Early anakinra treatment improves cardiac outcome of multisystem inflammatory syndrome in children, regardless of disease severity.

OBJECTIVE: The main aim of this study was to define the best treatment option for multisystem inflammatory syndrome in children (MIS-C) and to analyse the role of anakinra. METHODS: This is a multicentre retrospective cohort study. Patients were treated according to the attending physician's decision. The patients were divided into four groups on the basis of the first treatment at time of admittance: (i) IVIG, (ii) IVIG and methylprednisolone (≤2 mg/kg/day), (iii) IVIG with high-dose methylprednisolone (>2 mg/kg/day) and (iv) anakinra with or without IVIG and/or methylprednisolone. Primary outcomes were defined as the presence of at least one of the following features: death, the failure of initial treatment, meaning the need for additional treatment for clinical worsening and cardiac involvement at the end of follow-up. RESULTS: Two hundred thirty-nine patients were recruited. At univariate analysis, persistent heart involvement at discharge was more frequent in those not receiving anakinra as initial treatment (3/21 vs 66/189; P = 0.047). After comparisons between the four treatment regimens, adjusting for the propensity score, we observed that early treatment with anakinra was associated with a lower probability of developing persistent heart disease at the end of follow-up (odds ratio: 0.6; 95% CI: 0.4-1.0). CONCLUSION: We report that early treatment with anakinra is safe and very effective in patients with severe MIS-C. In addition, our study suggests that early treatment with anakinra is the most favourable option for patients with a higher risk of developing a severe disease outcome.

FGF10 Triggers De Novo Alveologenesis in a Bronchopulmonary Dysplasia Model: Impact on Resident Mesenchymal Niche Cells.

Bronchopulmonary dysplasia (BPD) is a neonatal lung disease developing in premature babies characterized by arrested alveologenesis and associated with decreased Fibroblast growth factor 10 (FGF10) expression. One-week hyperoxia (HYX) exposure of newborn mice leads to a permanent arrest in alveologenesis. To test the role of Fgf10 signaling to promote de novo alveologenesis following hyperoxia, we used transgenic mice allowing inducible expression of Fgf10 and recombinant FGF10 (rFGF10) protein delivered intraperitoneally. We carried out morphometry analysis, and IF on day 45. Alveolospheres assays were performed co-culturing AT2s from normoxia (NOX) with FACS-isolated Sca1Pos resident mesenchymal cells (rMC) from animals exposed to NOX, HYX-PBS, or HYX-FGF10. scRNAseq between rMC-Sca1Pos isolated from NOX and HYX-PBS was also carried out. Transgenic overexpression of Fgf10 and rFGF10 administration rescued the alveologenesis defects following HYX. Alveolosphere assays indicate that the activity of rMC-Sca1Pos is negatively impacted by HYX and partially rescued by rFGF10 treatment. Analysis by IF demonstrates a significant impact of rFGF10 on the activity of resident mesenchymal cells. scRNAseq results identified clusters expressing Fgf10, Fgf7, Pdgfra, and Axin2, which could represent the rMC niche cells for the AT2 stem cells. In conclusion, we demonstrate that rFGF10 administration is able to induce de novo alveologenesis in a BPD mouse model and identified subpopulations of rMC-Sca1Pos niche cells potentially representing its cellular target.

Time-resolved transcriptomic profiling of the developing rabbit's lungs: impact of premature birth and implications for modelling bronchopulmonary dysplasia.

BACKGROUND: Premature birth, perinatal inflammation, and life-saving therapies such as postnatal oxygen and mechanical ventilation are strongly associated with the development of bronchopulmonary dysplasia (BPD); these risk factors, alone or combined, cause lung inflammation and alter programmed molecular patterns of normal lung development. The current knowledge on the molecular regulation of lung development mainly derives from mechanistic studies conducted in newborn rodents exposed to postnatal hyperoxia, which have been proven useful but have some limitations. METHODS: Here, we used the rabbit model of BPD as a cost-effective alternative model that mirrors human lung development and, in addition, enables investigating the impact of premature birth per se on the pathophysiology of BPD without further perinatal insults (e.g., hyperoxia, LPS-induced inflammation). First, we characterized the rabbit's normal lung development along the distinct stages (i.e., pseudoglandular, canalicular, saccular, and alveolar phases) using histological, transcriptomic and proteomic analyses. Then, the impact of premature birth was investigated, comparing the sequential transcriptomic profiles of preterm rabbits obtained at different time intervals during their first week of postnatal life with those from age-matched term pups. RESULTS: Histological findings showed stage-specific morphological features of the developing rabbit's lung and validated the selected time intervals for the transcriptomic profiling. Cell cycle and embryo development, oxidative phosphorylation, and WNT signaling, among others, showed high gene expression in the pseudoglandular phase. Autophagy, epithelial morphogenesis, response to transforming growth factor ß, angiogenesis, epithelium/endothelial cells development, and epithelium/endothelial cells migration pathways appeared upregulated from the 28th day of gestation (early saccular phase), which represents the starting point of the premature rabbit model. Premature birth caused a significant dysregulation of the inflammatory response. TNF-responsive, NF-κB regulated genes were significantly upregulated at premature delivery and triggered downstream inflammatory pathways such as leukocyte activation and cytokine signaling, which persisted upregulated during the first week of life. Preterm birth also dysregulated relevant pathways for normal lung development, such as blood vessel morphogenesis and epithelial-mesenchymal transition. CONCLUSION: These findings establish the 28-day gestation premature rabbit as a suitable model for mechanistic and pharmacological studies in the context of BPD.

