HIV infects over 33 million people worldwide with a new infection occurring every 9 seconds. Sex is the primary mode of transmission and the majority of new infections occur during unprotected sexual contact between an HIV-infected individual and an uninfected sexual partner(s) since HIV infected individuals tend to shed virus in their genital secretions. The infectiousness of an individual is closely tied to the amount of virus in blood, which is closely associated with HIV levels shed in semen or vaginal fluid or rectal secretions. Although, Highly Active Antiretroviral Therapy (HAART) is associated with complete suppression of HIV RNA in blood to undetectable levels, the impact of HAART on semen HIV RNA levels is less clear. I evaluated the correlation between systemic and mucosal HIV-specific CD8+ T cell immune responses and HIV RNA levels in blood and semen. Overall, there was a strong positive correlation between HIV RNA levels in blood and semen. Neither systemic nor mucosal (in semen) HIV-specific CD8+ responses were associated with HIV RNA levels in blood or semen, in fact CD8+ T cell immune responses in semen correlated with increased HIV RNA levels in semen. Furthermore, inflammatory cytokines (IL-6, and IL-8) CMV levels in semen were associated with increased semen HIV RNA shedding. HAART initiation was associated with complete suppression of HIV viremia, but a significant proportion of individuals on suppressive HAART continue to shed HIV RNA in semen even after 6 months, and this isolated virus was infectious and often present at high levels (> 5000 copies/mL). Nevertheless, long-term HAART was associated with complete immune reconstitution of CD4+ T cells in the sigmoid colon of HIV-infected individuals on long-term therapy. These findings demonstrate that neither systemic nor mucosal HIV-specific CD8+ responses, when assayed with IFN- production as an endpoint, were associated with reduced HIV RNA levels in blood or semen. Semen HIV RNA levels did correlate with local inflammatory cytokines and CMV reactivation. Furthermore, despite effective HAART a significant proportion of HIV-infected men continued to shed HIV RNA in semen. However, long-term completely suppressive HAART was associated with complete immune reconstitution of the sigmoid colon.

HIV infects over 33 million people worldwide with a new infection occurring every 9 seconds. Sex is the primary mode of transmission and the majority of new infections occur during unprotected sexual contact between an HIV-infected individual and an uninfected sexual partner(s) since HIV infected individuals tend to shed virus in their genital secretions. The infectiousness of an individual is closely tied to the amount of virus in blood, which is closely associated with HIV levels shed in semen or vaginal fluid or rectal secretions. Although, Highly Active Antiretroviral Therapy (HAART) is associated with complete suppression of HIV RNA in blood to undetectable levels, the impact of HAART on semen HIV RNA levels is less clear. I evaluated the correlation between systemic and mucosal HIV-specific CD8+ T cell immune responses and HIV RNA levels in blood and semen. Overall, there was a strong positive correlation between HIV RNA levels in blood and semen. Neither systemic nor mucosal (in semen) HIV-specific CD8+ responses were associated with HIV RNA levels in blood or semen, in fact CD8+ T cell immune responses in semen correlated with increased HIV RNA levels in semen. Furthermore, inflammatory cytokines (IL-6, and IL-8) CMV levels in semen were associated with increased semen HIV RNA shedding. HAART initiation was associated with complete suppression of HIV viremia, but a significant proportion of individuals on suppressive HAART continue to shed HIV RNA in semen even after 6 months, and this isolated virus was infectious and often present at high levels (> 5000 copies/mL). Nevertheless, long-term HAART was associated with complete immune reconstitution of CD4+ T cells in the sigmoid colon of HIV-infected individuals on long-term therapy. These findings demonstrate that neither systemic nor mucosal HIV-specific CD8+ responses, when assayed with IFN- production as an endpoint, were associated with reduced HIV RNA levels in blood or semen. Semen HIV RNA levels did correlate with local inflammatory cytokines and CMV reactivation. Furthermore, despite effective HAART a significant proportion of HIV-infected men continued to shed HIV RNA in semen. However, long-term completely suppressive HAART was associated with complete immune reconstitution of the sigmoid colon.

