Wound Regeneration and Repair Methods and Protocols Methods in Molecular Biology 1037

Abstract

We describe an in vivo model system designed to evaluate the host response to implanted biomaterials:\rThe partial thickness rat abdominal wall defect model. The model allows for determination of the temporal\rand spatial distribution of the cellular and vascular response, the remodeling of the implanted material and\rsurrounding host soft tissue, and the function of the remodeled tissue over time. \r\rThis chapter reviews the use of urothelial cells as a means to enhance tissue regeneration and wound\rhealing in urinary tract system. It addresses the properties of urothelial cells, including their role as a permeability\rbarrier to protect underlying muscle tissue from the caustic effects of urine and as one of the\rmain cell types, along with smooth muscle cells, that are used in urethral or bladder tissue engineering\rtoday. This description includes a general overview of various isolation techniques and culture methods\rthat have been developed to improve urinary tract reconstruction in vivo and aid the characterization of\rgrowth factor expression in vitro. The chapter then describes various applications using urothelial cells,\rincluding production of multilayer urothelial sheets, tissue engineered bladder mucosa, tissue engineered\rurethra, and tissue engineered bladder. It also outlines the advantages of sandwich and layered coculture\rof these cells and the effects of epithelial–stromal cell interactions during tissue regeneration or wound\rhealing processes in the urinary tract. \rThe fi broblast-populated collagen lattice (FPCL) was intended to act as the dermal component for\r“skin- equivalent” or artifi cial skin developed for skin grafting burn patients. The “skin-equivalent” was\rclinically unsuccessful as a skin graft, but today it is successfully used as a dressing for the management of\rchronic wounds. The FPCL has, however, become an instrument for investigating cell–connective tissue\rinteractions within a three-dimensional matrix. Through the capacity of cell compaction of collagen fi brils,\rthe FPCL undergoes a reduction in volume referred to as lattice contraction. Lattice contraction proceeds\rby cell-generated forces that reduce the water mass between collagen fi bers, generating a closer relationship\rbetween collagen fi bers. The compaction of collagen fi bers is responsible for the reduction in the\rFPCL volume. Cell-generated forces through the linkage of collagen fi bers with fi broblast’s cytoskeletal\ractin- rich microfi lament structures are responsible for the completion of the collagen matrix compaction.\rThe type of culture dish used to cast FPCL as well as the cell number will dictate the mechanism for compacting\rcollagen matrices. Fibroblasts, at moderate density, cast as an FPCL within a petri dish and released\rfrom the surface of the dish soon after casting compact collagen fi bers through cell tractional forces.\rFibroblasts at moderate density cast as an FPCL within a tissue culture dish and not released for 4 days\rupon release show rapid lattice contraction through a mechanism of cell contraction forces. Fibroblasts at\rhigh density cast in an FPCL within a petri dish, released from the surface of the dish soon after casting,\rcompact a collagen lattice very rapidly through forces related to cell elongation. The advantage of the\rFPCL contraction model is the study of cells in the three-dimensional environment, which is similar to the\renvironment from which these cells were isolated. In this chapter methods are described for manufacturing\rcollagen lattices, which assess the three forces involved in compacting and/or organizing collagen fi brils\rinto thicker collagen fi bers. The clinical relevance of the FPCL contraction model is related to advancing\rour understanding of wound contraction and scar contracture.