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Forensic autopsy is an important tool for the proper management of non-natural deaths in minors. However, it seems that autopsy in minors is a practice which may not be performed routinely. In this framework, we conducted a study analyzing autopsies of minors (under 18 years of age in Italy) performed at the Institute of Forensic Medicine in Milan in the period 2001-2019. For the period 2015-2019, we extrapolated all deaths due to non-natural causes in minors to investigate how many and which of these deaths were not subjected to forensic autopsy. Of the total, 344 minors (235 males and 109 females) underwent autopsies, with an overall downward trend of about 80% since 2004. Most autopsies occurred between the ages of 0 and 1 year, and the fewest between the ages of 5 and 9 years. The place of death was home in most cases, and accidental death was most common, followed by natural death, suicide, and homicide, with prevalence varying by age group. Blunt force trauma predominated among accidental death in all age groups, followed by asphyxia. Similar findings were observed for suicides, although there was a more differentiated pattern for suicides between the ages of 15 and 17 years. Among homicides, blunt force trauma, asphyxia, and gunshot wounds were fairly evenly distributed across all age groups. Between 2015 and 2019, a total of 86 minors died of a non-natural cause, and a forensic autopsy was performed in only 33 cases (38%). Our data shows that fewer and fewer autopsies are being performed over the last years, which indicates a dangerous lack of forensic investigation of children and adolescent deaths, with enormous implications for prevention of child abuse., (© 2023. The Author(s).)

Background: The pathophysiology of mental illnesses is not fully understood, leading to insufficient remission, frequent adverse drug reactions, and treatment resistance. Pharmacogenetic (PGx) testing, a personalized approach recently adopted also in psychiatry, can guide effective drug therapy and minimize side effects. The objective of this study was to determine the perspective of Slovak clinicians regarding the integration of PGx testing in psychiatric clinical practice., Methods: Questionnaires covering various aspects such as prior experience with PGx testing, self-perceived competence, perceived utility, potential risks and challenges were distributed directly to attendees at two psychiatric conferences held in Slovakia in 2023 and their responses were statistically analysed., Results: Out of 54 respondents, only 7.4% had previous experience with PGx in clinical practice. The most clinicians felt that they should possess the skills to apply PGx testing in psychiatric clinical practice and were enthusiastic about increasing their expertise. They found PGx useful in medication selection, adverse effect management, and treatment-resistant depression. The primary concerns centered around the lack of well-defined guidelines and the financial considerations linked to the testing., Conclusions: Considering the participants' interest in PGx and its integration into clinical practice, educational programmes based on recommendations, guidelines, and convincing evidence could be organized (Tab. 4, Ref. 30). Text in PDF www.elis.sk Keywords: pharmacogenetics, pharmacogenomics, psychiatry, personalized treatment, Slovakia, perspective.

Most victims of physical violence sooner or later will access a hospital or medical cabinet because of that violence, and in particular emergency departments (EDs). This paper aims to analyze the performance of emergency ward clinicians in the forensic management of such victims by examining the activities carried out and the data reported. A total of 991 medical records were extrapolated from the database of the ED of the Policlinico of Milan in an average pre-pandemic 1-year activity. For each medical record, 16 parameters were analyzed in-depth including epidemiological data, information on the type of violent actions, injuries, and time between the infliction of the lesion and access to the ED. In the vast majority of cases, all the actions with medicolegal implications had been neglected by health professionals causing loss of data not only for the justice system but especially for correctly interpreting what happened and taking appropriate measures to protect the patient/victim. Hence, given that clinicians in EDs are busy with non-forensic clinical tasks (and rightly so), it should be ensured that there be specific forensic clinical personnel. However, it is crucial that when unfortunately there can be no forensic staff, at least the clinicians who work in the ED are properly trained to correctly apply essential medicolegal measures. Overall, timely and informed medical and forensic intervention is possible and necessary for the improvement and maintenance of the mental and physical health of victims of violence., (© 2023. The Author(s).)

