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Background Human immunodeficiency virus (HIV)–infected immunological nonresponders (INRs) fail to reconstitute their CD4+ T-cell pool after initiation of antiretroviral therapy, and their prognosis is inferior to that of immunological responders (IRs). A prevailing hypothesis is that the INR phenotype is caused by a persistently disrupted mucosal barrier, but assessments of gut mucosal immunology in different anatomical compartments are scarce. Methods We investigated circulating markers of mucosal dysfunction, immune activation, mucosal Th17 and Th22 cells, and mucosa-adherent microbiota signatures in gut mucosal specimens from sigmoid colon and terminal ileum of 19 INRs and 20 IRs in addition to 20 HIV-negative individuals. Results INRs had higher blood levels of the enterocyte damage marker intestinal fatty acid–binding protein than IRs. In gut mucosal biopsies, INRs had lower fractions of CD4+ T cells, higher fractions of interleukin 22, and a tendency to higher fractions of interleukin 17–producing CD4+ T cells. These findings were all restricted to the colon and correlated to circulating markers of enterocyte damage. There were no observed differences in gut microbial composition between INRs and IRs. Conclusions Restricted to the colon, enterocyte damage and mucosal immune dysfunction play a role for insufficient immune reconstitution in HIV infection independent of the gut microbiota.

IntroductionEicosanoids and intracellular signaling pathways are potential targets for host-directed therapy (HDT) in tuberculosis (TB). We have explored the effect of cyclooxygenase 2 inhibitor (COX-2i) treatment on eicosanoid levels and signaling pathways in monocytes.MethodsPeripheral blood mononuclear cells isolated from TB patients included in a randomized phase I clinical trial of standard TB treatment with (n=21) or without (n=18) adjunctive COX-2i (etoricoxib) were analyzed at baseline, day 14 and day 56. Plasma eicosanoids were analyzed by ELISA and liquid chromatography-mass spectrometry (LC-MS), plasma cytokines by multiplex, and monocyte signaling by phospho-flow with a defined set of phospho-specific antibodies.ResultsLipoxygenase (LOX)-derived products (LXA4 and 12-HETE) and pro-inflammatory cytokines were associated with TB disease severity and were reduced during TB therapy, possibly accelerated by adjunctive COX-2i. Phosphorylation of p38 MAPK, NFkB, Erk1/2, and Akt in monocytes as well as plasma levels of MIG/CXCL9 and procalcitonin were reduced in the COX-2i group compared to controls.ConclusionCOX-2i may reduce excess inflammation in TB via the LOX-pathway in addition to modulation of phosphorylation patterns in monocytes. Immunomodulatory effects of adjunctive COX-2i in TB should be further investigated before recommended for use as a HDT strategy.

Supplemental Digital Content is available in the text., OBJECTIVES: Increase of nucleated RBCs in peripheral blood has been shown to be predictive of mortality in ICU patients. The aim of this study was to explore the prognostic value of nucleated RBCs in the first blood sample taken at admission to the emergency department from patients with suspected sepsis. DESIGN: Single-center prospective cohort study. SETTING: Emergency department. PATIENTS: One-thousand two-hundred thirty-one consecutive adult patients with suspected sepsis were included in a prospective quality register-based cohort study. Inclusion criteria were as follows: patients received in rapid response team with blood cultures taken and immediate antibiotics given in the emergency department. Intervention: Not applicable. MEASUREMENT AND MAIN RESULTS: Nucleated RBCs, Sequential Organ Failure Assessment score, Quick Sequential Organ Failure Assessment, Charlson Comorbidity Index, and commonly used laboratory tests measured in the emergency department were compared with 30-day mortality. Nvaucleated RBC counts were divided into five groups, called “Nucleated RBC score,” according to nucleated RBC count levels and analyzed with logistic regression together with the Sequential Organ Failure Assessment score and Charlson Comorbidity Index. Of the 262 patients with nucleated RBCs equal to or higher than the detection limit (0.01 × 109/L), 26% died within 30 days, compared with 12% of the 969 patients with nucleated RBCs below the detection limit (p < 0.0001). Mortality was significantly higher for each increase in Nucleated RBC score, except from score 2 to 3, and was 62% in the highest group. In multivariate logistic regression, odds ratios for 30-day mortality were as follows: Nucleated RBC score: 1.33 (95% CI, 1.13–1.56), Sequential Organ Failure Assessment score: 1.32 (1.29–1.56), and Charlson Comorbidity Index: 1.17 (1.09–1.25). CONCLUSIONS: Most patients with suspected sepsis in emergency department had undetectable nucleated RBCs at admission to the emergency department. However, increased nucleated RBCs significantly predicted 30-day mortality. Nucleated RBCs may provide additional prognostic information to Sequential Organ Failure Assessment score and other laboratory tests.

Host-directed-therapy strategies are warranted to fight tuberculosis. Here we assess the safety and immunogenicity of adjunctive vaccination with the H56:IC31 candidate and cyclooxygenase-2-inhibitor treatment (etoricoxib) in pulmonary and extra-pulmonary tuberculosis patients in a randomized open-label phase I/II clinical trial (TBCOX2, NCT02503839). A total of 222 patients were screened, 51 enrolled and randomized; 13 in the etoricoxib-group, 14 in the H56:IC31-group, 12 in the etoricoxib+H56:IC31-group and 12 controls. Three Serious Adverse Events were reported in the etoricoxib-groups; two urticarial rash and one possible disease progression, no Serious Adverse Events were vaccine related. H56:IC31 induces robust expansion of antigen-specific T-cells analyzed by fluorospot and flow cytometry, and higher proportion of seroconversions. Etoricoxib reduced H56:IC31-induced T-cell responses. Here, we show the first clinical data that H56:IC31 vaccination is safe and immunogenic in tuberculosis patients, supporting further studies of H56:IC31 as a host-directed-therapy strategy. Although etoricoxib appears safe, our data do not support therapy with adjunctive cyclooxygenase-2-inhibitors., Modulating the host immune response during tuberculosis is an emerging and critical advance in the therapeutic approach. Here the authors present data from a first-in-human phase I/II randomised trial on the safety and immunogenicity of adjuvant therapy of the H56:IC31 vaccine and cyclooxygenase-2 inhibitors in patients with tuberculosis.

