The semi-seated position slightly reduces the effort to breathe during difficult weaning

Purpose

The influence of posture on breathing effort in patients with difficult weaning is unknown. We hypothesized that posture could modulate the breathing effort in difficult-to-wean patients.

Methods

A prospective, crossover, physiologic study was performed in 24 intubated patients breathing with pressure support who had already failed a spontaneous breathing trial or an extubation episode. Their median duration of mechanical ventilation before measurements was 25?days. Breathing pattern, occlusion pressure (P _0.1), intrinsic PEEP (PEEP_i), and inspiratory muscle effort evaluated by the pressure?Ctime product of the respiratory muscles and the work of breathing were measured during three postures: the seated position in bed (90°LD), simulating the position in a chair, the semi-seated (45°), and the supine (0°) positions consecutively applied in a random order. A comfort score was obtained in 17 cooperative patients. The influence of position on chest wall compliance was measured in another group of 11 sedated patients.

Results

The 45° position was associated with the lowest levels of effort (p????0.01) and occlusion pressure (p?<?0.05), and tended to be more often comfortable. Respiratory effort was the lowest at 45° in 18/24 patients. PEEP_i and PEEP_i-related work were slightly higher in the supine position (p????0.01), whereas respiratory effort, heart rate, and P _0.1 values were increased in the seated position (p?<?0.05).

Conclusion

A 45° position helps to unload the respiratory muscles, moderately reduces PEEP_i, and is often considered as comfortable. The semi-seated position may help the weaning process in ventilator-dependent patients.     

Exonucleolytic degradation of RNA by p53 protein in cytoplasm

p53 in cytoplasm displays an intrinsic 3'-->5' exonuclease activity. To understand the significance of p53 exonuclease activity in cytoplasm, cytoplasmic extracts of various cell lines were examined for exonuclease activity with different single-stranded RNA (ssRNA) substrates. Using an in vitro RNA degradation assay, we observed in cytoplasmic extracts of LCC2 cells, expressing high levels of endogenous wtp53, an efficient 3'-->5' exonuclease activity with RNA substrates, removing the 3'-terminal nucleotides. Interestingly, RNA containing AU-rich sequences (ARE) is the permissive substrate for exonucleolytic degradation. Evidence that exonuclease function with RNA detected in cytoplasmic extracts is attributed to the p53 is supported by several facts: (1) this activity closely parallels with status and levels of endogenous cytoplasmic p53; (2) the endogenous exonuclease exerts identical RNA substrate specificity and excision profile characteristic for purified baculovirus-or bacterially-expressed wtp53s; (3) the exonuclease activity with ARE RNA is competed out by the presence of ss or double-stranded DNA substrate utilized by p53 protein in cytoplasm; (4) immunoprecipitation by specific anti-p53 antibodies markedly reduced the exonuclease activity with both RNA and DNA substrates; and (5) transfection of the wtp53, but not exonuclease-deficient mutant p53-R175H, into p53-null H1299 or HCT116 cells induced high levels of exonuclease activity with ARE RNA substrate in cytoplasm with characteristic excision profile. The efficient ARE RNA degradation correlates with the efficient binding of p53 to ARE RNA in cytoplasm. The possible role of p53 exonuclease activity in ARE-mRNA destabilization in cytoplasm, which may be important for expression of proteins that control cell growth and/or apoptosis is discussed.     

