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Drannik AG, Nag K, Yao X-D, Henrick BM, Jain S, Ball BT, Plummer FA, Wachihi C, Kimani J, Rosenthal KL. "Anti-HIV-1 Activity of Elafin Is More Potent than Its Precursor's, Trappin-2, in Genital Epithelial Cells.". 2012. AbstractWebsite

Cervicovaginal lavage fluid (CVL) is a natural source of anti-HIV-1 factors; however, molecular characterization of the anti-HIV-1 activity of CVL remains elusive. In this study, we confirmed that CVLs from HIV-1-resistant (HIV-R) compared to HIV-1-susceptible (HIV-S) commercial sex workers (CSWs) contain significantly larger amounts of serine antiprotease trappin-2 (Tr) and its processed form, elafin (E). We assessed anti-HIV-1 activity of CVLs of CSWs and recombinant E and Tr on genital epithelial cells (ECs) that possess (TZM-bl) or lack (HEC-1A) canonical HIV-1 receptors. Our results showed that immunodepletion of 30% of Tr/E from CVL accounted for up to 60% of total anti-HIV-1 activity of CVL. Knockdown of endogenous Tr/E in HEC-1A cells resulted in significantly increased shedding of infectious R5 and X4 HIV-1. Pretreatment of R5, but not X4 HIV-1, with either Tr or E led to inhibition of HIV-1 infection of TZM-bl cells. Interestingly, when either HIV-1 or cells lacking canonical HIV-1 receptors were pretreated with Tr or E, HIV-1 attachment and transcytosis were significantly reduced, and decreased attachment was not associated with altered expression of syndecan-1 or CXCR4. Determination of 50% inhibitory concentrations (IC50) of Tr and E anti-HIV-1 activity indicated that E is ~130 times more potent than its precursor, Tr, despite their equipotent antiprotease activities. This study provides the first experimental evidence that (i) Tr and E are among the principal anti-HIV-1 molecules of CVL; (ii) Tr and E affect cell attachment and transcytosis of HIV-1; (iii) E is more efficient than Tr regarding anti-HIV-1 activity; and (iv) the anti-HIV-1 effect of Tr and E is contextual

Juno J, Tuff J, Choi R;, Card C, Kimani J, Wachihi C, Koesters-Kiazyk S, Ball BT, Farquhar C, Plummer FA, John-Stewart G, Luo M, Fowke KR. "The role of G protein gene GNB3 C825T Polymorphism in HIV-1 acquisition, progression and immune activation.". 2012. Abstract

The GNB3 C825T polymorphism is associated with increased G protein-mediated signal transduction, SDF-1α-mediated lymphocyte chemotaxis, accelerated HIV-1 progression, and altered responses to antiretroviral therapy among Caucasian subjects. The GNB3 825T allele is highly prevalent in African populations, and as such any impact on HIV-1 acquisition or progression rates could have a dramatic impact. This study examines the association of the 825T polymorphism with HIV-1 acquisition, disease progression and immune activation in two African cohorts. GNB3 825 genotyping was performed for enrolees in both a commercial sex worker cohort and a perinatal HIV transmission (PHT) cohort in Nairobi, Kenya. Ex vivo immune activation was quantified by flow cytometry, and plasma chemokine levels were assessed by cytokine bead array. Results GNB3 genotype was not associated with sexual or vertical HIV-1 acquisition within these cohorts. Within the Pumwani cohort, GNB3 genotype did not affect HIV-1 disease progression among seroconverters or among HIV-1-positive individuals after adjustment for baseline CD4 count. Maternal CD4 decline and viral load increase in the PHT cohort did not differ between genotypes. Multi-parametric flow cytometry assessment of T cell activation (CD69, HLA-DR, CD38) and Treg frequency (CD25+FOXP3+) found no differences between genotype groups. Plasma SDF-1α, MIP-1β and TRAIL levels quantified by cytokine bead array were also similar between groups. Conclusions In contrast to previous reports, we were unable to provide evidence to suggest that the GNB3 C825T polymorphism affects HIV-1 acquisition or disease progression within African populations. Ex vivo immune activation and plasma chemokine levels were similarly unaffected by GNB3 genotype in both HIV-1-negative and HIV-1-positive individuals. The paucity of studies investigating the impact of GNB3 polymorphism among African populations and the lack of mechanistic studies make it difficult to assess the true biological significance of this polymorphism in HIV-1 infection.

