Differential expression of genes regulated by the glucocorticoid receptor pathway in patients with pulmonary tuberculosis.

AIMS: Previous studies in TB patients showed an immuno-endocrine imbalance characterized by a disease-severity associated increase in plasma levels of proinflammatory cytokines and glucocorticoids (GCs). To analyze the potential immunomodulatory effect of circulating GCs over peripheral blood mononuclear cells (PBMC) from TB patients, we investigated the expression of positively (anti-inflammatory-related genes ANXA1; FKBP51; GILZ, NFKBIA, and NFKBIB) and negatively (inflammatory genes: IL-6, IL-1ß, and IFN-γ) Glucocorticoids Receptors (GR)-regulated genes. Plasma concentrations of cytokines and hormones, together with specific lymphoproliferation were also assessed. MATERIALS AND METHODS: Gene expression was quantified by RT-qPCR, specific lymphoproliferation by 3H-thymidine incorporation, whereas plasma cytokines and hormones levels by ELISA. KEY FINDINGS: Transcripts of ANXA1, GILZ, NFKBIB, and NFKBIA appeared significantly increased in patients, whereas FKBP51, IL-6, IL-1ß, and NF-κB remained unchanged. Upon analyzing according to disease severity, mRNA levels for ANXA1 and NFKBIB were even higher in moderate and severe patients. GILZ was increased in moderate cases, with NFKBIA and IL-1 ß being higher in severe ones, who also displayed increased GRß transcripts. TB patients had reduced plasma DHEA concentrations together with increased pro and anti-inflammatory cytokines (IFN-γ, IL-6, and IL-10) cortisol and cortisol/DHEA ratio, more evident in progressive cases, in whom their PBMC also showed a decreased mycobacterial-driven proliferation. The cortisol/DHEA ratio and GRα expression were positively correlated with GR-regulated genes mainly in moderate patients. SIGNIFICANCE: The increased expression of cortisol-regulated anti-inflammatory genes in TB patients-PBMC, predominantly in progressive disease, seems compatible with a relatively insufficient attempt to downregulate the accompanying inflammation.

Enhanced Migratory Capacity of T Lymphocytes in Severe Chagasic Patients Is Correlated With VLA-4 and TNF-α Expression.

Trypanosoma cruzi infection in humans leads to progression to chronic chagasic myocarditis (CCM) in 30% of infected individuals, paralleling T cell inflammatory infiltrates in the heart tissue. T-cell trafficking into the hearts of CCM patients may be modulated by in situ expression of chemotactic or haptotactic molecules, as the chemokine CXCL12, the cytokine tumor necrosis factor-alpha (TNF-α), and extracellular matrix proteins (ECM), such as fibronectin. Herein we evaluated the expression of fibronectin, CXCL12, and TNF-α in the myocardial tissue of T. cruzi seropositive (asymptomatic or with CCM), as well as seronegative individuals as healthy controls. Hearts from CCM patients exhibited enhanced expression of these three molecules. CXCL12 and TNF-α serum levels were also increased in CCM individuals. We then evaluated T lymphocytes from chronic chagasic patients by cytofluorometry, in terms of membrane expression levels of molecules involved in cell activation and cell migration, respectively, HLA-DR and the VLA-4 (very late antigen-4, being one integrin-type fibronectin receptor). Indeed, the expression of HLA-DR and VLA-4 was enhanced on T lymphocytes from chagasic patients, especially in the CCM group. To further approach the dynamics of T cell migratory events, we performed fibronectin-, TNF-α-, and CXCL12-driven migration. Peripheral blood mononuclear cells (PBMCs) and T cells from CCM patients presented an ex vivo enhanced migratory capacity driven by fibronectin alone when this ECM protein was placed in the membrane of transwell migration chambers. When TNF-α was previously placed upon fibronectin, we observed a further and significant increase in the migratory response of both PBMCs and T lymphocytes. Overall, these data suggest the existence in patients with chronic Chagas disease of a cardiac inflammatory infiltrate vector that promotes the recruitment and accumulation of activated T cells, driven in part by enhanced tissue expression of fibronectin and TNF-α, as well as the respective corresponding VLA-4 and TNF receptors.

Evidence for a More Disrupted Immune-Endocrine Relation and Cortisol Immunologic Influences in the Context of Tuberculosis and Type 2 Diabetes Comorbidity.

