Fate of melatonin orally administered in preterm newborns: Antioxidant performance and basis for neuroprotection.

Preterm infants cannot counteract excessive reactive oxygen species (ROS) production due to preterm birth, leading to an excess of lipid peroxidation with malondialdehyde (MDA) production, capable of contributing to brain damage. Melatonin (ME), an endogenous brain hormone, and its metabolites, act as a free radical scavenger against ROS. Unfortunately, preterms have an impaired antioxidant system, resulting in the inability to produce and release ME. This prospective, multicenter, parallel groups, randomized, double-blind, placebo-controlled trial aimed to assess: (i) the endogenous production of ME in very preterm infants (gestational age ≤ 29 + 6 WE, 28 infants in the ME and 26 in the placebo group); (ii) the exogenous hormone availability and its metabolization to the main metabolite, 6-OH-ME after 15 days of ME oral treatment; (iii) difference of MDA plasma concentration, as peroxidation marker, after treatment. Blood was collected before the first administration (T1) and after 15 days of administration (T2). ME and 6-OH-ME were detected by liquid chromatography tandem mass spectrometry, MDA was measured by liquid chromatograph with fluorescence detection. ME and 6-OH-ME were not detectable in the placebo group at any study time-point. ME was absent in the active group at T1. In contrast, after oral administration, ME and 6-OH-ME resulted highly detectable and the difference between concentrations T2 versus T1 was statistically significant, as well as the difference between treated and placebo groups at T2. MDA levels seemed stable during the 15 days of treatment in both groups. Nevertheless, a trend in the percentage of neonates with reduced MDA concentration at T2/T1 was 48.1% in the ME group versus 38.5% in the placebo group. We demonstrated that very preterm infants are not able to produce endogenous detectable plasma levels of ME during their first days of life. Still, following ME oral administration, appreciable amounts of ME and 6-OH-ME were available. The trend of MDA reduction in the active group requires further clinical trials to fix the dosage, the length of ME therapy and to identify more appropriate indexes to demonstrate, at biological and clinical levels, the antioxidant activity and consequent neuroprotectant potential of ME in very preterm newborns.

PEG 400 Ion Suppression in Busulfan Detection by High-Performance Liquid Chromatography-Tandem Mass Spectrometry.

BACKGROUND: Busulfan (Bu), an alkylating agent commonly used in chemotherapy and transplantation, exhibits high intraindividual pharmacokinetic variability and possible time-dependent variations in clearance, which complicate therapeutic drug monitoring. Numerous analytical methods have been developed to reduce analysis time and facilitate timely decision-making regarding treatment changes; however, the validation procedures rarely involve analysis of potentially interfering excipients. Macrogol 400 (PEG 400) should be considered as a possible interfering agent in the detection of plasma Bu levels, especially as an ionization suppressor. METHODS: Six intravenous formulations of Bu were compared with identify at least 1 common excipient (PEG 400). During the 176 therapeutic drug monitoring analyses of Bu, one of the PEG 400 specific mass-to-charge ratio transitions was determined using an instrumental method. After coelution with Bu and its internal standard (Bu-d8) was confirmed, all analyses were repeated using a different experimental setup free of ion suppression induced by PEG. The concentration-time profile of PEG 400 was also analyzed. RESULTS: The area under the curve obtained from the 2 data sets was compared and analyzed using Lin concordance correlation coefficient and Bland-Altman plot analysis. The results from the 2 analytical methods were comparable: PEG 400 negatively affected the Bu-d8 coefficient of variation but not the Bu/Bu-d8 ratio. CONCLUSIONS: The possible interference of PEG 400 should be thoroughly investigated, especially with respect to analytical methods that cannot be supported by correction of the stable isotopically labeled internal standard analog.

Prospective urinary albumin/creatinine ratio for diagnosis, staging, and organ response assessment in renal AL amyloidosis: results from a large cohort of patients.

