Histopathology of tumour associated sarcoid-like stromal reaction in breast cancer

Summary In 5 cases of invasive ductal and lobular carcinoma of the breast multiple epithelioid and giant cell containing granulomas were detected, localized mainly in circumferential regions, but also in the center of the carcinomas. These granulomas were interpreted as sarcoid-like stromal reactions, occurring as sarcoid-like lesions in uni- and bilateral primaries, in a recurrent tumour, and also in axillary lymph nodes. Histopathologically, these granulomas were not quite uniform, some of them corresponding to typical sarcoidosis, others showing marked proliferations of epithelioid or giant cells or containing fibrinoid exudate or necroses. The granulomas were surrounded by dense infiltrates of mononuclear cells. Tuberculosis and mycosis was excluded. There were no hints of generalized sarcoidosis. Pathogenetically, these are reactions in the tumour stroma of varying intensity, and are not caused by necroses of the tumour tissue nor by microbial infections. Such tumour-associated sarcoid-like stroma reactions are interpreted as a T-cell mediated immune response to an antigen expression of the carcinoma acting as the local trigger; in 2 cases they were connected with sarcoid-like lesions of the axillary lymph nodes. Their occurrence in bilateral carcinoma of the breast points to an immunological disposition for this special kind of host-versus-tumour response. The intensity of these changes in a recurrent tumour reflects an immunological hypersensitivity reaction.The pathogenetic and differential diagnostic aspects of epithelioid granulomas of the female breast in chronic granulomatous mastitis, panniculitis, foreign body reaction, rare infections, and in therapeutically induced sarcoidosis are described and discussed.Dedicated to Prof. Dr. K. Lennert, Kiel, in Honour of his 65th Birthday     

Heterogeneity in tuberculosis pathology,microenvironments and therapeutic responses

Tuberculosis (TB) lesions are extremely complex and dynamic. Here, we review the multiple types and fates of pulmonary lesions that form following infection by Mycobacterium tuberculosis and the impact of this spatial and temporal heterogeneity on the bacteria they harbor. The diverse immunopathology of granulomas and cavities generates a plethora of microenvironments to which M. tuberculosis bacilli must adapt. This in turn affects the replication, metabolism, and relative density of bacterial subpopulations, and consequently their respective susceptibility to chemotherapy. We outline recent developments that support a paradigm shift in our understanding of lesion progression. The simple model according to which lesions within a single individual react similarly to the systemic immune response no longer prevails. Host-pathogen interactions within lesions are a dynamic process, driven by subtle and local differences in signaling pathways, resulting in diverging trajectories of lesions within a single host. The spectrum of TB lesions is a continuum with a large overlap in the lesion types found in latently infected and active TB patients. We hope this overview will guide TB researchers in the design, choice of read-outs, and interpretation of future studies in the search for predictive biomarkers and novel therapies.     

Non-caseating granulomas in patients with hematologic malignancies. A report of three cases

Sarcoidosis or sarcoid-like lesions occurred in three patients with hematologic malignancies. In one patient, an overlap syndrome sarcoidosis/polycythemia vera was the most probable diagnosis. In the other two cases, the granulomas were probably secondary to myelofibrosis and Waldenstr?m's disease, respectively. Recent epidemiological studies have failed to show a causal relationship between sarcoidosis and malignancy. The co-existence of sarcoidosis and malignancy is probably fortuitous, but non-caseating granulomas may occur secondary to malignant diseases.     

