Evaluation of potential biomarkers for the diagnosis and monitoring of Systemic Lupus Erythematosus using the Cytometric Beads Array (CBA).

BACKGROUND: Systemic Lupus Erythematosus (SLE) is an autoimmune, multisystemic disease. Currently diagnosis depends on complex criteria developed by the American College of Rheumatology. Moreover, the lack of specific biomarkers also challenges the diagnosis. METHODS: Inflammatory biomarkers such as IL-8, IP-10, MIG, MIP-1α and RANTES were measured in serum samples from SLE patients and subjects in control groups (patients with other autoimmune diseases and healthy individuals). Forty-six SLE patients (22 patients with low activity, SLEDAI-2 K ≤ 4, 24 patients with moderate/high activity, SLEDAI-2 K > 4), 42 patients with other autoimmune diseases (OAD group), and 8 healthy volunteers participated in this study. RESULTS: MIG (p < .001) and RANTES (p < .001) concentrations in SLE patients and healthy controls, and IP-10 concentrations in SLE patients with different disease activities (low activity, p < .01, moderate/high activity, p < .05) differed significantly. IL-8 (p < .001) and MIP-1α (p < .001) concentrations in SLE patients differed from those in patients from the OAD group. IL-8 (p < .05), IP-10 (p < .01), MIG (p < .05), MIP-1α (p < .001), and RANTES (p < .05) were correlated with SLE activity; their concentrations in SLE patients with low and moderate/high activity differed significantly. CONCLUSIONS: Given the findings of this study, one can envision the possibility of future use of some of these cytokines to assist in the screening of SLE patients, or even in monitoring disease activity.

Evaluation of miltefosine for the treatment of dogs naturally infected with L. infantum (=L. chagasi) in Brazil.

Dogs naturally infected with Leishmania Infantum (=L. chagasi) were treated with miltefosine using different therapeutic regimens. The animals were evaluated for clinical evolution, biochemical parameters, parasite load (by real-time PCR), cytokine levels and humoral response. After treatment and during the following 24 months, there was progressive clinical improvement and complete recovery in 50% (7/14) of the treated animals. There was a decrease in the smear positivity of the bone marrow after treatment, and there was also a gradual and constant decrease in positive cultures at the end of the follow-up period. However, the PCR detection of parasite DNA remained positive. In general, all animals presented a significant increase in parasite load 6 months after treatment. The IFN-γ levels in all the groups tended to increase during follow-up period, regardless of the miltefosine dose administered. The IL-4 and IL-10 levels of the animals tended to decrease during follow-up, except after 300 days when only IL-10 increased. The serum antibodies identified antigens that ranged from 116 kDa to less than 29 kDa in the Western blot assay. Furthermore, 300 days after treatment, qualitative and quantitative differences in the antigen profiles were observed. Antigens of 97 and 46 kDa were the most intensely recognized. Higher levels of antigen-specific Leishmania IgG were detected before and 300 days after treatment in all groups. Taking together, the improvement in the clinical symptoms was not followed by parasitological clearance, suggesting that treatment with miltefosine is not recommended, especially in endemic areas like Brazil, where children are the major victims and dogs are involved in the maintenance of the parasite cycle.

Natural killer cell subpopulations in putative resistant individuals and patients with active Mycobacterium tuberculosis infection.

