Inflammatory profile of patients with tuberculosis with or without HIV-1 co-infection: a prospective cohort study and immunological network analysis

Background HIV-1 mediated dysregulation of the immune response to tuberculosis and its effect on the response to antitubercular therapy (ATT) is incompletely understood. We aimed to analyse the inflammatory profile of patients with tuberculosis with or without HIV-1 co-infection undergoing ATT, with specific focus on the effect of ART and HIV-1 viraemia in those co-infected with HIV-1. Methods In this prospective cohort study and immunological network analysis, a panel of 38 inflammatory markers were measured in the plasma of a prospective patient cohort undergoing ATT at Khayelitsha Site B clinic, Cape Town, South Africa. We recruited patients with sputum Xpert MTB/RIF-positive rifampicin-susceptible pulmonary tuberculosis. Patients were excluded from the primary discovery cohort if they were younger than 18 years, unable to commence ATT for any reason, pregnant, had unknown HIV-1 status, were unable to consent to study participation, were unable to provide baseline sputum samples, had more than three doses of ATT, or were being re-treated for tuberculosis within 6 months of their previous ATT regimen. Plasma samples were collected at baseline (1–5 days after commencing ATT), week 8, and week 20 of ATT. We applied network and multivariate analysis to investigate the dynamic inflammatory profile of these patients in relation to ATT and by HIV status. In addition to the discovery cohort, a validation cohort of patients with HIV-1 admitted to hospital with CD4 counts less than 350 cells per μL and a high clinical suspicion of new tuberculosis were recruited. Findings Between March 1, 2013, and July 31, 2014, we assessed a cohort of 129 participants (55 [43%] female and 74 [57%] male, median age 35·1 years [IQR 30·1–43·7]) and 76 were co-infected with HIV-1. HIV-1 status markedly influenced the inflammatory profile regardless of ATT duration. HIV-1 viral load emerged as a major factor driving differential inflammatory marker expression and having a strong effect on correlation profiles observed in the HIV-1 co-infected group. Interleukin (IL)-17A emerged as a key correlate of HIV-1-induced inflammation during HIV–tuberculosis co-infection. Interpretation Our findings show the effect of HIV-1 co-infection on the complexity of plasma inflammatory profiles in patients with tuberculosis. Through network analysis we identified IL-17A as an important node in HIV–tuberculosis co-infection, thus implicating this cytokine's capacity to correlate with, and regulate, other inflammatory markers. Further mechanistic studies are required to identify specific IL-17A-related inflammatory pathways mediating immunopathology in HIV–tuberculosis co-infection, which could illuminate targets for future host-directed therapies. Funding National Institutes of Health, The Wellcome Trust, UK Research and Innovation, Cancer Research UK, European and Developing Countries Clinical Trials Partnership, and South African Medical Research Council

Clinical, microbiologic, and immunologic determinants of mortality in hospitalized patients with HIV-associated tuberculosis: a prospective cohort study

