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Total phallic construction with radial artery forearm free-flap (RAFFF) is widely regarded as the gold standard approach for phalloplasty. However, donor-site morbidity remains a significant concern, which is typically managed by using a full-thickness skin graft (FTSG) on the forearm. Split thickness skin grafts (STSG) have been proposed as an alternative, along with the use of an acellular dermal matrix substitute. A retrospective comparative analysis was performed to assess the differences in operative, functional and cosmetic outcomes between FTSG (Group A) and the combination of acellular dermal matrix with STSG (Group B). A retrospective cohort study was conducted on all patients who underwent total phallic construction with RAFFF, between 2016 and 2021. Post-operative surgical and functional outcomes were evaluated using validated tools. A total of 34 patients were included in the study, with 18 patients (52.9%) in Group A and 16 patients (47.1%) in Group B. Group B demonstrated a significant advantage in terms of healing time (24 days vs. 30 days, p = 0.003) and complete graft take (93.8% vs. 27.8%, p = 0.001). Group B also had significantly shorter operative times (310 min vs. 447 min, p = 0.001) and a reduced median hospital stay (8 days vs. 10 days, p = 0.001). Satisfaction with cosmesis was significantly higher in Group B (93.8% vs. 66.7%, p = 0.048)., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

The construction history of a site is partially preserved underground and can be revealed through archaeological investigations, including excavations, integrated with earth observation (EO) methods and technologies that make it possible to overcome some operational limits regarding the areal dimensions and the investigation depths along with the invasiveness of the excavations themselves. An integrated approach based on EO and archaeological records has been applied to improve the knowledge of Machu Picchu. The attention has been focused on the first construction phase of Machu Picchu, and for this reason the investigations were directed to the imaging and characterization of the subsoil of the Plaza principal, considered the core of the whole archaeological area. Archaeological records and multiscale remote sensing (including satellite, UAS, and geophysical surveys) enabled the identification and characterization of the first construction phase of the site, including the preparation phases before building Machu Picchu. The interpretative hypothesis on the constructive history of Machu Picchu started from the identification and use of the quarry, followed by the planification and set of the drainage systems and by the next steps based on diverse reshaping phases of what would be the central plaza., (© 2023. Springer Nature Limited.)

Objective: The treatment of invasive penile cancer is based on partial or total penectomy and perineal urethrostomy configuration. 1-3 To report surgical and functional outcomes of penile amputation and perineal urethrostomy configuration in a consecutive series of patients with invasive penile cancer., Materials and Methods: A single-center retrospective analysis was conducted from January 2018 to December 2022. Data were extracted from clinical records and operative notes. As primary outcomes, surgical complications were described. Oncological outcomes through cancer-specific survival and overall survival were analyzed. Patient survival was estimated by a Kaplan-Meier analysis. Functional outcomes were assessed through the administration of the International Prostatic Symptoms Score (IPSS) questionnaire at 3 months follow-up., Results: Overall 10 patients were enrolled. The median follow-up was 16 months (Interquartile range (IQR 12-18)). The median age was 71 years (IQR 63-79). Operative time was set at 195 minutes (IQR 155-275). The median hospital stay was 8 days (IQR: 6-10). Postoperative complications occurred in 20% of patients, in 1 case surgical revision was necessary. Positive surgical margins were detected only in 1 patient. The median catheterization time was 15 days (IQR: 15-32). One patient developed local recurrence. At 1 year, the cancer-specific survival was 80% and overall survival was 60%. The median preoperative IPSS was 15 (IQR 12-19). The median postoperative IPSS was 6 (IQR 5-7)., Conclusion: Penile amputation and perineal urethrostomy configuration was demonstrated to be a safe and effective procedure to address invasive penile cancer., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background and Objectives : The peak of incidence of testicular cancer (TC) occurs among individuals in their reproductive age, emphasizing the importance of fertility preservation as an integral aspect of disease management. Sperm cryopreservation performed before orchiectomy is ineffective in azoospermic men, necessitating alternative approaches such as microdissection testicular sperm extraction (mTESE) at the time of orchiectomy (onco-mTESE) to obtain viable sperm. This study presents the findings from our institution's experience with onco-mTESE and critically discusses our results in light of the existing body of literature. Materials and Methods : This is a tertiary center retrospective analysis of onco-mTESE procedures performed at a single center between December 2011 and July 2022. The included patients were post-puberal men with testicular tumors requiring orchiectomy, along with concomitant severe oligozoospermia or azoospermia. Bilateral mTESE was performed in all cases. Surgical outcomes, sperm retrieval rates, the usage of preserved viable sperm, assistive reproductive techniques' results, and post-operative serum testosterone were recorded. Results : A total of nine patients were included, with a median age of 34 (IQR 29-36) years. All patients had germ cell tumors (GCTs), with seminomatous and non-seminomatous GCTs accounting for 44.4% ( n = 4) and 55.6% ( n = 5) of patients, respectively. Sperm retrieval occurred in three (33%) patients: one patient in the ipsilateral testis, one in the contralateral testis, and one in both testes. No complications were reported during the procedure, and no post-operative hypogonadism was observed. Among the three patients with successful sperm retrieval, an intracytoplasmic sperm injection (ICSI) was performed in two patients, resulting in two pregnancies, leading to one healthy live birth and one miscarriage. Conclusions : In the context of TC, it is essential to conduct a thorough evaluation of testicular function, including a semen analysis and cryopreservation. Onco-mTESE has proven its safety in preserving fertility in azoospermic cases while ensuring the efficacy of oncological treatment.

