Effect of Dietary Supplementation with Omega-3 Fatty Acid on the Generation of Regulatory T Lymphocytes and on Antioxidant Parameters and Markers of Oxidative Stress in the Liver Tissue of IL-10 Knockout Mice.

INTRODUCTION: chronic low-grade inflammation, or inflammaging, emerges as a crucial element in the aging process and is associated with cardiovascular and neurological diseases, sarcopenia, and malnutrition. Evidence suggests that omega-3 fatty acids present a potential therapeutic agent in the prevention and treatment of inflammatory diseases, mitigating oxidative stress, and improving muscle mass, attributes that are particularly relevant in the context of aging. The objective of the present study was to evaluate the effectiveness of supplementation with omega-3 fish oil in improving the immune response and oxidative stress in knockout mice for interleukin IL-10 (IL-10-/-). MATERIAL AND METHODS: female C57BL/6 wild-type (WT) and interleukin IL-10 knockout (IL-10-/-) mice were fed during 90 days with a standard diet (control groups), or they were fed/supplemented with 10% of the omega-3 polyunsaturated fatty acid diet (omega-3 groups). Muscle, liver, intestinal, and mesenteric lymph node tissue were collected for analysis. RESULTS: the IL-10-/-+O3 group showed greater weight gain compared to the WT+O3 (p = 0.001) group. The IL-10-/-+O3 group exhibited a higher frequency of regulatory T cells than the IL-10-/- group (p = 0.001). It was found that animals in the IL-10-/-+O3 group had lower levels of steatosis when compared to the IL-10-/- group (p = 0.017). There was even greater vitamin E activity in the WT group compared to the IL-10-/-+O3 group (p = 0.001) and WT+O3 compared to IL-10-/-+O3 (p = 0.002), and when analyzing the marker of oxidative stress, MDA, an increase in lipid peroxidation was found in the IL-10-/-+O3 group when compared to the IL-10-/- group (p = 0.03). Muscle tissue histology showed decreased muscle fibers in the IL-10-/-+O3, IL-10-/-, and WT+O3 groups. CONCLUSION: the findings show a decrease in inflammation, an increase in oxidative stress markers, and a decrease in antioxidant markers in the IL-10-/-+O3 group, suggesting that supplementation with omega-3 fish oil might be a potential intervention for inflammaging that characterizes the aging process and age-related diseases.

Asthma-associated bacterial infections: Are they protective or deleterious?

Eosinophilic, noneosinophilic, or mixed granulocytic inflammations are the hallmarks of asthma heterogeneity. Depending on the priming of lung immune and structural cells, subjects with asthma might generate immune responses that are TH2-prone or TH17-prone immune response. Bacterial infections caused by Haemophilus, Moraxella, or Streptococcus spp. induce the secretion of IL-17, which in turn recruit neutrophils into the airways. Clinical studies and experimental models of asthma indicated that neutrophil infiltration induces a specific phenotype of asthma, characterized by an impaired response to corticosteroid treatment. The understanding of pathways that regulate the TH17-neutrophils axis is critical to delineate and develop host-directed therapies that might control asthma and its exacerbation episodes that course with infectious comorbidities. In this review, we outline clinical and experimental studies on the role of airway epithelial cells, S100A9, and high mobility group box 1, which act in concert with the IL-17-neutrophil axis activated by bacterial infections, and are related with asthma that is difficult to treat. Furthermore, we report critically our view in the light of these findings in an attempt to stimulate further investigations and development of immunotherapies for the control of severe asthma.

IL-22 Is Deleterious along with IL-17 in Allergic Asthma but Is Not Detrimental in the Comorbidity Asthma and Acute Pneumonia.