OBJECTIVES: Quantification of 24 h-proteinuria is the gold standard for diagnosing, staging, and monitoring of patients with renal AL amyloidosis. However, 24 h-urine collection is cumbersome and may result in preanalytical error. In this prospective study, we investigated the role of urinary albumin/creatinine ratio (UACR) (cut-off: 300 mg/g) identifying renal involvement, evaluated a UACR-based staging system (UACR cut-off: 3,600 mg/g) and assessed whether UACR response (UACR decrease >30% without worsening in eGFR >25%) predicts renal outcome in 531 patients with newly-diagnosed AL amyloidosis. METHODS: From October 2013 paired 24 h-proteinuria and UACR (on first morning void) were measured in all newly-diagnosed patients with AL amyloidosis. Correlation between 24 h-proteinuria and UACR at baseline was assessed by Pearson's r test. Impact of UACR response on renal outcome was assessed in randomly created testing (n=354) and validation (n=177) cohorts. RESULTS: A strong linear correlation was found between 24 h-proteinuria and UACR at baseline (r=0.90; p<0.001). After a median follow-up of 31 months, 57 (11%) patients required dialysis. A UACR-based renal staging system identified three stages with significantly higher dialysis rate at 36 months comparing stage I with stage II and stage II with stage III. Achieving a renal response, according to a UACR-based criterion, resulted in lower dialysis rate in both testing and validation cohorts. CONCLUSIONS: UACR is a reliable marker for diagnosis, prognosis, and organ response assessment in renal AL amyloidosis and can reliably replace 24 h-proteinuria in clinical trials and individual patients' management.

Hemoperfusion with CytoSorb as Adjuvant Therapy in Critically Ill Patients with SARS-CoV2 Pneumonia.

We report a preliminary experience of adjuvant therapy with Hemoperfusion (HP) in patients with Severe Acute Respiratory Syndrome-CoronaVirus 2 (SARS-CoV2) pneumonia. Currently, there are no approved treatments for CoronaVirus Disease 19 (COVID-19); however, therapeutic strategies based on the preclinical evidence include supportive measures, such as oxygen supplementation, antiviral, and anticoagulant agents. Despite these treatments, 10% of patients worsen and develop severe acute respiratory distress syndrome (ARDS). Since the pathogenic mechanism of ARDS is an uncontrolled inflammatory state, we speculate that removing inflammation effectors from blood may contrast tissue injury and improve clinical outcome. In a scenario of dramatic medical emergency, we conducted an observational study on 9 consecutive patients hospitalized in COVID Intensive Care Unit, where 5 of 9 consecutive patients were treated with HP, due to the emergency overload made it impossible to deliver blood purification in the other 4 patients. COVID-19 was diagnosed through the identification of virus sequences by reverse transcription-PCR on respiratory specimens. All patients had severe pneumonia requiring continuous positive airway pressure. HP was started in all patients 6-7 days after hospital admission. The treated patients (T) received 2 consecutive sessions of HP using CytoSorb cartridge. Our results show a better clinical course of T compared to control patients (C), in fact all T except 1 survived, and only 2 of them were intubated, while all C required intubation and died. Lymphocytopenia worsened in C but not in T. C-reactive protein decreased in both patients, but to a greater extent in T. IL-6, IL-8, and TNF-α decreased after HP, IL-10 did not change. Respiratory function remained stable and did not worsen in T compared to C. The limited sample size and observational study design preclude a sound statement about the potential effectiveness of HP in COVID-19 patients, but our experience suggests a potential therapeutic role of adjuvant CytoSorb HP in the early course of CO-VID-19 pneumonia. A randomized clinical trial is ongoing.

Insulin resistance and potential modulators of ovarian reserve in young reproductive-aged women with obesity and type 1 diabetes.