Background: In high burden settings case fatality rates are reported to be between 11% and 32% in hospitalized patients with HIV-associated tuberculosis, yet the underlying causes of mortality remain poorly characterized. Understanding causes of mortality could inform development of novel management strategies to improve survival. We aimed to assess clinical and microbiologic determinants of mortality and to characterize the pathophysiological processes underlying death by evaluating host soluble inflammatory mediators and determined the relationship between these mediators and death as well as biomarkers of disseminated tuberculosis. Methods and Findings: Adult HIV-positive patients hospitalized with a new diagnosis of HIV-associated tuberculosis were enrolled in Cape Town between 2014-2016. Detailed tuberculosis diagnostic testing was performed. Biomarkers of tuberculosis dissemination and host soluble inflammatory mediators at baseline were assessed. Of 682 enrolled participants, 576 with tuberculosis (487/576, 84.5% microbiologically confirmed) were included in analyses. The median age was 37 years (IQR=31-43), 51.2% were female and the patients had advanced HIV with median CD4 count =58 cells/l (IQR= 21-120) and median HIV viral load=5.1 log10 copies/mL (IQR=3.3-5.7).Antituberculosis therapy was initiated in 566/576 (98.3%) and 487/576 (84.5%) started therapy within 48 hours of enrolment. Twelve-week mortality was 124/576 (21.5%) with 46/124 (37.1%) deaths occurring within 7 days of enrolment. Clinical and microbiologic determinants of mortality included disseminated tuberculosis (positive urine lipoarabinomannan, urine Xpert MTB/RIF or tuberculosis blood culture in 79.6% of deaths vs 60.7% of survivors, p=0.001), sepsis syndrome (high lactate in 50.8% of deaths vs 28.9% of survivors, p<0.001) and rifampicin resistant tuberculosis (16.9% of deaths vs 7.2% of survivors, p=0.002). Using non-supervised two-way hierarchical cluster and principal components analyses, we describe an immune profile dominated by mediators of the innate immune system and chemotactic signaling (IL-1Ra, IL-6, IL-8, MIP-1β/CCL4, IP-10/CXCL10, MIP-1α/CCL3) which segregated participants who died from those who survived. This immune profile was associated with mortality in a Cox proportional hazards model (adjusted hazard ratio = 2.2, 95%CI = 1.9-2.7, p<0.001) and with detection of biomarkers of disseminated tuberculosis. Clinicians attributed causes of death identified tuberculosis as a cause or one of the major causes of death in 89.5% of cases. We did not perform longitudinal sampling and did not have autopsy confirmed causes of death.Conclusions: In this study, we did not identify a major contribution from co-infections to these deaths. Disseminated tuberculosis, sepsis syndrome and rifampicin resistance were associated with mortality. An immune profile dominated by mediators of the innate immune system and chemotactic signaling was associated with both tuberculosis dissemination and mortality. These findings provide pathophysiologic insights into underlying causes of mortality and could be used to inform development of novel treatment strategies and develop methods to risk stratify patients to appropriately target novel interventions. Causal relationships cannot be established from this study

Inflammatory profile of HIV uninfected and co-infected patients with tuberculosis: a prospective cohort study and immunological network analysis

Background Human immunodeficiency virus type 1 (HIV-1) mediated dysregulation of the immune response to tuberculosis (TB) and its effect on the response to antitubercular treatment (ATT) is incompletely understood. Our aim was to perform an in-depth analysis of the inflammatory profile of HIV-1-uninfected and co-infected TB patients undergoing ATT, with sub-analysis of the effect of antiretroviral therapy and HIV-1 viraemia in the latter group. Methods An extensive panel of inflammatory markers were measured in plasma of a prospective patient cohort undergoing ATT at Khayelitsha Site B Clinic, South Africa between March 2013 and July 2014. Plasma samples were collected at baseline (1-5 days after commencing ATT), week 8 and week 20 of ATT. We applied network and multivariate analysis to investigate the dynamic inflammatory profile of these patients in relation to ATT and by HIV status. Findings HIV-1 status markedly influenced the inflammatory profile regardless of ATT duration. HIV-1 viral load emerged as a major factor driving differential inflammatory marker expression and impacting on correlation profiles observed in the HIV-1 co-infected group. Unexpectedly, IL-17A emerged as a key correlate of HIV-1 induced inflammation during HIV-TB co-infection. We validated these findings in a second cohort of hospitalized HIV-TB co-infected patients where the number of statistically significant correlations with IL-17A in network analysis predicted mortality. Interpretation Our findings demonstrate the effect of HIV-1 co-infection on the complexity of plasma inflammatory profiles in TB patients. Through network analysis we identified IL-17A as an important node in HIV-TB co-infection, thus implicating this cytokine’s capacity to correlate with, and regulate, other inflammatory markers. Further mechanistic studies are thus required to identify specific IL-17A related inflammatory pathways mediating immunopathology in HIV-TB co-infection, which may illuminate targets for future host directed therapies

Mycobacterium tuberculosis induction of heme Oxygenase-1 expression is Dependent on Oxidative stress and reflects Treatment Outcomes