There is evidence that IL-22 and IL-17 participate in the pathogenesis of allergic asthma. To investigate the role of IL-22, we used IL-22 deficient mice (IL-22 KO) sensitized and challenged with ovalbumin (OVA) and compared with wild type (WT) animals exposed to OVA. IL-22 KO animals exposed to OVA showed a decreased number and frequency of eosinophils, IL-5 and IL-13 in the airways, reduced mucus production and pulmonary inflammation. In addition, IL-22 KO animals exhibited a decreased percentage and number of lung CD11c+CD11b+ cells and increased apoptosis of eosinophils. Th17 cell transfer generated from IL-22 KO to animals previously sensitized and challenged with OVA caused a reduction in eosinophil frequency and number in the airways compared to animals transferred with Th17 cells generated from WT mice. Therefore, IL-22 is deleterious with concomitant secretion of IL-17. Our findings show a pro-inflammatory role for IL-22, confirmed in a model of allergen-free and allergen-specific immunotherapy. Moreover, during the comorbidity asthma and pneumonia that induces neutrophil inflammation, IL-22 was not detrimental. Our results show that targeting IL-22 would negatively affect the survival of eosinophils, reduce the expansion or migration of CD11c+CD11b+ cells, and negatively regulate allergic asthma.

Pioneering in vitro characterization of macrophage response induced by scorpion venoms from the Brazilian Amazon.

There are several scorpion species of medical relevance around the world. Some of them are well characterized by their toxins and clinical outcomes. Brazilian Amazon has a great amount of these arthropods that have an impact in the scorpionism events specifically in this region of Brazil. Recently, several studies pointed out the immune system activation during scorpion envenouming as an important facet of scorpionism, inducing a sepsis-like state that culminates in clinical severity and death. In this work, we characterized the macrophage response of three species of clinical relevance in Brazilian Amazon: Tityus silvestris, T. metuendus and T. obscurus and one specie with no toxic effects to humans, Brotheas amazonicus. All the four species analyzed were able to induce pro- and anti-inflammatory cytokine production in a J774.1 murine macrophage model. This activation was dependent on TLR2/TLR4/MyD88 activation and abolished by TLRs antagonists. These results suggest that the venoms of the four species analyzed were able to induce macrophage response in agreement to the well-established immune activation by T. serrulatus venom. Our findings provide new insights into the clinical repercussions of scorpionism of uncharacterized species and point to new biotechnological applications of these venoms and possible supportive therapies in scorpionism.

Xenogeneic mesenchymal stem cell biocurative improves skin wounds healing in diabetic mice by increasing mast cells and the regenerative profile.

Introduction: Diabetes mellitus (DM) is a chronic disease and a major cause of mortality and morbidity worldwide. The hyperglycemia caused by DM induces micro and macrovascular complications that lead, among other consequences, to chronic wounds and amputations. Cell therapy and tissue engineering constitute recent therapeutic alternatives to improve wound healing in diabetic patients. The current study aimed to analyze the effectiveness of biocuratives containing human mesenchymal stem cells (MSCs) associated with a hydrogel matrix in the wound healing process and related inflammatory cell profile in diabetic mice. Methods: Biocuratives containing MSCs were constructed by 3D bioprinting, and applied to skin wounds on the back of streptozotocin (STZ)-induced type 1 diabetic (T1D) mice. The healing process, after the application of biocuratives with or without MSCs was histologically analyzed. In parallel, genes related to growth factors, mast cells (MC), M1 and M2 macrophage profiles were evaluated by RT-PCR. Macrophages were characterized by flow cytometry, and MC by toluidine blue staining and flow cytometry. Results: Mice with T1D exhibited fewer skin MC and delayed wound healing when compared to the non-diabetic group. Treatment with the biocuratives containing MSCs accelerated wound healing and improved skin collagen deposition in diabetic mice. Increased TGF-ß gene expression and M2 macrophage-related markers were also detected in skin of diabetic mice that received MSCs-containing biocuratives. Finally, MSCs upregulated IL-33 gene expression and augmented the number of MC in the skin of diabetic mice. Conclusion: These results reveal the therapeutic potential of biocuratives containing MSCs in the healing of skin wounds in diabetic mice, providing a scientific base for future treatments in diabetic patients.

Timeline Kinetics of Systemic and Airway Immune Mediator Storm for Comprehensive Analysis of Disease Outcome in Critically Ill COVID-19 Patients.

