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The genetic monitoring of forest trees is of the utmost importance for securing the maintenance of species’ adaptive potential. Genetic monitoring, the quantification of temporal changes in a population’s genetic variation and structure, introduces prognosis and assists in defining tools for the management of genetic resources. Forest genetic monitoring (FGM) is imperative in biodiversity hotspots such as Greece, one of the most species-rich European countries, where >7000 native plant taxa (~20% endemics) exist and 28% of its land base is included in the Natura 2000 Special Areas of Conservation network. Moreover, (meta-)analyses of genetic data have shown that Greek forests’ tree populations present significantly higher values of polymorphism and differentiation compared to the average values reported in the international literature. FGM studies in Greece involve the following species: Abies borissi regis, Castanea sativa, Fagus sylvatica, Quercus petraea, Q. robur, Pinus nigra, and Prunus avium. FGM uses a genealogical approach and employs 11–26 Simple Sequence Repeat (SSR) loci in different cohorts (mature and juvenile trees), while the average time internal between assessments is 12.9 years. The results so far for A. borissi regis and F. sylvatica indicate a maintenance of genetic diversity over time, but with a slight drop of Ne in the juvenile cohort of hybrid firs compared to the mature trees.

Taxus baccata (European yew) is native to Greece, which hosts the most southeastern populations of the species. It is a medically interesting conifer due to its tetracyclic diterpenes, with the anticancer agent paclitaxel (taxol) being the most well-known. Yew trees with a high content of paclitaxel (taxol) have been identified in the natural population of Mt. Cholomon in Greece. The macropropagation of these plus trees is of high importance both for the establishment of future plantations and for the ex situ conservation of this valuable germplasm. A total of 600 cuttings from top and side shoots were taken from 10 highly producing paclitaxel trees and treated with a rooting solution containing the plant hormone indole-3-butyric acid potassium salt (K-IBA) concentrations of 6000 ppm and 12,000 ppm. Cuttings were placed in a greenhouse for rooting using a perlite and peat 2:1 rooting medium, under constant temperature and humidity conditions. After five months, the evaluation of rooting success showed that it varied between different genotypes (40–100%), but was generally considerable (average success of 79.5%). The higher K-IBA concentration resulted in a better rooting success, being only in 50% of the genotypes tested; however, root length was differentially affected, as use of the 12,000 ppm concentration resulted in an average root length on a per genotype basis of 2.76 cm, compared to an average of 2.69 cm when the 6000 ppm concentration was used. Overall, high-yielding paclitaxel (taxol) was successfully macropropagated, and these results can be applied to both ex situ conservation and plantation establishment.

Aleppo pine (Pinus halepensis) is widely distributed in the Mediterranean region and in other areas of the world, where it has been introduced due to its adaptive capacity to xerothermic conditions. The giant pine scale Marchalina hellenica often infests Aleppo pine, as well as other pines, in several southeastern European countries, causing pine declines. When combined with the expected intensified heat and drought events in eastern Mediterranean, the impact of this biotic parameter on the host pines may be exacerbated. The importance of understanding the defense mechanisms of Aleppo pine is emphasized by the recent invasion of the pine scale in new regions, like Australia, lacking the insect’s natural enemies, where more intense negative effects on pine species may occur. To date, Aleppo pine’s physiological responses to the infestation by M. hellenica are largely unknown. This study aimed at assessing the responses of Aleppo pine to the giant pine scale attack, both on an ecophysiological and a metabolic level. For this purpose, gas exchange, needle water status, and carbon and nitrogen content were measured during 1 year on healthy and infested adult trees. M etabolic profiling of Aleppo pine needles was also performed before, during, and after the high feeding activity of the insect. The maintenance of stable relative water content, δ13C signatures, and chlorophyll fluorescence in the needles of infested pines indicated that infestation did not induce drought stress to the host pines. At the peak of infestation, stomatal closure and a pronounced reduction in assimilation were observed and were associated with the accumulation of sugars in the needles, probably due to impaired phloem loading. At the end of the infestation period, tricarboxylic acids were induced and phenolic compounds were enhanced in the needles of infested pines. These metabolic responses, together with the recovery of photosynthesis after the end of M. hellenica intense feeding, indicate that in the studied region and under the current climate, Aleppo pine is resilient to the infestation by the giant pine scale. Future research should assess whether these promising defense mechanisms are also employed by other host pines, particularly in regions of the world recently invaded by the giant pine scale, as well as under more xerothermic regimes.

A curvature of a finger that bends inwards relative to the other fingers is a common observation during prenatal screening. When the angulation exceeds 10 degrees, it is known as “clinodactyly” and could suggest a variety of underlying issues. Even though it usually remains unnoticed during pregnancy, it may be a sign of serious fetal disease. We report the case of a fetus diagnosed with clinodactyly of the thumb accompanied by tachycardia, abnormal levels of maternal hormones in the first trimester and increased impedance to flow in the uterine arteries. Although non-invasive prenatal testing was normal, amniocentesis was carried out and two deviations at chromosome 20 were identified through molecular karyotyping. Our report aims to raise clinical suspicion regarding the potential association between genetic abnormalities and clinodactyly. A careful clinical and genetic consultation is required in order to achieve the most favorable outcome for both mother and child.

