Tryptophan Metabolites, Cytokines, and Fatty Acid Binding Protein 2 in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome.

Patients with Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) differ for triggers, mode of start, associated symptoms, evolution, and biochemical traits. Therefore, serious attempts are underway to partition them into subgroups useful for a personalized medicine approach to the disease. Here, we investigated clinical and biochemical traits in 40 ME/CFS patients and 40 sex- and age-matched healthy controls. Particularly, we analyzed serum levels of some cytokines, Fatty Acid Binding Protein 2 (FAPB-2), tryptophan, and some of its metabolites via serotonin and kynurenine. ME/CFS patients were heterogeneous for genetic background, trigger, start mode, symptoms, and evolution. ME/CFS patients had higher levels of IL-17A (p = 0.018), FABP-2 (p = 0.002), and 3-hydroxykynurenine (p = 0.037) and lower levels of kynurenine (p = 0.012) and serotonin (p = 0.045) than controls. Changes in kynurenine and 3-hydroxykynurenine were associated with increased kynurenic acid/kynurenine and 3-hydroxykynurenine/kynurenine ratios, indirect measures of kynurenine aminotransferases and kynurenine 3-monooxygenase enzymatic activities, respectively. No correlation was found among cytokines, FABP-2, and tryptophan metabolites, suggesting that inflammation, anomalies of the intestinal barrier, and changes of tryptophan metabolism may be independently associated with the pathogenesis of the disease. Interestingly, patients with the start of the disease after infection showed lower levels of kynurenine (p = 0.034) than those not starting after an infection. Changes in tryptophan metabolites and increased IL-17A levels in ME/CFS could both be compatible with anomalies in the sphere of energy metabolism. Overall, clinical traits together with serum biomarkers related to inflammation, intestine function, and tryptophan metabolism deserve to be further considered for the development of personalized medicine strategies for ME/CFS.

Response to a massive SARS-CoV-2 infection in a nursing home transformed into a caring center.

BACKGROUND: The best policy to follow when nursing homes are massively hit by SARS-CoV2 is unclear. AIM: To describe COVID-19 containment in a nursing home transformed into a caring center. METHODS: Physicians and nurses were recruited. The facility was reorganized and connected with the laboratory of the reference hospital. Ultrasound was used to diagnose pneumonia. Patients needing intensive care were transferred to the reference hospital. Hydroxychloroquine/azithromycin/enoxaparin were used initially, while amiodarone/enoxaparin were used at a later phase. Under both regimens, methylprednisolone was added for severe cases. Prophylaxis was done with hydroxychloroquine initially and then with amiodarone. PERIOD COVERED: March 22-July 31, 2020. RESULTS: The facility was reorganized in two days. Ninety-two guests of the 121 (76%) and 25 personnel of 118 (21.1%) became swab test positive. Seven swab test negative patients who developed symptoms were considered to have COVID-19. Twenty-seven patients died, 23 swab test positive, 5 of whom after full recovery. Four patients needing intensive care were transferred (3 died). Mortality, peaking in April 2020, was correlated with symptoms, comorbidities, dyspnea, fatigue, stupor/coma, high neutrophil to lymphocyte ratio, C-reactive protein, interleukin-6, pro-calcitonin, and high oxygen need (p ≤ 0.001 for all). Among swab-positive staff, 3 had pneumonia and recovered. Although no comparison could be made between different treatment and prophylaxis strategies, potentially useful suggestions emerged. Mortality compared well with that of nursing homes of the same area not transformed into care centers. CONCLUSION: Nursing homes massively hit by SARS-CoV-2 can become caring centers for patients not needing intensive care.

Approach to patients with COVID-19 disease: the procedure in Udine.

Coronavirus disease 2019 poses a serious threat to public health. The protocol developed at the Azienda Sanitaria Universitaria Friuli Centrale (Italy) is based on clinical data, laboratory tests, chest echography and HRCT. Several therapeutic options are considered, since patients vary in disease severity, evolution and co-morbidities and because so far there are no clear indications about therapeutic strategy based on randomized clinical trial. In this protocol chest echography has a central role in categorizing patient status, follow-up and decision-making.