Efficacy of synthetic surfactant (CHF5633) bolus and/or lavage in meconium-induced lung injury in ventilated newborn rabbits.

BACKGROUND: The pathogenesis of neonatal meconium aspiration syndrome (MAS) involves meconium-induced lung inflammation and surfactant inactivation. Bronchoalveolar lavage (BAL) with diluted surfactant facilitates the removal of meconium. CHF5633, one of the most promising synthetic surfactants, is effective in neonatal respiratory distress syndrome. Here we investigated its efficacy via BAL in an experimental MAS model. METHODS: Experimental MAS was induced at birth in near-term newborn rabbits by intratracheal instillation of reconstituted human meconium. First, undiluted CHF5633 was compared with a porcine-derived surfactant (Poractant alfa) via intratracheal bolus (200 mg/kg). Second, the efficacy of BAL with diluted CHF5633 (5 mg/mL, 20 ml/kg) alone, or followed by undiluted boluses (100 or 300 mg/kg), was investigated. RESULTS: Meconium instillation caused severe lung injury, reduced endogenous surfactant pool, and poor survival. CHF5633 had similar benefits in improving survival and alleviating lung injury as Poractant alfa. CHF5633 BAL plus higher boluses exerted better effects than BAL or bolus alone in lung injury alleviation by reversing phospholipid pools and mitigating proinflammatory cytokine mRNA expression, without fluid retention and function deterioration. CONCLUSIONS: CHF5633 improved survival and alleviated meconium-induced lung injury, the same as Poractant alfa. CHF5633 BAL plus boluses was the optimal modality, which warrants further clinical investigation. IMPACT: To explore the efficacy of a synthetic surfactant, CHF5633, in neonatal lung protection comparing with Poractant alfa in a near-term newborn rabbit model with meconium-induced lung injury. Similar effects on improving survival and alleviating lung injury were found between CHF5633 and Poractant alfa. Optimal therapeutic effects were identified from the diluted CHF5633 bronchoalveolar lavage followed by its undiluted bolus instillation compared to the lavage or bolus alone regimens. Animals with CHF5633 lavage plus bolus regimen exerted neither substantial lung fluid retention nor lung mechanics deterioration but a trend of higher pulmonary surfactant-associated phospholipid pools.

Relation of the work ability index to fitness for work in healthcare and public employees in a region of Northeastern Italy.

PURPOSE: Work ability indicates an individual's capacity to match job demands according to his/her physical and mental conditions and work circumstances. Occupational physicians should take into consideration the global health status of a worker in order to correctly assess if he/she is fit for the job. The aim of this study was to verify the association between fitness for work evaluation and Work Ability Index scores, as well as individual factors (age, gender, and anthropometric characteristics) and work-related variables (job type, years of working duration). METHODS: A cross-sectional study was conducted within the occupational health surveillance of health and public employers in the Friuli-Venezia Giulia region (2018-2022). The participants voluntarily agreed to answer the standard Work Ability Index questionnaire. Data were investigated by univariable as well as multivariable regression analysis. RESULTS: The Work Ability Index of the workers included in the study (N = 6893) resulted negatively associated with age, female sex, and body mass index. It was averagely lower in nurses and assistive personnel, and the highest in medical doctors and public employers. The fitness for work assessments was also statistically related to WAI scores. The results obtained from the univariable and the multivariable analysis were consistent. CONCLUSIONS: The Work Ability Index is an efficient tool to measure an individual's capability to sustain job demands, and can be taken into account to produce a correct fitness for work evaluation and consequently preserve workers' health status.

Chemotherapy after PD-1 inhibitors in relapsed/refractory Hodgkin lymphoma: Outcomes and clonal evolution dynamics.