In the present study, the levels of serum and airway soluble chemokines, pro-inflammatory/regulatory cytokines, and growth factors were quantified in critically ill COVID-19 patients (total n=286) at distinct time points (D0, D2-6, D7, D8-13 and D>14-36) upon Intensive Care Unit (ICU) admission. Augmented levels of soluble mediators were observed in serum from COVID-19 patients who progress to death. An opposite profile was observed in tracheal aspirate samples, indicating that systemic and airway microenvironment diverge in their inflammatory milieu. While a bimodal distribution was observed in the serum samples, a unimodal peak around D7 was found for most soluble mediators in tracheal aspirate samples. Systems biology tools further demonstrated that COVID-19 display distinct eccentric soluble mediator networks as compared to controls, with opposite profiles in serum and tracheal aspirates. Regardless the systemic-compartmentalized microenvironment, networks from patients progressing to death were linked to a pro-inflammatory/growth factor-rich, highly integrated center. Conversely, patients evolving to discharge exhibited networks of weak central architecture, with lower number of neighborhood connections and clusters of pro-inflammatory and regulatory cytokines. All in all, this investigation with robust sample size landed a comprehensive snapshot of the systemic and local divergencies composed of distinct immune responses driven by SARS-CoV-2 early on severe COVID-19.

Systemic Infection by Non-albicans Candida Species Affects the Development of a Murine Model of Multiple Sclerosis.

Candidiasis may affect the central nervous system (CNS), and although Candida albicans is predominant, non-albicans Candida species can also be associated with CNS infections. Some studies have suggested that Candida infections could increase the odds of multiple sclerosis (MS) development. In this context, we investigated whether systemic infection by non-albicans Candida species would affect, clinically or immunologically, the severity of experimental autoimmune encephalomyelitis (EAE), which is an animal model used to study MS. For this, a strain of C. glabrata, C. krusei, and C. parapsilosis was selected and characterized using different in vitro and in vivo models. In these analysis, all the strains exhibited the ability to form biofilms, produce proteolytic enzymes, and cause systemic infections in Galleria mellonella, with C. glabrata being the most virulent species. Next, C57BL/6 mice were infected with strains of C. glabrata, C. krusei, or C. parapsilosis, and 3 days later were immunized with myelin oligodendrocyte glycoprotein to develop EAE. Mice from EAE groups previously infected with C. glabrata and C. krusei developed more severe and more prevalent paralysis, while mice from the EAE group infected with C. parapsilosis developed a disease comparable to non-infected EAE mice. Disease aggravation by C. glabrata and C. krusei strains was concomitant to increased IL-17 and IFN-γ production by splenic cells stimulated with fungi-derived antigens and with increased percentage of T lymphocytes and myeloid cells in the CNS. Analysis of interaction with BV-2 microglial cell line also revealed differences among these strains, in which C. krusei was the strongest activator of microglia concerning the expression of MHC II and CD40 and pro-inflammatory cytokine production. Altogether, these results indicated that the three non-albicans Candida strains were similarly able to reach the CNS but distinct in terms of their effect over EAE development. Whereas C. glabrata and C. Krusei aggravated the development of EAE, C. parapsilosis did not affect its severity. Disease worsening was partially associated to virulence factors in C. glabrata and to a strong activation of microglia in C. krusei infection. In conclusion, systemic infections by non-albicans Candida strains exerted influence on the experimental autoimmune encephalomyelitis in both immunological and clinical aspects, emphasizing their possible relevance in MS development.

Protective Immunity against Gamma and Zeta Variants after Inactivated SARS-CoV-2 Virus Immunization.

The persistent circulation of SARS-CoV-2 represents an ongoing global threat due to the emergence of new viral variants that can sometimes evade the immune system of previously exposed or vaccinated individuals. We conducted a follow-up study of adult individuals that had received an inactivated SARS-CoV-2 vaccine, evaluating antibody production and neutralizing activity over a period of 6 months. In addition, we performed mice immunization with inactivated SARS-CoV-2, and evaluated the immune response and pathological outcomes against Gamma and Zeta variant infection. Vaccinated individuals produced high levels of antibodies with robust neutralizing activity, which was significantly reduced against Gamma and Zeta variants. Production of IgG anti-S antibodies and neutralizing activity robustly reduced after 6 months of vaccination. Immunized mice demonstrated cellular response against Gamma and Zeta variants, and after viral infection, reduced viral loads, IL-6 expression, and histopathological outcome in the lungs. TNF levels were unchanged in immunized or not immunized mice after infection with the Gamma variant. Furthermore, serum neutralization activity rapidly increases after infection with the Gamma and Zeta variants. Our data suggest that immunization with inactivated WT SARS-CoV-2 induces a promptly responsive cross-reactive immunity response against the Gamma and Zeta variants, reducing COVID-19 pathological outcomes.

