Abstract
BackgroundTissue engineering of vascularised skeletal muscle is a promising method for the treatment of soft tissue defects in reconstructive surgery. In this study we explored the characteristics of novel collagen and fibrin matrices for skeletal muscle tissue engineering. We analyzed the characteristics of newly developed hybrid collagen-I-fibrin-gels and collagen nanofibers as well as collagen sponges and OPLA®-scaffolds. Collagen-fibrin gels were also tested with genipin as stabilizing substitute for aprotinin.ResultsWhereas rapid lysis and contraction of pure collagen I- or fibrin-matrices have been great problems in the past, the latter could be overcome by combining both materials. Significant proliferation of cultivated myoblasts was detected in collagen-I-fibrin matrices and collagen nanofibers. Seeding cells on parallel orientated nanofibers resulted in strongly aligned myoblasts. In contrast, common collagen sponges and OPLA®-scaffolds showed less cell proliferation and in collagen sponges an increased apoptosis rate was evident. The application of genipin caused deleterious effects on primary myoblasts.ConclusionCollagen I-fibrin mixtures as well as collagen nanofibers yield good proliferation rates and myogenic differentiation of primary rat myoblasts in vitro In addition, parallel orientated nanofibers enable the generation of aligned cell layers and therefore represent the most promising step towards successful engineering of skeletal muscle tissue.
Highlights
Tissue engineering of vascularised skeletal muscle is a promising method for the treatment of soft tissue defects in reconstructive surgery
The gels with a fibrin concentration of 2,5 mg/ml showed a significant increase of cell number (p < 0,05) after 14 days when seeded with 100.000 cells
MyoD and MEF-2d were clearly over-expressed in gels with low cell concentration (100.000 cells, group 1 and 3) but in contrast, high cell concentration (500.000 cells, group 2 and 4) provoked down-regulation of desmin and MyoD and slight up-regulation of MEF-2d in case of group 4. These findings suggest that muscle differentiation and gene expression depend on cell-cell interactions and the initial cell number may have a considerable influence on myoblast differentiation and tissue development
Summary
Tissue engineering of vascularised skeletal muscle is a promising method for the treatment of soft tissue defects in reconstructive surgery. In this study we explored the characteristics of novel collagen and fibrin matrices for skeletal muscle tissue engineering. We analyzed the characteristics of newly developed hybrid collagen-I-fibrin-gels and collagen nanofibers as well as collagen sponges and OPLA®-scaffolds. Collagen-fibrin gels were tested with genipin as stabilizing substitute for aprotinin. Several attempts have been pursued to test suitable materials with various consistencies, such as gels [5,6] , sponges [7] e.g. and most recently electrospun nanofibers [8,9,10,11,12]. In this study our first aim was to develop and establish novel biocompatible matrices for myoblast cultivation. We intended to directly compare these novel matrices with varying different consistencies and micromorphologies to established tissue engineering matrices which had not yet been applied in skeletal muscle generation. We wanted to assess and identify their advantages and disadvantages as matrices for skeletal muscle tissue engineering
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