Bactericidal effect of tetracycline in E. coli strain ED1a may be associated with ribosome dysfunction.

Ribosomes translate the genetic code into proteins. Recent technical advances have facilitated in situ structural analyses of ribosome functional states inside eukaryotic cells and the minimal bacterium Mycoplasma. However, such analyses of Gram-negative bacteria are lacking, despite their ribosomes being major antimicrobial drug targets. Here we compare two E. coli strains, a lab E. coli K-12 and human gut isolate E. coli ED1a, for which tetracycline exhibits bacteriostatic and bactericidal action, respectively. Using our approach for close-to-native E. coli sample preparation, we assess the two strains by cryo-ET and visualize their ribosomes at high resolution in situ. Upon tetracycline treatment, these exhibit virtually identical drug binding sites, yet the conformation distribution of ribosomal complexes differs. While K-12 retains ribosomes in a translation-competent state, tRNAs are lost in the vast majority of ED1a ribosomes. These structural findings together with the proteome-wide abundance and thermal stability assessments indicate that antibiotic responses are complex in cells and can differ between different strains of a single species, thus arguing that all relevant bacterial strains should be analyzed in situ when addressing antibiotic mode of action.

High-confidence 3D template matching for cryo-electron tomography.

Visual proteomics attempts to build atlases of the molecular content of cells but the automated annotation of cryo electron tomograms remains challenging. Template matching (TM) and methods based on machine learning detect structural signatures of macromolecules. However, their applicability remains limited in terms of both the abundance and size of the molecular targets. Here we show that the performance of TM is greatly improved by using template-specific search parameter optimization and by including higher-resolution information. We establish a TM pipeline with systematically tuned parameters for the automated, objective and comprehensive identification of structures with confidence 10 to 100-fold above the noise level. We demonstrate high-fidelity and high-confidence localizations of nuclear pore complexes, vaults, ribosomes, proteasomes, fatty acid synthases, lipid membranes and microtubules, and individual subunits inside crowded eukaryotic cells. We provide software tools for the generic implementation of our method that is broadly applicable towards realizing visual proteomics.

Thinner is not always better: Optimizing cryo-lamellae for subtomogram averaging.

Cryo-electron tomography (cryo-ET) is a powerful method to elucidate subcellular architecture and to structurally analyze biomolecules in situ by subtomogram averaging, yet data quality critically depends on specimen thickness. Cells that are too thick for transmission imaging can be thinned into lamellae by cryo-focused ion beam (cryo-FIB) milling. Despite being a crucial parameter directly affecting attainable resolution, optimal lamella thickness has not been systematically investigated nor the extent of structural damage caused by gallium ions used for FIB milling. We thus systematically determined how resolution is affected by these parameters. We find that ion-induced damage does not affect regions more than 30 nanometers from either lamella surface and that up to ~180-nanometer lamella thickness does not negatively affect resolution. This shows that there is no need to generate very thin lamellae and lamella thickness can be chosen such that it captures cellular features of interest, thereby opening cryo-ET also for studies of large complexes.

AlphaFold: Research accelerator and hypothesis generator.

To celebrate the 50th anniversary of Cell Press and the Cell focus issue on structural biology, we discussed with scientists working across diverse fields how AlphaFold has changed their research and brought structural biology to the masses.

Understanding the cell: Future views of structural biology.

Determining the structure and mechanisms of all individual functional modules of cells at high molecular detail has often been seen as equal to understanding how cells work. Recent technical advances have led to a flush of high-resolution structures of various macromolecular machines, but despite this wealth of detailed information, our understanding of cellular function remains incomplete. Here, we discuss present-day limitations of structural biology and highlight novel technologies that may enable us to analyze molecular functions directly inside cells. We predict that the progression toward structural cell biology will involve a shift toward conceptualizing a 4D virtual reality of cells using digital twins. These will capture cellular segments in a highly enriched molecular detail, include dynamic changes, and facilitate simulations of molecular processes, leading to novel and experimentally testable predictions. Transferring biological questions into algorithms that learn from the existing wealth of data and explore novel solutions may ultimately unveil how cells work.