Checkpoint inhibitors (CPIs) are routinely employed in relapsed/refractory classical Hodgkin lymphoma. Nonetheless, persistent long-term responses are uncommon, and one-third of patients are refractory. Several reports have suggested that treatment with CPIs may re-sensitize patients to chemotherapy, however there is no consensus on the optimal chemotherapy regimen and subsequent consolidation strategy. In this retrospective study we analysed the response to rechallenge with chemotherapy after CPI failure. Furthermore, we exploratively characterized the clonal evolution profile of a small sample of patients (n = 5) by employing the CALDER approach. Among the 28 patients included in the study, 17 (71%) were primary refractory and 26 (92%) were refractory to the last chemotherapy prior to CPIs. Following rechallenge with chemotherapy, response was recorded in 23 (82%) patients experiencing complete remission and 3 (11%) patients experiencing partial remission. The tumour evolution of the patients inferred by CALDER seemingly occurred prior to the first cycle of therapy and was characterized either by linear or branching evolution patterns. Twenty-five patients proceeded to allogeneic stem cell transplantation. At a median follow-up of 21 months, median PFS and OS were not reached. In conclusion, patients who fail CPIs can be effectively rescued by salvage chemotherapy and bridged to allo-SCT/auto-SCT.

Dose-adjusted EPOCH and rituximab for the treatment of double expressor and double-hit diffuse large B-cell lymphoma: impact of TP53 mutations on clinical outcome.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease, including one-third of cases overexpressing MYC and BCL2 proteins (double expressor lymphoma, DEL) and 5-10% of patients with chromosomal rearrangements of MYC, BCL2 and/or BCL-6 (double/triple-hit lymphomas, DH/TH). TP53 mutations are detected in 20- 25% of DEL. We report the efficacy of dose-adjusted EPOCH and rituximab (DA-EPOCH-R) in a series of 122 consecutive patients, including DEL (n=81, 66%), DEL-MYC (n=9, 7%), DEL-BCL2 (n=13, 11%), or high-grade lymphomas (DH/TH) (n=19, 16%). Central nervous system (CNS) prophylaxis included intravenous methotrexate (n=66), intrathecal chemotherapy (IT) (n=40) or no prophylaxis (n=16). Sixty-seven patients (55%) had highintermediate or high International Prognostic Index (IPI) and 30 (25%) had high CNS-IPI. The 2-year progression-free survival (PFS) and overall survival (OS) for the entire study population were 74% and 84%, respectively. There was a trend for inferior OS for DH/TH (2-year OS: 66%, P=0.058) as compared to all the others. The outcome was significantly better for the IPI 0-2 versus IPI 3-5 (OS: 98% vs. 72%, P=0.002). DA-EPOCH-R did not overcome the negative prognostic value of TP53 mutations: 2-year OS of 62% versus 88% (P=0.036) were observed for mutated as compared to wild-type cases, respectively. Systemic CNS prophylaxis conferred a better 2-year OS (94%) as compared to IT or no prophylaxis (76% and 65%, respectively; P=0.008). DA-EPOCH-R treatment resulted in a favorable outcome in patients with DEL and DEL with single rearrangement, whereas those with multiple genetic alterations such as DEL-DH/TH and TP53 mutated cases still have an inferior outcome.

Diagnostic performance of deep learning algorithm for analysis of computed tomography myocardial perfusion.

PURPOSE: To evaluate the diagnostic accuracy of a deep learning (DL) algorithm predicting hemodynamically significant coronary artery disease (CAD) by using a rest dataset of myocardial computed tomography perfusion (CTP) as compared to invasive evaluation. METHODS: One hundred and twelve consecutive symptomatic patients scheduled for clinically indicated invasive coronary angiography (ICA) underwent CCTA plus static stress CTP and ICA with invasive fractional flow reserve (FFR) for stenoses ranging between 30 and 80%. Subsequently, a DL algorithm for the prediction of significant CAD by using the rest dataset (CTP-DLrest) and stress dataset (CTP-DLstress) was developed. The diagnostic accuracy for identification of significant CAD using CCTA, CCTA + CTP stress, CCTA + CTP-DLrest, and CCTA + CTP-DLstress was measured and compared. The time of analysis for CTP stress, CTP-DLrest, and CTP-DLStress was recorded. RESULTS: Patient-specific sensitivity, specificity, NPV, PPV, accuracy, and area under the curve (AUC) of CCTA alone and CCTA + CTPStress were 100%, 33%, 100%, 54%, 63%, 67% and 86%, 89%, 89%, 86%, 88%, 87%, respectively. Patient-specific sensitivity, specificity, NPV, PPV, accuracy, and AUC of CCTA + DLrest and CCTA + DLstress were 100%, 72%, 100%, 74%, 84%, 96% and 93%, 83%, 94%, 81%, 88%, 98%, respectively. All CCTA + CTP stress, CCTA + CTP-DLRest, and CCTA + CTP-DLStress significantly improved detection of hemodynamically significant CAD compared to CCTA alone (p < 0.01). Time of CTP-DL was significantly lower as compared to human analysis (39.2 ± 3.2 vs. 379.6 ± 68.0 s, p < 0.001). CONCLUSION: Evaluation of myocardial ischemia using a DL approach on rest CTP datasets is feasible and accurate. This approach may be a useful gatekeeper prior to CTP stress..

