[Evaluation of the quality of the human spermatozoon: comparison between spermatic DNA integrity and semen variables]. / Evaluación de la calidad del espermatozoide humano: comparación entre la integridad del ADN espermático y variables del semen.

Semen analysis does not have an absolute predictive value on fertility, however it is a reflection of male fertility potential, which is related to its spermatozoa quality and other semen variables. Great variability in human semen parameters has been demonstrated within a single individual, an observation that could explain why a male with low semen quality can successfully fertilize an egg. Although conventional semen analysis, such as sperm concentration, motility and morphology, provide important information about the clinical status of male fertility, new procedures to predict the sperm functional capability have been developed in the last decade, such as analysis of nuclear DNA integrity, which have improved considerably the clinical diagnosis of male infertility, and increased the knowledge about spermatozoa function. DNA fragmentation consist in interruptions, both in single and double DNA strains, that frequently occur in sperm samples from infertile patients. We have conducted a clinical study in semen samples from patients who have attended the Andrology laboratory of the University of Los Andes, between March 2007 and March 2009. The aim of this study was to compare sperm DNA integrity, analyzed by flow cytometry, with traditional semen parameters. Our results show remarkable correlations between conventional human semen variables and sperm chromatin integrity, contributing to asses an integral evaluation of sperm quality allowing the analysis of its fertilizing potential in clinical studies.

Evaluación de la calidad del espermatozoide humano: comparación entre la integridad del ADN espermático y variables del semen / Evaluation of the quality of the human spermatozoon: comparison between spermatic DNA integrity and semen variables

El análisis del semen no tiene valor predictivo absoluto de fertilidad, pero informa sobre el potencial de fertilidad del varón, el cual está relacionado con la calidad de sus espermatozoides y de otras variables del semen. Se ha comprobado que los valores del semen pueden mostrar gran variabilidad en un mismo individuo. Esto explica por que un hombre cuyas variables no son absolutamente normales, puede lograr un embarazo en su pareja. Dentro de los parámetros tradicionalmente utilizados en la evaluación clínica de la fertilidad masculina se encuentran: la concentración, la movilidad y la morfología espermática; además de medir estas variables, nuevos procedimientos han sido incorporados para evaluar la capacidad funcional de los espermatozoides, uno de los que ha alcanzado particular importancia en la última década es la medida de la integridad del ADN nuclear. La fragmentación del ADN consiste en interrupciones en las cadenas simples o dobles del ADN que ocurre frecuentemente en la muestra de pacientes no fértiles. Se ha llevado a cabo un estudio clínico, en muestras de semen provenientes de pacientes que acudieron al laboratorio de Andrología de la Universidad de los Andes, colectadas entre marzo del 2007 y marzo del 2009, a fin de establecer comparaciones entre los parámetros convencionales y la medición de la integridad de la cromatina espermática, mediante citometría de flujo. Los resultados obtenidos mostraron correlaciones evidentes entre los parámetros convencionales y la integridad del ADN espermático y aportan datos de gran utilidad en el estudio clínico integral de la infertilidad masculina.

Semen analysis does not have an absolute predictive value on fertility, however it is a reflection of male fertility potential, which is related to its spermatozoa quality and other semen variables. Great variability in human semen parameters has been demonstrated within a single individual, an observation that could explain why a male with low semen quality can successfully fertilize an egg. Although conventional semen analysis, such as sperm concentration, motility and morphology, provide important information about the clinical status of male fertility, new procedures to predict the sperm functional capability have been developed in the last decade, such as analysis of nuclear DNA integrity, which have improved considerably the clinical diagnosis of male infertility, and increased the knowledge about spermatozoa function. DNA fragmentation consist in interruptions, both in single and double DNA strains, that frequently occur in sperm samples from infertile patients. We have conducted a clinical study in semen samples from patients who have attended the Andrology laboratory of the University of Los Andes, between March 2007 and March 2009. The aim of this study was to compare sperm DNA integrity, analyzed by flow cytometry, with traditional semen parameters. Our results show remarkable correlations between conventional human semen variables and sperm chromatin integrity, contributing to asses an integral evaluation of sperm quality allowing the analysis of its fertilizing potential in clinical studies.

Altered T cell costimulation during chronic hepatitis B infection.