Background: Antibodies against cationic platelet chemokine, platelet factor 4 (PF4/CXCL4), have been described in heparin-induced thrombocytopenia (HIT), but also in patients positive for antiphospholipid antibodies (aPL) even in the absence of heparin treatment and HIT-related clinical manifestations. Anti-PF4 antibodies have been recently described also in subjects who developed thrombosis with thrombocytopenia syndrome (TTS) in association with adenoviral vector-based, but not with mRNA-based, COVID-19 vaccines., Objective: To investigate whether COVID-19 vaccination affects the production of anti-PF4 antibodies in aPL-positive patients and in control groups., Methods: Anti-PF4 immunoglobulins were detected in patients' and controls' serum samples by ELISA and their ability to activate normal platelets was assessed by the platelet aggregation test., Results: Anti-PF4 were found in 9 of 126 aPL-positive patients, 4 of 50 patients with COVID-19, 9 of 49 with other infections, and 1 of 50 aPL-negative patients with systemic lupus erythematosus. Clinical manifestations of TTS were not observed in any aPL patient positive for anti-PF4, whose serum failed to cause platelet aggregation. The administration of COVID-19 vaccines did not affect the production of anti-PF4 immunoglobulins or their ability to cause platelet aggregation in 44 aPL-positive patients tested before and after vaccination., Conclusions: Heparin treatment-independent anti-PF4 antibodies can be found in aPL-positive patients and asymptomatic carriers, but their presence, titre as well as in vitro effect on platelet activation are not affected by COVID-19 vaccination., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Mild asthma is prevalent in childhood and causes as many as 30%-40% asthma exacerbations requiring emergency visits. The management of "intermittent" and "mild persistent" asthma phenotypes is still a matter of debate, even if the role of inhaled corticosteroids, both continuous and intermittent, is a cornerstone in this field. Recent updates of the guidelines on the strategies to manage these patients are coming, since the role of inflammation in these asthma phenotypes is crucial, as well as the potential side effect and risks of short-acting beta 2 agonists overuse, prescribed as the only "as-needed" treatments. In this paper, we overview the new (r)evolution regarding intermittent and mild persistent asthma management., (© 2022 The Authors. Pediatric Allergy and Immunology published by European Academy of Allergy and Clinical Immunology and John Wiley & Sons Ltd.)

In the last few years much attention has been focused on research on severe asthma and the role of biologicals in its treatment, also in children. However, mild asthma is way more common in childhood and still causes as many as 30-40% of asthma exacerbations requiring emergency consultation. The management of "intermittent" and "mild persistent" asthma phenotypes is still a matter of debate, even if the role of inhaled corticosteroids, both continuous and intermittent, is a cornerstone in this field. Nevertheless, updates on the strategies to manage these patients are coming, since evidence emerged on the role of inflammation also in these asthma phenotypes as well as on the potential side effect and risks of short-acting beta 2 agonists overuse, which is common in patients for which they have been prescribed as the only as-needed treatment. Unsurprisingly, international guidelines, including GINA, are starting to recommend associating a corticosteroid when using a reliver. In this paper we overview the (r)evolution regarding the management of intermittent and mild persistent asthma. We also focus on the importance of knowing the chemical and physical characteristics of drugs and inhaler devices in order to optimize the treatment and reach the distal airways, as well as of trying to achieve a good compliance to treatments, especially in adolescents, for which it is currently possible to rely also on new digital health technologies.