INTRODUCTION: Both obesity and diabetes play a significant role in reproductive disorders in women and insulin resistance (IR) is a confirmed trait d'union. We evaluated the relationship between IR and an established ovarian reserve biomarker such as anti-mullerian hormone (AMH) together with other potential modulators of ovarian physiology (adiponectin and kisspeptin) in young reproductive-aged group women with obesity and type 1 diabetes (T1D). PATIENTS AND METHODS: We recruited 32 female youths: 14 of them presented with T1D (14.6 ± 2.6 years) and 18 with obesity (15.1 ± 2.6 years). The control group included 20 age-matched normal weight females. Each patient underwent physical examination and hormonal assessment. AMH, kisspeptin and adiponectin levels were also measured. IR was calculated as the homeostasis model assessment for insulin resistance (HOMA-IR) and the glucose disposal rate (eGDR) in patients with obesity and with T1D, respectively. RESULTS: adiponectin and kisspeptin levels were significantly different into groups (p ≤ .001), whereas AMH levels were not. Adiponectin values were higher in controls compared to patients with obesity (p < .001) and T1D (p = .02). Kisspeptin levels were lower in controls compared to patients with obesity (p = .001), without reaching statistical significance when compared to T1D (p = .06). IR was associated with lower adiponectin and higher kisspeptin levels (p < .001 and p = .02, respectively), but not with AMH. CONCLUSIONS: IR displays a relationship with adiponectin and kisspeptin in young reproductive-aged women with obesity and T1D. Interventions to correct IR in adolescents could be part of an early approach to prevent reproductive disorders and to promote factors associated with longevity in adult women.

Management of squamous cell carcinoma of the temporal bone: long-term results and factors influencing outcomes.

OBJECTIVE: Temporal bone squamous cell carcinoma (TBSCC) is a rare, aggressive tumor. Surgery, alone or combined with radiotherapy, represent the mainstay of treatment. To report our experience in the treatment of TBSCC and evaluate the disease-specific survival, identifying the factors influencing this outcome. MATERIALS AND METHODS: A retrospective study was performed on 66 patients between 1993 and 2018. Patients were staged according to the University of Pittsburgh-modified TNM staging system. Nine cases (13.6%) were Stage I, 7 cases (10.6%) Stage II, 20 cases (30.3%) Stage III and 30 cases (45.5%) Stage IV. Twenty-four patients underwent lateral temporal bone resection (LTBR) and 42 patients underwent subtotal temporal bone resection (STBR). RESULTS: One hundred percent of Stage I and II patients showed no evidence of disease (NED) after a median follow-up of 101 months (range 1-289 months). NED resulted in 88.2% of Stage III (mean follow-up 80.3 months; range 8-257) and 46.4% of stage IV (mean follow-up 50.6 months; range 3-217). Pittsburgh Stage or involvement of mastoid, facial nerve, medial wall of the middle ear, temporomandibular joint and middle fossa dura emerged as negative prognostic factors. The highest mortality rate occurred in the first 2 years after treatment, due to local recurrence. CONCLUSIONS: Prognosis of TBSCC can be excellent in early stage tumors, employing a LTBR. In more advanced cases, prognosis is poor. STBR with adjuvant radiotherapy represents the treatment of choice, offering acceptable survival rates. Given the rarity of the pathology, many controversies still exist concerning optimal management.

Early experience in tracheostomy and tracheostomy tube management in Covid-19 patients.

In Italy, we have experienced Europe's first and largest coronavirus outbreak. Based on our preliminary experience, we discuss the challenges in performing tracheotomy and tracheostoma care in the setting of a new pathogen.

Significance of serum Myostatin in hemodialysis patients.

BACKGROUND: Malnutrition and muscle wasting are common in haemodialysis (HD) patients. Their pathogenesis is complex and involves many molecules including Myostatin (Mstn), which acts as a negative regulator of skeletal muscle. The characterisation of Mstn as a biomarker of malnutrition could be useful in the prevention and management of this condition. Previous studies have reported no conclusive results on the actual relationship between serum Mstn and wasting and malnutrition. So, in this study, we evaluated Mstn profile in a cohort of regular HD patients. METHODS: We performed a cross-sectional study, enrolling 37 patients undergoing bicarbonate-HD (BHD) or haemodiafiltration (HDF) at least for six months. 20 sex-matched healthy subjects comprised the control group. Mstn serum levels were evaluated by ELISA before and after HD. We collected clinical and biochemical data, evaluated insulin resistance, body composition, malnutrition [by Malnutrition Inflammation Score (MIS)] and tested muscle function (by hand-grip strength, six-minute walking test and a questionnaire on fatigue). RESULTS: Mstn levels were not significantly different between HD patients and controls (4.7 ± 2.8 vs 4.5 ± 1.3 ng/ml). In addition, while a decrease in Mstn was observed after HD treatment, there were no differences between BHD and HDF. In whole group of HD patients Mstn was positively correlated with muscle mass (r = 0.82, p < 0.001) and inversely correlated with age (r = - 0.63, p < 0.01) and MIS (r = - 0.39, p = 0.01). No correlations were found between Mstn and insulin resistance, such as between Mstn levels and parameters of muscle strength and fatigue. In multivariate analysis, Mstn resulted inversely correlated with fat body content (ß = - 1.055, p = 0.002). CONCLUSIONS: Circulating Mstn is related to muscle mass and nutritional status in HD patients, suggesting that it may have a role in the regulation of skeletal muscle and metabolic processes. However, also considering the lack of difference of serum Mstn between healthy controls and HD patients and the absence of correlations with muscle function tests, our findings do not support the use of circulating Mstn as a biomarker of muscle wasting and malnutrition in HD.

