Recombinant 60-kDa heat shock protein from Paracoccidioides brasiliensis: is it a good antigen for serological diagnosis of paracoccidioidomycosis?

Paracoccidioides brasiliensis and P. lutzii are fungi that cause paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in South America. For serological diagnosis, although 43-kDa glycoprotein (gp43) is regarded as highly specific for PCM, the occurrence of false negative reactions in sera from patients infected with P. lutzii suggests that preparation with only one antigen is not recommended. Heat shock proteins are feasible alternatives as a second antigen because they are often highly immunogenic. In this study, we evaluated the usefulness of recombinant 60-kDa heat shock protein from P. brasiliensis (rPbHsp60) for the serological diagnosis of PCM. Using western blotting assay, we observed that 77.3% of the sera from PCM patients were positive to rPbHsp60, with 90.9% positivity to recombinant gp43 (rgp43). More importantly, sera from healthy subjects had 27% positivity to rPbHsp60 and none to rgp43. When rPbHsp60 was used in ELISA, we did not observe significant differences between the reactions with sera from PCM patients and healthy subjects, while the difference was clearly evident when the antigen was rgp43. Furthermore, rPbHsp60 was recognized by sera from patients with histoplasmosis, aspergillosis, sporotrichosis or tuberculosis in an ELISA test. These results show that rPbHsp60 is not a good antigen for PCM diagnosis.

B cells expressing IL-10 mRNA modulate memory T cells after DNA-Hsp65 immunization

In DNA vaccines, the gene of interest is cloned into a bacterial plasmid that is engineered to induce protein production for long periods in eukaryotic cells. Previous research has shown that the intramuscular immunization of BALB/c mice with a naked plasmid DNA fragment encoding the Mycobacterium leprae 65-kDa heat-shock protein (pcDNA3-Hsp65) induces protection against M. tuberculosis challenge. A key stage in the protective immune response after immunization is the generation of memory T cells. Previously, we have shown that B cells capture plasmid DNA-Hsp65 and thereby modulate the formation of CD8+ memory T cells after M. tuberculosis challenge in mice. Therefore, clarifying how B cells act as part of the protective immune response after DNA immunization is important for the development of more-effective vaccines. The aim of this study was to investigate the mechanisms by which B cells modulate memory T cells after DNA-Hsp65 immunization. C57BL/6 and BKO mice were injected three times, at 15-day intervals, with 100 µg naked pcDNA-Hsp65 per mouse. Thirty days after immunization, the percentages of effector memory T (TEM) cells (CD4+ and CD8+/CD44high/CD62Llow) and memory CD8+ T cells (CD8+/CD44high/CD62Llow/CD127+) were measured with flow cytometry. Interferon γ, interleukin 12 (IL-12), and IL-10 mRNAs were also quantified in whole spleen cells and purified B cells (CD43−) with real-time qPCR. Our data suggest that a B-cell subpopulation expressing IL-10 downregulated proinflammatory cytokine expression in the spleen, increasing the survival of CD4+ TEM cells and CD8+ TEM/CD127+ cells.

Antigen-presenting cells transfected with Hsp65 messenger RNA fail to treat experimental tuberculosis

In the last several years, the use of dendritic cells has been studied as a therapeutic strategy against tumors. Dendritic cells can be pulsed with peptides or full-length protein, or they can be transfected with DNA or RNA. However, comparative studies suggest that transfecting dendritic cells with messenger RNA (mRNA) is superior to other antigen-loading techniques in generating immunocompetent dendritic cells. In the present study, we evaluated a new therapeutic strategy to fight tuberculosis using dendritic cells and macrophages transfected with Hsp65 mRNA. First, we demonstrated that antigen-presenting cells transfected with Hsp65 mRNA exhibit a higher level of expression of co-stimulatory molecules, suggesting that Hsp65 mRNA has immunostimulatory properties. We also demonstrated that spleen cells obtained from animals immunized with mock and Hsp65 mRNA-transfected dendritic cells were able to generate a mixed Th1/Th2 response with production not only of IFN-γ but also of IL-5 and IL-10. In contrast, cells recovered from mice immunized with Hsp65 mRNA-transfected macrophages were able to produce only IL-5. When mice were infected with Mycobacterium tuberculosis and treated with antigen-presenting cells transfected with Hsp65 mRNA (therapeutic immunization), we did not detect any decrease in the lung bacterial load or any preservation of the lung parenchyma, indicating the inability of transfected cells to confer curative effects against tuberculosis. In spite of the lack of therapeutic efficacy, this study reports for the first time the use of antigen-presenting cells transfected with mRNA in experimental tuberculosis.