Background Seasonal influenza causes substantial numbers of hospitalizations annually, and there is a need to identify prognostic markers of clinical outcomes in hospitalized influenza patients. Methods Patients hospitalized with influenza were included during four influenza seasons (2014-2018). On admission, patients were assessed by the CRB65, CRB, SIRS and qSOFA risk scores. “Severe outcome” was defined as treatment in intensive care unit and/or all of the following three clinical parameters: pneumonia, O2 supplementation and hospital stay ≥ 5days. Regression analyses were used to study effects of scores, influenza (sub)types and age on outcomes and treatment. Results A total of 156 patients with median age of 70 years were included. Patients with influenza A(H3N2) were older than those with B and A(H1N1). Older age was associated with fewer symptoms. Severe outcome was observed in 22% of the cases. High CRB, CRB65, SIRS and qSOFA scores at admission was observed in 60%, 46%, 40% and 29% of patients with severe outcome, respectively. Influenza (sub)types were not associated with severe outcome. Antiviral treatment was given to 37% of the patients, while 78% received antibiotics. The use of antiviral treatment increased during the study period. Patients with influenza B received less antiviral treatment. Conclusion This is the first study describing adult patients hospitalized with seasonal influenza in Norway. Risk scores, and particularly CRB, may be useful to predict severe outcome in influenza disease. More patients might have profited from antiviral therapy; including patients with influenza B infection.

Here, we randomized 53 patients hospitalized with coronavirus disease 2019 (COVID-19) to hydroxychloroquine therapy (at a dose of 400 mg twice daily for seven days) in addition to standard care or standard care alone (ClinicalTrials.gov Identifier, NCT04316377). All severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) positive patients 18 years of age or older were eligible for study inclusion if they had moderately severe COVID-19 at admission. Treatment with hydroxychloroquine did not result in a significantly greater rate of decline in SARS-CoV-2 oropharyngeal viral load compared to standard care alone during the first five days. Our results suggest no important antiviral effect of hydroxychloroquine in humans infected with SARS-CoV-2., The use of hydroxychloroquine therapy for the treatment of Covid-19 is controversial. In this study, Lyngbakken and colleagues present a randomized controlled trial and show that the drug has no antiviral effects in humans infected with SARS-CoV-2.

Objectives The hypothesis of the study is that treatment with hydroxychloroquine sulphate in hospitalised patients with coronavirus disease 2019 (Covid-19) is safe and will accelerate the virological clearance rate for patients with moderately severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) when compared to standard care. Furthermore, we hypothesize that early treatment with hydroxychloroquine sulphate is associated with more rapid resolve of clinical symptoms as assessed by the National Early Warning Score 2 (NEWS2), decreased admission rate to intensive care units and mortality, and improvement in protein biomarker profiles (C-reactive protein, markers of renal and hepatic injury, and established cardiac biomarkers like cardiac troponin and B-type natriuretic peptide). Trial design The study is a two-arm, open label, pragmatic randomised controlled group sequential adaptive trial designed to assess the effect on viral loads and clinical outcome of hydroxychloroquine sulphate therapy in addition to standard care compared to standard care alone in patients with established Covid-19. By utilizing resources already paid for by the hospitals (physicians and nurses in daily clinical practice), this pragmatic trial can include a larger number of patients over a short period of time and at a lower cost than studies utilizing traditional randomized controlled trial designs with an external study organization. The pragmatic approach will enable swift initiation of randomisation and allocation to treatment. Participants Patients will be recruited from all inpatients at Akershus University Hospital, Lørenskog, Norway. Electronic real-time surveillance of laboratory reports from the Department of Microbiology will be examined regularly for SARS-CoV-2 positive subjects. All of the following conditions must apply to the prospective patient at screening prior to inclusion: (1) Hospitalisation; (2) Adults 18 years or older; (3) Moderately severe Covid-19 disease (NEWS2 of 6 or less); (4) SARS-CoV-2 positive nasopharyngeal swab; (5) Expected time of hospitalisation > 48 hours; and (6) Signed informed consent must be obtained and documented according to Good Clinical Practice guidelines of the International Conference on Harmonization, and national/local regulations. Patients will be excluded from participation in the study if they meet any of the following criteria: (1) Requiring intensive care unit admission at screening; (2) History of psoriasis; (3) Known adverse reaction to hydroxychloroquine sulphate; (4) Pregnancy; or (5) Prolonged corrected QT interval (>450 ms). Clinical data, including standard hospital biochemistry, medical therapy, vital signs, NEWS2, and microbiology results (including blood culture results and reverse transcriptase polymerase chain reaction [RT-PCR] for other upper airway viruses), will be automatically extracted from the hospital electronic records and merged with the study specific database. Intervention and comparator Included patients will be randomised in a 1:1 ratio to (1) standard care with the addition of 400 mg hydroxychloroquine sulphate (PlaquenilTM) twice daily for seven days or (2) standard care alone. Main outcomes The primary endpoint of the study is the rate of decline in SARS-CoV-2 viral load in oropharyngeal samples as assessed by RT-PCR in samples collected at baseline, 48 and 96 hours after randomization and administration of drug for the intervention arm. Secondary endpoints include change in NEWS2 at 96 hours after randomisation, admission to intensive care unit, mortality (in-hospital, and at 30 and 90 days), duration of hospital admission, clinical status on a 7-point ordinal scale 14 days after randomization ([1] Death [2] Hospitalised, on invasive mechanical ventilation or extracorporeal membrane oxygenation [3] Hospitalised, on non-invasive ventilation or high flow oxygen devices [4] Hospitalized, requiring supplemental oxygen [5] Hospitalised, not requiring supplemental oxygen [6] Not hospitalized, but unable to resume normal activities [7] Not hospitalised, with resumption of normal activities), and improvement in protein biomarker profiles (C-reactive protein, markers of renal and hepatic injury, and established cardiac biomarkers like cardiac troponin and B-type natriuretic peptide) at 96 hours after randomization. Randomisation Eligible patients will be allocated in a 1:1 ratio, using a computer randomisation procedure. The allocation sequence has been prepared by an independent statistician. Blinding (masking) Open label randomised controlled pragmatic trial without blinding, no active or placebo control. The virologist assessing viral load in the oropharyngeal samples and the statistician responsible for analysis of the data will be blinded to the treatment allocation for the statistical analyses. Numbers to be randomized (sample size) This is a group sequential adaptive trial where analyses are planned after 51, 101, 151 and 202 completed patients, with a maximum sample size of 202 patients (101 patients allocated to intervention and standard care and 101 patients allocated to standard care alone). Trial Status Protocol version 1.3 (March 26, 2020). Recruitment of first patient on March 26, 2020, and 51 patients were included as per April 28, 2020. Study recruitment is anticipated to be completed by July 2020. Trial registration ClinicalTrials.gov number, NCT04316377. Trial registered March 20, 2020. Full protocol The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.

