Flotillin-mediated stabilization of unfolded proteins in bacterial membrane microdomains.

The function of many bacterial processes depends on the formation of functional membrane microdomains (FMMs), which resemble the lipid rafts of eukaryotic cells. However, the mechanism and the biological function of these membrane microdomains remain unclear. Here, we show that FMMs in the pathogen methicillin-resistant Staphylococcus aureus (MRSA) are dedicated to confining and stabilizing proteins unfolded due to cellular stress. The FMM scaffold protein flotillin forms a clamp-shaped oligomer that holds unfolded proteins, stabilizing them and favoring their correct folding. This process does not impose a direct energy cost on the cell and is crucial to survival of ATP-depleted bacteria, and thus to pathogenesis. Consequently, FMM disassembling causes the accumulation of unfolded proteins, which compromise MRSA viability during infection and cause penicillin re-sensitization due to PBP2a unfolding. Thus, our results indicate that FMMs mediate ATP-independent stabilization of unfolded proteins, which is essential for bacterial viability during infection.

Postbiotics of Lacticaseibacillus paracasei CECT 9610 and Lactiplantibacillus plantarum CECT 9608 attenuates store-operated calcium entry and FAK phosphorylation in colorectal cancer cells.

Store-operated Ca2+ entry (SOCE) is a major mechanism for Ca2+ influx in colorectal cancer (CRC) cells. This mechanism, regulated by the filling state of the intracellular Ca2+ stores, is mediated by the endoplasmic reticulum Ca2+ sensors of the stromal interaction molecules (STIM) family [stromal interaction molecule 1 (STIM1) and STIM2] and the Ca2+-release-activated Ca2+ channels constituted by Orai family members, with predominance of calcium release-activated calcium channel protein 1 (Orai1). CRC cells exhibit enhanced SOCE due to remodeling of the expression of the key SOCE molecular components. The enhanced SOCE supports a variety of cancer hallmarks. Here, we show that treatment of the colorectal adenocarcinoma cell lines HT-29 and Caco-2 with inanimate Lacticaseibacillus paracasei (CECT9610) and Lactiplantibacillus plantarum (CECT9608) attenuates SOCE, although no detectable effect is seen on SOCE in normal colon mucosa cells. The effect of Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics was mediated by downregulation of Orai1 and STIM1, while the expression levels of Orai3 and STIM2 remained unaltered. Treatment of HT-29 and Caco-2 cells with inanimate Lacticaseibacillus paracasei and Lactiplantibacillus plantarum impairs in vitro migration by a mechanism likely involving attenuation of focal adhesion kinase (FAK) tyrosine phosphorylation. Cell treatment with the Orai1 inhibitor synta-66 attenuates SOCE and prevents any further effect of Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics. Together, our results indicate for the first time that Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics selectively exert negative effects on Ca2+ influx through SOCE in colorectal adenocarcinoma cell lines, providing evidence for an attractive strategy against CRC.

Imperfect match between radiation exposure times required for conidial viability loss and infective capacity reduction attenuate UV-B impact on Beauveria bassiana.

BACKGROUND: UV-B radiation represents a significant challenge for the widespread use of entomopathogenic fungi in pest management. This study focused on research of the asynchronous response between virulence and conidial viability against Ceratitis capitata adults using specific statistical models. Moreover, it was also investigated whether the observed differences in susceptibility to UV-B radiation in in vitro assays among three selected isolates of Beauveria bassiana were reflected in the above-mentioned asynchrony. RESULTS: While the irradiation of the three isolates of B. bassiana was associated with a significant loss of conidial viability, their virulence was not significantly affected compared to nonirradiated treatments when exposed to 1200 mW m-2 for 6 h before or after the inoculation of C. capitata. In fact, the irradiation time needed to reduce the mortality to 50% compared to the controls was 34.69 h for EABb 10/225-Fil, 16.36 h for EABb 09/20-Fil, and 24.59 h for EABb 09/28-Fil. Meanwhile, the irradiation time necessary to reduce conidial viability to 50% was 9.89 h for EABb 10/225-Fil, 8.74 h for EABb 09/20-Fil, and 4.71 h for EABb 09/28-Fil. CONCLUSION: These results highlight the importance of modeling the response of entomopathogenic fungi virulence and conidial susceptibility when exposed to UV-B radiation for the selection of environmentally competent isolates, regardless of the results obtained in previous in vitro assays on conidial germination. This strategic approach is critical in overcoming the challenges posed by UV-B radiation and holds the key to realizing the full potential of entomopathogenic fungi in pest management. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Unlocking New Avenues in Breast Cancer Treatment: The Synergy of Kinase Inhibitors and Immunotherapy.