Sarcoma sinovial primario de pericardio / Primary pericardial synovial sarcoma

Resumen El sarcoma sinovial primario del pericardio es un tumor muy raro y de mal pronóstico y se sabe poco en cuanto al manejo terapéutico. Presentamos el caso de una paciente de 51 años a quien se le realizó resección quirúrgica incompleta, quimioterapia y radioterapia. Hasta donde sabemos, este es el primer caso de un sarcoma sinovial primario de pericardio que luego de operado se mantuvo asintomático durante 5 años hasta que en una TAC de control se le detectaron metástasis cardiacas que comprometían las cavidades derechas y con quimioterapia, la ecocardiografía demostró la reso lución completa de las mismas.

Abstract Primary pericardial synovial sarcoma is an extraor dinarily very rare tumor with a poor prognosis, and little is known about its therapeutic management. We describe the case of a 51-year-old woman patient who underwent incomplete surgical resection, chemotherapy, and radiotherapy. To the best of our knowledge, no pri mary pericardial synovial sarcoma has been described which, after surgery, remains asymptomatic for 5 years, and until a control CT scan detects cardiac metastases that compromised the lumen of the right cavities and with chemotherapy, echocardiography demonstrated complete resolution of cardiac metastases.

Experiencia en endocarditis infecciosa a lo largo de 30 años en un hospital de Buenos Aires / Experience in infective endocarditis over 30 years in a hospital in Buenos Aires

Resumen La endocarditis infecciosa (EI) es una enfermedad que en las últimas décadas ha mostrado cambios en su presentación, diagnóstico y tratamiento. Se realizó un estudio prospectivo de 252 pacientes ingresados en un hospital de referencia en Buenos Aires, Argentina, con diagnóstico de EI, agrupados según década de ingreso: Grupo A: enero 1988 a diciembre 1997 (89 pacientes -35.3%-), Grupo B: enero 1998 a diciembre 2007 (88 pacientes -34.9%-), y Grupo C: enero 2008 a diciembre 2018 (75 pacientes -29.8%). Se analizaron y compararon las características: edad, sexo, cardiopatía de base, hemocultivos y gérmenes, presencia de vegetaciones, tratamiento quirúrgico y mortalidad intrahospitalaria. Durante las tres décadas, la cardiopatía predisponente mostró que la enfermedad valvular reumática disminuyó significativamente (p < 0.0001) mientras que la EI en los dispositivos cardíacos aumentó significativamente (p < 0.0001). El porcentaje de hemocultivos negativos disminuyó significativamente a lo largo de los años (p < 0.0001). La mortalidad hospitalaria mostró una reducción en la última década (p = 0.069). El desarrollo de complicaciones durante la hospitalización, la indicación de cirugía y la presencia de insuficiencia cardíaca al ingreso fueron predictores independientes de mortalidad hospitalaria. La presencia de vegetaciones y síndrome febril al ingreso fueron predictores independientes de menor mortalidad. La comparación a través de los años mostró cambios importantes en el perfil epidemiológico de la EI. Probablemente por el avance en las técnicas diagnósticas, el tratamiento y la implementación de equi pos interdisciplinarios de EI de la última década, la mortalidad intrahospitalaria marca una fuerte tendencia a la reducción.

