Genetic Variability of PXR in Saudi Arabians

Polymorphisms in the PXR gene play important roles in influencing the efficacy and toxicity of a large number of endogenous and exogenous substrates. Because of ethnic specificity, several studies have been directed toward the determination of PXR polymorphisms in various populations. In the current study, we determined the genotype and allele frequencies of 19 coding and regulatory polymorphisms in the PXR gene in Saudi Arabians by direct sequencing. Our results show that the frequencies of the regulatory PXR SNPs in Saudi Arabians differ from those in other ethnic groups, and the results endorse the commonly seen ethnic pattern of a paucity of the PXR coding SNPs.     

Antibodies to gp120 and PD-1 Expression on Virus-Specific CD8+ T Cells in Protection from Simian AIDS

We compared the relative efficacies against simian immunodeficiency virus (SIV) challenge of three vaccine regimens that elicited similar frequencies of SIV-specific CD4⁺ and CD8⁺ T-cell responses but differed in the level of antibody responses to the gp120 envelope protein. All macaques were primed with DNA plasmids expressing SIV gag, pol, env, and Retanef genes and were boosted with recombinant modified vaccinia Ankara virus (MVA) expressing the same genes, either once (1 × MVA) or twice (2 × MVA), or were boosted once with MVA followed by a single boost with replication-competent adenovirus (Ad) type 5 host range mutant (Ad5 h) expressing SIV gag and nef genes but not Retanef or env (1 × MVA/Ad5). While two of the vaccine regimens (1 × MVA and 1 × MVA/Ad5) protected from high levels of SIV replication only during the acute phase of infection, the 2 × MVA regimen, with the highest anti-SIV gp120 titers, protected during the acute phase and transiently during the chronic phase of infection. Mamu-A*01 macaques of this third group exhibited persistent Gag CD8⁺CM9⁺ effector memory T cells with low expression of surface Programmed death-1 (PD-1) receptor and high levels of expression of genes associated with major histocompatibility complex class I (MHC-I) and MHC-II antigen. The fact that control of SIV replication was associated with both high titers of antibodies to the SIV envelope protein and durable effector SIV-specific CD8⁺ T cells suggests the hypothesis that the presence of antibodies at the time of challenge may increase innate immune recruiting activity by enhancing antigen uptake and may result in improvement of the quality and potency of secondary SIV-specific CD8⁺ T-cell responses.     

Bacterial Cell Shape–Dependent Inflammatory Response in Mammary Epithelial Cells

An in vitro co-culture model of SCp2 mammary epithelial cells and Escherichia coli strains was established in bacterial non-CO₂ incubators. Co-culturing SCp2 cells with either the rod-shaped W3110 or spherical-shaped GC7378Tn10 strains of Escherichia coli led to an increase in interleukin-6 (IL-6) levels by SCp2 cells after 9 h. At a ratio of 1:100 (epithelial:bacterial), the rod-shaped W3110 strain induced almost double the amount of IL-6 induced by the spherical-shaped GC7378Tn10 strain. The effect of Escherichia coli morphology (rod versus spherical) on IL-6 production by SCp2 cells was further investigated by shifting GC7378Tn10 morphology to rod through introducing the pbpA gene by transduction and transformation. In both approaches, the generated rod strains elicited higher IL-6 levels in SCp2 cells compared to the spherical ones at 1:50 and 1:100 ratios (epithelial:bacterial). Our findings demonstrate the significance of cell shape in bacterial–host interactions with potential implications in bacterial pathogenesis in general.     

Angiotensin II mediates glutathione depletion, transforming growth factor-beta1 expression, and epithelial barrier dysfunction in the alcoholic rat lung