Immune distribution and localization of phosphoantigen-specific Vgamma2Vdelta2 T cells in lymphoid and nonlymphoid tissues in Mycobacterium tuberculosis infection

Little is known about the immune distribution and localization of antigen-specific T cells in mucosal interfaces of tissues/organs during infection of humans. In this study, we made use of a macaque model of Mycobacterium tuberculosis infection to assess phosphoantigen-specific Vγ2Vδ2 T cells regarding their tissue distribution, anatomical localization, and correlation with the presence or absence of tuberculosis (TB) lesions in lymphoid and nonlymphoid organs/tissues in the progression of severe pulmonary TB. Progression of pulmonary M. tuberculosis infection generated diverse distribution patterns of Vγ2Vδ2 T cells, with remarkable accumulation of these cells in lungs, bronchial lymph nodes, spleens, and remote nonlymphoid organs but not in blood. Increased numbers of Vγ2Vδ2 T cells in tissues were associated with M. tuberculosis infection but were independent of the severity of TB lesions. In lungs with apparent TB lesions, Vγ2Vδ2 T cells were present within TB granulomas. In extrathoracic organs, Vγ2Vδ2 T cells were localized in the interstitial compartment of nonlymphoid tissues, and the interstitial localization was present despite the absence of detectable TB lesions. Finally, Vγ2Vδ2 T cells accumulated in tissues appeared to possess cytokine production function, since granzyme B was detectable in the γδ T cells present within granulomas. Thus, clonally expanded Vγ2Vδ2 T cells appeared to undergo trans-endothelial migration, interstitial localization, and granuloma infiltration as immune responses to M. tuberculosis infection.     

Distribution of antibody titres against phenolic glycolipids from Mycobacterium tuberculosis in the sera from tuberculosis patients and healthy controls

Sera from tuberculosis (TB) patients and healthy controls were tested by ELISA for their antibody titres against the two major phenolic glycolipids (PGLs) of Mycobacterium tuberculosis, PGL-tbO (a 1:3 mixture of PGL-tb1 and its analogue whose phthiocerol moiety is phenolphthiotriol A) and PGL-tbK. Both PGL-tbs were shown to be specific to M. tuberculosis, and the profiles of serum anti-PGL-tbK titres revealed that PGL-tbK, like PGL-tb1, was fairly widely distributed among strains of M. tuberculosis. Even when these two PGL-tbs were used, however, the rate of ELISA-positives was not very high among TB patients, which is probably explained by the nature of the disease. Moreover, a considerable number of sera from healthy controls, especially from younger age groups, had high anti-PGL-tb titres, which implies that environmental exposure to M. tuberculosis is much higher than has been estimated from the actual TB cases. The ELISA system using these species-specific PGL-tb antigens may be useful for the survey of TB infection, since it gives more direct information on TB infection than the PPD skin test.     

Be alert to tuberculosis-mediated glomerulonephritis: a retrospective study

Mycobacterium tuberculosis infection causing glomerulonephritis is a rare disorder. This retrospective study analyzed the clinical characteristics of patients diagnosed with tuberculosis-mediated glomerulonephritis (TB-GN) between 2002 and 2009, as well as the diagnostic tools used. These findings were then compared with those of patients with primary glomerulonephritis (P-GN). The records of all patients were reviewed. The diagnosis of TB-GN was based on renal hematuria and/or proteinuria and cure after antituberculosis therapy alone plus urine culture positive for M. tuberculosis, demonstration of typical tubercle granulomas on renal biopsy specimens, or the detection of M. tuberculosis DNA by polymerase chain reaction (PCR) on renal specimens. Forty-six patients with TB-GN and 49 patients with P-GN were included. Compared with patients in the P-GN group, most (76%) patients with TB-GN had a history of TB. Systemic symptoms were much more frequent in patients with TB-GN than local genitourinary symptoms. Serological testing showed a statistical difference between the two groups. Immunoglobulin A nephropathy was found in the majority (72%) of patients with TB-GN. M. tuberculosis DNA detection was positive in 39 (84.8%) patients, a much higher positive rate of diagnosis than that with urine culture for M. tuberculosis. The manifestation of TB-GN is atypical and nonspecific. It warrants a high index of suspicion when patients with renal hematuria and proteinuria fail to respond to standard treatments for P-GN. Clinicians should pay close attention to the medical history and results of special laboratory tests. M. tuberculosis DNA detection on renal biopsy specimens should be considered in order to confirm the diagnosis of TB-GN.     

