Selective Transfection of a Transferrin Receptor-Expressing Cell Line with DNA-Lipid Nanoparticles.

Despite considerable progress in using lipid nanoparticle (LNP) vehicles for gene delivery, achieving selective transfection of specific cell types remains a significant challenge, hindering the advancement of new gene or gene-editing therapies. Although LNPs have been equipped with ligands aimed at targeting specific cellular receptors, achieving complete selectivity continues to be elusive. The exact reasons for this limited selectivity are not fully understood, as cell targeting involves a complex interplay of various cellular factors. Assessing how much ligand/receptor binding contributes to selectivity is challenging due to these additional influencing factors. Nonetheless, such data are important for developing new nanocarriers and setting realistic expectations for selectivity. Here, we have quantified the selective, targeted transfection using two uniquely engineered cell lines that eliminate unpredictable and interfering cellular influences. We have compared the targeted transfection of Chinese ovary hamster (CHO) cells engineered to express the human transferrin receptor 1 (hTfR1), CHO-TRVb-hTfR1, with CHO cells that completely lack any transferrin receptor, CHO-TRVb-neo cells (negative control). Thus, the two cell lines differ only in the presence/absence of hTfR1. The transfection was performed with pDNA-encapsulating LNPs equipped with the DT7 peptide ligand that specifically binds to hTfR1 and enables targeted transfection. The LNP's pDNA encoded for the monomeric GreenLantern (mGL) reporter protein, whose fluorescence was used to quantify transfection. We report a novel LNP composition designed to achieve an optimal particle size and ζ-potential, efficient pDNA encapsulation, hTfR1-targeting capability, and sufficient polyethylene glycol sheltering to minimize random cell targeting. The transfection efficiency was quantified in both cell lines separately through flow cytometry based on the expression of the fluorescent gene product. Our results demonstrated an LNP dose-dependent mGL expression, with a 5-fold preference for the CHO-TRVb-hTfR1 when compared to CHO-TRVb-neo. In another experiment, when both cell lines were mixed at a 1:1 ratio, the DT7-decorated LNP achieved a 3-fold higher transfection of the CHO-TRVb-hTfR1 over the CHO-TRVb-neo cells. Based on the low-level transfection of the CHO-TRVb-neo cells in both experiments, our results suggest that 17-25% of the transfection occurred in a nonspecific manner. The observed transfection selectivity for the CHO-TRVb-hTfR1 cells was based entirely on the hTfR1/DT7 interaction. This work showed that the platform of two engineered cell lines which differ only in the hTfR1 can greatly facilitate the development of LNPs with hTfR1-targeting ligands.

Rheumatoid arthritis associated vasculitis: a rare entity; case and review.

We present a case of a 60-year-old male with known seropositive rheumatoid arthritis and cerebral vasculitis who presented to the emergency room with abrupt onset lower back and abdominal pain. The patient developed peritonitis which led to an abdominal laparotomy where jejunal ischemia, necrosis, and perforation were found, requiring bowel resection. On pathology examination, the patient had mesenteric vessel intramural inflammation indicative of vasculitis. He developed an anastomotic leak on postoperative Day 4 and elected hospice care. A high index of suspicion for mesenteric vasculitis should be considered in patients presenting with abdominal pain in the setting of known rheumatoid arthritis associated vasculitis, especially patients with long-standing rheumatoid arthritis. The high mortality represented by gastrointestinal involvement in rheumatoid arthritis associated vasculitis warrants investigation in high-risk patients, despite its low prevalence. Treatment may consist of high-dose corticosteroids, immunosuppressive agents, biologic therapies that target the underlying autoimmune process, and in severe cases, bowel resection.

Epidemiology, Management, and Survival Outcomes of Germ Cell Cancer in Southern Portugal: A Population-Based Study (2008-2012).

INTRODUCTION: Building on previous suboptimal survival results, we aimed to perform a study of the epidemiological status, management, and outcomes of germ cell tumors (GCT) in the Portuguese population. MATERIALS AND METHODS: Retrospective populational study of GCT cases diagnosed between 2008 and 2012 in southern Portugal. Joinpoint regression was used to compute average annual percentage change (AAPC) in incidence rate. ESMO/EAU guidelines served as references to evaluate compliance. Association between compliance with guidelines and hospital GCT case load was performed by generalized estimating equation. Survival was calculated by Kaplan-Meier and prognostic factors by Cox models. RESULTS: The study included 401 GCT male cases. The AAPC was 5.4% (IC 95% 3.3-7.4, P < .001) from 1999 (an earlier cohort published) to 2012. The median time to diagnosis was 63 days (Q25 = 33 days; Q75 = 114 days; IQR = 81 days). For stage II/III the median time to start chemotherapy was 34 days (Q25 = 22 days; Q75 = 56 days; IQR = 22 days). In 86% cases there was noncompliance with guidelines for the orchiectomy report, 6% for staging, 38% for tumor markers evaluation, 20% for treatment and 25% for chemotherapy dose intensity. The 5-year overall survival was 93.8% (95% CI, 91.3%-96.4%). Hospitals that managed ≤ 3 GCT cases/ year had higher odds for noncompliance with guidelines of blood markers, treatment and dose intensity. None of GCT healthcare access and management factors studied were associated with prognosis. CONCLUSIONS: The burden of GCT is rising in Portugal. Although survival has improved, efforts must be made to nationally enhance training and expertise in GCT and support region adapted models of centralization of care.