Abstract In this research, fuzzy linear regression (FLR) method combined with three well-known fuzzy implications was implemented for evaluating the relation among the amount of fly ash in concrete mixture and the compressive strength of concrete. More specifically, 267 experimental data 40 of which were used for testing the validation of the process were subjected to FLR method for calculating the truth values, which indicated the degree of how the experimental outputs belong to the predicted ones. Also, the degree of fuzziness was calculated for performing the sensitivity analysis of the model. The truth values that emerged were used for applying three basic fuzzy implications such as Lukasiewicz, Reinchenbach, and Kleene-Dienes implication. By evaluating and comparing the results of every fuzzy implication, it was concluded that Lukasiewicz was the most appropriate implication method as it yielded the smallest deviation of truth values (σ = 4.00) in contrast to the theoretical ones (σ = 4.83 in Reinchenbach and σ = 12.31 in Kleene-Dienes fuzzy implication). The accuracy of the FLR method was also validated for calculating the coefficient of the mean absolute percentage error level (MAPE = 5.56%) of the blind prediction process, and the results revealed that the application of fuzzy linear regression method is suitable for evaluating the truth values of experimental data in order to be used in fuzzy implications. Thus, it is a satisfactory procedure for making inferences between concrete parameters.

Introduction: Acute hemiparesis is an emergency of various etiologies and possible fatal outcome. Aim: The aim of this study was to determine the etiology, clinical manifestations, and prognosis of acute hemiparesis in childhood. Materials and methods: This is a retrospective study of 52 children (28 boys and 24 girls) aged 4 months to 16 yrs 11 months with acute hemiparesis, admitted to the Clinic of Pediatrics at St George University Hospital between 2013 and 2020. After clinical examination, a modern diagnostic and therapeutic approach was used. Results: The identified causes of acute hemiparesis were ischemic stroke (21), postictal paresis after seizure (9), hemorrhagic stroke (6), venous sinus thrombosis (5), encephalitis (3), transient ischemic attacks (TIAs) (2), posterior reversible encephalopathy syndrome (PRES) (2), hemiplegic migraine (1), demyelinating CNS disease (1), ventriculoperitoneal shunt complication (1), glutaric aciduria type 1 (1), brain tumor (1), and CNS metastasis (1). The most common manifestations associated with acute hemiparesis were impaired consciousness (27), involvement of cranial nerves – central (25) and peripheral (1) facial nerve palsy, abducens (1), ocolomotorius (1), hypoglossus (1) nerve palsy, seizures (25), raised intracranial pressure (18), speech disorders (15), signs of infection (14), headache (12), and hemibody sensory disturbances (9). In 8 children (14%), the hemiparesis was permanent, in 2 recurrent, and in 7 (12%) fatal. Conclusions: Unlike in adult patients, acute hemiparesis in children has many causes, which requires a large number of etiological studies. With a quick diagnosis and adequate treatment, recovery of function in children is more common than in adults – it is achieved in two thirds of children.

Complete androgen insensitivity syndrome (CAIS) occurs in XY individuals due to androgen receptor dysfunction and presents with female phenotype and primary amenorrhea. Patients are typically raised and identify themselves as female and the absence of an upper vagina results in psychological and sexual problems. The cases of two sisters with CAIS that were managed differently regarding the neovagina formation are presented. Non-surgical neovagina formation results in sufficient vaginal depth and satisfying sexual life and may be preferred over the operation. Our findings highlight that the interaction of CAIS patients with each other and the continuous psychological and physical support by the medical team contribute significantly to the outcome.

Virilization is a rare condition in postmenopausal women, usually attributed to androgen excess of ovarian or adrenal origin. A 62-year-old woman presented with excessive hair loss of 3 months' duration and was investigated for an endocrine cause of alopecia. The hormonal evaluation revealed increased testosterone but normal levels of androstenedione and dehydroepiandrosterone sulfate, while the results of transvaginal ultrasonography and abdominal computed tomography were unremarkable. Based on these findings, the possibility of an adrenal androgen-secreting tumor was ruled out and suspicion of Leydig cell hyperplasia was raised. A bilateral laparoscopic salpingo-oophorectomy was performed due to the age of the patient and the diagnosis of Leydig cell hyperplasia was confirmed by histopathological examination. The postoperative course of the patient was uneventful and a repeat hormonal evaluation after the operation showed a normalization of androgen levels. In conclusion, Leydig cell hyperplasia should be considered as a likely cause of hyperandrogenism of ovarian origin in women who develop virilization. In postmenopausal women, bilateral oophorectomy will treat the disorder and provide a conclusive diagnosis via histopathological examination.

Background: Herpes simplex virus (HSV) seroprevalence in Bulgaria is higher than that in other countries but there is no information concerning involvement of these viruses in neuropathology. The aim of the present study was to determine the frequency of HSV DNA detection in cerebrospinal fluid (CSF) of patients with neurological diseases. Methods: This study is a retrospective survey of test results obtained from January 2015 to October 2019. During this period 617 CSF specimens were tested for the presence of HSV-1 and/or HSV-2 DNA by PCR. Results: Of all 612 CSF samples tested for HSV-1, 16.5% were positive. Of 547 CSF samples tested for HSV-2, 6.2% were positive. Co-infection with HSV-1 and HSV-2 was detected in 1.3% of tested 543 CSF samples. The difference of HSV-1 and/or HSV-2 prevalence in CSF of patients according to the gender and age was not statistically significant. The highest HSV-1 prevalence (25%) was detected in CSF from patients with multiple sclerosis (MS), followed by patients with encephalitis (20.6%). The highest HSV-2 prevalence (22.2%) was detected in CSF from patients with myelitis, followed by patients with encephalopathies (7.5%). Conclusion: Our results show a considerable prevalence of HSV-1 and HSV-2 in CSF of patients with neurological diseases indicating the important role of these viruses in neuropathology in Bulgaria. HSV-1 is with predominant role in development of both encephalitis and meningitis in different age groups. Testing of CSF for HSV should be indispensable part of diagnostic algoritm of these diseases.

