Human immunodeficiency virus type 1 quasi species that rebound after discontinuation of highly active antiretroviral therapy are similar to the viral quasi species present before initiation of therapy

In an effort to identify the sources of the viruses that emerge after discontinuation of therapy, analyses of human immunodeficiency virus (HIV) quasi species were done for 3 patients with sustained levels of HIV RNA of <50 copies/mL for 1-3 years. The sequences found in the rebounding plasma virus were closely related to those of the actively replicating form of viruses present before the initiation of combination therapy. All quasi species found in the rebounding plasma virus were also present in proviral DNA, cell-associated RNA in peripheral blood mononuclear cells (PBMC), and virion RNA derived from PBMC coculture during periods when plasma HIV RNA levels were <50 copies/mL. These findings suggest that the rapid resurgence of plasma viremia observed after discontinuation of therapy and the viruses cocultured from PBMC are derived from a relatively stable pool of the replicating form of virus rather than from activation of a previously latent pool.     

Four distinct antigenic regions are present in the primary structure of HIV-1 and HIV-2 proteinases.

OBJECTIVE: The purpose of this study was to assay reactivity of antibody-positive sera to different parts of the HIV-1 and HIV-2 proteinase (PR) proteins. DESIGN: Since the majority of HIV-1-antibody-positive sera react to the proteinase, but the antigenic determinants on the protein have not been identified, we attempted to identify these determinants. INTERVENTIONS: We synthesized 18 peptides representing the PR of HIV-1 and HIV-2 in order to map serum reactivity to the PR protein. RESULTS: Both HIV-1- and HIV-2-antibody-positive sera recognized four distinct antigenic regions in the HIV-1 and HIV-2 PR. CONCLUSIONS: Correlation between our results and the crystallographic structure of the protein revealed that the antigenic regions are positioned at the surface of the HIV-1 PR. Although the structure of HIV-2 PR has not yet been characterized, our results indicate that the folding of the HIV-1 and HIV-2 PR may be very similar.     

Omentin-1 and vaspin are present in the fetus and neonate, and perinatal concentrations are similar in normal and growth-restricted pregnancies

The aim of this study was to investigate circulating concentrations of omentin-1 and vaspin (adipocytokines predominantly secreted by visceral adipose tissue and not yet investigated in perinatal life) in maternal, fetal, and neonatal samples from intrauterine growth-restricted (IUGR; associated with altered development of adipose tissue) and appropriate-for-gestational-age (AGA) pregnancies and to correlate them with the respective insulin concentrations. Serum omentin-1 and vaspin concentrations were determined by enzyme immunoassay in 40 mothers and their 20 IUGR and 20 AGA singleton full-term fetuses and neonates on postnatal day 1 (N1) and day 4 (N4). Both hormones were detectable in fetal and neonatal blood (omentin-1 [mean ± SD, in nanograms per milliliter]: AGA vs IUGR group—fetal: 11.32 ± 1.88 vs 10.47 ± 1.30, N1: 10.74 ± 1.42 vs 10.46 ± 1.54, and N4: 10.90 ± 2.72 vs 11.35 ± 3.92; vaspin [median, minimum-maximum; in nanograms per milliliter]: AGA vs IUGR group—fetal: 0.39 [0.04-19.06] vs 0.40 [0.05-1.34], N1: 0.40 [0.04-16.70] vs 0.44 [0.23-3.34], and N4: 0.49 [0.02-8.89] vs 0.55 [0.06-3.92]). No significant differences in omentin-1 or vaspin concentrations were observed between IUGR and AGA groups, whereas fetal and N1 insulin concentrations were lower in the former (P = .025 and P = .027, respectively). In both groups, fetal omentin-1 concentrations were higher (P ≤ .018), whereas vaspin concentrations were lower (P ≤ .001), than maternal ones. Furthermore, maternal vaspin concentrations were higher in cases of cesarean delivery (P = .024). Omentin-1 and vaspin concentrations did not correlate with the respective insulin ones. In conclusion, omentin-1 and vaspin are present in the fetus and neonate. Perinatal concentrations of omentin-1 and vaspin are similar in IUGR cases and AGA controls—despite lower insulin concentrations in the former—and do not correlate with the respective insulin concentrations. Higher omentin-1 concentrations in the fetus may be crucial to enhance a growth-promoting effect, whereas lower maternal vaspin concentrations in cases of vaginal delivery may be attributed to spontaneous term delivery inflammation.     