Cancer is one of the world's most significant health problems today. Currently, breast cancer has globally surpassed lung cancer as the most commonly diagnosed cancer in women. In 2020, an estimated 2,261,419 new cases were diagnosed in women worldwide. Therefore, there is a need to understand the processes that can help us better treat this disease. In recent years, research in the fight against cancer has often been based on two treatment modalities. One of them is the use of protein kinase inhibitors, which have been instrumental in the development of new therapeutic strategies. Another crucial route is the use of immunotherapy, which has been touted as a great promise for cancer treatment. Protein kinase alterations can interfere with the effectiveness of other treatments, such as immunotherapy. In this review, we will analyze the role played by protein kinase alterations in breast cancer and their possible impact on the effectiveness of the response to immunotherapy treatments.

FACTORES PROMOTORES, DIFICULTADES Y ESTRATEGIAS PARA EMPRENDER Y CONTINUAR LA LACTANCIA MATERNA EN ESPAÑA: LA VOZ DE LAS MUJERES / FACTORS THAT PROMOTE, HINDER, AND SUPPORT PROMOTION, HINDERING AND STRATEGIC FACTORS TO BREASTFEEDING IN SPAIN: THE VOICE OF WOMEN

OBJETIVO: Identificar factores promotores, dificultades y estrategias para el inicio y continuación de la lactancia materna (LM), en base a la experiencia de mujeres lactantes. METODOLOGÍA: Estudio cualitativo fenomenológico realizado en la Región de Murcia (España) en el 2019. Se reclutaron a veintisiete mujeres que alimentaron a sus hijos/as con LM en el periodo de 2012-2018 mediante una asociación de LM y un muestreo de bola de nieve. Se realizaron entrevistas personales abiertas y un análisis temático de las transcripciones. RESULTADOS: Se estructuraron en tres temas: 1) Facilidades para el inicio y mantenimiento de la LM, 2) Dificultades ante la LM y 3) Estrategias utilizadas por las mujeres para solventar las barreras. Mientras que la motivación para amamantar y el apoyo recibido facilitaron la LM, las barreras más comunes se relacionaron con miedos e inseguridades personales, cansancio, problemas físicos y la respuesta social frente a la LM. Los motivos de abandono fueron el déficit de producción de leche, la incorporación laboral y la actitud del bebé. Las participantes desarrollaron autocuidados, búsqueda de información y asociaciones de LM y estrategias de conciliación familiar-laboral para paliar las dificultades. CONCLUSIÓN: Es necesaria una mayor implicación por parte de las/os profesionales de la salud para apoyar a las mujeres a través de la herramienta de educación para la salud, el apoyo por pares y medidas institucionales que favorezcan la LM en el lugar de trabajo.

