Prenatal diagnosis of Noonan syndrome in fetuses with increased nuchal translucency and a normal karyotype.

OBJECTIVE: Noonan syndrome (NS), one of the most common RASopathies, has an estimated incidence of 1 in 1,000-2,500 live births. In the prenatal period increased nuchal translucency, hygroma colli, hydrops fetus, congenital heart disease, kidney defects, larger amount of amniotic fluid can be observed in affected fetuses with this syndrome. In the fetuses with normal karyotype and no microdeletion/microduplication syndromes the examination of selected genes for RASopathies was added. The aim of the study was to evaluate the clinical benefit of massively parallel sequencing (MPS) of susceptible fetal DNA for NS, i.e., the diagnostic yield on the one hand and the proportion of detected variants of unknown significance (VOUS) on the other. DESIGN: Clinically diagnostic. SETTING: Centrum prenatální diagnostiky, Brno, s.r.o; Cytogenetická laboratoř Brno, s.r.o. METHODS: Samples of amniotic fluid or chorionic villus were analyzed. Selected genes associated with RASopathies were analyzed in case of the negative result of karyotype and array-CGH. A panel of twenty genes was investigated by MPS. RESULTS: In the two-years period, Noonan syndrome was detected in 10 from 95 investigated fetuses. This represents a 10.5% diagnostic efficiency of the method. DNA variants of unknown significance were detected in 10 fetuses. A segregation analysis helped to clarify their meaning in six fetuses. CONCLUSION: MPS allows fast molecular-genetic diagnosis of RASopathies already in the prenatal period. This method contributes to the clarification not only of phenotypic manifestations in already born individuals but also of ultrasound findings in fetuses with both normal karyotype and aCGH.

SARS-CoV-2 viral load is linked to remdesivir efficacy in severe Covid-19 admitted to intensive care.

Remdesivir therapy has been declared as efficient in the early stages of Covid-19. Of the 339 patients (males 55.8%, age 71(59;77) years) with a detectable viral load, 140 were treated with remdesivir (of those 103 in the ICU and 57 immunosuppressed) and retrospectively compared with 199 patients (of those 82 in the ICU and 28 immunosuppressed) who were denied therapy due to advanced Covid-19. The viral load was estimated by detecting nucleocapsid antigen in serum (n = 155, median 217(28;1524)pg/ml), antigen in sputum (n = 18, COI 18(4.6;32)), nasopharyngeal antigen (n = 44, COI 17(8;35)) and the real-time PCR (n = 122, Ct 21(18;27)). After adjustment for confounders, patients on remdesivir had better 12-month survival (HR 0.66 (0.44;0.98), p = 0.039), particularly when admitted to the ICU (HR 0.49 (0.29;0.81), p = 0.006). For the immunocompromised patients, the difference did not reach statistical significance (HR 0.55 (0.18;1.69), p = 0.3). The other most significant confounders were age, ICU admission, mechanical ventilation, leukocyte/lymphocyte ratio, admission creatinine and immunosuppression. The impact of monoclonal antibodies or previous vaccinations was not significant. Despite frequent immune suppression including haemato-oncology diseases, lymphopenia, and higher inflammatory markers in the remdesivir group, the results support remdesivir administration with respect to widely available estimates of viral load in patients with high illness severity.

Echocardiography predictors of sustained sinus rhythm after cardioversion of supraventricular arrhythmia in patients with septic shock.

PURPOSE: The echocardiography parameters may predict the maintenance of sinus rhythm after cardioversion of a supraventricular arrhythmia (SVA). MATERIALS AND METHODS: Patients in septic shock with onset of an SVA, normal to moderately reduced LV systolic function (EF_LV˃̳35%) and on a continuous noradrenaline of <1.0 µg/kg.min were included. Echocardiography was performed at the arrhythmia onset, 1 h and 4 h post cardioversion on an infusion of propafenone or amiodarone. RESULTS: Cardioversion was achieved in 96% of the 209 patients within a median time of 6(1.8-15.6)h, 134(64.1%) patients experienced at least one SVA recurrence after cardioversion. At 4 h the left atrial emptying fraction (LA_EF, cut-off 38.4%, AUC 0.69,p˂0.001), and transmitral A wave velocity-time-integral (Avti, cut-off 6.8 cm, AUC 0.65,p = 0.001) showed as limited predictors of a single arrhythmia recurrence. The LA_EF 44(36,49)%, (p = 0.005) and the Avti 8.65(7.13,9.50)cm, (p < 0.001) were associated with sustained sinus rhythm and decreased proportionally to increasing numbers of arrhythmia recurrences (p < 0.001 and p = 0.007, respectively). The enlarged left atrial end-systolic diameter at the arrhythmia onset (p = 0.04) and elevated systolic pulmonary artery pressure at 4 h (p = 0.007) were weak predictors of multiple(˃3) recurrences. CONCLUSION: The LA_EF and Avti are related to arrhythmia recurrences post-cardioversion suggesting potential guidance to the choice between rhythm and rate control strategies. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03029169, registered on 24th of January 2017.

Rapid isolation of lysosomal membranes from cultured cells.