Eficacia y seguridad en la vida real de amlodipino/irbesartán en combinación fija en el tratamiento de la hipertensión en América Latina: estudio PARCERIA / Real-life effectiveness and safety of fixed amlodipine/irbesartan in hypertension management in Latin America: the PARCERIA study

Resumen Introducción: Los estudios basados en la población latinoamericana han demostrado tasas de control de la presión arterial subóptimas. La combinación de medicamentos antihipertensivos en dosis fijas se asocia con mayor comodidad, adherencia y efectividad en comparación con la monoterapia. Objetivo: Evaluar la efectividad y seguridad en la vida real de la combinación fija de amlodipino/irbesartán en el tratamiento a largo plazo de la hipertensión en Argentina, Chile, Colombia, Guatemala y México. Materiales y método: estudio prospectivo, observacional, de cohorte única de 48 semanas, en el que se incluyeron adultos con hipertensión no controlada, tratados con una combinación fija de amlodipino/irbesartán según criterio médico y fueron seguidos en la consulta ambulatoria habitual. La presión arterial objetivo fue < 140/90 mmHg (< 130/80 mmHg para pacientes con diabetes o enfermedad renal). Resultados: Se incluyeron 509 pacientes (57.6% mujeres). La edad media (DE) y la puntuación de riesgo de Framingham a 10 años fueron 60.6 (12.5) años y 9.9 (8.78), respectivamente. Durante 48 semanas de seguimiento, 97.4% de los pacientes tomaron ≥ 80% de las dosis prescritas. Se observó mejoría en la presión arterial estadísticamente significativa y clínicamente importante (-25.7/-13.5 mmHg; p < 0.001). Se logró control en el 62.7% de los pacientes. El cumplimiento del tratamiento fue uno de los predictores significativos (p < 0.05) de la presión arterial objetivo. Ochenta y siete (17.1%) pacientes experimentaron 117 eventos adversos emergentes del tratamiento, incluidos 7 eventos graves en 5 (1.0%) pacientes. Los eventos adversos fueron generalmente leves (75.2%) y se consideró que no estaban relacionados con el tratamiento (76.1%). Los acontecimientos adversos más frecuentes fueron edema periférico (3.9% de los pacientes) y mareos (1.0%). Según las estimaciones de Kaplan-Meier, el tiempo medio (SE) hasta la interrupción del tratamiento por acontecimientos adversos fue de 32.85 (0.08) semanas. Conclusiones: El tratamiento con irbesartán/amlodipino en combinación de dosis fijas mostró efectividad clínica, con mejoría significativa de las cifras de presión, y mayor llegada a la meta en quienes cumplieron con el tratamiento. El tratamiento fue bien tolerado, con un bajo porcentaje de eventos adversos relacionados con el tratamiento, y pocos eventos graves. La combinación en dosis fija es una importante herramienta en el tratamiento de la hipertensión arterial.

Abstract Introduction: Latin American population-based studies have shown suboptimal blood pressure control rates. Fixed-dose anti-hypertensive combinations are associated with improved convenience, adherence and effectiveness compared with monotherapy. Objective: assessed the real-life effectiveness and safety of fixed amlodipine/irbesartan combination in long-term management of hypertension in Argentina, Chile, Colombia, Guatemala, and Mexico. Materials and method: this was a 48-week, prospective, observational, single-cohort study, which included adults with uncontrolled hypertension, treated with fixed amlodipine/irbesartan combination per the treating physician's judgment, were followed in routine care. Target blood pressure was < 140/90 mmHg (< 130/80 mmHg for patients with diabetes or renal disease). Results: 509 patients (57.6% females) were included. Mean (SD) age and Framingham 10-year risk-score were 60.6 (12.5) years and 9.9 (8.78), respectively. Over 48 weeks, 97.4% of patients took ≥ 80% of prescribed doses. Statistically significant and clinically important blood pressure improvements (-25.7/-13.5 mmHg; p < 0.001) were observed. Control was achieved by 62.7% of patients. Treatment compliance was one of the significant (p < 0.05) predictors of target blood pressure achievement. Eighty-seven (17.1%) patients experienced 117 treatment-emergent adverse events, including 7 serious events by 5 (1.0%) patients. Adverse events were generally mild (75.2%) and judged not to be treatment-related (76.1%). The most common adverse events were peripheral edema (3.9% of patients) and dizziness (1.0%). Based on Kaplan-Meier estimates, the mean (SE) time to adverse event-related discontinuation was 32.85 (0.08) weeks. Conclusion: Treatment with the fixed-dose combination of irbesartan/amlodipine demonstrated clinical effectiveness, with a significant improvement in blood pressure values and a higher rate of achieving the treatment goal in those who adhered to the regimen. The treatment was well-tolerated, with a low percentage of treatment-related adverse events, and few severe events. The fixed-dose combination is an important tool in the management of arterial hypertension.

