Effect of stretching on inflammation in a subcutaneous carrageenan mouse model analyzed at single-cell resolution.

Understanding the factors that influence the biological response to inflammation is crucial, due to its involvement in physiological and pathological processes, including tissue repair/healing, cancer, infections, and autoimmune diseases. We have previously demonstrated that in vivo stretching can reduce inflammation and increase local pro-resolving lipid mediators in rats, suggesting a direct mechanical effect on inflammation resolution. Here we aimed to explore further the effects of stretching at the cellular/molecular level in a mouse subcutaneous carrageenan-inflammation model. Stretching for 10 min twice a day reduced inflammation, increased the production of pro-resolving mediator pathway intermediate 17-HDHA at 48 h postcarrageenan injection, and decreased both pro-resolving and pro-inflammatory mediators (e.g., PGE2 and PGD2 ) at 96 h. Single-cell RNA sequencing analysis of inflammatory lesions at 96 h showed that stretching increased the expression of both pro-inflammatory (Nos2) and pro-resolution (Arg1) genes in M1 and M2 macrophages at 96 h. An intercellular communication analysis predicted specific ligand-receptor interactions orchestrated by neutrophils and M2a macrophages, suggesting a continuous neutrophil presence recruiting immune cells such as activated macrophages to contain the antigen while promoting resolution and preserving tissue homeostasis.

Establishment of a Novel Porcine Model to Study the Impact of Active Stretching on a Local Carrageenan-Induced Inflammation.

OBJECTIVE: Active stretching of the body is integral to complementary mind-body therapies such as yoga, as well as physical therapy, yet the biologic mechanisms underlying its therapeutic effects remain largely unknown. A previous study showed the impact of active stretching on inflammatory processes in rats. The present study tested the feasibility of using a porcine model, with a closer resemblance to human anatomy, to study the effects of active stretching in the resolution of localized inflammation. DESIGN: A total of 12 pigs were trained to stretch before subcutaneous bilateral Carrageenan injection in the back at the L3 vertebrae, 2 cm from the midline. Animals were randomized to no-stretch or stretch, twice a day for 5 mins over 48 hrs. Animals were euthanized for tissue collection 48 hrs postinjection. RESULTS: The procedure was well tolerated by the pigs. On average, lesion area was significantly smaller by 36% in the stretch group compared with the no-stretch group (P = 0.03). CONCLUSION: This porcine model shows promise for studying the impact of active stretching on inflammation-resolution mechanisms. These results are relevant to understanding the stretching-related therapeutic mechanisms of mind-body therapies. Future studies with larger samples are warranted.

Stretching Reduces Skin Thickness and Improves Subcutaneous Tissue Mobility in a Murine Model of Systemic Sclerosis.

OBJECTIVE: Although physical therapy can help preserve mobility in patients with systemic sclerosis (SSc), stretching has not been used systematically as a treatment to prevent or reverse the disease process. We previously showed in rodent models that stretching promotes the resolution of connective tissue inflammation and reduces new collagen formation after injury. Here, we tested the hypothesis that stretching would impact scleroderma development using a mouse sclerodermatous graft-versus-host disease (sclGvHD) model. METHODS: The model consists in the adoptive transfer (allogeneic) of splenocytes from B10.D2 mice (graft) into Rag2-/- BALB/c hosts (sclGvHD), resulting in skin inflammation followed by fibrosis over 4 weeks. SclGvHD mice and controls were randomized to stretching in vivo for 10 min daily versus no stretching. RESULTS: Weekly ultrasound measurements of skin thickness and subcutaneous tissue mobility in the back (relative tissue displacement during passive trunk motion) successfully captured the different phases of the sclGvHD model. Stretching reduced skin thickness and increased subcutaneous tissue mobility compared to no stretching at week 3. Stretching also reduced the expression of CCL2 and ADAM8 in the skin at week 4, which are two genes known to be upregulated in both murine sclGvHD and the inflammatory subset of human SSc. However, there was no evidence that stretching attenuated inflammation at week 2. CONCLUSION: Daily stretching for 10 min can improve skin thickness and mobility in the absence of any other treatment in the sclGvHD murine model. These pre-clinical results suggest that a systematic investigation of stretching as a therapeutic modality is warranted in patients with SSc.

