Generation of a vascularized organoid using skeletal muscle as the inductive source

Abstract

The technology required for creating an in vivo microenvironment and a neovasculature that can grow with and service new tissue is lacking, precluding the possibility of engineering complex three-dimensional organs. We have shown that when an arterio-venous (AV) loop is constructed in vivo in the rat groin, and placed inside a semisealed chamber, an extensive functional vasculature is generated. To test whether this unusually angiogenic environment supports the survival and growth of implanted tissue or cells, we inserted various preparations of rat and human skeletal muscle. We show that after 6 weeks incubation of muscle tissue, the chamber filled with predominantly well-vascularized recipient-derived adipose tissue, but some new donor-derived skeletal muscle and connective tissue were also evident. When primary cultured myoblasts were inserted into the chamber with the AV loop, they converted to mature striated muscle fibers. Furthermore, we identify novel adipogenesis-inducing properties of skeletal muscle. This represents the first report of a specific three-dimensional tissue grown on its own vascular supply.

Cardiac Tissue Engineering in an In Vivo Vascularized Chamber

“…16 This encapsulated tissue is supplied by its own vascular pedicle and is transplantable by microsurgical techniques to other parts of the body, 16 or possibly to an extracorporeal circulation in vitro. This model supports the survival and growth of adipose tissue, 17 implanted skeletal muscle myoblasts, 17 and fibroblasts. 18 In the present study, we examined the ability of this model to support the survival and growth of implanted neonatal rat cardiomyocytes.…”

“…17 The total construct volume consolidates between 1 and 10 weeks, although the absolute volume of cardiac tissue is maintained and the proportion of cardiac tissue increases in association with increased width of the cells. Positive immunostaining with Ki67 and PCNA in tissue constructs harvested at 1 to 10 weeks suggests that a small proportion of cardiomyocytes in the tissue constructs was produced by division of the implanted cardiomyocytes.…”

“…23 Here the cells grow in parallel with the newly developing capillary bed to form a vascularized interconnected network of cardiac tissue. We have previously shown in both rat 21 and mouse 38 chambers that other implanted tissues and cells, such as muscle, 17 myoblasts, 17 fat, 39 pancreatic islets, 40 fetal tissue, and fibroblasts, 16 can survive with this methodology. By seeding cardiomyocytes into a chamber in the rat, we have grown a living piece of cardiac tissue with a volume of Ϸ0.2 mL and thickness up to 1983 m, which easily exceeds the dimensions expected to be supported by diffusion alone (Figure 6).…”

Tissue Engineering of Skeletal Muscle

“…5) which can be implanted into various matrices (Polykandriotis et al 2008). Thus, vascularization in general as well as number and pattern of vessel ingrowth of different matrices can be analyzed (Arkudas et al 2010, Polykandriotis et al 2009) and the vascularized matrix offers a platform for tissue engineering for skeletal (Messina et al 2005) or cardiac (Morritt et al 2007) muscle in vivo. The feasibility of the AV-loop model in large animals ) has been demonstrated recently and poses another step towards a more clinical setting (Beier et al 2010).…”

Engineering vascularised tissues in vitro

“…Morrison et al further developed the concept by combining the AV-loop with polymer matrices and showed enhanced vascularization of the implant (Hofer et al, 2003;Cassell et al, 2001). By implanting the construct ectopically in a highly vascularized region, the AV-loop promoted vessels ingrowth and led to an in vivo prevascularized tissue engineering implant supporting the survival and differentiation of cells (Messina et al, 2005;Bach et al, 2006). This technique was clinically applied to skin tissue engineering (Tanaka et al, 1996(Tanaka et al, , 2000(Tanaka et al, , 2006 and, in combination with hard porous matrices, promoted the survival of transplanted osteoblasts (Kneser et al, 2006 a,b,c).…”