BACKGROUND Microbial translocation has been suggested as a driver of cardiovascular disease in HIV infection. We hypothesized that microbial translocation and the resulting monocyte activation would be associated with markers of endovascular dysfunction. METHODS In 60 HIV-infected patients on combination antiretroviral therapy, plasma levels of lipopolysaccharide, soluble CD14 (sCD14), asymmetric dimethylarginine (ADMA), and symmetric dimethylarginine (SDMA) were measured. RESULTS ADMA and SDMA were associated with sCD14 but not lipopolysaccharide. There was a significant increase in ADMA and SDMA through tertiles of sCD14, and both markers were associated with sCD14 in multivariate linear regression analyses. CONCLUSIONS Monocyte activation as measured by sCD14 is associated with endovascular dysfunction in HIV infection.

The current study examined the efficacy of a web-based intervention (WBI; Avanti) on symptoms of depression and well-being for patients diagnosed with HIV. A two-armed randomized trial recruited patients on antiretroviral therapy (ART) at an outpatient clinic. Thirty-six patients were allocated to Avanti and 31 patients to a control group. Primary outcomes were symptoms of depression and subjective well-being (SWB), and secondary outcomes included life satisfaction and affect balance. Paired tests showed that only patients following Avanti had significant improvements in SWB by 3 months as well as affect balance. No significant differences between groups were detected in any of the outcome parameters at baseline after 3 months, as expected from group size and variability in the parameters. However, time since HIV diagnosis and ART initiation moderated the effects of Avanti. In conclusion, our data show that patients with HIV infection may benefit from a WBI in adjunct to medical treatment.

Objectives The aim of the study was to test the hypothesis that microbial translocation, quantified by levels of lipopolysaccharide (LPS) and subsequent monocyte activation [soluble (sCD14)], is associated with hypertension in HIV-infected individuals. Methods In this exploratory substudy, 42 patients were recruited from a larger, longitudinal HIV-infected cohort study on blood pressure. LPS and sCD14 levels were measured retrospectively at the time of nadir CD4 cell count, selecting untreated HIV-infected patients with both advanced immunodeficiency and preserved immunocompetence at the time of nadir. Patients with later sustained hypertension (n = 16) or normotension (n = 26) throughout the study were identified. LPS was analysed using the Limulus Amebocyte Lysate colorimetric assay (Lonza, Walkersville, MD) and sCD14 using an enzyme-linked immunosorbent assay (ELISA). Nonparametric statistical tests were applied. Results In the HIV-infected patients [median (interquartile range) age 42 (32–46) years; 79% male and 81% Caucasian], LPS and sCD14 levels were both negatively correlated with nadir CD4 cell count. Plasma levels of LPS (P

Background Tuberculosis (TB) causes a major burden on global health with long and cumbersome TB treatment regimens. Host-directed immune modulating therapies have been suggested as adjunctive treatment to TB antibiotics. Upregulated cyclooxygenase-2 (COX-2)-prostaglandin E2 (PGE2) signaling pathway may cause a dysfunctional immune response that favors survival and replication of Mycobacterium tuberculosis (Mtb). Methods Blood samples were obtained from patients with latent TB (n = 9) and active TB (n = 33) before initiation of anti-TB chemotherapy. COX-2 expression in monocytes and ESAT-6 and Ag85 specific T cell cytokine responses (TNF-α, IFN-γ, IL-2), proliferation (carboxyfluorescein succinimidyl ester staining) and regulation (FOXP3+ T regulatory cells) were analysed by flow cytometry and the in vitro effects of the COX-1/2 inhibitor indomethacin were measured. Results We demonstrate that indomethacin significantly down-regulates the fraction of Mtb specific FOXP3+ T regulatory cells (ESAT-6; p = 0.004 and Ag85; p

Therapeutic immunization in chronic HIV infection aims to induce durable, HIV-specific immune responses capable of controlling disease progression, but immunological correlates of clinical efficacy are poorly defined. We have previously immunized 38 patients with a mixture of four short Gag p24-like conserved peptides (Vacc-4x) targeting skin dendritic cells. Antiretroviral treatment (ART) was initially stopped after completed immunizations and resumed post-protocol during regular clinical follow-up according to current guidelines. Four years after enrolment, Vacc-4x-specific cellular responses were evaluated in vivo by delayed-type hypersensitivity (DTH) skin test, and in vitro by a T-cell proliferation assay. Kaplan-Meier product-limit estimates were used to analyse time until ART was resumed. Peptide-specific cellular immune responses induced by Vacc-4x had persisted 4 years after the last immunization in terms of unchanged DTH independent of ART and detectable proliferative T-cell responses which correlated to the native peptides (R = 0.73, p = 0.01). Circulating bifunctional (IFN-γ and IL-10) Vacc-4x-specific T-cell clones were detected in 43% of patients. Subjects with the strongest post-immunization DTH responses appeared to start ART later compared with DTH low responders (p = 0.07). These data suggest that DTH responses should be further evaluated as a new and convenient tool for predicting clinical efficacy in trials testing therapeutic immunizations.