Background: Office hysteroscopy is a widely-accepted and useful tool in the every-day practice of gynaecologists. Methods: In this 20-year-retrospective study, data originating from 2675 patients who underwent vaginoscopic office hysteroscopy are presented. The Endoscopic Unit is located in the Department of Gynaecology, University Hospital of Ioannina and it is considered one of the busiest public institutions providing care for a large amount of Greek population in Epirus, North-western Greece. Our findings presented in this very first and nationally largest retrospective study, will contribute to the overall scientific knowledge by providing substantial data with regards to hysteroscopy and to epidemiology of endometrial pathology. Results: Common hysteroscopic indication across all age groups was Abnormal Uterine Bleeding (AUB). Predictive characteristics of hysteroscopy in the diagnosis of various conditions were evaluated for these patients in comparison with their histologic report as the gold standard. Comparing hysteroscopic findings with the respective histology reports revealed that in cases of normal endometrium, sensitivity of 60.9%, specificity of 92.1%, Positive Predictive Value (PPV) of 79.07% and Negative Predictive Value (NPV) 82.8% were estimated. Hysteroscopic detection of endometrial polyps demonstrated sensitivity of 92.04%, specificity of 89.1%, PPV of 73.5% and NPV 97.1%. For fibroids, sensitivity and specificity were calculated at 98.5% and 100% respectively, while PPV and NPV at 100% and 99.9%, respectively. For endometrial cancer, the predictive characteristics were estimated at 87.5% and 99.7% with regards to sensitivity and specificity, and 63.6% and 99.9% for PPV and NPV, respectively. Finally, for cases of hyperplasia, hysteroscopy showed sensitivity of 75.0%, specificity of 91.03%, PPV of 11.7% and NPV of 99.5%. Conclusions: To date, this is the largest retrospective study on office hysteroscopy with the use of vaginoscopic approach technique in Greece. This study has been conducted in one of the busiest public gynecologic endoscopic units across Greece. Our findings are consistent with the international scientific evidence, which has proven that hysteroscopy is an efficient and safe method to investigate pathologies within the uterine cavity and in general is accompanied by satisfactory patient acceptance.

Introduction: Adenomyosis is a common chronic disease in women of reproductive age, characterised by the presence of ectopic endometrial tissue within myometrium. Even though adenomyosis presents with chronic pelvic pain, menorrhagia or abnormal uterine bleeding, dysmenorrhoea, and dyspareunia and is often recognised after hysterectomies. However, the development of ultrasonography and magnetic resonance imaging has improved the pre-operative diagnosis of the disease. Hysteroscopy provides information in real time from the uterine cavity and the offers the possibility of obtaining direct biopsies. Material and Methods: The literature was searched via Pubmed and Embase with the following headings: diagnosis of adenomyosis or adenomyoma and office hysteroscopy, hysteroscopy findings of adenomyosis or adenomyoma, treatment of adenomyosis or adenomyoma with office hysteroscopy. Results: The literature showed that hysteroscopy can identify superficial adenomyosis. There are a variety of hysteroscopic images that can be connected with the disease. New equipment like the spirotome has been used to access deeper layers of myometrium and obtain biopsies under direct vision from the adenomyotic areas. Different methods of treatment have been also described, like enucleation of focal superficial adenomyoma, coagulation, evacuation of cystic adenomyosis when the lesion is smaller than 1.5 cm, and resection of adenomyotic nodules in case of bigger lesions (>1.5 cm). Diffuse superficial adenomyosis is also managed by resection. Conclusions: Hysteroscopy has revolutionised the approach to adenomyosis. It is a useful tool in assessing mainly superficial adenomyosis. The role of hysteroscopy in surgical management of adenomyosis need to be confirmed with further studies.

Most abdominal masses in the pediatric population derive from the ovaries. Ovarian masses can occur in all ages, although their incidence, clinical presentation and histological distribution vary among different age groups. Children and adolescents may develop non-neoplastic ovarian lesions, such as functional cysts, endometrioma, torsion, abscess and lymphangioma as well as neoplasms, which are divided into germ cell, epithelial, sex-cord stromal and miscellaneous tumors. Germ cell tumors account for the majority of ovarian neoplasms in the pediatric population, while adults most frequently present with epithelial tumors. Mature teratoma is the most common ovarian neoplasm in children and adolescents, whereas dysgerminoma constitutes the most frequent ovarian malignancy. Clinical manifestations generally include abdominal pain, palpable mass, nausea/vomiting and endocrine alterations, such as menstrual abnormalities, precocious puberty and virilization. During the investigation of pediatric ovarian masses, the most important objective is to evaluate the likelihood of malignancy since the management of benign and malignant lesions is fundamentally different. The presence of solid components, large size and heterogenous appearance on transabdominal ultrasonography, magnetic resonance imaging and computed tomography indicate an increased risk of malignancy. Useful tumor markers that raise concern for ovarian cancer in children and adolescents include alpha-fetoprotein, lactate dehydrogenase, beta subunit of human chorionic gonadotropin, cancer antigen 125 and inhibin. However, their serum levels can neither confirm nor exclude malignancy. Management of pediatric ovarian masses needs to be curative and, when feasible, function-preserving and minimally invasive. Children and adolescents with an ovarian mass should be treated in specialized centers to avoid unnecessary oophorectomies and ensure the best possible outcome.

