Comparación de la respuesta pro-inflamatoria de células endoteliales de arteria coronaria humana frente a Porphyromonas gingivalis w83 entre un modelo de cultivo celular tridimensional y uno bidimensional

Porphyromonas gingivalis (Pg) tiene capacidad de inducir respuesta inflamatoria celular sobre células endoteliales (CEs), lo cual puede conducir a procesos de disfunción endotelial relacionados con aterosclerosis. Los estudios in vitro para el estudio de los mecanismos de patogenia inducidos por periodontopatógenos se han hecho principalmente en modelos de cultivos celular sobre superficies plásticas bidimensionales (2D) los cuales no simulan el soporte natural sobre el cual las células endoteliales reposan en los tejidos. Sin embargo, se desconoce la respuesta de CE frente a P. gingivalis en modelos tridimensionales. Por tanto, se hace necesario el estudio en modelos de cultivo celular tridimensional mediante el uso de matrices extracelulares (MECs) biológicas como el colágeno que proporcionan el soporte tridimensional (3D). Por tal motivo el objetivo de este trabajo fue comparar la respuesta pro-inflamatoria de células endoteliales frente a Porphyromonas gingivalis entre un modelo de cultivo celular tridimensional y uno bidimensional. Para esto Células Endoteliales de Arteria Coronaria Humana (HCAEC) fueron cultivadas en placas de poliestireno hasta alcanzar confluencia (cultivo 2D) o por 7 días sobre matrices de colágeno tipo I (cultivo 3D). Los cultivos 2D y 3D fueron infectados a MOI 1 y MOI 100 con bacteria viva P.g de la cepa W83 durante 24 horas. Cultivos sin estimular se usaron como control negativo. Se evaluó la expresión génica de COX-2, eNOS y factor von Willebrand (vWF) mediante q-RT PCR. A partir de los sobrenadantes recolectados de ambos cultivos se cuantificó los niveles de citocinas pro-inflamatorias IL-1α, IL-1β, IL-6 y TNF-β y quimiocinas RANTES, MCP-1α, MIP-1α e IL-8 mediante citometría de flujo. Niveles de tromboxano-A2 y prostaglandina-I2 se midieron por ELISA. Cada tratamiento fue realizado por triplicado y se aplicó la prueba de Kruskal- Wallis con U de Mann-Whitney, considerándose las diferencias estadísticamente significativas cuando p<0.05. En el cultivo 3D P.g a ambos MOI indujo un incremento significativo en la expresión de COX-2 con respecto al control, el cual no fue observado en el cultivo 2D. Igualmente se observaron diferencias en la expresión de COX-2 entre los cultivos 2D y 3D a MOI 1, siendo mayor en 3D. La expresión de eNOS y vWF no fueron inducidos por P.g en ambos cultivos. Igualmente, no se observaron diferencias en la secreción de prostaglandinas. En cuanto a la secreción de citocinas, en el cultivo 2D se observó un incremento significativo de IL-8 a MOI 100 pero una disminución de MCP-1 a ambos MOI con respecto al control; mientras que en el cultivo 3D se observó una reducción en la secreción de MCP-1 a MOI 100 con respecto al control. Los resultados de este estudio indican que P.g W83 induce una débil respuesta celular endotelial pro-inflamatoria en ambos modelos celulares. Sin embargo debido a las diferencias observadas en el cultivo 3D no se descarta el uso de los modelos tridimensionales para comprender los mecanismos de patogenia relacionados con disfunción endotelial aterosclerótica inducida por periodontopatógenos en un entorno más natural.ColcienciasPorphyromonas gingivalis (Pg) has capability to induce cellular inflammatory response on endothelial cells (ECs, which can to lead to endothelial dysfunction processes related with atherosclerosis. In vitro studies to understand the periodontopathogen-induced mechanism of pathogenesis have been made mainly in cell culture models on bidimensional plastic surfaces (2D), which doesn´t simulate the natural extracellular matrix that surrounds the ECs in the tissue. However, the EC inflammatory response in three-dimensional model is unknown. Therefore, it is necessary to study this response in three-dimensional cell culture models using biologic extracellular matrix, like collagen that gives three-dimensional structure. Therefore, the aim of this study was to compare the endothelial cell culture inflammatory response against Porphyromonas gingivalis among a three-dimensional cell culture and a bidimensional cell culture. Human Coronary Artery Endothelial Cells (HCAEC) were cultured in polystyrene plates until they conflated (2D Culture) or for 7 days on type I collagen matrixes (3D Culture). The 3D and 2D cultures were infected at MOI 100 and MOI 1 with Pg strain W83 for 24 hours. Cultures without stimuli were taken as negative controls. Genic expression by qRT-PCR of COX-2, eNOS and von Willebrand factor (vWF) were evaluated. Supernatants of both cell cultures were taken to measure pro-inflammatory cytokines levels IL-1α, IL-1β, IL-6 y TNF-β and chemokine levels RANTES, MCP-1α, MIP-1α e IL-8 through flow cytometry. Levels of thromboxane A2 and prostaglandin I2 were measured by ELISA. Each treatment was performed in triplicate and the Kruskal-Wallis with U-Mann-Whitney test was applied, p<0.05 was considered significant. The 3D cell culture P.g induced a significant increase in the expression of COX-2 with regard to control at both MOI, such effect was not observed in the 2D cell. Likewise significant difference was observed at MOI 1 among both cell cultures, being greater in 3D cell culture. Differences in prostaglandin I2 levels were not observed. Concerning cytokine production, in the 2D cell culture, IL-8 levels showed significant increase at MOI 100 but MCP-1 levels decreased at both MOI with regard to control, while in 3D cell culture its levels decreased at MOI 100 in relation to control. The results of this work indicate that Pg W83 induce a weak cellular endothelial pro-inflammatory response in both cell cultures models. However, due the differences observed in the 3D cell culture we do not rule out the use of three-dimensional models to understand the mechanism of pathogenesis related to periodontopathogens-induced atherosclerotic endothelial dysfunction in a more natural environment.Magíster en Ciencias BiológicasMaestrí

