In vitro culture of primary human myoblasts by using the dextran microcarriers Cytodex3®.
BACKGROUND
Primary cells in vitro culture scale-up is a crucial issue in cell-based tissue and organ regeneration therapy. Reducing costs and space occupied by the cells cultured in vitro has been an important target. Cells cultured in vitro with the use of bioreactor with dextran microcarriers (Cytodex®) have potentially a chance to meet many of the cell therapy requirements.


METHODS
We used collagen-coated carriers (Cytodex3®) and a spinner flask bioreactor to develop environment suitable for human myoblast proliferation. In parallel, standard adherent in vitro culture conditions for myoblasts propagation (T-flask) were conducted. Cell cycle characterization, senescence, myogenic gene ex-pression and cell apoptosis were evaluated in order to find differences between two culture systems under study.


RESULTS
The number of cells obtained in bioreactor per 106 of starting cells population was approximately ten times lower in comparison with T-flask culture system. The microcarriers cultured adult myoblasts in compari-son with the regular T-flask culture showed faster and more advanced replicative aging and lower proliferative potential. Moreover, the percentage of the cells that entailed an irreversible cell arrest (G0 phase) was also significantly (p < 0.0001) increased.


CONCLUSIONS
Our results suggest that population of primary human myoblasts obtained from adult individuals and propagated on dextran microcarriers did not meet the requirements of the regenerative medicine regarding quantity and quality of the cells obtained. Nonetheless, further optimization of the cell scaling up process including both microcarriers and/or bioreactor program is still an important option.
Natalia
N
Rozwadowska
Agnieszka
A
Malcher
Ewa
E
Baumann
Tomasz Jan
TJ
Kolanowski
Marek
M
Rucinski
Tomasz
T
Mietkiewski
Katarzyna
K
Fiedorowicz
Department of Reproductive Biology and Stem Cells, Institute of Human Genetics Polish Academy of Sciences, Poznan, Poland. kurpimac@man.poznan.pl.
Maciej
M
Kurpisz
D000328:Adult; D019149:Bioreactors; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000502:physiology; D016922:Cellular Senescence; D003094:Collagen / Q000737:chemistry; D003911:Dextrans; D006801:Humans; D066298:In Vitro Techniques; D008863:Microspheres; D032446:Myoblasts / Q000166:cytology*
D003911:Dextrans; C052839:Cytodex; D003094:Collagen
T-flask culture; bioreactor; cell cycle; cell senescence; human myoblasts; myogenic gene expression; stem cell precursors; telomeres