Abstract Infective endocarditis (IE) is a disease that in recent decades has shown changes in its presentation, diagno sis and treatment. This is a prospective study of 252 patients admitted at a reference hospital in Buenos Aires, Argentina, with a diagnosis of IE and they were grouped according to the decade of admission: Group A: from January 1988 to December 1997 (89 patients -35.3%-), Group B: from January 1998 to December 2007 (88 patients -34.9%-), and Group C: from January 2008 to December 2018 (75 patients -29.8%). The characteristics were analyzed and compared: age, sex, underlying heart disease, blood cultures and germs, presence of veg etations, surgical treatment and in-hospital mortality. Over the three decades, the predisposing heart condition showed that rheumatic valve disease decreased significantly (p < 0.0001) while the IE in cardiac devices also increased significantly (p < 0.0001). The percentage of blood culture-negatives decreased significantly over the years (p < 0.0001). In-hospital mortality showed a downward trend in the last decade (p = 0.069). The devel opment of complications during hospitalization, the indication for surgery, and the presence of heart failure on admission were independent predictors of in-hospital mortality. The presence of vegetations and febrile syndrome on admission were independent predictor for lower mortality. The comparison over the years showed important changes in the epidemiological profile of IE. Probably due to advances in diagnostic techniques, treatment, and the implementation of interdisciplinary IE teams in the last decade, in-hospital mortality shows a strong tendency to decrease.

Examen clínico y maniobra de valsalva en la insuficiencia cardíaca / Clinical examination and the Valsalva maneuver in heart failure

La evaluación de la congestión en pacientes con insuficiencia cardíaca y fracción de eyección reducida (ICFEr) resulta relevante y estrechamente vinculada al curso clínico. El agregado del comportamiento de la presión arterial durante la maniobra de Valsalva en la cabecera del paciente (VAL) podría mejorar la evaluación clínica de congestión cuando la comparamos con los niveles de NT-proBNP y la estimación de la presión media en la aurícula izquierda por Doppler cardíaco, como subrogantes de congestión. Se realizó un examen clínico con el VAL, un examen de laboratorio y un Doppler cardíaco en 69 pacientes ambulatorios con insuficiencia cardíaca y fracción de eyección ≤ 40% en ritmo sinusal. El score de Framingham ≥ 2 (SFr ≥ 2) se empleó para evaluar congestión clínica. VAL fue clasificado como normal o anormal, el NT-proBNP como bajo (< 1000 pg/ml) o alto (≥ 1000 pg/ml) y la relación entre la velocidad del llenado ventricular rápido y la velocidad del Doppler tisular (E/e') como baja < 15 o alta ≥ 15. Se halló que 13/27 pacientes con SFr ≥ 2 tenían NT-proBNP alto (sensibilidad 62%, especificidad 70% y razón de verosimilitud positiva [LR+] de 2.08, p = 0.01). El agregado del VAL al SFr ≥ 2 mejoró la exactitud diagnóstica (sensibilidad 100%, especificidad 64% y LR+ 2.8 p = 0.0004). Comparado con la E/e', SFr ≥ 2 con VAL anormal mostró sensibilidad 86%, especificidad 56% y LR + 1.86 (p = 0.03). En conclusión, el agregado del comportamiento de la presión durante la maniobra de Valsalva podría mejorar la evaluación clínica de la congestión en la insuficiencia cardíaca.

Congestion in heart failure patients with reduced ejection fraction (HFrEF) is relevant and closely linked to the clinical course. Bedside blood pressure measurement during the Valsalva maneuver (Val) added to clinical examination may improve the assessment of congestion when compared to NT-proBNP levels and left atrial pressure (LAP) estimation by Doppler echocardiography, as surrogate markers of congestion in HFrEF. A clinical examination, LAP and blood tests were performed in 69 HFrEF ambulatory patients with left ventricular ejection fraction ≤ 40% and sinus rhythm. Framingham Heart Failure Score (HFS) was used to evaluate clinical congestion; Val was classified as normal or abnormal, NT-proBNP was classified as low (< 1000 pg/ml) or high (≥ 1000 pg/ml) and the ratio between Doppler early mitral inflow and tissue diastolic velocity was used to estimate LAP and was classified as low (E/e'< 15) or high (E/e' ≥ 15). A total of 69 patients with HFrEF were included; 27 had a HFS ≥ 2 and 13 of them had high NT-proBNP. HFS ≥ 2 had a 62% sensitivity, 70% specificity and a positive likelihood ratio of 2.08 (p=0.01) to detect congestion. When Val was added to clinical examination, the presence of a HFS ≥ 2 and abnormal Val showed a 100% sensitivity, 64% specificity and a positive likelihood ratio of 2.8 (p = 0.0004). Compared with LAP, the presence of HFS ≥ 2 and abnormal Val had 86% sensitivity, 54% specificity and a positive likelihood ratio of 1.86 (p = 0.03). In conclusion, an integrated clinical examination with the addition Valsalva maneuver may improve the assessment of congestion in patients with HFrEF.