Identification of Nef-HIV-1 domains involved in p22-phox interaction and superoxide production / Identificación del dominio de Nef-VIH-1 involucrado en la interacción con p22-phox y producción de superoxido

Nef -HIV-1 has been shown to be involved in NADPH complex interaction and superoxide production. The aim of this work was to study the domains involved in the interaction between Nef and p22-phox. Two approaches were used: 1) in silico modelling, to determine the potential binding motifs and design Nef truncated forms and 2) functional assays. The results showed that GFPVT 68-72, FPDW 121-124 and REVLE 179-183 on Nef are critical for p22-phox (RPQIG 142-146 and PGGP 181-184) docking. However, only the region containing FPDW 121-124 on Nef is able to induce superoxide production. Understanding the molecular mechanisms involved in generating oxidative stress during HIV infection, is critical for therapeutic intervention, in order to minimize viral replication and dissemination.

Se ha evidenciado que Nef-VIH-1 está involucrado en la interacción con el complejo NADPH y la producción de superóxido. El objetivo de este trabajo fue identificar los dominios implicados en la interacción entre Nef y p22-phox. Se utilizaron dos estrategias: 1) análisis in silico para determinar los posibles motivos de unión y el diseño Nef formas truncadas y 2) ensayos funcionales. Los resultados mostraron que GFPVT 68-72, FPDW 121 a 124 y 179 a 183 REVLE de Nef son críticos para su unión con p22-phox (RPQIG 142-146 y 181-184 PGGP). Sin embargo, sólo la región que contiene FPDW 121-124 en Nef, es capaz de inducir la producción de superóxido. La comprensión de los mecanismos moleculares implicados en la generación de estrés oxidativo durante la infección por VIH, es crítico para la intervención terapéutica, con el fin de minimizar la replicación y la propagación viral.

Activation-dependent mitochondrial translocation of Foxp3 in human hepatocytes.

Foxp3 is considered to be the master regulator for the development and function of regulatory T cells (Treg). Recently Foxp3, has been detected in extra lymphoid tissue, and in hepatocytes and has been associated with hepatocellular carcinoma (HCC), although its role has not been defined. Since it is expected that there is a relationship between protein localization, activity and cellular function, the aim of this study was to explore the subcellular localization of Foxp3 in resting and stimulated human hepatocytes. Foxp3 expression was measured by flow cytometry, subcellular fractioning, and immunofluorescence, and this data was used to track the shuttling of Foxp3 in different subcellular compartments in hepatocytes (HepG2 cell line), stimulated by using the PKC activators (PMA), core and preS1/2 antigen from hepatitis B virus (HBV). Our data shows that besides the nuclear location, mitochondrial translocation was detected after stimulation with PMA and at to a lesser extent, with preS1/2. In addition, Foxp3 is localizes at outer mitochondrial membrane. These results suggest a non-canonical role of Foxp3 in the mitochondrial compartment in human hepatocytes, and opens a new field about their role in liver damages during HBV infection.

Stretching Impacts Inflammation Resolution in Connective Tissue.

Acute inflammation is accompanied from its outset by the release of specialized pro-resolving mediators (SPMs), including resolvins, that orchestrate the resolution of local inflammation. We showed earlier that, in rats with subcutaneous inflammation of the back induced by carrageenan, stretching for 10 min twice daily reduced inflammation and improved pain, 2 weeks after carrageenan injection. In this study, we hypothesized that stretching of connective tissue activates local pro-resolving mechanisms within the tissue in the acute phase of inflammation. In rats injected with carrageenan and randomized to stretch versus no stretch for 48 h, stretching reduced inflammatory lesion thickness and neutrophil count, and increased resolvin (RvD1) concentrations within lesions. Furthermore, subcutaneous resolvin injection mimicked the effect of stretching. In ex vivo experiments, stretching of connective tissue reduced the migration of neutrophils and increased tissue RvD1 concentration. These results demonstrate a direct mechanical impact of stretching on inflammation-regulation mechanisms within connective tissue.