Computed tomography predictors of structural valve degeneration in patients undergoing transcatheter aortic valve implantation with balloon-expandable prostheses.

OBJECTIVES: Computed tomography (CT) provides excellent anatomy assessment of the aortic annulus (AoA) and is utilized for pre-procedural planning of transcatheter aortic valve implantation (TAVI). We sought to investigate if geometrical characteristics of the AoA determined by CT may represent predictors of structural valve degeneration (SVD) in patients undergoing TAVI with balloon-expandable valves. METHODS: This is a retrospective study on 124 consecutive patients (mean age: 79 ± 7 years; female: 61%) undergoing balloon-expandable TAVI prospectively enrolled in a registry. AoA maximum diameter (Dmax), minimum diameter (Dmin), and area were assessed using pre-procedural CT. SVD was identified during follow-up with transthoracic echocardiography documenting structural prosthetic valve abnormalities with or without hemodynamic changes. RESULTS: The mean follow-up was 5.9 ± 1.7 years. SVD was found in 48 out of 124 patients (38%). AoA Dmax, Dmin, and area were significantly smaller in patients with SVD compared to patients without SVD (25.6 ± 2.2 mm vs. 27.1 ± 2.8 mm, p = 0.012; 20.5 ± 2.1 mm vs. 21.8 ± 2.1 mm, p = 0.001 and 419 ± 77 mm2 vs. 467 ± 88 mm2, p = 0.002, respectively). At univariable analysis, female sex, BSA, 23-mm prosthetic valve size, Dmax < 27.1 mm, and a Dmin < 19.9 mm were associated with SVD, whereas at multivariable analysis, only Dmin < 19.9 mm (OR = 2.873, 95% CI: 1.191-6.929, p = 0.019) and female sex (OR = 2.659, 95% CI: 1.095-6.458, p = 0.031) were independent predictors of SVD. CONCLUSIONS: Female sex and AoA Dmin < 19.9 mm are associated with SVD in patients undergoing TAVI with balloon-expandable valves. When implanting large prostheses in order to avoid paraprosthetic regurgitation, caution should be observed due to the risk of excessive stretching of the AoA Dmin, which may play a role in SVD. KEY POINTS: • Long-term durability is a concern for transcatheter aortic valve bioprosthesis. • CT provides an excellent assessment of the aortic annulus's geometrical characteristics for prosthesis sizing before transcatheter aortic valve implantation (TAVI). • Female sex and a small minimum aortic annulus diameter measured with CT are independent predictors of structural valve degeneration in patients undergoing TAVI with balloon-expandable valves.

Nebulization of High-Dose Poractant Alfa in Newborn Piglets on Nasal Continuous Positive Airway Pressure Yields Therapeutic Lung Doses of Phospholipids.

OBJECTIVE: It is not known how much surfactant must be nebulized to reach a lung dose of phospholipids equivalent to that obtained by the instillation of 200 mg/kg of surfactant. We aimed to assess the feasibility of nebulizing a high-dose of poractant alfa with the eFlow-Neos investigational vibrating-membrane nebulizer in newborn piglets on nasal continuous positive airway pressure (nCPAP) and to determine whether this intervention would yield therapeutic lung doses of phospholipids. STUDY DESIGN: Twelve 1-day-old piglets on nCPAP received 600 mg/kg of poractant alfa admixed with technetium-99m via nebulization. Six piglets receiving 200 mg/kg of instilled synthetic surfactant served as controls. Lung deposition (percentage of the nominal dose) was determined by gamma scintigraphy, and the phospholipids' lung dose was calculated. RESULTS: The lung dose of phospholipids (mean ± standard deviation [SD]) was 138 ± 96 mg/kg with nebulization, and 172 ± 24 mg/kg with instillation (p = 0.42). Nebulization took 58 ± 12 minutes. The arterial partial pressure of carbon dioxide increased from 6.7 ± 1.1 to 7.2 ± 1.1 kPa during nebulization (p = 0.04). Cerebral oximetry remained stable, and there was no hemodynamic instability. CONCLUSION: Nebulization was well tolerated, and the mean lung dose of phospholipids was above 100 mg/kg, that is, not different from the instillation group. These experimental findings suggest that it may be feasible to reach therapeutic lung doses of phospholipids by surfactant nebulization during nCPAP. KEY POINTS: · It is not known if effective lung doses of surfactant can be delivered by nebulization.. · Nebulization of high-dose surfactant in newborn piglets on nCPAP was well tolerated.. · A high-dose of nebulized poractant alfa yielded therapeutic lung doses of phospholipids..