OBJECTIVE: To identify promoting factors, difficulties, and strategies related to the initiation and continuation of breastfeeding (BF), based on the experiences of breastfeeding women. METHODOLOGY: A phenomenological qualitative study conducted in the Region of Murcia (Spain) in 2019. Twenty-seven women who breastfed their children from 2012 to 2018 were recruited through a breastfeeding association and snowball sampling. Personal open interviews were conducted, and a thematic analysis of the transcriptions was performed. RESULTS: The findings were organized into three themes: 1) Facilitators for the initiation and maintenance of BF, 2) Difficulties faced during BF, and 3) Strategies employed by women to overcome the difficulties. While the motivation to breastfeed and provision of support facilitated BF, common challenges were noted, including personal fears and insecurities, fatigue, physical problems, and societal response to BF. Reasons for discontinuation included low milk production, the need to return to work, and the baby's attitude. Participants developed self-care practices, sought information, joined breastfeeding associations, and implemented work-family reconciliation strategies to address the challenges. CONCLUSION: Greater involvement from healthcare professionals is necessary to support women who breastfeed through health education, peer support, and institutional measures that promote BF in the workplace.

Assessment of high fidelity simulation with actors in palliative care in nursing students: a mixed methods study.

OBJECTIVE: The most effective training methods are experiential, including those focused on experiences and emotions. Clinical simulation, especially high-fidelity simulation, is one of the most effective methodologies for the acquisition of competencies in care like palliative care. The simulation with actors can train future healthcare science professionals: in technical, intellectual, or interpersonal skills. The objective is to evaluate high-fidelity simulation with actors as a tool in palliative care training for nursing students. METHOD: Over three years, the study was conducted in a Faculty of Nursing of the south of Spain with nursing students. A mixed methods study with sequential explanatory design in three moments was conducted: (1) Quasi-experimental study in a single group (n = 12) before and after attending the palliative care course with Clinical Simulation with actors to assess the communication skills (CICAA scale), (2) Qualitative study with phenomenological perspective after Clinical Simulation (174 reflective students' narratives), (3) Cross-sectional observational study, one year later, to assess the transfer of knowledge and skills to the clinical practice (71 students). RESULTS: Students who interacted with actors in Clinical Simulation improved their communication skills and the ability to establish an effective helping relationship with both end-of-life patients and their families. The students perceived the Clinical Simulation as an innovative learning methodology that is useful to encourage reflection and transfer of learning during their clinical internship. CONCLUSIONS: Standardization of the use of active learning methodologies is recommended for a better acquisition of transversal skills such as communication skills in palliative care.

Immunomodulatory effects of inactivated Ligilactobacillus salivarius CECT 9609 on respiratory epithelial cells.

The microbiota in humans and animals play crucial roles in defense against pathogens and offer a promising natural source for immunomodulatory products. However, the development of physiologically relevant model systems and protocols for testing such products remains challenging. In this study, we present an experimental condition where various natural products derived from the registered lactic acid bacteria Ligilactobacillus salivarius CECT 9609, known for their immunomodulatory activity, were tested. These products included live and inactivated bacteria, as well as fermentation products at different concentrations and culture times. Using our established model system, we observed no morphological changes in the airway epithelium upon exposure to Pasteurella multocida, a common respiratory pathogen. However, early molecular changes associated with the innate immune response were detected through transcript analysis. By employing diverse methodologies ranging from microscopy to next-generation sequencing (NGS), we characterized the interaction of these natural products with the airway epithelium and their potential beneficial effects in the presence of P. multocida infection. In particular, our discovery highlights that among all Ligilactobacillus salivarius CECT 9609 products tested, only inactivated cells preserve the conformation and morphology of respiratory epithelial cells, while also reversing or altering the natural immune responses triggered by Pasteurella multocida. These findings lay the groundwork for further exploration into the protective role of these bacteria and their derivatives.

Impact of Hospitalization of People With Dementia or Cognitive Impairment on Family Caregivers.