Artepillin C Reduces Allergic Airway Inflammation by Induction of Monocytic Myeloid-Derived Suppressor Cells.

Propolis is a natural product produced by bees that is primarily used in complementary and alternative medicine and has anti-inflammatory, antibacterial, antiviral, and antitumoral biological properties. Some studies have reported the beneficial effects of propolis in models of allergic asthma. In a previous study, our group showed that green propolis treatment reduced airway inflammation and mucus secretion in an ovalbumin (OVA)-induced asthma model and resulted in increased regulatory T cells (Treg) and polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC) frequencies in the lungs, two leukocyte populations that have immunosuppressive functions. In this study, we evaluated the anti-inflammatory effects of artepillin C (ArtC), the major compound of green propolis, in the context of allergic airway inflammation. Our results show that ArtC induces in vitro differentiation of Treg cells and monocytic MDSC (M-MDSC). Furthermore, in an OVA-induced asthma model, ArtC treatment reduced pulmonary inflammation, eosinophil influx to the airways, mucus and IL-5 secretion along with increased frequency of M-MDSC, but not Treg cells, in the lungs. Using an adoptive transfer model, we confirmed that the effect of ArtC in the reduction in airway inflammation was dependent on M-MDSC. Altogether, our data show that ArtC exhibits an anti-inflammatory effect and might be an adjuvant therapy for allergic asthma.

Kallikrein 5 Inhibition by the Lympho-Epithelial Kazal-Type Related Inhibitor Hinders Matriptase-Dependent Carcinogenesis.

Head and neck squamous cell carcinoma remains challenging to treat with no improvement in survival rates over the past 50 years. Thus, there is an urgent need to discover more reliable therapeutic targets and biomarkers for HNSCC. Matriptase, a type-II transmembrane serine protease, induces malignant transformation in epithelial stem cells through proteolytic activation of pro-HGF and PAR-2, triggering PI3K-AKT-mTOR and NFKB signaling. The serine protease inhibitor lympho-epithelial Kazal-type-related inhibitor (LEKTI) inhibits the matriptase-driven proteolytic pathway, directly blocking kallikreins in epithelial differentiation. Hence, we hypothesized LEKTI could inhibit matriptase-dependent squamous cell carcinogenesis, thus implicating kallikreins in this process. Double-transgenic mice with simultaneous expression of matriptase and LEKTI under the keratin-5 promoter showed a prominent rescue of K5-Matriptase+/0 premalignant phenotype. Notably, in DMBA-induced SCC, heterotopic co-expression of LEKTI and matriptase delayed matriptase-driven tumor incidence and progression. Co-expression of LEKTI reverted altered Kallikrein-5 expression observed in the skin of K5-Matriptase+/0 mice, indicating that matriptase-dependent proteolytic pathway inhibition by LEKTI occurs through kallikreins. Moreover, we showed that Kallikrein-5 is necessary for PAR-2-mediated IL-8 release, YAP1-TAZ/TEAD activation, and matriptase-mediated oral squamous cell carcinoma migration. Collectively, our data identify a third signaling pathway for matriptase-dependent carcinogenesis in vivo. These findings are critical for the identification of more reliable biomarkers and effective therapeutic targets in Head and Neck cancer.

Obesity-Induced Dysbiosis Exacerbates IFN-γ Production and Pulmonary Inflammation in the Mycobacterium tuberculosis Infection.