COVID-19 pandemic had a great impact on mental health, both in the general population and psychiatric patients. Little is known about the difference between these two populations in perceiving the pandemic as a traumatic event. The aim of the study was to compare psychiatric patients and healthy controls (HC) in terms of change over time of post-traumatic (PTSD) symptoms. Demographic and clinical variables were collected. Impact of Event Scale Revised (IES-R) scores were registered at T1 as lockdown period (March-April 2020) and T2 as restarting (May-June 2020). Descriptive analyses and linear regression models were performed. A total of 166 outpatients and 57 HC were recruited. Time (F = 15.76; p < 0.001) and diagnosis (F = 4.94; p < 0.001) had a significant effect on the change of IES-R scores, which resulted T1 > T2 (p < 0.001), except for subjects affected by Obsessive-Compulsive Disorder (OCD). Overall, IES-R scores were < in patients than in HC (p = 0.02), particularly in the schizophrenia (SKZ) subgroup (p < 0.001). IES-R scores of subjects with personality disorders (PDs) resulted to be > HC, although not statistically significant. The lockdown period was perceived as more traumatic than the reopening phase by both groups, with the exception of OCD patients, probably because of the clinical worsening associated with the urge of control against risks of contamination. Overall, HC reported more PTSD symptoms than psychiatric patients did, particularly SKZ ones. PD patients, in contrast, may be more vulnerable to PTSD symptoms probably as a result of poor coping skills. Together with OCD patients, subjects with PDs may need closer monitoring during the different phases of the pandemic. Trial Registration: ClinicalTrials.gov Identifier: NCT04694482.

The European Search Catalogue for Plant Genetic Resources (EURISCO) is a central entry point for information on crop plant germplasm accessions from institutions in Europe and beyond. In total, it provides data on more than two million accessions, making an important contribution to unlocking the vast genetic diversity that lies deposited in >400 germplasm collections in 43 countries. EURISCO serves as the reference system for the Plant Genetic Resources Strategy for Europe and represents a significant approach for documenting and making available the world's agrobiological diversity. EURISCO is well established as a resource in this field and forms the basis for a wide range of research projects. In this paper, we present current developments of EURISCO, which is accessible at http://eurisco.ecpgr.org., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

During the COVID-19 pandemic, not only crowded refugee camps and immigration detention centers, but also receptions were places in which outbreaks occurred. To date there has been no report of the application of a COVID-19 surveillance system in reception centers for unaccompanied foreign minors only, who most of all deserve the utmost attention. Aware of this critical issue, we implemented a pilot COVID-19 surveillance program at the Zendrini center in Milan. It was started in September 2021 and was carried out for 4 months. Nasopharyngeal antigenic swabs were adopted. One day a week, two forensic physicians performed the first antigenic swab to minors who had just entered the center, or a monitoring swab after 15 days to those who were still hosted at the center. Operators were also swabbed for surveillance. A total of 80 subjects were enrolled and divided into 68 (72.5%) unaccompanied foreign minors and 22 (27.5%) operators. A total of 178 antigenic nasopharyngeal swabs were performed and tested negative. Regarding the monitoring activities, it was found that the minimum number of swabs per subject was 1 and the maximum number was 7, with an average value of 2.2 per individual. Having been able to confirm the absence of SARS-CoV-2 within the community represented a way to protect individual and collective health that could not have been pursued otherwise. Only inclusive approaches can allow communities and societies to respond more effectively to this crisis, and reduce the risk of future ones, intended as both upcoming COVID-19 waves and new infectious diseases.

Older adults are subject to vulnerability and abusive behaviors have serious negative health consequences. Since the phenomenon is underestimated, several challenges are faced to prevent, diagnose, and treat abuses on older adults. In this context, clinical forensic medicine practitioners can play a pivotal role. A retrospective monocentric analysis of all the files concerning victims of violence observed in the SVSeD Center in Milan over the age of 65 for a five-year period was performed. For each medical report, data regarding the variables of the victims, the type of violence, and the variables of the perpetrators were analysed. Older adult victims of abuses were 166. Abuses against women were more frequent than against men (less than 6 % of consultations). In one third of cases, subjects also suffered from disabling diseases and reported signs of physical violence. Perpetrators were male known individuals in almost 90 % of cases. Violence was perpetrated by a partner or a family member. The present study may help in identifying risk factors for domestic violence against older adults. Consequently, preventive policies should be designed to interrupt the vicious circle of violence. Older adult abuse deserves additional focus and better education for healthcare professionals and further research should be carried out to better understand the actual epidemiology of the phenomenon and to develop therapeutical and caring strategies., Competing Interests: Conflict of interest Not applicable., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Introduction: Sexual violence against young girls is a pervasive multifaceted phenomenon which embraces several different forced sexual acts, including attempted and/or completed rape, sexual coercion and harassment, and sexual contact with force or threat of force. The aim of this study is to evaluate the characteristics of sexual violence in adolescent girls, by a retrospective analysis of 731 consecutive cases of sexually abused girls., Materials and Methods: We analyzed demographic characteristics, risk factors for rape, and the eventual subsequent presence and type of ano-genital lesions., Results: We found that sexual violence with penetration was perpetrated in 591 (80%) cases. Vulnerability factors related with a major risk of rape were age >17 years old and consumption of alcohol and/or other drugs. About 196 (55%) victims had at a least one genital lesion., Conclusion: A prompt identification of red flags of sexual violence may help physicians in suspecting and managing cases of sexual assault in adolescent girls, even in the absence of typical lesions., Competing Interests: The authors report no conflicts of interest for this work., (© 2022 Barbara et al.)