Background: Ovarian teratomas are the most common type of ovarian mass during childhood and adolescence. There is no clear guidance for the management of the ovarian teratomas in children and adolescents. It is necessary, however, to understand the feature of these tumours and the indications for operating on them, and to put subjective criteria for the management in elective or emergency presentations. Methods: Medical records of patients between the age of 12 and 20 years old that underwent surgery for ovarian teratomas at the Department of Obstetrics and Gynaecology in University Hospital of Ioannina, between January 2000 and August 2022, were reviewed. A medical database was searched between January 2000 and August 2022 with the following keywords: ovarian teratoma, paediatrics and adolescents. Results: We present the results of forty patients, with a mean age of 11.8 years of age. All patients had surgery, where three out of four underwent oophorectomy and only one out of four underwent ovary-sparing operation; similar results were found in the literature. Conclusion: Ovary-sparing treatment is more common recently compared with the past, such as MIS compared with laparotomy. Better-organised and -planned prospective multi-centre studies are necessary to gain a deeper knowledge of the physiology and prognostic factors of teratomas in children.

Neonatal thrombosis is a relatively common disease/condition and most often associated with predisposing genetic factors, underlying medical disorders and acquired trigger factors, such as iatrogenic interventions. Perinatal femoral artery thrombosis with no underlying risk factors has not been presented in the international literature. There has been no evidence of perinatal artery thrombosis with no underlying risk factors in the international literature. In this case presentation, a neonate was born via normal vaginal delivery at 39+4 weeks of gestation, in an otherwise uncomplicated pregnancy without risk factors, with marked discoloration of right lower abdomen and right lower limb, initially pale and eventually evolving to cyanotic. At the time of birth, such clinical presentation was a challenge to clinicians regarding diagnosis. The absence of right-sided palpable femoral pulses raised suspicion and led to a diagnostic approach primarily including ultrasonography. A right femoral blood clot blocking blood flow and resulting in the pale right lower limb was revealed via Doppler ultrasound. Screening test for thrombophilia was negative and neonatal arterial patency was achieved after administration of Low Molecular Weight Heparin (LMWH). After a 6-week course of treatment, vascular latency was fully restored and confirmed by ultrasound on day 10 post-partum. Clinicians should be aware of this unusual event that can be detrimental and endanger limb survival if immediate further action is not taken. Neonatal thrombosis can occur even at the time of birth; thus, clinicians should be alert in cases of neonates born with pale extremities. Low-risk cases could still pose a great threat to the survival of an extremity if appropriate action is not taken.

Objective: Fruits of lemon pepper (Zanthoxylum acanthopodium DC., Rutaceae) have been traditionally used as a spice and in folk medicine for treatment of diarrhea and stomachache. Stomachache could be associated with mycobacterial infection. The present study was designed to investigate the activity of Z. acanthopodium fruits against a non-infectious Mycobacterium smegmatis and to identify the important phytochemical constituent that is toxic towards mycobacteria. Materials and Methods: The broth microdilution method was used to determine the minimal inhibitory concentration (MIC) of ethyl acetate or hexane extract of green, young fruits of Z. acanthopodium. Effect of active extract (hexane) on cell membranee integrity was studied by measuring sodium and potassium leakage into extracelullar liquid using Atomic Absorbtion Spectrophotometer (AAS). Next, cell morphology was observedby using Scanning Microscope Electron (SEM). Column chromatography was used for fractionation and purification of hexane extract while the chemical structure of the active compound was determined using NMR technique. Rifampicin, an antimycobacterial compound, was used as positive control. Results: Hexane extract was active against M. smegmatis with an MIC of 64 µg/ml. Plant extract at the concentration of 128 µg/ml caused ions leakage. Concentration of sodium in extracellular liquid of cells treated with plant extract was significantly higher than that of untreated cells. SEM observation revealed cell wall deformation in cultures treated with the extract. NMR spectroscopy analysis of the most active fraction revealed that the compound that exerted toxicity on M. smegmatis was geranyl acetate. Conclusion: Geranyl acetate was an important constituent of Z. acanthopodium fruit that has antimycobacterial activity. Possibly, Z. acanthopodium fruit exert its toxic effects against M. smegmatisthrough damaging cell membrane.

Audiovisual content is highly employed in modern education curricula. This study aimed to identify students’ perceptions regarding the use of video in the courses they attend. Quantitative research was conducted among students of journalism and communication studies. Four courses were used for this study that employ video-based and video-assisted teaching strategies for active learning. Students used videos in different educational settings for flipped learning, blended learning, and autonomous self-paced learning. Their perceptions were examined through an extended form of the Technology Acceptance Model, encompassing more parameters (such as self-efficacy, perceived enjoyment, satisfaction, attitude, and intention of use) to identify causal relationships. In addition, the roles of technical parameters, such as internet connection, audio quality, and video quality, along with the duration of a video, were explored. The findings reveal students’ positive attitudes towards videos, and all factors were significantly related to the intention of use. In addition, the technical aspects of the video did not constitute a problem in the educational process, most likely due to contemporary quality in internet connections and audiovisual productions along with playback equipment. The duration of the videos was also not presented as an issue as there are flexible ways to embed videos in the learning process. Overall, videos can be used in multiple active learning scenarios to enhance students’ motivation and engagement and provide a joyful, collaborative, and hospitable learning environment. Students are familiar with digital tools and technologies and seem to enjoy audiovisual material in the learning process.