Consultorio de Atención Inmediata como estrategia de gestión de calidad en el Servicio de Emergencia de un hospital de Lima, Perú

Objetivo: Determinar la eficacia de la implementación del Consultorio de Atención Inmediata como estrategia de gestión de calidad en el Servicio de Emergencia de un hospital público de Lima, Perú. Materiales y métodos: Estudio analítico, cuasi experimental de antes y después, con grupos diferentes, que se realizó en 338 usuarios externos atendidos en el Servicio de Emergencia del Hospital María Auxiliadora. Se evaluó el tiempo de espera antes y después de la implementación del Consultorio de Atención Inmediata, así como la satisfacción a través del cuestionario SERVQUAL modificado -validado y recomendado por el Ministerio de Salud (Minsa) y aplicado en el grupo de posimplementación-, además de su relación con el tiempo de espera obtenido. El análisis se realizó a través del software de IBM SPSS S25.0 mediante medidas de frecuencias y porcentajes, diferencia de medias en grupos distintos con el test de Levene y la medida no paramétrica del coeficiente de correlación de Spearman con un nivel de significancia p < 0,05. Resultados: Los resultados mostraron predominio del sexo femenino (60,95 %), en el rango de edad de 14 a 29 años (24,56 %), en la prioridad IV (67,16 %); el tiempo de espera para la atención tuvo una media de 17,70 previo a la implementación y una media de 4,27 posterior a esta, por lo tanto, hubo una diferencia significativa después de la estrategia de gestión (p < 0,00). La satisfacción del Consultorio de Atención Inmediata se obtuvo en el 56,21 % de los usuarios externos, con énfasis en la dimensión empatía (76,33 %) y capacidad de respuesta (69,23 %), mientras que la dimensión con menor satisfacción fue la fiabilidad (48,52 %), además de obtener una correlación significativa inversa entre el tiempo de espera y la satisfacción (p < 0,01 y rho: -0,39). Conclusiones: La implementación del Consultorio de Atención Inmediata en el Servicio de Emergencia fue eficaz; en consecuencia, el tiempo de espera disminuyó, lo cual, a su vez, generó satisfacción en el usuario externo.

Objective: To determine the effectiveness of the implementation of the Immediate Care Office as a quality management strategy at the Emergency Service of a public hospital in Lima, Peru. Materials and methods: An analytical, quasi-experimental, before-and-after study conducted with 338 outpatients from different groups treated at the Emergency Service of Hospital María Auxiliadora. Before and after the implementation of the Immediate Care Office, waiting time, satisfaction-assessed through the modified SERVQUAL questionnaire, which was validated and recommended by the Ministry of Health and administered to the postimplementation group-as well as the relationship between satisfaction and waiting time were evaluated. The analysis was performed using IBM SPSS statistics V25.0, frequencies and percentages, the mean difference of both groups obtained through the Levene's test, and the nonparametric measurement of the Spearman's correlation coefficient with a significance level of p < 0.05. Results: The results showed a predominance of the female sex (60.95 %), the 14-to-29-year age range (24.56 %) and the Emergency Severity Index level IV (67.16 %). The average waiting time accounted for 17.70 and 4.27 before and after the office implementation, respectively. Therefore, there was a significant difference after the management strategy (p < 0.00). Out of all outpatients, 56.21 % were satisfied with the implementation of the Immediate Care Office, mainly with the empathy (76.33 %) and responsiveness (69.23 %) dimensions, while reliability was the dimension with the lowest satisfaction score (48.52 %). Additionally, there was a significant inverse correlation between waiting time and satisfaction (p < 0.01 and rho: -0.39). Conclusions: The implementation of the Immediate Care Office at the Emergency Service was effective since it reduced the waiting time, which in turn brought satisfaction to the outpatients.

The Long Shadow: A Historical Perspective on Racism in Medical Education.

To dismantle racism in U.S. medical education, people must understand how the history of Christian Europe, Enlightenment-era racial science, colonization, slavery, and racism shaped modern American medicine. Beginning with the coalescence of Christian European identity and empire, the authors trace European racial reasoning through the racial science of the Enlightenment into the White supremacist and anti-Black ideology behind Europe's global system of racialized colonization and enslavement. The authors then follow this racist ideology as it becomes an organizing principle of Euro-American medicine and examine how it manifests in medical education in the United States today. Within this historical context, the authors expose the histories of violence underlying contemporary terms such as implicit bias and microaggressions. Through this history, they also gain a deeper appreciation of why racism is so prevalent in medical education and how it affects admissions, assessments, faculty and trainee diversity, retention, racial climate, and the physical environment. The authors then recommend 6 historically informed steps for confronting racism in medical education: (1) incorporate the history of racism into medical education and unmask institutional histories of racism, (2) create centralized reporting mechanisms and implement systematic reviews of bias in educational and clinical activities, (3) adopt mastery-based assessment in medical education, (4) embrace holistic review and expand its possibilities in admissions, (5) increase faculty diversity by using holistic review principles in hiring and promotions, and (6) leverage accreditation to combat bias in medical education. These strategies will help academic medicine begin to acknowledge the harms propagated throughout the history of racism in medicine and start taking meaningful steps to address them. Although the authors have focused on racism in this paper, they recognize there are many forms of bias that impact medical education and intersect with racism, each with its particular history, that deserve their own telling and redress.