T-cell response to hepatitis B virus (HBV) is vigorous, polyclonal and multi-specific in patients with acute hepatitis who ultimately clear the virus, whereas it is narrow and inefficient in patients with chronic disease, where inappropriate early activation events could account for viral persistence. We investigated the induction of activation receptors and cytokine production in response to HBcAg and crosslinking of CD28 molecules, in CD4+ cells from a group of chronically infected patients (CIP) and naturally immune subjects (NIS). We demonstrated that CD4+ cells from CIP did not increase levels of CD40L and CD69 following stimulation with HBcAg alone or associated to CD28 crosslinking, in contrast to subjects that resolved the infection (p<0.01). Furthermore, CD4+ cells from CIP produced elevated levels of IL-10 in response to HBcAg. These results suggest that a predominant inhibitory environment may be responsible for altered T cell costimulation, representing a pathogenic mechanism for viral persistence.

[Impaired activation and costimulation of T CD4+ lymphocytes during chronic hepatitis C infection]. / Déficit en señales de activación y coestimulación en linfocitos T CD4+ de pacientes infectados con el virus de la hepatitis C.

Hepatitis C chronic infection occurs in 80% of the cases and eventually leads to cirrhosis and hepatocellular carcinoma. A deficient adaptive immune response has been described during chronic infection which contributes to viral persistence. This altered T cell response could be associated to deficient costimulation signals during priming of T cells. We have conducted an in vitro study to explore the activation phenomenon of CD4+ T cells focusing on costimulation via the CD28 receptor, associated to stimulation with purified Hepatitis C (HCV) core antigen. Our study involved the induction of CD69, CD25 and CD40L activation receptors, along with detection of intracellular cytokines such as IFN-gamma, TGF-beta and IL-10. Analysis was performed in chronically HCV infected patients, intrafamilial members of HCV-infected patients and healthy individuals. HCV core antigen induced CD40L expression in CD4+ cells from intrafamilial members, in contrast to chronically infected patients and control individuals. Association of CD28 crosslinking increased CD69 and IFN-gamma expression in chronically infected patients, suggesting a detriment in this signaling pathway. Additionally, an increased TGF-beta expression was observed in CD4+ cells from HCV-infected patients, which was corrected by addition of CD28 crosslinking. Our results may contribute to understand the underlying mechanism of T cell tolerance against HCV during chronic infection, and to provide new targets for the designing of therapeutic strategies to control the infection and to offer protective immunity against the virus.

Déficit en señales de activación y coestimulación en linfocitos T CD4+ de pacientes infectados con el virus de la hepatitis C / Impaired activation and costimulation of T CD4+ lymphocytes during chronic hepatitis C infection

La infección crónica por el virus de la Hepatitis C ocurre en 80% de los casos y conduce eventualmente a cirrosis y carcinoma hepatocelular. Se ha descrito que una respuesta inmunitaria adaptativa deficiente pudiera contribuir a la persistencia viral. Dicha alteración pudiera ser atribuida a defectos en señales de co-estimulación durante la activación de los linfocitos CD4+. En este estudio exploramos la respuesta de esta población linfoide en presencia del antígeno del core del VHC asociado con la co-estimulación vía CD28, midiendo la expresión de moléculas de superficie: CD69, CD25, CD40L y de citocinas intracitoplasmáticas tales como: IFN-g, TGF-b e IL-10. El análisis fue realizado en individuos infectados crónicos, familiares contactos y controles sanos. El antígeno del core del VHC indujo la expresión de CD40L en los linfocitos CD4+ de familiares contactos, a diferencia de lo observado en paciente infectados crónicos e individuos controles sanos. Al asociar la estimulación vía CD28, los pacientes infectados con VHC incrementaron la expresión de CD69 e IFN-g, sugiriendo un déficit en la señalización a través de esta vía de coestimulación. Adicionalmente, se observó un incremento en la producción de TGF-b en los pacientes infectados en respuesta al antígeno, el cual fue corregido al coestimular vía CD28. Nuestros resultados pueden ayudar a comprender los mecanismos de tolerancia generados durante la infección crónica por el VHC y ofrecer una aproximación que pudiera contribuir al diseño de estrategias terapéuticas para el control de la infección crónica y al desarrollo de inmunidad protectora contra el virus.

CD40/CD40L expression in leukocytes from chronic granulomatous disease patients.