Pulmonary tuberculosis (PTB), caused by Mycobacterium tuberculosis (Mtb), is a major health problem worldwide, further aggravated by the convergence of type 2 diabetes mellitus (DM) which constitutes an important risk factor for TB development. The worse scenario of patients with PTB and DM may be partly related to a more unbalanced defensive response. As such, newly diagnosed PTB patients with DM (TB+DM, n = 11) or not (TB, n = 21), as well as DM (n = 18) patients and pair matched controls (Co, n = 22), were investigated for the circulating immuno-endocrine-metabolic profile (ELISA), along with studies in peripheral blood mononuclear cells (PBMC) analyzing transcript expression (RT-qPCR) of mediators involved in glucocorticoid functionality. Given the hyperglycemic/hypercortisolemic scenario of TB+DM patients, PBMC were also exposed to stress-related cortisol concentrations (0.1 and 1 µM) and supraphysiologic glucose doses (10, 20, and 40 mM) and assessed for the specific response against Mtb stimulation (lymphoproliferation, -thymidine incorporation-, and cytokine production -bead-cytometry). All TB patients displayed increased plasma amounts of cortisol, growth hormone -hGH-, and proinflammatory mediators. In turn, TB+DM showed even higher levels of interferon gamma -IFN-γ- and hGH (vs. TB), or IL-6, C reactive protein, cortisol and hGH (vs. DM). Both DM groups had equally augmented values of IL-10. All TB patients showed decreased dehydroepiandrosterone- sulfate concentrations, even more in TB+DM cases. Leptin was also decreased in both TB cases, particularly in the TB group, revealing a lower body mass index, as well. Unlike PBMC from TB cases showing a decreased relationship between the glucocorticoids receptor (GR) isoforms (GRα/GRß; functional isoform/negative isoform), cells from TB+DM patients had no changes in this regard, along with an increased expression of 11-beta hydroxysteroid dehydrogenase type-1, the enzyme facilitating intracellular cortisone to cortisol conversion. TB+DM patients also showed an increased Mtb antigen-driven lymphoproliferation. Compared to TB, DM and HCo counterparts, PBMC from TB+DM patients had a biased Th1 response to Mtb stimulation (increased IL-2 and IFN-γ production), even when exposed to inhibitory cortisol doses. TB+DM patients show a more unbalanced immuno-endocrine relationship, respect the non-diabetic counterparts, with a relative deficiency of cortisol immunomodulatory influences, despite their more favorable microenvironment for cortisol-mediated immune effects.

Dysregulated Network of Immune, Endocrine and Metabolic Markers is Associated to More Severe Human Chronic Chagas Cardiomyopathy.

Individuals who are infected with Trypanosoma cruzi develop chronic Chagas cardiomyopathy (CCC), which is a complication involving a series of immune pathogenetic mechanisms, although an association between immune and metabolic alterations was more recently proposed. Accordingly, we investigated the immuno-metabolic response in chagasic patients and their possible influence on CCC pathogenesis. To this end, T. cruzi-seropositive (asymptomatic or with CCC) and sero-negative individuals were studied. Serum tumour necrosis factor (TNF)-α, interleukin (IL)-6, adipocytokines and the expression of their receptors in peripheral blood mononuclear cell (PBMC) were evaluated, together with other factors influencing the immune response. CCC patients showed major metabolic and hormonal abnormalities, in parallel with increased IL-6 and leptin serum levels. TNF-α receptor s, leptin and adiponectin receptors (ObR and Adipo-Rs respectively), as well as PPAR-γ expression in PBMCs from CCC patients were compatible with a counteracting response leading to an unfavourable immune-metabolic profile. These results suggest that persistently increased levels of immune-metabolic pro-inflammatory mediators along with the adverse endocrine anti-inflammatory response of CCC individuals, may contribute to the underlying mechanisms dealing with myocardial tissue damage.

The clinical recovery of tuberculosis patients undergoing specific treatment is associated with changes in the immune and neuroendocrine responses.

Tuberculosis (TB) caused by Mycobacterium tuberculosis is a health problem worldwide. Patients with pulmonary TB show a neuro-immune-endocrine imbalance characterized by an impaired cellular immunity together with increased plasma levels of cortisol, pro- and anti-inflammatory cytokines and markedly decreased dehydroepiandrosterone (DHEA) levels. Extending these findings, we now investigated the immune-endocrine profile of TB patients undergoing specific treatment. Patients (n = 24) were bled at diagnosis (T0), 2, 4, 6 months after treatment initiation and 3 months following its completion. At T0, TB patients showed increased plasma levels of interleukin-6 (IL-6), C reactive protein, interferon-gamma (IFN-γ) and transforming growth factor beta (TGF-ß). These mediators decreased during treatment, reaching levels similar to those from healthy controls (n = 26). Specific treatment led to an increased lymphoproliferative response along with clinical improvement. Newly diagnosed patients had low levels of DHEA, with increased cortisol amounts and cortisol/DHEA ratio, which normalized upon specific treatment. As regards glucocorticoid receptors (GR), TB patients at diagnosis presented a reduced mRNA GRα/GRß ratio in their peripheral blood mononuclear cells. Furthermore, multivariate analysis showed that cortisol/DHEA ratio was positively associated with inflammatory mediators for which this ratio may constitute a disease biomarker. Anti-mycobacterial treatment results in a better immune-endocrine scenario for the control of physiopathological processes accompanying disease development and hence implied in clinical recovery.

