Effect of cholecalciferol as adjunctive therapy with insulin on protective immunologic profile and decline of residual ß-cell function in new-onset type 1 diabetes mellitus.

OBJECTIVE: To evaluate the effect of vitamin D3 on cytokine levels, regulatory T cells, and residual ß-cell function decline when cholecalciferol (vitamin D3 administered therapeutically) is given as adjunctive therapy with insulin in new-onset type 1 diabetes mellitus (T1DM). DESIGN AND SETTING: An 18-month (March 10, 2006, to October 28, 2010) randomized, double-blind, placebo-controlled trial was conducted at the Diabetes Center of São Paulo Federal University, São Paulo, Brazil. PARTICIPANTS: Thirty-eight patients with new-onset T1DM with fasting serum C-peptide levels greater than or equal to 0.6 ng/mL were randomly assigned to receive daily oral therapy of cholecalciferol, 2000 IU, or placebo. MAIN OUTCOME MEASURE: Levels of proinflammatory and anti-inflammatory cytokines, chemokines, regulatory T cells, hemoglobin A1c, and C-peptide; body mass index; and insulin daily dose. RESULTS: Mean (SD) chemokine ligand 2 (monocyte chemoattractant protein 1) levels were significantly higher (184.6 [101.1] vs 121.4 [55.8] pg/mL) at 12 months, as well as the increase in regulatory T-cell percentage (4.55% [1.5%] vs 3.34% [1.8%]) with cholecalciferol vs placebo. The cumulative incidence of progression to undetectable (≤0.1 ng/mL) fasting C-peptide reached 18.7% in the cholecalciferol group and 62.5% in the placebo group; stimulated C-peptide reached 6.2% in the cholecalciferol group and 37.5% in the placebo group at 18 months. Body mass index, hemoglobin A1c level, and insulin requirements were similar between the 2 groups. CONCLUSIONS: Cholecalciferol used as adjunctive therapy with insulin is safe and associated with a protective immunologic effect and slow decline of residual ß-cell function in patients with new-onset T1DM. Cholecalciferol may be an interesting adjuvant in T1DM prevention trials.

HIV-1 induces NALP3-inflammasome expression and interleukin-1ß secretion in dendritic cells from healthy individuals but not from HIV-positive patients.

OBJECTIVE: NALP3-inflammasome is an innate mechanism, alternative to type-1 interferon, which is able to recognize nucleic acids and viruses in the cytoplasm and to induce pro-inflammatory response. Here, we hypothesized the involvement of inflammasome in the early defense against HIV-1 and in the full maturation of dendritic cells: for this, we evaluated the response of dendritic cells pulsed with HIV-1 in terms of inflammasome activation in healthy donors. Moreover, inflammasome response to HIV was evaluated in HIV-infected individuals. DESIGN AND METHODS: Monocyte-derived dendritic cells isolated from 20 healthy individuals (HC-DC) and 20 HIV-1-infected patients (HIV-DC) were pulsed with alditrithiol-2-inactivated HIV-1. We then analyzed inflammasome genes expression and interleukin-1ß (IL-1ß) secretion. RESULTS: In HC-DC, HIV-1 induced higher NLRP3/NALP3 mRNA expression compared with other inflammasome genes such as NALP1/NLRP1 or IPAF/NLRC4 (P < 0.001). This augmented expression was accompanied by CASP1-increased and IL1B-increased mRNA levels and by a significant increment of IL-1ß secretion (P < 0.05). Otherwise, HIV-1 failed to activate inflammasome and cytokine production in HIV-DC. HIV-DC showed an increased NLRP3/NALP3 basal expression, suggesting a chronic inflammatory profile of patients' immune cells. CONCLUSION: HIV-1 was able to induce a NALP3-inflammasome response in healthy individuals, indicating that this inflammasome could play a role in the first steps of HIV-1 infection; the consequent inflammatory process may be important for directing host immune response against the virus and/or disease progression. HIV-DC seemed to be chronically activated, but unresponsive against pathogens. Our findings could be of interest considering the ongoing research about dendritic cell manipulation and therapeutic strategies for AIDS involving dendritic cell-based immune-vaccines.

Geração in vitro de células T efetoras e células T reguladoras mediada por células dendríticas pulsadas com vírus autólogo de pacientes infectados pelo HIV-1 / In vitro generation of effector T cells and regulatory T cells by monocyte-derived dendritic cells from HIV-1-infected patients pulsed with autologous virus