Análisis clínico-tomográfico en obstrucción de intestino delgado por adherencias según el sitio de obstrucción en la tomografía / Clinical-tomographic analysis in adhesive small bowel obstruction according to obstruction site in the tomography

Introducción. Las adherencias postoperatorias son la causa más frecuente de obstrucción de intestino delgado. La clínica sugiere el diagnóstico, pero de manera poco precisa la causa y el sitio de la obstrucción. La tomografía computarizada contrastada es el estudio óptimo y permite identificar de manera oportuna a los pacientes que requieren intervención quirúrgica. El objetivo de este estudio fue analizar la correlación entre la clínica y el sitio de obstrucción detectado en la tomografía computarizada contrastada de abdomen, en pacientes con sospecha diagnóstica de obstrucción de intestino delgado por adherencias. Métodos. Estudio prospectivo, transversal y analítico de pacientes con sospecha clínica de obstrucción de intestino delgado por adherencias y antecedentes quirúrgicos y su correlación con el sitio de obstrucción detectado en la tomografía computarizada de abdomen contrastada, de pacientes atendidos entre marzo de 2016 y febrero de 2019 en un hospital de segundo nivel. Resultados. Se incluyeron 41 pacientes, la media de edad fue de 59 años y el género masculino el más comprometido (68,3 %, n=28); la ausencia de evacuaciones estuvo presente en 97,5 % (p=0,026). La tomografía computarizada contrastada mostró el sitio de obstrucción en 73 % de los pacientes y la localización de la obstrucción más prevalente fue en íleon distal (31,7 %, n=13). Se asoció a leucocitosis (p=0,041) y a dolor más intenso (p=0,049), sin presentar irritación peritoneal. Conclusión. La obstrucción localizada en el íleon distal se caracterizó por presentar más dolor y mayor recuento leucocitario, sin correlación como factor de riesgo para requerir tratamiento quirúrgico.

Introduction. Postoperative adhesions are the most common cause of small bowel obstruction. The clinical presentation suggests the diagnosis, but imprecisely the cause and the site of the obstruction. Contrast computed tomography is the optimal study and allows the timely identification of patients requiring surgical intervention. The objective of this study was to analyze the correlation between the symptoms and the obstruction site detected in the abdominal contrasted computed tomography in patients with suspected diagnosis of small bowel obstruction due to adhesions. Methods. Prospective, cross-sectional and analytical study of patients with clinical suspicion of small bowel obstruction due to adhesions and surgical history, and its correlation with the obstruction site detected in the abdominal contrasted computed tomography, during March 2016 to February 2019 in a secondary level hospital. Results. Forty-one patients were included, the mean age was 59 years and the male gender was the most frequent (68.3%, n=28); the absence of evacuations was present in 97.5% (p=0.026). Contrast computed tomography showed the obstruction site in 73% of the patients. The most prevalent location of the obstruction was in the distal ileum (31.7%, n=13). It was associated with leukocytosis (p=0.041) and more intense pain (p=0.049), without presenting peritoneal irritation. Conclusion. The obstruction located in the distal ileum was characterized by more pain and a higher white blood cell count, without correlation as a risk factor for requiring surgical treatment

SNAP23 is essential for platelet and mast cell development and required in connective tissue mast cells for anaphylaxis.

Degranulation, a fundamental effector response from mast cells (MCs) and platelets, is an example of regulated exocytosis. This process is mediated by SNARE proteins and their regulators. We have previously shown that several of these proteins are essential for exocytosis in MCs and platelets. Here, we assessed the role of the SNARE protein SNAP23 using conditional knockout mice, in which SNAP23 was selectively deleted from either the megakaryocyte/platelet or connective tissue MC lineages. We found that removal of SNAP23 in platelets results in severe defects in degranulation of all three platelet secretory granule types, i.e., alpha, dense, and lysosomal granules. The mutation also induces thrombocytopenia, abnormal platelet morphology and activation, and reduction in the number of alpha granules. Therefore, the degranulation defect might not be secondary to an intrinsic failure of the machinery mediating regulated exocytosis in platelets. When we removed SNAP23 expression in MCs, there was a complete developmental failure in vitro and in vivo. The developmental defects in platelets and MCs and the abnormal translocation of membrane proteins to the surface of platelets indicate that SNAP23 is also involved in constitutive exocytosis in these cells. The MC conditional deletant animals lacked connective tissue MCs, but their mucosal MCs were normal and expanded in response to an antigenic stimulus. We used this mouse to show that connective tissue MCs are required and mucosal MCs are not sufficient for an anaphylactic response.

Aspectos relevantes de la enfermedad por hígado graso no alcohólico / Relevant aspects of nonalcoholic fatty liver disease

