IGERT-Lifechips student research report from Matthew Coblyn
NSF Funded IGERT-Lifechips program at the University of California, Irvine presents a research project from Matthew Coblyn, a Lifechips student from the department of Chemical Engineering and Material Science. Matthew conducted his research project in his advisor’s lab, Professor Young Jik Kwon, where he investigated dendritic cells as a target in immunotherapy. Dendritic cells (DCs) are the most potent antigen presenting cells in our immune system. DCs are able to initiate primary T-cell responses and can stimulate/modulate both B and T lymphocytes during an immune response. Therefore, they play a key role in immunotherapy as a target of transfection. Unfortunately, they are present in very low quantities in the body, requiring other methods of acquisition for use in research. One method is to terminally differentiate murine bone marrow cells (BMs) into dendritic cells by growing BMs in media containing Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF).
Matthew’s research looked into the effect of GM-CSF concentration on BM differentiation into dendritic cells by monitoring overall cell densities and cell densities of the cells that present CD11c and CD40 transmembrane proteins over the course of a 10-day growth/differentiation period. Both CD11c and CD40 transmembrane proteins are found in high levels on dendritic cells. Cell densities were measured using flow cytometry and fluorescence markers for CD11c and CD40. The second part of Matthew’s work looked into the effectiveness of nanoparticles with DNA encoding green fluorescence protein (GFP) at transfecting these differentiated dendritic cells. The nanoparticle was composed of a prototype monomer synthesized in Professor Kwon’s lab that was acid degradable. Various dilutions of GM-CSF supernatant from hybridoma cells were created. The GM-CSF concentration in the undiluted supernatant was determined using ELISA but that information is not readily available.
Results showed the the 10-1x dilution was the most effective for differentiating murine bone marrow cells into dendritic cells. At this concentration, peak dendritic cell density occurred on day six of experiment. In the transfection study, dendritic cells were successfully transfected with naked GFP DNA but no transfection occurred using the nanoparticle delivery system. Based on previous work with other cell types and zeta potential measurements, Matthew and his lab believe that the nanoparticles were lost during purification and concentration steps. Matthew proposes that further work must be performed to address this issue and optimize post-synthesis processing.
Lifechips student Matthew Coblyn’s research was a multidisciplinary research because it involved traditional organic chemical synthesis and the more recent fields of tissue engineering and immunotherapies. Lifechips program has played a significant role in his decision and opportunity to work on this project because he was inspired by the idea of joint researches and also Lifechips fellowships allowed him the freedom to meet and work with professors who otherwise might not have had the resources to bring him into their labs.