Natural killer (NK) cells represent a crucial component of the innate immune system and are primarily regulated by the interactions of their activation and inhibitory receptors with ligands available on target cells. The genetically linked Ly49 and NKR-P1 family of receptors constitute two of the major regulatory receptor systems used by NK cells and have been shown to bind different ligands. Whereas the Ly49 receptors survey MHC-I ligands on target cells, the NKR-Pl receptor family members bind to various members of the C-type lectin-related (Clr) family. Interestingly, NKR-P1 and Clr haplotypes possess a stable genomic polymorphism across multiple mouse strains, suggesting that this inhibitory receptor:ligand relationship has an important role in the maintenance of host cellular cognate specificities. The NKR-P1 and Clr receptor-ligand pairs identified in mice include the NKR-P1B:Clr-b and the NKR-P1G:Clr-f interacting pairs. Previous RT-PCR and in situ RNA hybridization data generated by our laboratory determined that kidney tubular epithelium as well as the small and large intestinal epithelial cells specifically and highly expresses the Clr-f transcripts. Contrarily, the Clr-b transcripts were only detected on hematopoietic cells of various lymphoid organs and kidneys. Moreover, foregoing studies revealed that the loss of Clr-b following viral or chemical induced stress mediates NK cell killing of the target cell, suggesting a tissue-specific immune-surveillance mechanism in parallel with the global MHC-I-dependent missing-self model. However, the role of the NKR-P1B:Clr-b recognition-system have never been examined in the intestine. Additionally, the role of Clr-f in the kidney and intestines, where they are highly expressed, has not been investigated. For these reasons, I aimed in my thesis to provide a better understanding of the functional aspect of the NKR-P1B:Clr-b and NKR-P1G:Clr-f recognition systems in mediating gut mucosal and renal homeostasis, respectively. First, in order to determine the role of NKR-P1B and Clrb receptor:ligand pair as a “missing-self” immunosurveillance system in the gut, I started by identifying the expression pattern of both the receptor and ligand on various intestinal cells. My results demonstrate that NK cells do not represent the major NKR-P1B-expressing cells in the gut lamina propria. Instead, ILC3 subsets constituted the predominant cell population expressing the receptor, whereas γδT cells composed a small fraction of NKR-P1B+ lymphocytes. In addition, the NKR-P1B expression on myeloid cells was exclusive to colon macrophages and DC subsets. Interestingly, the highest percentage of NKR-P1B+ immune cells was found in the gut, which suggests the dominant role of NKR-P1B in regulating immune functions at the level of intestinal mucosa. As expected, the expression of the NKR-P1B ligand, Clr-b, appeared on all innate immune cell types in the gut. Next, using oral infection models of Salmonela typhimurium and Citrobacter rodentium, I showed that NKR-P1B-deficient NK cells, ILC3 and γδ T cells are hyporesponsive compared to their WT counterparts. In particular, gut NKR-P1B-deficient NK cells and γδT cells secreted low levels of IFNγ cytokine while infected with S.typhimurium. Importantly, the decreased IFNγ secretion by NK and γδT cells was associated with an increased dissemination of the bacterium into the knockout spleens at day 5 post-infection. Likewise, I detected a significant decrease in IL-22 cytokine production by NKR-P1B-deficient ILC3 compared to their WT counterparts at both steady state and following C.rodentium infection. Next, I address the potential role of Clr-f in the kidney. Renal tubular epithelial cells have been shown to express high levels of Clr-f transcripts. Epithelial cells constitute the major cellular component of kidney tubules and are well known to mediate metabolic waste excretion, reabsorption of essential molecules as well as other physiological functions, such as ions exchange and water retention. To determine the role of Clr-f in renal epithelial cells, I generated a Clr-f-deficient mouse with the help of two of my previous lab colleagues. Importantly, chemical analysis on urine and serum samples from knockout and WT littermates indicated that Clr-f-deficient kidneys display a decreased filtration capacity. In particular, higher creatinine levels were detected in the Clr-f deficient serum. In addition, Clr-f-deficient mice appeared to have a lower fractional excretion of sodium (FENa) in their urine filtrates in comparison to WT excreted urine. Blood pressure measurements on the same mice at 12 and 24 weeks of age revealed a hypotensive phenotype in the Clr-f-deficient mice. Furthermore, pathological assessment of Clr-f-deficient kidneys exhibited moderate and aggravated lesions of the tubular epithelium along with marked glomerular mesangiolysis. Lastly, flow cytometry analysis on isolated lymphocytes from Clr-f-deficient and WT mice demonstrated comparable immune infiltrates between the two mouse genotypes. Altogether, our data shows that the absence of Clr-f results in the development of glomerular and tubular lesions in an immune-independent manner leading to an abnormal kidney function. Additionally, the disruption of NKR-P1B:Clr-b recognition system results in abnormal innate immune cell number and function in the mouse intestine. These novel findings sheds light on the important role of Clr-f in maintaining healthy kidney morphology and function, as well as the crucial role for NKR-P1B:Clr-b interactions in mediating intestinal homeostasis at steady and infected states.