Identification of Nef-HIV-1 domains involved in p22-phox interaction and superoxide production.

Nef -HIV-1 has been shown to be involved in NADPH complex interaction and superoxide production. The aim of this work was to study the domains involved in the interaction between Nef and p22-phox. Two approaches were used: 1) in silico modelling, to determine the potential binding motifs and design Nef truncated forms and 2) functional assays. The results showed that GFPVT 68-72, FPDW 121-124 and REVLE 179-183 on Nef are critical for p22-phox (RPQIG 142-146 and PGGP 181-184) docking. However, only the region containing FPDW 121-124 on Nef is able to induce superoxide production. Understanding the molecular mechanisms involved in generating oxidative stress during HIV infection, is critical for therapeutic intervention, in order to minimize viral replication and dissemination.

[Effects of occupational exposure to pesticides on semen quality of workers in an agricultural community of Merida state, Venezuela]. / Efectos de la exposición ocupacional a plaguicidas sobre la calidad del semen en trabajadores de una comunidad agrícola del estado Merida, Venezuela.

Numerous studies report adverse effects of pesticides on male reproductive health. The objectives of this study were to investigate whether there is a relationship between occupational exposure to pesticides and semen quality, and to determine whether chronic exposure to pesticides differentially affects semen quality in men of different ages. A comparative study of 64 farmers and 64 control men was performed. The farmers were interviewed to determine their occupational history and particularly, activities that may involve exposure to pesticides. Semen parameters were evaluated and a comparative analysis of semen variables between exposed and control groups, as well as between age groups: 18-29, 30-37 and 38-60 years was done. Significant alterations of some semen parameters in the exposed group were found, such as: decreases in sperm concentration, slow progressive motility and sperm membrane integrity; at the same time, increases in eosin Y positive and sperm DNA fragmentation index. The results obtained by age groups showed significant differences between exposed and control groups for the parameters of membrane integrity, eosin Y positive and sperm DNA fragmentation index, being the exposed group between 18-29 years that showed the highest altered cases of these parameters. Our results prove that occupational pesticide exposure is associated with alterations in sperm quality, creating a risk to farm workers in their reproductive capacity.

Efectos de la exposición ocupacional a plaguicidas sobre la calidad del semen en trabajadores de una comunidad agrícola del estado Mérida, Venezuela. / Effects of occupational exposure to pesticides on semen quality of workers in an agricultural community of Merida state, Venezuela.

Numerosos estudios informan de los efectos adversos de plaguicidas sobre la salud reproductiva masculina. Los objetivos de este estudio fueron investigar si existe una relación entre exposición ocupacional a plaguicidas y la calidad del semen, y determinar si la exposición crónica a plaguicidas afecta diferencialmente la calidad del semen de trabajadores de diferentes edades. Se realizó un estudio comparativo entre 64 agricultores y 64 hombres control. Los trabajadores agrícolas fueron entrevistados para determinar su historia ocupacional, particularmente las actividades que pueden involucrar exposición a plaguicidas. Se evaluaron los parámetros seminales y se hizo un análisis comparativo entre el grupo expuesto y control, así como entre los grupos de edad 18-29, 30-37 y 38-60 años. Se encontraron alteraciones significativas de algunos parámetros del semen en el grupo expuesto, tales como: disminuciones en la concentración, motilidad lenta progresiva e integridad de membrana espermática; a su vez, incrementos en eosina Y positiva e índice de fragmentación del DNA espermático. Los resultados obtenidos por grupo de edad mostraron diferencias significativas entre los grupos expuesto y control, para los parámetros de integridad de membrana, eosina Y positiva e índice de fragmentación del DNA espermático, siendo el grupo expuesto entre 18-29 años el que mostró mayores casos alterados de estos parámetros. Los resultados de este estudio comprueban que la exposición ocupacional a plaguicidas está asociada con alteraciones en la calidad espermática, creando riesgo para la capacidad reproductiva de los trabajadores del campo.

