that of group A3 (P lt; 0.05), groups B1 and B2 were significantly higher than group B3 (P lt; 0.05). Collagen type II and Aggrecan gene expressions of the experimental groups at each time point were significantly higher than those of the control groups (P lt; 0.05); at 14 and 21 days, collagen type II and Aggrecan gene expressions of groups A1 and A2 were significantly higher than those of group A3, and groups B1 and B2 were also significantly higher than group B3 (P lt; 0.05). At 14 and 21 days, the GAG contents of groups A1 and A2 were significantly higher than those of group A3 (P lt; 0.05); groups B1 and B2 were also significantly higher than group B3 (P lt; 0.05). At 21 days, toluidine blue staining showed that obvious blue-staining and even cartilage lacunae were seen in groups A2 and B2, but light and quite rare blue-staining in groups A1, A3, B1, and B3. Conclusion The rolling compression loading bioreactor has great promotion effect on chondrogenesis of rabbit BMSCs with rolling parameters of 0.4 Hz, 3 hours, and 20% compression.
Objective Collagen type II is a characteristic molecular of chondrocyte. With continuous subculture of chondrocytes, they progressively lose the abil ity to express collagen type II. To observe the effect of collagen type IIon redifferentiation of dedifferentiated rabbit chondrocytes so as to lay a experimental foundation for use of chondrocytes in cartilage tissue engineering. Methods Cartilage was harvested under sterile conditions from tibio-femoral joints of 7-monthold New Zealand white rabbit. The rabbit articular chondrocytes were subcultured in vitro to the 7th generation (named P1-P7).Dedifferentiated rabbit chondrocytes were chosen by RT-PCR, real-time PCR, and 1, 9-dimethylmethylene blue (DMMB) assay. Then dedifferentiated rabbit chondrocytes were treated with various concentrations (0, 0.5%, 1.0%, and 1.5%) of exogenous collagen type II. The redifferentiation of dedifferentiated chondrocytes was measured by RT-PCR and real-time PCR, and the glycosaminoglycan content was determined by DMMB assay. Results The glycosaminoglycan content of P1-P7 chondrocytes were (12.20 ± 0.17), (11.20 ± 0.24), (11.18 ± 0.16), (10.89 ± 0.50), (8.73 ± 0.19), (9.39 ± 0.32), and (8.18 ± 0.20) μg, respectively, showing no significant difference (P gt; 0.05) among P2, P3, and P4, and showing significant differences (P lt; 0.05) among other generations. The mRNA of collagen type I, collagen type II, and aggrecan expressed at P4-P7, showing no significant difference in the mRNA expression of collagen type I (P gt; 0.05) and significant differences in the mRNA expressions of collagen type II and aggrecan (P lt; 0.05) among P4-P7. The glycosaminoglycan content at concentrations of 0, 0.5%, 1.0%, and 1.5% were (8.20 ± 0.16), (14.61 ± 0.33), (13.93 ± 0.25), and (19.59 ± 0.46) μg, showing significant differences among different concentrations (P lt; 0.05). With exogenous collagen type II concentrations increased, the mRNA expressions of collagen type II and aggrecan gene were up-regulated gradually, but collagen type I gene was down-regulated, showing significant differences (P lt; 0.05). Conclusion Collagen type II can promote redifferentiation and activation of dedifferentiated rabbit chondrocytes.
Assessment of Real World Observational Studies (ArRoWS) is a tool developed by the Leicester Real World Evidence (LRWE) Unit of the Diabetes Research Centre of the University of Leicester in the United Kingdom to assess the quality of real world evidence research, and has been reported to have good practicability. ArRoWS can be used to quickly and specifically assess the quality of real world evidence research that uses electronic health record information. The tool contains 16 items, nine of which are common items, and seven of which are related to specific research designs. The current study introduces the development background, development process, assessment items, assessment criteria, and application methods of ArRoWS and other related aspects, to provide references for real world researchers in China.