Diet and Lifestyle Role in Homocysteine Metabolism in Turner Syndrome.

OBJECTIVE: Patients with Turner syndrome (TS) have an unfavorable cardiometabolic profile. Hyperhomocysteinemia is a potential cardiovascular risk factor influenced by genetic and environmental factors, therapies, unbalanced diets and other lifestyle factors. We retrospectively studied the relationship between total plasma homocysteine (Hcy), serum vitamin B12 (B12) and folate concentration in TS patients, taking into account the genetic profile, diet, smoking habits, hormonal therapies and dietary supplements of the subjects. PATIENTS AND METHODS: We evaluated 50 TS patients (31.5 ± 12.5 years). Medication, including vitamin supplementation, was obtained. Eating habits, cigarette smoking, alcohol and coffee consumption were investigated using phone interviews. Levels of Hcy metabolism parameters were classified by using the relevant cutoff value for an adult population and compared with a reference sample drawn from the general population. RESULTS: Inadequate Hcy and B12 levels were noted, despite vitamin supplementation. Holotranscobalamin (HoloTC) was above the relevant cutoff in the population, and supplemented subjects showed mean levels lower than nonsupplemented subjects (p = 0.005). Dietary supplementation (p = 0.038), lifestyle (coffee consumption, p = 0.01) and hormonal replacement therapy (p = 0.02) are important factors for Hcy metabolism. No genetic influence on Hcy levels was noted. Multivariable regression analysis identified vitamin supplementation (p = 0.045) as the only independent predictor of increased Hcy levels. CONCLUSION: Cardiovascular risk in TS can be reduced using educational approaches to a healthy lifestyle with dietary guidelines. Besides this, we also recommend measuring HoloTC for the prompt detection of B12 deficiency and to consider hormone replacement therapy in the biochemical assessment of homocysteine in TS.

Epicardial Fat Thickness in Non-Obese Neurologically Impaired Children: Association with Unfavorable Cardiometabolic Risk Profile.

BACKGROUND: Cardiovascular risk is reported in disabled children and epicardial fat (EF) is considered an independent predictor of cardiovascular disease (CVD). No data on the EF thickness (EFT) evaluation in disabled children have been published. OBJECTIVE: We investigated EFT in neurologically impaired (NI) children; its relationship with their metabolic profile was also considered. METHODS: Clinical data, body composition estimation, biochemical profile, and ultrasound-measured EFT were performed in 32 disabled patients (12.4 ± 6.3 years). Pathological parameters were defined using the following criteria: waist circumference >95th percentile, waist to height ratio (WHtR) >0.5, total cholesterol and triglycerides (TG) values >95th percentile, high density lipoprotein cholesterol <5th percentile, fasting blood glucose >100 mg/dL, homeostasis model assessment for insulin resistance (HOMA) >97.5th percentile, and EFT >3.6 mm. RESULTS: EFT values in NI children were higher compared with control group values (p = 0.02). EFT correlated with gender (p < 0.001), age (p = 0.02), pubertal stage (p = 0.04), as well as WHtR (p = 0.03). A correlation between EFT and leptin was also noted (p = 0.04). EFT levels significantly correlated with pathological TG (p = 0.01) and HOMA-IR (p = 0.04). CONCLUSIONS: Higher EFT was observed in NI children compared with controls. EFT values correlated with clinical, metabolic, and endocrinological parameters. Ultrasound-measured EFT could be used to promptly detect subclinical CVD and to prevent adverse outcomes in disabled children.