Femicide constitutes a leading cause of premature deaths for women, yet it has been the subject of limited research until recently. Enhanced data collection and analysis on killings of women and girls are necessary to understand and address this unrelenting phenomenon. This study examines all cases of female homicide encountered at the Institute of Legal Medicine of Milan (Italy) spanning from 1999 to 2019; data from 2020/2021 were shown separately given the bias that the forced cohabitation and stay at home during the lockdowns of the SARS-CoV-2 pandemic may represent regarding violence against women and femicide. In this study, specific factors were considered, including the age and nationality of the victims, the place of recovery of the bodies, the victim's relationship to the perpetrator and the injuries they suffered. As a result, 200 female killings were found among the over 15,000 autopsies and 535 homicides investigated at the Institute of Legal Medicine of Milan from 1999 to 2019, representing an average of 9.5 femicides yearly. The majority of victims were Italian (74%) and half were aged between 18 and 49 years old. The killings were overwhelmingly committed in the domestic setting (78.5%) by male perpetrators (at least 85%), related to the victims as intimate or ex-intimate partners and members of the family (73.5%). The homicides were mainly perpetrated with sharp (32%) or blunt instruments (21.5%), shooting (18.5%) and asphyxiation (16.5%). This study is part of a growing effort to enhance data collection and analysis on femicide. Studying and monitoring the rates of femicide (or "femicide watch") will permit to better understand, reduce, and finally end femicide globally., (Copyright © 2022 The Chartered Society of Forensic Sciences. Published by Elsevier B.V. All rights reserved.)

SARS-CoV-2 infection causes a pulmonary disease (COVID-19) which spread worldwide generating fear, anxiety, depression in the general population as well as among subjects affected by mental disorders. Little is known about which different psychopathological changes the pandemic caused among individuals affected by different psychiatric disorders, which represents the aim of the present study. Specific psychometric scales were administered at three time points: T0 as outbreak of pandemic, T1 as lockdown period, T2 as reopening. Descriptive analyses and linear regression models were performed. A total of 166 outpatients were included. Overall, psychometric scores showed a significant worsening at T1 with a mild improvement at T2. Only psychopathology in schizophrenia (SKZ) patients and obsessive-compulsive (OC) symptoms did not significantly improve at T2. Subjects affected by personality disorders (PDs) resulted to be more compromised in terms of general psychopathology than depressed and anxiety/OC ones, and showed more severe anxiety symptoms than SKZ patients. In conclusion, subjects affected by PDs require specific clinical attention during COVID-19 pandemic. Moreover, the worsening of SKZ and OC symptoms should be strictly monitored by clinicians, as these aspects did not improve with the end of lockdown measures. Further studies on larger samples are needed to confirm our results. ClinicalTrials.gov Identifier: NCT04694482., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Nowadays, the clinical forensic medical management of migration flows comprises the age assessment of unaccompanied minors. The process of age estimation is a fundamental pillar for legally ensuring the minors' rights and their protection needs. The procedure is complex and involves different phases and actors, from medical doctors to law enforcement officers. The present study aimed to investigate the performance of Greulich and Pyle, Demirjian, and Mincer methods when performed by raters both trained and without training. Also, the interrater reliability within groups of raters from different areas of expertise was evaluated. A total of 36 participants were enrolled for this study, divided in two groups according to their level of experience with age estimation methods. Each rater was asked to allocate stages and standards for age assessment, evaluating ten orthopantomograms and ten hand-wrist roentgenograms. The interrater reliability expressed through the Fleiss Kappa coefficient and the agreement with the reference standard were calculated. The results showed that none of the categories analyzed could reach a good interrater reliability (ĸ > 0.8) for both methods. The study results highlighted variation and disagreement in the interpretation of the sample among raters and in the subsequent stages and standards allocation. In conclusion, the results of this study highlight that expertise does influence the reliability of the most utilized methods of age estimation of living individuals and stress the importance of proper training and practice, which could greatly increase the accuracy of age assessments., (© 2021. The Author(s).)