La enfermedad por hígado graso no alcohólico (EHGNA) es una condición que incluye desde la esteatosis hepática simple y la esteatohepatitis, hasta la cirrosis hepática y eventualmente el carcinoma hepatocelular. La diabetes tipo 2 y la obesidad son los principales factores asociados a la EHGNA. Su prevalencia en la población general se ha descrito entre el 20% y el 30%. Estos pacientes tienen un riesgo aumentado de mortalidad y presentan mayor incidencia que la población general de complicaciones hepáticas y cardiovasculares. La asociación de diferentes factores promueve la acumulación de ácidos grasos en el parénquima hepático, generando un estado de estrés, con formación de radicales de oxígeno y liberación de citoquinas inflamatorias que determinan la progresión de la enfermedad. Aunque existen diferentes pruebas no invasivas para el diagnóstico y estadificación de esta entidad, la biopsia hepática es la única prueba que permite identificar de manera fiable la presencia de inflamación, además del grado de fibrosis. El tratamiento actual de la EHGNA se basa en los cambios de estilo de vida del paciente, que han demostrado ser efectivos, incluso para revertir la fibrosis. Desafortunadamente, la adherencia a las medidas generales es muy pobre, de ahí la necesidad de contar con estrategias farmacológicas. Hasta el momento, no contamos con medicamentos aprobados por las agencias regulatorias para esta entidad, y los únicos fármacos recomendados por las sociedades internacionales son la pioglitazona y la vitamina E, que no están exentas de efectos adversos. Actualmente se encuentran bajo investigación diferentes medicamentos que buscan reducir la actividad inflamatoria sin aumento de la fibrosis, o mejoría de la fibrosis sin deterioro de la esteatohepatitis.

Nonalcoholic fatty liver disease (NAFLD) is a condition that ranges from simple hepatic steatosis and steatohepatitis, to liver cirrhosis and eventually hepatocellular carcinoma. Type 2 diabetes and obesity are the main factors associated with NAFLD. The prevalence in the general population has been described between 20% and 30%. These patients are at increased risk of mortality and have a higher incidence than the general population of liver and cardiovascular complications. The association of different factors promotes the accumulation of fatty acids in the liver parenchyma, generating a state of stress, with the formation of oxygen radicals and the release of inflammatory cytokines that determine the progression of the disease. Although there are different non-invasive tests for the diagnosis and staging of this condition, liver biopsy is the only test that reliably identifies the presence of inflammation, in addition to the degree of fibrosis. The current treatment of NAFLD is based on changes in the patient's lifestyle, which have been shown to be effective, including in reversing fibrosis. Unfortunately, adherence to general measures is very poor, hence the need for pharmacological strategies. So far, we do not have drugs approved by the regulatory agencies for this disease, and the only drugs recommended by international societies are pioglitazone and vitamin E, which are not exempt from adverse effects. Currently, different drugs are under investigation that seek to reduce inflammatory activity without increasing fibrosis, or improvement of fibrosis without deterioration of steatohepatitis.

Aspectos relevantes en hemocromatosis hereditaria / Relevant features in hereditary hemochromatosis

La hemocromatosis hereditaria es una enfermedad que se caracteriza por la sobrecarga sistémica de hierro y se asocia a múltiples mutaciones genéticas que conducen a una producción inadecuadamente baja de la hormona hepcidina o a una alteración en la unión de la hepcidina a la ferroportina. Esto tiene como resultado un aumento de la absorción intestinal y el depósito de cantidades excesivas de hierro en las células, lo cual, a su vez, si no se corrige, genera daño tisular. La expresión clínica puede variar desde individuos completamente asintomáticos, hasta pacientes con cirrosis hepática a temprana edad, y eventualmente carcinoma hepatocelular. Habitualmente, el diagnóstico no es invasivo e incluye el examen clínico, la evaluación de los parámetros de hierro plasmático, imágenes y pruebas genéticas. El principal tratamiento es la flebotomía, pero terapias alternativas como la suplementación con hepcidina son un tema de investigación actual.

Hereditary hemochromatosis is a disease characterized by systemic iron overload of genetic origin, that leads to an inadequately low production of the hormone hepcidin or a reduction in hepcidinferroportin binding. This results in an increased intestinal absorption and the deposit of excessive amounts of iron in cells, which in turn results in tissue damage if not treated. The clinical expression can vary from completely asymptomatic individuals, to patients with liver cirrhosis at an early age, and eventually hepatocellular carcinoma. Diagnosis is usually noninvasive and includes clinical examination, assessment of plasma iron levels, imaging studies, and genetic testing. The main medical treatment is phlebotomy, but alternative therapies such as hepcidin supplementation are the subject of current research.

Caracterización de pacientes con cirrosis hepática y bacteriemia de un hospital universitario en Medellín, Colombia / Characterization of patients with liver cirrhosis and bacteremia treated at a university hospital in Medellín-Colombia

Resumen Introducción: la bacteriemia en pacientes cirróticos es frecuente y se asocia con una alta mortalidad y hospitalización prolongada. Este estudio describe las características demográficas, clínicas y de laboratorio en pacientes con cirrosis hepática y bacteriemia en un hospital de cuarto nivel. Métodos: estudio observacional de cohorte retrospectiva. Incluyó pacientes con cirrosis hepática y bacteriemia entre el 1 de enero de 2010 y el 31 de diciembre de 2017 en el Hospital Pablo Tobón Uribe de Medellín, Colombia. Se recogieron variables demográficas, clínicas y de laboratorio. Se estimó la supervivencia durante el tiempo de hospitalización y hasta 30 días desde el diagnóstico de bacteriemia. Resultados: se hallaron 78 pacientes con cirrosis y bacteriemia. La media de edad fue de 65 años, 66,7 % fueron mujeres. Las principales etiologías de la cirrosis fueron: criptogénica (30,8 %) y esteatohepatitis no alcohólica (EHNA; 19,3 %). La principal fuente de infección fue la vía urinaria (24 %), seguida de colangitis (23 %) y la bacteriemia espontánea (19 %). Los bacilos gramnegativos (BGN) representaron la mayoría de los aislamientos (67,9 %). La prevalencia de multidrogorresistentes (MDR) fue de 25,6 % y el uso adecuado de antibiótico empírico fue de 80,8 %. La mortalidad a 30 días fue de 11,5 %. Como mejores predictores de mortalidad se encontraron la puntuación Child-Pugh y Model for End-stage Liver Disease (MELD) al ingreso con área bajo la curva ROC (AUROC) de 0,79 (p = 0,008) y 0.72 (p = 0,042), respectivamente. Conclusiones: los hallazgos permiten conocer las principales características de los pacientes con cirrosis que desarrollan bacteriemia en nuestro medio. Se encontró un número considerable de infecciones MDR. Los pacientes con un grado avanzado de la cirrosis son los que presentan un mayor riesgo de mortalidad.

