Associations of cytokines, sleep patterns, and neurocognitive function in youth with HIV infection.

Youth infected with HIV at birth often have sleep disturbances, neurocognitive deficits, and abnormal psychosocial function which are associated with and possibly resulted from elevated blood cytokine levels that may lead to a decreased quality of life. To identify molecular pathways that might be associated with these disorders, we evaluated 38 HIV-infected and 35 uninfected subjects over 18-months for intracellular cytokine levels, sleep patterns and duration of sleep, and neurodevelopmental abilities. HIV infection was significantly associated with alterations of intracellular pro-inflammatory cytokines (TNF-α, IFN-γ, IL-12), sleep factors (total time asleep and daytime sleep patterns), and neurocognitive factors (parent and patient reported problems with socio-emotional, behavioral, and executive functions; working memory-mental fatigue; verbal memory; and sustained concentration and vigilance. By better defining the relationships between HIV infection, sleep disturbances, and poor psychosocial behavior and neurocognition, it may be possible to provide targeted pharmacologic and procedural interventions to improve these debilitating conditions.

Impaired restoration of plasmacytoid dendritic cells in HIV-1-infected patients with poor CD4 T cell reconstitution is associated with decrease in capacity to produce IFN-alpha but not proinflammatory cytokines.

We analyzed reconstitution characteristics of plasmacytoid dendritic cells (PDCs) and myeloid DCs-1 in 38 HIV-1-infected patients with impaired restoration of CD4 T cell counts despite prolonged suppression of plasma viremia (discordant) and compared them with 42 patients showing good immunological and virological responses following highly active antiretroviral therapy (HAART). While myeloid DCs showed spontaneous recovery following HAART in both the groups, the discordant patients demonstrated poor peripheral reconstitution of PDCs as compared with concordant patients. The ability of PDCs to produce IFN-alpha following stimulation with TLR7 ligand imiquimod and TLR9 ligand CpG ODN-2216 was also impaired in discordant patients even after 2 years following initiation of HAART. Lower IFN-alpha expression in the PDCs following TLR stimulation was further associated with lower expression of transcription factor, IFN regulatory factor-7. In contrast, production of TNF-alpha and IL-6 following TLR stimulation was comparable in both groups of patients, indicating that impaired reconstitution characteristics do not affect the capacity of PDCs to produce proinflammatory cytokines. The discordant patients had significantly lower baseline CD4 T cell counts and higher baseline viral load at the initiation of HAART implying that lower baseline CD4 T cell counts and higher plasma viral load are associated with impaired restoration of CD4 T cells and PDCs, thus, increasing the susceptibility of discordant patients toward opportunistic infections despite virological control.

The influence of different types of alcoholic beverages on disrupting highly active antiretroviral treatment (HAART) outcome.

AIMS: Studies have yielded conflicting results regarding alcohol's influence on HIV outcomes, particularly after highly active antiretroviral treatment (HAART). Discrepant findings may be related to confounding variables, including gender, patterns of alcohol abuse and type of alcohol beverage beyond the amount consumed. METHODS: Using a cohort study, differences in HAART effectiveness after 24 weeks of therapy were compared as a function of amount and preference for alcohol, drinking only liquor (LI, n = 55) or only wine or beer (BW, n = 110). Given the critical role of thymus on HAART response, changes in thymus size, CD4s, naïve lymphocytes and viral loads were assessed. RESULTS: After HAART, positive increases in both CD4s (+12 cell counts/mm(3)) and thymus size (+0.7 mm(3)) were evident in the BW group. In contrast, the LI subgroup exhibited a decline in both parameters (-4 CD4 cells/mm(3) and -0.6 mm(3) in thymus size). Women in the LI group exhibited significantly lower CD4 (163.4 +/- 46.2) and naïve counts (178 +/- 69.5) than LI men (CD4: 281.6 +/- 203, P = 0.05; lymphocytes: 301.4 +/- 198, P = 0.04). In adjusted regression models, the LI compared to the BW subgroup had greater odds of maintaining detectable viral loads (RR = 1.35, 95% CI 1.04-1.75; P = 0.03), increased thymus volumes (RR = 3.8, P = 0.04) and replenished naïve cells (RR = 13, P = 0.02). CONCLUSIONS: Liquor was associated with thymus deterioration and thus with poorer viro-immune outcomes after HAART. Subtyping participants by alcohol consumption patterns seems to be clinically relevant and needs to be accounted for in future studies.