Numerous studies report adverse effects of pesticides on male reproductive health. The objectives of this study were to investigate whether there is a relationship between occupational exposure to pesticides and semen quality, and to determine whether chronic exposure to pesticides differentially affects semen quality in men of different ages. A comparative study of 64 farmers and 64 control men was performed. The farmers were interviewed to determine their occupational history and particularly, activities that may involve exposure to pesticides. Semen parameters were evaluated and a comparative analysis of semen variables between exposed and control groups, as well as between age groups: 18-29, 30-37 and 38-60 years was done. Significant alterations of some semen parameters in the exposed group were found, such as: decreases in sperm concentration, slow progressive motility and sperm membrane integrity; at the same time, increases in eosin Y positive and sperm DNA fragmentation index. The results obtained by age groups showed significant differences between exposed and control groups for the parameters of membrane integrity, eosin Y positive and sperm DNA fragmentation index, being the exposed group between 18-29 years that showed the highest altered cases of these parameters. Our results prove that occupational pesticide exposure is associated with alterations in sperm quality, creating a risk to farm workers in their reproductive capacity.

Programmed hepatocytes cell death associated with FLIP downregulation in response to extracellular preS1/2.

Chronic hepatitis B virus (HBV) infection involves liver damage resulting in continuous cell injury and death. During HBV infection, hepatocytes exhibit changes in death receptor expression and in their susceptibility to death. These changes are observed not only in infected cells but also in bystander cells. Because excess viral surface protein (HBsAg) is secreted in large amounts as soluble particles containing preS proteins, the role of soluble preS1/2 in hepatocyte (HepG2) death modulation is an important issue to be explored. An increase of cell death induced by preS1/2 was observed. Also, cell death was associated with the down-regulation of FLIP and activation of caspase 8, caspase 9, and BID. Additionally, hepatocytes exhibited a sensitization to death mediated by the Fas receptor. These results, may contribute to understanding the role of envelope proteins (preS1/2) in the pathogenesis of HBV infection.

A deficient translocation of CD3ζ, ZAP-70 and Grb2 to lipid raft, as a hallmark of defective adaptive immune response during chronic hepatitis B infection.

Hepatitis B is considered to be a worldwide public health problem. An immunosuppressor microenvironment has been proposed to contribute to viral persistence during chronic disease. Understanding the intracellular signaling cascade in T-cells from HBV-infected patients, will contribute to unravel the mechanisms that control the development of immune response during hepatitis B. We analyze lipid rafts formation and early activation signals in chronic HBV infected patients, compared to naturally immune subjects (NIS). Patients show: (1) diminished GM1 clustering, (2) A deficient lipid rafts recruitment of CD3ζ/ZAP-70/Grb2, and (3) these proteins do not merge with GM1 within the lipid rafts. Finally, immunoprecipitation assays proved that ZAP-70 does not associate to CD3ζ. These results show for the first time, defects regarding early key events in T-cell activation, in chronically infected HBV patients, which may contribute not only to understand HBV immune tolerance, but to reveal new potential therapeutic targets to control the infection.

Occupational exposure to organophosphate and carbamate pesticides affects sperm chromatin integrity and reproductive hormone levels among Venezuelan farm workers.