Imunização terapêutica utilizando células dendríticas derivadas de monócitos (MoDCs) pulsadas com antígenos de HIV constitui um meio promissor de potencializar a resposta imune específica anti-HIV em pacientes infectados. Neste contexto, é importante ressaltar que células dendríticas além de estimular a resposta imune específica, podem ser capazes de promover a tolerância periférica em linfócitos T CD4+ e T CD8+ ao induzir deleção, anergia ou através da expansão de células T reguladoras (T regs). Experimentos in vitro foram conduzidos para avaliar a capacidade de MoDCs pulsadas com HIV autólogo inativado em induzir apoptose celular, respostas celulares específicas e a geração de T regs. Os pacientes avaliados neste estudo foram indivíduos infectados pelo HIV, sem uso de tratamento antirretroviral (n = 14) com número de células T CD4+ acima de 350 células/L. MoDCs foram geradas a partir de células mononucleares de sangue periférico e em seguida foram pulsadas com vírus autólogo inativado por Aldrithiol-2, tratadas com estímulo para maturação e então cultivadas com linfócitos autólogos. A apoptose de linfócitos T e MoDCs e a frequência de células efetoras e reguladoras foram avaliadas por citometria de fluxo. Os resultados obtidos mostraram que não houve diferença nos níveis de apoptose de células T CD4+, T CD8+ ou MoDCs entre os grupos pulsadas e não pulsadas com HIV inativado. Foi observado que tanto MoDCs pulsadas quanto aquelas não pulsadas com o vírus autólogo inativado foram capazes de induzir células T CD4+ secretoras de IFN-, enquanto que apenas MoDCs pulsadas levou a um aumento no percentual de células T CD8+ efetoras. Pacientes com contagem de células T CD4+ acima de 500 células/L apresentaram um percentual maior de células T CD4+ secretoras de IFN após estimulo de MoDCs pulsadas. Esta diferença não foi observada em células T CD8 +. T regs também foram induzidas in vitro após cocultivo com MoDCs. Níveis basais mais elevados de T regs foram encontrados...

Therapeutic immunization using inactivated autologous HIVpulsed dendritic cells (DCs) is a promising strategy to enhance specific anti-HIV immune responses in infected patients. In this context, it is important to note that DC besides stimulate a specific immune response, may be able to promote tolerance in peripheral CD4 + and CD8 + T cells inducing deletion, anergy or through expansion of regulatory T cells (T reg). In vitro experiments were conducted to evaluate the capacity of autologous HIVstimulated DC to induce apoptosis, effector cellular T cell responses and T reg generation. For these purposes, we used peripheral blood from HAARTnaïve HIVinfected patients (n=14) with CD4+ T cell counts above 350 cell/L for generation of monocytederived DC (MoDC). MoDC were pulsed with aldrithiol-2 (AT-2)-inactivated autologous virus and matured. MoDC were then cocultured with autologous lymphocytes and the apoptosis, production of IFN- and T reg cell frequency were evaluated by flow cytometry. There was no difference in the rate of apoptosis of CD4, CD8 T cells or MoDC between the groups pulsed and not pulsed with inactivated HIV. MoDC pulsed or not with inactivated autologous virus induced IFN- +CD4+ T cells, whereas only pulsed MoDC were able to increase effector CD8+ T cells percentage along the time culture. Patients with CD4+ T cell counts above 500 had an increased percentage of CD4 + T cells secreting IFN- upon DC pulsed with HIV. This difference was not observed in CD8 + T cells. Interestingly, T reg were also induced in vitro after MoDC cocultivation. Higher baseline T reg counts were found in patients with lower plasma viral loads. These results show that MoDC pulsed with HIV-1 are able to induce effector lymphocyte but also elevate the frequency of T reg in vitro. Patients with higher CD4+ T cell counts are able to respond more efficiently to the stimulation with pulsed MoDC, and persistent viremia in chronic HIV infection is associated with significant...(AUA)

Enhanced removal from the plasma of LDL-like nanoemulsion cholesteryl ester in trained men compared with sedentary healthy men.

The objective of this study was to evaluate the effects of exercise training on plasma removal of a cholesterol-rich nanoemulsion (LDE) that mimics low-density lipoprotein (LDL) lipid structure and binds to LDL receptors. LDE-derived cholesteryl ester plasma kinetics was studied in 24 exercise-trained and 20 sedentary male subjects. LDE labeled with [(14)C]cholesteryl ester was injected intravenously, and plasma samples were collected over a 24-h period to determine radioisotope decay curves. LDL cholesterol concentration was similar in both groups. Fractional clearance rate (FCR) of the nanoemulsion label was greater in the exercise-trained group compared with the sedentary group (0.138 +/- 0.152 and 0.0261 +/- 0.023 h(-1), respectively). A positive correlation was found (r = 0.60, P < 0.01) between FCR and peak O(2) consumption in trained subjects. Circulating oxidized LDL levels were lower in trained subjects compared with the sedentary group (9.0 +/- 2.0 and 16.0 +/- 3.0 mU/l). LDE was also injected into control and LDL receptor gene knockout mice submitted and not submitted to training. Muscle LDE uptake percentage was increased in the trained mice compared with the untrained mice (1.1 +/- 0.8 and 0.2 +/- 0.1, respectively, P < 0.0001) in the control group but not in the knockout animals, indicating that the LDL receptor is involved in the increased uptake elicited by exercise. These results show that exercise training increases LDE plasma removal, which in turn suggests that it also increases LDL receptors or LDL receptor activity.