HIV-1 viremia during the first 28 weeks of pregnancy is not associated with mother-to-child transmission

It is currently recommended that antiretroviral prophylaxis to prevent mother-to-child transmission (MTCT) of HIV be initiated at 14 weeks of gestation. However, the relevance of early-gestation HIV viral load level for intrauterine MTCT is unknown. The objective of this study was to determine the relationship between prenatal maternal viral load and intrauterine MTCT. Records of HIV-infected pregnant women in two centers in Brazil, from 1999 to 2004 were analyzed. Three pregnancy periods were considered: earlier than 14 weeks, 14 to 27 6/7 weeks, and 28 weeks of gestation or more. Peripartum HIV exposure was also computed. Maximum viral load in each period was the measure of HIV exposure. Four hundred fifty-seven HIV-infected pregnant women were evaluated, but 53 were excluded. The MTCT rate was 0.49 percent (2/404-95 percent confidence interval (CI95) = 0.14-1.79 percent). Newborns were not breast-fed. Median viral load for the earlier-than-14-week period was 9,900 copies/mL (P25-75 1,000-50,775 copies/mL), 8,350 copies/mL (P25-75 707-42,000 copies/mL) for the 14 to 27 6/7-week period, and 435 copies/mL (P25-75 90-7,775 copies/mL) after the 28-week period. The peripartum median viral load was 400 copies/mL (P25-75 80-500 copies/mL). MTCT in mothers with VL > 1,000 copies/mL during the first 14 weeks (0.67 percent, 2/298) was not different from those with VL =1,000 copies/mL (0.0 percent, 0/96, P=1). Analogously, in the 14 to 27 6/7-week period, MTCT was similar in groups with VL higher (0.68 percent, 2/292) or lower (0 percent, 0/106) than 1,000 copies/mL (P=1). Regarding VL >1,000 copies/mL at 28-weeks-or-later and at peripartum periods, MTCT rates were 1.15 percent (2/173, P = 0.18) and 2.8 percent (2/71, P = 0.03), respectively. Intrauterine transmission does not seem to be influenced by HIV viremia during the first 28 weeks of pregnancy.

Risk factors for poor immune response to influenza vaccination in elderly people

Influenza vaccination of elderly people is efficacious and cost effective for the prevention of influenza and its complications. Some studies have pointed out low immunogenicity in this group. Health status has been poorly investigated as a risk factor that may influence the immune response to influenza vaccine. We established an immunization response study of a highly-matched elderly population in a nursing home. One-hundred-twenty subjects of Ashkenazian origin had their vaccine-induced antibody response assessed. Good response was obtained in 30.8 percent (37/120), and 31.7 percent (38/120) did not react. A lack of good response was found to be associated with dementia (P=0.016) in a multivariate analysis. In addition to dementia, malnutrition was frequently observed among poor responders, suggesting that these factors should be considered in vaccination studies. Chemoprophylaxis in addition to vaccination for elderly presenting dementia should be considered, particularly for those people living nursing homes.

Desenvolvimento de uma estratégia para otimizar a indicação de endoscopia digestiva alta em pacientes com dispepsia atendidos em nível primário de saúde / Development of a strategy to optimize the indication to the upper gastrointestinal endoscopy in patients with dyspepsia at the primary health care level

OBJETIVO: Desenvolver uma estratégia capaz de otimizar a indicação de endoscopia digestiva alta (esofagogastroduodenoscopia-EGD) em pacientes com dispepsia e sem sinal de alarme para doença orgânica, baseada em variáveis clínicas e sociais com maior valor em discriminar indivíduos com EGD com alteração de indivíduos com EGD normal. CASUÍSTICA E MÉTODO: 200 pacientes (idade: 16-76 anos) com dispepsia e sem evidência de doença orgânica, atendidos em nível primário, foram entrevistados com um questionário estruturado e submetidos a EGD. Análise de regressão logística múltipla identificou variáveis com maior valor em discriminar indivíduos com EGD com alteração de indivíduos com EGD normal, bem como indivíduos com EGD com úlcera péptica daqueles com EGD normal. RESULTADOS: As variáveis com maior valor em discriminar indivíduos com EGD com alteração daqueles com EGD normal foram: idade igual ou superior a 45 anos e sexo masculino. Para discriminar indivíduos com EGD com úlcera péptica daqueles com EGD normal, as variáveis foram: idade igual ou superior a 45 anos; sexo masculino; tabagismo; empachamento e absenteísmo. Um modelo prático foi desenvolvido, visando a apoiar a decisão de indicar ou não EGD, apresentando sensibilidade de 78,9 por cento, especificidade de 51,7 por cento, valor preditivo positivo (VPP) de 39,4 por cento e preditivo negativo (VPN) de 86 por cento, para a identificação de indivíduos com EGD com alteração; e sensibilidade de 84 por cento, especificidade de 70 por cento, VPP de 38 por cento e VPN de 95 por cento, para a identificação de indivíduos com EGD com úlcera péptica. CONCLUSÃO: Uma estratégia baseada em variáveis clínicas e sociais de pacientes com dispepsia poderia potencialmente diminuir o número de EGD desnecessárias.