Body mass index is not associated with survival outcomes and immune-related adverse events in patients with Hodgkin lymphoma treated with the immune checkpoint inhibitor nivolumab.

BACKGROUND: Overweight and obese patients with solid tumors receiving anti-programmed cell death-1 (PD-1)/PD-ligand-1(PD-L1) immune checkpoint inhibitors exhibit improved survival and higher risk of immune-related adverse events (irAEs) than those with a normal body mass index (BMI). In classic Hodgkin lymphoma (cHL), the impact of BMI on survival and immune-related toxicity is unknown. We evaluated for the first time associations of BMI with survival and irAEs in patients with relapsed/refractory (RR)-cHL undergoing PD-1 blockade. METHODS: Data from a multicenter study on 133 patients treated with the anti-PD1 antibody nivolumab (July 2015-December 2016) were retrieved from a prospective database. Progression-free (PFS), overall survival (OS), incidence and severity of irAEs according to BMI categories were estimated by Kaplan-Meier method, landmark-analyses and Cox regressions. RESULTS: Patients, mostly males (63%, n = 84) with a median age of 35 years (range, 15-82), advanced stage (75%), B symptoms (63%), bulky disease (24%), a median of 4 previous treatments (range, 1-9), received a median of 18 nivolumab doses (range, 1-57). No statistically significant differences across BMI subgroups emerged as to PFS, with 1-year rates of 67.1% for both normal weight (n = 66; 49.6%) and overweight (n = 31; 23.3%) patients. Underweight (n = 12; 9%) and obese (n = 24; 18%) patients had a 1-year PFS of 54.5% and 49%, respectively. In survival analyses, BMI either as a continuous (P = 0.5) or categorical (P for trend = 0.63) variable failed to associate with PFS. Response rates and time-to-response did not cluster in any BMI subset. No BMI-related differences in OS emerged across normal, overweight and obese patients but underweight patients had the worst survival. Occurrence of irAEs of whatever severity did not statistically associate with BMI. CONCLUSIONS: In patients with RR-cHL receiving nivolumab, no statistically significant differences emerged in response rates, PFS and OS across BMI categories of normal weight, overweight and obese. Overweight/obese patients did not display an increased risk of irAEs. The exquisite sensitivity to anti-PD-1 antibodies, the unique cytokine milieu and effector pathways triggered by nivolumab in cHL, may represent biologic 'equalizers' counteracting the immunoregulatory effects of adiposity. Differently from solid tumors, BMI is not associated with treatment efficacy and immune-related toxicity and does not represent a predictive tool for PD-1-targeted immunotherapies in cHL.

Dose-dense ABVD as first-line therapy in early-stage unfavorable Hodgkin lymphoma: results of a prospective, multicenter double-step phase II study by Fondazione Italiana Linfomi.

We investigated the feasibility and activity of an intensified dose-dense ABVD (dd-ABVD) regimen in patients with early-stage unfavorable Hodgkin lymphoma (HL). This prospective, multicenter, phase II study enrolled 96 patients with newly diagnosed, unfavorable stage I or II classical HL. The patients received four cycles of dd-ABVD followed by radiotherapy. Interim PET (PET-2) was mandatory after two courses. Primary endpoints were the evaluation of dd-ABVD feasibility and activity (incidence of PET-2 negativity). The feasibility endpoint was achieved with 48/52 (92.3%) patients receiving > 85% of the programmed dose. The mean dose intensity in the overall patient population (n = 96) was 93.7%, and the median duration of dd-ABVD was 85 days (range, 14-115) versus an expected duration of 84 days. PET-2 was available for 92/96 (95.8%) patients, of whom 79 were PET-2 negative (85.9%). In total, 90 (93.8%) patients showed complete response at the end of treatment. With a follow-up of 80.9 months (3.3-103.2), the median progression-free survival (PFS) and overall survival (OS) were not reached. At 84 months, PFS and OS rates were 88.4% and 95.7%, respectively. No evidence for a difference in PFS or OS was observed for PET-2-negative and PET-2-positive patients. Infections were documented in 8.3% and febrile neutropenia in 6.2% of cases. Four patients died: one had alveolitis at cycle 3, one death was unrelated to treatment, and two died from a secondary cancer. dd-ABVD is feasible and demonstrates activity in early-stage unfavorable HL. The predictive role of PET-2 positivity in early-stage unfavorable HL remains controversial. The study was registered in the EudraCT (reference number, 2011-003,191-36) and the ClinicalTrials.gov (reference number, NCT02247869) databases.