Cytokine quantitation: technologies and applications.

Assessment of cytokines in body fluids, cells or tissues provides important information in understanding of disease process and designing treatment strategies. Today, wide range of cytokine assays are available, including; measurement of levels of cytokines (direct) or cytokine soluble receptor levels (indirect) in body fluids or cellular supernatants (immunoassays and cytokine bioassays), measurement of cytokines produced by population of cells (multiparametric flow cytometry, magnetic beads based quantitation of cytokine producing cells, mRNA based assays), measurement of cytokines produced by single cells (Enzyme-linked immunospot (ELISPOT), Intra-cytoplasmic cytokine staining (ICC), mRNA based assays) and detection of cytokines in tissues (immunostaining). Improved understanding of cytokine interactions has led to a consensus that simultaneous assessment of many cytokines in a biological sample provides more comprehensive information rather than assessing a single cytokine. Thus, technologies that measure one cytokine at a time are being gradually replaced by multiplex-type formats (DNA and protein microarrays).

Increase in frequencies of circulating Th-17 cells correlates with microbial translocation, immune activation and exhaustion in HIV-1 infected patients with poor CD4 T-cell reconstitution.

We analyzed the association of circulating Th-17 cells (cTh-17) with immune activation (IA), immune exhaustion (IE) and regulatory T-cells (T-regs) in 20 human immunodeficiency virus-1 (HIV-1) infected patients with impaired restoration of CD4 T-cell counts despite prolonged suppression of plasma viremia (discordant) and compared it with 20 HIV-1 infected patients showing good immunologic and virologic responses (concordant) following highly active antiretroviral therapy (HAART). Discordant HIV-1 infected patients showed significantly higher frequencies of cTh-17 cells compared to concordant patients and healthy controls after PMA+Ionomicin stimulation. Discordant patients also showed higher CD4 T-cell immune activation (HLA-DR+CD38+) than concordant patients which directly correlated with microbial translocation. Additionally, CD4 T-cells of discordant patients showed higher frequencies of CD4 T-cells expressing multiple immune exhaustion markers (Tim3+PD-1+) which correlated with immune activation indicating that combined analysis of inhibitory molecules along with PD-1 might be a better predictor for immune exhaustion of CD4 T-cells. Increased cTh-17 cell frequency correlated inversely with CD4 T-cell percentages and absolute counts and directly with CD4 T-cell immune activation and T-reg frequencies. Persistent CD4 T-cell immune activation might favor differentiation of activated CD4 T-cells toward cTh-17 phenotype in discordant patients. Discordant patients had significantly lower baseline CD4 T-cell counts and higher viral load at the initiation of HAART and higher immune activation and immune exhaustion after being on HAART for long time indicating that these factors might be associated with an increase in cTh-17 cell frequency, thus, increasing the risk of disease progression despite virologic control.

Older age and plasma viral load in HIV-1 infection.

BACKGROUND: The purpose of the study was to examine the relationship between age and plasma viral load in HIV-1-infected individuals. DESIGN: The experimental method was to recruit older (> 50 years of age) and younger (18-39 years of age) HIV-1-infected individuals. The plasma viral load was measured using the Roche Molecular Systems UltraSensitive Roche HIV-1 Monitor test reflexively with the standard Amplicor HIV Monitor test to quantify viral load in the range of 50-750,000 copies of HIV-1 RNA/ml plasma. SUBJECTS: A total of 135 HIV-1-seropositive individuals (at Centers for Disease Control and Prevention early symptomatic stage B or late symptomatic stage/AIDS C) were enrolled as part of a larger cohort also consisting of HIV-1-seronegative individuals. RESULTS: A generalized linear models statistical analysis was conducted in order to evaluate age category as a predictor of plasma viral load. The result was a significant effect of age category, with older age associated with a lower plasma viral load. The association held controlling for antiretroviral therapy usage, disease stage, antiretroviral medication adherence, HIV-1 serostatus duration, alcohol and substance use, recent sexually transmitted disease, and sociodemographics (except income). CONCLUSION: Older age was associated with lower levels of HIV-1 replication in this sample, independent of antiretroviral therapy usage, regimen adherence, and disease stage. It is suggested that the effect may be caused by changes in viral evolution or immunological monitoring specific to older individuals with HIV-1 infection.