Human regulatory T cells (Tregs) are essential in maintaining immunological tolerance and suppress effector T cells. Tregs are commonly up-regulated in chronic infectious diseases such as tuberculosis (TB) and human immunodeficiency virus (HIV) infection and thereby hamper disease-specific immune responses and eradication of pathogens. The MEK/ERK signaling pathway is involved in regulation of the FoxP3 transcription factor, which directs a lineage-specific transcriptional program to define Tregs and control their suppressive function. Here, we aimed to target activation of disease-specific Tregs by inhibition of the MEK/ ERK signaling pathway based on the hypothesis that this would improve anti-HIV and anti- TB immunity. Stimulation of T cells from untreated TB (n = 12) and HIV (n = 8) patients with disease-specific antigens in vitro in the presence of the MEK inhibitor (MEKI) trametinib (GSK1120212) resulted in significant down-regulation of both FoxP3 levels (MFI) and fractions of resting (CD45RA+FoxP3+) and activated (CD45RA−FoxP3++) Tregs. MEKI also reduced the levels of specific T effector cells expressing the pro-inflammatory cytokines (IFN-γ, TNF-α and IL-2) in both HIV and TB patients. In conclusion, MEKIs modulate disease antigen-specific Treg activation and may have potential application in new treatment strategies in chronic infectious diseases where reduction of Treg activity would be favorable. Whether MEKIs can be used in current HIV or TB therapy regimens needs to be further investigated.

BACKGROUND Microbial translocation and chronic inflammation may contribute to non-AIDS morbidity in patients with HIV. This study assessed the impact of probiotic intervention on microbial translocation and inflammation in patients on antiretroviral therapy with viral suppression and subnormal CD4 count. METHODS Thirty-two patients receiving antiretroviral therapy (CD4

HIV-infected patients who fail to reconstitute their CD4 T-cell counts during suppressive antiretroviral therapy (ART) have increased risk of both AIDS-related and non-AIDS-related morbidity and mortality. Improved understanding of immunological nonresponse (INR) is necessary to enable earlier clinical intervention.In a cohort of 112 HIV-infected patients starting ART, we performed a serial analysis of 32 plasma-soluble markers, assessed by multiplex cytokine and enzyme immunoassay. Samples were drawn pre-ART and during the first 3 years of treatment, with a final observation time of 8.4 years (interquartile range, 7.0-10.7 years) on ART. Long-term INR (LT-INR) was defined as failure to reach a CD4 T-cell count350 cells per microliter. Marker stability was evaluated by parallel analysis of samples from ART-naïve and HIV-seronegative controls.Baseline CD4 T-cell counts predicted subsequent LT-INR (n = 15) [odds ratio, 1.10 (95% confidence interval: 1.01 to 1.19) pr. 10 cells/μL reduction in CD4 count, P = 0.030] in the cohort as a whole, but not in patients with baseline CD4 counts200 cells per microliter (n = 78). LT-INR was best characterized by elevated plasma levels of the CC chemokine macrophage inflammatory protein 1β (MIP-1β), both at baseline (pre-ART) and during ART. In patients with baseline CD4 counts200 cells per microliter, baseline MIP-1β predicted LT-INR [odds ratio 1.23 (95% confidence interval: 1.02 to 1.47) per 10 pg/mL increase in MIP-1β, P = 0.029].Elevated pre-ART levels of MIP-1β identified LT-INR patients who started ART at CD4 counts200. INR was characterized by persistently high MIP-1β during suppressive ART. Thus, MIP-1β may be of use for early identification of LT-INR.

Long-term HIV-specific immune responses and clinical outcomes were evaluated in HIV-infected patients previously immunized with p24-like peptides (Vacc-4x) targeting dendritic cells (DC). Vacc-4x-induced cellular immune responses were unchanged 1.5 years after completing immunization, and 62% were still off combined antiretroviral treatment (CART). The magnitude of early Vacc-4x responses determined whether the resumption of CART was clinically indicated 2 years after enrollment. These observations encourage further exploration of both Vacc-4x and other HIV peptide-based immunization regimens targeting DC.

Objective The Vacc-4x immunotherapy candidate is composed of four modified peptides corresponding to conserved domains of the HIV-1 protein p24 that preferentially include HLA-A2 restricted elements. Dose-dependent safety and immunogenicity of Vacc-4x and the significance of a HLA-A2 haplotype were examined. Design Non-AIDS, HIV-1 infected healthy patients (n = 40) stable on HAART with CD4 counts > 300 x 10 cells/l were randomized to receive either low-dose or high-dose Vacc-4x over 26 weeks in an open, prospective phase II clinical trial. Methods Patients received a total of 10 intradermal injections, using recombinant granulocyte-macrophage colony stimulating factor as a local adjuvant. Vacc-4x-specific cellular responses were monitored in vivo by delayed-type hypersensitivity (DTH) skin test infiltrates and in vitro by both T-cell proliferation, and induction /secretion of cytokines. Results Most patients developed Vacc-4x-specific DTHs (90%) and proliferative T-cell responses (80%) that were inter-related in magnitude. High-dose Vacc-4x generally induced stronger specific immune responses than low dose in terms of DTH areas and CD4 and CD8 T-cell proliferation. Only HLA-A2 negative patients had a definite dose advantage, and this subgroup had in fact the best overall DTH and proliferative responses. In contrast, no significant dose difference was observed for HLA-A2 positive patients. No serious adverse events were reported. Conclusions HIV-associated specific responses were safely induced in most patients by Vacc-4x in a dose-dependent manner and were also influenced by the HLA haplotype.