Pattern of interleukin-1beta secretion in response to lipopolysaccharide and ATP before and after interleukin-1 blockade in patients with CIAS1 mutations

OBJECTIVE: To examine the synthesis, processing, and secretion of interleukin-1beta (IL-1beta), as well as the clinical and biologic effects of IL-1 blockade, in patients with chronic infantile neurologic, cutaneous, articular (CINCA) syndrome and Muckle-Wells syndrome (MWS), in an effort to understand the molecular mechanisms linking mutations of the CIAS1 gene and IL-1beta hypersecretion, and the underlying response to IL-1 receptor antagonist (IL-1Ra). METHODS: Six patients with CINCA syndrome or MWS were treated with IL-1Ra and followed up longitudinally. Monocytes obtained from the patients and from 24 healthy donors were activated with lipopolysaccharide (LPS) for 3 hours, and intracellular and secreted IL-1beta levels were determined by Western blotting and enzyme-linked immunosorbent assay before and after exposure to exogenous ATP. RESULTS: LPS-induced IL-1beta secretion was markedly increased in monocytes from patients with CIAS1 mutations. However, unlike in healthy subjects, secretion of IL-1beta was not induced by exogenous ATP. Treatment with IL-1Ra resulted in a dramatic clinical improvement, which was paralleled by an early and strong down-regulation of LPS-induced IL-1beta secretion by the patients' cells in vitro. CONCLUSION: Our results showed that the requirements of ATP stimulation for IL-1beta release observed in healthy individuals are bypassed in patients bearing CIAS1 mutations. This indicates that cryopyrin is the direct target of ATP and that the mutations release the protein from the requirement of ATP for activation. In addition, the dramatic amelioration induced by IL-1Ra treatment is at least partly due to the strong decrease in IL-1beta secretion that follows the first injections of the antagonist. These findings may have implications for other chronic inflammatory conditions characterized by increased IL-1beta.     

The PNH phenotype cells that emerge in most patients after CAMPATH-1H therapy are present prior to treatment

Paroxysmal nocturnal haemoglobinuria (PNH) cells are deficient in glycosylphosphatidylinositol (GPI) linked antigens due to a somatic mutation of the PIG-A gene in a haemopoietic stem cell. It appears that a PNH clone reaches detectable proportions only when there is selection in its favour. GPI-deficient T lymphocytes have been identified in patients treated with CAMPATH-1H, a monoclonal antibody against the GPI-linked CD52 molecule. CAMPATH-1H selects for cells that are deficient in CD52 (such as PNH-like cells) promoting the development of a PNH-like clone (analogous to PNH). We report that 10/15 patients with chronic lymphocytic leukaemia developed PNH-like lymphocytes after therapy with CAMPATH-1H. The remaining five patients developed no PNH-like cells at any stage, including one patient who received 12 weeks of therapy. The inactivating PIG-A mutation has been identified in one patient. This mutation was detectable by an extremely sensitive mutation-specific PCR-based analysis in the patient's mononuclear cells prior to CAMPATH-1H therapy. The frequency and phenotype of GPI-deficient lymphocytes after CAMPATH-1H and the detection of a PIG-A mutation in the lymphocytes prior to CAMPATH-1H therapy indicated that such mutations were present in a very small proportion of cells prior to selection in their favour by CAMPATH-1H. This suggests that a large proportion of individuals have cells with PIG-A mutations that are not detectable by flow cytometry and thus may have the potential to develop PNH.     