Chronic granulomatous disease (CGD) is an inherited disorder caused by defects in the NADPH oxidase complex, which generates superoxide, the precursor of hydrogen peroxide (H(2)O(2)) and other reactive oxygen derivatives with microbicidal activity. Because CGD patients are at risk of chronic inflammatory manifestations, including inflammatory bowel disease and autoimmune diseases, and it is not clear whether these pathologies are exclusively secondary to altered superoxide production, or whether distinct immunologic defects are involved, we explored cell proliferation, lymphocyte cell counts, immunoglobulin levels, presence of autoimmune antibodies and expression of costimulatory molecules in leukocytes from CGD patients. We found that CGD patients have a diminished phytohemagglutinin-induced proliferation of blood mononuclear cells. Following stimulation with PMA plus ionomycin, a reduced percentage of CD40L expression in T lymphocytes and a diminished expression of CD40 molecules in neutrophils were observed on leukocytes from these patients. Our results suggest an altered interplay between elements of innate and adaptive immunity in CGD patients, which may be reflected in an increased susceptibility to opportunistic infections.

Antigen-induced regulatory T cells in HBV chronically infected patients.

T cell response against HBV is vigorous in patients with acute hepatitis who clear the virus, whereas it is weak and narrowly focused in patients with chronic disease. We report that following incubation with HBcAg, a population of CD4+FoxP3+ cells expressing phenotypic markers of both natural and induced Tregs, can be antigen-induced from peripheral mononuclear cells. Conversely, naive and naturally immune subjects did not increase CD4+FoxP3+ Tregs following stimulation with HBcAg, supporting the idea that natural Tregs are able to respond specifically to HBV antigen. Furthermore, increased frequencies of antigen-induced CD4+FoxP3+IL-10+ Tregs correlated with viral load, suggesting that antigen-induced Tregs could contribute to an inadequate response against the virus, leading to chronic infection and support the view that specific natural Tregs may be implicated in host immune tolerance during HBV infection.

[Pattern of T cell activation in absence of protective immunity against hepatitis B virus. Review]. / Patrón de activación de linfocitos T en ausencia de respuesta protectora contra el virus dela hepatitis B. Revisión.

Hepatitis B is an important cause of morbidity and mortality around the world. One-third of the world's population has been estimated to be infected with hepatitis B virus (HBV). A significant amount of evidence suggests that both humoral and cellular immune responses are important to eliminate the virus and that, cellular immunity is involved in the pathogenesis of the disease. Vaccination with HBsAg is considered as the main strategy for effective control of the infection and viral transmission. However, approximately 5-10% of immunized individuals fail to elicit detectable specific antibodies and remain at risk for hepatitis B infection. In this work we have reviewed the current status in the pathogenesis of the disease and the mechanisms described to explain nonresponsiveness to the vaccine as well. Since nonresponders to the vaccine are at risk for the infection, a common mechanism to explain the absence or inappropriate immune response to virus components is proposed. Within the suggested model an impaired activation of T lymphocytes against viral antigens, both in nonresponders to vaccination and chronically infected patients, is described. These observations could be consistent with potential differences in the MHC/Ag presentation; therefore contributing to our understanding of the altered T helper response as an underlying mechanism for the lack of protective immunity against VHB.

Patrón de activación de linfocitos T en ausencia de respuesta protectora contra el virus de la hepatitis B: revisión / Pattern of T cell activation in absence of protective immunity against hepatitis B virus: review

La hepatitis B es una causa importante de morbilidad y mortalidad en el mundo. Un cúmulo importante de evidencias sugiere que tanto la respuesta humoral como la celular son importantes para la eliminación del virus y que la respuesta celular se ha involucrado en la patogénesis de la enfermedad. La vacunación con el antígeno de superficie (HBsAg) es considerada como la estrategia principal para el control de la infección. Sin embargo, aproximadamente 5 a 10 por ciento de los individuos fallan en producir anticuerpos específicos. En este trabajo se han revisado aspectos fundamentales en la inmunopatogenia de la Hepatitis B, así como también fenómenos que explican la ausencia de respuesta a la vacuna. Partiendo de la premisa que los individuos no respondedores a la vacuna contra la Hepatitis B, están en riesgo de infección, se propone un mecanismo común para explicar la ausencia de respuesta frente al virus. En el contexto del modelo planteado se describe una alteración en la activación de los linfocitos T, tanto en no respondedores como en infectados crónicos. Estas observaciones son consistentes con diferencias potenciales en el eje de presentación MHC/Ag, contribuyendo al entendimiento sobre la alteración de la respuesta T cooperadora como un mecanismo de base para la ausencia de inmunidad protectora contra el virus de la hepatitis B (VHB)

[Role of human immunodeficiency virus in leukocytes apoptosis from infected patients]. / Papel del virus de la inmunodeficiencia humana en 1a apoptosis de leucocitos de pacientes infectados.