Objectives: Biomarkers for diagnosis and therapy efficacy in tuberculosis (TB) are requested. We have studied biomarkers that may differentiate between active and latent TB infection (LTBI), the influence of HIV infection and changes during anti-TB chemotherapy. Methods: Thirty-eight plasma cytokines, assessed by multiplex and enzyme immunoassays, were analyzed in patients with active TB before and during 24 weeks of anti-TB chemotherapy (n = 65), from individuals with LTBI (n = 34) and from QuantiFERON-TB (QFT) negative controls (n = 65). The study participants were grouped according to HIV status. Results: Plasma levels of the CXC chemokine IP-10 and soluble TNF receptor type 2 (sTNFr2) significantly differentiated active TB from the LTBI group, irrespective of HIV status. In the HIV-infected group the sensitivity and specificity was 100% for IP-10 with a cut-off of 2547 pg/mL. Plasma IP-10 declined gradually during anti-TB chemotherapy (12–24 weeks, p = 0.002) to a level comparable to LTBI and QFT negative control groups. sTNFr2 fluctuated throughout therapy, but was decreased after 12–24 weeks (p = 0.006). Conclusions: IP-10 distinguished with high accuracy active TB from LTBI irrespective of HIV infection and declined during anti-TB chemotherapy. Plasma IP-10 may serve as a diagnostic biomarker to differentiate between the stages of TB infection and for monitoring therapy efficacy. ª 2015 The Authors. Published by Elsevier Ltd on behalf of the The British Infection Association. This is an open access article under the CC BY-NC-SA license (http://creativecommons.org/licenses/by-nc-sa/4.0/).

SUMMARYTo examine the potential pathogenic role of IL-10 in HIV infection, we measured serum IL-10 levels in 51 HIV-infected patients and 23 healthy controls both on cross-sectional and longitudinal testing. All clinical groups (Centers for Disease Control (CDC) categories) of HIV-infected patients had significantly higher circulating IL-10 levels than controls, with the highest levels among the AIDS patients, particularly in patients with ongoing Mycobacterium avium complex (MAC) infection. Among 32 HIV-infected patients followed with longitudinal testing (median observation time 39 months), patients with disease progression had increasing IL-10 levels in serum, in contrast to non-progressing patients where levels were stable. While both IL-10 and tumour necrosis factor-alpha (TNF-α) increased in patients with disease progression, the IL-10/TNF-α ratio decreased in these patients, suggesting imbalance between these two cytokines. Finally, we found that highly active anti-retroviral therapy (HAART) induced a significant, gradual decrease in IL-10 levels but without normalization. These findings suggest a pathogenic role for IL-10 in HIV infection, and may suggest a possible role for immunomodulating therapy which down-regulates IL-10 activity in addition to concomitant potent anti-retroviral therapy in HIV-infected patients.

Because persistent tumor necrosis factor (TNF)-alpha activation may play a pathogenic role in human immunodeficiency virus infection, TNF component levels were assessed over 78 weeks in plasma and peripheral blood mononuclear cells (PBMC) during highly active antiretroviral therapy (HAART) in 40 HIV-infected patients. HAART induced a significant decline in plasma levels of TNF-alpha and soluble TNF receptors and was associated with a fall in the abnormally increased unstimulated and a rise in the abnormally low Mycobacterium avium complex-purified-protein derivative-stimulated TNF-alpha released from PBMC. However, concentrations of these TNF components were not normalized. Patients with virologic and immunologic treatment failure after 52 weeks had higher levels of several TNF components than other patients early after initiation of therapy, also during periods with adequate virologic response. Although TNF components significantly decreased during HAART, these results support data indicating that full immunologic normalization is not achieved during such therapy. The persistent activation of the TNF system in a subgroup of persons may be involved in treatment failure.

Objectives: Therapeutic vaccination in chronic HIV-infection aims to attenuate disease progression by promoting new HIV-specific T cell-clones. Most clinical trials with therapeutic vaccines are conducted on patients receiving antiretroviral therapy (ART). However, studies of vaccination in untreated individuals are limited. We present the first data from a phase I/II clinical trial with a peptide-based therapeutic vaccine (Vacc-5q) consisting of five short, modified consensus peptides from p17, p24, and Tat. In addition to evaluating safety and immunogenicity of Vacc-5q, we compared responses induced in patients on effective ART with those of ART-naive patients. Methods: HIV-infected patients stable on ART (n=10) and treatment naive patients (n=10) received 11 intradermal injections of Vacc-5q over 26 weeks, using GM-CSF as an adjuvant. Immunogenicity was assessed both in vivo by delayed type hypersensitivity (DTH) skin tests, and in vitro by T cell activation assays. Persistence of immune responses was retested after 3.5 years. Results: Vacc-5q was found to be immunogenic. Specific T cell-responses both in vivo and in vitro increased significantly from baseline to week 4 (p

Background Biomarkers to differentiate between active tuberculosis (TB) and latent TB infection (LTBI) and to monitor treatment responses are requested to complement TB diagnostics and control, particularly in patients with multi-drug resistant TB. We have studied soluble markers of the Toll-like-receptor 4 (TLR-4) pathway in various stages of TB disease and during anti-TB treatment. Methods Plasma samples from patients with culture confirmed drug-sensitive TB (n = 19) were collected before and after 2, 8 and 24 weeks of efficient anti-TB treatment and in a LTBI group (n = 6). Soluble (s) CD14 and myeloid differentiation-2 (MD-2) were analyzed by the Enzyme-linked immunosorbent assay (ELISA). Lipopolysaccharide (LPS) was analyzed by the Limulus Amebocyte Lysate colorimetric assay. Nonparametric statistics were applied. Results Plasma levels of sCD14 (p

