Translation, cultural adaptation to Brazil and validation of the venous leg ulcer quality of life questionnaire (VLU-QoL-Br)

Objetivo:traduzir o instrumento Venous legulcer quality of life questionnaire (VLU-QoL), adaptá-lo culturalmente para o português do Brasil e validá-lo com pacientes do Hospital das Clínicas da Faculdade de Medicina de Botucatu (FMB) da Universidade Estadual Paulista (Unesp). Métodos:o questionário foi traduzido por um tradutor profissional e por dois dermatologistas especialistas na área de úlceras venosas (UV), sendo reformulado em reunião com os três tradutores. O constructo (VLU-QoL-Br) foi submetido a pré-entrevista com 10 portadores de UV para a adaptação da linguagem. Posteriormente, foi aplicado em pacientes do HC-Unesp, e como teste-reteste para verificação de sua reprodutibilidade. Resultados:foram avaliados 82 pacientes, sendo 56 (68%) do sexo feminino. A idade média foi de 67,3 anos. O questionário foi traduzido, adaptado e aplicado aos pacientes. O constructo apresentou alta consistência interna (alfa= 0,94) e adequada correlação item-total. Quando avaliados os 32 retestes, observou-se correlação intraclasse para concordância de 0,78 (p < 0,01), indicando boa reprodutibilidade do constructo. A análise fatorial confirmatória corroborou as dimensões do questionário original: atividades, psicológico e sintomas. Escores do VLU-QoL-Br se associaram, independentemente, à área total das úlceras e a menor escolaridade dos sujeitos (p < 0,01). Conclusão:a tradução, a adaptação e a validação do questionário VLU-Qol-Br demonstrou boa performance psicométrica, permitindo seu uso clínico no Brasil. É importante avaliar seu desempenho em outras regiões e em diferentes amostras de indivíduos.Objective:translating the Venous leg ulcer quality of life questionnaire (VLU-QoL), and culturally adapting it to Brazilian Portuguese and validate it with patients at the Hospital das Clínicas of the Botucatu Medical School - Unesp. Methods:the questionnaire was translated by a professional translator and two dermatologists specialized in the area of venous ulcers (VU), reformulated in a meeting of the three translators. The construct (VLU-QoL-Br) was submitted to pre-interviews with ten VU patients for adaptation of the language. Subsequently, it was applied to patients at the HC-Unesp, and for test-retest reliability for verification of its reproducibility. Results:82 patients were evaluated, with 56 (68%) women. The age average was 67.3 years. The questionnaire was translated, adapted and applied to the patients. The construct presented high internal consistency (alpha = 0.94) and adequate item-total correlation. When the 32 retests were evaluated, an intra-class correlation was noted for concordance of 0.78 (p<0.01), indicating good reproducibility of the construct. The confirmatory factor analysis corroborated the dimensions of the original questionnaire: activities, psychologies, and symptoms. VLU-QoL-Br scores were associated, independently, to the total area of the ulcers and a lower education level of the subjects (p<0.01). Conclusion:the translation, adaptation and validation of the VLU-QoL-Br questionnaire were concluded, demonstrating good psychometric performance, and enabling its clinical use in Brazil. It is important to evaluate its performance in other regions and different samples of individuals.Unesp FMBUnesp FMB Dermatology DepartmentUnesp FMBUnesp FMB Dermatology Departmen

Granulocyte macrophage colony-stimulating factor enhances the modulatory effect of cytokines on monocyte-derived multinucleated giant cell formation and fungicidal activity against Paracoccidioides brasiliensis

Multinucleated giant cells (MGC) are cells present in characteristic granulomatous inflammation induced by intracellular infectious agents or foreign materials. The present study evaluated the modulatory effect of granulocyte macrophage colony-stimulating factor (GM-CSF) in association with other cytokines such as interferon-gamma (IFN-γ), tumour necrosis factor-alpha, interleukin (IL)-10 or transforming growth factor beta (TGF-β1) on the formation of MGC from human peripheral blood monocytes stimulated with Paracoccidioides brasiliensis antigen (PbAg). The generation of MGC was determined by fusion index (FI) and the fungicidal activity of these cells was evaluated after 4 h of MGC co-cultured with viable yeast cells of P. brasiliensis strain 18 (Pb18). The results showed that monocytes incubated with PbAg and GM-CSF plus IFN-γ had a significantly higher FI than in all the other cultures, while the addition of IL-10 or TGF-β1 had a suppressive effect on MGC generation. Monocytes incubated with both pro and anti-inflammatory cytokines had a higher induction of foreign body-type MGC rather than Langhans-type MGC. MGC stimulated with PbAg and GM-CSF in association with the other cytokines had increased fungicidal activity and the presence of GM-CSF also partially inhibited the suppressive effects of IL-10 and TGF-β1. Together, these results suggest that GM-CSF is a positive modulator of PbAg-stimulated MGC generation and on the fungicidal activity against Pb18

Phenotypic and functional evaluations of peripheral blood monocytes from chronic-form paracoccidioidomycosis patients before and after treatment

