Treatment of volumetric muscle loss in mice using nanofibrillar scaffolds enhances vascular organization and integration

Abstract

Traumatic skeletal muscle injuries cause irreversible tissue damage and impaired revascularization. Engineered muscle is promising for enhancing tissue revascularization and regeneration in injured muscle. Here we fabricated engineered skeletal muscle composed of myotubes interspersed with vascular endothelial cells using spatially patterned scaffolds that induce aligned cellular organization, and then assessed their therapeutic benefit for treatment of murine volumetric muscle loss. Murine skeletal myoblasts co-cultured with endothelial cells in aligned nanofibrillar scaffolds form endothelialized and aligned muscle with longer myotubes, more synchronized contractility, and more abundant secretion of angiogenic cytokines, compared to endothelialized engineered muscle formed from randomly-oriented scaffolds. Treatment of traumatically injured muscle with endothelialized and aligned skeletal muscle promotes the formation of highly organized myofibers and microvasculature, along with greater vascular perfusion, compared to treatment of muscle derived from randomly-oriented scaffolds. This work demonstrates the potential of endothelialized and aligned engineered skeletal muscle to promote vascular regeneration following transplantation.

Pre-Innervated Tissue Engineered Muscle Promotes a Pro-Regenerative Microenvironment Following Volumetric Muscle Loss

“…Aligned nanofiber scaffolds are the preferred biomaterial for muscle reconstruction since they not only promote myofibers alignment but can also be stacked to provide bulk to the engineered tissue. Aligned nanofiber scaffolds have been shown to facilitate NMJ formation in vitro as well as promote alignment of regenerating myofibers in vivo as compared to randomly oriented nanofibers 19,27,37 .…”

“…Most tissue engineering strategies towards VML repair are evaluated in small animal models 44 . Cell based approaches generally comprise of cell lines (mouse/human) or primary cells and hence are carried out in athymic animals to allow in vivo survival and maturation of the constructs 19,27,44,45 .…”

“…Inadequate re-vascularization remains one of the major challenges to engineer thick skeletal muscle limiting nutrient exchange and survival of implanted cells 23 . Pre-vascularized constructs comprising of preformed vascular networks have been shown to promote microvasculature, vascular perfusion of the graft and inosculation with host vascular system thereby significantly improving muscle regeneration following VML 23,27,47 . Neurotrophic factors like NGF, BDNF, GDNF, NT-3 have been reported to enhance angiogenesis in different tissues like skin, heart and cartilage through receptor mediated activation or recruitment of proangiogenic precursor cells [53][54][55] .…”

“…Acellular scaffolds cannot address such multidimensional challenges and reinforces the necessity of incorporating multiple cell types in a scaffold 18 . Pre-vascularized constructs fabricated by coculture of myoblasts and endothelial cells on aligned nanofibrous scaffolds have been found to promote organized myofiber regeneration and vascular integration in murine VML model 27 . Although angiogenic and immunomodulatory strategies have been explored in VML repair, lack of innervation in an engineered muscle remains one of the major impediments to its success as a functional muscle replacement 28 .…”

Aligned and Conductive 3D Collagen Scaffolds for Skeletal Muscle Tissue Engineering

“…In an effort to address this important unmet medical need research has focused on tissue engineering/regenerative medicine approaches, specifically the use of acellular extracellular matrix (ECM) scaffolds, seeded with myogenic cell sources for subsequent implantation [11][12][13]. Other approaches have employed rationally engineered biomaterial platforms that provide chemical and physical cues to direct cell survival and more accurately mimic healthy ECM microenvironments [14][15][16][17]. Numerous studies also combine biomaterials with isolated muscle cells as well as supporting cells including neurons, endothelial cells, and fibroblasts to augment the repair process [11,[18][19][20][21][22].…”