Reference ranges of lymphocyte subsets in healthy adults and adolescents with special mention of T cell maturation subsets in adults of South Florida.

BACKGROUND: Analysis of peripheral blood lymphocyte subsets has become an essential tool in the evaluation of outcome of diagnostic and research related questions in immunological and pathological conditions. Periodic evaluation and establishment of normal lymphocyte reference ranges are required in clinical and research settings of various immunodeficiency disorders for evaluation of the significance of observations. It is also important that age and gender specific lymphocyte subset reference ranges should be locally established for meaningful comparison and accurate result interpretation as age plays a significant role in the development of immune system. METHODS: We performed dual platform flow cytometry to determine reference ranges for lymphocyte subsets (CD3, CD4, CD8, CD19 [B cells] and CD16+CD56+ [Natural Killer - NK cells]) in 50 adolescents (age range: 12-18) and 100 adults (age range: 21-67) along with T cell maturation, activation and co-stimulatory molecules in healthy multiracial adult population of South Florida. RESULTS: The lymphocyte reference ranges percentages [absolute counts - Abs, cells/µl] unadjusted for gender differences for adolescents are: CD3: 49-83 [939-2959]; CD4: 27-53 [467-1563]; CD8: 16-40 [259-1262]; CD19+ B cells: 8-31 [169-1297] and CD16+CD56+ NK cells: 3-30 [59-1178] and for adults are: CD3: 65-88 [983-3572]; CD4: 26-62 [491-2000]; CD8: 14-44 [314-2,087]; CD19+ B cells: 2-27 [64-800] and CD16+CD56+ NK cells: 2-27 [27-693]. The ranges for CD4:CD8 ratio for adolescents and adults are 0.7-2.6 and 0.6-4.4, respectively. Gender based analysis of relative percentages of lymphocyte subsets showed no significant differences between adult and adolescent males and females. The mean CD4:CD8 ratio was significantly higher in adult females than males (P=0.04) and in adolescents this difference was not significant between genders. The mean CD3 and CD4 T cell percentages were higher and CD19 cell percentages were lower in adults compared to adolescents (P<0.0001). Absolute lymphocyte counts showed a positive correlation with the absolute counts of CD3+, CD4+, CD8+, CD19+, CD16+CD56+, CD45RO+ and CD45RA+ cells (all correlations with P<0.0001 except CD45RO [P=0.01] and CD45RA [P=0.03]). CONCLUSION: The reference values of peripheral blood lymphocyte subsets were analyzed in healthy adolescent and adult population of South Florida. This study indicates the need for periodic evaluation and establishment of lymphocyte reference ranges for patient population served based on gender and age since these could influence immune status and treatment outcome.

Tobacco smoking increases immune activation and impairs T-cell function in HIV infected patients on antiretrovirals: a cross-sectional pilot study.

