OBJETIVO: Avaliar o estado funcional dos linfócitos T CD4+ e CD8+ de pacientes pediátricos venezuelanos infectados pelo HIV-1. MÉTODOS: As crianças foram agrupadas como progressoras rápidas (PRs) ou progressoras lentas (PLs), com base no quadro clínico. Para determinar a funcionalidade dos linfócitos T CD4+ e CD8+, foram utilizadas técnicas de citometria de fluxo e caracterizados parâmetros de funcionalidade dessas células por meio de testes ex vivo como expressão de CD95/Fas e de CD127 e frequência de apoptose. Além disso, determinamos, em células mononucleares de sangue periférico, a proliferação do HIV e a produção de interleucina-10 (IL-10), do fator de necrose tumoral alfa (TNF-α) e de interferon gama (IFN-γ), e também estimamos o IFN-γ intracelular em células T CD4+. RESULTADOS: Nossos resultados indicam que vários mecanismos moleculares e celulares dos linfócitos T CD4+ e CD8+ tiveram piora nos PRs em comparação com PLs e controles. Ambos os tipos de linfócitos T dos PRs apresentaram aumento na expressão de CD95/Fas (p < 0,01), redução na expressão de CD127 (p < 0,01) e elevação na frequência de apoptose (p < 0,01). Além disso, as células T desses pacientes apresentaram diminuição na capacidade de proliferação mitogênica (p < 0,05), redução na porcentagem de linfócitos T CD4+ produtores de IFN-γ (p < 0,05) e menor capacidade de produção de IL-10, TNF-α e IFN-γ (p < 0,01) em comparação com PLs e controles. CONCLUSÃO: Nossos resultados indicam que o declínio das respostas moleculares e celulares dos linfócitos T está relacionado a uma rápida progressão e à diminuição na resistência à infecção pelo HIV-1 em crianças.

OBJECTIVE: To evaluate simultaneously the functional state of CD4+ and CD8+ T lymphocytes from Venezuelan HIV-1-infected pediatric patients. METHODS: Children were assigned to subgroups of rapid progressors (RPs) and slow progressors (SPs), based on clinical features. To determine the degree of CD4+ and CD8+ T-lymphocyte functionality, flow cytometry techniques were used, and diverse parameters of the functionality of these cells were characterized by ex vivo tests, such as expression of CD95/Fas and CD127, and frequency of apoptosis. In addition, we determined, in cultured peripheral blood mononuclear cells, HIV-specific proliferation and the production of interleukin-10 (IL-10), tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ), besides measuring intracellular IFN-γ in CD4+ T cells. RESULTS: Our results indicate that several molecular and cellular mechanisms of CD4+ and CD8+ T lymphocytes are deteriorated in RPs in comparison with SPs and controls. Indeed, both types of T lymphocytes from RPs exhibited an increased expression of CD95/Fas (p < 0.01), a significantly reduced expression of CD127 (p < 0.01), and an augmented frequency of apoptosis (p < 0.01). Furthermore, T cells from these patients displayed a diminished capacity of mitogen proliferation (p < 0.05), a reduced percentage of IFN-γ producing CD4+ T lymphocytes (p < 0.05) and a smaller capacity of IL-10, TNF-α and IFN-γ production (p < 0.01) in comparison with SP and control patients. CONCLUSION: Our findings indicate that the decline of the normal T lymphocyte molecular and cellular responses is related to a rapid progression and a decreased resistance to HIV-1 infection in children.