The human and simian immunodeficiency viruses HIV-1, HIV-2 and SIV interact with similar epitopes on their cellular receptor, the CD4 molecule

The cellular receptor for HIV-1 is the leucocyte differentiation antigen, CD4. Blocking of HIV-1 infectivity can be achieved with monoclonal antibodies (MAbs) to some, but not all epitopes of this antigen. We demonstrate here, by inhibition of virus infection, blocking of syncytium formation and inhibition of pseudotype infection with a panel of CD4 MAbs, that HIV-1, HIV-2 and simian immunodeficiency virus (SIV) isolates share the same cellular receptor, the CD4 glycoprotein. It is also shown that very similar epitopes of this molecule are involved in virus binding. We infer from these data that the binding sites on these viruses are highly conserved regions, and may therefore make good targets for potential vaccines. In addition, we show that cell surface expression of CD4 is similarly modulated after infection of cell lines by all the viruses.     

Dynamics of drug-resistant HIV-1 in plasma and peripheral blood cells in patients during and after enfuvirtide therapy

Genetic analysis of HIV-1 gp41 in plasma and peripheral blood mononuclear cell (PBMC) compartments was performed longitudinally in 10 HIV-infected patients treated with enfuvirtide. The appearance of enfuvirtide resistance mutations in PBMCs (DNA) generally occurred after (from 23 to 157 weeks) being recognizable in plasma (RNA). The disappearance of drug-resistant HIV-1 mutations to enfuvirtide in seven patients who discontinued the drug was directly dependent of the exposure time failing enfuvirtide, being associated mainly with the selection of secondary/compensatory mutations recognizable in proviral DNA.     

HIV-1 alters T helper cytokines, interleukin-12 and interleukin-18 responses to the protozoan parasite Leishmania donovani

OBJECTIVE: To investigate the in vitro and in vivo effect of HIV-1 on lymphoproliferative and T helper (Th) cytokine responses in leishmaniasis. METHODS: Th1 [interleukin (IL)-2 and interferon (IFN)-gamma] and Th2 (IL-4 and IL-10) as well as IFN-gamma-inducing cytokines (IL-12 and IL-18) were measured in antigen and mitogen-stimulated culture supernatants of peripheral blood mononuclear cells (PBMC) of healthy donors, HIV-infected and visceral leishmaniasis (VL) patients with or without HIV co-infection. RESULTS: Proliferative responses to phytohaemagglutinin (PHA) were significantly lower in PBMC from VL and asymptomatic HIV-infected persons compared with responses in healthy individuals. VL-HIV co-infected patients showed the lowest responses. Although there was no significant difference in the Leishmania-induced proliferative responses among the healthy group and those infected with HIV only, VL patients (with or without HIV) exhibited very low proliferation. When cultured with PHA or Leishmania, PBMC from healthy donors produced high levels of a Th1 cytokine (IFN-gamma) and low levels of Th2 cytokines (IL-4 and IL-10). In addition, co-culturing PBMC from healthy donors with a killed HIV preparation abrogated the production of IFN-gamma induced by Leishmania and augmented IL-4 and IL-10 production. Cells from HIV-infected patients produced low levels of IFN-gamma, but high levels of IL-10. The addition of anti-IL-10 did not increase Leishmania-induced proliferative responses or IFN-gamma production. Both IL-12 and/or IL-18 responses were lower in VL patients, HIV-infected, or VL-HIV co-infected patients as compared with those of healthy donors. CONCLUSION: The data suggest that the inhibitory effect of HIV and VL on proliferation and IFN-gamma production is not due to IL-10 alone, but that the defect induced by HIV and VL probably operates at the level of regulation of IFN-gamma-inducing factors, such as IL-12 and IL-18.     

Morbidity in HIV-1-infected individuals before and after the introduction of antiretroviral therapy: a longitudinal study of a population-based cohort in Uganda

Background

We compared morbidities in HIV‐1‐infected patients before and after the introduction of antiretroviral therapy (ART) in a rural Ugandan cohort followed from 1990 to 2008. ART was introduced in 2004.

Methods

Random‐effects Poisson regression models were used to estimate incidence rates of World Health Organization (WHO) stage‐defining diseases in HIV‐infected individuals aged 13 years or older with known seroconversion dates, and in an age‐stratified sample of HIV‐negative individuals.