Surfactant lung delivery with LISA and InSurE in adult rabbits with respiratory distress.

BACKGROUND: In preterm infants, InSurE (Intubation-Surfactant-Extubation) and LISA (less invasive surfactant administration) techniques allow for exogenous surfactant administration while reducing lung injury associated with mechanical ventilation. We compared the acute pulmonary response and lung deposition of surfactant by LISA and InSurE in surfactant-depleted adult rabbits. METHODS: Twenty-six spontaneously breathing surfactant-depleted adult rabbits (6-7 weeks old) with moderate RDS and managed with nasal continuous positive airway pressure were randomized to 3 groups: (1) 200 mg/kg of surfactant by InSurE; (2) 200 mg/kg of surfactant by LISA; (3) no surfactant treatment (Control). Gas exchange and lung mechanics were monitored for 180 min. After that, surfactant lung deposition and distribution were evaluated monitoring disaturated-phosphatidylcholine (DSPC) and surfactant protein C (SP-C), respectively. RESULTS: No signs of recovery were found in the untreated animals. After InSurE, oxygenation improved more rapidly compared to LISA. However, at 180' LISA and InSurE showed comparable outcomes in terms of gas exchange, ventilation parameters, and lung mechanics. Neither DSPC in the alveolar pool nor SP-C signal distributions in a frontal lung section were significantly different between InSurE and LISA groups. CONCLUSIONS: In an acute setting, LISA demonstrated efficacy and surfactant lung delivery similar to that of InSurE in surfactant-depleted adult rabbits. IMPACT: Although LISA technique is gaining popularity, there are still several questions to address. This is the first study comparing LISA and InSurE in terms of gas exchange, ventilation parameters, and lung mechanics as well as surfactant deposition and distribution. In our animal study, three hours post-treatment, LISA method seems to be as effective as InSurE and showed similar surfactant lung delivery. Our findings provide some clarifications on a fair comparison between LISA and InSurE techniques, particularly in terms of surfactant delivery. They should reassure some of the concerns raised by the clinical community on LISA adoption in neonatal units.

Sample preparation strategy for the detection of steroid-like compounds using MALDI mass spectrometry imaging: pulmonary distribution of budesonide as a case study.

Corticosteroids as budesonide can be effective in reducing topic inflammation processes in different organs. Therapeutic use of budesonide in respiratory diseases, like asthma, chronic obstructive pulmonary disease, and allergic rhinitis is well known. However, the pulmonary distribution of budesonide is not well understood, mainly due to the difficulties in tracing the molecule in lung samples without the addition of a label. In this paper, we present a matrix-assisted laser desorption/ionization mass spectrometry imaging protocol that can be used to visualize the pulmonary distribution of budesonide administered to a surfactant-depleted adult rabbit. Considering that budesonide is not easily ionized by MALDI, we developed an on-tissue derivatization method with Girard's reagent P followed by ferulic acid deposition as MALDI matrix. Interestingly, this sample preparation protocol results as a very effective strategy to raise the sensitivity towards not only budesonide but also other corticosteroids, allowing us to track its distribution and quantify the drug inside lung samples.

Radiological pitfalls associated with the diagnosis of usual interstitial pneumonia pattern on high-resolution computed tomography and associated findings: experience from a single Italian center.

BACKGROUND: The diagnostic algorithm for idiopathic pulmonary fibrosis (IPF) based on high-resolution computed tomography (HRCT) findings and multidisciplinary discussion (MDD) has been well established. PURPOSE: To identify the causes of disagreement between non-thoracic and thoracic radiologist involved in MDD for the imaging diagnosis of usual interstitial pneumonia (UIP) patterns and associated findings on HRCT and to improve the understanding of IPF by non-expert radiologists through a more systematic approach to HRCT. MATERIAL AND METHODS: This study included 68 patients who underwent MDD for suspected IPF. We compared the first reports generated before MDD by non-expert radiologists with the CT pattern and associated findings of IPF reported by thoracic radiologist involved in MDD. RESULTS: Regarding the diagnosis of CT pattern by non-expert radiologists, 30/68 patients received a discordant diagnosis, and in another 28 reports, all features of the CT pattern were described without reaching a diagnostic conclusion. The first report was concordant in only 10 patients. For 63 cases in which associated findings were reported by expert radiologists in MDD, we documented discrepancies in 47 cases where associated findings were considered absent by the first non-thoracic radiologist. CONCLUSION: We found significant discrepancies in the imaging diagnosis of UIP patterns and associated findings on HRCT between non-expert and thoracic radiologists included in MDD. Therefore, in this study, we analyzed and suggested diagnostic strategies to improve non-expert radiologists' approach to HRCT.

