a)Trans-differentiation of Adult Stem cells
Differentiation of Stem cells into different lineages has immense potential in cell replacement therapy. Our group has focused on improving rates of differentiation of adult stem cells using 3-Dimensional culture systems, improving isolation of pure populations of stem cells from bone marrow and other sources such as Umbilical cord blood and the Heart. One of the major objectives we are currently pursuing is cardiac regeneration using stem cells, which includes the resident stem cell population and through differentiation of Bone marrow derived stem cells. Cells when grown in culture adhere to the surface of a treated plastic flask or dish, which is very different from the 3 Dimensional interactive environments in which they reside in vivo. While recreating an exact environment akin to the natural matrix is a considerable feat, a 3 dimensional space can be fabricated that can support proliferation and induce differentiation spontaneously or by directed measures. Scaffolds can be biological or artificial in origin. Biological scaffolds tend to have some advantages such as; being biological in nature they are much more compatible and are usually biodegradable, easily synthesized and fabricated. The focus of our study is achieving differentiation of stem cells in vitro and maintaining a considerably sized culture of the same for cell-based therapy.
The scaffold we have employed is fabricated from PLA and we study the proliferation and maintenance of stem cells over PLA microspheres. We investigate the possibility of the creation of niche like structures inside these cell-PLA aggregates. We have attempted to study the differentiation of bone marrow derived Mesenchymal stem cells into specific lineages by using PLA microspheres with specific differentiation agents such as 5- Azacytidine, Oxytocin and other inducers.
Human bone marrow derived mesenchymal stem cells (BMSCs) have got tremendous potential to differentiate into many cell kinds of the mesodermal, ectodermal and endodermal lineages. Hepatic transdifferentiation of hBMSCs by various hepatogenic growth factors, serum components as well as by genetic manipulations have been reported both in vitro and in vivo. A culture system containing cholestatic serum (obtained from obstructedbile duct) is an innovative way of proliferating mesenchymal stem cells and differentiating them selectively towards hepatocyte like cells. In such a hepatogenic environment, BMSCs are fine tuned towards hepatic lineage due to presence of liver specific activating factors, as is happening in vivo in liver injury conditions to maintain hepatic homeostasis. The primary objective of this project is to evaluate the effectiveness of cholestatic serum of a unique kind (obtained from heart failure patients having etilogy of ischemic hepatitis or cardiac jaundice) in inducing hepatic transdifferentiation of hBMSCs on various scaffolds of natural origin. Human BMSCs derived can be used as a novel in vitro model for hepatotoxicity testing. Also, in future, it can be used as effective cell source fornoninvasive therapeutic transplantation in treating injured livers. However, a more detailed biochemical profiling and molecular characterization of cholestatic serum obtained from heart failure patients is needed before taking it for clinical applications.
b) Development of a natural Biomaterial for Cardiac regeneration
The growing importance of mesenchymal stem cell isolation and culture for stem cell based therapies in cardiac regeneration emphasizes the need for mammalian models to assess efficacy and procedure standardization. We have developed four putative immortalized BMSC cell lines which were characterized and found to be of MSC origin along with variability in cell surface marker expression. Absence of polyploidy, chromosomal aberrations and translocations was confirmed in these putative cell lines. The BMSC cell lines showed characteristics of mesenchymal stem cell line and therefore, could be specifically induced to differentiate in to cardiomyogenic lineage using 5-azacytidine. A Ventricular CMG cell line was thus generated from these BMSC cell lines. We have also developed an efficient biodegradable cardiac biomaterial which supports better growth of BMSC, enhances cardiomyogenic differentiation of BMSC, possesses protective properties against hypoxia conditions and can be used as a scaffold with potential therapeutic applications in cardiac tissue regeneration. We are currently involved in molecular and functional characterization of the cardiac biomaterial to validate its application as a cardiac regenerative scaffold.
c) Development and Characterization of Tissue Engineered Xenografts
Acellular Tissue-engineered or decellularized xenografts and homografts have already been implanted in humans or are currently approaching the clinical setting. Efficiently decellularized, tissue-engineered homografts might prolong durability by reducing recipient inflammation, fibrous scarring, and calcification, ultimately decreasing the number of patients requiring reconstructive cardiac surgery. The postulated proof of immune reactions towards acellular xenografts and growth potential being translated to humans lacks any scientific evidence. The objective of this project is to examine host immune response toward acellular tissue engineered xenografts and further examines the effects of cell seeding on acellular scaffold.
Intuitions