Europe is very active in terms of conserving plant genetic resources, with hundreds of genebanks and thousands of dedicated people involved. However, the resulting infrastructure is, along with being very expensive, far from efficient and not very reliable. In this opinion paper, the authors describe how this situation arose, and why the European Cooperative Programme for Plant Genetic Resources (ECPGR), the collaborative umbrella organization of the European countries involved, has not been able to improve this situation so far significantly. The principles of the decentralized virtual genebank (AEGIS) are described, and an analysis is made of the reasons for its lack of success. Possible changes for making AEGIS a success, or at least steps in the right direction, are proposed. These changes center around the creation of a system of certified genebanks with proper quality management, guaranteeing the long-term conservation of, and immediate access to the plant genetic resources conserved in it.

The identification of decedents has always been a central issue in forensic pathology, for ethical, criminal, and administrative reasons, but today, it needs more attention due to issues related not only to migration but also to the weakening of family ties. This article presents a descriptive study discussing the Italian regulatory situation developed in the last decade to face the many identification issues, with all its improvements and flaws. Hence, data gathered in 25 years of at the Institute of Legal Medicine of Milan and the epidemiology of unidentified decedents are illustrated. Briefly, from 1995 to 2019, the number of unidentified human remains with no identity or requesting verification of identity amounts to 726, i.e., 3% circa of all autopsies performed at the Institute, with an average of 29 individuals per year. In total, 528 (72.7%) individuals were successfully identified, 100 (13.8%) remained without an identity, and 98 (13.5%) individuals remained with suspected yet unconfirmed identities. Percentages for each identification technique are displayed, with insight into the role of forensic anthropology and odontology compared to genetics, and into the misuse of non-scientific methods allowed by Public Prosecutors. All the data is compared, as much as possible, with the very few recent studies concerning the problem worldwide. Finally, the article aims to show the Italian experience in dealing with unidentified bodies, in order to provide food for thought for other countries toward a discussion regarding a global issue which is sometimes taken for granted and underestimated., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Over the past years medical centres specifically addressed in gender-based violence have developed protocols for the collections of evidence useful in the courtroom, including accurate documentation of physical and psychological states of the victim and collection of samples. Previous studies showed an association between documented physical trauma and conviction but unfortunately, few studies in the recent literature analysed the factors that influence the legal outcome and final judgement. The present study focused on the elements that appeared of significance in the legal outcome, including medico-legal evaluation, source of the crime report and circumstance of the assault., Methods: It was conducted a retrospective analysis of all the judgments issued by the Public Prosecutor's Office at a Court of a Metropolitan Italian city regarding sexual and domestic violence, from January 1st 2011 to 31st December 31st 2015. Examination regarded the demographic information of the victim and of the defendant, information on the crime, the circumstances of the aggression and medical information retrieved. Sentences were subsequently divided into two categories based on the legal outcome (conviction vs acquittal) and the different characteristics of the two sub-populations were compared to verify if there were variables significantly associated to the judge's final judgment., Results: Over the 5 years taken into consideration, there have been 1342 verdicts regarding crimes of sexual violence (374 cases) and regarding abuses against family members or cohabitants (875): other 93 cases regarded both sexual violence and abuse. 66.3% ended in conviction of the offender and 33.7% in acquittal of the accused. Cases of conviction were more frequent when they involved: use of a weapon by the assailant, as well as if the assailant had a criminal record and had a history of drug abuse or other addictions; duration of proceeding less 22 months and a civil party involved; presence of clinical documentation together with other deposition in addition to victim's deposition; also frequent episodes of violence and application of precautionary measures were associated to conviction., Conclusions: Many factors seem able to influence the judge's judgment, although clearly each case must be singularly evaluated. The mere presence of medical documentation, without the support of other sources of evidence, such as the victim's statement or further declarations, however, is almost always not definitive for the verdict. Despite so, in cases where there are multiple sources of evidence, clinical documentation can provide useful elements and can give clues on the consistency between the history told and injuries observed., Competing Interests: The authors have declared that no competing interests exist.

It is commonly accepted that crime scene recovery and recording are key moments of any judicial inspection in which investigators must decide on the correct strategies to put into place. Complex outdoor scenarios, presenting partially or entirely skeletonised remains, can benefit more than others by the intervention of environmental specialists (forensic anthropologists, archaeologists, entomologists and botanists). These experts are capable of singling out, correctly recording and recovering environmental evidence that can lead to a more comprehensive reconstruction of a given criminal episode. If human remains are discovered in an outdoor scenario, the on-site presence of a botanist will guarantee a correct approach to the identification, recording and recovery of any botanical evidence. If an on-site botanist is not available, the operators must be capable of both the botanical evaluation of a scene and the implementation of correct botanical sampling protocols.The following collection of unusual case histories that aim at underlining the efficacy of forensic botany will examine the determination of post mortem or the post depositional interval, evidence for a victim's post mortem transfer, evidence for the identification of a primary crime scene and evidence for the identification of a victim's dismemberment site. In another two cases, one, we will illustrate the important role that forensic botany played in the discrimination between botanical material used to voluntarily conceal a victim and vegetation that had grown naturally above a disposal site, whereas the other will highlight the protocols implemented for the identification of a murder weapon.