OBJECTIVES: Several reports suggest that chronic pesticide exposure may affect semen quality and male fertility in humans. The objective of this study was to evaluate the association between occupational exposure to organophosphate (OP) and carbamate (CB) pesticides and semen quality, as well as levels of reproductive and thyroid hormones of Venezuelan farm workers. METHODS: Thirty-five healthy men (unexposed group) and 64 male agricultural workers (exposed group) were recruited for clinical evaluation of fertility status. Fresh semen samples were evaluated for sperm quality and analyzed for DNA fragmentation index (DFI) by flow cytometry. Pesticide exposure was assessed by measuring erythrocyte acetylcholinesterase (AChE) and plasma butyrylcholinesterase (BuChE) with a Test-mate ChE field kit. Serum levels of total testosterone (Tt), follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), thyroid stimulating hormone (TSH) and free thyroxine (FT4) were analyzed using enzyme immunoassay kits. RESULTS: Evidence of pesticide exposure was found in 87.5% of farmers based on AChE and BuChE inhibition. Significant increments were observed in sperm DFI with significant decreases in some semen parameters. DFI was negatively correlated with BuChE, sperm concentration, morphology and vitality in these workers. The levels of Tt, PRL, FT4 and TSH appeared to be normal; however, there was a tendency for increased LH and FSH levels in exposed workers. CONCLUSIONS: Our results confirm the potential impact of chronic occupational exposure to OP/CB pesticides on male reproductive function, which may cause damage to sperm chromatin, decrease semen quality and produce alterations in reproductive hormones, leading to adverse reproductive health outcomes.

Muscarinic antibodies and heart rate responses to dynamic exercise and to the Valsalva maneuver in chronic chagasic patients / Anticorpos muscarínicos e resposta da frequência cardíaca ao exercício dinâmico e a manobra de Valsalva na doença de Chagas crônica

We have studied the cardiac chronotropic responses to the Valsalva maneuver and to dynamic exercise of twenty chronic chagasic patients with normal left ventricular function and no segmental wall abnormalities by two-dimensional echocardiogram. The absolute increase in heart rate of the patients (Δ = 21.5 ± 10 bpm, M±SD) during the maneuver was significantly diminished when compared to controls (Δ = 31.30 ± 70, M±SD, p = 0.03). The minimum heart rate (58.24 ± 8.90 vs. 62.80 ± 10, p = 0.68) and the absolute decrease in heart rate at the end of the maneuver (Δ = 38.30 ± 13 vs. Δ = 31.47 ± 17, p = 0.10) were not different from controls. The initial heart rate acceleration during dynamic exercise (Δ = 12 ± 7.55 vs. Δ = 19 ± 7.27, M±SD, p = 0.01) was also diminished, but the heart rate recovery during the first ten seconds was more prominent in the sero-positive patients (Median: 14, Interquartile range: (9.75-17.50 vs. 5(0-8.75, p = 0.001). The serum levels of muscarinic cardiac auto-antibodies were significantly higher in the chagasic patients (Median: 34.58, Interquartile Range: 17-46.5, Optical Density) than in controls (Median: 0, Interquartile Range: 0-22.25, p = 0.001) and correlated significantly and directly (r = 0.68, p = 0.002) with early heart rate recovery during dynamic exercise. The results of this investigation indirectly suggest that, the cardiac muscarinic auto-antibodies may have positive agonist effects on parasympathetic heart rate control of chagasic patients.

Foram estudadas as respostas cronotrópicas cardíacas à manobra de Valsalva e ao exercício dinâmico de vinte pacientes chagásicos com função ventricular esquerda normal e sem alterações da contractilidade segmentar por ecocardiografia bidimensional. O aumento absoluto da frequência cardíaca dos pacientes (Δ = 21,5 ± 10 bpm, M ± DP) durante a manobra de Valsalva foi significativamente menor quando se comparava ao grupo controle (Δ = 31,30 ± 70, p = 0,03). A frequência cardíaca mínima (58,24 ± 8,90 vs 62,80 ± 10, p = 0,68) e a diminuição da frequência cardíaca absoluta no final da manobra (Δ = 38,30 ± 13 vs Δ = 31,47 ± 17, p = 0,10) não foram diferentes em comparação com o grupo controle. A aceleração inicial da frequência cardíaca durante o exercício dinâmico (Δ = 12 ± 7,55 vs Δ = 19 ± 7,27, p = 0,01) também foi menor, mas a recuperação da frequência cardíaca, durante os primeiros dez segundos, foi maior no grupo sero-positivos [mediana:14 (intervalo interquartil: 9,75-17,50) vs 5 (0 - 8,75), p = 0,001]. Os níveis séricos de auto-anticorpos muscarínicos cardíacos foram significativamente maiores nos pacientes chagásicos do que no grupo controle [(mediana: 34,58 densidade óptica (intervalo interquartil 17 - 46,5) vs (mediana: 0, intervalo interquartil 0 - 22,25) p = 0,001] e a correlação é significativa e direta (r = 0,68, p = 0,002) com o início da recuperação da frequência cardíaca durante o exercício dinâmico. Os resultados desta investigação sugerem que indiretamente, os auto-anticorpos muscarínicos cardíacos, podem ter ação agonista positiva sobre o controle parassimpático da frequência cardíaca dos pacientes chagásicos.