Importancia de la aplicación de normas de Bioseguridad en el área de Radiología.

La bioseguridad vista desde su misma composición debe garantizar la seguridad y protección de la población a la cual está dirigida, en este artículo se hace énfasis en el área de radiología invitando a establecer parámetros de convivencia normativa, donde lo vital debe ser la garantía de un espacio y ambiente digno para quienes, por sus funciones, han de velar por la seguridad de otros. De acuerdo a la normatividad se busca establecer, desarrollar, aplicar y generar ambientes libres de riesgos de contagio, especialmente en los espacios laborales, donde es muy frecuente la posibilidad de estar en contacto con infecciones que dan paso a la aparición de enfermedades, afectando la salud del entorno; es por ello, que surge la necesidad de cumplir los protocolos normativos que ayuden a preservar y conservar el medio ambiente en condiciones óptimas, procurando con ello, salvaguardar el bienestar y la salud de quienes están expuestos, a través de la ejecución de las normas establecidas para ello, en las cuales está plasmada de manera específica, la forma en que se debe dar uso a las herramientas y los medios de barrera; gracias a ello, se minimiza el riesgo de contagio y se generan mejores ambientes en el ámbito laboral.Los riesgos de contaminación, no pueden eliminarse totalmente, pero se puedenprevenir. La autodisciplina, puede garantizar un trabajo seguro, por lo tanto, se deben desarrollar estrategias de capacitación del autocuidado de la salud, prevención y del mejor cumplimiento de las reglamentaciones establecidas

Risk of developing checkpoint immune pneumonitis and its effect on overall survival in non-small cell lung cancer patients previously treated with radiotherapy

Introduction:Immune checkpoint inhibitor-related pneumonitis (ICIP) is a potentially lifethreatening immune-related adverse event (irAE), especially in non-small cell lung cancer(NSCLC) patients. Currently, the potential for increased irAE in patients who receiveradiotherapy is scarcely known, although a connection between antitumor immuneresponses and irAEs has been suggested. In this study, we evaluated the developmentof ICIP in non-small cell lung cancer patients with prior radiotherapy, treated withimmunotherapy in the second-line.Methods:In this retrospective trial, we included patients treated with second-lineimmunotherapy at the National Cancer Institute in Mexico City from February 2015 toFebruary 2018. Clinical, radiological and treatment variables were evaluated accordingto the presence of ICIP as defined by the Common Terminology Criteria for AdverseEvents (4.0) in patients with or without a previous (≥months) history of radiotherapy.Results:Among 101 NSCLC patients who received treatment with ICIs, 22 patients(21.8%) were diagnosed with ICIP, of which 73% (16/22) had a history of radiotherapy(OR 6.04, 95% CI 2.03−18.0,p<0.001). Median progression free survival and overallsurvival were similar in patients who developed ICIP compared with those who did not,however, patients who presented grade≥2 ICIP had an increased risk of mortality (HR2.54, 95% CI 1.20−5.34,p= 0.014).Conclusion:In this real-world cohort of NSCLC patients treated with ICI, the historyof prior radiotherapy was associated with increased risk for ICIP development. Unlikeother irAEs, grade≥2 ICIP is an independent prognostic factor for decreased survivalin NSCLC patients

