Kenneth J. Gollob

Up-regulation of Th1-type responses in mucosal leishmaniasis patients.

ABSTRACT The cytokine profile produced by peripheral blood mononuclear cells (PBMC) in response to leishmania antigens and the ability of interleukin-10 (IL-10) and transforming growth factor β (TGF-β) to modulate the immune response were evaluated in 21 mucosal leishmaniasis patients. Patients with mucosal disease exhibited increased gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) secretion and decreased IL-10 secretion compared to patients with classical cutaneous leishmaniasis. CD4+ Th1 cells were the main source of IFN-γ and TNF-α production in mucosal leishmaniasis patients. Evaluation of cytokine gene expression in PBMC of these patients showed that there was strong up-regulation of IFN-γ transcripts upon stimulation with leishmania antigen, in contrast to the baseline levels of IL-10 mRNA. IL-10 suppressed IFN-γ production by 48% in cell cultures from cutaneous leishmaniasis patients and by 86% in cell cultures from healthy subjects stimulated with purified protein derivative, whereas in similar conditions IL-10 suppressed IFN-γ production by 19% in cell cultures from mucosal leishmaniasis patients stimulated with leishmania antigen. TGF-β suppressed IFN-γ levels to a greater extent in healthy subjects than in mucosal leishmaniasis and cutaneous leishmaniasis patients. These data indicate that a poorly modulated T-cell response in mucosal leishmaniasis patients leads to production of high levels of proinflammatory cytokines, such as IFN-γ and TNF-α, as well as a decreased ability of IL-10 and TGF-β to modulate this response. These abnormalities may be the basis for the pathological findings observed in this disease.

Decreased In Situ Expression of Interleukin-10 Receptor Is Correlated with the Exacerbated Inflammatory and Cytotoxic Responses Observed in Mucosal Leishmaniasis

ABSTRACT Human infection with Leishmania braziliensis can lead to cutaneous leishmaniasis (CL) or mucosal leishmaniasis (ML). We hypothesize that the intense tissue destruction observed in ML is a consequence of an uncontrolled exacerbated inflammatory immune response, with cytotoxic activity. For the first time, this work identifies the cellular sources of inflammatory and antiinflammatory cytokines, the expression of effector molecules, and the expression of interleukin-10 (IL-10) receptor in ML and CL lesions by using confocal microscopy. ML lesions displayed a higher number of gamma interferon (IFN-γ)-producing cells than did CL lesions. In both ML and CL, CD4+ cells represented the majority of IFN-γ-producing cells, followed by CD8+ cells and CD4− CD8− cells. The numbers of tumor necrosis factor alpha-positive cells, as well as those of IL-10-producing cells, were similar in ML and CL lesions. The effector molecule granzyme A showed greater expression in ML than in CL lesions, while inducible nitric oxide synthase did not. Finally, the expression of IL-10 receptor was lower in ML than in CL lesions. Thus, our data identified distinct cytokine and cell population profiles for CL versus ML patients and provide a possible mechanism for the development of ML disease through the demonstration that low expression of IL-10 receptor is present in conjunction with a cytotoxic and inflammatory profile in ML.

Flow Cytometric Determination of Cellular Sources and Frequencies of Key Cytokine-Producing Lymphocytes Directed against Recombinant LACK and Soluble Leishmania Antigen in Human Cutaneous Leishmaniasis

ABSTRACT Leishmaniasis, caused by infection with the protozoan parasiteLeishmania, affects millions of individuals worldwide, causing serious morbidity and mortality. This study directly determined the frequency of cells producing key immunoregulatory cytokines in response to the recombinant antigen Leishmania homolog of receptors for activated kinase C (LACK) and soluble leishmania antigen (SLA), and it determined relative contributions of these antigens to the overall cytokine profile in individuals infected for the first time with Leishmania braziliensis. All individuals presented with the cutaneous clinical form of leishmaniasis and were analyzed for proliferative responses to LACK antigen and SLA, frequency of lymphocyte subpopulations (analyzed ex vivo), and antigen-induced (LACK and SLA) cytokine production at the single-cell level (determined by flow cytometry). The following were determined. (i) The Th1-type response previously seen in patients with cutaneous leishmaniasis is due to gamma interferon (IFN-γ) production by several different sources, listed in order of contribution: CD4+ T lymphocytes, CD4−, CD8− lymphocytes, and CD8+ T lymphocytes. (ii) SLA induced a higher frequency of lymphocytes producing IFN-γ and tumor necrosis factor alpha (TNF-α) than did LACK. (iii) LACK induced an activation of monocyte populations as reflected by an increased percentage of CD14-positive cells. (iv) Neither SLA nor LACK induced detectable frequencies of cells producing interleukin-4 (IL-4) or IL-5. These data demonstrated a multifaceted immune response to SLA in human leishmaniasis involving Th1 CD4+ T lymphocytes (IFN-γ+ and IL-10−/IL-4−), Tc1 CD8+ T cells (IFN-γ+, and IL-10−/IL-4−), and a high frequency of TNF-α-producing lymphocytes. Moreover, it was determined that the recombinant antigen LACK acts as a weak inducer of Th1-type lymphocyte responses compared to SLA.