People with dementia (PWD) have a higher risk of hospitalization than people without dementia. Hospitalizations are stressful events for PWD and their caregivers, representing a considerable change to their routines. The current descriptive longitudinal study aimed to identify the positive and negative reactions, experiences related to health and social integrated care, resource use, and work status of family caregivers of PWD or cognitive impairment admitted to the hospital with a proximal femur fracture undergoing surgery. Findings indicated that family caregivers (N = 174) are fully committed to providing assistance in activities of daily living and supervision, showing positive attitudes on self-esteem and negative attitudes toward lack of family support and impact on finances, schedule, and health. Overall caregiver experiences with integrated health and social care improved after hospitalization but decreased after discharge. One month after hospitalization, family caregivers maintained the same work hours but used fewer health care resources. Hospitalization represents a good opportunity to approach family caregivers and determine their needs to provide them with interventions to minimize their burden and improve their well-being. [Research in Gerontological Nursing, 16(6), 283-290.].

Soluble mutant huntingtin drives early human pathogenesis in Huntington's disease.

Huntington's disease (HD) is an incurable inherited brain disorder characterised by massive degeneration of striatal neurons, which correlates with abnormal accumulation of misfolded mutant huntingtin (mHTT) protein. Research on HD has been hampered by the inability to study early dysfunction and progressive degeneration of human striatal neurons in vivo. To investigate human pathogenesis in a physiologically relevant context, we transplanted human pluripotent stem cell-derived neural progenitor cells (hNPCs) from control and HD patients into the striatum of new-born mice. Most hNPCs differentiated into striatal neurons that projected to their target areas and established synaptic connexions within the host basal ganglia circuitry. Remarkably, HD human striatal neurons first developed soluble forms of mHTT, which primarily targeted endoplasmic reticulum, mitochondria and nuclear membrane to cause structural alterations. Furthermore, HD human cells secreted extracellular vesicles containing mHTT monomers and oligomers, which were internalised by non-mutated mouse striatal neurons triggering cell death. We conclude that interaction of mHTT soluble forms with key cellular organelles initially drives disease progression in HD patients and their transmission through exosomes contributes to spread the disease in a non-cell autonomous manner.

Label-Free Identification of Carbonaceous Particles Using Nonlinear Optical Microscopy.

The adverse health effects of ambient carbonaceous particles (CPs) such as carbon black (CB), black carbon (BC), and brown carbon (BrC) are becoming more evident and depend on their composition and emission source. Therefore, identifying and quantifying these particles in biological samples are important to better understand their toxicity. Here, we report the development of a nonlinear optical approach for the identification of CPs such as CB and BrC using imaging conditions compatible with biomedical samples. The unique visible light fingerprint of CB and BrC nanoparticles (NPs) upon illumination with a femtosecond (fs) pulsed laser at 1300 nm excitation wavelength is an effective approach for their identification in their biological context. The emission from spectral features of these CPs was investigated with time-domain fluorescence lifetime imaging (FLIM) to further support their identification. This study is performed for different types of CPs embedded in agarose gel as well as in in vitro mammalian cells. The unique nonlinear emissive behavior of CP NPs used for their label-free identification is further complementary with fluorophores typically used for specific staining of biological samples thus providing the relevant bio-context.

"Proteotranscriptomic analysis of advanced colorectal cancer patient derived organoids for drug sensitivity prediction".

BACKGROUND: Patient-derived organoids (PDOs) from advanced colorectal cancer (CRC) patients could be a key platform to predict drug response and discover new biomarkers. We aimed to integrate PDO drug response with multi-omics characterization beyond genomics. METHODS: We generated 29 PDO lines from 22 advanced CRC patients and provided a morphologic, genomic, and transcriptomic characterization. We performed drug sensitivity assays with a panel of both standard and non-standard agents in five long-term cultures, and integrated drug response with a baseline proteomic and transcriptomic characterization by SWATH-MS and RNA-seq analysis, respectively. RESULTS: PDOs were successfully generated from heavily pre-treated patients, including a paired model of advanced MSI high CRC deriving from pre- and post-chemotherapy liver metastasis. Our PDOs faithfully reproduced genomic and phenotypic features of original tissue. Drug panel testing identified differential response among PDOs, particularly to oxaliplatin and palbociclib. Proteotranscriptomic analyses revealed that oxaliplatin non-responder PDOs present enrichment of the t-RNA aminoacylation process and showed a shift towards oxidative phosphorylation pathway dependence, while an exceptional response to palbociclib was detected in a PDO with activation of MYC and enrichment of chaperonin T-complex protein Ring Complex (TRiC), involved in proteome integrity. Proteotranscriptomic data fusion confirmed these results within a highly integrated network of functional processes involved in differential response to drugs. CONCLUSIONS: Our strategy of integrating PDOs drug sensitivity with SWATH-mass spectrometry and RNA-seq allowed us to identify different baseline proteins and gene expression profiles with the potential to predict treatment response/resistance and to help in the development of effective and personalized cancer therapeutics.

