Generation of Human Lung Organoid Cultures from Healthy and Tumor Tissue to Study Infectious Diseases.

Respiratory viruses cause mild to severe diseases in humans every year, constituting a major public health problem. Characterizing the pathogenesis in physiologically relevant models is crucial for developing efficient vaccines and therapeutics. Here, we show that lung organoids derived from human primary or lung tumor tissue maintain the cellular composition and characteristics of the original tissue. Moreover, we show that these organoids sustain viral replication with particular infection foci formation, and they activate the expression of interferon-associated and proinflammatory genes responsible for mediating a robust innate immune response. All together, we show that three-dimensional (3D) lung organoids constitute a relevant platform to model diseases and enable the development of drug screenings. IMPORTANCE Three-dimensional (3D) human lung organoids reflect the native cell composition of the lung as well as its physiological properties. Human 3D lung organoids offer ideal conditions, such as timely availability in large quantities and high physiological relevance for reassessment and prediction of disease outbreaks of respiratory pathogens and pathogens that use the lung as a primary entry portal. Human lung organoids can be used in basic research and diagnostic settings as early warning cell culture systems and also serve as a relevant platform for modeling infectious diseases and drug development. They can be used to characterize pathogens and analyze the influence of infection on, for example, immunological parameters, such as the expression of interferon-associated and proinflammatory genes in the context of cancer. In our study, we found that cancer-derived lung organoids were more sensitive to influenza A virus infection than those derived from healthy tissue and demonstrated a decreased innate immune response.

Pan-cancer analysis distinguishes transcriptional changes of aneuploidy from proliferation.

Patterns of gene expression in tumors can arise as a consequence of or result in genomic instability, characterized by the accumulation of somatic copy number alterations (SCNAs) and point mutations (PMs). Expression signatures have been widely used as markers for genomic instability, and both SCNAs and PMs could be thought to associate with distinct signatures given their different formation mechanisms. Here we test this notion by systematically investigating SCNA, PM, and transcriptome data from 2660 cancer patients representing 11 tumor types. Notably, our data indicate that similar expression signatures can be derived from correlating gene expression with either SCNA or PM load. Gene sets related to cell growth and proliferation generally associated positively, and immunoregulatory gene sets negatively, with variant burden. In-depth analyses revealed several genes whose de-regulation correlates with SCNA but not with PM burden, yielding downstream effectors of TP53 and MYC signaling unique to high-SCNA tumors. We compared our findings to expression changes observed in two different cancer mouse models with persistent mitotic chromosomal instability, observing a decrease in proliferative expression signatures. Our results suggest that overexpression of cell-cycle-related genes are a characteristic of proliferation, and likely tumor evolution, rather than ongoing genomic instability.

Acute expression of human APOBEC3B in mice results in RNA editing and lethality.

BACKGROUND: RNA editing has been described as promoting genetic heterogeneity, leading to the development of multiple disorders, including cancer. The cytosine deaminase APOBEC3B is implicated in tumor evolution through DNA mutation, but whether it also functions as an RNA editing enzyme has not been studied. RESULTS: Here, we engineer a novel doxycycline-inducible mouse model of human APOBEC3B-overexpression to understand the impact of this enzyme in tissue homeostasis and address a potential role in C-to-U RNA editing. Elevated and sustained levels of APOBEC3B lead to rapid alteration of cellular fitness, major organ dysfunction, and ultimately lethality in mice. Importantly, RNA-sequencing of mouse tissues expressing high levels of APOBEC3B identifies frequent UCC-to-UUC RNA editing events that are not evident in the corresponding genomic DNA. CONCLUSIONS: This work identifies, for the first time, a new deaminase-dependent function for APOBEC3B in RNA editing and presents a preclinical tool to help understand the emerging role of APOBEC3B as a driver of carcinogenesis.

Acquisition of chromosome instability is a mechanism to evade oncogene addiction.