Circulating free light chain measurement in the diagnosis, prognostic assessment and evaluation of response of AL amyloidosis: comparison of Freelite and N latex FLC assays.

BACKGROUND: The measurement of circulating free light chain (FLC) is essential in the diagnosis, prognostic stratification and evaluation of response to therapy in light chain (AL) amyloidosis. For more than 10 years, this has been done with an immunonephelometric assay based on polyclonal antibodies (Freelite), and cutoffs for staging and response assessment have been validated with this method. Recently, a new assay based on monoclonal antibodies (N latex FLC) has been marketed in Europe. METHODS: We evaluated and compared the clinical performance of the two assays in 426 patients with newly diagnosed AL amyloidosis. RESULTS: We found suboptimal agreement between the two methods, with differences between values obtained with the Freelite and N latex FLC assays increasing with the concentration of clonal FLC. The diagnostic sensitivity of the Freelite (82%) and N latex FLC (84%) assays was similar, and both improved to 98% in combination with serum and urine immunofixation. The concentration of FLC measured with both methods had prognostic significance. Less pronounced decreases in FLC best predicted improved survival with the N latex FLC assay (33% vs. 50%), and there was poor concordance (84%) in discrimination of responders. CONCLUSIONS: The two assays have similar diagnostic and prognostic performance. However, they are not interchangeable, and follow-up should be done with either one. New response criteria are needed for the N latex FLC assay.

A staging system for renal outcome and early markers of renal response to chemotherapy in AL amyloidosis.

The kidney is involved in 70% of patients with immunoglobulin light-chain (AL) amyloidosis, but little is known on progression or reversibility of renal involvement, and criteria for renal response have never been validated. Newly diagnosed patients from the Pavia (n = 461, testing cohort) and Heidelberg (n = 271, validation cohort) centers were included. Proteinuria >5 g/24 h and estimated glomerular filtration rate (eGFR) <50 mL/min predicted progression to dialysis best. Proteinuria below and eGFR above the thresholds indicated low risk (0 and 4% at 3 years in the testing and validation cohorts, respectively). High proteinuria and low eGFR indicated high risk (60% and 85% at 3 years). At 6 months, a ≥25% eGFR decrease predicted poor renal survival in both cohorts and was adopted as criterion for renal progression. A decrease in proteinuria by ≥30% or below 0.5 g/24 h without renal progression was the criterion for renal response, being associated with longer renal survival in the testing and validation populations. Hematologic very good partial or complete remission at 6 months improved renal outcome in both populations. We identified and validated a staging system for renal involvement and criteria for early assessment of renal response and progression in AL amyloidosis that should be used in clinical practice and trial design.

The impact of renal function on the clinical performance of FLC measurement in AL amyloidosis.

BACKGROUND: The measurement of circulating free light chains (FLC) is of utmost importance in immunoglobulin light chain (AL) amyloidosis, being a fundamental part of the diagnostic workup, prognostic stratification and assessment of response to therapy. Renal failure is a common feature of AL amyloidosis and can considerably affect the concentration of FLC. METHODS: We assessed the impact of renal failure on the clinical performance of the Freelite assay in 982 consecutive, newly diagnosed patients with AL amyloidosis, 822 with estimated glomerular filtration rate (eGFR) ≥30 mL/min/1.73 m2, and 160 with eGFR <30 mL/min/1.73 m2. RESULTS: The diagnostic sensitivity of the κ/λ FLC ratio was lower for λ amyloidogenic FLC in patients with renal failure (81% vs. 60%, p<0.001) and the FLC concentration had no independent prognostic significance in patients with severe renal dysfunction. However, FLC response to chemotherapy could still discriminate patients with better outcome. CONCLUSIONS: Renal failure is a relevant interference factor when using the Freelite assay for the identification of the amyloidogenic light chain and for prognostic assessment in patients with AL amyloidosis and renal failure.