Low-dose aspirin (ASA) is used to prevent cardiovascular events. The most commonly used formulation is enteric-coated ASA (EC-ASA) that may be absorbed more slowly and less efficiently in some patients. To uncover these "non-responders" patients, the availability of proper analytical methods is pivotal in order to study the pharmacodynamics, the pharmacokinetics and the metabolic fate of ASA. We validated a high-throughput, isocratic reversed-phase, negative MRM, LC-MS/MS method useful for measuring circulating ASA and salicylic acid (SA) in blood and plasma. ASA-d4 and SA-d4 were used as internal standards. The method was applied to evaluate: (a) the "in vitro" ASA degradation by esterases in whole blood and plasma, as a function of time and concentration; (b) the "in vivo" kinetics of ASA and SA after 7 days of oral administration of EC-ASA or plain-ASA (100 mg) in healthy volunteers (three men and three women, 37-63 years). Parameters of esterases activity were V max 6.5 ± 1.9 and K m 147.5 ± 64.4 in plasma, and V max 108.1 ± 20.8 and K m 803.2 ± 170.7 in whole blood. After oral administration of the two formulations, t max varied between 3 and 6 h for EC-ASA and between 0.5 and 1.0 h for plain-ASA. Higher between-subjects variability was seen after EC-ASA, and one subject had a delayed absorption over eight hours. Plasma AUC was 725.5 (89.8-1222) for EC-ASA, and 823.1(624-1196) ng h/mL (median, 25-75% CI) for plain ASA. After the weekly treatment, serum levels of TxB 2 were very low (< 10 ng/mL at 24 h from the drug intake) in all the studied subjects, regardless of the formulation or the t max . This method proved to be suitable for studies on aspirin responsiveness.

Introduction: Enthesitis-related Arthritis (ERA) is a specific category of juvenile idiopathic arthritis (JIA) characterized by axial and/or peripheral arthritis, and enthesitis, although other different extra-articular manifestations may encompass its clinical spectrum. Materials and Methods: In order to examine if ERA-JIA with extra-articular involvement may represent a different entity from ERA without extra-articular involvement, we performed a retrospective, observational, monocentric study, in a cohort of ERA patients followed between 2001 and September 2020 at the Pediatric Rheumatology Unit of Meyer Children Hospital of Florence. We analyzed the demographic, clinical, laboratory and imaging data at the disease onset, as well as after 3, 6, and 12 months follow up. Results: We have enrolled 53 patients, 33 males. At the time of diagnosis, average age was 10.9 years, 53 patients had active arthritis and 25 active enthesitis. The middle foot involvement was present in 20 patients. Twenty-five children achieved clinical remission on medication. Extra-articular manifestations were observed in 14 patients, of whom 3 had inflammatory bowel disease, 5 uveitis, one uveitis associated with Crohn disease, 4 SAPHO syndrome, one celiac disease. The cohort was stratified according to the presence/absence of extra-articular manifestations. It was observed that middle foot involvement was more frequent in patients with no extra-articular manifestations (18/39 vs. 2/14; χ 2 = 4.45, p = 0.05). Additionally, patients presenting extra-articular manifestation needed more frequently (12/14 vs. 21/39, χ 2 = 4.45, p = 0.05), and preciously (months: 3.7 ± 5.4 vs. 16.7 ± 26.5, p = 0.02), treatment with biologic agents. Finally, these patients achieved belatedly (months: 31.6 ± 32.3 vs. 22.9 ± 18.3, p = 0.01) and less frequently (3/14 vs. 22/39; χ 2 = 5.50, p = 0.03) the clinical remission on medication. Eventually, extra-articular involvement inversely correlated with the middle-foot arthritis (ρ s -0.29, p = 0.03), the chance to achieve remission on medication (ρ s -0.31 e p = 0.02), as well as the chance to keep overall remission, with and without medication (ρ s -0.28, p = 0.04). Conclusion: In our cohort, children diagnosed with ERA-JIA at the onset of disease and then developed extra-articular manifestations show the absence of middle foot involvement and worse prognosis with an early need for the use of biologic agents, and overall low chance to achieve remission., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Pagnini, Scavone, Maccora, Mastrolia, Marrani, Bertini, Lamot and Simonini.)

Background: Congenital hypothyroidism (CH) is reported to be more common in preterm infants than in term infants, especially in sick preterm infants. Though a frequent possibility of transitory thyroidal alterations in this category of neonates, the evolution of CH to transient or permanent forms is unpredictable., Methods: We retrospectively analyzed medical records of 28 preterm infants (<37 weeks gestation) who had exhibited a positive screening for CH at birth during the period 2000-2015 followed in our Center. Children were divided into three groups: permanent CH (PCH) with thyroid dysgenesis, PCH with eutopic normal-sized thyroid gland, and transient CH (TCH) with eutopic normal-sized thyroid gland. In all groups we described clinical and biochemical characteristics. Secondly, we analyzed the differences between patients with thyroid dysgenesis and patients with eutopic normal-sized gland and we compared PCH and TCH groups with normal-sized thyroid gland in order to identify clinical or biochemical data for early detection of transient forms., Results: Of all patients, 21.4% showed thyroid dysgenesis while 78.6% presented eutopic normal-sized gland. Infants with thyroid dysgenesis had higher median (IQR) baseline s-TSH and levothyroxine (L-T4) dose per weight at 12 months (12 m-dose) than patients with eutopic normal-sized gland. At re-evaluation of the patients with eutopic normal-sized gland, 36% showed PCH and 64% had TCH. The age of the patients at the beginning of L-T4 treatment, gestational age (GA), birth weight, blood thyroid stimulating hormone levels (b-TSH) at first newborn screening (NBS), baseline serum thyroid stimulating hormone (s-TSH), and L-T4 12 m-dose were statistically different between the two groups., Conclusions: Our results demonstrate that factors as GA, birth weight, b-TSH levels at first NBS, baseline s-TSH, L-T4 12 m-dose and age at the start of the treatment may be considered useful predictive elements for the evolution of CH., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest relevant to this article., (Copyright © 2020. Published by Elsevier B.V.)