Chromosome instability (CIN) has been associated with therapeutic resistance in many cancers. However, whether tumours become genomically unstable as an evolutionary mechanism to overcome the bottleneck exerted by therapy is not clear. Using a CIN model of Kras-driven breast cancer, we demonstrate that aneuploid tumours acquire genetic modifications that facilitate the development of resistance to targeted therapy faster than euploid tumours. We further show that the few initially chromosomally stable cancers that manage to persist during treatment do so concomitantly with the acquisition of CIN. Whole-genome sequencing analysis revealed that the most predominant genetic alteration in resistant tumours, originated from either euploid or aneuploid primary tumours, was an amplification on chromosome 6 containing the cMet oncogene. We further show that these tumours are dependent on cMet since its pharmacological inhibition leads to reduced growth and increased cell death. Our results highlight that irrespective of the initial CIN levels, cancer genomes are dynamic and the acquisition of a certain level of CIN, either induced or spontaneous, is a mechanism to circumvent oncogene addiction.

Plk1 overexpression induces chromosomal instability and suppresses tumor development.

Polo-like kinase 1 (Plk1) is overexpressed in a wide spectrum of human tumors, being frequently considered as an oncogene and an attractive cancer target. However, its contribution to tumor development is unclear. Using a new inducible knock-in mouse model we report here that Plk1 overexpression results in abnormal chromosome segregation and cytokinesis, generating polyploid cells with reduced proliferative potential. Mechanistically, these cytokinesis defects correlate with defective loading of Cep55 and ESCRT complexes to the abscission bridge, in a Plk1 kinase-dependent manner. In vivo, Plk1 overexpression prevents the development of Kras-induced and Her2-induced mammary gland tumors, in the presence of increased rates of chromosome instability. In patients, Plk1 overexpression correlates with improved survival in specific breast cancer subtypes. Therefore, despite the therapeutic benefits of inhibiting Plk1 due to its essential role in tumor cell cycles, Plk1 overexpression has tumor-suppressive properties by perturbing mitotic progression and cytokinesis.

Mejora de conocimientos y satisfacción en una acción formativa sobre el servicio de dispensación de medicamentos / Improvement of knowledge and satisfaction in a training action on drug dispensing service

Introducción y objetivo: Evaluar la mejora de los conocimientos, las habilidades y la satisfacción de los alumnos en un curso sobre el servicio de dispensación. Material y métodos: Diseño: estudio cuasi experimental (antes/después) con intervención educativa. Participantes: farmacéuticos comunitarios de la provincia de Pontevedra. Se realizaron siete ediciones (octubre de 2018-octubre de 2019). Intervención educativa: curso semipresencial de 26 horas + 6 horas de taller presencial con simulaciones de casos y entrenamiento mediante role-playing. Evaluación: se evaluaron los dos primeros niveles de Kirkpatrik, reacción (satisfacción) y competencia (conocimientos y actuación clínica), mediante un cuestionario con 20 preguntas y cuatro casos prácticos. Para valorar la satisfacción se utilizó un cuestionario de tipo Likert con 12 preguntas. Los datos cualitativos se expresan como porcentajes y los cuantitativos como media -desviación estándar (DE)-. Resultados: Participaron 114 alumnos, un 78% mujeres, con 39 años de edad media, un 77% adjuntos. Conocimientos: el porcentaje medio de respuestas correctas por participante al inicio fue del 44,5% (DE = 14,1%) y al final del 69,3% (DE = 16,8%; p < 0,0001). Resolución de casos: el porcentaje medio de aciertos pasó del 73,2% (DE = 9,8%) al inicio al 85,6% (DE = 7%) al final (p < 0,0001). Satisfacción: el grado de satisfacción medio fue de 3,5 puntos sobre 4 (DE = 0,12), equivalente al 87,2% (DE = 3,1%). La pregunta mejor valorada fue ‘El nivel de conocimientos sobre el tema impartido por los ponentes fue bueno’: 3,7; y la menos valorada fue ‘Las condiciones del local y el material audiovisual me parecieron idóneos’: 3,3. Conclusiones: El curso ha conseguido mejorar los conocimientos y las habilidades de los alumnos, lo que, unido a la satisfacción obtenida, puede tener influencia efectiva en su ejercicio profesional.(AU)

