Htin Aung

Impact of tuberculosis (TB) on HIV-1 activity in dually infected patients

Active TB in HIV‐1‐infected subjects is associated with increased HIV‐1‐related immunodeficiency and mortality. We assessed plasma viral load in HIV‐1‐infected patients with pulmonary TB (HIV/TB) and non‐TB symptomatic HIV‐1‐infected patients (HIV). HIV‐1 load was higher in HIV/TB compared with HIV at higher CD4 counts (> 500/μl) (P < 0·01), but not at lower CD4 counts (< 500/μl). We also evaluated the status of HIV‐1 gene expression in peripheral blood mononuclear cells (PBMC) and serum from HIV/TB and CD4‐matched healthy HIV‐infected patients (HIV/C) by reverse transcriptase‐polymerase chain reaction over a range of CD4 (> 900/μl to < 200/μl). HIV‐1 RNA in serum and PBMC correlated to one another, and both were markedly higher in HIV/TB compared with HIV/C with higher CD4 counts. Also, during a longitudinal study of anti‐tuberculous chemoprophylaxis in HIV‐1‐infected patients, 10 subjects who developed TB had serologies before, at the time, and after the diagnosis of TB. These HIV/TB patients had an increase in viral load (average 2·5‐fold) at the time of diagnosis of TB (P < 0·05). Overall, these data indicate that the transcriptional activity of HIV‐1 is enhanced in HIV‐1‐infected patients with active TB, especially during early HIV‐1 disease. As TB often is an early HIV‐1 opportunistic infection, it may particularly favour early viral replication and dissemination, and therefore contribute to progression of HIV‐1 disease.

Increased Replication of HIV-1 at Sites of Mycobacterium tuberculosis Infection: Potential Mechanisms of Viral Activation

&NA; Tuberculosis (TB) enhances HIV‐1 replication and the progression to AIDS in dually infected patients. We employed pleural TB as a model to understand the interaction of the host with HIV‐1 during active TB, at sites of Mycobacterium tuberculosis (MTB) infection. HIV‐1 replication was enhanced both in the cellular (pleural compared with blood mononuclear cells) and acellular (pleural fluid compared with plasma) compartments of the pleural space. Several potential mechanisms for expansion of HIV‐1 in situ were found, including augmentation in expression of tumor necrosis factor (TNF)‐&agr; and the HIV‐1 noninhibitory &bgr;‐chemokine (MCP‐1), low presence of HIV‐1 inhibitory &bgr;‐chemokines (MIP‐1&agr;, MIP‐1&bgr;, and RANTES [regulated on activation, normal T expressed and secreted]), and upregulation in expression of the HIV‐1 coreceptor, CCR5, by pleural fluid mononuclear cells. Thus, at sites of MTB infection, conditions are propitious both for transcriptional activation cf HIV‐1 in latently infected mononuclear cells, and facilitation of viral infection of newly recruited cells. These mechanisms may contribute to enhanced viral burden and dissemination during TB infection.

Induction of monocyte expression of tumor necrosis factor alpha by the 30-kD alpha antigen of Mycobacterium tuberculosis and synergism with fibronectin.

Native 30-kD antigen, also known as alpha antigen, is a fibronectin-binding protein that is secreted by live Mycobacterium tuberculosis. This antigen may play an important biological role in the host-parasite interaction since it elicits delayed type hypersensitivity response and protective immunity in vivo and T lymphocyte blastogenesis and IFN-gamma production in vitro. In the present study, we show that, TNF-alpha protein is produced in monocyte culture supernatants in response to 30-kD antigen and the level is as high as that to purified protein derivative of M. tuberculosis. This stimulatory effect was not due to contamination with either bacterial lipopolysaccharide or mycobacterial lipoarabinomannan. The preincubation of monocytes with plasma fibronectin significantly enhanced the release of TNF-alpha into the culture supernatants in response to 30-kD antigen. This effect was blocked by polygonal antibody to plasma fibronectin. In contrast, the monocytic cell line U937 failed to release TNF-alpha protein in the culture supernatants in response to 30-kD antigen with or without preincubation with plasma fibronectin. To determine whether this observation was due to differential binding of the 30-kD to fibronectin on these cells, a cell based ELISA was used. Pretreatment of monocytes with fibronectin enhanced their binding of the 30-kD antigen. U937 cells bound the 30-kD antigen weakly with or without fibronectin pretreatment. These results indicate that 30-kD antigen which is a known secretary antigen of M. tuberculosis is a stimulus for human monocytes to express TNF-alpha and that stimulatory effect may be mediated through plasma fibronectin.