Diagnosis of autism spectrum disorder (ASD) before the age of three years is a challenge. Analyzing the present practice may help reaching that goal. Aim: To investigate developmental abnormalities and diagnostic pathway of ASD patients in pediatric practice. Methods: Retrospective cross-sectional study of 192 children aged 13 months to 17 years 11 months (average 4 years 9 months), investigated in an outpatient and hospital setting from January 2015 to June 2018 by a semi-structured history and clinical examination, and diagnosed with ASD by Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-5) criteria. Results: Behavioral peculiarities were detected in the history of the first two years of life in 74.8% of the subjects. The first developmental abnormalities were noticed by the parents at ages from 8 to 36 months (mean 15.6 months) and were predominantly in speech (in 94.6%) and non-verbal communication (11.3%). Developmental regression was reported in 42.1% of the patients occurring between the ages of 6 and 50 months (mean 17.9 months), affecting most commonly speech (88.4% of cases), non-verbal communication (29.2%), and behavior (12.8%). By history, the first manifestations of ASD were noticed at ages from 8 months to 84 months (mean 18.5 months), and were disorders of expressive speech (in 66.7% of cases), receptive speech (in 45.8%), non-verbal communication (35.4%), behavior (27.6%), play (8.9%), socialization (5.7%), and joint attention (2.1%). The most common motive for specialized consultation was delay in language development—in 84.6% of children. The age of ASD diagnosis varied between 12 and 132 months (mean 39.7 months), and the time period between first ASD manifestations and diagnosis was in the range of 0 to 79 months (mean 23.3 months). Many symptoms of abnormal social communication, unnoticed by parents, were detected objectively in more than 95% of the cases—absent or rare spontaneous or reciprocal smile; lack of sharing of interest or affect; abnormal eye contact; lack of finger pointing; lack of gaze to a pointed object; poor facial expressions; lack of imaginary play, etc. Conclusions: Almost two years are needed for diagnosing abnormal development in other domains besides speech in ASD patients. Diagnosis before the age of three years can be achieved by focusing parents’ and pediatricians’ attention on social communication and behavior in patients with speech delay or developmental regression.

Food waste represents 25−35% of the European Municipal Solid Waste (MSW) [1], thus its diversion into innovative utilization streams is critical for sustainable waste management and the achievement of circularity. [...]

The comorbidity of autistic spectrum disorder (ASD) and epilepsy has been widely discussed but many questions still remain unanswered. The aim of this study was to establish the occurrence of epilepsy among children with ASD to define the type of epileptic seizures and syndromes, the age of onset of epilepsy, EEG abnormalities, the used antiepileptic drugs and the therapeutic responses for seizures and autistic behavior, as well as to find some correlations between epilepsy and gender, etiology and intellectual disability (ID). A retrospective study of medical files of 59 patients (aged 1–18 years) with ASD during a 5-year period was performed. ASD diagnosis was based on the DSM-5 diagnostic criteria. The patients were examined with a detailed medical history, physical and neurological examination, as well as some additional functional, imaging, laboratory and genetic investigations ASD etiology was syndromic in 9, probable syndromic in 9, and idiopathic in 41 children. ID was established in 90% of ASD children, and epilepsy in 44.4%. The onset of epilepsy prevailed before 7 years of age. The most common seizure types were focal with or without secondary generalization (53.4%). Focal epileptiform EEG abnormalities prevailed. Therapeutic response to seizures was good: 58% were seizure-free, while 27% had >50% seizure reduction but no improvement in autistic behavior. There was no correlation between epilepsy and either occurrence or degree of ID. There was a correlation between the frequency of epileptic seizures and the degree of ID. There was no significant difference among epilepsy rates in different etiologic, gender, and ID groups, probably because of the high percentage of ID and because this was a hospital-based study. Our study showed a significant percentage of epilepsy in ASD population and more than 1/4 were of symptomatic etiology. Those could be managed with specific treatments based on the pathophysiology of the gene defect.

Background: Acute generalized exanthematous pustulosis (AGEP) is a rare and severe cutaneous reaction usually triggered by drugs. Other causative factors such as viral infections are rarely involved. In this study, we report a case of AGEP caused by a spider bite. Case Summary: A 56-year-old woman was referred to the allergy unit after a spider bite at the left popliteal fossa, while gardening, 5 days earlier. The offending spider was captured and identified by an entomologist as belonging to the Loxosceles rufescens species. No acute reaction was observed; however, after 24 hours, due to the occurrence of typical dermonecrotic skin lesions associated with erythema and edema, Cefuroxime and Clindamycin were administered intramuscularly after medical advice was given. Almost 72 hours after the spider bite, an erythematous and partly edematous eruption appeared locally in the gluteus area bilaterally, which progressively expanded to the trunk, arms and femors. Within 24 hours dozens of small, pinhead sized, non- follicular pustules were present, mainly in the folds. The patient complained of a burning sensation of the skin in addition to pruritus; and simultaneously had a fever of 38-39 °C as the eruption expanded. Discussion: A spider bite may represent a possible causative factor of AGEP. A spider's venom contains sphingomyelinase that stimulates the release of IL8 and GM-CSF, which are involved in AGEP pathogenesis. Whether or not the con-current use of antibiotics has an effect in AGEP appearance when combined with a spider's venom, cannot be excluded.

Advances in solid-state nuclear magnetic resonance (ssNMR) spectroscopy and cryogenic electron microscopy (cryoEM) have revealed the polymorphic nature of the amyloid state of proteins. Given the association of amyloid with protein misfolding disorders, it is important to understand the principles underlying this polymorphism. To address this problem, we combined computational tools to predict the specific regions of the sequence forming the β-spine of amyloid fibrils with the availability of 30, 83 and 24 amyloid structures deposited in the Protein Data Bank (PDB) and Amyloid Atlas (AAt) for the amyloid β (Aβ) peptide, α-synuclein (αS), and the 4R isoforms of tau, associated with Alzheimer's disease, Parkinson's disease, and various tauopathies, respectively. This approach enabled a statistical analysis of sequences forming β-sheet regions in amyloid polymorphs. We computed for any given sequence residue n the fraction of PDB/AAt structures in which that residue adopts a β-sheet conformation (Fβ(n)) to generate an experimental, structure-based profile of Fβ(n) vs n, which represents the β-conformational preference of any residue in the amyloid state. The peaks in the respective Fβ(n) profiles of the three proteins, corresponding to sequence regions adopting more frequently the β-sheet structural core in the various fibrillar structures, align very well with the peaks identified with five predictive algorithms (ZYGGREGATOR, TANGO, PASTA, AGGRESCAN, and WALTZ). These results indicate that, despite amyloid polymorphism, sequence regions most often forming the structural core of amyloid have high hydrophobicity, high intrinsic β-sheet propensity and low electrostatic charge across the sequence, as rationalised and predicted by the algorithms., (© 2025 The Author(s).)