BACKGROUND: The influence of tobacco smoking on the immune system of HIV infected individuals is largely unknown. We investigated the impact of tobacco smoking on immune activation, microbial translocation, immune exhaustion and T-cell function in HIV infected individuals. METHOD: HIV infected smokers and non-smokers (n = 25 each) with documented viral suppression on combination antiretroviral therapy and HIV uninfected smokers and non-smokers (n = 15 each) were enrolled. Markers of immune activation (CD38 and HLA-DR) and immune exhaustion (PD1, Tim3 and CTLA4) were analyzed in peripheral blood mononuclear cells (PBMCs) by flow cytometry. Plasma markers of microbial translocation (soluble-CD14 - sCD14 and lipopolysaccharide - LPS) were measured. Antigen specific functions of CD4+ and CD8+ T-cells were measured, by flow cytometry, in PBMCs after 6 hours stimulation with Cytomegalovirus, Epstein-Barr virus and Influenza Virus (CEF) peptide pool. RESULTS: Compared to non-smokers, smokers of HIV infected and uninfected groups showed significantly higher CD4+ and CD8+ T-cell activation with increased frequencies of CD38+HLA-DR+ cells with a higher magnitude in HIV infected smokers. Expressions of immune exhaustion markers (PD1, Tim3 and CTLA4) either alone or in combinations were significantly higher in smokers, especially on CD4+ T-cells. Compared to HIV uninfected non-smokers, microbial translocation (sCD14 and LPS) was higher in smokers of both groups and directly correlated with CD4+ and CD8+ T-cell activation. Antigen specific T-cell function showed significantly lower cytokine response of CD4+ and CD8+ T-cells to CEF peptide-pool stimulation in smokers of both HIV infected and uninfected groups. CONCLUSIONS: Our results suggest that smoking and HIV infection independently influence T-cell immune activation and function and together they present the worst immune profile. Since smoking is widespread among HIV infected individuals, studies are warranted to further evaluate the cumulative effect of smoking on impairment of the immune system and accelerated disease progression.

Gender differences in the association of hazardous alcohol use with hypertension in an urban cohort of people living with HIV in South Florida.

OBJECTIVE: Industrialized countries are currently experiencing an epidemic of high blood pressure (HBP) extending to people living with HIV (PLWH). Given the prevalence of hazardous alcohol use (HAU), this study examines the relationship between alcohol consumption and hypertension in PLWH. Including a gender analysis is critical, given the high rates of HAU and HIV among females. METHOD: We followed PLWH including both HAU and non-HAU (200 each). Participants were assessed twice for body weight, blood pressure, alcohol consumption, and other BP-associated lifestyle factors. High blood pressure (defined as systolic/diastolic blood pressure above 140/90 mmHg and/or treatment of HBP) was the primary outcome. RESULTS: Overall prevalence of hypertension was 38% and higher among HAU compared to non-HAU (42% vs. 34%, p = 0.02). Less than half with HBP (42%) were receiving treatment for hypertension. Overall, males had a 50% higher risk of HBP than women (odds ratio: 1.5, 95% CI: 1-2.6, p = 0.05). However among HAU, females were twice as likely to suffer HBP as their male counterparts (95% CI: 1-3.9, p = 0.02). Those HAU who preferred liquor, versus wine, had higher adjusted mean BP (132.6±18 vs. 122.3±14 mm Hg, p = 0.05). Additional analyses indicated that consumption of >1 standard drink of liquor or beer/day was associated with HBP. Risk of hypertension was noted in those with daily consumption of >3 glasses of wine. For those reporting <1 drink per day, the odds ratio of having HBP was 0.97 (CI: 0.6-0.99, p = 0.05). Factors associated with hypertension in the multivariate model included increased age, gender, BMI, HAU particularly of liquor, and smoking. CONCLUSIONS: Excessive hypertension burden in this population and its association with HAU and sub-optimal care indicate the need for preventive and educational intervention in PLWH. Analyses highlight the necessity of gender and type-of-beverage specific approaches.

Beyond the Brain: The Role of Brain-Derived Neurotrophic Factor in Viroimmune Responses to Antiretroviral Therapy among People Living with HIV with and without Alcohol Use.