Herein, we intended to perform flow-cytometric analyses of peripheral blood NK-cell subsets in patients with active tuberculosis (TB) and those putative resistant subjects displaying positive tuberculin skin test (TST+) and compared with TST- healthy controls. Our findings demonstrated distinct phenotypic features in TST+ as compared with TB. While lower values of NK-cells with increased frequency of CD3-CD16+ CD56- and CD3-CD16-CD56+ subsets besides lower frequency of CD3-CD16+ CD56+ NK-cells was observed in TST+, unaltered levels of NK-cells with increased levels of CD3-CD16+ CD56- NK-cells with lower frequency of CD3-CD16+ CD56+ NK-cells was found in TB. Additional analysis highlighted a shift towards increased levels of CD3-CD16-/+CD56bright NK-cells as the hallmark of TST+, whereas unaltered frequency was observed in TB. Increased levels of CD3+CD56+ cells were observed in both TST+ and TB. Further focusing on the monocyte/NK-cell network, we have reported that enhanced frequency of CD14+ CD16+ monocytes particularly observed in TST+. Outstanding were the distinct correlation profiles observed between CD3-CD16-CD56+ NK-cells and CD3+ CD56+ cells CD14+ CD16+ monocytes for TST+ and TB. These data suggested that high levels of CD3-CD16-CD56+ NK-cells aside CD14+ CD16+ monocytes as well as non-concurrent increment of CD3+ CD56+ cells, may be involved in protective mechanisms in putative tuberculosis-resistant individuals. On the other hand, the basal levels of macrophage-like monocytes despite its positive correlation with increased levels of CD3+ CD56+ cells may count for the lack of the protective immunity in patients with active tuberculosis. Further studies focusing on the cytokine profiling of peripheral blood innate immunity cells before and after chemotherapeutic treatment are currently under evaluation.

Pulmonary tuberculosis: evaluation of interferon-gamma levels as an immunological healing marker based on the response to the Bacillus Calmette-Guerin.

Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis whose interaction with the host may lead to a cell-mediated protective immune response. The presence of interferon gamma (IFN-gamma) is related to this response. With the purpose of understanding the immunological mechanisms involved in this protection, the lymphoproliferative response, IFN-gamma and other cytokines like interleukin (IL-5, IL-10), and tumor necrosis factor alpha (TNF-alpha) were evaluated before and after the use of anti-TB drugs on 30 patients with active TB disease, 24 healthy household contacts of active TB patients, with positive purified protein derivative (PPD) skin tests (induration > 10 mm), and 34 asymptomatic individuals with negative PPD skin test results (induration < 5 mm). The positive lymphoproliferative response among peripheral blood mononuclear cells of patients showed high levels of IFN-gamma, TNF-alpha, and IL-10. No significant levels of IL-5 were detected. After treatment with rifampicina, isoniazida, and pirazinamida, only the levels of IFN-gamma increased significantly (p < 0.01). These results highlight the need for further evaluation of IFN-gamma production as a healing prognostic of patients treated.

Pulmonary tuberculosis: evaluation of interferon-gamma levels as an immunological healing marker based on the response to the Bacillus Calmette-Guerin

Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis whose interaction with the host may lead to a cell-mediated protective immune response. The presence of interferon-gamma is related to this response. With the purpose of understanding the immunological mechanisms involved in this protection, the lymphoproliferative response, IFN-gamma and other cytokines like interleukin (IL-5, IL-10), and tumor necrosis factor alpha (TNF-alfa) were evaluated before and after the use of anti-TB drugs on 30 patients with active TB disease, 24 healthy household contacts of active TB patients, with positive purified protein derivative (PPD) skin tests (induration > 10 mm), and 34 asymptomatic individuals with negative PPD skin test results (induration < 5 mm). The positive lymphoproliferative response among peripheral blood mononuclear cells of patients showed high levels of IFN-gamma, TNF-alfa, and IL-10. No significant levels of IL-5 were detected. After treatment with rifampicina, isoniazida, and pirazinamida, only the levels of IFN-gamma increased significantly (p < 0.01). These results highlight the need for further evaluation of IFN-gamma production as a healing prognostic of patients treated.

Technetium-99m labeling anti-amastigote polyclonal antibodies of Leishmania amazonensis.

Anti-amastigote polyclonal antibody (IgG) was incubated with solutions of stannous chloride and sodium borohidride. After that, 3.7 MBq of technetium-99m (99mTc) was added. A labeling yield of the antibody about 84% was obtained. After filtration of 99mTc-IgG, the radiochemical purity increased from 84 to 95%. The labeling of IgG with 99mTc did not modify the immunoreactivity of the antibody, since it was able to identify in vitro and in vivo the specific antigen of Leishmania amazonensis.