Mentor: Dr. Glyn Palmer
College of Medicine
"I first became involved in research to explore a possible career option as a Biology undergrad. I am now involved with Dr. Palmer's experiments out of genuine curiosity of what information we can learn from equine mesenchymal stem cells."
- Stem Cells
- Common Good Award -UF Catalyst 2014
- Certificate of Achievement -Innovation Academy Internship Program
- Florida Bright Futures Scholarship
- American Red Cross
- St. Francis House
Hobbies and Interests
Generation of Novel Reporter Constructs For Quantification of Bone Cell Differentiation
Mesenchymal Stem Cells (MSCs) are a type of adult stem cell found in many tissues and have the ability to form cells of the mesenchymal lineage including cartilage (chondrocytes) and bone cells (osteoblasts). Their high capacity for expansion in cell culture and accessibility make them ideally suited for autologous cell-based therapies for the repair of bone and cartilage lesions. Improved healing of experimentally-induced cartilage lesions and bone fractures has been reported using MSCs in a variety of approaches, and in cell culture, numerous induction protocols have been established for MSC differentiation into the osteoblast and chondrocyte cell lineages. However, despite their widespread use, preclinical studies using MSCs have not progressed much beyond proof-of-concept. We propose that this can be attributed to a combination of factors, including the lack of a definitive MSC marker as well as variations among the selection of MSC cell source and specific methods for induction of differentiation. Finally, methods that are used to assess differentiation often rely on endpoint histological staining which is primarily qualitative and represents an average response across all cells. With current techniques, it is difficult to determine if MSCs within a population uniformly undergo differentiation, or following in vivo application, the extent to which they participate in tissue regeneration. In an effort to establish a more quantitative method for assessment of MSC differentiation our studies are directed towards developing molecular tools that can be used to standardize and compare outcomes of MSC differentiation protocols used in bone and cartilage repair therapies. To this end, we have previously designed and characterized reporter constructs that enable quantification of chondrocyte differentiation in adult MSCs. Complementing these studies, the proposed research project aims to construct novel transcriptional reporters and test each for the ability to quantify osteoblast differentiation in MSC cultures. The constructs will contain fluorescent (GFP) or bioluminescent (luciferase) proteins under the control of an osteoblast-specific gene promoter and be used to quantify osteoblast differentiation in MSC populations and assess cell to cell variations in osteogenic activity. To identify a suitable osteogenic-responsive reporter construct, we will first test candidate promoter sequences of 3 genes associated with osteogenesis: Osteocalcin (OCN),and Indian Hedgehog (IHH). These genes were selected based on their association with the osteoblast phenotype, and upregulation of their expression during osteoblast differentiation of MSCs in cell cultures. Using lentiviral-based reporter constructs encoding luciferase or GFP, we will compare the expression profiles obtained with each promoter with conventional assessments of osteogenesis including cell mineralization and expression of osteogenic markers. The specificity of each promoter-reporter will also be tested by measuring reporter gene expression in MSCs that are differentiated into alternate cell lineages such as fat cells (adipocytes). Promoter constructs will be used in further studies if they demonstrate high levels luciferase and GFP induction during differentiation, good correlation with conventional osteogenic assays, and osteoblast lineage specificity. Having validated one or more of these promoter constructs as an osteoblast-specific reporter, we will be able to use the constructs as tools to compare osteogenic responses among different MSC cell types, and the effects of specific osteoblast differentiation factors.