Abstract Introduction: Bacteremia in cirrhotic patients is frequent and associated with high mortality and prolonged hospital stays. This study describes the demographic, clinical, and laboratory characteristics of patients with liver cirrhosis and bacteremia treated at a quaternary care hospital. Methodology: Observational, retrospective cohort study. The sample consisted of patients with liver cirrhosis and bacteremia treated between January 1, 2010, and December 31, 2017, at the Hospital Pablo Tobon Uribe of Medellín, Colombia. Demographic, clinical, and laboratory variables were collected. Survival was estimated during the time of hospitalization and up to 30 days following the diagnosis of bacteremia. Results: 78 patients had cirrhosis and bacteremia. The average age was 65 years; 66.7% were women. Cirrhosis was labeled cryptogenic in 30.8% of the cases and NASH in 19.3%. The main source of infection was the urinary tract (24%), followed by cholangitis (23%) and spontaneous bacteremia (19%). Gram-negative bacteria were observed in most of the isolates (67.9%). The prevalence of MDR was 25.6%, and the adequate use of empirical antibiotics was 80.8%. The 30-day mortality rate was 11.5%. The best mortality predictors were the Child-Pugh and MELD scores on admission with AUROC of 0.79 (P=0.008) and 0.72 (P=0.042), respectively. Conclusions: The findings allow describing the main characteristics of patients with cirrhosis who develop bacteremia in our environment. A considerable number of MDR infections were found. Patients with an advanced degree of cirrhosis are at the highest risk of mortality.

Phylogenetic relationships of Strongyloides species in carnivore hosts.

Strongyloides stercoralis is a parasitic nematode and a major pathogen responsible for human strongyloidiasis. The presence of this species in the dog population has led to an interest in studying the phylogenetic relationships among Strongyloides spp. in carnivore hosts. In the present study, Strongyloides spp. from various carnivore hosts (raccoon, Japanese badger, Siberian weasel, raccoon dog, masked palm civet, and domestic cat) were sought. Except for civets, Strongyloides spp. were identified in all host species. Based on 18S rDNA sequences, nine OTUs (operational taxonomy units) were identified. Molecular phylogenetic analyses using 18S28S rDNA and mitochondrial cox1 (cytochrome c oxidase subunit 1) sequences clustered them into two groups. The first group (named the stercoralis/procyonis group) was comprised of six OTUs and occurred in cats, raccoon dogs, raccoons (S. procyonis), Siberian weasels, and Japanese badgers and included S. stercoralis from humans and dogs. The second group (named the planiceps group) was made up of Strongyloides spp. from raccoon dogs (two OTUs) and one OTU from Siberian weasels. Subsequent analysis using almost the full-length nucleotide sequences of protein-coding genes in their mitochondrial genomes placed Strongyloides spp. of cats in a sister taxon position to S. stercoralis, whereas S. procyonis from raccoons was more distantly related to them. The presence of Strongyloides spp. from various carnivore hosts, which are close relatives of S. stercoralis, suggests this group of Strongyloides (the stercoralis/procyonis group) essentially evolved as parasites of carnivores, although more data on Strongyloides spp. from primate hosts are needed.

Disruption of PCDH10 and TNRC18 Genes due to a Balanced Translocation.

Balanced chromosomal rearrangements are usually associated with a normal phenotype, although in some individuals, phenotypic alterations are observed. In these patients, molecular characterization of the breakpoints can reveal the pathogenic mechanism, providing the annotation of disease-associated loci and a better genotype-phenotype correlation. In this study, we describe a patient with a balanced reciprocal translocation between 4q27 and 7p22 associated with neurodevelopmental delay. We performed cytogenetic evaluation, next-generation sequencing of microdissected derivative chromosomes, and Sanger sequencing of the junction points to define the translocation's breakpoints at base pair resolution. We found that the PCDH10 and TNRC18 genes were disrupted by the breakpoints at chromosomes 4 and 7, respectively, with the formation of chimeric genes at the junction points. Gene expression studies in the patient's peripheral blood showed reduced expression of TNRC18, a gene with unknown function and clinical significance. PCDH10 plays a role in the development of the nervous system and might be involved with the patient's neurodevelopmental delay. In this study, the full molecular characterization of the junction points was shown as an efficient tool for fine breakpoint mapping in balanced translocations in order to unmask gene disruptions and investigate the potential pathogenic role of the disrupted genes.