Essential thrombocythemia (ET) patients are treated with aspirin (acetylsalicylic acid [ASA]) to prevent thrombosis. Previous studies showed that serum thromboxane (Tx) B 2 was high 24 hours after enteric-coated (EC)-ASA in ET patients, due to increased number of noninhibited reticulated platelets (RPs), consequent to high platelet turnover, and that ASA should be given twice a day to ET patients. We studied ET patients ( n  = 17) and healthy subjects ( n  = 10) on 100 mg EC-ASA once daily; experiments were repeated after 14-day treatment with 100 mg plain-ASA once daily. Serum TxB 2 , plasma ASA, and salicylic acid (SA) were measured before the morning dose and up to 8 hours thereafter. Blood activity of ASA-deacethylating esterases, in vitro inhibition of collagen-induced TxB 2 production by ASA (10-1,000 µM), and number of RP were measured. TxB 2 inhibition by ASA in vitro and esterases activities were normal in all subjects. EC-ASA elicited highly variable responses; 6 ET patients were poor responders, as their serum TxB 2 was high after EC-ASA; their plasma levels of ASA and SA were low/undetectable. In contrast to EC-ASA, plain ASA decreased serum TxB 2 and increased plasma ASA and SA in all subjects. Serum TxB 2 was high in ET patients at 24 hours and significantly correlated with RP count (but not RP percentage) and platelet count. Plain ASA should be used in ET patients to inhibit platelets efficiently. The identification of ET patients who might benefit from twice a day ASA could simply be based on their platelet count: since their platelet turnover is not increased, ET patients with normalized platelet count should not need twice a day ASA treatment., Competing Interests: None declared., (Thieme. All rights reserved.)

Congenital hypothyroidism (CH) is the most common endocrine disease in children, according to literature, infants with CH have an increased risk of associated congenital malformations (CM), especially cardiac defects (CD), compared to the general population. We retrospectively analyzed medical records of 255 patients with a positive screening result for CH in the period 1991-2016 followed at our Center. At the time of enrollment, the clinical examination included looking for the presence of heart murmurs and dysmorphic features. In all patients an echocardiography with cardiological evaluation were performed. Of all patients, 191 were included in the final analysis. Of these, 51.3% (98/191) presented an eutopic normally sized thyroid gland while 48.7% (93/191) showed a thyroid dysgenesis. Among the studied infants, 13.6% (26/191) presented CD. The most frequent cardiac anomaly was atrial septal defect (ASD) which was found in 65.4% (17/26) of patients with CD. Other defects were ventricular septal defect (VSD), patent ductus arteriosus (PDA), pulmonary valve stenosis (PvS), transposition of the great vessels (TGV), aortic valve stenosis (AvS). Six patients had multiple defects. In the analysed group, there was no significant relation with sex, type of CH, median blood-TSH (b-TSH) and serum-TSH (s-TSH) values and frequency of CD. There is a high prevalence of CD in CH, indicating the need of routine echocardiography in these patients to achieve an early diagnosis and management of CD., (Copyright 2020 Biolife Sas. www.biolifesas.org.)