OBJECTIVE: Given the emerging data suggesting the key role of brain-derived neurotrophic factor (BDNF) in the immune system, we assessed longitudinally whether BDNF depletions induced by hazardous alcohol use (HAU) would impact a response to antiretroviral therapy (ART). METHODS: In a prospective single-site cohort, virological and immunological responses to ART in 200 hazardous and 200 nonhazardous users were obtained, along with plasma BDNF levels. RESULTS: Hazardous drinkers were more likely to have BDNF levels <4000 pg/mL (odds ratio [OR] = 1.6, P = .01). Participants with BDNF <4000 pg/mL were less likely to have CD4 counts of more than 500 cells/mm(3) (P = .02) and to achieve viral suppression over the follow-up period (OR = 1.5, P = .03). Multivariate analysis confirmed the significant role of HAU and low BDNF in predicting viroimmune responses. CONCLUSION: Hazardous alcohol use was associated with BDNF alterations, which in turn were linked to a limited response to ART in terms of viral suppression and CD4 count improvements.

Poor lymphoproliferative responses with low proportion of Gag-specific CD8 TEMRA cells in HIV-1-infected patients showing immunological and virological discordance despite prolonged suppression of plasma viremia.

We investigated in-vitro lymphoproliferative responses and T-cell subsets in 38 HIV-1-infected patients showing impaired restoration of CD4 T cells despite prolonged viral suppression (discordant), and in 42 HIV-1-infected patients showing positive immunological and virological responses to highly active antiretroviral therapy (HAART) (concordant). In comparison to concordant patients, discordant patients showed poor lymphocyte proliferation, lower secretion of IL-2 and IFN-gamma, a lower percentage of perforin and granzyme-B-producing CD8 T cells, and poor differentiation of effector memory CD8 T(EM) cells into CD8 T(EMRA) cells in in-vitro stimulation assays, especially against HIV-1 Gag p24 and one of its peptide pools. Functional CD8 T-cell responses of discordant patients after stimulation with recall antigens, Candida albicans, and tetanus toxoid, were also inferior to concordant patients, but comparable to normal healthy controls. Examination of the multifunctional roles of T cells is imperative in describing the overall magnitude of immune responses to HIV-1. Our results suggest that prolonged suppression of plasma viremia alone does not warrant good qualitative and quantitative CD8 T-cell responses to HIV-1, implying that CD4 T cells are required for maintenance of protective CD8 T-cell responses.

Low cholesterol? Don't brag yet ... hypocholesterolemia blunts HAART effectiveness: a longitudinal study.

BACKGROUND: In vitro studies suggest that reducing cholesterol inhibits HIV replication. However, this effect may not hold in vivo, where other factors, such as cholesterol's immunomodulatory properties, may interact. METHODS: Fasting blood samples were obtained on 165 people living with HIV at baseline and after 24 weeks on highly active antiretroviral therapy (HAART). Participants were classified as hypocholesterolemic (HypoCHL; <150 mg/dl) or non-HypoCHL (>150 mg/dl) and were compared on viro-immune outcomes. RESULTS: At baseline, participants with HypoCHL (40%) exhibited lower CD4 (197 +/- 181 vs. 295 +/- 191 cells/mm3, p = 0.02) and CD8 (823 +/- 448 vs. 1194 +/- 598 cells/mm3, p = 0.001) counts and were more likely to have detectable viral loads (OR = 3.5, p = 0.01) than non-HypoCHL controls. After HAART, participants with HypoCHL were twice as likely to experience a virological failure >400 copies (95% CI 1-2.6, p = 0.05) and to exhibit <200 CD4 (95% CI 1.03-2.9, p = 0.04) compared with non-HypoCHL. Low thymic output was related to poorer CD4 cell response in HypoCHL subjects. Analyses suggest a dose-response relationship with every increase of 50 mg/dl in cholesterol related to a parallel rise of 50 CD4 cells. CONCLUSIONS: The study implicates, for the first time, HypoCHL with impaired HAART effectiveness, including limited CD4 repletion by the thymus and suboptimal viral clearance.

A randomized controlled trial of Structural Ecosystems Therapy for HIV medication adherence and substance abuse relapse prevention.