Increased frequency of CD4+ CD25+ FoxP3+ T regulatory cells in pulmonary tuberculosis patients undergoing specific treatment and its relationship with their immune-endocrine profile.

Tuberculosis (TB) is a major health problem requiring an appropriate cell immune response (IR) to be controlled. Since regulatory T cells (Tregs) are relevant in IR regulation, we analyzed Tregs variations throughout the course of TB treatment and its relationship with changes in immune-endocrine mediators dealing with disease immunopathology. The cohort was composed of 41 adult patients, 20 of them completing treatment and follow-up. Patients were bled at diagnosis (T0) and at 2 (T2), 4 (T4), 6 (T6), and 9 months following treatment initiation. Twenty-four age- and sex-matched healthy controls (HCo) were also included. Tregs (flow cytometry) from TB patients were increased at T0 (versus HCo P < 0.05), showing even higher values at T2 (versus T0 P < 0.01) and T4 (versus T0 P < 0.001). While IL-6, IFN-γ, TGF-ß (ELISA), and Cortisol (electrochemiluminescence, EQ) were augmented, DHEA-S (EQ) levels were diminished at T0 with respect to HCo, with cytokines and Cortisol returning to normal values at T9. Tregs correlated positively with IFN-γ (R = 0.868, P < 0.05) at T2 and negatively at T4 (R = -0.795, P < 0.05). Lowered levels of proinflammatory cytokines together with an increased frequency of Tregs of patients undergoing specific treatment might reflect a downmodulatory effect of these cells on the accompanying inflammation.

Vitamin D levels and bone mass in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic systemic inflammatory autoimmune disease with high prevalence of osteoporosis. Previous evidence indicates an association between vitamin D deficiency and autoimmune diseases. The aim of this study was to evaluate serum 25 hydroxyvitamin D [25(OH)D] levels, bone mineral density (BMD) and disease activity in RA patients living in Argentina. We studied 34 RA women and 41 healthy women as a control group. RA patients had lower 25(OH)D levels (20.4 ± 0.9 ng/ml) than controls (26.3 ± 1.9 ng/ml; p < 0.05). No significant differences were found in lumbar spine BMD between premenopausal (preM) or postmenopausal (postM) patients, but femoral neck BMD was significantly lower in postM RA patients (T score -2.5 ± 0.4) than in postM control subjects (T score -0.9 ± 0.3, p = 0.014). Although no linear correlation between 25(OH)D levels and disease activity (DAS-28) was found, patients with moderate-high disease activity had lower 25(OH)D levels than those with low disease activity: DAS-28 >3.2: 19.5 ± 0.88 ng/ml; DAS-28 ≤3.2: 23.7 ± 2.8 ng/ml (p = 0.047). After 1 year of vitamin D treatment 25(OH)D levels were increased while DAS-28 were decreased (n = 25; p < 0.05). We conclude that patients with RA had lower 25(OH)D levels than the control group. Low levels of 25(OH)D were associated with moderate-high disease activity suggesting the importance of optimal 25(OH)D levels in RA patients. Femoral neck BMD was lower in postM RA patients. No differences in lumbar BMD were found between preM and postM RA patients, suggesting that bone mass evaluation in RA patients should include femoral neck BMD regardless of age.

Assessment of cross-reactive host-pathogen antibodies in patients with different stages of chronic Chagas disease.

INTRODUCTION AND OBJECTIVES: Trypanosoma cruzi infection has been shown to induce humoral autoimmune responses against host antigens tissues. Particularly, antibodies cross-reacting with myocardial antigens may play a role in the development of the severe forms of chronic Chagas disease. The aim of this study was to determine the association between clinical stage of the disease and the presence of autoantibodies in patients with chronic Chagasic disease. METHODS: We performed a cross-sectional study in T. cruzi-seropositive patients divided into 3 groups according to the classic classification of chronic Chagas heart of Storino et al. All participants underwent complete clinical examination and their sera were used to measure autoantibody levels. RESULTS: All patients had detectable levels of anti-p2ß and anti-B13 autoantibodies but none had anti-Na-K-ATPase antibodies. No association was observed between electrocardiographic conduction disturbances and autoantibody levels. Patients with chronic Chagas disease stage III had the highest levels of anti-B13 antibodies and a high risk of mortality score, showing a clear association between disease stage and this score. CONCLUSIONS: Anti-B13 antibodies were significantly higher in chronic Chagas disease stage III patients, suggesting that these antibodies may be involved in disease progression and that they might be a useful marker of poor prognosis in terms of heart compromise. Our results also reveal an important correlation between the level of anti-B13 autoantibodies and symptomatic heart failure and/or dilated cardiomyopathy.