Muscarinic antibodies and heart rate responses to dynamic exercise and to the Valsalva maneuver in chronic chagasic patients.

We have studied the cardiac chronotropic responses to the Valsalva maneuver and to dynamic exercise of twenty chronic chagasic patients with normal left ventricular function and no segmental wall abnormalities by two-dimensional echocardiogram. The absolute increase in heart rate of the patients (Δ = 21.5 ± 10 bpm, M±SD) during the maneuver was significantly diminished when compared to controls (Δ = 31.30 ± 70, M±SD, p = 0.03). The minimum heart rate (58.24 ± 8.90 vs. 62.80 ± 10, p = 0.68) and the absolute decrease in heart rate at the end of the maneuver (Δ = 38.30 ± 13 vs. Δ = 31.47 ± 17, p = 0.10) were not different from controls. The initial heart rate acceleration during dynamic exercise (Δ = 12 ± 7.55 vs. Δ = 19 ± 7.27, M±SD, p = 0.01) was also diminished, but the heart rate recovery during the first ten seconds was more prominent in the sero-positive patients (Median: 14, Interquartile range: (9.75-17.50 vs. 5(0-8.75, p = 0.001). The serum levels of muscarinic cardiac auto-antibodies were significantly higher in the chagasic patients (Median: 34.58, Interquartile Range: 17-46.5, Optical Density) than in controls (Median: 0, Interquartile Range: 0-22.25, p = 0.001) and correlated significantly and directly (r = 0.68, p = 0.002) with early heart rate recovery during dynamic exercise. The results of this investigation indirectly suggest that, the cardiac muscarinic auto-antibodies may have positive agonist effects on parasympathetic heart rate control of chagasic patients.

Cardiac autonomic control mechanisms in the pathogenesis of chagas' heart disease.

Primary abnormalities of the autonomic nervous system had been postulated as the pathogenic mechanisms of myocardial damage, in patients with Chagas disease. However, recent investigations indicate that these abnormalities are secondary and amenable to treatment with beta-adrenergic blockers. Moreover, muscarinic cardiac autoantibodies appear to enhance parasympathetic activity on the sinus node. Therefore, the purpose of this paper is to analyze how knowledge on Chagas' disease evolved from being initially considered as a primary cardioneuromyopathy to the current status of a congestive cardiomyopathy of parasitic origin.

Diabetes mellitus tipo 1 y factores ambientales: La gran emboscada

La Diabetes Mellitus Tipo 1 (DM1) es una de las patologías más estudiadas en la actualidad, no solo por el aumento de su incidencia, sino también por su aparición a edades cada vez más tempranas. La DM1 es una enfermedad autoinmune de una alta complejidad genética y donde la susceptibilidad a factores ambientales parece jugar un papel preponderante. Elementos como parto por cesárea, deficiencia de vitamina D, exposición temprana a proteínas de la leche de vaca, exposición limitada a microorganismos durante la infancia y el incremento en la incidencia de obesidad infantil han sido relacionados con el desarrollo de esta entidad, convergiendo todos estos factores en un punto clave: la pérdida de la tolerancia inmunológica intestinal y la participación de células T auto-reactivas en pacientes susceptibles. Por su parte, la leche materna ofrece una serie de factores de crecimiento, inmunológicos, e incluso insulina, que son capaces de inducir una respuesta tolerogénica en el microambiente intestinal con la subsecuente disminución de la autoinmunidad. En esta revisión se expondrá la evidencia y los mecanismos fisiopatológicos propuestos por medio de los cuales los elementos mencionados desencadenarían una alteración de la inmunomodulación intestinal y un incremento en la predisposición al desarrollo de DM1.