Multigene mutation profiling and clinical characteristics of small-cell lung cancer in never-smokers vs. seavy smokers (Geno1.3-CLICaP)

Objectives: Lung cancer is a heterogeneous disease. Presentation and prognosis are known to vary according to several factors, such as genetic and demographic characteristics. Small-cell lung cancer incidence is increasing in never-smokers. However, the disease phenotype in this population is different compared with patients who have a smoking history. Material and Methods: To further investigate the clinical and genetic characteristics of this patient subgroup, a cohort of small cell lung cancer patients was divided into smokers (n = 10) and never/ever-smokers (n = 10). A somatic mutation profile was obtained using a comprehensive NGS assay. Clinical outcomes were compared using the Kaplan-Meier method and Cox proportional models. Results: Median age was 63 years (46–81), 40% were men, and 90% had extended disease. Smoker patients had significantly more cerebral metastases (p = 0.04) and were older (p = 0.03) compared to their non-smoker counterparts. For never/ever smokers, the main genetic mutations were TP53 (80%), RB1 (40%), CYLD (30%), and EGFR (30%). Smoker patients had more RB1 (80%, p = 0.04), CDKN2A (30%, p = 0.05), and CEBPA (30%, p = 0.05) mutations. Response rates to first-line therapy with etoposide plus cisplatin/carboplatin were 50% in smokers and 90% in never/ever smokers (p = 0.141). Median overall survival was significantly longer in never smokers compared with smokers (29.1 months [23.5–34.6] vs. 17.3 months [4.8–29.7]; p = 0.0054). Never/ever smoking history (HR 0.543, 95% CI 0.41–0.80), limited-stage disease (HR 0.56, 95% CI 0.40–0.91) and response to first-line platinum-based chemotherapy (HR 0.63, 95% CI 0.60–0.92) were independently associated with good prognosis. Conclusion: Our data supports that never/ever smoker patients with small-cell lung cancer have better prognosis compared to their smoker counterparts. Further, patients with never/ever smoking history who present with small-cell lung cancer have a different mutation profile compared with smokers, including a high frequency of EGFR, MET, and SMAD4 mutations. Further studies are required to assess whether the differential mutation profile is a consequence of a diverse pathological mechanism for disease onset

Genotyping squamous cell lung carcinoma in Colombia (Geno1.1-CLICaP)

Background: Lung cancer is a public health problem, and squamous cell carcinoma (SCC) is the second most prevalent subtype of this neoplasm. Compared to other subtypes, including adenocarcinoma, SCC is less well understood in terms of molecular pathogenesis, limiting therapeutic options among targeted agents approved for other disease subgroups. In this study, we sought to characterize the SCC genomic profile using a validated Next Generation Sequencing (NGS) platform. Methods: The comprehensive NGS assay (TruSight Tumor 170) was used in order to target the full coding regions of 170 cancer-related genes on SCC samples. PD-L1 expression in tumor cells (TCs) was assessed using clone 22C3 (Dako). Clinical outcomes were correlated with molecular profile, including progression free survival (PFS), overall response rate (ORR), and overall survival (OS). Results: A total of 26 samples were included, median age was 67 years (r, 33–83) and 53.8% were men. Tobacco consumption was identified in all subjects (mean 34-year package). For first-line treatment 80.8% of patients received cisplatin or carboplatin plus gemcitabine. In terms of molecular profile, we identified a high prevalence of inactivating mutations in TP53 (61.5%), PIK3CA (34.6%), MLL2 (34.6%), KEAP1 (38.4%), and NOTCH1 (26.9%). PD-L1 expression ranged from negative, 1, 2–49, and ≥50% in 23.1, 38.5, 26.9, and 11.5%, respectively. Interestingly, the genetic alterations did not have an effect in PFS, OS or ORR in this study. However, PDL1 expression was higher among those who had mutations in TP53 (p = 0.037) and greater expression of PDL1 was related to PIK3CA alterations (p = 0.05). Conclusions: The genomic profile of SCC encompasses important genes including TP53, PIK3CA and KEAP1. TP53 mutations could be associated with PDL1 expression, generating hypothesis regarding specific treatment options

