Composite Tissue Regeneration

Broad objectives in this area are i) to understand the influence of structural features, chemical signals, and mechanical signals on the proliferation and differentiation of stem cells from various sources and ii) to utilize these conditions in forming composite tissue grafts.  Successful completion of this project will have significant impact on i) the regeneration and transplantation of high quality tissues on-demand, ii) the development of synthetic surrogates to test disease states (mechanism of wound healing), toxicology studies and effect of pathogens i.e., as real time sensors for detecting biological agents, and iii) importantly changes the way cell culture experiments will be performed in the future worldwide.

Background.  With the advent of bone marrow transplantation to cure various hematologic disorders, cell based therapies have see significant attention.  In particular stem cells from various have been harvested and explored for use.  In addition, the plastic nature of stem cells is well demonstrated in a variety of models; bone marrow cells can differentiate into liver cells or muscle cells based on the location of their recruitment.  Based on these success, many cell-based therapies have been investigated in clinical trials.  For example, bone, cardiac tissue and cartilage. However, stem cell based therapies have not seen significant success due to a major problem of poor regeneration attributed to attrition of injected cells.  Non-invasive approaches using injectable hydrogels have shown marginal improvements as cells could be dispersed instead of injecting pellets.  However, significant loss of cells remains a problem and hydrogels are also very weak relative to native tissues.  In addition, current products obtainable by tissue engineering are limited to tissues/organs without blood vessels due to the technological limitations in developing organized vasculature.  Using stem cells to differentiate to diverse cell types, including endothelial cells i.e., the cells forming the vascular networks is an option.  However, the primary challenge is to transcribe the in vivo microenvironment into in vitro conditions so that vascularized tissues can be regenerated in 3D configuration.  Evaluating 3D configurations are critical a) to understand the spatio-temporal effects, b) to evaluate the reorganization of various compartments and organ formation, and c) to develop devices that can be used in clinical applications.  Thus, exploring the possibility of differentiating cells in 3D will significantly help as an alternative cell source in tissue regeneration.  A number of 3D models are explored using matrigels obtained from a mouse sarcoma.  However, matrigels do not accurately reflect the diversity of proteins of most tissues; for example, Laminin-1 is not present at high quantities in most adult tissues.  Our innovation is that we mimic by introducing tissue-specific matrix elements without using any cross-linkers.

Our approach.  With the advent of genetically inducing pluripotency in mature cells (called induced pluripotent stem cells), we explore differentiating adult cells by controlling the scaffold architecture and signaling.  We utilize adult stem cells from different tissues such as bone marrow, cord blood and adipose tissue.  We evaluate the regeneration patterns and compare how stem cells would perform at two levels a) microscale and b) nanoscale using novel technologies.  Human foreskin fibroblasts are explored to form cartilage.  We alter hydrogel formulation to suite a required tissue, chemically crosslink one component selectively within the hydrogel to improve mechanical properties, and incorporate nutrients including oxygen releasing molecules.  We use anisotropic injectable hydrogels that can be used with minimally invasive surgical procedure.  In addition, we evaluate bioreactor configurations that could apply different stress to different types of cells within the scaffold.  We evaluate cell differentiation using various cell specific markers.  We perform many evaluations including cell growth, attachment, and assembly and maturation of matrix.  We use the injectable hydrogels to form vasculatures and other types of tissues within the same template.