OBJECTIVE Microbial translocation has been suggested to be a driver of immune activation and inflammation. It is hypothesized that microbial translocation may be related to dyslipidemia, insulin resistance, and the risk of coronary heart disease in HIV-infected individuals. DESIGN Cross-sectional study of 60 HIV-infected patients on combination antiretroviral therapy with viral suppression >2 years and 31 healthy age-matched controls. METHODS Lipopolysaccharide (LPS) was analyzed by limulus amebocyte lysate colorimetric assay. Lipids, including cholesterol, low-density lipoprotein (LDL), and triglycerides, were measured. Glucose metabolism was determined using an oral glucose tolerance test. Body composition was determined using whole-body dual-energy x-ray absorptiometry scans and magnetic resonance imaging. The Framingham risk score was used to assess risk of cardiovascular disease and myocardial infarction. RESULTS HIV-infected patients had higher level of LPS compared with controls (64 pg/mL vs. 50 pg/mL, P = 0.002). Likewise, HIV-infected patients had higher triglycerides, LDL, and fasting insulin as well as evidence of lower insulin sensitivity compared with controls. Among HIV-infected patients, high LPS was associated with a higher level of triglycerides and LDL and with lower insulin sensitivity. Importantly, among HIV-infected patients, high LPS was associated with a higher Framingham risk score. CONCLUSIONS HIV-infected patients with suppressed viral replication had increased level of microbial translocation as measured by LPS. LPS was associated with cardiometabolic risk factors and increased Framingham risk score. Hence, the gastrointestinal mucosal barrier may be a potential therapeutic target to prevent dyslipidemia and future cardiovascular complications in HIV infection.

Purpose of the study: One of the most prevalent mental conditions in people with HIV is depression as uniquely characterized by low positive affect. This study examined the effect of a web-based intervention (Avanti) on overall mood and depressive symptoms among patients with HIV infection. Methods: Patients treated with effective antiretroviral treatment were included in a two-armed trial with substance abuse as an exclusion criterion and randomized to Avanti (n=36) or control (n=31). Patients were surveyed at baseline, as well as 1 and 3 months after the initiation of a 5-week intervention period. Outcomes were Center for Epidemiological Studies-Depression scale and the Positive and Negative Affect Schedule which was combined into an overall mood index. Changes within groups were tested by Wilcoxon matched pairs test and baseline differences between groups by chi-square and Mann-Whitney independent samples test. Summary of results: Baseline scores for both groups were similar. However, patients in the intervention group had an improvement in median (M) overall mood from baseline (M=67.6) to 1 month (M=71, p=0.02) which was also maintained from 1 to 3 months (M=71.9). Moreover, these patients had a favorable reduction in negative affect from 1 (M=24) to 3 months (M=22, p=0.01) and a transient improvement in positive affect from baseline (M=31.7) to 1 month after intervention onset (M=35, p

Summary Objectives The objectives of the study were to assess the utility of dried blood spots (DBS) for the detection of cytomegalovirus (CMV) antibody and viremia in a resource-poor setting, to study the prevalence of CMV antibody and viremia in HIV-infected patients with access to antiretroviral therapy (ART) in Tanzania, and to relate CMV viremia to outcome. Methods DBS were prepared from 168 ART-naive patients at baseline. Demographic, clinical, and laboratory data were obtained from patient records. CMV antibody was analyzed by chemiluminescent microparticle immunoassay and viremia by quantitative PCR. Results All patients were CMV-seropositive. At baseline 38 (22.6%) had detectable CMV viremia and 14 (8.3%) had a CMV viral load ≥200 copies/ml. In 135 patients available for follow-up, CMV ≥200 copies/ml was an independent risk factor for death with a hazard ratio of 5.0 (95% confidence interval 2.1–11.9) after adjusting for confounders. Symptoms compatible with CMV disease were common with viremia ≥200 copies/ml and CD4+ T cell counts 3 , but confirmatory diagnostic procedures were unavailable. Conclusions DBS are suitable for the detection of CMV antibody and viremia in HIV patients in resource-poor areas. CMV viremia was frequent and associated with an increased risk of death. Improved diagnosis and treatment of CMV may improve the prognosis for HIV-infected patients in developing countries and should be addressed in future studies.

Vacc-4x contains 4 HIV p24-like short peptides. In a previous phase II trial this immunized 90% of 38 patients on antiretroviral treatment (ART) after intradermal delivery in conjunction with local granulocyte-macrophage colony-stimulating factor (GM-CSF) as adjuvant. In this study, 22 responders were retested for cellular memory at a median 7.3 y after their last immunization. All had resumed effective ART after an interspersed ART-free median interval of 2.2 y.Vacc-4x as 15-mer overlapping by 2 amino acid panels and Vacc-4x consensus peptide sequences (4xCP) were used as antigens. Proliferation was determined as percentages of CFSE(dim)HLA-DR(+) 7AAD(-) CD3+ T cells of the CD4+ and CD8+ subsets after 6 days of culture. Frequencies of specific T cells in 6-h cultures were determined by interferon-γ (IFN)(+) CD4+ and IFN(+) CD8+, as well as degranulating bifunctional CD107a + IFN(+) CD8 + subsets.Proliferative CD4+ and CD8+ responses to Vacc-4x as well as 4xCP were still present in 95% and 68%, respectively. Proliferation correlated with the Vacc-4x delayed-type hypersensitivity test (DTH) obtained after completed immunizations (CD4 + r = 0.63 (p = 0.002) and CD8 + r = 0.47 (p = 0.03)), suggesting that they represent T cell memory recall responses. The proliferative CD8+ and possibly CD4 + subset responses to 4xCP peptides correlated with Vacc-4x (r = 0.46 (p = 0.03) and r = 0.38 (p = 0.08), respectively). Forty-one percent still had Vacc-4x-specific IFN + CD4 + T cells, which correlated to corresponding frequencies of 4xCP peptides (r = 0.50, p = 0.02). CD107a(+) IFN(+) CD8 + T cell responses against Vacc-4x were found in 55%.Evidence of long-lasting T cell memory recall responses to a peptide-based immunotherapeutic candidate for HIV-infected patients should enhance the focus on peptide-based intradermal vaccine delivery.