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

CD8 T cells contribute to protective immunity against Mycobacterium tuberculosis. In humans, M. tuberculosis reactive CD8 T cells typically recognize peptides associated to classical MHC class Ia molecules, but little information is available on CD8 T cells recognizing M. tuberculosis Ags presented by nonclassical MHC class Ib molecules. We show here that CD8 T cells from tuberculosis (TB) patients recognize HLA‐E‐binding M. tuberculosis peptides in a CD3/TCR αβ mediated and CD8‐dependent manner, and represent an additional type of effector cells playing a role in immune response to M. tuberculosis during active infection. HLA‐E‐restricted recognition of M. tuberculosis peptides is detectable by a significant enhanced ex vivo frequency of tetramer‐specific circulating CD8 T cells during active TB. These CD8 T cells produce type 2 cytokines upon antigenic in vitro stimulation, help B cells for Ab production, and mediate limited TRAIL‐dependent cytolytic and microbicidal activity toward M. tuberculosis infected target cells. Our results, together with the finding that HLA‐E/M. tuberculosis peptide specific CD8 T cells are detected in TB patients with or without HIV coinfection, suggest that this is a new human T‐cell population that participates in immune response in TB.     

Mycobacterial DNA in recurrent sarcoidosis in the transplanted lung – a PCR-based study on four cases

Sarcoidosis is a systemic granulomatous inflammation, which may be caused by mycobacteria other than M. tuberculosis complex (MOTT) in one-third of cases. A few cases of recurrent sarcoidosis in the transplanted lung have been reported. However, mycobacteria have been excluded by acid-fast stains only. We investigated four cases of recurrent sarcoidosis in lung transplant patients. Using PCR for the insertion sequence 6110 of Mycobacterium tuberculosis complex and a second PCR for the mycobacterial chaperonin (65-kDa antigen coding sequence), we looked for mycobacterial DNA. In three cases sequence analysis was also performed. One patient was negative for mycobacterial DNA in explanted, but positive for M. tuberculosis DNA in transplanted lung, qualifying this case as M. tuberculosis infection in the transplant. Three patients were negative for M. tuberculosis DNA, but were positive for MOTT-DNA in both explanted and transplanted lungs. In these three patients sequence identity of the amplified sequences before and after transplantation was proven, which rules out mycobacteriosis. Recurrent sarcoidosis does occur, but can only be proven by the exclusion of mycobacterial DNA. In cases of recurrent MOTT-DNA-positive sarcoidosis the diagnosis cannot be confirmed except by proof of sequence identity. Probably MOTT-DNA-positive sarcoidosis is more likely to recur in a transplanted lung. Received: 19 August 1999 / Accepted: 11 November 1999     

Revisiting the Natural History of Tuberculosis

Once Mycobacterium tuberculosis infects a person it can persist for a long time in a process called latent tuberculosis infection (LTBI). LTBI has traditionally been considered to involve the bacilli remaining in a non-replicating state (dormant) in old lesions but still retaining their ability to induce reactivation and cause active tuberculosis (TB) once a disruption of the immune response takes place. The present review aims to challenge these concepts by including recent experimental data supporting LTBI as a constant endogenous reinfection process as well as the recently introduced concepts of damage-response and tolerance frameworks to explain TB induction. These frameworks highlight the key role of an exaggerated and intolerant host response against M. tuberculosis bacilli which induces the classical TB cavity in immunocompetent adults once the constant endogenous reinfection process has resulted in the presence of bacilli in the upper lobes, where they can grow faster and the immune response is delayed. This essay intends to provide new clues to understanding the induction of TB in non-immunosuppressed patients.     