The structure of the Shiga toxin 2a A-subunit dictates the interactions of the toxin with blood components.

Hemolytic uremic syndrome (eHUS) is a severe complication of human infections with Shiga toxins (Stxs)-producing Escherichia coli. A key step in the pathogenesis of eHUS is the interaction of Stxs with blood components before the targeting of renal endothelial cells. Here, we show that a single proteolytic cleavage in the Stx2a A-subunit, resulting into two fragments (A1 and A2) linked by a disulfide bridge (cleaved Stx2a), dictates different binding abilities. Uncleaved Stx2a was confirmed to bind to human neutrophils and to trigger leukocyte/platelet aggregate formation, whereas cleaved Stx2a was ineffective. Conversely, binding of complement factor H was confirmed for cleaved Stx2a and not for uncleaved Stx2a. It is worth noting that uncleaved and cleaved Stx2a showed no differences in cytotoxicity for Vero cells or Raji cells, structural conformation, and contaminating endotoxin. These results have been obtained by comparing two Stx2a batches, purified in different laboratories by using different protocols, termed Stx2a(cl; cleaved toxin, Innsbruck) and Stx2a(uncl; uncleaved toxin, Bologna). Stx2a(uncl) behaved as Stx2a(cl) after mild trypsin treatment. In this light, previous controversial results obtained with purified Stx2a has to be critically re-evaluated; furthermore, characterisation of the structure of circulating Stx2a is mandatory to understand eHUS-pathogenesis and to develop therapeutic approaches.

Prevalence of cranial involvement in a cohort of Italian patients with chronic non-bacterial osteomyelitis.

OBJECTIVES: Chronic non-bacterial osteomyelitis (CNO) is a non-infectious inflammatory disease characterised by uni- or multi-focal bone lytic lesions. CNO mainly affects metaphysis of long bones, pelvis and shoulder girdle. Neurocranium lesions are extremely rare. The objective of the study is to describe the prevalence and clinical manifestations of CNO patients with neurocranium involvement in an Italian cohort of CNO patients. METHODS: This is a retrospective study. Medical records of patients with CNO admitted to eight paediatric rheumatology centres were reviewed. RESULTS: Among 86 patients with CNO enrolled in the study, three of them were female and presented neurocranium involvement - multifocal lesions. Two out of the 3 patients were completely asymptomatic for cranial involvement, while one of the 3 complained of cranial bossing. Cranial involvement was detected with bone scintigraphy and then confirmed by magnetic resonance imaging and/or computed tomography. Two patients presented fever and two with skin manifestations. Laboratory inflammatory markers were increased in two of them. All patients underwent bone biopsy confirming the diagnosis. They all received NSAIDs. Two patients received corticosteroids and then methotrexate and achieved clinical remission, while one patient received pamidronate. CONCLUSIONS: This is the first report of neurocranium involvement in a cohort of patients affected by CNO. In our cohort no patient showed significant signs attributable to cranial involvement.

Dose-Response Study on Surfactant Nebulization Therapy During Nasal Continuous Positive Airway Pressure Ventilation in Spontaneously Breathing Surfactant-Deficient Newborn Piglets.