A laboratory score in the diagnosis of autoimmune atrophic gastritis: a prospective study.

BACKGROUND: Several biomarkers have been proposed for the diagnosis of autoimmune atrophic gastritis (AAG), but at the present there is no appropriate testing strategy for the disease. GOALS: The aim of this study was to develop and validate a laboratory score able to address the diagnosis of AAG in a general practice setting. STUDY: We prospectively evaluated a number of serum biomarkers (vitamin B12, mean corpuscular volume, hemoglobin, gastrin, and chromogranin A levels) in a case-control population and built 2 biochemical scores, the first with all the parameters [Global Score (GS)], and the second as the best statistical combination of them [Simple Score (SS)]. In the second phase we validated the score that proved to be more efficient on a random population referred to our center (Gastroenterology Outpatient Clinic). RESULTS: Both models turned out to be reliable in detecting patients with suspected AAG, showing excellent accuracy [area under the receiver operating curve (AUC-ROC) 0.94; 95% confidence interval (CI), 0.91-0.97 for GS and AUC-ROC 0.93; 95% CI, 0.89-0.86 for SS]. The SS proved to be more convenient because of its accessibility and availability in a general setting and its low cost. The validation of the SS showed a sensitivity of 85.7% (95% CI, 57.2-98.2) and a specificity of 83.7% (95% CI, 74.2-90.89). CONCLUSIONS: Herein, we describe 2 nonexpensive and reliable score models, particularly the SS, that can be applied in daily medical practice for identifying patients potentially affected by AAG.

Epidemiological characteristics of bloodstream infections in patients with different degrees of liver disease.

Observational retrospective study to evaluate the etiology, the outcome and the risk factors of bloodstream infections (BSIs) in patients with liver disease. One hundred and forty-eight BSIs were diagnosed (infection rate: 0.60 per 100 days of hospital stay), 62 BSIs (41.9 %) were associated with Gram-positive bacteria (infection rate: 0.25 per 100 days of hospital stay) and 80 (54.4 %) with Gram-negative bacteria (infection rate: 0.32 per 100 days of hospital stay). Admission-associated mortality was higher in patients with BSI than in those without BSI (20.6 versus 5.0 %, p < 0.001). Patients with cirrhosis had an increased risk to develop a BSI compared with patients with chronic hepatitis, specifically for Gram-positive (and Staphylococcus spp)-related BSI.

Biochemical markers in early diagnosis and management of systemic amyloidoses.

Systemic amyloid diseases are characterized by widespread protein deposition as amyloid fibrils. Precise diagnostic framing is the prerequisite for a correct management of patients. This complex process is achieved through a series of steps, which include detection of the tissue amyloid deposits, identification of the amyloid type, demonstration of the amyloidogenic precursor, and evaluation of organ dysfunction/damage. Laboratory medicine plays a central role in the diagnosis and management of systemic amyloidoses, through the quantification of the amyloidogenic precursor and evaluation of end-organ damage using biomarkers.

Inappropriately low hepcidin levels in patients with myelodysplastic syndrome carrying a somatic mutation of SF3B1.

Somatic mutations of the RNA splicing machinery have been recently identified in myelodysplastic syndromes. In particular, a strong association has been found between SF3B1 mutation and refractory anemia with ring sideroblasts, a condition characterized by ineffective erythropoiesis and parenchymal iron overload. We studied the relationship between SF3B1 mutation, erythroid activity and hepcidin levels in myelodysplastic syndrome patients. Erythroid activity was evaluated through the proportion of marrow erythroblasts, soluble transferrin receptor and serum growth differentiation factor 15. Significant relationships were found between SF3B1 mutation and marrow erythroblasts (P=0.001), soluble transferrin receptor (P=0.003) and serum growth differentiation factor 15 (P=0.033). Serum hepcidin varied considerably, and multivariable analysis showed that the hepcidin to ferritin ratio, a measure of adequacy of hepcidin levels relative to body iron stores, was inversely related to the SF3B1 mutation (P=0.013). These observations suggest that patients with SF3B1 mutation have inappropriately low hepcidin levels, which may explain their propensity to parenchymal iron loading.