Purpose: To explore the relations between insulin resistance, plasma lactate, and mitochondrial (mt) DNA alterations in skeletal muscle in HIV-infected patients treated with nucleoside reverse transcriptase inhibitors (HIV+NRTI+). Method: Insulin resistance was estimated using the homeostatic model assessment (HOMA-IR). Mitochondrial dysfunction was determined by plasma lactate at rest and after subanaerobic exercise, mitochondrial/nuclear DNA (mt/nDNA) ratio, and mtDNA deletions in skeletal muscle. Results: HIV+NRTI+ patients (n = 27) had higher levels of HOMA-IR, higher lactate at rest as well as after exercise, and more frequent mtDNA deletions and decreased mt/nDNA ratios compared with controls (n = 15). Only in HIV+NRTI+ patients, HOMA-IR correlated with resting lactate (r = 0.5, p = .02) and probably also lactate 3, 5, and 8 minutes after exercise (r = 0.4; p = .075, p = .048, and p = .056, respectively). In contrast, neither HOMA-IR nor the lactate levels correlated with mt/nDNA ratio and mtDNA deletions in skeletal muscle in HIV+NRTI+ patients (r .6), whereas resting lactate correlated with mt/nDNA ratio in HIV seronegative controls (r = –0.7, p = .02). Conclusion: In HIV+NRTI+ patients, both resting and postexercise levels of lactate were related to insulin resistance rather than mtDNA alterations in skeletal muscle. Key words: HIV-1 infection, insulin resistance, lactate, mitochondrial dysfunction

OBJECTIVES To examine the immune modulating effects of cyclooxygenase type 2 (COX-2) inhibitors (COX-2i) in HIV-infected patients on combination antiretroviral treatment (CART). DESIGN In-depth substudy from an approved, open, controlled, randomized study comparing the immune modulating effects of CART in combination with COX-2i after 12 weeks. METHODS Patients (n = 38) on long-term CART with stable viral load (VL) 100/microl were randomized to CART and rofecoxib 25 mg bid (n = 12) or celecoxib 400 mg bid (n = 12), or CART only without placebo (n = 14). Routine clinical chemistry, CD4+ and CD8+ counts and VL were safety parameters. Immunological parameters included C-reactive protein, beta2-microglobulin, Ig isotypes and IgG subclasses as well as several T-lymphocyte subsets. Non-parametric analyses were used throughout. RESULTS Prestudy experiments showed higher median intracellular expression of COX-2 in CD4+ (P = 0.048) and possibly CD8+ (P = 0.09) T cells from patients on CART compared with uninfected controls. In the clinical study, increased CD4+ T-cell counts were observed only in patients on COX-2i with VL < 50 copies/ml (P = 0.02). Decreased expression of CD38+ on CD8+ T cells and subsets as well as reductions in IgA and IgM (P < 0.03) were most pronounced in patients on COX-2i who had detectable VL (n = 6). COX-2i treatment enhanced the perforin content particularly in the differentiated CD27-/CD8+ T-cell subsets compared with controls (P = 0.05). CONCLUSIONS COX-2i together with CART improved markers for persistent immune activation, particularly in patients with viraemia, as well as enhanced perforin expression, and thereby strengthened COX-2 as a potential therapeutic target in HIV infection.

We have previously shown that HIV p24-like peptides (Vacc-4x) via activation of skin dendritic cells induced immune responses in 90% of HIV patients on highly active antiretroviral treatment (HAART). These patients (n = 38) were here subjected to a final 14-week interruption of HAART. Patients with the highest delayed type hypersensitivity (DTH) responses to Vacc-4x-peptides before treatment interruption tended to achieve lower actual HIV RNA levels at the end of the study compared to Vacc-4x DTH low-responders (p = 0.08) and significantly so in terms of viral loads relative to their individual pre-HAART HIV RNA set-points (p = 0.04). CD4+ lymphocyte counts were maintained only among DTH high responders but decreased in the other patients during recurrent viremia (p ≤ 0.02). No other pre-study differences in HIV history or p24-responses were found.

Background Vacc-4x, a Gag p24-based therapeutic HIV vaccine, has been shown to reduce viral load set-points after intradermal administration. In this randomized controlled pilot study we investigate intranasal administration of Vacc-4x with Endocine as adjuvant. Methods Safety and immunogenicity were tested in patients on effective ART. They were randomized to low, medium or high dose Vacc-4x or adjuvant alone, administered four times at weekly intervals with no booster. Vacc-4x-specific T cell responses were measured in vitro by proliferation and in vivo by a single DTH skin test at the end of study. Nasal and rectal mucosal secretions were analyzed for Vacc-4x-specific antibodies by ELISA. Immune regulation induced by Vacc-4x was assessed by functional blockade of the regulatory cytokines IL-10 and TGF-β. Results Vacc-4x proliferative T cell responses increased only among the vaccinated (p≤0.031). The low dose group showed the greatest increase in Vacc-4x CD8+T cell responses (p = 0.037) and developed larger DTH (p = 0.005) than the adjuvant group. Rectal (distal) Vacc-4x IgA and IgG antibodies also increased (p = 0.043) in this group. In contrast, the high dose generated higher nasal (local) Vacc-4x IgA (p = 0.028) and serum IgG (p = 0.030) antibodies than the adjuvant. Irrespective of dose, increased Vacc-4x CD4+T cell responses were associated with low proliferation (r = −0.82, p