Type 1 Diabetes Mellitus (T1DM) is one of the most studied pathologies to date, not only due to the elevating incidence but also because it is being diagnosed at even earlier ages. T1DM is an autoimmune disease with a very complex genetic background where environmental factors seem to play a very important triggering role. Elements like cesarean section, vitamin D deficiency, early exposure to cow milk proteins, limited exposure to microorganisms during infancy, and the increase of childhood obesity have been related to the development of this disease, converging all the factors into one key feature: loss of intestinal immunological tolerance and the participation of auto-reactive T cells from a susceptible patient. On the other hand, breast milk offers a series of growth and immunological factors, even insulin, which are able to induce a tolerogenic response in the intestinal microenvironment, lowering the probability of any autoimmune phenomena. The following review will expose evidence of the pathophysiological mechanisms involved in each environmental element associated with intestinal immunomodulation and the increase risk of T1DM.

Fas and FasL expression in leukocytes from chronic granulomatous disease patients.

Abstract. Chronic Granulomatous Disease (CGD) is a primary immunodeficiency characterized by defects in superoxide (O2-) production, which result from mutations in one of the four NADPH oxidase components, predisposing to bacterial and fungal infections. Besides the O2-defect, it has been described that neutrophils from CGD patients are resistant to cell death, a phenomenon that has been connected to chronic inflammation and predisposition to autoimmune diseases. A diminished expression of Fas and its counterpart FasL, molecules known to play a major role in cell death, has been described in lymphocytes depleted of O2-reactive oxygen species (ROS), suggesting an involvement of ROS in Fas/FasL expression. In this work, Fas and FasL expressions were analyzed in T cells and neutrophils from two CGD families, previously known to harbor two different molecular defects: absence of either p47-phox or p67-phox. We found that T lymphocytes from CGD patients express low levels of Fas and FasL, while a diminished FasL expression was observed on neutrophils from a CGD A470 patient. These defects may contribute to understand altered cell death in CGD patients.

[Analysis of p22-phox and p47-phox subcellular localization and distribution in neutrophils from human immunodeficiency virus (HIV) infected patients]. / Localización y distribución celular de p22-phox y p47-phox en neutrófilos humanos de pacientes infectados con VIH.

INTRODUCTION: During human immunodeficiency virus (HIV) infection a dysfunction of polymorphonuclear (PMN) cells has been described including a progressively altered superoxide production as disease progression. The NADPH oxidase has been described as a major source of superoxide. The neutrophil NADPH oxidase comprises a plasma membrane-bound cytochrome b558 (which is a heterodimer of one p22-phox and one gp91-phox subunit) and cytosolic subunits, namely p47-phox, p67-phox and p40-phox. During neutrophil activation in response to various agonists, the cytosolic subunits translocate to and associate with the cytochrome b558, a process that results in oxidase activation. Therefore, an altered superoxide production could be a consequence of abnormal distribution or translocation of NADPH oxidase components in response to HIV infection. MATERIAL AND METHODS: We used several strategies including: confocal microscopy, subcellular fractionation and sucrose gradients, to analyze the cellular distribution of two of the NADPH oxidase components (p22-phox and p47-phox). RESULTS: We observed that in resting cells, a substantial proportion of p22-phox from HIV positive patients is distributed in regions close to the cytoplasmic membrane, sediment in high density sucrose fractions and is located in the cytoplasmic insoluble fraction. Additionally, a diffuse cytosolic distribution of p47-phox was observed in neutrophils from HIV infected patients. The results demonstrate an inappropriate cell distribution of NADPH-complex in PMN from HIV positive patients.