Exosomes carrying mycobacterial antigens can protect mice against Mycobacterium tuberculosis infection

Approximately 2 billion people are infected with Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), and an estimated 1.5 million individuals die annually from TB. Presently, Mycobacterium bovis BCG remains the only licensed TB vaccine; however, previous studies suggest its protective efficacy wanes over time and fails in preventing pulmonary TB. Therefore, a safe and effective vaccine is urgently required to replace BCG or boost BCG immunizations. Our previous studies revealed that mycobacterial proteins are released via exosomes from macrophages infected with M. tuberculosis or pulsed with M. tuberculosis culture filtrate proteins (CFP). In the present study, exosomes purified from macrophages treated with M. tuberculosis CFP were found to induce antigen‐specific IFN‐γ and IL‐2‐expressing CD4⁺ and CD8⁺ T cells. In exosome‐vaccinated mice, there was a similar T_H1 immune response but a more limited T_H2 response compared to BCG‐vaccinated mice. Using a low‐dose M. tuberculosis mouse aerosol infection model, exosomes from CFP‐treated macrophages were found to both prime a protective immune response as well as boost prior BCG immunization. The protection was equal to or superior to BCG. In conclusion, our findings suggest that exosomes might serve as a novel cell‐free vaccine against an M. tuberculosis infection.     

T regulatory cells and Th1/Th2 cytokines in peripheral blood from tuberculosis patients

About 10% of people infected with Mycobacterium tuberculosis develop active tuberculosis (TB), and Th1 effector cells and Th1 cytokines play key roles in controlling M. tuberculosis infection. Here, we hypothesise that this susceptibility to M. tuberculosis infection is linked to increased T regulatory (Treg) cells and Th2 cytokines in TB patients. To test this, we recruited 101 participants (71 TB patients, 12 non-TB pulmonary diseases and 18 healthy subjects) and investigated Treg cells and Th1/Th2 cytokines in peripheral blood. CD4⁺CD25⁺ T cells and CD4⁺CD25⁺FoxP3⁺ T cells significantly increased and IL-5 dramatically decreased in TB patients relative to healthy subjects. CD8⁺CD28⁻ T cells, IFN-γ, TNF-α, IL-10 and IL-4 significantly increased in patients with culture and sputum smear-positive pulmonary TB (PTB(+)) compared with healthy subjects. CD4⁺CD25⁺FoxP3⁺ and CD8⁺CD28⁻ T cells significantly decreased in PTB(+) after one month of chemotherapy. CD4⁺, CD4⁺CD25⁺ and CD8⁺CD28⁺ T cells significantly increased in extra-pulmonary TB patients after one month of chemotherapy. These findings suggest that M. tuberculosis infection induces circulating CD4⁺CD25⁺FoxP3⁺ and CD8⁺CD28⁻ T cell expansion, which may be related to the progression of M. tuberculosis infection, and that the balance between effector immune responses and suppression immune responses is essential to control M. tuberculosis infection.     

Different types of pulmonary granuloma necrosis in immunocompetent vs. TNFRp55-gene-deficient mice aerogenically infected with highly virulent Mycobacterium avium

The immunopathogenesis of mycobacterial infections frequently involves the formation of caseating granulomas which cause tissue destruction and, in the case of tuberculosis (TB), may lead to cavity formation. Both intravenous and aerosol models of Mycobacterium tuberculosis infection in mice do not reflect the pulmonary lesions characteristic of TB patients. Using both low-dose (10² colony-forming units, cfu) and high-dose (10⁵ cfu) aerosol infection with a highly virulent strain of Mycobacterium avium (TMC724) in C57BL/6 mice, it is now shown that these mice are capable of developing centrally caseating necrosis in lung granulomas after approximately 4 months of infection. In contrast, mice infected intravenously with the high dose never developed this type of lesion, although bacterial counts in their lungs reached levels comparable to those attained by aerosol-infected mice (10¹⁰ cfu). To study the relevance of events signalled by tumour necrosis factor (TNF) in this model, TNFRp55 gene-deficient and syngeneic C57BL/6 immunocompetent mice were infected with 10⁵ cfu M. avium via aerosol. In gene-deficient mice, newly formed pulmonary granulomas acutely disintegrated, showing signs of apoptotic cell death and neutrophil influx, and TNFRp55 knock-out mice all succumbed to infection just beyond the stage of granuloma initiation. Aerogenic infection with M. avium in mice is a suitable model to study the immunopathogenesis of granuloma necrosis because it closely mimicks the histopathology of mycobacterial infections in humans, including TB. Furthermore, the use of TNFRp55 gene-deficient mice in this model establishes a role for TNF in maintaining the integrity of a developing pulmonary granuloma. Copyright © 1999 John Wiley & Sons, Ltd.     