Rapid detection of blaKPC carbapenemase genes by real-time PCR

Carbapenem resistance among Enterobacteriaceae is an emerging problem worldwide. Klebsiella pneumoniae carbapenemase (bla_KPC) enzymes are among the most common β-lactamases described. In this study, we report the development and validation of a real-time PCR (q-PCR) assay for the detection of bla_KPC genes using TaqMan chemistry. The q-PCR amplification of bla_KPC DNA was linear over 7 log dilutions (r² = 0.999; slope, 3.54), and the amplification efficiency was 91.6%. The q-PCR detection limit was 1 CFU, and there was no cross-reaction with DNA extracted from several multidrug-resistant bacteria. Perianal/rectal swabs (n = 187) collected in duplicate from 128 patients admitted to Sheba Medical Center surgical intensive care units were evaluated for the presence of carbapenem-resistant bacteria by culturing on MacConkey agar-plus-carbapenem disks and for bla_KPC genes by q-PCR. Carbapenem-resistant organisms, all K. pneumoniae, were isolated from 47 (25.1%) of the 187 samples collected, while bla_KPC genes were detected in 54 (28.9%) of the patient samples extracted by the NucliSENS easyMAG system. Of these, seven samples were positive for bla_KPC genes by q-PCR but negative for carbapenem resistance by culture, while all samples in which no carbapenem-resistant bacteria were detected by culture also tested negative by q-PCR. Thus, the sensitivity and specificity of the q-PCR assay after extraction by the NucliSENS easyMAG system were 100% and 95%, respectively. Similar values were obtained after DNA extraction by the Roche MagNA Pure LC instrument: 97.9% sensitivity and 96.4% specificity. Overall, the bla_KPC q-PCR assay appears to be highly sensitive and specific. The utilization of q-PCR will shorten the time to bla_KPC detection from 24 h to 4 h and will help in rapidly isolating colonized or infected patients and assigning them to cohorts.     

Epidemiology of carbapenem resistant Klebsiella pneumoniae colonization in an intensive care unit

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has emerged during recent years in several intensive care units. The objective of our study was to determine the incidence of CRKP and the risk factors associated with acquisition during intensive care unit (ICU) stay. This prospective cohort study was conducted between May 2007 and April 2008 in a medical-surgical ICU at a tertiary medical center. Rectal surveillance cultures were obtained from patients on admission and twice weekly. Of screened patients, 7.0% (21/299) were CRKP colonized on admission to the ICU. One hundred eighty (81%) patients were screened at least twice. Of these, 48 (27%) patients acquired CRKP during ICU stay. Of the 69 CRKP colonized patients (both imported and ICU acquired), 29% (20/69) were first identified by microbiologic cultures, while screening cultures identified 49 patients (71%). Of these, 23 (47%) subsequently developed clinical microbiological cultures. Independent risk factors for CRKP acquisition included recent surgery (OR 7.74; CI 3.42-17.45) and SOFA score on admission (OR 1.17; CI 1-1.22). In conclusion, active surveillance cultures detected a sizable proportion of CRKP colonized patients that were not identified by clinical cultures. Recent surgical procedures and patient severity were independently associated with CRKP acquisition.     

Platelets selectively enhance lymphocyte adhesion on subendothelial matrix under arterial flow conditions

Developing novel anti-platelet therapies is an important clinical strategy for the prevention of arterial thromboses which cause heart attacks and most strokes. Thrombin activates platelets via protease-activated receptors (PARs), and PAR antagonists are currently under investigation as antithrombotics. Yet despite these clinical advances, the importance of PARs to platelet activation during thromboses formed under pathological conditions has not been investigated. To this end, we examined the role of PAR-dependent platelet activation in thrombus formation in the presence of elevated blood shear rates. We used two in vivo thrombosis models and an ex vivo whole blood flow approach in PAR4(-/-) mice, whose platelets are unresponsive to thrombin, to show that the contribution of PAR-mediated platelet activation to thrombosis is diminished at pathological blood shear rates as a direct result of decreased incorporation of thrombin-activated platelets into growing thrombi. Our ex vivo observations were replicated in human whole blood treated with a PAR1 antagonist. These results define a novel, shear-regulated role for thrombin/PAR-dependent platelet activation during thrombosis and provide important insights into the conditions under which PAR antagonists may best be used for the prevention of acute coronary syndromes.     