Clonal chromosomal mosaicism and loss of chromosome Y in elderly men increase vulnerability for SARS-CoV-2

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, COVID-19) had an estimated overall case fatality ratio of 1.38% (pre-vaccination), being 53% higher in males and increasing exponentially with age. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, we found 133 cases (1.42%) with detectable clonal mosaicism for chromosome alterations (mCA) and 226 males (5.08%) with acquired loss of chromosome Y (LOY). Individuals with clonal mosaic events (mCA and/or LOY) showed a 54% increase in the risk of COVID-19 lethality. LOY is associated with transcriptomic biomarkers of immune dysfunction, pro-coagulation activity and cardiovascular risk. Interferon-induced genes involved in the initial immune response to SARS-CoV-2 are also down-regulated in LOY. Thus, mCA and LOY underlie at least part of the sex-biased severity and mortality of COVID-19 in aging patients. Given its potential therapeutic and prognostic relevance, evaluation of clonal mosaicism should be implemented as biomarker of COVID-19 severity in elderly people. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, individuals with clonal mosaic events (clonal mosaicism for chromosome alterations and/or loss of chromosome Y) showed an increased risk of COVID-19 lethality

EGFR exon 20 insertions in advanced non-small cell lung cancer: A new history begins

Although targeted therapy is standard of care in a large subset of oncogenic addicted non-small cell lung cancers (NSCLC), until recently, this therapeutic approach has not been feasible for all genomic alterations such as for those tumors harboring Epidermal Growth Factor Receptor (EGFR) exon 20 insertion (ex20ins) mutations. Despite being the third most common EGFR mutation, a limited efficacy of first- and second-generation EGFR tyrosine kinase inhibitors (TKI) exists. This is related to the heterogeneity at the molecular level in EGFR ex20ins mutation variants and the finding that this mutation promotes active kinase conformation but does not increase the affinity for EGFR TKI. As a result, the prognosis of this population is diminished. Therefore, chemotherapy remained the most suitable strategy in this subset of EGFR mutant NSCLC patients. Recently, new treatment strategies have been reported in this landscape, either with new EGFR TKI or bispecific antibodies, which may establish a new standard of care in the coming future for these patients. Future research should focus on elucidating the oncogenic degree of all EGFR ex20ins variants, the potential role of combination strategies either with chemotherapy or immune checkpoint inhibitors, and the most appropriate first-line treatment strategy in this subgroup. Finally, the knowledge of mechanisms of acquired resistance to these new agents upon progression is a priority for personalising treatment at that time. It is in this framework, that we provide a thorough overview on this subject

Squamous cell lung cancer: genomic evolution and personalized therapy

Objetivo. Revisar el estado del arte en relación con la información actual sobre el cáncer de pulmón de células escamosas (CPCE) y describir las anomalías genéticas reportadas, su efecto y los agentes terapéuticos más prometedores. Material y métodos. Se realizó una revisión de artículos publicados en revistas indizadas, así como las guías de tratamiento publicadas por instancias locales e internacionales. Resultados. El CPCE representa una proporción menor de la carga mundial de la enfermedad por cáncer pulmonar en comparación con su presentación más frecuente, el adenocarcinoma. Sin embargo, más de 400 000 casos son reportados anualmente, una población sustancial para quienes las opciones terapéuticas son escasas y con una eficacia limitada. Diversos grupos se han dado a la tarea de elucidar los mecanismos que conllevan al desarrollo del CPCE, incluyendo anomalías moleculares que puedan servir como blancos para el diseño de fármacos. Conclusiones. Existen blancos terapéuticos potenciales para el CPCE que deben ser estudiados en ensayos clínicos para ser validados.Objective. To review the state-of-the-art in relation to the current information on squamous cell lung cancer (SCLC). We describe the genetic anomalies reported, their effect, and fi­nally the most promising therapeutic agents. Materials and methods. We reviewed published articles in peer-reviewed journals as well as current treatment guidelines from local and international resources. Results. SCLC represents a smaller proportion of the global burden of disease for lung cancer compared to its more frequent presentation, the ade­nocarcinoma. However, more than 400 000 cases are reported annually, a substantial population for whom therapeutic op­tions are scarce and with limited efficacy. Several groups have been given the task of elucidating the mechanisms that lead to the development of SCLC, including molecular anomalies that can be used as targets for drug design. Conclusion. There are potential therapeutic targets for SCLC, which must be studied in clinical trials for validation