Influence of diabetes mellitus on immunity to human tuberculosis

Type 2 diabetes mellitus(DM) is a major risk factor for the development of active pulmonary tuberculosis (TB), with development of DM pandemic in countries where TB is also endemic. Understanding the impact of DM on TB and the determinants of co‐morbidity is essential in responding to this growing public health problem with improved therapeutic approaches. Despite the clinical and public health significance posed by the dual burden of TB and DM, little is known about the immunological and biochemical mechanisms of susceptibility. One possible mechanism is that an impaired immune response in patients with DM facilitates either primary infection with Mycobacterium tuberculosis or reactivation of latent TB. Diabetes is associated with immune dysfunction and alterations in the components of the immune system, including altered levels of specific cytokines and chemokines. Some effects of DM on adaptive immunity that are potentially relevant to TB defence have been identified in humans. In this review, we summarize current findings regarding the alterations in the innate and adaptive immune responses and immunological mechanisms of susceptibility of patients with DM to M. tuberculosis infection and disease.     

Neutrophils express pro- and anti-inflammatory cytokines in granulomas from Mycobacterium tuberculosis-infected cynomolgus macaques

Neutrophils are implicated in the pathogenesis of tuberculosis (TB), a disease caused by Mycobacterium tuberculosis infection, but the mechanisms by which they promote disease are not fully understood. Neutrophils can express cytokines that influence TB progression, and so we compared neutrophil and T-cell expression of the Th1 cytokines IFNγ and TNF, the Th2 cytokine IL-4, and regulatory cytokine IL-10 in M. tuberculosis-infected macaques to determine if neutrophil cytokine expression contributes to dysregulated immunity in TB. We found that peripheral blood neutrophils produced cytokines after stimulation by mycobacterial antigens and inactive and viable M. tuberculosis. M. tuberculosis antigen-stimulated neutrophils inhibited antigen-specific T-cell IFNγ production. In lung granulomas, neutrophil cytokine expression resembled T-cell cytokine expression, and although there was histologic evidence for neutrophil interaction with T cells, neutrophil cytokine expression was not correlated with T-cell cytokine expression or bacteria load. There was substantial overlap in the spatial arrangement of cytokine-expressing neutrophils and T cells, but IL-10-expressing neutrophils were also abundant in bacteria-rich areas between caseum and epithelioid macrophages. These results suggest that neutrophils contribute to the cytokine milieu in granulomas and may be important immunoregulatory cells in TB granulomas.     

Old age-associated enrichment of peripheral T regulatory cells and altered redox status in pulmonary tuberculosis patients

Aging influences the susceptibility and prognosis to various infectious diseases including tuberculosis (TB). Despite the impairment of T-cell function and immunity in older individuals, the mechanism for the higher incidence of TB in the elderly remains largely unknown. Here, we evaluated the age-associated immune alterations, particularly in effector and Treg responses in pulmonary TB patients. We also evaluated the impact of redox status and its modulation with N-acetyl-cysteine (NAC) in elderly TB. Higher frequency of Treg cells and reduced IFN-γ positive T cells were observed among older TB patients. The elevated number of Treg cells correlated tightly with bacillary load (i.e. disease severity); which declined significantly in response to successful anti-tubercular treatment. We could rescue Myobacterium tuberculosis-specific effector T cell (Th1) responses through various in vitro approaches, for example, Treg cell depletion and co-culture experiments, blocking experiments using antibodies against IL-10, TGF-β, and programmed death-1 (PD-1) as well as NAC supplementation. We report old age-associated enrichment of Treg cells and suppression of M. tuberculosis-specific effector T (Th1) cell immune responses. Monitoring these immune imbalances in older patients may assist in immune potentiation through selectively targeting Treg cells and/or optimizing redox status by NAC supplementation.     