The correct interpretation of skin lesions is a crucial issue in forensic medicine. Many macroscopic and microscopic vital reaction markers and molecules have been studied in the past years in order to estimate the timing of injuries. However, literature lacks information regarding how to perform sampling to obtain a more reliable and accurate response. The present study investigates 25 bruises and analyses the different histological asset in different areas (central or marginal) and the different layers (epidermis, dermis, hypodermis and muscle) of the wound. All wounds were sampled in several areas and analysed under a 200× magnification optical microscope, simply counting the degree of haemorrhaging (i.e hematic infiltration) visible in each optical field (0 = 0%-5%; 1 = 6%-25%; 2 = 26%-50%; 3 = 51%-75%; 4 = 76%-100%). All the injuries presented some kind of infiltration in at least one area, nonetheless only 56% of the wounds were infiltrated in the entirety of the areas. Finally, in 28% the dermal layer did not show any trace of infiltration. Therefore, considering such results it seems that depth or area of sampling of a bruise may largely influence the evaluation of vitality and hence of age of a wound. Though this may seem intuitive, no studies have examined this issue in depth., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Violence perpetrated on male victims is a phenomenon that is currently underestimated by both national and international scientific communities, since males are historically (and stereotypically) considered the perpetrators rather than the victims of violence. As a consequence, the available literature lacks data which would allow a better understanding of this issue and its presenting features. We undertook a retrospective analysis of 231 medical files of male victims of violence over a five-year-period (2014-2018) at the Soccorso Violenza Sessuale e Domestica Centre in Milan, Italy. The sample included 112 victims of sexual violence and 119 victims of domestic violence, most of whom were younger than 18 years of age. The main aim of this study was to describe the presentations of male victims of violence in order to understand the phenomenon better, increase awareness of the issue and enable improved health-care management. The need to pay more attention to this vulnerable part of the population is mandatory in health-care services, and it includes: the provision of management guidelines, training to attending physicians and a supportive service to male survivors. Understanding which risk factors are related to male abuse can help with the development of programmes that identify, prevent and minimise violence - this being especially useful for primary-care clinicians. This is the first Italian study to deal with domestic and sexual violence involving male victims.

Introduction: Changing patterns of migration has required states and governments to respond to the specific medical and legal needs of asylum seekers. Based on medical assessments undertaken at the University Institute of Legal Medicine, the present study aims to describe the cases of asylum applicants who have suffered from physical violence, including torture, and the variables involved., Methods: Over a 10-year period, 225 survivors were examined by clinical forensic professionals from the University Institute of Legal Medicine., Results: 85% of asylum applicants came from Africa, 87% were male, and the most common age group was 26-40 years old. 46% of applicants fled their country for political reasons. Blunt force injuries were reported in 45% of cases, the trunk was the most affected area of the body (40%), and applicants presented with an average of two different mechanisms of lesions and an average of four lesions each., Discussion/conclusion: Assessment of physical violence on asylum seekers requires the cooperation of professionals with different skillsets and training.

Recent experimental data indicate a pathogenic role of complement activation in congestive heart failure (CHF). The aim of this study was to evaluate contact and complement systems activation in patients hospitalized for an acute episode of CHF. Forty-two of 80 consecutive patients admitted at our hospital with confirmed diagnosis of acute CHF were enrolled. They underwent blood sampling within 24 h from admission (T0) and at clinical stability (T1). Patients were stratified for ejection fraction (EF) based on echocardiographic test. We measured plasma levels of C3, C4, sC5b-9 and cleaved high molecular weight kininogen (contact activation marker). At T1, C3 levels increased significantly compared to T0 (97 ± 2 versus 104 ± 3% of total pooled plasma, P < 0·01). Classifying patients according to EF, only patients with preserved EF presented a significant increase of C3 from T0 to T1 (99 ± 3 versus 108 ± 4%, P = 0·03). When the sample was stratified according to clinical outcome, C3 (98 ± 3 versus 104 ± 4%, P = 0·03) and sC5b-9 levels (204 ± 10 versus 230 ± 11 ng/ml, P = 0·03) were increased in patients who had positive outcome after hospitalization. CHF patients with preserved EF and positive outcome after hospitalization showed higher levels of sC5b-9 in the T1 period compared with T0 (211 ± 14 versus 243 ± 14 ng/ml, P = 0·04). Our results suggest that the complement system reacts differently if CHF occurs with preserved or reduced EF. This finding is interesting if we consider the difference in epidemiology, pathogenesis and possible therapeutic approaches of these two clinical entities., (© 2017 British Society for Immunology.)