Phylogenetic origin of Helicobacter pylori pathogenicity island and risk of stomach cancer and high-grade premalignant gastric lesions.

Infection by Helicobacter pylori (Hp) has been causally linked to risk of gastric cancer (GC). The coevolution of Hp and humans shaped the risk of GC as our species left Africa and migrated to the other continents. Latin America (LatAm) is a high GC incidence region where Hp evolved uniquely in the 500 years since European colonization. Differential virulence of the Hp cagA -pathogenicity island (cagPAI) by ancestral origin has been reported. We hypothesized that Hp phylogenetic origin might play a role in determining GC risk in LatAm. We used genotypes of 50 Hp genetic variants mapping to the Hp cagPAI, studied in 1220 subjects from Venezuela, Colombia, Mexico and Paraguay, who were infected with cagA-positive Hp, including 150 GC, 177 high-grade premalignant lesions (HGPMLs) and 893 low-grade premalignant lesions. We estimated the phylogenetic origin of Hp cagPAI in all study subjects by use of the STRUCTURE software and principal component analysis (PCA) and tested whether the estimated African ancestry percentage was associated with the risk of GC or HGPML. African ancestral component estimates by STRUCTURE and PCA were highly correlated. STRUCTURE-based African origin estimate was not significantly associated with the risk of HGPML, but it was inversely associated with GC risk: the OR associated with the continuous values of African component was 0.09 (95% CI, 0.01-0.85; P = 0.035). Similar trends were observed for GC with PCA-based estimates, but the association was not statistically significant. These results suggest that Hp ancestral origin may play a role in gastric carcinogenesis.

Palbociclib in advanced stage hormone receptor-positive breast cancer: real-world data from a Chilean multicentre registry.

Background: The addition of cyclin-dependent kinases inhibitors (CDKi) to endocrine therapy (ET) as the first- or second line treatment improves progression-free and overall survival (OS) in hormone receptor-positive, HER2 negative (HR+/HER2-) advanced stage breast cancer (ABC). Our study compared survival rates and prognostic factors in Chilean patients that used palbociclib as first or subsequent (≥second) lines of treatment in a real-world setting. Methods: Our retrospective population-cohort study included HR+/HER2- ABC patients. We calculated 5-year OS and performed a multivariate analysis to determine prognostic factors. Results: A total of 106 patients were included. Median age was 49 years (19-86), 28.3% (30) had de novo stage IV disease; 63% received palbociclib with ET as first line, 54% of them with aromatase inhibitor over fulvestrant. Median OS for the entire cohort was 99 months and 5-year OS was 69%. Patients that received first line palbociclib had a 5-year OS of 89% versus 43% for ET monotherapy or ≥second line palbociclib (p = 0.0062). Multivariate analysis showed that the year at diagnosis and CDKi timing (first line versus ≥second line) were significantly associated with OS. Conclusion: Our real-world data show that first-line CDKi + ET provides a statistically significant benefit in OS versus ≥second line in HR+/HER2- ABC patients.

"This looks like a movie": a case report of post-surgical amnesia.