The microbiota of the gut-lung axis affects local and far-reaching immune responses and might also trigger chronic and inflammatory diseases. We hypothesized that gut dysbiosis induced by obesity, which coexists in countries with a high tuberculosis burden, aggravates the host susceptibility and the pulmonary damage tolerance. To assess our hypothesis, we used a model of high-fat diet (HFD)-induced obesity, followed by infection of C57BL/6 mice with Mycobacterium tuberculosis. We showed that obesity increased the susceptibility, the pulmonary inflammation and IFN-γ levels in M. tuberculosis-infected mice. During the comorbidity obesity and tuberculosis, there is an increase of Bacteroidetes and Firmicutes in the lungs, and an increase of Firmicutes and butyrate in the feces. Depletion of gut microbiota by antibiotic treatment in the obese infected mice reduced the frequencies of CD4+IFN-γ+IL-17- cells and IFN-γ levels in the lungs, associated with an increase of Lactobacillus. Our findings reinforce the role of the gut-lung axis in chronic infections and suggest that the gut microbiota modulation may be a potential host-directed therapy as an adjuvant to treat TB in the context of IFN-γ-mediated immunopathology.

Interactions of Extracellular Vesicles from Pathogenic Fungi with Innate Leukocytes.

Several studies have shown the immunomodulatory effects of extracellular vesicles (EVs) released by pathogenic fungi. Herein, we discuss the data regarding the immunomodulatory properties of fungal EVs, but also of EVs produced by infected leukocytes. This characterizes a two-way path, in which both host and fungal EVs could coexist and play crucial roles in disease progression or protection in fungal infections. We suggest that EVs can dictate the progress of fungal diseases, and their potential as therapeutic targets.

Interplay between alveolar epithelial and dendritic cells and Mycobacterium tuberculosis.

The innate response plays a crucial role in the protection against tuberculosis development. Moreover, the initial steps that drive the host-pathogen interaction following Mycobacterium tuberculosis infection are critical for the development of adaptive immune response. As alveolar Mϕs, airway epithelial cells, and dendritic cells can sense the presence of M. tuberculosis and are the first infected cells. These cells secrete mediators, which generate inflammatory signals that drive the differentiation and activation of the T lymphocytes necessary to clear the infection. Throughout this review article, we addressed the interaction between epithelial cells and M. tuberculosis, as well as the interaction between dendritic cells and M. tuberculosis. The understanding of the mechanisms that modulate those interactions is critical to have a complete view of the onset of an infection and may be useful for the development of dendritic cell-based vaccine or immunotherapies.

Mycobacterium tuberculosis-infected alveolar epithelial cells modulate dendritic cell function through the HIF-1α-NOS2 axis.

Tuberculosis kills more than 1 million people every year, and its control depends on the effective mechanisms of innate immunity, with or without induction of adaptive immune response. We investigated the interaction of type II alveolar epithelial cells (AEC-II) infected by Mycobacterium tuberculosis with dendritic cells (DCs). We hypothesized that the microenvironment generated by this interaction is critical for the early innate response against mycobacteria. We found that AEC-II infected by M. tuberculosis induced DC maturation, which was negatively regulated by HIF-1α-inducible NOS2 axis, and switched DC metabolism from an early and short peak of glycolysis to a low energetic status. However, the infection of DCs by M. tuberculosis up-regulated NOS2 expression and inhibited AEC-II-induced DC maturation. Our study demonstrated, for the first time, that HIF-1α-NOS2 axis plays a negative role in the maturation of DCs during M. tuberculosis infection. Such modulation might be useful for the exploitation of molecular targets to develop new therapeutic strategies against tuberculosis.

COVID-19: Integrating the Complexity of Systemic and Pulmonary Immunopathology to Identify Biomarkers for Different Outcomes.

In the last few months, the coronavirus disease 2019 (COVID-19) pandemic has affected millions of people worldwide and has provoked an exceptional effort from the scientific community to understand the disease. Clinical evidence suggests that severe COVID-19 is associated with both dysregulation of damage tolerance caused by pulmonary immunopathology and high viral load. In this review article, we describe and discuss clinical studies that show advances in the understanding of mild and severe illness and we highlight major points that are critical for improving the comprehension of different clinical outcomes. The understanding of pulmonary immunopathology will contribute to the identification of biomarkers in an attempt to classify mild, moderate, severe and critical COVID-19 illness. The interface of pulmonary immunopathology and the identification of biomarkers are critical for the development of new therapeutic strategies aimed to reduce the systemic and pulmonary hyperinflammation in severe COVID-19.