Background: Acquired angioedema due to C1-inhibitor deficiency (C1-INH-AAE) is a rare disease with no prevalence data or approved therapies., Objective: To report data on patients with C1-INH-AAE followed at Angioedema Center, Milan (from 1976 to 2015)., Methods: Diagnostic criteria included history of recurrent angioedema without wheals; decreased C1-INH antigen levels and/or functional activity of C1-INH and C4 antigen less than 50% of normal; late symptom onset (>40 years); no family history of angioedema and C1-INH deficiency., Results: In total, 77 patients (58% females; median age, 70 years) were diagnosed with C1-INH-AAE and 675 patients with hereditary angioedema due to C1-INH deficiency (C1-INH-HAE) (1 patient with C1-INH-AAE/8.8 patients with C1-INH-HAE). Median age at diagnosis was 64 years. Median time between symptom onset and diagnosis was 2 years. Sixteen patients (21%) died since diagnosis, including 1 because of laryngeal edema. Angioedema of the face was most common (N = 63 [82%]), followed by abdomen (N = 51 [66%]), peripheries (N = 50 [65%]), and oral mucosa and/or glottis (N = 42 [55%]). Forty-eight of 71 patients (68%) had autoantibodies to C1-INH. In total, 56 patients (70%) used on-demand treatment for angioedema including intravenous pdC1-INH 2000 U (Berinert, CSL Behring, Marburg, Germany) (N = 49) and/or subcutaneous icatibant 30 mg (Firazyr, Shire; Milano, Italy) (N = 27). Eventually, 8 of 49 patients receiving pdC1-INH became nonresponsive; all had autoantibodies. Thirty-four patients received long-term prophylaxis with tranexamic acid (effective in 29) and 20 with androgens (effective in 8)., Conclusions: The incidence of C1-INH-AAE was 1 for every 8.8 patients with C1-INH-HAE. Thirty percent of the deaths were related to the disease. Treatments approved for C1-INH-HAE are effective in C1-INH-AAE, although with minimal differences., (Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Diabetes is a complex metabolic disease that exposes patients to the deleterious effects of hyperglycemia on various organs. Achievement of normoglycemia with exogenous insulin treatment requires the use of high doses of hormone, which increases the risk of life-threatening hypoglycemic episodes. We developed a gene therapy approach to control diabetic hyperglycemia based on co-expression of the insulin and glucokinase genes in skeletal muscle. Previous studies proved the feasibility of gene delivery to large diabetic animals with adeno-associated viral (AAV) vectors. Here, we report the long-term (∼8 years) follow-up after a single administration of therapeutic vectors to diabetic dogs. Successful, multi-year control of glycemia was achieved without the need of supplementation with exogenous insulin. Metabolic correction was demonstrated through normalization of serum levels of fructosamine, triglycerides, and cholesterol and remarkable improvement in the response to an oral glucose challenge. The persistence of vector genomes and therapeutic transgene expression years after vector delivery was documented in multiple samples from treated muscles, which showed normal morphology. Thus, this study demonstrates the long-term efficacy and safety of insulin and glucokinase gene transfer in large animals and especially the ability of the system to respond to the changes in metabolic needs as animals grow older.