Inflammation and the coagulation system in tuberculosis: Tissue Factor leads the dance

Mycobacterium tuberculosis, the causative agent of tuberculosis, drives the formation of granulomas, structures in which both immune cells and the bacterial pathogen cohabit. The most abundant cells in granulomas are macrophages, which contribute as both cells with bactericidal activity and as targets for M. tuberculosis infection and proliferation during the entire course of infection. The mechanisms and factors involved in the regulation and control of macrophage microenvironment‐specific polarization and plasticity are not well understood, as some granulomas are able to control bacteria growth and others fail to do so, permitting bacterial spread. In this issue of the European Journal of Immunology, Venkatasubramanian et al. [Eur. J. Immunol. 2016. 46: 464–479] show that mice lacking the tissue factor gene in myeloid cells have augmented M. tuberculosis growth and increased inflammation in the lungs. This suggests that tissue factor, an initiator of coagulation, is important for the generation of fibrin, which supports granuloma formation. This article demonstrates for the first time the involvement of tissue factor in inducing effective immunity against M. tuberculosis, and sheds new lights on the complex interplay between host inflammatory response, the coagulation system, and the control of M. tuberculosis infection.     

Robust immune response elicited by a novel and unique Mycobacterium tuberculosis protein using an optimized DNA/protein heterologous prime/boost protocol

An efficacious tuberculosis (TB) vaccine will probably need to induce both CD4 and CD8 T‐cell responses specific to a protective Mycobacterium tuberculosis antigen(s). To achieve this broad cellular immune response we tested a heterologous DNA/protein combination vaccine strategy. We used a purified recombinant protein preparation of a unique M. tuberculosis antigen (rMT1721) found in the urine of TB patients, an optimized plasmid DNA expressing this protein (DNA‐MT1721), and a Toll‐like receptor 4 agonist adjuvant. We found that priming mice with DNA‐MT1721 and subsequently boosting with rMT1721 elicited high titres of specific IgG1 and IgG2a antibodies as well as high magnitude and polyfunctional CD4⁺ T‐cell responses. However, no detectable CD8⁺ T‐cell response was observed using this regimen of immunization. In contrast, both CD4⁺ and CD8⁺ T‐cell responses were detected after a prime/boost vaccination regimen using rMT1721 as the priming antigen and DNA‐MT1721 as the boosting immunogen. These findings support the exploration of heterologous DNA/protein immunization strategies in vaccine development against TB and possibly other infectious diseases.     

Results from 5 Years of Nationwide DNA Fingerprinting of Mycobacterium tuberculosis Complex Isolates in a Country with a Low Incidence of M. tuberculosis Infection

Results from DNA fingerprint analyses of Mycobacterium tuberculosis complex isolates from tuberculosis (TB) patients diagnosed during 5 years in Denmark are presented. The lack of success in eradicating TB in this low-incidence country may be explained by an unrecognized high frequency of active TB transmission (57%) among native Danes. Only two strains of M. tuberculosis are responsible for 40% of all clustered cases of TB among Danes.     

Drug resistance and genotypic analysis of Mycobacterium tuberculosis strains from Thai tuberculosis patients

The aim of this study was the molecular characterization of primary drug‐resistant Mycobacterium tuberculosis strains in Thailand. We examined a group of M. tuberculosis isolates from newly registered tuberculosis (TB) cases, collected at the largest university hospital, the Siriraj Hospital, in Thailand. Of 76 selected drug‐resistant M. tuberculosis strains recovered from previously untreated pulmonary TB patients whose sputum samples were sent to this hospital, 29 (38%) were single‐drug resistant, 26 (34%) multidrug resistant and two (2.6%) extensively drug resistant. Fifty (66%) strains belonged to Beijing genotype. The study demonstrate a severe problem of drug resistance among recently detected TB patients, and two large clusters of genetically similar strains indicated ongoing transmission of drug‐resistant TB.