A 52-year-old male patient with a background of adaptive personality disorder was admitted for mitral valve repair and cardiac ablation for atrial fibrillation. He suffered intraoperative complications with severe mitral insufficiency that suffered ischemia.. Post-operatively, he demonstrated acute loss of retrograde autobiographical memory, prosopagnosia and a loss of public semantic memory. His CT scan was normal and MRI was not possible due to intra-cardiac leads. An initial diagnosis of hypoxic-ischemic encephalopathy was considered. A neuropsychological examination undertaken 20 days after his surgery showed a severe alteration of retrograde autobiographical memory, marked alteration of semantic knowledge and prosopagnosia. He demonstrated an average performance in tasks measuring constructional praxis, visuospatial ability, and executive functions. 34 days after surgery, and after a short nap, the patient "returns" to the day before admission and consequently recovers his memory. Repeat neuropsychological assessment demonstrated performance within the normal range across all previously tested domains. This sudden recovery of memory, together with a normal MRI, led to a rethinking of the diagnosis of dissociative amnesia. This case illustrates the long-standing discussion about the organic or functional origin of some memory disorders, in which, despite advances in neuroimaging techniques, it is still difficult to know their etiology .

Intención de aceptación a la vacunación contra la COVID-19 en sectores vulnerables del Ecuador

Introducción: La COVID-19 causó que varios sectores profesionales hayan tenido que enfrentarlo en primera línea, viéndose afectados ante la vulnerabilidad de contraer el virus. A pesar de la baja tasa de mortalidad de los momentos actuales y la poca saturación de pacientes con COVID-19 en los centros de salud, la aplicación de una cuarta dosis de inoculación ha generado posturas diferentes entre varios países. Objetivo: Determinar si el personal considerado con altos riesgos de vulnerabilidad de la ciudad de Santo Domingo de los Colorados, en Ecuador, tiene intenciones favorables para la aplicación de la cuarta dosis de la vacuna contra la COVID-19. Método: Se desarrolló un estudio cuantitativo de alcance correlacional y diseño transversal. Un cuestionario conformado por 16 preguntas midió las variables: riesgo de contagio, conocimiento percibido sobre la vacuna, confianza sobre la vacuna e intención de vacunarse; el cual fue aplicado a 375 participantes. Los análisis estadísticos fueron desarrollados a través de Excel y Statistical Package for Social Sciences 21 (SPSS 21). Resultados: Los análisis estadísticos evidenciaron que el riesgo de contagio (β=0,178**), el conocimiento percibido sobre la vacuna (β=0,218**) y la confianza sobre la vacuna (β=0,192**) se correlacionan significativamente con la intención de vacunarse, ante lo cual se evidencia la necesidad de recibir una cuarta dosis de inoculación por parte de los sectores vulnerables. Conclusiones: Esta es la primera investigación que expone resultados respecto a la intención de vacunación en las personas vulnerables y pone en evidencia la intención de acceder a una cuarta dosis de inoculación.

Introduction: COVID-19 caused healthcare professional workers have faced the pandemic on the frontline at the risk of being infected with the virus. Despite the low mortality rate at present and the low presence of patients with COVID-19 in health care centers, the application of a fourth booster dose has generated different positions among several countries. Objective: To determine whether personnel considered being at high risk of vulnerability in the city of Santo Domingo de los Colorados, Ecuador, have favorable intentions for receiving the fourth booster dose of the COVID-19 vaccine. Method: A quantitative study of correlational scope and cross-sectional design was developed. A questionnaire consisting of 16 questions measured the following variables: risk of infection, perceived knowledge of the vaccine, confidence in the vaccine and intention to be vaccinated; this questionnaire was applied to 375 participants. Statistical analyses were developed using the microsoft Excel spreadsheed and Statistical Packagefor Social Sciences 21 (SPSS 21). Results: Statistical analyses showed that the risk of infection (β=0.178**), perceived knowledge about the vaccine (β=0.218**) and confidence about the vaccine (β=0.192**) are significantly correlated with the intention to be fully vaccinated, thus showing the need for a fourth booster dose by vulnerable sectors. Conclusion: This is the first research that presents results regarding the intention to vaccinate vulnerable people and highlights the intention to access a fourth booster dose.

