关键词: 脱细胞基质, 脱细胞骨骼肌, 丝素蛋白, 复合支架, 凝胶支架, 组织工程, 物种差异

Abstract:

Objective In order to investigate physicochemical properties between decellularized matrix (dECM) from different species sources and to select dECM from appropriate species sources for tissue engineering applications, three types of composite scaffolds consisting of decellularized skeletal muscle (DSM) from three different animal sources, namely, bovine, murine, and porcine, composited with silk protein (SF), were designed.
Method DSM from different animal sources of bovine, murine, and porcine were complexed with SF. Histological observations and mechanical properties of natural skeletal muscle tissues were tested before and after decellularization. The mechanical properties, hydrophilicity, infrared spectra and microstructure of the composite gel scaffolds were tested to determine the influence dECM from different species on the gel scaffold properties.
Results Skeletal muscles from different animal sources were decellularized to remove most of the cells and retain the extracellular matrix, and the DNA content of skeletal muscle tissues from bovine, murine, and porcine sources were (25.50±0.75) ng/mg, (40.75±3.77) ng/mg, and (27.00±9.37) ng/mg, which were less than the generally accepted minimum standard of 50 ng/mg. The mechanical properties of skeletal muscle tissues of bovine, murine and porcine origin were tested before and after decellularization, respectively. The results showed that the Young's modulus of fresh skeletal muscle tissues from bovine, murine and porcine animal sources were (1.54±1.05) kPa, (1.21±0.17) kPa, (1.88±1.83) kPa, respectively, and that of decellularized skeletal muscle tissues from bovine, murine and porcine animal sources were (1.05±0.77)kPa, (0.52±0.13) kPa, (0.89±0.61) kPa, and the results of the one-way analysis of variance showed no significant difference in the mechanical properties of skeletal muscle from bovine and porcine animal sources, and no significant difference was observed in murine skeletal muscle either, before and after decellularization treatment. The mechanical properties of three groups of SF-DSM scaffolds from bovine, murine, and porcine sources were tested separately. The results showed that the Young's modulus of SF-DSM scaffolds from bovine, murine, and porcine sources were (5.68±0.49) kPa, (7.24±0.38) kPa, and (5.48±0.44) kPa. The results of the one-way analysis of variance showed significant differences between bovine and murine scaffolds and between porcine and murine scaffolds, but no significant differences were observed between bovine and porcine scaffolds. And the compressive strength of SF-DSM scaffolds from bovine, murine, and porcine sources at 50% compression were (1 391.80±548.72) kPa, (1 316.02±321.86) kPa, and (1 093.69±285.98) kPa, respectively. The porosity of SF-DSM scaffolds from bovine, murine, and porcine sources were measured to be (89.49±3.73)%, (85.30±7.87)%, and (84.67±4.16)%, respectively, and the three groups of scaffolds were porous materials, which were favorable to the cellular adhesion growth. The hydrophilicity of the three groups of scaffolds was tested, and the water absorption rate of SF-DSM scaffolds from bovine, murine and porcine sources were (4.91±1.40) g/g、(6.60±0.04) g/g and (4.69±0.22) g/g, respectively, and the water retention rate were (3.31±0.84) g/g, (3.77±0.21) g/g, (3.02±0.38) g/g, respectively. The results of infrared spectroscopy of SF-DSM scaffolds showed successful cross-linking of SF and DSM from different sources.
Conclusion The mechanical properties of skeletal muscle tissue obtained from bovine, murine, and porcine sources were tested and did not differ significantly between species before and after decellularization. There was no difference in the compressive capacity of SF-DSM scaffolds prepared from bovine, murine, and porcine skeletal muscle decellularized matrices, and there was a significant difference in Young's modulus. No significant differences existed in swelling rate, water retention, and porosity; while the water absorption rate of murine-derived scaffolds was significantly higher than that of porcine-derived ones, but showed no difference with bovine-derived scaffolds. In conclusion the overall performance of SF-DSM scaffolds was not affected by the source of DSM.

Key words: decellularized matrix, decellularized skeletal muscle, silk protein, composite scaffold, gel scaffold, tissue engineering, species difference