Y Chromosome Detection of Three-Dimensional Tissue-Engineered Skeletal Muscle Constructs in a Syngeneic Rat Animal Model

Abstract

Surgical reconstruction of muscle tissue lost by trauma or tumor ablation is limited by the lack of availability of functional native tissue substitution. Moreover, so far most inherited or acquired muscle diseases are lacking sufficient treatment, because only few alternatives exist to provide functional restoration of lost muscle tissues. Engineering those tissues and transplantation into sites of dysfunction may be an alternative approach and may allow replacement of such damaged or failing skeletal muscle tissues. Techniques attempting reconstruction of some human tissues and organs (tissue engineering) have been introduced into clinical practice recently. One major problem that previous transplantation studies were facing is the ability of detection of transplanted cells after integration. Using the Y chromosome in situ hybridization technique in a syngeneic rat model allows transplantation of cell constructs orthotopically, without manipulation of the cells, with no rejection or immunosuppression being implied, but providing a nondilutable genetic marker to identify transplanted cells. The purpose of our study was to create functional skeletal muscle tissue in vivo using the transplantation of primary myoblasts precultivated within a three-dimensional (3D) fibrin matrix and to determine the fate of the transplanted cells using the Y chromosome detection technique. 3D myoblast cultures were established derived from male donor rats and after 7 days of cultivation we performed an orthotopic transplantation of 3D cell constructs into a created muscle defect within the gracilis muscle of syngeneic female rats. Anti-desmin immunostaining and Y chromosome in situ hybridization indicated the survival and integration of transplanted male myoblasts into the female recipient animal, thus demonstrating the feasibility of this approach in tissue engineering and the research of cell transplantation in general.