Circulating CXCR5+PD-1+ Response Predicts Influenza Vaccine Antibody Responses in Young Adults but not Elderly Adults

Although influenza vaccination is recommended for all adults annually, the incidence of vaccine failure, defined as weak or absent increase in neutralizing Ab titers, is increased in the elderly compared with young adults. The T follicular helper cell (Tfh) subset of CD4 T cells provides B cell help in germinal centers and is necessary for class-switched Ab responses. Previous studies suggested a role for circulating Tfh cells (cTfh) following influenza vaccination in adults, but cTfh have not been studied in elderly adults in whom weak vaccine responses are often observed. In this study, we studied cTfh expressing CXCR5 and programmed death-1 (PD-1). cTfh from elderly adults were present at reduced frequency, had decreased in vitro B cell help ability, and had greater expression of ICOS compared with young adults. At 7 d after inactivated influenza vaccination, cTfh correlated with influenza vaccine–specific IgM and IgG responses in young adults but not in elderly adults. In sum, we have identified aging-related changes in cTfh that correlated with reduced influenza vaccine responses. Future rational vaccine design efforts should incorporate Tfh measurement as an immune correlate of protection, particularly in the setting of aging.

Activation of β-chemokines and CCR5 in persons infected with human immunodeficiency virus type 1 and tuberculosis

Tuberculosis (TB) in human immunodeficiency virus type 1 (HIV-1)-infected persons is associated with progression of HIV-1 disease. The expression of macrophage inflammatory protein (MIP)-1alpha and CCR5 was assessed in HIV-1-infected patients with pulmonary TB (HIV-1/PTB) and without PTB (HIV-1/C), PTB patients not infected with HIV-1 (PTB), and control subjects. Mycobacterium tuberculosis (MTB)-induced MIP-1alpha production was lower in peripheral blood mononuclear cells (PBMC) of HIV-1/PTB patients than in those of PTB patients (P< .05) and was lower in PBMC of HIV-1/C patients than in those of control subjects (P< .005). However, MIP-1alpha production was higher in PBMC of HIV/PTB patients than in those of HIV-1/C patients (P< .01). The pattern of MTB-induced RANTES production was similar to that of MIP-1alpha. However, MTB induced greater expression of mRNA for CCR5 in PBMC of HIV-1/PTB patients than in those of HIV-1/C patients (P< .04). Furthermore, the MTB-induced HIV p24 antigen level in PBMC of HIV-1/PTB patients with a CD4 cell count <500 cells/microL was higher (P< .05) than that in HIV-1/C patients. Thus, perturbations in chemokine pathways in HIV-1/PTB patients may accelerate HIV-1 disease.

Macrophage-Activating Cytokines in Human Immununodeficiency Virus Type 1–Infected and –Uninfected Patients with Pulmonary Tuberculosis

Tuberculosis (TB) is the most common opportunistic infection in human immunodeficiency virus type 1 (HIV-1)-infected patients globally and occurs throughout the course of HIV-1 disease. Here the production of interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha by peripheral blood mononuclear cells (PBMC) of HIV-1-infected versus -uninfected patients with newly diagnosed pulmonary TB (PTB) was compared. Findings were correlated with cytokine profiles, clinical presentation, and expression of inducible nitric oxide (iNOS). Most HIV-1/PTB patients with a CD4 cell count of 200-500 cells/microL had high IFN-gamma production and radiographic evidence of atypical PTB. Low IFN-gamma production and radiographic evidence of reactivated PTB characterized both HIV-1/PTB patients with a CD4 cell count >or=500 cells/microL and HIV-1-uninfected patients. TNF-alpha levels were similar in all HIV-1/PTB patients, regardless of CD4 cell count. Induction of iNOS in PBMC was low and was associated with low IFN-gamma production. These data underscore the potential pathogenic role of macrophage-activating cytokines in TB in HIV-1-infected patients.

Persistent Replication of Human Immunodeficiency Virus Type 1 despite Treatment of Pulmonary Tuberculosis in Dually Infected Subjects

ABSTRACT Tuberculosis (TB) is the most common life-threatening infection in human immunodeficiency virus (HIV)-infected persons and frequently occurs before the onset of severe immunodeficiency. Development of TB is associated with increased HIV type 1 (HIV-1) viral load, a fall in CD4 lymphocyte counts, and increased mortality. The aim of this study was to examine how treatment of pulmonary TB affected HIV-1 activity in HIV-1/TB-coinfected subjects with CD4 cell counts of >100 cells/μl. HIV-1/TB-coinfected subjects were recruited in Kampala, Uganda, and were monitored over time. Based upon a significant (0.5 log10 copies/ml) decrease in viral load by the end of treatment, two patient groups could be distinguished. Responders (n = 17) had more rapid resolution of anemia and pulmonary lesions on chest radiography during TB treatment. This group had a significant increase in viral load to levels not different from those at baseline 6 months after completion of TB treatment. HIV-1 viral load in nonresponders (n = 10) with TB treatment increased and at the 6 month follow-up was significantly higher than that at the time of diagnosis of TB. Compared to baseline levels, serum markers of macrophage activation including soluble CD14 decreased significantly by the end of TB treatment in responders but not in nonresponders. These data further define the impact of pulmonary TB on HIV-1 disease. HIV-1 replication during dual HIV-1/TB infection is not amenable to virologic control by treatment of TB alone. Concurrent institution of highly active antiretroviral treatment needs to be evaluated in patients dually infected with pulmonary TB and HIV-1.