Hallazgos ecocardiográficos en la esclerodermia sin antecedentes cardiovasculares / Echocardiographic Findings in Scleroderma without History of Cardiovascular Disease

Introducción La esclerodermia es una enfermedad del tejido conectivo que se caracteriza por extensa fibrosis cutánea y de diversos órganos. La insuficiencia cardíaca y la hipertensión pulmonar son síndromes cardiovasculares asociados con esta entidad que determinan pronóstico. La ecocardiografía Doppler cumple un papel fundamental en el examen anatómico y funcional de las estructuras cardíacas en la esclerodermia y permite el diagnóstico de alteraciones incipientes, en la relajación o la mecánica segmentaria ventricular, entre otras, además de la evaluación de la presencia y el grado de hipertensión pulmonar. Objetivo Describir los hallazgos ecocardiográficos relevantes en pacientes con diagnóstico de esclerodermia sin antecedentes cardiovasculares. Material y métodos Estudio de corte transversal y descriptivo de 46 pacientes con diagnóstico establecido de esclerodermia, que asistieron al laboratorio de ecocardiografía por control de rutina. Se realizó el análisis del ventrículo izquierdo (VI) que comprendió función sistólica (FSVI) a través de la fracción de eyección (FEy) y se definió deterioro de la FSVI ante una FEy < 50%, determinación de diámetro sistólico (DS) y diastólico (DD), espesores parietales, asinergias regionales (AR), función diastólica (ondas E y A, relación del flujo transmitral), que se definió como normal, tipo I (alteración de relajación), tipo II (seudonormal) y tipo III (restrictivo), y Doppler tisular septal y lateral con ondas S, E y A, relación E/E´. En la aurícula izquierda (AI) se cuantificó el área en cm²; se consideró dilatación ante valores > 20 cm². En el ventrículo derecho se evaluó función sistólica (FSVD) cualitativa, TAPSE (excursión del anillo tricuspídeo), diámetro diastólico (DDVD) a nivel basal, Doppler tisular lateral (DTVD) con determinación de ondas S, E y A, función diastólica (FDVD) por ondas E y A y relación del flujo tricuspídeo. Se calcularon la presión pulmonar sistólica (PPS) a través de la suma del gradiente del reflujo tricuspídeo y la presión de la aurícula derecha (diámetro de la vena cava inferior y colapso inspiratorio). Se evaluó la presencia de valvulopatías significativas. Resultados La edad media de los pacientes fue de 50 ± 7,7 años, el 84% de sexo femenino, el 26% tenía hipertensión arterial y el 24%, tabaquismo. Entre los hallazgos ecocardiográficos se observaron: FSVI conservada en el 98%, FEy promedio del 62 ± 9,95%, septum interventricular de 9,7 ± 2 mm, DDVI de 43 ± 8,5 mm, AR en el 2% (n = 1); la FDVI fue normal en el 48%, tipo I en el 44%, tipo II en el 2% y tipo III en el 4%. El DTVI presentó una onda S promedio de 11 ± 0,16 cm/seg y valores < 11 en el 45%, con una relación E/E' de 5 (0% > 10). El área promedio de la AI fue de 18 ± 3,1 cm² y se observó dilatación en el 24%. VD: FSVD normal en el 96% (n = 44), DDVD de 32 ± 3,6 mm, TAPSE 24 ± 3,1 mm, trastorno de relajación del VD en el 26%, onda S DTVD 14 ± 2,1 cm/seg; la PPS promedio fue de 40 mm ± 11,7 Hg con un rango de 25-110 mm Hg y con diagnóstico de hipertensión pulmonar en el 48% (n = 22). El incremento de los valores de PPS = 50 mm Hg (moderada-grave) alcanzó el 15% (n = 7). Se halló derrame pericárdico en 4 pacientes. No se observaron valvulopatías significativas. Conclusiones Entre los hallazgos ecocardiográficos de los pacientes con esclerodermia sin antecedentes cardiovasculares se observó una prevalencia elevada de disfunción diastólica leve del VI y del VD, dilatación de la AI, disminución de la onda S en el DTVI (menor contracción longitudinal regional) y un porcentaje elevado de hipertensión pulmonar (48%).