The European Search Catalogue for Plant Genetic Resources, EURISCO, provides information about 1.8 million crop plant accessions preserved by almost 400 institutes in Europe and beyond. EURISCO is being maintained on behalf of the European Cooperative Programme for Plant Genetic Resources. It is based on a network of National Inventories of 43 member countries and represents an important effort for the preservation of world's agrobiological diversity by providing information about the large genetic diversity kept by the collaborating collections. Moreover, EURISCO also assists its member countries in fulfilling legal obligations and commitments, e.g. with respect to the International Treaty on Plant Genetic Resources, the Second Global Plan of Action for Plant Genetic Resources for Food and Agriculture of the United Nation's Food and Agriculture Organization, or the Convention on Biological Diversity. EURISCO is accessible at http://eurisco.ecpgr.org., (© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Mucopolysaccharidosis type II (MPSII) is an X-linked lysosomal storage disease characterized by severe neurologic and somatic disease caused by deficiency of iduronate-2-sulfatase (IDS), an enzyme that catabolizes the glycosaminoglycans heparan and dermatan sulphate. Intravenous enzyme replacement therapy (ERT) currently constitutes the only approved therapeutic option for MPSII. However, the inability of recombinant IDS to efficiently cross the blood-brain barrier (BBB) limits ERT efficacy in treating neurological symptoms. Here, we report a gene therapy approach for MPSII through direct delivery of vectors to the CNS. Through a minimally invasive procedure, we administered adeno-associated virus vectors encoding IDS (AAV9- Ids ) to the cerebrospinal fluid of MPSII mice with already established disease. Treated mice showed a significant increase in IDS activity throughout the encephalon, with full resolution of lysosomal storage lesions, reversal of lysosomal dysfunction, normalization of brain transcriptomic signature, and disappearance of neuroinflammation. Moreover, our vector also transduced the liver, providing a peripheral source of therapeutic protein that corrected storage pathology in visceral organs, with evidence of cross-correction of nontransduced organs by circulating enzyme. Importantly, AAV9- Ids -treated MPSII mice showed normalization of behavioral deficits and considerably prolonged survival. These results provide a strong proof of concept for the clinical translation of our approach for the treatment of Hunter syndrome patients with cognitive impairment.

Gene therapy is an attractive tool for the treatment of monogenic disorders, in particular for lysosomal storage diseases (LSD) caused by deficiencies in secretable lysosomal enzymes in which neither full restoration of normal enzymatic activity nor transduction of all affected cells are necessary. However, some LSD such as Mucopolysaccharidosis Type IIIB (MPSIIIB) are challenging because the disease's main target organ is the brain and enzymes do not efficiently cross the blood-brain barrier even if present at very high concentration in circulation. To overcome these limitations, we delivered AAV9 vectors encoding for α-N-acetylglucosaminidase (NAGLU) to the Cerebrospinal Fluid (CSF) of MPSIIIB mice with the disease already detectable at biochemical, histological and functional level. Restoration of enzymatic activity in Central Nervous System (CNS) resulted in normalization of glycosaminoglycan content and lysosomal physiology, resolved neuroinflammation and restored the pattern of gene expression in brain similar to that of healthy animals. Additionally, transduction of the liver due to passage of vectors to the circulation led to whole-body disease correction. Treated animals also showed reversal of behavioural deficits and extended lifespan. Importantly, when the levels of enzymatic activity were monitored in the CSF of dogs following administration of canine NAGLU-coding vectors to animals that were either naïve or had pre-existing immunity against AAV9, similar levels of activity were achieved, suggesting that CNS efficacy would not be compromised in patients seropositive for AAV9. Our studies provide a strong rationale for the clinical development of this novel therapeutic approach as the treatment for MPSIIIB., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objectives: To evaluate whether reduced activity of the anti-inflammatory HSP70 pathway correlates with nonalcoholic fatty liver disease (NAFLD) progression and with markers of oxidative stress because obesity activates inflammatory JNKs, whereas HSP70 exerts the opposite effect., Methods: Adult obese patients (N = 95) undergoing bariatric surgery were divided into steatosis (ST), steatohepatitis (SH), and fibrosis (SH+F) groups. The levels of HSP70, its major transcription factor, HSF1, and JNKs were assessed by immunoblotting hepatic and visceral adipose tissue; data were confirmed by immunohistochemistry. Plasma biochemistry (lipids, HbA1c , HOMA, hepatic enzymes, and redox markers) was also evaluated., Results: In both liver and adipose tissue, decreased HSP70 levels, paralleled by similar reductions in HSF1 and reduced plasma antioxidant enzyme activities, correlated with insulin resistance and with NAFLD progression (expression levels were as follows: ST > SH > SH + F). The immunohistochemistry results suggested Kupffer cells as a site of HSP70 inhibition. Conversely, JNK1 content and phosphorylation increased., Conclusions: Decreased HSF1 levels in the liver and fat of obese patients correlated with impairment of HSP70 in an NAFLD stage-dependent manner. This impairment may affect HSP70-dependent anti-inflammation, with consequent oxidative stress and insulin resistance in advanced stages of NAFLD. Possible causal effects of fat cell senescence are discussed., (© 2014 The Obesity Society.)