Comparación entre dos esquemas de inmunosupresión en pacientes con trasplante hepático en un hospital universitario en Medellín, Colombia / Comparison of two immunosuppressive regimens for liver transplant patients in a university hospital in Medellín, Colombia

Introducción. El desarrollo de terapias inmunosupresoras en trasplante hepático ha sido uno de los aspectos fundamentales que ha permitido disminuir la presencia de rechazos y mejorar la supervivencia del injerto y de los pacientes. El presente estudio se hizo para conocer la efectividad de dos esquemas de tratamiento inmunosupresor en una cohorte de pacientes con trasplante hepático, entre 2006 y 2017, en un hospital universitario en Medellín, Colombia. Metodología. Se realizó un estudio observacional retrospectivo donde se compararon dos esquemas de tratamiento inmunosupresor con ciclosporina (CsA) y azatioprina (AZA) versus tacrolimus (TAC) y micofenolato (MMF). Resultados. Se incluyeron 147 pacientes al estudio, 79 mujeres y 68 hombres, con una mediana de edad de 55 años. La tasa de incidencia de rechazo agudo en el grupo CsA/AZA fue de 7,3 y para el grupo TAC/MMF fue de 13,8, con una razón de tasas de 0,53 (IC95%=0,31-0,89) por cada 100 personas/año, siendo esta diferencia estadísticamente significativa (p=0,02). No hubo diferencias significativas entre los grupos con respecto a la presencia de rechazo crónico, supervivencia del injerto o de los pacientes. Con respecto a los efectos adversos asociados al tratamiento, solo hubo diferencia significativa en una mayor presencia de diarrea en el grupo TAC/MMF. Conclusión. Solo se encontró diferencia significativa en cuanto a un número mayor de rechazos agudos en el grupo tratado con TAC/MMF. Estos hallazgos están en concordancia con la experiencia local, en la que en pacientes seleccionados se puede utilizar este esquema, con buenos resultados clínicos y menores costos para el sistema de salud. Hasta el momento, esta es la primera cohorte retrospectiva de Colombia y Latinoamérica que realiza una comparación, como la expuesta.

Introduction. The development of immunosuppressive therapies in liver transplantation has been one of the major contributing factors that have reduced the presence of rejections and improved graft and patient survival. The present study was conducted to determine the effectiveness of two immunosuppressive schemes in a cohort of liver transplant patients, between 2006 and 2017, at a university hospital in Medellin, Colombia. Methodology. A retrospective observational study was performed to compare two immunosuppressive treatment schemes with cyclosporine (CsA) and azathioprine (AZA) versus tacrolimus (TAC) and mycophenolate (MMF). Results. A total of 147 patients were included in the study, 79 women and 68 men, with a median age of 55 years. The incidence rate of acute rejection in the CsA/AZA group was 7.3 while in the TAC/MMF group was 13.8, with a rate ratio of 0.53 (95%CI=0.31-0.89) for every 100 person-year, this difference being statistically significant (p=0.02). There were no significant differences between the groups regarding the presence of chronic rejection, graft or patient survival. With respect to adverse effects associated with the treatment, there was only a significant difference in the presence of diarrhea in the TAC/MMF group. Conclusion. A significant difference was only found in terms of a higher number of acute rejections in the group treated with TAC/MMF. These findings are in agreement with local experience, in which this scheme can be used in selected patients, with good clinical results and lower costs for the health system. So far, this is the first retrospective study in Colombia and Latin America to make a comparison such as the one presented.

Prevalência de las alteraciones del metabolismo óseo-mineral asociadas a enfermedad renal crónica no en diálisis / Prevalence of bone mineral metabolism alterations associated with chronic kidney disease not on dialysis

Resumen Introducción: la enfermedad renal crónica (ERC) es un problema de salud pública, siendo el trastorno del metabolismo óseo mineral una de sus principales complicaciones y que contribuye directamente a la morbilidad y mortalidad. Varios estudios previos han demostrado un aumento de su prevalencia a medida que disminuye la tasa de filtración glomerular (TFG), sin embargo, no contamos con datos en nuestro país ni en América Latina. Métodos: realizamos un estudio transversal unicéntrico en un servicio de consulta de nefrología, en adultos con ERC G1 a 5 que no estuvieran en terapia de reemplazo renal, evaluados entre enero de 2014 y marzo de 2015. La recolección de datos se realizó con un instrumento predefinido que incluía datos demográficos, alteraciones de los parámetros del metabolismo mineral y óseo, y su manejo. Resultados: se incluyeron 2026 pacientes, de los cuales 1756 tenían medición de hormona paratiroidea, la edad promedio fue 74 años, el 62 % eran mujeres. La distribución por grados de ERC fue: G1:4,9 %, G2:22,8 %, G3: 57,4 %, G4: 12,5 % y G5:2,4 %. Las principales causas fueron la nefropatía hipertensiva y diabética. Encontramos deficiencia de vitamina D en el 78,16 %, hiperparatiroidismo secundario en el 63,67 % e hiperfosfatemia en el 12,38 %, con aumento de la prevalencia a medida que la TFG empeoraba. Conclusiones: encontramos que las alteraciones del metabolismo mineral y óseo son frecuentes en los pacientes con enfermedad renal crónica e inician desde estadios tempranos, como se ha demostrado en otros estudios. Consideramos que estos resultados llevarán a nuevas investigaciones de manejo en pacientes con ERC.