Introdução: O COVID-19 fez com que diversos setores profissionais o enfrentassem na linha de frente, sendo afetados pela vulnerabilidade de contrair o vírus. Apesar da baixa taxa de mortalidade atual e da baixa saturação de pacientes com COVID-19 nos centros de saúde, a aplicação de uma quarta dose de inoculação gerou posições diferentes entre vários países. Objetivo: Determinar se o pessoal considerado de alto risco de vulnerabilidade na cidade de Santo Domingo de los Colorados, no Equador, tem intenções favoráveis para a aplicação da quarta dose da vacina contra COVID-19. Método: Foi desenvolvido um estudo quantitativo com escopo correlacional e delineamento transversal. Um questionário composto por 16 questões mediu as variáveis: risco de contágio, conhecimento percebido sobre a vacina, confiança sobre a vacina e intenção de se vacinar; que foi aplicado a 375 participantes. As análises estatísticas foram realizadas no Excel e no Statistical Package for the Social Sciences 21 (SPSS 21). Resultados: As análises estatísticas mostraram que o risco de contágio (β=0,178**), o conhecimento percebido sobre a vacina (β=0,218**) e a confiança sobre a vacina (β=0,192**) estão significativamente correlacionados com a intenção de ser vacinado, o que evidencia a necessidade de receber uma quarta dose de inoculação por setores vulneráveis. Conclusões: Esta é a primeira pesquisa que expõe resultados sobre a intenção de vacinação em pessoas vulneráveis e evidencia a intenção de acessar uma quarta dose de inoculação.

Lack of EGFR catalytic activity in hepatocytes improves liver regeneration following DDC-induced cholestatic injury by promoting a pro-restorative inflammatory response.

Despite the well-known hepatoprotective role of the epidermal growth factor receptor (EGFR) pathway upon acute damage, its specific actions during chronic liver disease, particularly cholestatic injury, remain ambiguous and unresolved. Here, we analyzed the consequences of inactivating EGFR signaling in the liver on the regenerative response following cholestatic injury. For that, transgenic mice overexpressing a dominant negative mutant human EGFR lacking tyrosine kinase activity (ΔEGFR) in albumin-positive cells were submitted to liver damage induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC), an experimental model resembling human primary sclerosing cholangitis. Our results show an early activation of EGFR after 1-2 days of a DDC-supplemented diet, followed by a signaling switch-off. Furthermore, ΔEGFR mice showed less liver damage and a more efficient regeneration following DDC injury. Analysis of the mechanisms driving this effect revealed an enhanced activation of mitogenic/survival signals, AKT and ERK1/2-MAPKs, and changes in cell turnover consistent with a quicker resolution of damage in response to DDC. These changes were concomitant with profound differences in the profile of intrahepatic immune cells, consisting of a shift in the M1/M2 balance towards M2 polarity, and the Cd4/Cd8 ratio in favor of Cd4 lymphocytes, overall supporting an immune cell switch into a pro-restorative phenotype. Interestingly, ΔEGFR livers also displayed an amplified ductular reaction, with increased expression of EPCAM and an increased number of CK19-positive ductular structures in portal areas, demonstrating an overexpansion of ductular progenitor cells. In summary, our work supports the notion that hepatocyte-specific EGFR activity acts as a key player in the crosstalk between parenchymal and non-parenchymal hepatic cells, promoting the pro-inflammatory response activated during cholestatic injury and therefore contributing to the pathogenesis of cholestatic liver disease. © 2022 The Pathological Society of Great Britain and Ireland.