Soluble oligomers arising from the aggregation of the amyloid beta peptide (Aβ) have been identified as the main pathogenic agents in Alzheimer's disease (AD). Prefibrillar oligomers of the 42-residue form of Aβ (Aβ 42 O) show membrane-binding capacity and trigger the disruption of Ca 2+ homeostasis, a causative event in neuron degeneration. Since bioactive lipids have been recently proposed as potent protective agents against Aβ toxicity, we investigated the involvement of sphingosine 1-phosphate (S1P) signalling pathway in Ca 2+ homeostasis in living neurons exposed to Aβ 42 O. We show that both exogenous and endogenous S1P rescued neuronal Ca 2+ dyshomeostasis induced by toxic Aβ 42 O in primary rat cortical neurons and human neuroblastoma SH-SY5Y cells. Further analysis revealed a strong neuroprotective effect of S1P 1 and S1P 4 receptors, and to a lower extent of S1P 3 and S1P 5 receptors, which activate the G i -dependent signalling pathways, thus resulting in the endocytic internalization of the extrasynaptic GluN2B-containing N-methyl-D-aspartate receptors (NMDARs). Notably, the S1P beneficial effect can be sustained over time by sphingosine kinase-1 overexpression, thus counteracting the down-regulation of the S1P signalling induced by Aβ 42 O. Our findings disclose underlying mechanisms of S1P neuronal protection against harmful Aβ 42 O, suggesting that S1P and its signalling axis can be considered promising targets for therapeutic approaches for AD., (© 2022 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Trodusquemine is an aminosterol with a variety of biological and pharmacological functions, such as acting as an antimicrobial, stimulating body weight loss and interfering with the toxicity of proteins involved in the development of Alzheimer's and Parkinson's diseases. The mechanisms of interaction of aminosterols with cells are, however, still largely uncharacterized. Here, by using fluorescently labeled trodusquemine (TRO-A594 and TRO-ATTO565), we show that trodusquemine binds initially to the plasma membrane of living cells, that the binding affinity is dependent on cholesterol, and that trodusquemine is then internalized and mainly targeted to lysosomes after internalization. We also found that TRO-A594 is able to strongly and selectively bind to myelinated fibers in fixed mouse brain slices, and that it is a marker compatible with tissue clearing and light-sheet fluorescence microscopy or expansion microscopy. In conclusion, this work contributes to further characterize the biology of aminosterols and provides a new tool for nerve labeling suitable for the most advanced microscopy techniques., (© 2022 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Alzheimer's disease is characterized by the accumulation in the brain of the amyloid β (Aβ) peptide in the form of senile plaques. According to the amyloid hypothesis, the aggregation process of Aβ also generates smaller soluble misfolded oligomers that contribute to disease progression. One of the mechanisms of Aβ oligomer cytotoxicity is the aberrant interaction of these species with the phospholipid bilayer of cell membranes, with a consequent increase in cytosolic Ca 2+ levels, flowing from the extracellular space, and production of reactive oxygen species (ROS). Here we investigated the relationship between the increase in Ca 2+ and ROS levels immediately after the exposure to misfolded protein oligomers, asking whether they are simultaneous or instead one precedes the other. Using Aβ 42 -derived diffusible ligands (ADDLs) and type A HypF-N model oligomers (OAs), we followed the kinetics of ROS production and Ca 2+ influx in human neuroblastoma SH-SY5Y cells and rat primary cortical neurons in a variety of conditions. In all cases we found a faster increase of intracellular Ca 2+ than ROS levels, and a lag phase in the latter process. A Ca 2+ -deprived cell medium prevented the increase of intracellular Ca 2+ ions and abolished ROS production. By contrast, treatment with antioxidant agents prevented ROS formation, did not prevent the initial Ca 2+ flux, but allowed the cells to react to the initial calcium dyshomeostasis, restoring later the normal levels of the ions. These results reveal a mechanism in which the entry of Ca 2+ causes the production of ROS in cells challenged by aberrant protein oligomers., (© 2022. The Author(s).)

Background: This study aimed to evaluate the effect of local nasal desmopressin premedication on blood loss and the quality of surgical field in Functional Endoscopic Sinus Surgery (FESS)., Material and Methods: In a randomized clinical trial, patients referred for FESS to treat their bilateral chronic rhinosinusitis were recruited. The participants were adults (≥18 years). They were randomly assigned (1:1:1) to receive low-dose (20 μg) or high-dose (40 μg) intranasal desmopressin (DDAVP) or placebo 60 min before the induction of general anesthesia. Standard FESS was performed by the same surgeon. The primary outcomes were volume of intraoperative bleeding and the quality of surgical field. Clean surgical field was defined as a score ≤2 on the Boezaart grading system., Results: A total of 120 patients were included on an intention-to-treat basis (mean age: 41.0 years; 40 women, 80 men). There were no significant differences in primary outcomes between low-dose DDAVP and placebo. As for the volume of blood loss, however, there was a significant difference between high-dose DDAVP and placebo (mean difference: 29.6 ml; adjusted Cohen's d: -1.02; p  < .001). Also, in the high-dose DDAVP, the probability of having a good surgical field over time was about two times higher than in the placebo group (RRs for first and second surgical sides: 1.89 and 2.18). The number needed to treat for the two time points was 1.6 and 1.3, respectively., Conclusion: The present study showed that the use of desmopressin at a dose of 40 μg 1 h before surgery can reduce bleeding and improve the quality of the surgical field. Further studies are recommended to be able to generalize these findings to other ENT surgeries., Level of Evidence: 1b., Competing Interests: The authors declare that no competing interests exist., (© 2022 The Authors. Laryngoscope Investigative Otolaryngology published by Wiley Periodicals LLC on behalf of The Triological Society.)