As key cellular elements of hemostasis, platelets represent a primary target for thrombosis and bleeding management. Currently, therapeutic manipulations of platelet function (antithrombotic drugs) and count (platelet transfusion) are performed with limited or no real-time monitoring of the desired outcome at the point-of-care. To address the need, we have designed and fabricated an easy-to-use, accurate, and portable impedance aggregometer called "MICELI" (MICrofluidic, ELectrical, Impedance). It improves on current platelet aggregation technology by decreasing footprint, assay complexity, and time to obtain results. The current study aimed to optimize the MICELI protocol; validate sensitivity to aggregation agonists and key blood parameters, i.e., platelet count and hematocrit; and verify the MICELI operational performance as compared to commercial impedance aggregometry. We demonstrated that the MICELI aggregometer could detect platelet aggregation in 250 μL of whole blood or platelet-rich plasma, stimulated by ADP, TRAP-6, collagen, epinephrine, and calcium ionophore. Using hirudin as blood anticoagulant allowed higher aggregation values. Aggregation values obtained by the MICELI strongly correlated with platelet count and were not affected by hematocrit. The operational performance comparison of the MICELI and the Multiplate ® Analyzer demonstrated strong correlation and similar interdonor distribution of aggregation values obtained between these devices. With the proven reliability of the data obtained by the MICELI aggregometer, it can be further translated into a point-of-care diagnostic device aimed at monitoring platelet function in order to guide pharmacological hemostasis management and platelet transfusions., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Microfluidic flow chambers (MFCs) allow the study of platelet adhesion and thrombus formation under flow, which may be influenced by several variables. We developed a new MFC, with which we tested the effects of different variables on the results of platelet deposition and thrombus formation on a collagen-coated surface., Methods: Whole blood was perfused in the MFC over collagen Type I for 4 min at different wall shear rates (WSR) and different concentrations of collagen-coating solutions, keeping blood samples at room temperature or 37 °C before starting the experiments. In addition, we tested the effects of the antiplatelet agent acetylsalicylic acid (ASA) (antagonist of cyclooxygenase-1, 100 µM) and cangrelor (antagonist of P2Y 12 , 1 µM)., Results: Platelet deposition on collagen (I) was not affected by the storage temperature of the blood before perfusion (room temperature vs. 37 °C); (II) was dependent on a shear rate in the range between 300/s and 1700/s; and (III) was influenced by the collagen concentration used to coat the microchannels up to a value of 10 µg/mL. ASA and cangrelor did not cause statistically significant inhibition of platelet accumulation, except for ASA at low collagen concentrations., Conclusions: Platelet deposition on collagen-coated surfaces is a shear-dependent process, not influenced by the collagen concentration beyond a value of 10 µg/mL. However, the inhibitory effect of antiplatelet drugs is better observed using low concentrations of collagen., Competing Interests: The authors declare no conflict of interest.

Yolk sac tumor (YST) is a rare tumor that usually occurs in the first two decades of life. It is considered the second most common malignant germ cell tumor of the ovary, characterized by a rapid growth and a bad prognosis due to the frequent metastasis. We report the case of a 12-year-old girl who came to our observation for an acute abdominal pain. Clinical examination evidenced a vague mass in the suprapubic region and a lower abdomen tenderness, the US imaging revealed a complex lesion of the left ovary (19 x 13 cm) and the alpha-fetoprotein (AFP) resulted high (5858 ng/mL). Computed tomography (CT) revealed a large pelvic mass. The treatment consisted of debulking surgery of yolk sac tumor followed by 4 cycles of BEP protocol (Bleomycin, Etoposide, Cisplatin). After 3 years of follow-up there was no evidence of disease recurrence. (www.actabiomedica.it).

Background: Loss-of-function mutations of fibroblast growth factor receptor 1 gene (FGFR1) have been reported so far. These mutations have been described in the extracellular domain, consisting of three Ig-like domains in the single transmembrane helix and in the intracellular region, containing a tyrosine kinase domain and cause about 10% of all cases of Kallmann syndrome. FRGR1 mutations could be associated with non reproductive phenotype such as cleft palate and dental agenesis and a wide spectrum of reproductive phenotype., Case Report: The patient, 17 years and 11 months old, was a Bulgarian male referred to our Pediatric Endocrinology Unit for pubertal failure and hyposmia. Clinical evaluation revealed a highpitched voice, gynecomastia and obesity. Hormonal study revealed hypogonadotropic hypogonadism. Molecular analysis, performed by Next Generation Sequencing and confirmed by Sanger sequencing, led to the identification of a novel and previously undescribed mutation c.1058 C>G (p. S353C) in heterozygous state on exon 8 of the FGFR1 gene., Conclusion: The novel mutation, that we found in a boy with Kallman syndrome, could destabilize the D3 immunoglobulin like receptor domain that is crucial for the FGF-FGFR interaction. (www.actabiomedica.it).

We present a case of an 11-month-old girl who was referred to our unit for an erythematous rash that appeared on the face and extremities. Personal and family history was not relevant. Laboratory tests were normal. During recovery, diameter and colour intensity of the cutaneous lesions increased, but after some weeks, lesions had a self-limited resolution without any treatment. Based on clinical and laboratory findings, a diagnosis of acute hemorrhagic edema of infancy (AHEI) was made.  AHEI is a rare cutaneous leukocytoclastic vasculitis that usually affects children aged between 4 and 24 months. Etiology is unknown but almost of 75% of cases are preceded by infectious episodes, vaccinations or use of medications. In contrast to the dramatic cutaneous eruption, clinical conditions are usually optimal. Classically, AHEI is characterized by a triad of symptoms: fever, edema and purpura. Skin lesions are erythematous, annular, medallion-like, purpuric plaques that have a rapid onset and appear on the face and extremities, sparing trunk and mucosal membranes. Initially interpreted as a variant of Henoch-Schönlein purpura, now it is considered a distinct disease. In the majority of cases the disease is benign and self-limited without a visceral involvement, so a conservative approach is most often chosen., Competing Interests: No competing interests were disclosed., (Copyright: © 2019 Carboni E et al.)