Validez diagnóstica de cinco escalas de predicción clínica para la trombosis venosa profunda (TVP)

Introducción: la trombosis venosa profunda (TVP) es una entidad común que afecta principalmente el sistema venoso profundo de los miembros inferiores, para el cual se han desarrollado múltiples escalas de predicción clínica, las cuales han sido construidas y validadas en pacientes ambulatorios y hospitalizados. Objetivos: validar cinco escalas de predicción clínica para TVP en pacientes atendidos en un centro de tercer nivel en la sabana de Bogotá, Colombia. Métodos: se llevó a cabo un estudio de corte transversal con análisis de prueba diagnóstica en sujetos con sospecha de TVP, incluyendo aquellos que contaran con la realización de ecografía Doppler venosa de miembros inferiores. Se calculó el rendimiento de cinco escalas de predicción clínica para TVP (Wells clásico y modificado, Oudega, CEBI y Constans) para pacientes ambulatorios u hospitalizados, individualizando la población en la que fueron validadas. Resultados: ingresaron al análisis 974 pacientes, de estos 485 (49,7 %) presentaron TVP. La escala de Constans tuvo un mejor rendimiento diagnóstico entre los pacientes hospitalizados y ambulatorios, con un área bajo la curva ROC de 0,73 (95 % 0,70-0,78) al compararla con Wells clásico, Wells modificado, Oudega y CEBI. Al comparar el rendimiento de Constans en ambos grupos de pacientes por separado, también se observó un mejor rendimiento con respecto a las demás escalas. Conclusión: la escala de Constans presenta un mejor rendimiento diagnóstico comparado con las demás escalas al ser aplicada en paciente hospitalizados y ambulatorios.

Summary Introduction: The deep vein thrombosis (DVT) is a common entity that mainly affects the deep venous system of the lower limbs, for which multiple clinical prediction scales have been developed, which have been constructed and validated in outpatients and inpatients. Objetives: We aimed to validated five clinical prediction scores for the diagnosis of lower limb DVT in patients from La Sabana de Bogota, Colombia. Methods: A cross-sectional study with analysis of a diagnostic test was carried out in patiens with suspected deep vein thrombosis, including those who had venous Doppler ultrasound of the lower limbs for suspected DVT. The performance of five clinical prediction scales for DVT (classic and modified Wells, Oudega, CEBI and Constans) for outpatients and inpatients was calculated in those scores who are validated in both populations and only in ambulatory or hospitalized patients for those that are specific scores. Results: Nine hundred seventy-four patients were entered into the analysis, of which 485 (49.7%) presented DVT. The Constans scale had a better diagnostic performance among inpatients and outpatients with an area under the ROC curve of 0.73 (95% 0.70-0.78) when compared with classic Wells, modified Wells, Oudega and CEBI. When we compared Constans performance in both groups of patients separately, we observed better performance with respect to the other scores. Conclusion: The Constans scale presents a better diagnostic performance compared to the other scales when applied to inpatients and outpatients.

Efficient and Fast Generation of Relevant Disease Mouse Models by In Vitro and In Vivo Gene Editing of Zygotes.

Knockout mice for human disease-causing genes provide valuable models in which new therapeutic approaches can be tested. Electroporation of genome editing tools into zygotes, in vitro or within oviducts, allows for the generation of targeted mutations in a shorter time. We have generated mouse models deficient in genes involved in metabolic rare diseases (Primary Hyperoxaluria Type 1 Pyruvate Kinase Deficiency) or in a tumor suppressor gene (Rasa1). Pairs of guide RNAs were designed to generate controlled deletions that led to the absence of protein. In vitro or in vivo ribonucleoprotein (RNP) electroporation rendered more than 90% and 30% edited newborn animals, respectively. Mice lines with edited alleles were established and disease hallmarks have been verified in the three models that showed a high consistency of results and validating RNP electroporation into zygotes as an efficient technique for disease modeling without the need to outsource to external facilities.