Results

The most common morbid event was bacterial pneumonia, with an incidence of 7.4/100 person‐years (pyr) among 309 HIV seroconverters and 1.3/100 pyr among 348 HIV‐negative participants [hazard ratio (HR) 5.64; 95% confidence interval (CI) 3.6–8.8]. Among seroconverters, the incidence of the acquisition of any WHO stage‐defining disease rose from 14.4/100 pyr (95% CI 11.1–18.6) in 1990–1998 to 46.0/100 pyr (95% CI 37.7–56.0) in 1999–2003. Following the introduction of ART, the incidence among seroconverters declined to 36.4/100 pyr (95% CI 27.1–48.9) in 2004–2005 and to 28.3/100 pyr (95% CI 21.2–37.8) in 2006–2008. At the individual level, a higher rate of acquiring any WHO stage‐defining disease was independently associated with lower CD4 cell count, longer duration of HIV infection and older age. In addition, individuals who had been on ART for longer than 12 months had a substantially lower rate of any WHO stage disease than those not yet on ART (adjusted HR 0.35; 95% CI 0.2–0.6).

Conclusion

Morbidity in HIV‐positive participants decreased following the introduction of ART, and this decline was more marked with increasing duration on ART. The benefits of decreased HIV‐related morbidity from ART lend support to urgent efforts to ensure universal access to early diagnosis of HIV infection and to ART, especially in rural Africa.     

Both baseline HIV-1 drug resistance and antiretroviral drug levels are associated with short-term virologic responses to salvage therapy

BACKGROUND: To determine the impact of HIV-1 drug resistance at baseline and antiretroviral drug levels (DL) during follow-up on virologic response to the next antiretroviral regimen. METHODS: Baseline genotypic and phenotypic susceptibility was obtained for plasma virus from patients failing a protease inhibitor-containing regimen. Untimed plasma antiretroviral DL were performed and the distribution of DL after 12 weeks of follow-up was classified as above (DLHigh) or below (DLLow) the median. Inhibitory quotients [IQ = (DL at week 12)/(fold change in IC50 to wild-type)] were determined for each drug in the regimen. Primary outcome was change in log10 plasma HIV-1 RNA viral load (DeltaVL) from baseline to 12 weeks. RESULTS: There were 137 patients who had baseline resistance data available for the antiretroviral drugs used in the salvage regimen, and DL at week 12. Each drug with DLHigh was associated with DeltaVL = -0.40 (P = 0.0002) while each drug with DLLow had DeltaVL = -0.16 (P = 0.11). In multivariate models DeltaVL associated with each active drug (defined by genotype) with DLHigh was -0.48 log10 (P < 0.0001), and with each active drug with DLLow was -0.22 (P = 0.03). The DeltaVL was -0.18 if no drugs in the regimen had an IQ > median, compared to -0.58 for one drug, -1.06 for two drugs, -0.86 for three drugs, and -1.44 for four or five drugs with IQ > median (P < 0.0001 for trend). CONCLUSIONS: In salvage therapy, both the number of active drugs and the DL for each drug in the new regimen determine the antiviral response.     

Diminished in vitro production of interleukin-1 and tumor necrosis factor-alpha during acute visceral leishmaniasis and recovery after therapy.

Disturbance of T cell-mediated immunity has been reported in acute visceral leishmaniasis (AVL). In a study of 16 patients with AVL, defective production of interleukin-1 (IL-1) by peripheral blood mononuclear cells was demonstrated in response to leishmania antigens, heat-killed Listeria organisms, and lipopolysaccharide when compared to posttherapy values or controls. This global defect in IL-1 production was corrected after successful therapy. Twelve of 16 patients responded with a greater than or equal to 2.5-fold increase in IL-1 production that correlated with clinical cure, P less than .01. Depressed production of tumor necrosis factor (TNF) was leishmania antigen-specific and similarly recovered after therapy. In vitro TNF production during the follow-up period did not correlate with clinical status but high serum levels were associated with AVL. Since T cells are activated by processed antigens presented on class II major histocompatibility molecules and by newly synthesized IL-1, defective IL-1 production may contribute to the immunosuppression observed in AVL.     