Bioactivation of Latent Transforming Growth Factor β1 by Mycobacterium tuberculosis in Human Mononuclear Phagocytes

Biologically active transforming growth factor beta 1 (TGFβ1) has been identified at sites of Mycobacterium tuberculosis (MTB) infection in the lung; however, the underlying mechanism(s) for its activation is not clear. Here using an enzyme‐linked immunospot assay for TGFβ1, we show that human blood monocytes (MN) and alveolar macrophages (AM) produce bioactive TGFβ1 upon stimulation by MTB. However, only MTB‐stimulated MN increased TGFβ1 production on a per cell basis. The frequency of TGFβ1‐producing MN was reduced by an inhibitor of plasmin, bdellin, indicating a role for plasmin pathways in the bioactivation of cytokine. The expression of urokinase plasminogen activator receptor (uPAR) mRNA and both surface and soluble uPAR (CD87) was increased in MTB‐activated MN. However, antibody neutralization of uPAR suppressed bioactive TGFβ1 in MN alone. Thus, the more immature MN, which are continuously recruited to the lung during tuberculosis (TB), have a higher capacity to bioactivate TGFβ1 by expression of components of the plasmin pathway. Excess production and bioactivation of TGFβ1 at sites of MTB infection may undermine host immune responses during TB.

Role of Cellular Activation and Tumor Necrosis Factor—α in the Early Expression of Mycobacterium tuberculosis 85B mRNA in Human Alveolar Macrophages

BACKGROUND Infection of alveolar macrophages (AMs), which constitute the first line of defense against Mycobacterium tuberculosis, initiates an intense interaction between the hosts innate immune response and mycobacteria that may assist in the successful intracellular parasitism of M. tuberculosis. METHODS Expression of tumor necrosis factor (TNF)- alpha and M. tuberculosis 85B mRNA was studied in M. tuberculosis-infected AMs, to better delineate the role of macrophages in the early events in initiation of infection. RESULTS Both TNF- alpha mRNA and M. tuberculosis 85B were induced in AMs; at 24 h, the time point of maximum TNF- alpha induction, the mRNA levels for TNF- alpha and M. tuberculosis 85B correlated with one another, and induction of either gene correlated strongly with their protein levels. Inhibition of endogenous TNF- alpha by soluble (s) TNF receptor (R) I and sTNFRII reduced expression of both TNF- alpha and M. tuberculosis 85B. The activation of nuclear factor- kappa B was found to underlie expression of both TNF- alpha and M. tuberculosis 85B. Exogenous TNF- alpha was slightly more potent than interleukin (IL)-6 and granulocyte-macrophage colony-stimulating factor and was significantly stronger than IL-1 in inducing expression of M. tuberculosis 85B. Interestingly, inhibition of bactericidal mediators, reactive oxygen intermediates (ROIs) and reactive nitrogen intermediates (RNIs), reduced expression of TNF- alpha and M. tuberculosis 85B genes in M. tuberculosis-infected AMs. CONCLUSION Activation of AMs by M. tuberculosis initiates a cascade of events whereby TNF- alpha, ROI, and RNI enhance the expression of the M. tuberculosis 85B gene.

Polyfunctional Mycobacterium tuberculosis-specific effector memory CD4+ T cells at sites of pleural TB

Pleural tuberculosis (TB) is a common presentation of Mycobacterium tuberculosis (MTB) infection, and despite spontaneous resolution remains a strong risk factor for reactivation pulmonary TB in a majority of individuals. This study was undertaken to further understand the characteristics of immune cells at sites of pleural TB. A significant shift toward memory CD4+ T cells with an effector phenotype and away from naïve CD4+ T cells in pleural fluid as compared to blood mononuclear cells was found. These data suggest that effector T cells are capable of migrating to sites of active TB infection and/or the differentiation to effector phenotype T cells in situ is highly amplified. Using multi-parameter flow cytometry analysis, a significant portion of MTB-specific CD4+ T cells in the pleural space were polyfunctional demonstrating two, three or four simultaneous functions including IFN-gamma, IL-2, TNF-alpha, and or MIP-1 alpha production. A greater proportion of these polyfunctional cells were of effector memory rather than central memory phenotype. The role of these polyfunctional MTB-specific CD4+ T cells at sites of pleural TB requires further study.