Competing Interests: None

Accumulation of ubiquitin-positive, tau- and α-synuclein-negative intracellular inclusions of TDP-43 in the central nervous system represents the major hallmark correlated to amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U). Such inclusions have variably been described as amorphous aggregates or more structured deposits having amyloid properties. Here we have purified full-length TDP-43 (FL TDP-43) and its C-terminal domain (Ct TDP-43) to investigate the morphological, structural and tinctorial features of aggregates formed in vitro by them at pH 7.4 and 37 °C. AFM images indicate that both protein variants show a tendency to form filaments. Moreover, we show that both FL TDP-43 and Ct TDP-43 filaments possess a largely disordered secondary structure, as ascertained by far-UV circular dichroism and Fourier transform infra-red spectroscopy, do not bind Congo red and induce a very weak increase of thioflavin T fluorescence, indicating the absence of a clear amyloid-like signature.

Alzheimer's disease, which is the most common form of dementia, is characterized by the aggregation of the amyloid β peptide (Aβ) and by an impairment of calcium homeostasis caused by excessive activation of glutamatergic receptors (excitotoxicity). Here, we studied the effects on calcium homeostasis caused by the formation of Aβ oligomeric assemblies. We found that Aβ oligomers cause a rapid influx of calcium ions (Ca 2+ ) across the cell membrane by rapidly activating extrasynaptic N -methyl-d-aspartate (NMDA) receptors and, to a lower extent, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. We also observed, however, that misfolded oligomers do not interact directly with these receptors. Further experiments with lysophosphatidylcholine and arachidonic acid, which cause membrane compression and stretch, respectively, indicated that these receptors are activated through a change in membrane tension induced by the oligomers and transmitted mechanically to the receptors via the lipid bilayer. Indeed, lysophosphatidylcholine is able to neutralize the oligomer-induced activation of the NMDA receptors, whereas arachidonic acid activates the receptors similarly to the oligomers with no additive effects. An increased rotational freedom observed for a fluorescent probe embedded within the membrane in the presence of the oligomers also indicates a membrane stretch. These results reveal a mechanism of toxicity of Aβ oligomers in Alzheimer's disease through the perturbation of the mechanical properties of lipid membranes sensed by NMDA and AMPA receptors.

Alzheimer's disease (AD) is the most prevalent form of dementia and soluble amyloid β (Aβ) oligomers are thought to play a critical role in AD pathogenesis. Cellular prion protein (PrP C ) is a high-affinity receptor for Aβ oligomers and mediates some of their toxic effects. The N -terminal region of PrP C can interact with Aβ, particularly the region encompassing residues 95-110. In this study, we identified a soluble and unstructured prion-derived peptide (PrP 107-120 ) that is external to this region of the sequence and was found to successfully reduce the mitochondrial impairment, intracellular ROS generation and cytosolic Ca 2+ uptake induced by oligomeric Aβ 42 ADDLs in neuroblastoma SH-SY5Y cells. PrP 107-120 was also found to rescue SH-SY5Y cells from Aβ 42 ADDL internalization. The peptide did not change the structure and aggregation pathway of Aβ 42 ADDLs, did not show co-localization with Aβ 42 ADDLs in the cells and showed a partial colocalization with the endogenous cellular PrP C . As a sequence region that is not involved in Aβ binding but in PrP self-recognition, the peptide was suggested to protect against the toxicity of Aβ 42 oligomers by interfering with cellular PrP C and/or activating a signaling that protected the cells. These results strongly suggest that PrP 107-120 has therapeutic potential for AD.

The involvement of transactivation response (TAR) DNA-binding protein 43 (TDP-43) in neurodegenerative diseases was revealed in 2006, when it was first reported to be the main component of the intracellular inclusions in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration. After 12 yr it is not yet possible to purify to a reasonable yield and in a reproducible manner a stable full-length protein, which has limited so far the characterization of its structure, function, molecular interactors, and pathobiology. Using a novel protocol we have achieved the purification of the full-length TDP-43, with both a short pectate lyase B tag and a glutathione S -transferase tag, which consisted in its expression in bacteria, solubilization from inclusion bodies, purification under denaturing conditions, refolding, and a final size exclusion chromatography (SEC) step. Differential scanning fluorimetry was used to find the best buffers and combination of additives to increase both its solubility and its stability. The protein is pure, as determined with electrophoresis, Western blotting, and mass spectrometry; properly refolded, as revealed by circular dichroism and fluorescence spectroscopies; functional, because it binds to DNA and protein partners; and stable to degradation and aggregation in a physiologic solution. Analyses with dynamic light scattering and SEC revealed that the protein is a dimer.-Vivoli Vega, M., Nigro, A., Luti, S., Capitini, C., Fani, G., Gonnelli, L., Boscaro, F., Chiti, F. Isolation and characterization of soluble human full-length TDP-43 associated with neurodegeneration.