Current understanding of immunity to Trypanosoma cruzi infection and pathogenesis of Chagas disease.

Chagas disease caused by Trypanosoma cruzi remains an important neglected tropical disease and a cause of significant morbidity and mortality. No longer confined to endemic areas of Latin America, it is now found in non-endemic areas due to immigration. The parasite may persist in any tissue, but in recent years, there has been increased recognition of adipose tissue both as an early target of infection and a reservoir of chronic infection. The major complications of this disease are cardiomyopathy and megasyndromes involving the gastrointestinal tract. The pathogenesis of Chagas disease is complex and multifactorial involving many interactive pathways. The significance of innate immunity, including the contributions of cytokines, chemokines, reactive oxygen species, and oxidative stress, has been emphasized. The role of the components of the eicosanoid pathway such as thromboxane A2 and the lipoxins has been demonstrated to have profound effects as both pro- and anti-inflammatory factors. Additionally, we discuss the vasoconstrictive actions of thromboxane A2 and endothelin-1 in Chagas disease. Human immunity to T. cruzi infection and its role in pathogen control and disease progression have not been fully investigated. However, recently, it was demonstrated that a reduction in the anti-inflammatory cytokine IL-10 was associated with clinically significant chronic chagasic cardiomyopathy.

Current concepts in immunoregulation and pathology of human Chagas disease.

Purpose of review Chagas disease is a complex ailment caused by infection with Trypanosoma cruzi. It afflicts millions in Latin America. Years of studies have focused on the development of pathology in Chagas disease and recent studies have helped us understand the cellular mechanisms behind differential clinical evolution of Chagas disease. Recent findings We discuss recent findings concerning the cellular immune response in human Chagas disease focusing on immunoregulation and the development of pathology. We seek to put several findings into the context of a disease that initially controls an extreme and patent infection, and later progresses to a chronic phase marked by the presence (cardiac and digestive forms), or not (indeterminate form), of associated pathology. Summary Several theories exist to explain differential clinical evolution of Chagas disease. A coherent understanding of these theories will certainly aid in determining what combination of them approximates the true development of chagasic pathology. For achieving the goal of developing a successful therapy or intervention, it is critical that no theory be excluded at this point, but. Rather, rather that a thoughtful analysis and assimilation of the best components of each system into a central theory that best fits the reality of human Chagas disease is desirable.

Exacerbated inflammatory cellular immune response characteristics of HAM/TSP is observed in a large proportion of HTLV-I asymptomatic carriers

BackgroundA small fraction of Human T cell Leukemia Virus type-1 (HTLV-I) infected subjects develop a severe form of myelopathy. It has been established that patients with HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP) show an exaggerated immune response when compared with the immunological response observed in HTLV-I asymptomatic carriers. In this study the immunological responses in HAM/TSP patients and in HTLV-I asymptomatic carriers were compared using several immunological assays to identify immunological markers associated with progression from infection to disease.MethodsImmunoproliferation assays, cytokine levels of unstimulated cultures, and flow cytometry analysis were used to evaluate the studied groups. Nonparametric tests (Mann-Whitney U test and Wilcoxon matched-pairs signed ranks) were used to compare the difference between the groups.ResultsAlthough both groups showed great variability, HAM/TSP patients had higher spontaneous lymphoproliferation as well as higher IFN-γ levels in unstimulated supernatants when compared with asymptomatic carriers. Flow cytometry studies demonstrated a high frequency of inflammatory cytokine (IFN-γ and TNF-α) producing lymphocytes in HAM/TSP as compared to the asymptomatic group. This difference was accounted for mainly by an increase in CD8 cell production of these cytokines. Moreover, the HAM/TSP patients also expressed an increased frequency of CD28-/CD8+ T cells. Since forty percent of the asymptomatic carriers had spontaneous lymphoproliferation and IFN-γ production similar to HAM/TSP patients, IFN-γ levels were measured eight months after the first evaluation in some of these patients to observe if this was a transient or a persistent situation. No significant difference was observed between the means of IFN-γ levels in the first and second evaluation.ConclusionsThe finding that a large proportion of HTLV-I carriers present similar immunological responses to those observed in HAM/TSP, strongly argues for further studies to evaluate these parameters as markers of HAM/TSP progression.