BACKGROUND: Substance abuse in women with HIV/AIDS overshadows other priorities, including health care. Substance abuse may cause women to avoid health care systems and not adhere to their medication regimen. METHODS: A randomized controlled trial tested the efficacy of Structural Ecosystems Therapy (SET) relative to a psychoeducational Health Group (HG) in 126 HIV+ women in recovery. SET, a 4-month intervention, focused on building family support for relapse prevention and HIV medication adherence. Over 12-month follow-up, women were assessed for drug use and medication adherence every 2 months; CD4 T-cell count and HIV viral load were assessed every 4 months. RESULTS: Levels of drug use did not differ by condition. There was a significant difference in curvature of the rates of change in drug use with SET increasing and then decreasing and HG decreasing and then increasing. Women in SET were more likely to increase substance abuse services in response to relapse and separate from drug using household members than were women in HG. These two changes explained the decline in drug use observed within SET between 6 and 12 months. SET showed declines in medication adherence but increases in CD4 T-cell count relative to HG. The increase in CD4 T-cell count in SET was related to increasing proportions of women in SET taking antiretroviral medications. CONCLUSION: The results of the trial were mixed. Women in SET did not show better drug use or medication adherence outcomes, but did show improvement in CD4 T-cell count and theoretical mechanisms of action on drug relapse.

Analysis of immunological markers associated with pregnancy and HIV-1 infection: relevance in perinatal transmission in HIV-1-infected pregnant women with low plasma viral load.

PROBLEM: In HIV-1-infected pregnant women with low plasma viral load, risk factors associated with perinatal HIV-1 transmission are not clearly understood. METHOD OF STUDY: We analyzed distribution of peripheral CD8 T-cell subsets, plasma cytokines and measured secretory leukocyte peptidase inhibitor (SLPI) and myeloid-related protein (MRP)-8 levels in whole-blood and cervico-vaginal fluid (CVF) specimens obtained from 35 HIV-1-infected pregnant women (group 1), 12 HIV-1-infected non-pregnant women (group 2) and 15 HIV-1 uninfected pregnant women (group 3). RESULTS: The group 1 women had higher expression of CD38, human leukocyte antigen-DR and CD95 on CD8 T-cells and higher levels of plasma tumor necrosis factor-alpha and epidermal growth factor. CVF-SLPI levels were the highest in group-3, while MRP-8 levels were the highest in group 1 women in plasma and CVF (P < 0.01). Although there were no cases of perinatal HIV-1 transmission, group 1 women undergoing HIV-1-indicated cesarean section had lower levels of CVF-SLPI as compared with those undergoing normal vaginal delivery. CONCLUSION: Pregnancy contributes to the activation of peripheral CD8 T cells and increase in pro-inflammatory cytokines. Production of protective mucosal secretory factors such as SLPI is affected by HIV-1 infection in pregnant women and down-regulated SLPI levels may indirectly indicate a higher possibility of perinatal HIV-1 transmission.

Cerebrospinal and peripheral human immunodeficiency virus type 1 load in a multisite, randomized, double-blind, placebo-controlled trial of D-Ala1-peptide T-amide for HIV-1-associated cognitive-motor impairment.

D-Ala1-peptide T-amide (DAPTA) has shown neuroprotection in vitro against gp120-induced loss of dendritic arborization and is promulgated as a CCR5 antagonist. A multisite, randomized, double-blind clinical trial of DAPTA versus placebo prior to combination antiretroviral therapy conducted with human immunodeficiency virus (HIV)-1 seropositive participants having cognitive impairment showed no overall cognitive effect, though subgroups with greater impairment and CD4 cell counts of 201 to 500 cells/mm3 at baseline showed significant improvement. The objective of this study was to examine whether intranasal administration of DAPTA at a dose of 2 mg three times per day (tid) was associated with a reduction of cerebrospinal fluid (CSF) and peripheral (plasma and serum) viral load among a subgroup of participants completing 6 months of treatment. Baseline and 6-month CSF (n = 92) and peripheral (plasma n = 33; serum n = 24) viral load were measured by the Roche Ultrasensitive assay, version 1.5, with reflexive use of the AMPLICOR assay and preservation of the blind. A DAPTA treatment indicator variable was tested using generalized linear models on change in viral load. Peripheral load (combined plasma and serum) was significantly reduced in the DAPTA-treated group. No group differences in CSF viral load were found. This retrospective study on a limited subgroup of the original trial sample indicated that DAPTA treatment may reduce peripheral viral load without concomitant CSF effects. Future studies should be undertaken to confirm the existence of this result and the CSF-periphery dissociation observed with respect to HIV-1-associated cognitive-motor impairment.

