1. The Role of Mast Cells and CD117 in the Spread of HIV-1 Particles in Vitro Stacey Baker, Department of Biology, York College Project Summary Little is known about the relationship between HIV-1, mast cells, and CD117. When HIV-1 infects mast cells, CD117 receptors increase and cause apoptosis. This allows for release and spread of the virus. Studies have shown that HIV-1 may be stored in mast cells and at the point of apoptosis the virus particles are released and may spread rapidly to surrounding cells. The intention of this proposal is to determine if individuals with mast cell disorders have a lower susceptibility to HIV infection than do individuals with normal mast cell levels. To determine the susceptibility of HIV-1 infection, tissue samples will be grown and infected with different concentrations of HIV. The level of apoptosis and CD117 markers will be measured by the TUNEL ASSAY and flow cytometry. Results will be analyzed to determine if the tissues that have lower concentrations of mast cells will have a lower concentration of virus particles and apoptosis. This would be useful in determining if a treatment can be created to lower the susceptibility to HIV-1 infection in individuals. Introduction  CD117 is a cytokine receptor found mainly on mast cells at moderate levels. It is responsible for cell survival, differentiation, and proliferation. When CD117 is found in high concentrations it is linked to various carcinomas.  Mast cells are found mainly in mucosal areas of the body. Their main role in the organism is immune responses through degranulation and inflammation.  HIV-1 is a human immunodeficiency virus that affects many different types of cells in the body. Review of Literature  Human Immunodeficiency Virus causes apoptosis in many cells that are infected with this virus (Ahr et al. 2004).  Recent studies have shown that mast cells infected with HIV-1 may possibly be a reservoir for virus particles (Sundstrom et al.2004). This is due to the fact that when the mast cells undergo degranulation, they release the virus to surrounding mucosal tissues which can be the primary site of HIV infection (Petrow, Ruscetti, Taub, and Mikovits 1999).  Other studies have identified that high expression levels of CD117 are linked to cell apoptotic death and CD117 may serve several functions in the organism (He et al. 1997). Objective To determine if mast cells, and their expression of CD117, affect the spread of HIV-1 in infected individuals. To determine if a mast cells disorder can in some way make an individual less susceptible to becoming infected with HIV. H o : Individuals with mast cell disorders have a lower susceptibility to HIV-1 infection. Prediction I predict to see a decrease in the number of virus particles and infected tissue cells for tissue with a decrease in mast cells and a decreased level of CD117 expression. ~With a low number of mast cells the virus won’t replicate as quickly. ~There will be a decrease in cells that undergo apoptosis and spread the virus particles. ~The initial infection will need a higher concentration of virus particles to infect the tissue. Figure 1: Comparing the effect of mast cells concentration and release of HIV virions from tissues. Amount of HIV released from the tissue Concentration of mast cells in tissues Methods  Obtain tissue in vitro 1. Lung will be used for tissue with high levels of mast cells.  IHC procedures to locate CD117  Determine CD117 expressions levels by: 1. Staining Intensity 2. Flow Cytometry  Separate Tissue into two categories: 1. High concentration of mast cells 2. Low concentration of mast cells ~ A single observer blind to the study will count the mast cells  Grow tissue cultures ~ 25 samples from each category  Different Concentrations of HIV-1 applied to tissue  Monitored every 12 hours and samples collected 1. Apoptosis measured by TUNEL assay 2. Virus particles examined under the microscope  Data Analyzed Literature Cited Sundstrom, J. Bruce, Dawn M.Little, Francois Villinger, Jane E. Ellis, and Aftab A. Ansari. "Signaling through Toll-Like Receptors Triggers HIV-1 Replication in Latently Infected Mast Cells." The Journal of Immunology 172(2004): 4391-4401. Petrow, Cl, Ruscetti FW, Taub DD, Mikovits JA.. "In vitro infection of human mast cells by HIV-1 stimulates degranulation and apoptosis.." American Society of Microbiology 99(1999): He, Jianglin, Carlos M. DeCastro, George R. Vandenbark, Jorge Busciglio, and Dana Gabuzda. "Astrocyte apoptosis induced by HIV-1 transactivation of the c-kit proto-oncogene.." The national academy of Science 94(1997): 3954-3959. Acknowledgments I would like to thank Dr. Ander Pindzola for his advice and guidance in understanding the information. I would also like to thank Dr. Carolyn Mathur for her support in helping me during senior thesis and creating the poster. Prediction Continued I will also predict to see an increased number of virus particles in tissue samples that have a increased number of CD117 expression and apoptosis. ~The more mast cells that break apart due to the increased level of CD117, the more virus particles that will be spread to surrounding tissue. ~A lower concentration of virus will be needed to begin the initial infection. Conclusion Individuals with a mast cell disorder will be less susceptible of contracting HIV-1 when exposed to the virus at low concentrations. This is due to the idea that the virus won’t be replicated as quickly in tissue and the low concentration can be destroyed by natural immune system cells. This information would be helpful in HIV research to see if it is possible to contain HIV-1 infected cells in certain locations of the body and avoid spreading HIV throughout the organism. CD117 expression in tissues Amount of HIV released from the tissue Figure 2: Comparing the expression of CD117 on mast cells and release of HIV virions from tissues. Taken from: http://moon.ouhsc.edu Taken from: http://www.cihr-irsc.gc.ca/images/PHIL_2.jpg Figure 3: Comparing the level of apoptosis in mast cells and release of HIV virions from tissue. Amount of HIV released from the tissue Level of apoptosis in tissue