Amiodarone affects Ebola virus binding and entry into target cells.

Ebola Virus Disease is one of the most lethal transmissible infections characterized by a high fatality rate. Several research studies have aimed to identify effective antiviral agents. Amiodarone, a drug used for the treatment of arrhythmias, has been shown to inhibit filovirus infection in vitro by acting at the early step of the viral replication cycle. Here we demonstrate that amiodarone reduces virus binding to target cells and slows down the progression of the viral particles along the endocytic pathway. Overall our data support the notion that amiodarone interferes with Ebola virus infection by affecting cellular pathways/ targets involved in the viral entry process.

Influence of the type of congenital heart defects on epithelial lining fluid composition in infants undergoing cardiac surgery with cardiopulmonary bypass.

BackgroundIn children with congenital heart disease (CHD), altered pulmonary circulation compromises gas exchange. Moreover, pulmonary dysfunction is a complication of cardiac surgery with cardiopulmonary bypass (CPB). No data are available on the effect of different CHDs on lung injury. The aim of this study was to analyze epithelial lining fluid (ELF) surfactant composition in children with CHD.MethodsTracheal aspirates (TAs) from 72 CHD children (age 2.9 (0.4-5.7) months) were obtained before and after CPB. We measured ELF phospholipids, surfactant proteins A and B (SP-A, SP-B), albumin, and myeloperoxidase activity. TAs from 12 infants (age 1.0 (0.9-2.9) months) with normal heart/lung served as controls.ResultsHeart defects were transposition of great arteries (19), tetralogy of Fallot (TOF, 20), atrial/ventricular septal defect (ASD/VSD, 22), and hypoplastic left heart syndrome (11). Increased levels of ELF SP-B were found in all defects, increased myeloperoxidase activity in all except the TOF, and increased levels of ELF albumin and SP-A only in ASD/VSD patients. Postoperatively, ELF findings remained unchanged except for a further increase in myeloperoxidase activity.ConclusionELF composition has distinctive patterns in different CHD. We speculate that a better knowledge of the ELF biochemical changes may help to prevent respiratory complications.

Chest ultrasound findings in pulmonary tuberculosis.

In resource-limited countries, the diagnosis of pulmonary tuberculosis (TB) is based on clinical findings, chest radiography and the demonstration of acid-fast bacilli in sputum. Few data are available on the use of ultrasound (US) to diagnose pulmonary TB. Chest US was performed in patients with lung TB from a rural African setting, to look for signs of the disease and to clarify the role US may have in the diagnosis of pulmonary TB. Sixty adult patients diagnosed with lung TB underwent chest US. All patients had abnormal findings. The most frequent was a subpleural nodule (SUN), which was mostly multiple and also found in radiologically normal areas. Other findings were lung consolidations, cavitations, miliary patterns made of miniature SUNs, and pleural and pericardial effusions. Chest US is a complementary tool in evaluating patients with suspected lung TB in resource-limited settings where the disease has high prevalence.

Antiviral activity of cationic amphiphilic drugs.

INTRODUCTION: Emerging and reemerging viral infections represent a major concern for human and veterinary public health and there is an urgent need for the development of broad-spectrum antivirals. Areas covered: A recent strategy in antiviral research is based on the identification of molecules targeting host functions required for infection of multiple viruses. A number of FDA-approved drugs used to treat several human diseases are cationic amphiphilic drugs (CADs) that have the ability to accumulate inside cells affecting several structures/functions hijacked by viruses during infection. In this review we summarized the CADs' chemical properties and effects on the cells and reported the main FDA-approved CADs that have been identified so far as potential antivirals in drug repurposing studies. Expert commentary: Although there have been concerns regarding the efficacy and the possible side effects of the off-label use of CADs as antivirals, they seem to represent a promising starting point for the development of broad-spectrum antiviral strategies. Further knowledge about their mechanism of action is required to improve their antiviral activity and to reduce the risk of side effects.