The antioxidant enzyme heme oxygenase-1 (HO-1) is implicated in the pathogenesis of tuberculosis (TB) and has been proposed as a biomarker of active disease. Nevertheless, the mechanisms by which Mycobacterium tuberculosis (Mtb) induces HO-1 as well as how its expression is affected by HIV-1 coinfection and successful antitubercular therapy (ATT) are poorly understood. We found that HO-1 expression is markedly increased in rabbits, mice, and non-human primates during experimental Mtb infection and gradually decreased during ATT. In addition, we examined circulating concentrations of HO-1 in a cohort of 130 HIV-1 coinfected and uninfected pulmonary TB patients undergoing ATT to investigate changes in expression of this biomarker in relation to HIV-1 status, radiological disease severity, and treatment outcome. We found that plasma levels of HO-1 were elevated in untreated HIV-1 coinfected TB patients and correlated positively with HIV-1 viral load and negatively with CD4+ T cell count. In both HIV-1 coinfected and Mtb monoinfected patients, HO-1 levels were substantially reduced during successful TB treatment but not in those who experienced treatment failure or subsequently relapsed. To further delineate the molecular mechanisms involved in induction of HO-1 by Mtb, we performed a series of in vitro experiments using mouse and human macrophages. We found that Mtb-induced HO-1 expression requires NADPH oxidase-dependent reactive oxygen species production induced by the early-secreted antigen ESAT-6, which in turn triggers nuclear translocation of the transcription factor NRF-2. These observations provide further insight into the utility of HO-1 as a biomarker of both disease and successful therapy in TB monoinfected and HIV-TB coinfected patients and reveal a previously undocumented pathway linking expression of the enzyme with oxidative stress

Lipid mediators of inflammation and Resolution in individuals with tuberculosis and tuberculosis-Diabetes.

Individuals with concurrent tuberculosis (TB) and Type 2 diabetes (DM) have a higher risk of adverse outcomes. To better understand potential immunological differences, we utilized a comprehensive panel to characterize pro-inflammatory and pro-resolving (i.e., mediators involved in the resolution of inflammation) lipid mediators in individuals with TB and TB-DM. A nested cross-sectional study of 40 individuals (20 newly diagnosed DM and 20 without DM) was conducted within a cohort of individuals with active drug-susceptible treatment-naïve pulmonary TB. Lipid mediators were quantified in serum samples through lipid mediator profiling. We conducted correlation-based analysis of these mediators. Overall, the arachidonic acid-derived leukotriene and prostaglandin families were the most abundant pro-inflammatory lipid mediators, while lipoxins and maresins families were the most abundant pro-resolving lipid mediators in individuals with TB and TB-DM. Individuals with TB-DM had increased correlations and connectivity with both pro-inflammatory and pro-resolving lipid mediators compared to those with TB alone. We identified the most abundant lipid mediator metabolomes in circulation among individuals with TB and TB-DM; in addition, our data shows a substantial number of significant correlations between both pro-inflammatory and pro-resolving lipid mediators in individuals with TB-DM, delineating a molecular balance that potentially defines this comorbidity.This work was supported primarily by the United States National Institutes of Health (NIH), Bethesda, MD, USA (R01AI097494 to JG). Additional support for this work was obtained through Federal funds from the Government of India’s (GOI’s) Department of Biotechnology (DBT; New Delhi), the Indian Council of Medical Research (ICMR; New Delhi, India), the United States NIH, National Institute of Allergy and Infectious Diseases (NIAID), Office of AIDS Research (OAR), and distributed in part by CRDF Global (Arlington, VA, USA) (USB1-31147-XX13 CRDF/NIH to AG), and the NIH-funded Johns Hopkins Baltimore-Washington-India Clinical Trials Unit for NIAID Networks (U01AI069465 to VM, NG, AG). RS was supported by NIH/National Institute of Child Health and Human Development grant R00HD089753. RL was supported by the BJGMC JHU HIV TB Program funded by the Fogarty International Center, Bethesda, MD, USA (NIH grant D43TW009574). ANG was supported by NIH Research Training Grant # D43 TW009340 funded by the NIH Fogarty International Center, the National Institute of Neurological Disorders and Stroke, the National Institute of Mental Health, the National Heart, Lung, and Blood Institute and the National Institute of Environmental Health Sciences (Bethesda, MD, USA). BBA was supported by Intramural research program from FIOCRUZ and from the National Institutes of Health (U01AI115940) and NIH/CRDF RePORT International Supplemental Funds. BBA is a senior investigator from the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Brazil. KFF was supported by a postdoctoral fellowship from the CNPq, Brazil. MBA was supported by a PhD fellowship from the Fundação de Amparo à Pesquisa do Estado da Bahia (FAPESB). JD is funded by European Research Council under the European Union’s Horizon 2020 research and innovation program (Grant 677542), a Sir Henry Dale Fellowship jointly funded by the Wellcome Trust and the Royal Society (Grant 107613/Z/15/Z), and the Barts Charity (Grant MGU0343). The content of this paper is solely the responsibility of the authors and does not necessarily represent the official views of the funders