The formation of misfolded protein oligomers during early stages of amyloid aggregation and the activation of neuroinflammatory responses are two key events associated with neurodegenerative diseases. Although it has been established that misfolded oligomers are involved in the neuroinflammatory process, the links between their structural features and their functional effects on the immune response remain unknown. To explore such links, we took advantage of two structurally distinct soluble oligomers (type A and B) of protein HypF-N and compared the elicited microglial inflammatory responses. By using confocal microscopy, protein pull-down, and high-throughput mass spectrometry, we found that, even though both types bound to a common pool of microglial proteins, type B oligomers-with a lower solvent-exposed hydrophobicity-showed enhanced protein binding, correlating with the observed inflammatory response. Furthermore, the interactome associated with inflammatory-mediated neurodegeneration revealed previously unidentified receptors and signaling molecules likely to be involved in the oligomer-elicited innate immune response.

Frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS) are progressive and fatal neurodegenerative disorders showing mislocalization and cytosolic accumulation of TDP-43 inclusions in the central nervous system. The decrease in the efficiency of the clearance systems in aging, as well as the presence of genetic mutations of proteins associated with cellular proteostasis in the familial forms of TDP-43 proteinopathies, suggest that a failure of these protein degradation systems is a key factor in the aetiology of TDP-43 associated disorders. Here we show that the internalization of human pre-formed TDP-43 aggregates in the murine neuroblastoma N2a cells promptly resulted in their ubiquitination and hyperphosphorylation by endogenous machineries, mimicking the post-translational modifications observed in patients. Moreover, our data identify mitochondria as the main responsible sites for the alteration of calcium homeostasis induced by TDP-43 aggregates, which, in turn, stimulates an increase in reactive oxygen species and, finally, caspase activation. The inhibition of TDP-43 proteostasis in the presence of selective inhibitors against the proteasome and macroautophagy systems revealed that these two systems are both severely involved in TDP-43 accumulation and have a strong influence on each other in neurodegenerative disorders associated with TDP-43.

Amyotrophic lateral sclerosis and frontotemporal lobar degeneration with ubiquitin-positive inclusions are neurodegenerative disorders that share the cytosolic deposition of TDP-43 (TAR DNA-binding protein 43) in the CNS. TDP-43 is well known as being actively degraded by both the proteasome and macroautophagy. The well-documented decrease in the efficiency of these clearance systems in aging and neurodegeneration, as well as the genetic evidence that many of the familial forms of TDP-43 proteinopathies involve genes that are associated with them, suggest that a failure of these protein degradation systems is a major factor that contributes to the onset of TDP-43-associated disorders. Here, we inserted preformed human TDP-43 aggregates in the cytosol of murine NSC34 and N2a cells in diffuse form and observed their degradation under conditions in which exogenous TDP-43 is not expressed and endogenous nuclear TDP-43 is not recruited, thereby allowing a time zero to be established in TDP-43 degradation and to observe its disposal kinetically and analytically. TDP-43 degradation was observed in the absence and presence of selective inhibitors and small interfering RNAs against the proteasome and autophagy. We found that cytosolic diffuse aggregates of TDP-43 can be distinguished in 3 different classes on the basis of their vulnerability to degradation, which contributed to the definition-with previous reports-of a total of 6 distinct classes of misfolded TDP-43 species that range from soluble monomer to undegradable macroaggregates. We also found that the proteasome and macroautophagy-degradable pools of TDP-43 are fully distinguishable, rather than in equilibrium between them on the time scale required for degradation, and that a significant crosstalk exists between the 2 degradation processes.-Cascella, R., Fani, G., Capitini, C., Rusmini, P., Poletti, A., Cecchi, C., Chiti, F. Quantitative assessment of the degradation of aggregated TDP-43 mediated by the ubiquitin proteasome system and macroautophagy., (© FASEB.)

Amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin positive inclusions (FTLD-U) are two clinically distinct neurodegenerative conditions sharing a similar histopathology characterized by the nuclear clearance of TDP-43 and its associated deposition into cytoplasmic inclusions in different areas of the central nervous system. Given the concomitant occurrence of TDP-43 nuclear depletion and cytoplasmic accumulation, it has been proposed that TDP-43 proteinopathies originate from either a loss-of-function (LOF) mechanism, a gain-of-function (GOF) process, or both. We have addressed this issue by transfecting murine NSC34 and N2a cells with siRNA for endogenous murine TDP-43 and with human recombinant TDP-43 inclusion bodies (IBs). These two strategies allowed the depletion of nuclear TDP-43 and the accumulation of cytoplasmic TDP-43 aggregates to occur separately and independently. Endogenous and exogenous TDP-43 were monitored and quantified using both immunofluorescence and Western blotting analysis, and nuclear functional TDP-43 was measured by monitoring the sortilin 1 mRNA splicing activity. Various degrees of TDP-43 cytoplasmic accumulation and nuclear TDP-43 depletion were achieved and the resulting cellular viability was evaluated, leading to a quantitative global analysis on the relative effects of LOF and GOF on the overall cytotoxicity. These were found to be ∼55% and 45%, respectively, in both cell lines and using both readouts of cell toxicity, showing that these two mechanisms are likely to contribute apparently equally to the pathologies of ALS and FTLD-U., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

Purpose: To compare spectrophotometric vs. visual color identification of tooth shade on the basis of three different commercially available shade guides., Methods: On 54 subjects, two operators (with >10 years experience) independently selected the best match of the middle third of one of the patient's upper central incisors, using consecutively three shade guides (Vita Lumin Vacuum, Chromascop, Vita 3D Master). After each individual shade match, the same operator performed a shade measurement with a SpectroShade spectrophotometer., Results: The Vita Lumin shade guide appeared to provide the most consistent results between human and spectrophotometrical evaluation, although Vita 3D Master offered a greater variety of colors. In approximately 47% of the cases, the spectrophotometer provided more accurate results than visual selection. From a clinical point of view, this is a noteworthy improvement in the complex process of color match for indirect restoration.