Tissue Regeneration of Human Mesenchymal Stem Cells on Porous Gelatin Micro-Carriers by Long-Term Dynamic In Vitro Culture

BACKGROUND: Tissue engineering is a multidisciplinary field which attracted much attention in recent years. One of the most important issue in tissue engineering is how to obtain high cell numbers and tissue regeneration while maintaining appropriate cellular characteristics in vitro for restoring damaged or dysfunctional body tissues and organs. These demands can be achieved by the use of three dimensional (3D) dynamic cultures of cells combined with cell-adhesive micro-carriers. METHODS: In this study, human mesenchymal stem cells (hMSCs) were cultured in a wave-bioreactor system for up to 100 days, after seeding on Cultisphere-S porous gelatin micro-carriers. Cell counting was performed at the time points of 7, 12, 17, 31 days and compared to those of hMSCs cultured under static condition. Higher growth and proliferation rates was achieved in wave-type dynamic culture, when cell culture continued to day 31. A scanning electron microscope (SEM) photographs, both live and dead and MTT assays were taken to confirm the survival and distribution of cells on porous gelatin micro-carrier surfaces. The results of histological stains such as hematoxylin and eosin, Masson’s trichrome, Alcian blue and Alizarin red S also showed improved proliferation and tissue regeneration of hMSCs on porous gelatin micro-carriers. CONCLUSION: The experimental results demonstrated the effect and importance of both micro-carriers and bioreactor in hMSC expansion on cell proliferation and migration as well as extracellular matrix formation on the superficial and pore surfaces of the porous gelatin micro-carriers, and then their inter-connections, leading to tissue regeneration.

The optimization of culture conditions of MRC -5 human diploid cell / 预防医学

Objective To optimize the culture conditions of MRC -5 human diploid cell.Methods To compare the growth status of MRC -5 cells,three kinds of culture medium with T25 bottles and Spinner cultivation system Cytodex1 micro carrier were used.Morphology,cell counting,growth curve,glucose -lactic acid value were observed and detected daily for screening a kind of suitable medium.Cell proliferation was compared with different levels of the bovine serum.Results There were no significant differences among the three kinds of culture medium.There were significant differences among MEM((43.25 ±0.60)×104 cells/mL,(12.98 ±1.27)×105 cells /mL),M199 ((35.40 ±1.41 )×104 cells/mL, (10.76 ±1.31)×105 cells /mL)and DMEM/F12 ((36.75 ±1.59)×104 cells/mL,(11.22 ±1.42)×105 cells /mL)(P<0.01).The cell proliferation of MEM cultures was 5.17 and 6.49 times better than those of M199 and DMEM/F12 cultures.Imported fetal bovine serum cell proliferation ((4.55 ±0.51)×105 cells /mL)was better than the other three bovine serum ((4.12 ±1.03,3.59 ±0.48,3.53 ±0.52)×105 cells /mL).Conclusion Tree kinds of culture medium can be used to culture MRC -5 human diploid cell.The MEMculture is better.Imported fetal bovine serum is better than other kinds of serum.