Recruitment of CD8+ T cells expressing granzyme A is associated with lesion progression in human cutaneous leishmaniasis

Human infection with Leishmania braziliensis leads to the establishment of cutaneous leishmaniasis (CL), characterized by the appearance of skin lesions that progress from nonulcerated to ulcerated forms. Our goal was to characterize the immunological kinetics associated with this progression, comparing the cellular composition, cytokines and granzyme expression between lesions of patients with early (E‐CL) and late stages (L‐CL) of CL. Histopathological analysis showed that lesions from L‐CL had more exuberant inflammatory infiltrate as compared to E‐CL. Although E‐CL and L‐CL lesions were predominantly mononuclear, lesions from E‐CL patients presented higher neutrophil and eosinophil counts than L‐CL. While percentages of CD4+ and of CD68+ cells were slightly higher in L‐CL, a fivefold increase of CD8+ cells was observed in L‐CL, as compared to E‐CL. Moreover, CD8+ T‐cells from L‐CL expressed significantly higher levels of granzyme A than E‐CL. Interestingly, granzyme A expression was positively correlated with intensity of the inflammatory infiltrate in L‐CL but not E‐CL. Lastly, percentages of IFN‐γ+ and IL‐10+ cells were higher in L‐CL as compared to E‐CL, with CD4+ T‐cells and CD68+ monocytes as the main sources of these cytokines, respectively. These results suggest that recruitment of CD8+ granzyme A+ T cells is involved in lesion progression in human CL.

Interleukin 17 Production among Patients with American Cutaneous Leishmaniasis

Interleukin 17 (IL-17) plays a critical role in inflammation and autoimmunity. Very little is known about IL-17 in protozoa infection. Here, we show that lymphocytes obtained from patients with mucosal leishmaniasis and cutaneous leishmaniasis produce higher levels of IL-17 than do lymphocytes obtained from uninfected control subjects (P<.01). There was a tendency for tissue obtained from patients with mucosal leishmaniasis to contain a higher number of cells expressing IL-17, compared with tissue obtained from patients with cutaneous leishmaniasis, and there was a direct correlation between the number of cells expressing IL-17 and the presence of cellular inflammation at the lesion site (r2 = 0.86; P<.001) These data support the role of IL-17 in the pathogenesis of the inflammatory reaction in leishmaniasis.

Norepinephrine, dopamine and dexamethasone modulate discrete leukocyte subpopulations and cytokine profiles from human PBMC

The interplay between the immune and neuroendocrine systems is intense, with the cross-talk between these two systems increasing during stress circumstances. Stress events culminate with hormonal pathway activation elevating the plasma levels of glucocorticoids and catecholamines. The majority of the works evaluating the effects of stress hormones on immune cells have utilized in vivo animal models or clinical studies. This work evaluates the effects of norepinephrine, dopamine, dexamethasone, and the combination of norepinephrine and dexamethasone on cellular activation and expression of immunoregulatory cytokines and chemokines by human PBMC in vitro. Norepinephrine and dopamine increased lymphocyte activation accompanied by augmented Th1 and Th2 type cytokine production. Dexamethasone reduced cell activation and decreased frequencies of cytokine producing cells and chemokine production. The action of norepinephrine together with dexamethasone resulted in immunosupression. The observed effects of hormones and neurotransmitters on leukocyte subsets likely underlie their immunomodulatory action in vivo.

Differential immune regulation of activated T cells between cutaneous and mucosal leishmaniasis as a model for pathogenesis

Cutaneous (CL) and mucosal leishmaniasis (ML) are characterized by a predominant type 1 immune response (IFN‐γ and TNF‐α production) and strong inflammatory response in the lesions with few parasites. This exacerbated type 1 response is more evident in ML as compared to CL. Our main hypothesis is that a differential immune regulation of T cell activation leads to over reactive T cells in ML. In the present study, we investigated immunological factors that could explain the mechanisms behind it by comparing some immune regulatory mechanisms between ML and CL patients: frequency of cells expressing co‐stimulatory molecules, apoptotic markers, T cell activation markers; and ability of neutralizing antibodies to IL‐2, IL‐12 and IL‐15 do down‐regulate IFN‐γ production in leishmania antigen‐stimulated peripheral blood mononuclear cells (PBMC). Interestingly, in CL anti‐IL‐2 and anti‐IL‐15 significantly suppressed antigen‐specific IFN‐γ production, while in ML only anti‐IL‐2 suppressed IFN‐γ production. Finally, higher frequency of CD4+ T cells expressing CD28−, CD69+ and CD62L low were observed in ML as compared to CL. These data indicate that an exacerbated type 1 response in ML is differentially regulated and not appropriately down modulated, with increased frequencies of activated effectors T cells, maintaining the persistent inflammatory response and tissue damage observed in ML.