Introduction and objective: To evaluate the improvement of students’ knowledge, skills, and satisfaction in a course on dispensation service.Material and methods: Design: quasi-experimental study (before/after) with educational intervention. Participants: Pontevedra community pharmacists. Seven editions (October 2018-October 2019). Educational intervention: hybrid-learning course 26-hour e-learning + 6-hour workshop with case simulations and training by role-playing. Assessment: the first two levels of Kirkpatrik were evaluated: reaction (satisfaction) and competence (knowledge/clinical performance). Questionnaire with 20 questions and 4 case studies. A Likert-type questionnaire with 12 questions was used to assess satisfaction. Qualitative data are expressed as percentages and quantitative data are expressed as average (SD). Results: 114 students, 78% female, 39 years (average age), 77% relief pharmacists. Knowledge: average percentage of correct questionnaire answers per participant at the beginning 44.5% (SD-14.1%), at the end 69.3% (SD-16.8%), (p < 0,0001). Case-study results: average percentage of correct answers increased from 73.2% (SD-9.8%) at the beginning to 85.6% (SD-7.0%) at the end (p < 0.0001). Satisfaction: average satisfaction rating of 3.5/4 (SD-0.12); equivalent to 87.2% (SD-3.1%). Best-rated question: ‘The knowledge level of the subject taught by the speakers was good’: 3.7. Less valued: ‘The conditions of the premises and audiovisual material seemed suitable to me’: 3.3. Conclusions: The course has managed to improve the knowledge and skills of the students, which, together with the satisfaction obtained, can have an effective influence on their professional exercise.(AU)

Percepción de los usuarios de la farmacia comunitaria sobre la COVID-19 / Community pharmacy users' perception of COVID-19

OBJETIVOS: evaluar la percepción de los usuarios de las farmacias sobre la repercusión de la pandemia COVID-19 en su salud, el conocimiento sobre su posible inclusión en grupos de riesgo y su actitud ante una posible vacuna. MATERIAL Y MÉTODOS: estudio observacional transversal aleatorizado, en farmacias de Pontevedra y Ourense, desde marzo a junio de 2020 al comienzo y después de la restricción de movimientos. Sujetos: usuarios ≥ 18 años que acuden a las farmacias participantes en demanda de medicamentos y/o material de higiene y protección. Procedimiento: el usuario cumplimentaba un cuestionario anónimo, depositándolo hasta la noche en una bandeja de plástico desinfectada diariamente. El procedimiento se repetirá tras cesar el confinamiento. Resultados de la primera fase: se realizaron 706 encuestas. 415 (58,8 %) mujeres, edad media 48,9 (DE=16,9) años. 100 participantes (14,2 %) viven solos.637 (90,2 %) creen que la COVID-19 es más peligrosa que la gripe, 189 (26,8 %) desconocen estar en algún grupo de riesgo. 107 (15,1 %) dicen sentirse mal o muy mal con el aislamiento. Los aspectos más afectados son: familiar 350 (49,6 %) y emocional 338 (47,9 %). En 2019/20 se vacunaron de la gripe 172 (24,4 %) y en 2020/21 piensan vacunarse 243 (34,4 %). 448 (63,5 %) se vacunarán frente a la COVID-19 cuando exista vacuna, esté o no financiada por el Sistema Nacional de Salud, 183 (25,9 %) se lo pensarán. CONCLUSIONES: los encuestados consideran la COVID-19 más peligrosa que la gripe estacional. En esta primera fase la afectación sobre bienestar y salud no parece elevada. Un alto porcentaje se vacunará frente a la COVID-19