Decreased exposure to saquinavir in HIV-1-infected patients after long-term antiretroviral therapy including ritonavir and saquinavir

OBJECTIVE: To explore whether steady-state plasma pharmacokinetics of ritonavir and saquinavir change during long-term treatment in HIV-1-infected patients on antiretroviral treatment including ritonavir and saquinavir. METHODS: The pharmacokinetics of ritonavir and saquinavir were assessed during an 8-h period on two occasions in six HIV-1 infected patients on stable twice daily treatment with ritonavir 400 mg, saquinavir 400 mg and stavudine 40 mg with or without lamivudine 150 mg twice daily. RESULTS: The first study day was 4-12 months (median 7 months) after the start of the current regimen. The second study day was 9-15 months (median 10 months) later. No significant differences were observed for the ritonavir pharmacokinetics between the first and second study day. However, median change in plasma trough level of saquinavir between the two study days was -30% (range -79 to +11%; P = 0.06). Median change in maximum plasma concentration was -40% (range -62 to +34%; P = 0.09). The median change in area under the plasma concentration versus time curve over 0-8 h was -33% (range -53 to +21%; P = 0.06). CONCLUSION: The exposure to saquinavir decreased over time in HIV-infected patients on stable antiretroviral therapy. These data suggest that regular monitoring of plasma drug concentrations should become part of routine patient care even in apparently compliant patients.     

Sexual risk behaviour relates to the virological and immunological improvements during highly active antiretroviral therapy in HIV-1 infection

OBJECTIVES: To evaluate the effect of highly active antiretroviral therapy (HAART) on the sexual behaviour of homosexual men, we conducted (i) an ecological study of time trends in sexual behaviour and sexually transmitted diseases; (ii) a HAART-effect study focused on the practice of unprotected anogenital sex. DESIGN: Subjects were participants in the ongoing Amsterdam Cohort Studies (ACS) among homosexual men, initiated in 1984. Data for (i) represented all ACS visits by HIV-1-positive and -negative participants who entered ACS at or below 30 years of age and were followed until 35 years (n = 1062). Data for (ii) represented all ACS visits of HIV-1-positive men from 1992 to 2000 (n = 365), of whom 84 were HAART recipients with at least 2 months of behavioural follow-up. RESULTS: (i) After HAART became generally available in July 1996, unprotected sex was practised more frequently and the incidence of gonorrhoea was higher compared to March 1992-June 1996 among HIV-1-negative and -positive men, respectively. (ii) Among HIV-1-positive men, a higher level of unprotected sex with casual partners was observed after HIV-1 RNA became undetectable and CD4 cell counts increased with the use of HAART. Notably, in individuals who did not receive HAART, high HIV-1-RNA levels (above 10(5) copies/ml) were likewise related to unprotected sex with casual partners. CONCLUSION: Data support the need for the reinforcement of safe sex prevention messages among HIV-1-negative men, and our data also provide a lead for redirecting and tailoring current prevention strategies to the needs of HIV-1-positive men.     

Contribution of interleukin-1 to activation of coagulation and fibrinolysis, neutrophil degranulation, and the release of secretory- type phospholipase A2 in sepsis: studies in nonhuman primates after interleukin-1 alpha administration and during lethal bacteremia