Abstract Background: chronic kidney disease (CKD) is a public health problem, and bone mineral metabolism disorder is one of its main complications that directly contributes to morbidity and mortality. Several previous studies have shown an increase in its prevalence as the glomerular filtration rate (GFR) decreases, however, we do not have data from our country or Latin America. Methods: We conducted a unicentric cross-sectional study in a nephrology consultation service in adults with CKD G1 to 5 who were not in renal replacement therapy, evaluated between January 2014 and March 2015. Data collection was performed with an instrument predefined that included demographic data, alterations of the mineral and bone metabolism parameters, and their management. Results: 2026 patients were included, of whom 1756 had parathyroid hormone measurement, the average age was 74 years, 62% were women. The distribution by degrees of CKD was: G1: 4.9%, G2: 22.8%, G3: 57.4%, G4: 12.5% and G5: 2.4%. The main causes were hypertensive and diabetic nephropathy. We found vitamin D deficiency in 78.16%, secondary hyperparathyroidism in 63.67% and hyperphosphatemia in 12.38%, with an increase in prevalence as GFR worsened. Conclusions: We found that mineral and bone metabolism alterations are frequent in patients with chronic kidney disease and start from early stages, as has been demonstrated in other studies. We believe that these results will lead to new management investigations in patients with CKD.

Munc18-2, but not Munc18-1 or Munc18-3, regulates platelet exocytosis, hemostasis, and thrombosis.

Platelet degranulation, a form of regulated exocytosis, is crucial for hemostasis and thrombosis. Exocytosis in platelets is mediated by SNARE proteins, and in most mammalian cells this process is controlled by Munc18 (mammalian homolog of Caenorhabditis elegans uncoordinated gene 18) proteins. Platelets express all Munc18 paralogs (Munc18-1, -2, and -3), but their roles in platelet secretion and function have not been fully characterized. Using Munc18-1, -2, and -3 conditional knockout mice, here we deleted expression of these proteins in platelets and assessed granule exocytosis. We measured products secreted by each type of platelet granule and analyzed EM platelet profiles by design-based stereology. We observed that the removal of Munc18-2 ablates the release of alpha, dense, and lysosomal granules from platelets, but we found no exocytic role for Munc18-1 or -3 in platelets. In vitro, Munc18-2-deficient platelets exhibited defective aggregation at low doses of collagen and impaired thrombus formation under shear stress. In vivo, megakaryocyte-specific Munc18-2 conditional knockout mice had a severe hemostatic defect and prolonged arterial and venous bleeding times. They were also protected against arterial thrombosis in a chemically induced model of arterial injury. Taken together, our results indicate that Munc18-2, but not Munc18-1 or Munc18-3, is essential for regulated exocytosis in platelets and platelet participation in thrombosis and hemostasis.

Syntaxin 3, but not syntaxin 4, is required for mast cell-regulated exocytosis, where it plays a primary role mediating compound exocytosis.

Mast cells (MCs) participate in allergy, inflammation, and defense against pathogens. They release multiple immune mediators via exocytosis, a process that requires SNARE proteins, including syntaxins (Stxs). The identity of the Stxs involved in MC exocytosis remains controversial. Here, we studied the roles of Stx3 and -4 in fully developed MCs from conditional knockout mice by electrophysiology and EM, and found that Stx3, and not Stx4, is crucial for MC exocytosis. The main defect seen in Stx3-deficient MCs was their inability to engage multigranular compound exocytosis, while leaving most single-vesicle fusion events intact. We used this defect to show that this form of exocytosis is not only required to accelerate MC degranulation but also essential to achieve full degranulation. The exocytic defect was severe but not absolute, indicating that an Stx other than Stx3 and -4 is also required for exocytosis in MCs. The removal of Stx3 affected only regulated exocytosis, leaving other MC effector responses intact, including the secretion of cytokines via constitutive exocytosis. Our in vivo model of passive systemic anaphylaxis showed that the residual exocytic function of Stx3-deficient MCs was sufficient to drive a full anaphylactic response in mice.

Biological Activities of Extracts from Aerial Parts of Salvia pachyphylla Epling Ex Munz.

The antioxidant, antimicrobial, antiproliferative, and enzyme inhibitory properties of five extracts from aerial parts of Salvia pachyphylla Epling ex Munz were examined to assess the prospective of this plant as a source of natural products with therapeutic potential. These properties were analyzed by performing a set of standard assays. The extract obtained with dichloromethane showed the most variety of components, as they yielded promising results in all completed assays. Furthermore, the extract obtained with ethyl acetate exhibited the greatest antioxidant activity, as well as the best xanthine oxidase inhibitory activity. Remarkably, both extracts obtained with n-hexane or dichloromethane revealed significant antimicrobial activity against the Gram-positive bacteria; additionally, they showed greater antiproliferative activity against three representative cell lines of the most common types of cancers in women worldwide, and against a cell line that exemplifies cancers that typically develop drug resistance. Despite that, other extracts were less active, such as the methanolic or aqueous; their results are promising for the isolation and identification of novel bioactive molecules.