Echocardiographic Findings in Scleroderma without History of Cardiovascular Disease Introduction Scleroderma is a connective tissue disease characterized by extensive fibrosis of the skin and various organs. Heart failure and pulmonary hypertension are cardiovascular syndromes associated with this entity that determine prognosis. Doppler echocardiography plays a key role in anatomical and functional assessment of scleroderma and allows the diagnosis of early alterations in ventricular relaxation or segmental contraction, as well as assessing the presence and degree of pulmonary hypertension. Objective The aim of the study was to describe relevant echocardiographic findings in patients with scleroderma without history of cardiovascular disease. Methods A cross-sectional, descriptive study was conducted in 46 patients with confirmed diagnosis of scleroderma, who underwent a routine echocardiographic study. Left ventricular systolic function (LVSF) was assessed through ejection fraction (EF). Left ventricular systolic function impairment was defined as EF < 50%, systolic diameter (LVSD) and diastolic diameter (LVDD), wall thickness, regional assynergies (RA), diastolic function (E and A waves and transmitral flow ratio, defined as normal, type I (impaired relaxation), type II (pseudonormal), and type III (restrictive), and septal and lateral tissue Doppler (LVTD) with S, E and A waves and the E/E' ratio. The left atrial (LA) área was assessed in cm² and LA dilation was defined as área > 20 cm². Right ventricular assessment included: qualitative systolic function (RVSF), TAPSE (tricuspid annular plañe systolic excursión), diastolic diameter (RVDD) at the basal level, lateral tissue Doppler (RVTD) with measurement of S, E and A waves, and diastolic function (RVDF) by means of E and A waves and tricuspid flow. Systolic pulmonary pressure (SPP) was measured from the sum of the tricuspid reflux gradient and right atrial pressure (inferior vena cava diameter and inspiratory collapse). Presence of significant valve disease was quantified. Results Forty-six patients with mean age of 50 ± 7.7 years were include in the study. Eighty-four percent of patients were female, 26% had hypertension and 24% smoking habits. Echocardiographic findings showed preserved LVSF in 98% of patients, mean EF of 62±9.95%, interventricular septum 9.7±2 mm, LVDD 43±8.5 mm, and RA in 2% of patients (n = 1). Normal LVDF was present in 48% of patients, type I in 44%, type II in 2% and type III in 4%. Mean left ventricular tissue Doppler S wave was 11±0.16 cm/s and valúes < 11 were found in 45% of cases, with an E/E' ratio of 5 (0% > 10). The LA área was 18±3.1 cm² and 24% of patients presented dilation. Normal RVSF was found in 96% of patients (n = 44), RVDD was 32±3.6 mm, TAPSE 24±3.1 mm, impaired RV relaxation was found in 26% of cases and RVTD S wave was 14 cm/s. Mean SPP was 40 ±11.7 mm Hg (range 25-110 mm Hg), with diagnosis of pulmonary hypertension in 48% of patients (n = 22). An increase in SPP > 50 mm Hg (moderate-severe) was present in 15% of patients (n = 7). Pericardial effusion was found in 4 patients. There was no significant valve disease. Conclusions The echocardiographic findings of patients with scleroderma without history of cardiovascular disease showed a high prevalence of mild LV and RV diastolic dysfunction, LA dilation, decreased LVTD S wave (lower regional longitudinal contraction) and high incidence of pulmonary hypertension (48%).