Schellenberg JJ, Dumonceaux TJ, Hill JE, Kimani J, Jaoko W, Wachihi C, Mungai JN, Lane M, Fowke KR, Ball BT. "Selection, phenotyping and identification of acid and hydrogen peroxide producing bacteria from vaginal samples of canadian and East African women.". 2012.
Plummer FA, Luo M, Ball TB, Kimani J, Wachihi C, Tuff J, Lacap P, Price H. "A Trim5alpha Exon 2 Polymorphism is Associated with Protection from HIV-1 Infection in Pumwani Sexworker Cohort.". 2010. Abstract

The innate immune component TRIM5α has the ability to restrict retrovirus infection in a species-specific manner. TRIM5α of some primate species restricts infection by HIV-1, while huTRIM5α lacks this specificity. Previous studies have suggested that certain polymorphisms in huTRIM5 may enhance or impair the proteins affinity for HIV-1. This study investigates the role of TRIM5 polymorphisms in resistance/susceptibility to HIV-1 within the Pumwani sex worker cohort in Nairobi, Kenya. A group of women within this cohort remain HIV-1 seronegative and PCR negative despite repeated exposure to HIV-1 through active sex work. Design A 1 kb fragment of Trim5alpha gene, including exon 2, from 1032 women enrolled in the Pumwani sex worker cohort was amplified and sequenced. SNPs and haplotypes were compared between HIV-1 positive and resistant women. Methods The TRIM5 exon 2 genomic fragment was amplified, sequenced and genotyped. Pypop32-0.6.0 was used to determine SNP and haplotype frequencies and statistical analysis was carried out using SPSS-13.0 for windows. Results A TRIM5 SNP (rs10838525) resulting in the amino acid change from Arginine to Glutamine at codon 136, was enriched in HIV-1 resistant individuals (p=1.104E-05; OR:2.991; CI95%:1.806–4.953) and women with 136Q were less likely to seroconvert (p=0.002; Log Rank: 12.799). Wild type TRIM5α exon 2 was associated with susceptibility to HIV-1 (p=0.006; OR:0.279; 95%CI:0.105–0.740) and rapid seroconversion (p=0.001; Log Rank: 14.475). Conclusions Our findings suggest that a shift from arginine to glutamine at codon 136 in the coiled-coil region of TRIM5α confers protection against HIV-1 in the Pumwani sex worker cohort. Keywords: TRIM5α, Single nucleotide polymorphism, HIV-1, Sex Workers, Taxonomy-based Sequence Analysis, Disease Association, Disease Resistance

McKinnon LR, Ball TB, Wachihi C, Chinga N, Maingi A, Luo M, Fowke KR, Plummer FA. "Substantial Intrapatient Differences In The Breadth And Specificity Of Hiv-specific Cd8+ T-cell Interferon-gamma And Proliferation Responses.". 2008. Abstract

HIV vaccine design and evaluation require a better understanding of protective immune responses. HIV-specific CD8+ T-cell responses have been characterized extensively using interferon-gamma (IFN-gamma) enzyme-linked immunosorbent spot (ELISPOT) assays, which do not always correlate with control of viral replication or disease progression. Alternative aspects of CD8+ T-cell responses, in particular those associated with a central memory (Tcm) phenotype, may be more protective against disease progression. To determine the extent that the breadth and specificity of HIV-specific CD8+ T-cell responses differ based on immunological readout, we screened in HIV-infected Kenyan sex workers for responses to HIV Env using IFN-gamma ELISPOT and 6-day carboxyfluorescein succinimidyl ester-based proliferation assays. This comparison revealed substantial differences in the epitopes recognized when the assay readout was IFN-gamma versus proliferation. Although 24 and 41 IFN-gamma and proliferative responses were identified, overlapping specificity was observed for only 5 responses. Breadth also differed between assays in several patients. Env-specific IFN-gamma breadth was found to correlate inversely with CD4 count (r = -0.66, P = 0.005), although this was not the case for proliferation. These data suggest that efforts to define HIV-specific CD8+ T-cell responses may need to be revisited using additional immunological readouts.