We present a simple method for enrichment of lysosomal membranes from HEK293 and HeLa cell lines taking advantage of selective disruption of lysosomes by methionine methyl ester. Organelle concentrate from postnuclear supernatant was treated with 20 mmol/l methionine methyl ester for 45 min to lyse the lysosomes. Subsequently, lysosomal membranes were resolved on a step sucrose gradient. An enriched lysosomal membrane fraction was collected from the 20%/35% sucrose interface. The washed lysosomal membrane fraction was enriched 30 times relative to the homogenate and gave the yield of more than 8%. These results are comparable to lysosomal membranes isolated by magnetic chromatography from cultured cells (Diettrich et al., 1998). The procedure effectively eliminated mitochondrial contamination and minimized contamination from other cell compartments. The enriched fractions retained the ability to acidify membrane vesicles through the activity of lysosomal vacuolar ATPase. The method avoids non-physiological overloading of cells with superparamagnetic particles and appears to be quite robust among the tested cell lines. We expect it may be of more general use, adaptable to other cell lines and tissues.

The impact of obesity on the outcome of severe SARS-CoV-2 ARDS in a high volume ECMO centre: ECMO and corticosteroids support the obesity paradox.

PURPOSE: The aim was to verify the impact of obesity on the long-term outcome of patients with severe SARS-CoV-2 ARDS. MATERIALS AND METHODS: The retrospective study included patients admitted to the high-volume ECMO centre between March 2020 and March 2022. The impact of body mass index (BMI), co-morbidities and therapeutic measures on the short and 90-day outcomes was analysed. RESULTS: 292 patients were included, of whom 119(40.8%) were treated with veno-venous ECMO cannulated mostly (73%) in a local hospital. 58.5% were obese (64.7% on ECMO), the ECMO was most frequent in BMI > 40(49%). The ICU mortality (36.8% for obese vs 33.9% for the non-obese, p = 0.58) was related to ECMO only for the non-obese (p = 0.04). The 90-day mortalities (48.5% obese vs 45.5% non-obese, p = 0.603) of the ECMO and non-ECMO patients were not significantly influenced by BMI (p = 0.47, p = 0.771, respectively). The obesity associated risk factors for adverse outcome were age <50 (RR 2.14) and history of chronic immunosuppressive therapy (RR 2.11, p = 0.009). The higher dosage of steroids (RR 0.57, p = 0.05) associated with a better outcome. CONCLUSIONS: The high incidence of obesity was not associated with worse short and long-term outcomes. ECMO in obese patients together with the use of steroids in the later stage of ARDS may improve survival.

Nonlinear phenomena in oscillatory systems of homogeneous reactions - experimental observations.

An oscillating system of homogeneous chemical reactions (oxidation of malonic acid by bromate with ceric/cerous ion system as the catalyst) was followed experimentally in several reactor configurations, operated both as close and open systems. Characteristic nonlinear phenomena observed include: damped oscillations of concentrations of reaction components in a well stirred batch reactor, undampted oscillations and sudden changes of oscillatory behaviour (jump to a new steady state) in CSTR and interaction of oscillations in the two connected CSTRs. In a tubular reactor stationary and travelling concentration waves were observed. In the system of CSTR connected with a tubular reactor spreading of waves from CSTR to the tubular reactor, and vice versa was followed. In the flow through tubular reactor the formation of non-monotomic concentration profiles was measured, which were basically stationary with respect to changing flow-rate. The course of oscillations in CSTR was simulated on the basis of a semiempirical reaction kinetics model. An attempt to simulate quantitatively the behaviour of the distributed system with the common type of Fickian diffusion balance equations failed. Difficulties in the proper description of the transport of ions through the concentration gradient and the gradient of the electrical potential found in this system are discussed.

[Neurilemmoma of the esophagus imitating carcinoma of the thyroid isthmus]. / Neurilemom jícnu imitující istmický karcinom stítné zlázy.

Fine needle aspiration cytology of a "stone hard" nodule in isthmic thyroid region of a 56-year old woman yielded unusual spindle and polyploid cells in a relatively monomorphic inflammatory background. Finding was suspicious of malignancy. Differential diagnosis of reparative inflammatory changes, anaplastic or medullary carcinoma was considered. During the operation a tight connection of the tumour with oesophageal wall as well as with thyroid was discovered. Biopsy proved the tumour to be a neurilemmoma originating in the oesophageal wall. Most of the tumour cells were S-100 protein positive. Comparison of the cytologic smears and biopsy slides proved the unusual cells to be of tumour origin.

[A nosocomial epidemic caused by adenovirus type 3 at a clinic for children and adolescents]. / Nozokomální epidemie vyvolaná adenovirem typu 3 na klinice detí a dorostu.

The authors describe an epidemic of acute viral respiratory diseases with a nosocomial character at a clinic for children and adolescents, caused by adenovirus type 3. Of 80 exposed children 26 fell ill (attack rate 32.5%). The disease took a clinical course typical for adenovirus type 3. The authors discuss the problem of respiratory infections with a viral aetiology at paediatric departments and their possible prevention.