OBJECTIVES: The current clinical treatment of neonates with respiratory distress syndrome includes endotracheal intubation and intratracheal instillation of exogenous surfactant. Nebulization of surfactant offers an attractive alternative. The aims of this study were to test nebulization as a noninvasive method of administering surfactant and determine the optimal dose for the treatment of respiratory distress syndrome-associated pathophysiology of the neonatal lungs. DESIGN: Prospective, randomized, animal model study. SETTING: An experimental laboratory. SUBJECTS: Thirty-six newborn piglets. INTERVENTIONS: Different doses (100, 200, 400, and 600 mg/kg) of poractant alfa were administered via a vibrating membrane nebulizer (eFlow-Neos; Pari Pharma GmbH, Starnberg, Germany) or a bolus administration using the intubation-surfactant-extubation (Insure) technique (200 mg/kg) to spontaneously breathing newborn piglets (n = 6/group) with bronchoalveolar lavage-induced respiratory distress syndrome during nasal continuous positive airway pressure (180 min). MEASUREMENTS AND MAIN RESULTS: Pulmonary, hemodynamic, and cerebral effects were assessed. Histologic analysis of lung and brain tissue was also performed. After repeated bronchoalveolar lavage, newborn piglets developed severe respiratory distress syndrome. Rapid improvement in pulmonary status was observed in the Insure group, whereas a dose-response effect was observed in nebulized surfactant groups. Nebulized poractant alfa was more effective at doses higher than 100 mg/kg and was associated with similar pulmonary, hemodynamic, and cerebral behavior to that in the Insure group, but improved lung injury scores. CONCLUSIONS: In newborn piglets with severe bronchoalveolar lavage-induced respiratory distress syndrome, our results demonstrate that the administration of nebulized poractant alfa using an investigational customized eFlow-Neos nebulizer is an effective and safe noninvasive surfactant administration technique.

Computed tomography (CT)-derived radiomic features differentiate prevascular mediastinum masses as thymic neoplasms versus lymphomas.

OBJECTIVES: We aimed to assess the ability of radiomics, applied to not-enhanced computed tomography (CT), to differentiate mediastinal masses as thymic neoplasms vs lymphomas. METHODS: The present study was an observational retrospective trial. Inclusion criteria were pathology-proven thymic neoplasia or lymphoma with mediastinal localization, availability of CT. Exclusion criteria were age < 16 years and mediastinal lymphoma lesion < 4 cm. We selected 108 patients (M:F = 47:61, median age 48 years, range 17-79) and divided them into a training and a validation group. Radiomic features were used as predictors in linear discriminant analysis. We built different radiomic models considering segmentation software and resampling setting. Clinical variables were used as predictors to build a clinical model. Scoring metrics included sensitivity, specificity, accuracy and area under the curve (AUC). Wilcoxon paired test was used to compare the AUCs. RESULTS: Fifty-five patients were affected by thymic neoplasia and 53 by lymphoma. In the validation analysis, the best radiomics model sensitivity, specificity, accuracy and AUC resulted 76.2 ± 7.0, 77.8 ± 5.5, 76.9 ± 6.0 and 0.84 ± 0.06, respectively. In the validation analysis of the clinical model, the same metrics resulted 95.2 ± 7.0, 88.9 ± 8.9, 92.3 ± 8.5 and 0.98 ± 0.07, respectively. The AUCs of the best radiomic and the clinical model not differed. CONCLUSIONS: We developed and validated a CT-based radiomic model able to differentiate mediastinal masses on non-contrast-enhanced images, as thymic neoplasms or lymphoma. The proposed method was not affected by image postprocessing. Therefore, the present image-derived method has the potential to noninvasively support diagnosis in patients with prevascular mediastinal masses with major impact on management of asymptomatic cases.

Multiple Skin Squamous Cell Carcinomas in Junctional Epidermolysis Bullosa Due to Altered Laminin-332 Function.

Variably reduced expression of the basement membrane component laminin-332 (α3aß3γ2) causes junctional epidermolysis bullosa generalized intermediate (JEB-GI), a skin fragility disorder with an increased susceptibility to squamous cell carcinoma (SCC) development in adulthood. Laminin-332 is highly expressed in several types of epithelial tumors and is central to signaling pathways that promote SCC tumorigenesis. However, laminin-332 mutations and expression in individuals affected by JEB-GI and suffering from recurrent SCCs have been poorly characterized. We studied a JEB-GI patient who developed over a hundred primary cutaneous SCCs. Molecular analysis combined with gene expression studies in patient skin and primary keratinocytes revealed that the patient is a functional hemizygous for the p.Cys1171* mutant allele which is transcribed in a stable mRNA encoding for a ß3 chain shortened of the last two C-terminal amino acids (Cys1171-Lys1172). The lack of the Cys1171 residue involved in the C-terminal disulphide bond to γ2 chain did not prevent assembly, secretion, and proteolytic processing of the heterotrimeric molecule. Immunohistochemistry of SCC specimens revealed accumulation of mutant laminin-332 at the epithelial-stromal interface of invasive front. We conclude that the C-terminal disulphide bond is a structural element crucial for laminin-332 adhesion function in-vivo. By saving laminin-332 amount, processing, and signaling role the p.Cys1171* mutation may allow intrinsic pro-tumorigenic properties of the protein to be conveyed, thus contributing to invasiveness and recurrence of SCCs in this patient.