AIMS: to assess the pharmacy users' perception on the impact of the COVID-19 pandemic on their health, the knowledge of their possible inclusion in risk groups and their attitude towards a potential vaccine. MATERIAL AND METHODS: Randomized, cross-sectional, observational study in pharmacies of Pontevedra and Ourense, from March to June 2020 at the beginning and after movement restrictions. Subjects: users ≥ 18 years old who go to the participating pharmacies to purchase medicines and/or hygiene and protection material. Procedure: the user completed an anonymous questionnaire, which was then placed until night in a plastic tray that was disinfected daily. The procedure will be repeated after the confinement has been terminated. Results of the first phase: 706 surveys were conducted. 415 (58.8%) women, aged 48.9 9 years on average (SD=16.9). 100 participants (14.2%) live alone. 637 (90.2%) believe that COVID-19 is more dangerous than influenza, 189 (26.8%) don't know if they belong to any risk group. 107 (15.1%) say they feel bad or very bad about isolation. The most affected aspects are: familiar 350 (49.6%) and emotional 338 (47.9%). In 2019/20, 172 (24.4%) were vaccinated against influenza and in 2020/21 243 (34.4%) are planning to get vaccinated. 448 (63.5%) will be vaccinated against COVID-19 when the vaccine is available, whether or not it is financed by the National Health System, and 183 (25.9%) will think about it. CONCLUSIONS: respondents consider COVID-19 more dangerous than seasonal influenza. In this first phase, the impact on well-being and health does not seem to be high. A high percentage of people will be vaccinated against COVID-19

Preferencias de formación de los farmacéuticos comunitarios de la provincia de Pontevedra / Training preferences of the community pharmacics of the province of Pontevedra (Spain)

OBJETIVOS: Conocer las preferencias de los farmacéuticos comunitarios (FC) de Pontevedra en relación con la formación continuada, qué materias despiertan mayor interés y posibles relaciones entre preferencias y características demográficas. MÉTODOS: Estudio observacional transversal realizado en diciembre 2017 y enero 2018. Participaron farmacéuticos colegiados en Pontevedra ejercientes en farmacias comunitarias. El tamaño muestral se calculó para una precisión del 10,0%, un intervalo de confianza para poblaciones finitas al 95% bilateral, y una población de 495 titulares y 768 adjuntos, en 72 titulares y 75 adjuntos. Se elaboró una encuesta que se pilotó con seis FC, se hicieron las modificaciones resultantes del pilotaje y se difundió a todos los FC colegiados, que la cumplimentaron en la web del Colegio o impresa. RESULTADOS: Se recibieron 261 cuestionarios (77 titulares, 29,5% y 184, 70,5% adjuntos), correspondientes al 20,7% de los colegiados ejercientes en farmacia comunitaria. El 62,5% cree que no tiene suficiente formación. Las materias formativas más demandadas fueron Farmacología/Farmacoterapia, 206 FC (78,9%), Nutrición, 175 (67,0%) y Servicios Profesionales Farmacéuticos, 161 (61,7). La modalidad semipresencial (contenidos teóricos online y talleres presenciales) (48,3%), frecuencia trimestral (47,9%), duración de 20-30 horas/año (48,7%), y obligatoriedad de un mínimo de créditos anuales (46,0%) son las opciones preferidas. Para el 69,0% la duración de los talleres presenciales debe ser entre 1 y 4 horas. La organización de la formación continuada debe estar a cargo del Colegio. CONCLUSIONES: El estudio permitirá elaborar programas formativos dirigidos a los colegiados ejercientes en farmacias comunitarias mejor adaptados a sus necesidades y preferencias