The objective was to study the progression of HIV infection in relation to immunological and virological variables with emphasis on the role of CD8+ lymphocytes. Prospective follow-up from October 1991 of patients observed for at least 18 months allowing nucleoside analogue monotherapy. Peripheral CD4+ and CD8+ lymphocyte counts HIV RNA and soluble CD8 were analysed by statistics allowing the evaluation of serial data avoiding time points with concurrent infections. Setting: Tertiary university clinic. Patients: Forty-nine patients were followed for 52.6 months baseline CD4+ count of 300 x 10/6/l sample interval of 5.9 months (medians). Main outcome measures: AIDS death and CDC groups B- or C-related events. AIDS developed in 28% of patients. Baseline CD8+ counts above the median were significantly associated with AIDS development; the best Cox model included CD8+ cells and the log10RNA/CD4 ratio. A decline in CD8+ counts relative to baseline most significantly predicted AIDS along with higher baseline RNA and actual CD4+ counts of less than 200 x 10/6/l. Levels of soluble CD8 in the blood relative to total CD8+ cells significantly increased in patients developing AIDS. Death occurred in 16% of the patients and was only predicted by high CD8+ cell counts at baseline. CDC B- and C-related events occurred in 35% of the patients and were best predicted by high baseline CD8+ counts and high RNA levels. The serial quantitation of CD8+ lymphocytes gave highly significant predictive information on the natural progression of HIV infection in patients with moderate to severe immune deficiency. Our data suggest that the hyperactivation of CD8+ lymphocytes is an important factor leading to a numerical decrease of CD8+ lymphocytes in progressive HIV infection. (authors)

Background Since its implementation more than a decade ago combination Antiretroviral Therapy (ART) has had a dramatic effect on HIV-1 induced mortality and morbidity. However, 15-20% of the patients initiating ART do not regain CD4 T-cells despite an effective suppression of HIV1mRNA levels in blood. The incomplete therapeutic effect leaves these patients at elevated risk of disease complications and death. Attempts to reduce clinical events through IL-2 induced stimulation of CD4 T-cells in the ESPRIT and SILCAAT studies have not proven to be successful. Until further research has given better opportunities for a successful direct induction of functional CD4 cells a logical alternative may be to increase CD4 counts indirectly by improving anti-HIV effect while on ART using antigen-specific immune stimulation with an effective HIV-vaccine.

Preoperative serum levels of secretory IgA (SIgA) and secretory IgM (SIgM) were quantified by an enzyme-linked immunosorbent assay in 100 patients with large bowel carcinoma. The values obtained were related to tumor characteristics such as Dukes' stage, differentiation, DNA ploidy pattern, expression of secretory component (SC), volume, and plasma level of carcinoembryonic antigen (CEA). Statistical comparison was made with matched controls. Only patients with Stage D tumors showed significantly increased serum levels of SIgA and SIgM (P less than 0.004). The combined diagnostic sensitivity of SIg and CEA in Stage D was 0.94. In patients with liver involvement, serum SIgA and SIgM were elevated in 67% and 53%, respectively. However, circulating SIg was neither correlated with tumor SC expression nor with any other studied variable. Thus, the raised serum SIg levels were apparently not caused by release of SC from the tumors but, instead, by hampered liver function due to hepatic metastasis.

One hundred patients operated for colorectal carcinoma were followed clinically and with serial blood samples from 5 to 8 years. Levels of secretory component (SC) associated with IgA and IgM in serum were measured and related to Dukes' stage, histological differentiation, tumour expression of SC, and circulating carcinoembryonic antigen (CEA). On the whole, elevated levels of SC in serum were found in 15 of the 20 patients who already had (n = 15), or later developed (n = 5), liver metastasis. Four of the latter 5 patients showed raised SC levels with a 5.5 months median lead time from the first positive serum sample to clinically manifest liver disease. These data are interesting in view of the promising results reported for liver resection in patients with colorectal carcinoma.

The clinical significance of cytomegalovirus (CMV) DNA detection in post-kidney transplantation infection surveillance was examined by comparing the performance of three assays for detection of CMV in blood: the test for CMV-pp65-antigen in leukocytes, which is routinely employed in our laboratory, the quantitative plasma CMV-DNA-polymerase chain reaction (PCR; Cobas Amplicor CMV Monitor test®) and the qualitative plasma CMV-DNA-PCR (Amplicor CMV test®). Thirteen kidney transplant recipients were monitored with serial samples taken over a period of 3 months following transplantation. The quantitative CMV-PCR was the test with highest sensitivity, 95.9%, vs. 88.9% and 76.9% for the CMV-pp65 antigen assay and qualitative CMV-PCR, respectively. The virus load in the first positive specimens, assessed as DNA-copies/mL, was significantly associated with CMV disease because five of the six patients who developed disease, but only one of the seven who did not develop disease, had more than 3000 CMV-DNA-copies/mL. The number of CMV-pp65 antigen-positive cells in the first positive specimens did not have predictive value for development of CMV disease. Assessment of CMV in plasma by the quantitative CMV-PCR is especially useful since it has a high sensitivity and the amount of CMVDNA in plasma is a good predictor of CMV disease.

AIMS: To evaluate serum secretory component in relation to early detection and clinical management of liver metastasis in patients with colorectal cancer. METHODS: Secretory component and carcinoembryonic antigen (CEA) were analysed in serial serum samples from 23 patients who had liver metastases as the only apparent recurrence, and in sera from 54 matched controls. Results of surgical treatment of recurrences were classified peroperatively as radical when no residual tumour was apparent and resection margins were free of disease. RESULTS: In total, 18 (78%) patients had increased secretory component during the whole follow up period (median 16 months); 12 (52%) had raised secretory component concentrations before clinical recurrence (median lead time 5.2 months). There was no difference before recurrence between circulating secretory component and CEA in sensitivity and lead times. Seventeen patients underwent surgery for hepatic metastasis; seven had radical hepatic resection of which only two (29%) showed increased secretory component concentrations before clinical recurrence; both had concurrent raised CEA values. By contrast, secretory component was raised in 83% of those cases considered inoperable. CONCLUSIONS: Although serum secretory component clearly increases in most patients with liver metastases, its clinical value seems questionable because secretory component apparently indicates mainly inoperable hepatic metastases.