Changes in the immune and endocrine responses of patients with pulmonary tuberculosis undergoing specific treatment.

We evaluated immune and endocrine status following antituberculosis treatment in HIV-negative patients with newly diagnosed tuberculosis (TB). Treatment led to a decrease in IL-6, IL-1ß, and C-reactive protein levels. Cortisol levels decreased throughout the anti-TB treatment, particularly after 4 months, but changes were less pronounced than those seen in proinflammatory mediators. Specific therapy resulted in increased dehydroepiandrosterone (DHEA) levels, which peaked after 4 months and started to decline after 6 months of treatment, reaching levels below those detected at inclusion. In contrast, in most patients, dehydroepiandrosterone sulfate (DHEAS) levels remained unchanged, although a trend toward increased concentrations was observed in a few cases 3 months after the treatment was finished. Specific therapy also resulted in more balanced cortisol/DHEA and cortisol/DHEAS ratios. Etiologic treatment involves favorable immune and endocrine changes, which may account for its beneficial effects.

Effect of rhGH on the synthesis and secretion of VLDL to lymph and plasma from the intestine of the female rat.

The intravenous administration of rhGH (recombinant human Growth Hormone) to fasting female rats causes an increase in the rate of synthesis and secretion of VLDL (very low density lipoproteins). This phenomenon has three striking characteristics: (1) the demonstration of an unexpected lipogenic effect of rhGH, (2) its rapid occurrence after intravenous injection of the hormone and (3) the apparent dependence on the levels of circulating estrogens, as deduced by the lack of effect of rhGH on males and castrated females. The target tissue for the lipogenic effect was traced to the intestine by means of perfusion experiments of isolated duodenal loops. Impairment of liver blood supply discarded this tissue as the source of VLDL induced by rhGH. After a single dose of rhGH (T(1/2)=16min), the increase in plasma TAG (triacylglycerides) levels followed a positive exponential course that lasted ca. 3h. The same phenomenon (with no significant differences in kinetic parameters) was observed in three other experimental circumstances: fasting intact virgin female rats with impaired hepatic circulation, perfusion of isolated duodenum and sampling of mesenteric lymph. It is assumed that rhGH stimulates the synthesis of TAG and VLDL by the physiological mechanisms already present in enterocytes. Because increased plasma levels of VLDL and GH have been demonstrated in the last week of rat pregnancy, we believe that the reported phenomenon has physiological implications, hypothetically associated with fetal lung maturation. As an hypothesis, we suggest that the effect of growth hormone (of pituitary or placental origin) on the synthesis and secretion of VLDL by enterocytes uses a nongenomic pathway.

Perfil lipídico en niños y adolescentes de una población escolar / Lipid profile in a school population of children and adolescents

Introducción. La relación existente entre las alteraciones del metabolismo lipídico y el riesgo de enfermedades cardiovasculares, unida a otras condiciones como la obesidad, lleva a considerar la importancia del estudio del perfil lipídico desde edades tempranas. El objetivo de este trabajo fue estudiar el perfil lipídico de niños y adolescentes para aportar datos según edad y sexo comparándolos con los obtenidos por otros autores.Población, material y métodos. Se realizó un estudio descriptivo transversal de una muestra de 960 niños y adolescentes escolarizados, en distintas escuelas públicas de Rosario y alrededores, pertenecientes a una situación socioeconómica media o baja. Se incluyeron varones y mujeres de 5 a 18 años, sin antecedentes patológicos conocidos; se excluyeron aquellos con sobrepeso (6,6 por ciento).Resultados. Se determinaron parámetros antropométricos y bioquímicos. Los valores obtenidos fueron:colesterol: 164,9 más o menos 23,6 mg/dl; triglicéridos: 72,3 más menos 31,7 mg/dl; colesterol LDL: 96,1 más menos 23,6 mg/dl; colesterol HDL: 55,1 más menos 5,6 mg/dl; colesterol no HDL: 110,1 más menos 25,4mg/dl; colesterol/colesterol HDL: 3,03 más menos 0,59; colesterol LDL/colesterol HDL: 1,77 más menos 0,59. En la muestra estudiada, no se observaron diferencias estadísticamente significativas en los parámetros entre varones y mujeres ni entre los distintos grupos etarios, a excepción de los triglicéridos (significativamente mayores en mujeres de 5 a 11 años, p mayor 0,001).Conclusión. La comparación de nuestros resultados con los de otros autores nos permitió establecer diferencias con distinto grado de significación. El aporte del estudio del perfil lipídico en una población de niños y adolescentes podría ser de utilidad como referente para identificar factores de riesgo modificables y la necesidad de intervenciones conducentes a adoptar medidas de prevención desde edades tempranas.