DNA Vaccine for the Prevention and Treatment fo Tuberculosis

Tradicional systems for developing drugs and vaccines are failing spectaculary to deliver the goods in the fight against tuberculosis (TB). The disease that afflicts the developing world defies the imagination in its scale. One third of the world's population - 2 billion people - is infected with Mycobacterium tuberculosis, and 16 million have active TB. Shockingly, TB hit an all-time high in 1999 with 8 million new cases - 95 per cent of them in developing countries - and 2 million deaths. The disease is spreading rapidly throughout the world. The toll is set to rise; AIDS activates the dormant form of the disease, while multidrug resistance is spreading across the planet. The last new drug for TB was introduced over thirty years ago and industry has been reluctant to invest in discovering new families of drugs because of the financial risks in investing in products destined largely for developing country markets. If global health is left to market forces, historians will remember this era as one in which humanity stood idly by while half the planet languished in sickness. Fortunately some researchers have realized this, and are driving forward new models for TB therapy and vaccine discovery. One of the latest sign of this trend is the development of a DNA vaccine for the prevention and treatment of TB by our research group. Over the last few years, some of our experiments in wich mycobacterial antigens were presented to the immune system, as of they were viral antigens (DNA vaccine), have had a significant impact on our understanding of protective immunity against tuberculosis. They also markedly enhanced the prospects for new vaccines. We now know that individual mycobacterial-protein antigens expressed from DNA-vaccine constructs can confer protection equal to that from live BCG vaccine in mice. A critical determinant of the outcome of immunization appears to be the degree to which antigen-specific cytotoxic T cells are generated by the immune response. We have demonstrated that DNA vaccination is an affective way of establishing long lasting cytotoxic T-cell memory and protection against tuberculosis. Moreover, our new preclinical work shows that DNA vaccines, initially designed to prevent infection, can also have a dramatic therapeutic action. In infected mice, the immune response can be caused to switch from one that is relatively inefficient and gives bacterial stasis to one that kills the bacteria, eliminating...

Estudo caso-controle da infecçäo pelo HIV em pacientes em programa de hemodiálises / Control case study of HIV infection in dialysis patients

O Brasil é o quarto país no mundo em número de casos de AIDS, apresentando elevada proporçäo da doença adquirida através de transfusäo de sangue. Uma vez que os pacientes com insuficiência renal crônica em programa de hemodiálise säo freqüentemente submetidos a transfusöes sanguíneas, constituem-se em grupo com risco potencial de exposiçäo ao HIV. Estudou-se a prevalência de infecçäo pelo HIV em 117 pacientes em programa de hemodiálise na Casa de Saúde Santa Marcelina, identificando-se nove (7,7%) pacientes soropositivos pelos métodos ELISA e Western Blot. Nenhum paciente tinha outro fator de risco para infecçäo, além das transfusöes. Cada pacientes soropositivo foi comparado com três controles soronegativos, pareados por sexo. O número médio de transfusöes foi de 29,3 unidades para os casos e 26,7 unidades para os controles. O tempo médio de diálise foi de 28,2 e 27,2 meses para casos e controles, respectivamente. O risco de infecçäo pelo HIV näo foi associado ao número de transfusöes ou ao tempo de diálise. A relaçäo OKT/OKT foi menor ou igual a um em todos os pacientes infectados

Miosite lúpica pseudogranulomatosa / Pseudogranulomatoues lupic myositis

Os autores relatam um caso de LES, cuja sintomatologia principal era dada por uma importante miopatia. Pela primeira vez na literatura, estudou-se a biópsia muscular por técnicas histoquímicas, concluindo-se por uma miosite lúpica pseudogranulomatosa. Discute-se a raridade do achado, bem como os diagnósticos diferenciais com a PAN, dermatopolimiosite e polimiosite, tendo em vista critérios histológicos