Surfactant protein B and A concentrations are increased in neonatal pneumonia.

BACKGROUND: Term newborns with pneumonia show a reduced pulmonary compliance due to multiple and ill-defined factors. Surfactant proteins' (SPs) changes could have a role in the reduced compliance but the matter is still unsettled. The aim of this study was to clarify the meaning of SPs changes during pneumonia in term newborns. METHODS: In 28 term ventilated newborns, 13 with pneumonia and 15 with no lung disease, we measured SP-B, SP-A, disaturated-phosphatidylcholine (DSPC), and total phospholipids (PL) concentrations in tracheal aspirates at intubation and close to extubation. We also measured DSPC kinetics using (U-(13)C-PA)dipalmitoyl-phosphatidylcholine. RESULTS: At baseline, SP-B, expressed as % of PL, was significantly different between the groups, being 3.5-fold higher in pneumonia than controls. Conversely, SP-A did not vary between the groups. At extubation, SP-B and SP-A concentrations had decreased significantly in newborns with pneumonia, while there was no significant change in controls. DSPC t1/2 was significantly shorter in the pneumonia group (11.8 (5.5-19.8) h vs. 26.6 (19.3-63.6) h, P = 0.011). CONCLUSION: In term newborns with pneumonia, SP-B increases with respect to PL, and DSPC is turned over at a faster rate. Disease's resolution is associated with the restoration of the normal ratio between SP-B and PL.

Amiodarone and metabolite MDEA inhibit Ebola virus infection by interfering with the viral entry process.

Ebola virus disease (EVD) is one of the most lethal transmissible infections characterized by a high fatality rate, and a treatment has not been developed yet. Recently, it has been shown that cationic amphiphiles, among them the antiarrhythmic drug amiodarone, inhibit filovirus infection. In the present work, we investigated how amiodarone interferes with Ebola virus infection. Wild-type Sudan ebolavirus and recombinant vesicular stomatitis virus, pseudotyped with the Zaire ebolavirus glycoprotein, were used to gain further insight into the ability of amiodarone to affect Ebola virus infection. We show that amiodarone decreases Ebola virus infection at concentrations close to those found in the sera of patients treated for arrhythmias. The drug acts by interfering with the fusion of the viral envelope with the endosomal membrane. We also show that MDEA, the main amiodarone metabolite, contributes to the antiviral activity. Finally, studies with amiodarone analogues indicate that the antiviral activity is correlated with drug ability to accumulate into and interfere with the endocytic pathway. Considering that it is well tolerated, especially in the acute setting, amiodarone appears to deserve consideration for clinical use in EVD.

Are adaptive randomised trials or non-randomised studies the best way to address the Ebola outbreak in west Africa?

The Ebola outbreak that has devastated parts of west Africa represents an unprecedented challenge for research and ethics. Estimates from the past three decades emphasise that the present effort to contain the epidemic in the three most affected countries (Guinea, Liberia, and Sierra Leone) has been insufficient, with more than 24,900 cases and about 10,300 deaths, as of March 25, 2015. Faced with such an exceptional event and the urgent response it demands, the use of randomised controlled trials (RCT) for Ebola-related research might be both unethical and infeasible and that potential interventions should be assessed in non-randomised studies on the basis of compassionate use. However, non-randomised studies might not yield valid conclusions, leading to large residual uncertainty about how to interpret the results, and can also waste scarce intervention-related resources, making them profoundly unethical. Scientifically sound and rigorous study designs, such as adaptive RCTs, could provide the best way to reduce the time needed to develop new interventions and to obtain valid results on their efficacy and safety while preserving the application of ethical precepts. We present an overview of clinical studies registered at present at the four main international trial registries and provide a simulation on how adaptive RCTs can behave in this context, when mortality varies simultaneously in either the control or the experimental group.

Surfactant protein C metabolism in human infants and adult patients by stable isotope tracer and mass spectrometry.