Background: Paracoccidioidomycosis (PCM) is systemic mycosis caused by the thermal dimorphic fungus of genus Paracoccidioides, leading to either acute/subacute (AF) or chronic (CF) clinical forms. Numerous CF patients after treatment exhibit sequels, such as pulmonary and adrenal fibrosis. Monocytes are cells that are involved in the inflammatory response during active infection as well as in the fibrogenesis. These cells comprise a heterogeneous population with distinct phenotypic and functional activities. The scope of this study was to identify changes regarding functional and phenotypical aspects in monocytes comparing CF PCM patients on antifungal treatment versus non-treated patients (PMC-p).Methods: Twenty-three CF PCM composed of 11 non-treated patients (NTG) and 12 patients in apparent cure (ACG) were studied. Sixteen healthy individuals were used as control group (CG). Monocyte subsets were determined by immunophenotyping based on CD14 and CD16 expression. Cellular function was measured in vitro with and without stimulation with lipopolysaccharide (LPS) and P. brasiliensis exoantigen (PbAg) for 24 hours. Independent samples were compared using unpaired t tests, dependent samples were analyzed by paired t-test. Groups of more than two independent samples were analyzed using an ANOVA, with Tukey's post-test. Significance was set up at p < 0.05.Results: Our results showed high counts of peripheral blood CD14(+)CD16(+) and CD14(+)CD16(++) monocytes in untreated PCM-p accompanied by intense production of pro-inflammatory cytokines (IL-1 beta and TNF-alpha) and profibrotic growth factors (TGF-beta 1 and bFGF) by monocytes challenged with P. brasiliensis antigens. After the introduction of antifungal therapy, the counts of CD14(+)CD16(+) cells returned to baseline while CD14(+)CD16(++) counts remained high. Interestingly, counts of CD14(+)CD16(++) monocytes remained elevated even 52 +/- 7 months after successful antifungal treatment. Furthermore, the ACG-patients showed preserved pro-inflammatory activity in the presence of specific antigen stimuli and high spontaneous production of TNF-a by monocytes.Conclusions: Infection with Paracoccidioides leads to initiation of a specific proinflammatory response by monocytes of PCM-p during active disease and in the apparent cure. A profibrotic profile by monocytes was observed only at admission. Furthermore, PCM-p with apparent cure showed high spontaneous production of TNF-alpha and high counts of CD14(+)CD16(++) monocytes, probably induced by hypoxia duo to fibrotic sequelae.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Univ Estadual Paulista, UNESP, Fac Med Botucatu, BR-18618970 Botucatu, SP, BrazilUniv Estadual Paulista, UNESP, Dept Ciencias Biol, Fac Ciencias,Lab Imunopatol Expt LIPE, BR-17047001 Bauru, SP, BrazilUniv Estadual Paulista, UNESP, Fac Med Botucatu, BR-18618970 Botucatu, SP, BrazilUniv Estadual Paulista, UNESP, Dept Ciencias Biol, Fac Ciencias,Lab Imunopatol Expt LIPE, BR-17047001 Bauru, SP, BrazilFAPESP: 09/51105-

Therapeutic Administration of Recombinant Paracoccin Confers Protection against Paracoccidioides brasiliensis Infection: Involvement of TLRs.

Paracoccin (PCN) is an N-acetylglucosamine-binding lectin from the human pathogenic fungus Paracoccidioides brasiliensis. Recombinant PCN (rPCN) induces a T helper (Th) 1 immune response when prophylactically administered to BALB/c mice, protecting them against subsequent challenge with P. brasiliensis. In this study, we investigated the therapeutic effect of rPCN in experimental paracoccidioidomycosis (PCM) and the mechanism accounting for its beneficial action. METHODOLOGY/PRINCIPAL FINDINGS: Four distinct regimens of rPCN administration were assayed to identify which was the most protective, relative to vehicle administration. In all rPCN-treated mice, pulmonary granulomas were less numerous and more compact. Moreover, fewer colony-forming units were recovered from the lungs of rPCN-treated mice. Although all therapeutic regimens of rPCN were protective, maximal efficacy was obtained with two subcutaneous injections of 0.5 microg rPCN at 3 and 10 days after infection. The rPCN treatment was also associated with higher pulmonary levels of IL-12, IFN-gamma, TNF-alpha, nitric oxide (NO), and IL-10, without IL-4 augmentation. Encouraged by the pulmonary cytokine profile of treated mice and by the fact that in vitro rPCN-stimulated macrophages released high levels of IL-12, we investigated the interaction of rPCN with Toll-like receptors (TLRs). Using a reporter assay in transfected HEK293T cells, we verified that rPCN activated TLR2 and TLR4. The activation occurred independently of TLR2 heterodimerization with TLR1 or TLR6 and did not require the presence of the CD14 or CD36 co-receptors. The interaction between rPCN and TLR2 depended on carbohydrate recognition because it was affected by mutation of the receptor\u27s N-glycosylation sites. The fourth TLR2 N-glycan was especially critical for the rPCN-TLR2 interaction. CONCLUSIONS/SIGNIFICANCE: Based on our results, we propose that PCN acts as a TLR agonist. PCN binds to N-glycans on TLRs, triggers regulated Th1 immunity, and exerts a therapeutic effect against P. brasiliensis infection