• Characterization of a high-affinity, highly selective tryptophan transport system in the human macrophage and the effects of overexpression of tryptophanyl t-RNA synthetase on Jurkat proliferation

      Seymour, Robert L.; Department of Pediatrics (2004-04)
      Suppression of T cell activation by macrophages/dendritic cells via tryptophan degradation has been shown to play an important role in immunotolerance. Tryptophan degradation is carried out by the enzyme indolamine-2,3-dioxegenase (IDO). This model raises many questions. This study addresses two of these questions. First, how does the macrophage gain access to and degrade tryptophan to a level below 50 nM in culture medium? This is achieved despite the fact that the known high affinity tryptophan transport systems accept other amino acids and have Km values for tryptophan ranging fi-om 10-100 pM. In this study we show that the macrophage possesses a high-affinity, highly selective, and Na-independent tryptophan transport system with a Km for tryptophan of about 300 nM. This would allow the macrophage to have effective access to tryptophan at concentrations in the nanomolar range. We also show T cells do not possess this transport system. Second, how does the T cell sense the level of intracellular free tryptophan? It has been shown in the past that if T cells are stimulated in medium containing less than SOOnM tryptophan that they attempt to activate but arrest in mid-Gl of the cell cycle. The enzyme tryptophanyl t-RNA S5mthetase (WRS) charges tRNA*'^ with tryptophan. This enzjmie has two protein isoforms, with one having a non-canonical N-terminal kinase domain. WRS is also upregulated by interferon gamma (IFNy). These characteristics put WRS in a position to be an intracellular free tryptophan sensor. Here we show that transient transfection of the T cell line, Jurkat, with cDNA encoding the kinase-containing isoform of WRS inhibits proliferation. In addition, to the above we have ereated a subline of Jurkat, which stably arrests in the absence of tryptophan. We also show that this new subline is resistant to the drug G418 but is sensitive to hygromycin. When treated with rapamycin the Jurkat sub line will stably arrest in the presence or absence of tryptophan. Rapamycin is a known immunosuppressive agent, which inhibits T cell proliferation. This leads to the speculation of a possible link between the signaling pathways involved in tryptophan sensing and those involved in the effects of rapamycin.
    • EFFECTS OF CHRONIC ALOHOL AND GLUCOSE EXPOSURE ON VIABILITY OF ALVEOLAR MACROPHAGES

      Keller, Elizabeth; College of Science and Mathematics; College of Allied Health; Curry-McCoy, Tiana; Thomas, Amanda; Augusta University (2019-02-13)
      The adverse health risks associated with alcohol abuse and obesity are widely known by the general population. Although lesser known, studies have presented the lungs as secondary organs affected by such lifestyle factors. Healthy lungs are protected against infection and harmful airborne particles by macrophages, the working entities of the immune system which fight potential sources of infection. When these immune-responsive cells are compromised and unable to perform their functions, lung health may deteriorate. Therefore, a healthy pulmonary alveolar macrophage population is vital for adequate lung function. Chronic alcohol abuse and obesity have been shown to suppress alveolar macrophage function, thus lowering the lungs� first line of defense. The objective of this study is to determine the effects of exogenous ethanol and increased glucose concentration on macrophage size and viability via an�in vitro�study on NR8383 rat alveolar macrophages. The study measures macrophage viability under treatment conditions.
    • Role of Splenic Macrophages in the Initiation of Tolerance to Apoptotic Cell Associate Antigens

      Ravishankar, Buvana; Department of Biochemistry and Molecular Biology (2014-03)
      Systemic Autoimmune disease occurs due to the breakdown of tolerance to self-antigens caused in part by impaired clearance of apoptotic cells. The spleen is a primary site for generation of the tolerogenic response to selfantigens in the periphery. The marginal zone (MZ), which contains the specialized macrophage (MΦs) populations: Marginal Zone Macrophages (MZMs) and Metallophilic Marginal zone Macrophages (MMMs), as well as B cells and dendritic cells (DCs) play a requisite role in capture of apoptotic cells and the initiation of tolerance to associated self antigens. Moreover, defective MZ cellular architecture may lead to increased auto-reactivity that exacerbates autoimmune like disease progression. MZMs are specialized to recognize and capture apoptotic cells and promote peripheral tolerance to apoptotic cells and associated antigens. However, the mechanism by which MZMs enforce this tolerance is not known. Thus, the overall goal of our project is to fill the lapse in the scientific knowledge and understanding of the mechanism(s) by which the splenic stromal MΦs drive immune tolerance to apoptotic cell associated antigens.