Antiphospholipid syndrome (APS) is a chronic and disabling condition characterized by recurrent thrombosis and miscarriages mediated by antibodies against phospholipid-binding proteins (aPL), such as beta 2 glycoprotein I (β 2 GPI). Complement is involved in APS animal models and complement deposits have been documented in placenta and thrombotic vessels despite normal serum levels. Analysis of circulating blood cells coated with C4d displays higher sensitivity than the conventional assays that measure soluble native complement components and their unstable activation products in systemic lupus erythematosus (SLE). As C4d-coated blood cell count has been reported to be more sensitive than serum levels of complement components and their activation products in systemic lupus erythematosus (SLE) patients, we decided to evaluate the percentage of C4d positive B lymphocytes (BC4d), erythrocytes (EC4d), and platelets (PC4d) in primary APS patients and asymptomatic aPL positive carriers as marker of complement activation in APS. We assessed by flow cytometry the percentages of BC4d, EC4d, and PC4d in primary APS (PAPS; n. 23), 8 asymptomatic aPL positive carriers, 11 APS-associated SLE (SAPS), 17 aPL positive SLE, 16 aPL negative SLE, 8 aPL negative patients with previous thrombosis, 11 immune thrombocytopenia (ITP) patients, and 26 healthy subjects. In addition, we used an in vitro model to evaluate the ability of a monoclonal anti-β 2 GPI antibody (MBB2) to bind to normal resting or activated platelets and fix complement. EC4d and PC4d percentages were significantly higher in PAPS and aPL carriers as well as aPL positive SLE and SAPS than in aPL negative controls. The highest values were found in PAPS and in SAPS. The EC4d and PC4d percentages were significantly correlated with serum C3/C4 and anti-β 2 GPI/anti-cardiolipin IgG. In vitro studies showed that MBB2 bound to activated platelets only and induced C4d deposition. The detection of the activation product C4d on circulating erythrocytes and platelets supports the role of complement activation in APS. Complement may represent a new therapeutic target for better treatment and prevention of disability of APS patients.

Objective: To identify factors that discriminate between transient and permanent congenital hypothyroidism., Methods: Retrospective evaluation of 58 children with congenital hypothyroidism and eutopic thyroid gland. Gender, gestational age, birth weight, TSH and serum thyroxine levels at diagnosis and L-thyroxine dose at 12 and 24 months of age were analyzed., Results: Median (IQR) initial TSH levels were 73.3 (276.5) µIU/mL in permanent hypothyroidism and 24.24 (52.7) µU/mL in transient hypothyroidism (P =0.0132). The optimum cut-off value of initial TSH to predict transient hypothyroidism was 90 µIU/mL. Mean (SD) L-thyroxine doses at 24 months of age were 2.64 (0.98) µg/kg/day in permanent hypothyroidism and 1.91 (0.65) µg/kg/day in transient hypothyroidism. Requirement of L-thyroxine dose at 24 months of ≤0.94 µg/kg/day had the highest sensitivity (100%) to predict transient hypothyroidism., Conclusions: L-thyroxine doses at 24 months can predict transient hypothyroidism in patients with eutopic thyroid gland earlier than at 36 months.

A variety of laboratory tests have been developed, which can diagnose a number of both congenital and acquired disorders of platelet function. Many tests of platelet function measure the ability of platelets to adhere to each other, forming platelet aggregates, which represent the major constituents of hemostatic plugs and of arterial thrombi. Light transmission aggregometry (LTA) is still considered the gold standard of platelet aggregation tests, but other platelet aggregation-based tests are also available. Among them, the flow cytometry-based methods may be more convenient than LTA for the study of patients with very low or very high platelet counts. The use of platelet aggregation tests has also been advocated to monitor the treatment with antiplatelet agents (mostly the P2Y 12 antagonist clopidogrel) of patients with thrombotic arterial occlusions, with the aim of improving their efficacy and safety. However, randomized clinical trials failed to show any advantage of this strategy; as a consequence, international guidelines now recommend against laboratory monitoring of antiplatelet therapy.