La preeclampsia (PE) es una enfermedad que se presenta en la segunda mitad de embarazo. Es una condición caracterizada por hipertensión, proteinuria y frecuentemente cierto grado de restricción del crecimiento intrauterino (RCIU). El óxido nítrico (ON) regula el flujo de sangre en la placenta humana, induce vasodilatación, inhibe la agregación plaquetaria, y previene la adhesión de plaquetas a las células endoteliales. En este trabajo se evaluó, en sangre periférica de mujeres embarazadas normales (n = 46) y con PE (n = 50), los niveles séricos de nitritos y se analizó la expresión de las óxido nítrico sintasas constitutiva endotelial (eNOS) e inducible (iNOS) en tejido placentario. Se observó un aumento significativo en la concentración sérica de nitritos en las pacientes con PE al compararlas con mujeres embarazadas normales (150,64 ± 8,94 vs 40,62 ± 1,65 µM, p < 0,00001) y un aumento significativo en la expresión de las enzimas NOS (eNOS e iNOS) en las placentas de esas mismas pacientes, con respecto a las mujeres con embarazo normal (iNOS 4,29 ± 1,51 vs 0,59 ± 0,13; eNOS 1,78 ± 0,74 vs 0,46 ± 0,22, p < 0,005). Nuestros resultados muestran que existe una relación entre los valores de nitritos en suero de sangre periférica y el análisis de la expresión de las enzimas eNOS e iNOS en extracto de proteínas de tejido de placenta.

Preeclampsia (PE) is a disease that onsets in the second half of pregnancy. This condition is characterized by hypertension, proteinuria and, frequently, intrauterine growth restriction (IUGR). Nitric oxide (NO) regulates blood flow in the human placenta, it induces vasodilatation, inhibition of platelet aggregation and prevents adhesion of platelets to endothelial cells. In this work, nitrite levels were evaluated in the sera of peripheral blood of normal pregnant women (n = 46) and women with PE (n = 50); additionally, the expression of endothelial constitutive nitric oxide and inducible synthases (eNOS and iNOS, respectively) of placental tissues, were determined. An increased concentration of serum nitrites from patients with PE, in relation to normal pregnant women (150.64 ± 8. 94 vs 40, 62 ± 1, 65 µM, p < 0. 00001) was observed. An increased expression of nitric oxide synthases (eNOS and iNOS), in the placental tissues of (PE) patients, as compared to that of normal pregnant women (iNOS 4.29 ± 1.51 vs 0. 59 ± 0.13; eNOS 1.78 ± 0.74 vs 0.46 ± 0.22, p < 0.005) was also observed. Our results show that there exists a relationship between serum nitrites concentration and the expression of eNOS and iNOS, as analyzed in protein extracts of placental tissues.

En el presente trabajo, estudiamos la expresión ex vivo e in vitro de los receptores CD95/Fas y CD127 en linfocitos T CD8+ totales, vírgenes y de memoria de pacientes infectados con VIH con diferentes concentraciones sanguíneas de linfocitos T CD4+. Además, a estas células se les determinó la apoptosis espontánea e inducida por antígenos virales (Env) y su capacidad proliferativa específica. Los resultados mostraron que los pacientes con una baja densidad de linfocitos T CD4 sanguíneos (CD4 < 250/µL), manifestaban ex vivo una alta expresión de CD95/Fas y una baja expresión de CD127 en todas las subpoblaciones de linfocitos T CD8+ en relación a los pacientes con una densidad mayor de linfocitos T CD4+ en sangre (CD4 > 250/µL). En ensayos in vitro usando el antígeno Env, los linfocitos T CD8+ totales presentaron una expresión similar de ambos receptores con una mayor incidencia de apoptosis espontánea y una disminuida capacidad proliferativa en relación a los controles. Estos resultados señalan como el desarrollo progresivo de la infección por VIH en pacientes no tratados está asociado con una disminución de linfocitos T CD8+ sanguíneos, caracterizados por fallas en su capacidad proliferativa y en la frecuencia de apoptosis, lo cual se evidencia por una expresión alterada de CD127 y CD95/Fas.