Interleukin-6 and platelet protagonists in T lymphocyte and virological response.

The present cross-sectional study evaluated the status and relationship of interleukin-6, a platelet growth factor, with platelet counts, viral load, CD4 counts, and antiretroviral treatment in 75 HIV-infected subjects with thrombocytopenia and 50 gender-, race-, age- and antiretroviral treatment-matched controls without thrombocytopenia. Mean IL-6 production was significantly higher in thrombocytopenic participants (13 432+/-8596) than in non-thrombocytopenic subjects (12 859+/-3538 pg/10(5) Lym). Univariate analyses indicated, however, that thrombocytopenic patients were more likely to have <3000 pg of IL-6 than non-thrombocytopenic patients (OR=7 95% CI 1.3-12; P=0.01). For additional analyses, participants were dichotomized above and below 3000 pg of IL-6. Despite similar age, gender, drug use and antiretroviral treatment, thrombocytopenic participants had lower CD4 counts (186.5+/-149 vs. 401+/-286, P=0.005) than non-thrombocytopenic subjects. Thrombocytopenic participants with elevated IL-6, with or without HAART, were more likely to have higher HIV-replication (496 273+/-210 416; 34 656+/-25 332) than thrombocytopenic individuals with low IL-6 levels (105 332+/-42 699; 19 015+/-14 296 P=0.05). Non-thrombocytopenic patients with high IL-6 levels exhibited the highest CD4s (466.7+/-333) and the lowest viral burden (63 094+/-53 300) of the groups. Two distinct categories of HIV-associated thrombocytopenia exist: one accompanied by low IL-6, and another with compensatory elevations of IL-6. In thrombocytopenic individuals, the latter was associated with the poorest immunological and virological responses.

Intracellular substrates for the primer-unblocking reaction by human immunodeficiency virus type 1 reverse transcriptase: detection and quantitation in extracts from quiescent- and activated-lymphocyte subpopulations.

Treatment of human immunodeficiency virus type 1 (HIV-1)-infected patients with 3'-azido-3'-deoxythymidine (AZT) selects for mutant forms of viral reverse transcriptase (RT) with increased ability to remove chain-terminating nucleotides from blocked DNA chains. We tested various cell extracts for the presence of endogenous acceptor substrates for this reaction. Cell extracts incubated with HIV-1 RT and [(32)P]ddAMP-terminated DNA primer/template gave rise to (32)P-labeled adenosine 2',3'-dideoxyadenosine 5',5'''-P(1),P(4)-tetraphosphate (Ap(4)ddA), ddATP, Gp(4)ddA, and Ap(3)ddA, corresponding to the transfer of [(32)P]ddAMP to ATP, PP(i), GTP, and ADP, respectively. Incubation with [(32)P]AZT monophosphate (AZTMP)-terminated primer/template gave rise to the analogous (32)P-labeled AZT derivatives. Based on the rates of formation of the specific excision products, ATP and PP(i) levels were determined: ATP was present at 1.3 to 2.2 mM in H9 cells, macrophages, and unstimulated CD4(+) or CD8(+) T cells, while PP(i) was present at 7 to 15 microM. Under these conditions, the ATP-dependent reaction predominated, and excision by the AZT-resistant mutant RT was more efficient than wild type RT. Activated CD4(+) or CD8(+) T cells contained 1.4 to 2.7 mM ATP and 55 to 79 microM PP(i). These cellular PP(i) concentrations are lower than previously reported; nonetheless, the PP(i)-dependent reaction predominated in extracts from activated T cells, and excision by mutant and wild-type RT occurred with similar efficiency. While PP(i)-dependent excision may contribute to AZT resistance in vivo, it is likely that selection of AZT-resistant mutants occurs primarily in an environment where the ATP-dependent reaction predominates.