OBJECTIVES: To know the preferences of the community pharmacists (CP) of Pontevedra in relation to continued education, which subjects arouse the greatest interest and possible relationships between preferences and demographic characteristics. METHODS: Cross-sectional observational study in December 2017 and January 2018. Collegiate community pharmacists participated in Pontevedra. The sample size was calculated for an accuracy of 10.0%, a confidence interval for finite populations at 95% bilateral and a population of 495 holders and 768 attached, in 72 holders and 75 attached. A survey was prepared and piloted with six CPs, the modifications resulting from the piloting were made and disseminated to all collegiate CPs, who completed it on the College website or printed. Outcomes: 261 questionnaires (77 holders, 29.5% and 184, 70.5% attached) were received, corresponding to 20.7% of the collegiate pharmacists working in a community pharmacy. 62.5% think they don't have enough training. The most demanded training subjects were Pharmacology/Pharmacotherapy, 206 CP (78.9%), Nutrition, 175 (67.0%) and Professional Pharmaceutical Services, 161 (61.7). Semi-presencial modality (online theoretical content and face-to-face workshops) (48.3%), quarterly frequency (47.9%), duration of 20-30 hours/year (48.7%), and mandatory minimum annual credits (46.0%) are the preferred options. For 69.0% the duration of the face-to-face workshops should. be between 1 and 4 hours. The organization of the countinuated education must be in charge of the College. Conclussions: The study will allow to develop training programs targeted to the CPs who are members of the College and practice in community pharmacies. These programs will be better adapted to their needs and preferences

Evaluación de la no adherencia al tratamiento hipoglucemiante en la farmacia comunitaria / Evaluation of non-adherence to hipoglucemiary treatment in community pharmacy

Introducción: La adherencia al tratamiento en enfermedades crónicas como la diabetes mellitus (DM) es de vital importancia. El incumplimiento está directamente implicado en el aumento del número de medicamentos prescritos en enfermos crónicos y es una de las causas más importantes de fracaso de los tratamientos. Objetivos: Evaluar en la farmacia comunitaria la prevalencia de no adherencia al tratamiento en DM tipo 2 mediante la realización del cuestionario MMAS-8. Material y métodos: Estudio observacional transversal realizado en mayo y junio de 2018. Se seleccionaron pacientes mayores de 45 años, en tratamiento con hipoglucemiantes. Se midió la adherencia farmacoterapéutica mediante la administración del cuestionario (MMAS-8) según la puntuación obtenida en el test y la percepción que tienen sobre su tratamiento. Resultados: Participaron 64 pacientes, 29 (45,3%) mujeres y 35 (54,7) hombres. Las mujeres tenían 65,5 años de edad media (DE=15,3) y los hombres 65,6 años (DE=12,3). La puntuación media del test de MMAS-8 fue de 6,2±2,1, no encontrando diferencias significativas entre sexos. Se encontró que el número de pacientes con baja adherencia fueron 21 (32,8%), con media adherencia 19 (29,7%) y con alta adherencia 24 (37,5%). No se encontró relación entre el porcentaje de adherencia al tratamiento con el sexo, edad, nivel de estudios y condición laboral. La creencia de que el tratamiento mejora la enfermedad y que los medicamentos prescritos son los adecuados aumenta la adherencia. Conclusiones: El cuestionario MMAS-8 se ha mostrado como una herramienta rápida y sencilla para la evaluación de la adherencia del tratamiento en la farmacia comunitaria

Background: The treatment adherence in chronic diseases such as diabetes mellitus is of vital importance. Noncompliance is directly implicated in the increase of the prescribed drugs number in chronic patients and is one of the most important causes of treatment failure. Objectives: To evaluate in the community pharmacy the prevalence of non-adherence to treatment in type 2 diabetes mellitus by completing the MMAS-8 questionnaire. Methods: Cross-sectional observational study conducted in May and June 2018. Patients older than 45 years were selected, in treatment with hypoglycemic agents. Pharmacotherapeutic adherence was measured by administering the questionnaire (MMAS-8) according to the score obtained in the test and the perception they have of their treatment. Results: 64 patients participated, 29 (45.3%) women and 35 (54.7) men. Women ware 65.5 years of age (SD = 15.3) and men 65.6 years (SD = 12.3). The mean score of the MMAS-8 test was 6.2±2.1, not finding significant differences between sexes. It was found that the number of patients with low adherence was 21 (32.8%), with medium adherence 19 (29.7%) and with high adherence 24 (37.5%). No relation was found between the percentage of adherence to treatment with sex, age, level of education and work condition. The belief that the treatment improves the disease and that the prescribed medications are adequate, increases adherence. Conclusions: The MMAS-8 questionnaire has been shown as a quick and simple tool for evaluating the adherence of the treatment in the community pharmacy