Surfactant protein C (SP-C) is deemed as the surfactant protein most specifically expressed in type II alveolar epithelial cells and plays an important role in surfactant function. SP-C turnover in humans and its meaning in the clinical context have never been approached. In this study, we used mass spectrometry to investigate SP-C turnover in humans. We studied four infants and eight adults requiring mechanical ventilation. All patients had no lung disease. Patients received a 24-h continuous infusion of (13)C-leucine as precursor of SP-C, and serial tracheal aspirates and plasma samples were obtained every 6 h till 48 h. SP-C was isolated from tracheal aspirates by sorbent-phase chromatography. (13)C-leucine SP-C enrichment could be successfully measured in three infant and in four adult samples by using mass spectrometry coupled with a gas chromatographer. Median SP-C fractional synthesis rate, secretion time, and peak time were 15.7 (14.1-27.5)%/day, 6.0 (4.7-11.5) h, and 24 (20-27) h. In conclusion, this study shows that it is feasible to accurately determine SP-C turnover in humans by stable isotopes.

Impaired surfactant protein B synthesis in infants with congenital diaphragmatic hernia.

Pulmonary hypoplasia and hypertension account for significant morbidity and mortality in neonates with congenital diaphragmatic hernia (CDH). Whether CDH is associated with surfactant dysfunction remains controversial. Therefore, we measured disaturated phosphatidylcholine (DSPC) and surfactant protein (SP)-B concentration in tracheal aspirates and their synthesis rate in infants with CDH compared to infants without lung disease. (2)H2O as a precursor of DSPC and 1-(13)C-leucine as a precursor of SP-B were administered to 13 infants with CDH and eight controls matched for gestational age. DSPC and SP-B were isolated from tracheal aspirates, and their fractional synthesis rate was derived from (2)H and (13)C enrichment curves obtained by mass spectrometry. DSPC and SP-B amounts in tracheal aspirates were also measured. In infants with CDH, SP-B fractional synthesis rate and amount were 62±27% and 57±22% lower, respectively, than the value found in infants without lung disease (p<0.01 and p<0.05, respectively). There were no significant group differences in DSPC fractional synthesis rate and amount. Infants with CDH have a lower rate of synthesis of SP-B and less SP-B in tracheal aspirates. In these infants, partial SP-B deficiency could contribute to the severity of respiratory failure and its correction might represent a therapeutic goal.

Surfactant protein B amount and kinetics in newborn infants: an optimized procedure.

Surfactant protein B (SP-B) plays a key role in surfactant homeostasis affecting its biophysical properties and physiological function. Recently, a method to measure SP-B amount and kinetics from tracheal aspirates (TAs) became available. The main objective of this study was to improve the critical steps of the procedure to obtain a better SP-B sensitivity. We administered a 24 h continuous infusion of 1 mg/kg/h of 1(13)C-leucine to ten newborn infants. SP-B was isolated from serial TAs and its fractional synthesis rate, secretion time, peak time and half life were derived from (13)C enrichment curves obtained by gas chromatography mass spectrometry. SP-B amount in TAs was also assessed. During the extraction step, acidification and organic solvent ratio optimization doubled the recovery of SP-B from TAs, so did the elongation of the propylation time (from 20 min to 1 h) with enhanced leucine derivatization yield. Measurement of (13)C leucine enrichments, and therefore all SP-B kinetics parameters, were successfully calculated in all TAs samples due to the increase of SP-B yield. SP-B amount was 0.29 (0.16-0.41) % of total phospholipids with a minimum value of 0.08% belonging to one of the respiratory distress syndrome (RDS) patients. In conclusion, this new procedure enables accurate determination of SP-B kinetics even in the presence of low protein amount like in preterm RDS patients.

Simultaneous measurement of phosphatidylglycerol and disaturated-phosphatidylcholine palmitate kinetics from alveolar surfactant. Study in infants with stable isotope tracer, coupled with isotope ratio mass spectrometry.