Alcohol abuse markedly increases the risk of sepsis-mediated acute lung injury. In a rat model, ethanol ingestion alone (in the absence of any other stress) causes pulmonary glutathione depletion, increased expression of transforming growth factor-beta1 (TGF-beta1), and alveolar epithelial barrier dysfunction, even though the lung appears grossly normal. However, during endotoxemia, ethanol-fed rats release more activated TGF-beta1 into the alveolar space where it can exacerbate epithelial barrier dysfunction and lung edema. Ethanol ingestion activates the renin-angiotensin system, and angiotensin II is capable of inducing oxidative stress and TGF-beta1 expression. We determined that lisinopril, an angiotensin-converting enzyme inhibitor that decreases angiotensin II formation, limited lung glutathione depletion, and treatment with either lisinopril or losartan, a selective angiotensin II type 1 receptor blocker, normalized TGF-beta1 expression. The glutathione precursor procysteine also prevented TGF-beta1 expression, suggesting that TGF-beta1 may be induced indirectly by angiotensin II-mediated oxidative stress and glutathione depletion. Importantly, lisinopril treatment normalized barrier function in alveolar epithelial cell monolayers from ethanol-fed rats, and treatment with either lisinopril or losartan normalized alveolar epithelial barrier function in ethanol-fed rats in vivo, as reflected by lung liquid clearance of an intratracheal saline challenge, even during endotoxemia. In parallel, lisinopril treatment limited TGF-beta1 protein release into the alveolar space during endotoxemia. Together, these results suggest that angiotensin II mediates oxidative stress and the consequent TGF-beta1 expression and alveolar epithelial barrier dysfunction that characterize the alcoholic lung.     

Role of RNA in facilitating Gag/Gag-Pol interaction

We have examined the influence of RNA upon the interaction of Gag-Pol with Gag during human immunodeficiency virus type 1 (HIV-1) assembly. COS7 cells were transfected with protease-negative HIV-1 proviral DNA, and Gag/Gag-Pol complexes were detected by coimmunoprecipitation with anti-integrase. In COS7 cells, Gag/Gag-Pol is found almost entirely in pelletable, membrane-bound complexes. Exposure of cells to 1% Triton X-100 releases Gag/Gag-Pol from bulk membrane, but the complexes remain pelletable. The role of RNA in facilitating the interaction between Gag and Gag-Pol was examined in these bulk membrane-free, pelletable complexes. The specific presence of viral genomic RNA is not required to maintain the Gag/Gag-Pol interaction, but some type of RNA is, since exposure to RNase destabilized the Gag/Gag-Pol complex. When present only in Gag, the nucleocapsid mutation R7R10K11S, which inhibits Gag binding to RNA, inhibits the formation of both Gag and Gag/Gag-Pol complexes. When present only in Gag-Pol, this mutation has no effect upon complex formation. This result indicates that Gag-Pol may not interact directly with RNA but rather requires RNA-facilitated Gag multimerization for its interaction with Gag.     

In vivo efficacy of antimicrobe-impregnated saline-filled silicone implants

Bacterial colonization of mammary implants is a prelude to clinical infection and has been implicated in the etiology of capsular contracture. Antimicrobial impregnation of a variety of medical devices with the combination of minocycline and rifampin has recently emerged as a potentially effective method for preventing device colonization and device-related infection. The objective of this animal study was to examine in vivo the antimicrobial efficacy of minocycline/rifampin-impregnated, saline-filled silicone implants. A rabbit model of Staphylococcus aureus colonization and infection of subcutaneously placed implants was used. A total of 48 saline-filled silicone implants (24 antimicrobe-impregnated and 24 control unimpregnated implants) were suspended in a 106 colony-forming units/ml bacterial suspension of S. aureus for 30 minutes at room temperature, allowed to dry for 60 minutes, and then implanted subcutaneously in the back of 12 rabbits (two antimicrobe-impregnated and two control implants were placed in each rabbit). Rabbits were monitored daily, then killed either at 2 weeks (10 rabbits) or at 4 weeks (two rabbits) and cultured. The antimicrobe-impregnated implants were 12 times less likely to be colonized than control unimpregnated implants (two of 24 versus 23 of 24; p < 0.001), and they were a significantly less likely cause of implant-related infection (0 of 24 versus 22 of 24; p < 0.001) and implant-related abscess (0 of 24 versus 21 of 24; p < 0.001) than control implants. The minocycline/rifampin-impregnated implants routinely demonstrated zones of inhibition against S. aureus at the time of explantation. These results indicate that minocycline/rifampin-impregnated implants can significantly decrease the rate of bacterial colonization, implant-related infection, and implant-related abscess. Antimicrobe-impregnated implants also have the potential of reducing the likelihood of capsular contracture.