We studied the ex vivo and in vitro expression of CD95/Fas and CD127 receptors in total, naïve and memory CD8+ T cells from VIH infected patients with different blood counts of CD4+ T cells. In addition, spontaneous and induced apoptosis were determined in vitro using a viral antigen (Env), along with an evaluation of their specific proliferative capacity. The obtained results demonstrated that patients with low counts of CD4+ T cells (CD4 < 250/µL), showed ex vivo, a high expression of CD95/Fas and a low expression of CD127 in all CD8+ T cell subgroups, as compared with patients with bigger counts of CD4+ T cells in blood (CD4> 250/µL). In vitro analyses using Env antigen showed that CD8+ T cells displayed a similar expression of both receptors, with a higher incidence of spontaneous and induced apoptosis and a diminished proliferative capacity as compared with controls. Results indicate how the progression of VIH infection in non-treated patients is related to a decrease of CD8+ T cells in blood, characterized by failures in their proliferative capacity and apoptosis frequency, which is demonstrated by the altered expression of CD127 and CD95/Fas expression.

La enfermedad de Chagas constituye un importante problema de salud pública en América Latina por presentar elevados índices de prevalencia y por la gravedad de sus cuadros clínicos, tales como la Miocardiopatía Chagásica Crónica (MCC), en la cual los pacientes pueden presentar diferentes grados de alteración cardíaca, siendo los pacientes chagásicos tipo 3 los que presentan mayor compromiso. Por otra parte, se conoce que diversos polimorfismos en el gen del angiotensinógeno (AGT) están correlacionados con el desarrollo de ciertas miocardiopatías hipertróficas. En este trabajo se estudió la posible correlación entre el polimorfismo -6A/G del promotor del gen del AGT con la susceptibilidad al desarrollo de una MCC en pacientes chagásicos tipo 3. Para esto, se estudió una población de 22 pacientes chagásicos tipo 3 (confirmado por seguimientos y parámetros clínicos) y 19 voluntarios seronegativos a Trypanosoma cruzi y sin ningún tipo de alteraciones cardíacas. Los genotipos derivados del polimorfismo -6A/G fueron determinados mediante la técnica de MS-PCR (Mutagenically Separated PCR), utilizando dos iniciadores alelo-específicos para el promotor del gen del AGT. Los resultados obtenidos indican que la distribución de los genotipos GG, AA y AG derivados del polimorfismo -6A/G no difieren significativamente entre pacientes y controles (1, 3, 18 y 1, 2, 16; respectivamente) con un valor de ÷2 obtenido de 0,0983 (p<0,005), lo cual sugiere que el polimorfismo -6A/G en el promotor del gen del AGT no está correlacionado con el desarrollo de MCC presente en estos pacientes.

Chagas´disease constitutes an important problem of public health in Latin America for its high prevalence indexes and for the severity of its clinical manifestations, such as the cardiomyopathy, in which the patients can present different grades of heart alteration, being the Chagas’ patients type 3 those that present greater heart compromise. On the other hand, it is known that diverse polymorphisms in the gene of the angiotensinogen (AGT), are correlated with the development of certain hypertrophic cardiomyopathies. In this work, we studied the correlation between the polymorphism -6A/G in the promoter of the gene AGT, with the susceptibility to the development a hypertrophic cardiomyopathy in Chagas patients’ type 3. For this, a sample of 22 Chagasic patients type 3 was studied (confirmed by evaluations and clinical parameters) and 19 seronegative controls to Trypanosoma cruzi without any type of heart alterations. The derivative genotypes of the polymorphism -6A/G were determined by the technique of MS-PCR (Mutagenically Separated PCR), using two allele-specific initiators (primers) for the promoter of the gene AGT. Our results indicate that the distribution of the genotypes GG, AA and AG of the polymorphism -6A/G do not differ significantly among patients and controls (1, 3, 18 and 1, 2, 16; respectively) with a value of ÷2 obtained of 0, 0983 (p < 0,005), that which suggests that the polymorphism -6A/G in the promoter of the gene AGT is not correlated with the development of cardiomyopathy in these patients.