Loss-of-function mutations of the the ATP-binding cassette-1 ( ABCA1 ) gene are the cause of Tangier disease (TD) in homozygous subjects and familial HDL deficiency (FHD) in heterozygous subjects. These disorders are characterized by reduced plasma HDL-cholesterol (HDL-C) and altered efflux of cholesterol from cells. Previous studies in TD patients and ABCA1 -/- murine models reported defects in platelet count, morphology, and function, but the issue is still controversial. We analyzed three subjects with low to very low HDL-C levels due to the loss-of-function mutations of the ABCA1 gene. Two related patients with FHD were heterozygous carriers of two mutations on the same ABCA1 allele; one, with TD, was homozygous for a different mutation. Mild to moderate thrombocytopenia was observed in all the patients. No morphological platelet abnormalities were detected under optical or EM. History of moderate bleeding tendency was recorded only in one of the FHD patients. Only limited alterations in platelet aggregation and activation of the integrin αIIbβ3 were observed in one FHD patient. While α-granule secretion (P-selectin), content, and secretion of platelet δ-granules (serotonin, ATP, and ADP) and thromboxane (TX) A 2 synthesis were normal in all the patients, the expression of lysosomal CD63, in response to some agonists, was reduced in TD patients. In conclusion, three patients carrying ABCA1 genetic variants had low platelet count, with the lowest values observed in TD, not associated with major alterations in platelet morphology and response to agonists or bleeding., (© 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Aims: Patients with acute coronary syndromes (ACSs) are treated with acetylsalicylic acid (ASA) and antagonists of the P2Y 12 receptor (P2Y 12 R) for adenosine diphosphate (ADP). Based on the demonstration that P2Y 12 R antagonists inhibit thromboxane A 2 (TxA 2 ) production (target of ASA), it was surmised that ACS patients might be treated with P2Y 12 R antagonists only. However, this demonstration contrasts with the results of previous studies. The aim of this study was to test whether P2Y 12 R antagonists have off-target/indirect inhibitory effects on platelet TxA 2 production., Methods and Results: We studied 3 patients with inherited P2Y 12 R deficiency and 33 healthy subjects. Serum TxB 2 (TxA 2 metabolite) levels were similar in P2Y 12 R-deficient patients and healthy subjects and were not decreased by P2Y 12 R antagonists in vitro. Serum TxB 2 levels did not decrease in 20 patients treated with prasugrel (10 mg q.i.d.) or placebo for 14 days. Arachidonic acid- and collagen-induced platelet aggregation (PA) and TxB 2 production in platelet-rich plasma (PRP) of healthy subjects were inhibited in vitro by P2Y 12 R antagonists. However, P2Y 12 R antagonists did not inhibit TxB 2 production when PA was prevented by avoiding the stirring of PRP in the aggregometer. The P2Y 1 ADP-receptor antagonist MRS2500 had similar effects on PA and TxB 2 production as P2Y 12 R antagonists. Acetylsalicylic acid inhibited TxB 2 production more effectively than a P2Y 12 R antagonist; only the combination of ASA and a P2Y 12 R antagonist inhibited PA induced by high concentration of collagen., Conclusion: Inherited deficiency or pharmacological inhibition of P2Y 12 R does not affect the platelet capacity to synthesize TxA 2 . There is no pharmacological evidence that ACS patients may be safely treated with P2Y 12 R antagonists without ASA., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2015. For permissions please email: journals.permissions@oup.com.)

The purpose of this study was to investigate the antimicrobial properties of multifunctional nanocomposites based on poly(dl-Lactide-co-Glycolide) (PLGA) and increasing concentration of silver (Ag) nanoparticles and their effects on cell viability for biomedical applications. PLGA nanocomposite films, produced by solvent casting with 1 wt%, 3 wt% and 7 wt% of Ag nanoparticles were investigated and surface properties were characterized by atomic force microscopy and contact angle measurements. Antibacterial tests were performed using an Escherichia coli RB and Staphylococcus aureus 8325-4 strains. The cell viability and morphology were performed with a murine fibroblast cell line (L929) and a human osteosarcoma cell line (SAOS-2) by cell viability assay and electron microscopy observations. Matrix protein secretion and deposition were also quantified by enzyme-linked immunosorbent assay (ELISA). The results suggest that the PLGA film morphology can be modified introducing a small percentage of silver nanoparticles, which induce the onset of porous round-like microstructures and also affect the wettability. The PLGA/Ag films having silver nanoparticles of more than 3 wt% showed antibacterial effects against E. coli and S. aureus . Furthermore, silver-containing PLGA films displayed also a good cytocompatibility when assayed with L929 and SAOS-2 cells; indicating the PLGA/3Ag nanocomposite film as a promising candidate for tissue engineering applications.

