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Characterization of human myoblast cultures for tissue engineering
Skeletal muscle tissue engineering, a promising specialty, aims at the reconstruction of skeletal muscle loss. In vitro tissue engineering attempts to achieve this goal by creating differentiated, functional muscle tissue through a process in which stem cells are extracted from the patient, e.g. by...
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Published in: | International journal of molecular medicine 2008-01, Vol.21 (1), p.49-56 |
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Main Authors: | , , , , , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that cite this one |
Online Access: | Get full text |
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Summary: | Skeletal muscle tissue engineering, a promising specialty, aims at the
reconstruction of skeletal muscle loss. In vitro tissue engineering attempts to
achieve this goal by creating differentiated, functional muscle tissue through
a process in which stem cells are extracted from the patient, e.g. by muscle biopsies,
expanded and differentiated in a controlled environment, and subsequently re-implanted.
A prerequisite for this undertaking is the ability to cultivate and differentiate
human skeletal muscle cell cultures. Evidently, optimal culture conditions must
be investigated for later clinical utilization. We therefore analysed the proliferation
of human cells in different environments and evaluated the differentiation potential
of different culture media. It was shown that human myoblasts have a higher rate
of proliferation in the alamarBlue® assay when cultured on gelatin-coated culture
flasks rather than polystyrene-coated flasks. We also demonstrated that myoblasts
treated with a culture medium with a high concentration of growth factors [growth
medium (GM)] showed a higher proliferation compared to cultures treated with a
culture medium with lower amounts of growth factors [differentiation medium (DM)].
Differentiation of human myoblast cell cultures treated with GM and DM was analysed
until day 16 and myogenesis was verified by expression of MyoD, myogenin, α-sarcomeric
actin and myosin heavy chain by semi-quantitative RT-PCR. Immunohistochemical
staining for desmin, Myf-5 and α-sarcomeric actin was performed to verify the
myogenic phenotype of extracted satellite cells and to prove the maturation of
cells. Cultures treated with DM showed positive staining for α-sarcomeric actin.
Notably, markers of differentiation were also detected in cultures treated with
GM, but there was no formation of myotubes. In the enzymatic assay of creatine
phosphokinase, cultures treated with DM showed a higher activity, evidencing a
higher degree of differentiation. In this study, we obtained detailed information
regarding the cultivation and differentiation of human myoblast cultures in different
environments. By exploring optimal culture conditions for skeletal muscle tissue
engineering, we acquired culture data for comparison with other sources of stem
cells in order to find the most applicable stem cell for focussed clinical usage. |
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ISSN: | 1107-3756 1791-244X |
DOI: | 10.3892/ijmm.21.1.49 |