The hallmark of the immunodeficiency virus infection is a progressive detriment of the immune response which has been associated to a gradual loss of its responsible components, in particularly, CD4 positive T cells. Although this cell population is considered the main target of the virus, there is a recent deal of interest in studying other components that may not be targets of the virus, but are important elements to control infectious microorganisms and that have been demonstrated to be altered during HIV infection. Neutrophils (PMN) are innate immune components that play a fundamental role against HIV infection and these cells have been described as functionally altered during AIDS. It has been suggested that such a dysfunction could be attributed to an increased susceptibility of these cells to accelerated spontaneous apoptosis. However, the underlying mechanisms that induce programmed cell death of neutrophils remain unknown. In previous works we have explored some events involved during cell death of neutrophils from HIV infected patients. It is the purpose of this work to review the current knowledge of apoptosis signals in neutrophils and to discuss our own data about some mechanisms involved in spontaneous and Fas mediated apoptosis, which may contribute to understand neutrophils dysfunction during HIV infection.

Papel del virus de la inmunodeficiencia humana en la apoptosis de leucocitos de pacientes infectados / Role of human immunodeficiency virus in leukocytes apoptosis from infected patients

La infección por el virus de la inmunodeficiencia humana (VIH) se caracteriza por el deterioro progresivo de la respuesta inmune, asociado a una pérdida gradual de los elementos que la generan, especialmente de los linfocitos T CD4+. Aunque estas células son el principal blanco del virus, en la actualidad existe gran interés por el estudio de otros elementos celulares, que se encuentran alterados y que contribuyen al deterioro del paciente. Dentro de este grupo de células se encuentran los polimorfonucleares (PMN), células fundamentales en la defensa innata contra el VIH y cuya función está alterada en el curso de la enfermedad. Probablemente la causa de este deterioro se debe, en parte, a un incremento en su susceptibilidad para sufrir apoptosis espontánea; sin embargo, no han sido dilucidados los mecanismos involucrados en generar muerte celular programada en los neutrófilos de pacientes VIH positivos. En trabajos previos, hemos explorado algunos eventos asociados con la muerte de los neutrófilos de pacientes infectados. Este artículo tiene como objetivo principal profundizar en los eventos involucrados en la apoptosis espontánea e inducida vía Fas, que pudieran estar implicados en el deterioro de los PMN durante la infección por el VIH

The nonresponse to hepatitis B vaccination is associated with impaired lymphocyte activation.

Nonresponsiveness against hepatitis B vaccination has been described in 4-10% of immunized subjects. We have explored the specific cell response to hepatitis B surface antigen by analyzing: PBMC proliferation, cytokine production (Th1, Th2 profiles, and TGF-beta), and activation molecules on Th cells. A poor proliferative response was demonstrated in nonresponders (P < 0.05). T cells from responders produced all tested cytokines (P < 0.01), in contrast with nonresponders subjects (P < 0.05). Expression of CD69 and CD25 was diminished in T cells from nonresponders (P < 0.01). A reduced expression of CD40L was also detected in T cells from nonresponders (P < 0.01). An elevated correlation coefficient was observed between CD40L on CD4+ cells and antibody production. These results suggest an overall inability of T cells to be activated which could be consistent with potential differences in antigen presentation. In conclusion, our results suggest that an altered Th response may be a consequence of inappropriate early activation events.

Risk factors for complications after performance of ERCP.