Disaturated-phosphatidylcholine (DSPC) and phosphatidylglycerol (PG) are respectively the first and the third most abundant phospholipid in human alveolar surfactant. Their concentration decreases in airway surfactant of adults and infants with respiratory distress syndrome and cystic fibrosis. In this study, we used mass spectrometry (IRMS) to investigate the turnover of DSPC and PG in tracheal aspirates (TA) obtained from infants with normal or diseased lungs. We studied eight infants requiring mechanical ventilation: two with no lung disease, four with diaphragmatic hernia, one with ATP-binding cassette sub-family A member 3 heterozygote mutation and one with sepsis. Patients received deuterated water for 48 h as metabolic precursors of palmitate-DSPC and palmitate-PG. Serial TAs were obtained every 6 h for five days or until extubation. DSPC and PG were isolated from TA by column and high-performance thin layer chromatography. Deuterium enrichments of palmitate-DSPC and PG residues were measured by IRMS coupled with a gas chromatographer. Median secretion time (ST), peak time (PT) and fractional synthesis rate (FSR) were 3.7 [0.9- 13.4] h, 71.0 [52.2 - 85.2] h and 6.6 [6.3 - 11.1] %/day for DSPC and 19.3 [6.4 - 22.8] h, 49.0 [33.0 - 52.5] h and 5.8 [4.8 - 10.9] %/day for PG. This study shows that it is feasible to use deuterium derived from body water to trace simultaneously airway surfactant DSPC and PG in humans. When compared within the same patient, DSPC and PG had similar fractional synthesis rates, but PG had a shorter PT, suggesting differences in the life cycle of these essential surfactant components.

Amiodarone impairs trafficking through late endosomes inducing a Niemann-Pick C-like phenotype.

Patients treated with amiodarone accumulate lysobisphosphatidic acid (LBPA), also known as bis(monoacylglycero)phosphate, in airway secretions and develop in different tissues vacuoles and inclusion bodies thought to originate from endosomes. To clarify the origin of these changes, we studied in vitro the effects of amiodarone on endosomal activities like transferrin recycling, Shiga toxin processing, ESCRT-dependent lentivirus budding, fluid phase endocytosis, proteolysis and exosome secretion. Furthermore, since the accumulation of LBPA might point to a broader disturbance in lipid homeostasis, we studied the effect of amiodarone on the distribution of LBPA, unesterified cholesterol, sphingomyelin and glycosphyngolipids. Amiodarone analogues were also studied, including the recently developed derivative dronedarone. We found that amiodarone does not affect early endosomal activities, like transferrin recycling, Shiga toxin processing and lentivirus budding. Amiodarone, instead, interferes with late compartments of the endocytic pathway, blocking the progression of fluid phase endocytosis and causing fusion of organelles, collapse of lumenal structures, accumulation of undegraded substrates and amassing of different types of lipids. Not all late endocytic compartments are affected, since exosome secretion is spared. These changes recall the Niemann-Pick type-C phenotype (NPC), but originate by a different mechanism, since, differently from NPC, they are not alleviated by cholesterol removal. Studies with analogues indicate that basic pKa and high water-solubility at acidic pH are crucial requirements for the interference with late endosomes/lysosomes and that, in this respect, dronedarone is at least as potent as amiodarone. These findings may have relevance in fields unrelated to rhythm control.

Disaturated-phosphatidylcholine and surfactant protein-B turnover in human acute lung injury and in control patients.

