Upscaled Skeletal Muscle Engineered Tissue with In Vivo Vascularization and Innervation Potential

Abstract

Engineering functional tissues of clinically relevant size (in mm-scale) in vitro is still a challenge in tissue engineering due to low oxygen diffusion and lack of vascularization. To address these limitations, a perfusion bioreactor was used to generate contractile engineered muscles of a 3 mm-thickness and a 8 mm-diameter. This study aimed to upscale the process to 50 mm in diameter by combining murine skeletal myoblasts (SkMbs) with human adipose-derived stromal vascular fraction (SVF) cells, providing high neuro-vascular potential in vivo. SkMbs were cultured on a type-I-collagen scaffold with (co-culture) or without (monoculture) SVF. Large-scale muscle-like tissue showed an increase in the maturation index over time (49.18 ± 1.63% and 76.63 ± 1.22%, at 9 and 11 days, respectively) and a similar force of contraction in mono- (43.4 ± 2.28 µN) or co-cultured (47.6 ± 4.7 µN) tissues. Four weeks after implantation in subcutaneous pockets of nude rats, the vessel length density within the constructs was significantly higher in SVF co-cultured tissues (5.03 ± 0.29 mm/mm2) compared to monocultured tissues (3.68 ± 0.32 mm/mm2) (p < 0.005). Although no mature neuromuscular junctions were present, nerve-like structures were predominantly observed in the engineered tissues co-cultured with SVF cells. This study demonstrates that SVF cells can support both in vivo vascularization and innervation of contractile muscle-like tissues, making significant progress towards clinical translation.

Cultivation of Bovine MSCs on Plant-Based Scaffolds in a macrofluidic Single-Use Bioreactor for Cultured Meat

“…Reliable and cost-effective methods to create tailor-made bioreactor systems are vital for advancing biotechnology, for proliferating cells in suspension and cultivating cells on scaffolds. In addition there is a great need for versatility and the ability to culture cells on unconventional scaffolds [26][27][28][29][30][31][32][33][34][35][36][37][38][39][40]. This is the case for existing industries such as pharma but also for the emerging cultured meat industry [31,41].…”