Lester RT, Yao X-D, Ball BT, McKinnon LR, Kaul R, Wachihi C, Jaoko W, Plummer FA, Rosenthal KL. "Toll-like receptor expression and responsiveness are increased in viraemic HIV-1 infection.". 2008. Abstract

Toll-like receptors (TLR) are important in pathogen recognition and may play a role in HIV disease. We evaluated the effect of chronic untreated and treated HIV-1 infection on systemic TLR expression and TLR signalling.Together, these data indicate that chronic viraemic HIV-1 is associated with increased TLR expression and responsiveness, which may perpetuate innate immune dysfunction and activation that underlies HIV pathogenesis, and thus reveal potential new targets for therapy.

Alimonti JB, Koesters SA, Kimani J, Matu L, Wachihi C, Plummer FA, Fowke KR. "Cd4+ T Cell Responses In Hiv-exposed Seronegative Women Are Qualitatively Distinct From Those In Hiv-infected Women.". 2005. AbstractWebsite

The immune response of human immunodeficiency virus (HIV)-exposed seronegative (ESN) women may be qualitatively different from that in those infected with HIV (HIV(+)). In a cohort of female commercial sex workers in Nairobi, Kenya, we found significantly lower (P< or =.01) levels of CD4(+)-specific immune activation and apoptosis in the ESN women compared with those in the HIV(+) women. Compared with the HIV(+) women, a lower proportion of the ESN women showed p24 peptide pool responses by the short-term, CD4(+)-specific, interferon (IFN)- gamma intracellular cytokine staining assay, whereas the proportion showing responses by the long-term, CD8(+)-depleted T cell proliferation assay was similar. Interestingly, the ESN responders had a 4.5-fold stronger proliferation response (P=.002) than the HIV(+) group. These data suggest that, compared with those in HIV(+) women, CD4(+) T cells in ESN women have a much greater ability to proliferate in response to p24 peptides.

WACHIHI DRWAIGWACHARLES, WANGUI DRGITAURUTH. "Quantitative ex vivo analysis of functional virus-specific CD8 T lymphocytes in the blood and genital tract of HIV-infected women. Kaul R, Thottingal P, Kimani J, Kiama P, Waigwa CW, Bwayo JJ, Plummer FA, Rowland-Jones SL.AIDS. 2003 May 23;17(8):1139-44.". In: AIDS. 2003 May 23;17(8):1139-44. Doctoral Thesis; 2003. Abstract
BACKGROUND: CD8 T lymphocytes are important in HIV-1 control and mediate virus-specific immunity in the blood and genital tract. The induction and monitoring of mucosal CD8 cell responses will be an important component of HIV-1 vaccine trials, but information regarding the frequency, phenotype and function of genital tract CD8 cell responses is lacking. METHODS: Simultaneous blood and cervical cytobrush samples were obtained from 16 HIV-1-infected Kenyan sex workers. Epitope-specific CD8 T lymphocyte frequencies in the blood and genital tract were analysed after short-term peptide incubation and intracellular cytokine staining for interferon-gamma (IFN gamma). RESULTS: Cervical sampling resulted in adequate cell numbers for analysis in 10/16 women. Background IFN gamma production was higher in CD3+/CD8+ lymphocytes from the genital tract than from blood (0.48% versus 0.1%; P < 0.01). Responses to staphylococcal enterotoxin B were detected in cervical CD8 lymphocytes from 10/10 women, at a similar frequency to blood (16.7% in cervix and 13.3% in blood; P = 0.4). HIV-1-specific responses were detected the cervix of 8/10 women, with a trend to higher response frequencies in the genital tract than blood (2.1% versus 0.8%; P = 0.09). Co-expression of integrin CD103 (alpha E beta 7), a mucosal marker, was used to confirm the mucosal origin of cervical responses. CONCLUSIONS: Cytobrush sampling and intracellular cytokine staining is well suited to the analysis of cervical CD8 cell responses. The frequency of functional virus-specific CD3+/CD8+ T cells is similar in the genital tract and blood of HIV-1-infected women. The role of genital tract CD8 cell responses in HIV-1 control warrants further investigation.

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