Although studies with interleukin-1 receptor antagonist (IL-1ra) in animal models have shown that IL-1 contributes to mortality in sepsis, the mechanisms whereby IL-1 mediates lethal effects are not well established. A possible mechanism is that IL-1 enhances the activation and release of other inflammatory mediator systems such as coagulation, fibrinolysis, neutrophils, and secretory-type phospholipase A2 (sPLA2). We investigated this possibility by assessing the effect of intravenously injected recombinant human IL-1 alpha (rhIL-1 alpha) on these plasma parameters in baboons. In addition, we examined the course of these inflammatory parameters in baboons after a challenge with a lethal dose of Escherichia coli and while receiving a 24-hour constant infusion of IL-1ra or placebo. Intravenous administration of IL-1 alpha (10 micrograms/kg) induced the formation of thrombin, as evidenced by the appearance of thrombin-antithrombin III (TAT) complexes into the circulation (peak levels, 188 +/- 92 ng/mL at 2 hours), as well as the activation of fibrinolysis, assessed by circulating plasmin-alpha 2- antiplasmin complexes (PAP complexes; peak levels, 0.4% +/- 0.03% of fully activated plasma at 1 hour), the release of tissue-type plasminogen activator (t-PA; peak levels, 6 +/- 2 ng/mL at 2 hours), and its inhibitor, plasminogen activator inhibitor (PAI; peak levels, 724 +/- 246 ng/mL at 4 hours). Il-1 alpha administration also induced the release of sPLA2 (maximal levels, 336 +/- 185 ng/mL at 8 hours), but not degranulation of neutrophils. In the septic baboons, a significant reduction of the formation of thrombin (peak TAT levels decreased from 582 +/- 78 ng/mL to 219 +/- 106 ng/mL; P < .005), the release of t-PA (peak levels decreased from 37 +/- 11 ng/mL to 17 +/- 2 ng/mL; P < .001), and its inhibitor, PAI (peak levels decreased from 2,639 +/- 974 ng/mL to 1,110 +/- 153 ng/mL; P <.001), was observed in the group receiving IL-1ra compared to that receiving placebo. The release of neutrophilic elastase was also significantly attenuated in IL-1a-treated animals (peak levels, 1,024 +/- 393 and 655 +/- 104 ng/mL in control and treatment groups, respectively; P < .05). The difference between sPLA2 levels in both groups, although higher in the controls (maximal levels, 3,140 +/- 1,435 ng/mL in control v 2,217 +/- 1,375 ng/mL in IL-1ra-treated group), was not significant. Thus, IL-1 contributes to activation of various other mediator systems in severe sepsis in nonhuman primates. We propose that these effects may explain the lethal actions of IL-1 in this sepsis model and suggest a similar role for IL-1 in severe human sepsis.     

Influence of RANTES, SDF-1 and TGF-beta levels on the value of interleukin-7 as a predictor of virological response in HIV-1-infected patients receiving double boosted protease inhibitor-based therapy

OBJECTIVES: Interleukin-7 (IL-7), RANTES (regulated on activation, normal T cell expressed and secreted), stromal cell-derived factor-1 (SDF-1) and transforming growth factor-beta (TGF-beta) appear to share certain biological properties in vitro and all are involved in HIV-1 disease progression. Our earlier observations indicated that IL-7 levels decrease upon CD4 T-cell recovery and represent a new, independent predictor of virological response. Here, we examine associations among circulating levels of IL-7, RANTES, SDF-1 and TGF-beta in hopes of gaining insight into their contribution to the predictive value of IL-7. METHODS: Levels of IL-7, RANTES, SDF-1 and TGF-beta, and immune and viral parameters were assessed in HIV-1-infected patients. RESULTS: Cross-sectional (n=148) and longitudinal (n=36) analyses showed that levels of IL-7, but not RANTES, SDF-1 or TGF-beta, were increased in HIV-1-infected adults compared with those of healthy controls. In the cross-sectional study, levels of IL-7 were correlated with RANTES (r=0.31, P=0.002) and TGF-beta (r=0.53, P<0.001) but not with SDF-1 (r=0.12, P=0.22), and these associations were more pronounced in patients with CD4 T-cell counts >200 cells/microL. In contrast to IL-7, levels of RANTES, SDF-1 and TGF-beta were not correlated with CD4 T-cell counts. Longitudinal analysis revealed a marked decline in IL-7 levels accompanied by an increase in CD4 T-cell count following antiretroviral therapy (ART), but no changes in RANTES, SDF-1 or TGF-beta levels. Multivariate regression analysis showed no influence of baseline RANTES, SDF-1 or TGF-beta levels on the value of IL-7 as a predictor of virological response at 48 weeks. CONCLUSIONS: Collectively, these results indicate that changes in IL-7 levels did not induce changes in RANTES, SDF-1 or TGF-beta. Furthermore, they indicate that RANTES, SDF-1 or TGF-beta levels do not explain the predictor value of IL-7 in patients receiving ART.