Fungal Extracellular Vesicles as Potential Targets for Immune Interventions.

The release of extracellular vesicles (EVs) by fungi is a fundamental cellular process. EVs carry several biomolecules, including pigments, proteins, enzymes, lipids, nucleic acids, and carbohydrates, and are involved in physiological and pathological processes. EVs may play a pivotal role in the establishment of fungal infections, as they can interact with the host immune system to elicit multiple outcomes. It has been observed that, depending on the fungal pathogen, EVs can exacerbate or attenuate fungal infections. The study of the interaction between fungal EVs and the host immune system and understanding of the mechanisms that regulate those interactions might be useful for the development of new adjuvants as well as the improvement of protective immune responses against infectious or noninfectious diseases. In this review, we describe the immunomodulatory properties of EVs produced by pathogenic fungi and discuss their potential as adjuvants for prophylactic or therapeutic strategies.

Systemic Immunological changes in patients with distinct clinical outcomes during Mycobacterium tuberculosis infection.

BACKGROUND: The lung lesions in an individual infected with tuberculosis (TB) are surprisingly variable and independent of each other. However, there is no circulating biomarker yet able to segregate patients according to the extent of lung lesions. MATERIALS AND METHODS: In this study, the phenotypic and functional profile of leukocytes of patients with active pulmonary tuberculosis (TB) and controls (CO) were fully scrutinized by immunophenotyping assays and in vitro short-term whole blood culture. The TB group was subdivided according to the extent of lung lesions as unilateral (UNI) and bilateral (BI). RESULTS: The results show that TB group display an altered leukocyte profile in the peripheral blood with significant lower counts of NK-cells, CD3+CD56+CD16+/- NKT-cells, CD4+T-cells, CD19+B-cells when compared to CO. Increased CD4+T-cells and CD8+T-cell activation was observed by the upregulation of activation markers (HLA-DR) as well as of chemokine receptors (CCR2, CCR3, and CXCR4). In addition, TB group presented a significant decrease proportion of CD14LowCD16+ monocytes despite the increase in HLA-DR expression. Regarding the severity of the disease, in the BI group a reduction in frequency of CD19+CD5+ B-cells and expression of HLA-DR in CD14LowCD16+ monocytes was observed. Furthermore, the extent of lung lesions influences the production of molecules as observed by significantly larger production of IL-4 by neutrophils, total T-cells, CD4+T-cells, CD8+T-cells and CD19+B-cells in UNI as compared to BI. By contrast, in BI group the frequency of high producers of both IL-17+CD4+T-cells and IL-17+CD8+T-cells were significantly increased than UNI, suggesting the deleterious role of these subsets during active pulmonary Mtb infection. CONCLUSION: The immunophenotypic characterization of unilateral and bilateral active TB performed in the present study indicates that the extent of lung lesion could be associated with a fine-tuning between immunological responses during untreated Mtb infection.

Mycobacterial Hsp65 antigen upregulates the cellular immune response of healthy individuals compared with tuberculosis patients.

Previously we showed that 65-kDa Mycobacterium leprae heat shock protein (Hsp65) is a target for the development of a tuberculosis vaccine. Here we evaluated peripheral blood mononuclear cells (PBMC) from healthy individuals or tuberculosis patients stimulated with two forms of Hsp65 antigen, recombinant DNA that encodes Hsp65 (DNA-HSP65) or recombinant Hsp65 protein (rHsp65) in attempting to mimic a prophylactic or therapeutic study in vitro, respectively. Proliferation and cytokine-producing CD4+ or CD8+ cell were assessed by flow cytometry. The CD4+ cell proliferation from healthy individuals was stimulated by DNA-HSP65 and rHsp65, while CD8+ cell proliferation from healthy individuals or tuberculosis patients was stimulated by rHSP65. DNA-HSP65 did not improve the frequency of IFN-gamma+ cells from healthy individuals or tuberculosis patients. Furthermore, we found an increase in the frequency of IL-10-producing cells in both groups. These findings show that Hsp65 antigen activates human lymphocytes and plays an immune regulatory role that should be addressed as an additional antigen for the development of antigen-combined therapies.