BACKGROUND: ERCP has become widely available for the diagnosis and treatment of benign and malignant pancreaticobiliary diseases. In this prospective study, the overall complication rate and risk factors for diagnostic and therapeutic ERCP were identified. METHODS: Data were collected prospectively on patient characteristics and endoscopic techniques from 1223 ERCPs performed at a single referral center and entered into a database. Univariate and multivariate analyses were used to identify risk factors for ERCP-associated complications. RESULTS: Of 1223 ERCPs performed, 554 (45.3%) were diagnostic and 667 (54.7%) therapeutic. The overall complication rate was 11.2%. Post-ERCP pancreatitis was the most common (7.2%) and in 93% of cases was self-limiting, requiring only conservative treatment. Bleeding occurred in 10 patients (0.8%) and was related to a therapeutic procedure in all cases. Nine patients had cholangitis develop, most cases being secondary to incomplete drainage. There was one perforation (0.08%). All other complications totaled 1.5%. Variables derived from cannulation technique associated with an increased risk for post-ERCP pancreatitis were precut access papillotomy (20%), multiple cannulation attempts (14.9%), sphincterotome use to achieve cannulation (13.1%), pancreatic duct manipulation (13%), multiple pancreatic injections (12.3%), guidewire use to achieve cannulation (10.2%), and the extent of pancreatic duct opacification (10%). Patient characteristics associated with an increased risk of pancreatitis were sphincter of Oddi dysfunction (21.7%) documented by manometry, previous ERCP-related pancreatitis (19%), and recurrent pancreatitis (16.2%). Pain during the procedure was an important indicator of an increased risk of post-ERCP pancreatitis (27%). Independent risk factors for post-ERCP pancreatitis were identified as a history of recurrent pancreatitis, previous ERCP-related pancreatitis, multiple cannulation attempts, pancreatic brush cytology, and pain during the procedure. CONCLUSIONS: The most frequent ERCP-related complication was pancreatitis, which was mild in the majority of patients. The frequency of post-ERCP pancreatitis was similar for both diagnostic and therapeutic procedures. Bleeding was rare and mostly associated with sphincterotomy. Other complications such as cholangitis and perforation were rare. Specific patient- and technique-related characteristics that can increase the risk of post-ERCP complications were identified.

Nongranulomatous anterior uveitis associated with alendronate therapy.

We describe a case of acute nongranulomatous anterior uveitis associated with alendronate therapy, in an adult woman, without medical history of previous diseases, except for intercurrent problems of osteoporosis. The symptoms disappeared abruptly after anti-inflammatory therapy and discontinuation of alendronate. Side effects associated with ocular inflammation have been recently documented in 3 patients under alendronate therapy. Clinical and laboratory diagnosis of ocular inflammation syndromes are also reviewed.

Nongranulomatous anterior uveitis associated with alendronate therapy

We describe a case of acute nongranulomatous anterior uveitis associated with alendronate therapy, in an adult woman, without medical history of previous diseases, except for intercurrent problems of osteoporosis. The symptoms disappeared abruptly after anti-inflammatory therapy and discontinuation of alendronate. Side effects associated with ocular inflammation have been recently documented in 3 patients under alendronate therapy. Clinical and laboratory diagnosis of ocular inflammation syndromes are also reviewed.

Anticuerpos anti-VHC en insuficientes renales crónicos / Anti-HCV antibodies in patients with chronic renal failure

El virus de la hepatitis C es la principal causa de hepatitis post-transfusional en el mundo, pero poco se conoce acerca de la transmisión en los pacientes con insuficiencia renal crónica y en las unidades de hemodialisis. El presente trabajo estimó la prevalencia de los anticuerpos anti-VHC en los pacientes con insuficiencia renal crónica del Departamento de Nefrología del Hospital Universitario de los Andes, en Mérida Venezuela. Se admitieron 22 pacientes al estudio, todos con insuficiencia renal crónica. Los datos médicos referentes a factores de riesgo (trasnfuciones, drogadicción y promiscuidad sexual) fueron obtenidos de las historias clínicas y las encuestas aplicadas a los pacientes. Un ELISA anti-VHC de segunda generación fue usado como prueba serológica. De los 22 pacientes estudiados, 5 fueron anti-VHC positivos y 17 fueron negativos. Los 5 pacientes seropositivos eran hemodializados, lo cual arrojó una prevalencia del 22,7 por ciento en el grupo de estudio. Todos los pacientes insuficientes renales crónicos no hemodializados fueron anti-VHC (p= 0.0047). No hubo asociación significativa entre transfuciones y positividad al anti-VHC (p= 0.36). La hemodiálisis podría ser un factor de riesgo en la trasmisión del virus de la hepatitis C

Presentación de un caso de síndrome de sweet tratado con dapsona / Presentation of a case sweet sydrome tratment with dapsone

El Síndrome de Sweet se carateriza por fiebre, placas eritematosas, edematosas y dolorosas, de distribución asimétrica, leucocitosis en sangre periférica e infiltración de la dermis por neutrófilos. Puede ser de origen idiopático o estar asociado a enfermedades mieloproliferativas y a tumores sólidos. Responde rápidemente a la administración de corticosteroides pero las enfermedades cardiovasculares y metabólicas limitan su uso y frecuentemente se observan recaídas. Presentamos y caso típico de Síndrome de Sweet que presentó artritis, episcleritis y proteinuria que evolucionó satisfactoriamente con el uso de Dapsona, sin recidiva luego de dos años de seguimiento