Atherosclerotic pattern of coronary myocardial bridging assessed with CT coronary angiography

The aim of our study was to evaluate the atherosclerotic pattern of patients with coronary myocardial bridging (MB) by means of CT Coronary Angiography (CT-CA). 254 consecutive patients (166 male, mean age 58.6 ± 10.3) who underwent 64-slice CT-CA according to current clinical indications were reviewed for the presence of MB and concomitant segmental atherosclerotic pattern. Coronary plaques were assessed in all patients enrolled. 73 patients (29%) presented single (90%) or multiple (10%) MB, frequently (93%) localized in the mid-distal left anterior descending artery. The MB segment was always free of atherosclerosis. Segments proximal to the MB presented: no atherosclerotic disease (n = 37), positive remodeling (n = 23), <50% (n = 14), or >50% stenoses (n = 7). Distal segments presented a different atherosclerosis pattern (P < 0.0001): absence of disease (n = 73), no significant lesions (n = 8). No significant differences were found between segments proximal to MB and proximal coronary segments apart from left main trunk. Pattern of atherosclerotic lesions located in segments 6 and 7 significantly differs between patients with MB and patients without MB (P < 0.05). CT-CA is a reliable method to non-invasively demonstrate MB and related atherosclerotic pattern. CT-CA provides new insight regarding atherosclerosis distribution in segments close to MB.     

Trypanosoma cruzi Calreticulin Topographical Variations in Parasites Infecting Murine Macrophages

Trypanosoma cruzi calreticulin (TcCRT), a 47-kDa chaperone, translocates from the endoplasmic reticulum to the area of flagellum emergence. There, it binds to complement components C1 and mannan-binding lectin (MBL), thus acting as a main virulence factor, and inhibits the classical and lectin pathways. The localization and functions of TcCRT, once the parasite is inside the host cell, are unknown. In parasites infecting murine macrophages, polyclonal anti-TcCRT antibodies detected TcCRT mainly in the parasite nucleus and kinetoplast. However, with a monoclonal antibody (E2G7), the resolution and specificity of the label markedly improved, and TcCRT was detected mainly in the parasite kinetoplast. Gold particles, bound to the respective antibodies, were used as probes in electron microscopy. This organelle may represent a stopover and accumulation site for TcCRT, previous its translocation to the area of flagellum emergence. Finally, early during T. cruzi infection and by unknown mechanisms, an important decrease in the number of MHC-I positive host cells was observed.     

The Sheba Medical Center healthcare workers' children's school: can we open schools safely?

ObjectiveThe role of school closure in mitigating coronavirus disease 2019 (COVID-19) transmission has been questioned. In our medical centre, during a 9-week national lockdown, an alternative school was opened for health-care workers' (HCW) children with a small number of children per class and strict symptom surveillance. After lockdown was lifted we screened children and their parents for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serology.MethodsWe conducted a cross-sectional study of HCW parents and their children after one teacher contracted COVID-19 following exposure at home and 53 children were exposed, isolated and tested by RT-PCR. We compared families with children attending the alternative school with families whose children who remained at home during the 9-week lockdown. Epidemiological and medical data were collected using a short questionnaire; nasopharyngeal and oropharyngeal swabs were obtained and tested for SARS-CoV-2 by RT-PCR, and blood was collected for SARS-CoV-2 IgA and IgG titres.ResultsA total of 435 children attended the Sheba alternative school. Among the 53 children exposed to the infected teacher, none tested positive by RT-PCR. Of these, 18 children–parent pairs were tested for serology and all were negative. A total of 106/435 (24%) children and their 78 parents were recruited for the cross-sectional study; 70 attended the Sheba school and 36 did not. Approximately 16% of children in either group reported symptoms (11/70 in the school group and 6/36 in the ‘stay home’ group), but SARS-CoV-2 was not detected by PCR in any, and previous exposure, as determined by serological tests, was low and not significantly different between the groups.ConclusionIn an alternative school for children of HCWs, active during COVID-19 national outbreak, we found no evidence of increased infection compared with children that stayed home.