New strategy for testing efficacy of immunotherapeutic compounds for diabetes in vitro.

BACKGROUND: The valuable role of immunotherapy in treating autoimmune diseases is increasingly recognized by those involved in the research and clinical application of new biopharmaceuticals products. However, many aspects related to the mechanisms of immune-modulated therapies remain to be elucidated in order to explore fully the emerging opportunities. The non-obese diabetic NOD mouse develops insulin-dependent diabetes mellitus spontaneously as a consequence of an autoimmune process in the presence of pathogenic CD4(+) T cells that typically exhibit Th17 cell phenotypes. The change of a Th17 phenotype into a pattern of regulatory T cells (Treg) is extremely important in controlling autoimmune diseases. Heat shock proteins (HSPs) are stress-induced proteins with immunoregulatory properties. In the current study, the capacity of Hsp65 and Hsp70 mycobacterial HSPs and a constructed DNA encoded Hsp65 (DNAhsp65) to transform the pattern of the immune response from Th17 into Treg cells has been studied in vitro using co-cultures of antigen presenting cells (APCs) and T cells in NOD mice. RESULTS: Cells harvested from NOD mice and cultured for 48 h (without immunoregulatory compounds) presented with Th1/Th17 patterns and secretions of IL-6, IFN-γ, IL-10 and IL-17 cytokines. The cultured cells from the non-diabetic BALB/C mice exhibited a Th1 pattern and the production of IL 6 and IFN-γ secretions. An up-regulation was observed in the supernatants from the co-cultures of NOD cells that were stimulated with DNAhsp65, Hsp65 or Hsp70 through increased levels of IL-10 secretion and the suppression of IL-6, IFN-γ and IL-17 production. In addition, immunoregulation was demonstrated through IL-17 suppression in the co-culture stimulated by the specific insulin antigen. Moreover, an increase of immunoregulatory compounds were observed in the co-culture through the expression of CD11b(+)CD86(+) activation markers on APCs, as well as the frequency of Treg cells expressing CD4(+)CD3(+) and CD4(+)CD25(hi). CONCLUSIONS: The in vitro observation of Th17 cells differentiating into Tregs in NOD mice could raise the hypothesis that the immune regulatory activity of HSPs could be an efficient strategy for diabetes prevention and treatment.

Genetic background affects the expansion of macrophage subsets in the lungs of Mycobacterium tuberculosis-infected hosts.

M1 macrophages are more effective in the induction of the inflammatory response and clearance of Mycobacterium tuberculosis than M2 macrophages. Infected C57BL/6 mice generate a stronger cellular immune response compared with BALB/c mice. We hypothesized that infected C57BL/6 mice would exhibit a higher frequency and function of M1 macrophages than infected BALB/c mice. Our findings show a higher ratio of macrophages to M2 macrophages in the lungs of chronically infected C57BL/6 mice compared with BALB/c mice. However, there was no difference in the functional ability of M1 and M2 macrophages for the two strains in vitro. In vivo, a deleterious role for M2 macrophages was confirmed by M2 cell transfer, which rendered the infected C57BL/6, but not the BALB/c mice, more susceptible and resulted in mild lung inflammation compared with C57BL/6 mice that did not undergo cell transfer. M1 cell transfer induced a higher inflammatory response, although not protective, in infected BALB/c mice compared with their counterparts that did not undergo cell transfer. These findings demonstrate that an inflammation mediated by M1 macrophages may not induce bacterial tolerance because protection depends on the host genetic background, which drives the magnitude of the inflammatory response against M. tuberculosis in the pulmonary microenvironment. The contribution of our findings is that although M1 macrophage is an effector leucocyte with microbicidal machinery, its dominant role depends on the balance of M1 and M2 subsets, which is driven by the host genetic background.