• Morphological and Functional Characterization of GPR109A in Mammalian Retina

      Gambhir, Deeksha; Department of Biochemistry and Molecular Biology (2013)
      Vision is considered as one of the most important of the five human senses. This was reflected by a poll that was based on human psychology that revealed that the fear of losing vision is much higher than the fear of losing any of the other senses (Leo et al., 1999). Diabetic retinopathy (DR) is a leading cause of blindness worldwide. Diabetic retinopathy is defined as damage to retina caused by the complications of diabetes. Hyperglycemia is a major factor in the pathogenesis of diabetic retinopathy (Davis et al., 1998). The diabetes-induced metabolic and physiologic abnormalities in the retina are consistent with chronic inflammation (Kern et al., 2007). G-protein coupled receptors (GPCRs) comprise a large protein family of transmembrane receptors that play central roles in several biological processes (Wettschureck and Offermanns, 2005). GPCRs have the ability to bind to chemically distinct ligands, and as such are widely used as targets for pharmaceuticals aimed at treating pain, inflammation, and a broad spectrum of diseases (Vassilatis et al., 2003; Offermanns et al., 2006). GPR109A is a newly discovered Grlinked GPCR known for its anti-lipidemic and anti-inflammatory properties in several cell types. Here, my major goal was to study the expression and functional role of GPR109A in retina an its relevance to diabetes-associated retinal implications.
    • The Role of Tumor Necrosis Factor Alpha (TNF-a) In the Eyes and Brain of HSV-1 Infected Euthymic BALB/C Mice

      Fields, Mark A; Department of Biochemistry and Molecular Biology (2007-12)
      After uniocular anterior chamber (AC) inoculation of HSV-1, virus and TNFalpha (TNF-a) are detected in the eyes and brain o f HSV-1 infected euthymic BALB/c mice. The overall goal o f this study was to investigate the role o f TN F-a in the eyes and brain o f HSV-1 infected BALB/c mice. Mice were treated with thalidomide for TN F-a inhibition or injected with clodronate liposomes to deplete macrophages, and the AC of one eye (ipsilateral) was injected with HSV-1 (KOS). In thalidomide-treated mice, both suprachiasmatic nuclei (SCN) were infected by day 5 p.i. and the titer of virus in the SCN contralateral to the side o f injection was increased. In macrophage depleted mice, both SCN were infected at day 6 p.i. and the titer o f virus in the SCN o f these mice was increased at day 6 and 7 p.i. compared with controls. The titer o f virus in the contralateral (uninoculated) eye o f macrophage depleted mice was increased at day 7 p.i. The results o f these studies suggest that TN F-a plays a role in limiting virus replication in the SCN o f euthymic BALB/c mice and that one source of TN F-a is macrophages. In order to further investigate the role of TNF-a, a recombinant o f HSV-1 (KOS77VF) was constructed that produces TNF-a constitutively. Euthymic BALB/c mice were injected in one anterior chamber with the TNF-a recombinant, with a recombinant containing the pCI plasmid, with a recombinant rescue virus, or with the parental virus. Mice from each group were sacrificed on day 1-9 p.i. and the uninjected eyes were removed. In the uninjected eye o f KOSTNF -infected mice, TN F-a expression was increased and there were more viral antigen positive cells and immune inflammatory cells. There was earlier microscopic evidence o f retinal infection and destruction in these mice. In addition, the titer o f virus in the uninjected eye was significantly increased in KOSTiVF-infected mice on day 7 p.i. compared with KOSpCI, KOS6{irescue, or with KOS6/? infected mice. These results suggest that overproduction o f TN F-a by HSV-1 (KOS7WF) facilitates spread o f virus infection in the eye through increased inflammation and TNF-a-mediated damage to retinal cells.