BACKGROUND: Patients with adult respiratory distress syndrome (ARDS) and acute lung injury (ALI) have low concentrations of disaturated-phosphatidylcholine and surfactant protein-B in bronchoalveolar lavage fluid. No information is available on their turnover. OBJECTIVES: To analyze disaturated-phosphatidylcholine and surfactant protein-B turnover in patients with ARDS/ALI and in human adults with normal lungs (controls). METHODS: 2H2O as precursor of disaturated-phosphatidylcholine-palmitate and 113C-Leucine as precursor of surfactant protein-B were administered intravenously to 12 patients with ARDS/ALI and to 8 controls. Disaturated-phosphatidylcholine and surfactant protein-B were isolated from serial tracheal aspirates, and their fractional synthetic rate was derived from the 2H and 13C enrichment curves, obtained by gas chromatography mass spectrometry. Disaturated-phosphatidylcholine, surfactant protein-B, and protein concentrations in tracheal aspirates were also measured. RESULTS: 1) Surfactant protein-B turned over at faster rate than disaturated-phosphatidylcholine both in ARDS/ALI patients and in controls. 2) In patients with ARDS/ALI the fractional synthesis rate of disaturated-phosphatidylcholine was 3.1 times higher than in controls (p < 0.01), while the fractional synthesis rate of surfactant protein-B was not different. 3) In ARDS/ALI patients the concentrations of disaturated-phosphatidylcholine and surfactant protein-B in tracheal aspirates were markedly and significantly reduced (17% and 40% of the control values respectively). CONCLUSIONS: 1) Disaturated-phosphatidylcholine and surfactant protein-B have a different turnover both in healthy and diseased lungs. 2) In ARDS/ALI the synthesis of these two surfactant components may be differently regulated.

Dosing of porcine surfactant: effect on kinetics and gas exchange in respiratory distress syndrome.

OBJECTIVE: The goal was to study exogenous surfactant disaturated phosphatidylcholine (DSPC) kinetics in preterm infants with respiratory distress syndrome (RDS) who were treated with 100 or 200 mg/kg porcine surfactant. METHODS: Sixty-one preterm infants with RDS undergoing mechanical ventilation received, within 24 hours after birth, 100 mg/kg (N = 40) or 200 mg/kg (N = 21) porcine surfactant mixed with [U-(13)C]dipalmitoylphosphatidylcholine. Clinical and respiratory parameters were recorded, and DSPC half-life and pool size and endogenous DSPC synthesis rate were calculated. RESULTS: Clinical characteristics and short-term outcomes did not differ between groups. In the 100 mg/kg group, 28 infants (70%) received a second dose after 25 +/- 11 hours and 9 (22.5%) a third dose after 41 +/- 11 hours; in the 200 mg/kg group, 6 infants (28.6%) received a second dose after 33 +/- 8 hours and 1 a third dose. The DSPC half-life was longer in the 200 mg/kg group (first dose: 32 +/- 19 vs 15 +/- 15 hours [P = .002]; second dose: 43 +/- 32 vs 21 +/- 13 hours [P = .025]). DSPC synthesis rates and pool sizes before the first and second doses did not differ between the groups. The 200 mg/kg group exhibited a greater reduction in the oxygenation index than did the 100 mg/kg group after the first (P = .009) and second (P = .018) doses. CONCLUSIONS: Porcine surfactant given to preterm infants with RDS at a dose of 200 mg/kg resulted in a longer DSPC half-life, fewer retreatments, and better oxygenation index values.

Continuous subcutaneous insulin infusion (CSII) 30 years later: still the best option for insulin therapy.

Thirty years after its introduction, the use of continuous subcutaneous insulin infusion (CSII) keeps increasing, especially among children and adolescents. The technique, when used properly, is safe and effective.Compared with traditional NPH-based multiple daily injections (MDI), CSII provides a small but clinically important reduction of HbA(1c) levels, diminishes blood glucose variability, decreases severe hypoglycaemic episodes and offers a better way to cope with the dawn phenomenon.Insulin analogues have improved the treatment of diabetes, eroding part of the place previously occupied by CSII, but CSII still remains the first option for patients experiencing severe hypoglycaemic episodes, high HbA(1c) values or marked glucose variability while being treated with optimized MDI. Furthermore CSII is better than MDI considering the effects on quality of life and the possibility to adjust insulin administration according to physical activity or food intake.CSII may be limited by cost. Present estimates suggest that CSII may be cost-effective just for patients experiencing a marked improvement in HbA(1c) or a decrease in severe hypoglycaemic episodes, but the effects on quality of life are difficult to measure.CSII does not merely imply wearing an external device; it requires a multidisciplinary team, intensive patient education and continuous follow up.