Introduction: The platelet function analyzer (PFA)-100 is used in clinical practice to screen patients with bleeding diathesis and suspected defects of primary hemostasis. A new cartridge, INNOVANCE PFA P2Y, has been specifically developed to monitor patients' response to drugs inhibiting the platelet P2Y₁₂ receptor for ADP. In this study, we compared the ability of INNOVANCE PFA P2Y to detect congenital defects of the platelet P2Y₁₂ receptor to that of standard cartridge formulations currently in clinical use., Materials and Methods: We studied two patients with severe P2Y₁₂ deficiency, one patient with heterozygous P2Y₁₂ deficiency and one with dysfunctional P2Y₁₂ receptor. Closure times were measured using 3 cartridges: collagen/ADP, collagen/epinephrine, and INNOVANCE PFA P2Y. The results obtained in the four patients with P2Y₁₂ defects were compared to those obtained for 20 healthy controls., Results: In 2 patients with severe P2Y₁₂ deficiency, closure times of INNOVANCE PFA P2Y and collagen/ADP cartridges were >300 s, while those of collagen/epinephrine cartridge were variable (186s and >300 s). In the patient with dysfunctional P2Y₁₂, closure time of INNOVANCE PFA P2Y was >300 s, while closure times of collagen/ADP and collagen/epinephrine were normal. Closure times of all cartridges were normal in the patient with heterozygous P2Y₁₂ deficiency., Conclusion: Our study provides the first evidence that INNOVANCE PFA P2Y cartridge is sensitive to congenital severe and moderate defects of the platelet P2Y₁₂ receptors., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Staphylococcus epidermidis is a leading cause of nosocomial infections, and its virulence is attributable to formation of biofilm, especially on implanted devices. Photodynamic treatment (PDT) has been actively investigated for the eradication of bacterial biofilm growing on dental plaques and oral implants. In this study, we used Tri-meso (N-methyl-pyridyl), meso (N-tetradecyl-pyridyl) porphine (C14) for inactivation of two structurally distinct S. epidermidis biofilms grown on Ti6Al4V alloy and compared its photosensitizing efficiency with that of the parent molecule, tetra-substituted N-methyl-pyridyl-porphine (C1). A more significant reduction in bacterial survival was observed when both bacterial biofilms were exposed to a lower dose of C14, and simultaneously to visible light in comparison with C1. The different responses of both staphylococcal biofilms to C1- or C14-treatment appeared to depend on photosensitizer endocellular concentration. C14 bound to both biofilms to a greater extent than C1. Moreover, C14 penetrates deeper into the bacterial membranes, as determined by fluorescence quenching experiments with methylviologen, allowing for better bacterial killing photoefficiency. Confocal laser scanning microscope (CLSM) analysis indicated damage to bacterial cell membranes in both photodynamically treated biofilms, while disruption of PDT-treated biofilm was confirmed by scanning electron microscopy (SEM). In summary, C14 may be a potential photosensitizer for the inactivation of staphylococcal biofilms for many device-related infections which are accessible to visible light.

The synaptonemal complex (SC) of specimens of Bos taurus taurus from the Holstein-Friesian, Piemontese, and Simmental breeds, was analysed. The analysis included quantification of the frequency of various types of abnormalities in the SC, and the frequency of cells with SC abnormalities. All animals had 29 autosomal bivalents and one sexual bivalent and the most frequently recorded abnormality was pairing failure. The number of cells with abnormalities in the Holstein-Friesian breed was 29.41%, in the Piemontese breed was 30.00% and in the Simmental breed it was 29.54%. The subspecies Bos taurus taurus had 29.63% of cells showing abnormalities with 57.33% of these abnormalities occurring in zygotene and 42.67% occurring in pachytene. Statistical analyses showed that there were no significant differences in the number of cells with SC abnormalities among the breeds studied. The frequency of cells with abnormalities, and the efect on the fertility of the Holstein-Friesian, Piemontese and Simmental breeds are discussed.

We report fluorescence in-situ hybridization (FISH) and somatic cell hybrid mapping data for 13 different horse genes (ANP, CD2, CLU, CRISP3, CYP17, FGG, IL1RN, IL10, MMP13, PRM1, PTGS2, TNFA and TP53). Primers for PCR amplification of intronic or untranslated regions were designed from horse-specific DNA or mRNA sequences in GenBank. Two different horse bacterial artificial chromosome (BAC) libraries were screened with PCR for clones containing these 13 Type I loci, nine of which were found in the libraries. BAC clones were used as probes in dual colour FISH to confirm their precise chromosomal origin. The remaining four genes were mapped in a somatic cell hybrid panel. All chromosomal assignments except one were in agreement with human-horse ZOO-FISH data and revealed new and more detailed information on the equine comparative map. CLU was mapped by synteny to ECA2 while human-horse ZOO-FISH data predicted that CLU would be located on ECA9. The assignment of IL1RN permitted analysis of gene order conservation between HSA2 and ECA15, which identified that an event of inversion had occurred during the evolution of these two homologous chromosomes.

The synaptonemal complex (SC) was analyzed in four F1 hybrids of Bos taurus taurus and B. taurus indicus including Gyr-Simmental (G-S), Nelore Simmental (N-S), Gyr-Holstein-Friesian (G-H) and Nelore-Piemontese (N-P). We analysed the frequency of various types of SC abnormalities and the frequency of cells with SC abnormalities. The results were compared with similar observations made on purebred animals. All the animals studied possessed 29 autosomal and one sex bivalent. The frequency of cells with abnormalities in the hybrids were 28.0% in the N-P, 29.1% in the G-S, 33.3% in the N-S and 40.0% in the G-H. The frequency of cells with abnormalities in the four hybrids was 31.5%; 57.9% of these abnormalities occurred in zygotene and 42.0% occurred in pachytene. The comparisons among the hybrids and among the hybrids and their parental breeds showed that the only significant difference was between Gyr and Gyr-Holstein-Friesian animals. Some aspects of the relationship between the frequency of cells with anomalies and the fertility of hybrids are discussed.