Background: The inherited deficiency of C1-inhibitor (C1-INH), which can be quantitative (type I) or qualitative (type II), is characterized by recurrent attacks of oedema, and it is known as hereditary angioedema due to C1-INH deficiency (HAE-C1-INH). The frequency of symptoms varies widely among patients and in the same patient during life., Objective: To identify laboratory markers of disease severity in HAE-C1-INH patients., Methods: We studied 162 patients with differently severe HAE-C1-INH during remission, 31 HAE-C1-INH patients during attacks, and 81 normal controls, evaluating complement parameters, spontaneous plasma kallikrein activity, the capacity of plasma to inhibit exogenous kallikrein activity, and cleavage of high-molecular-weight kininogen (HK). Sixty-five HAE-C1-INH patients were screened for mutations in the C1-INH gene., Results: As expected, plasma C1-INH levels and activity and C4 levels were low in the HAE-C1-INH patients. Spontaneous plasma kallikrein activity in patients in remission was higher than in controls (P = 0.001) and increased during acute attacks (P = 0.01), whereas the capacity of inhibiting kallikrein activity was lower in patients in remission than in controls (P = 0.001) and further reduced during attacks (P = 0.001). HAE-C1-INH patients in remission had higher levels of cleaved HK than controls (P = 0.001), and these further increased during acute attacks (P = 0.001). Cleaved HK levels were higher in highly symptomatic HAE-C1-INH patients than in those with less frequent attacks (P = 0.001). Thirty-five different mutations in the C1-INH gene were equally distributed in patients with different attack frequencies., Conclusions: Measuring plasma levels of cleaved HK may be a sensitive mean of assessing disease severity in HAE-C1-INH patients., (© 2014 John Wiley & Sons Ltd.)

Local varieties of leafy kales (Brassica oleracea L.) are grown in home gardens in Calabria and Sicily for self-consumption, in the same area where the wild relative Brassica rupestris Raf. also grows. With the use of AFLP markers, comparisons were made of the genetic diversity and population structure of ten wild and 22 cultivated populations, as well as of a hybrid population and of four commercial cultivars of different B. oleracea crops. The level of genetic diversity was higher in leafy kales than in wild populations and this diversity was mainly distributed within populations. Wild populations remained distinct from cultivated material. Additionally, most wild populations were distinctively isolated from each other. On the other hand, it was not possible to molecularly distinguish even geographically distant leafy kale populations from each other or from different B. oleracea crops. It was possible to detect inter-crossing between leafy kales and B. rupestris. Findings from this study illustrate the existing level of genetic diversity in the B. oleracea gene pool. Individual populations (either wild or leafy kales) with higher levels of genetic diversity have been identified and suggestions are given for an informed conservation strategy. Domestication hypotheses are also discussed., (© 2015 The Authors.)

Pancreatic gene transfer could be useful to treat several diseases, such as diabetes mellitus, cystic fibrosis, chronic pancreatitis, or pancreatic cancer. Helper-dependent adenoviral vectors (HDAds) are promising tools for gene therapy because of their large cloning capacity, high levels of transgene expression, and long-term persistence in immunocompetent animals. Nevertheless, the ability of HDAds to transduce the pancreas in vivo has not been investigated yet. Here, we have generated HDAds carrying pancreas-specific expression cassettes, that is, driven either by the elastase or insulin promoter, using a novel and convenient plasmid family and homologous recombination in bacteria. These HDAds were delivered to the pancreas of immunocompetent mice via intrapancreatic duct injection. HDAds, encoding a CMV-GFP reporter cassette, were able to transduce acinar and islet cells, but transgene expression was lost 15 days postinjection in correlation with severe lymphocytic infiltration. When HDAds encoding GFP under the control of the specific elastase promoter were used, expression was detected in acinar cells, but similarly, the expression almost disappeared 30 days postinjection and lymphocytic infiltration was also observed. In contrast, long-term transgene expression (>8 months) was achieved with HDAds carrying the insulin promoter and the secretable alkaline phosphatase as the reporter gene. Notably, transduction of the liver, the preferred target for adenovirus, was minimal by this route of delivery. These data indicate that HDAds could be used for pancreatic gene therapy but that selection of the expression cassette is of critical importance to achieve long-term expression of the transgene in this tissue.

For most lysosomal storage diseases (LSDs) affecting the CNS, there is currently no cure. The BBB, which limits the bioavailability of drugs administered systemically, and the short half-life of lysosomal enzymes, hamper the development of effective therapies. Mucopolysaccharidosis type IIIA (MPS IIIA) is an autosomic recessive LSD caused by a deficiency in sulfamidase, a sulfatase involved in the stepwise degradation of glycosaminoglycan (GAG) heparan sulfate. Here, we demonstrate that intracerebrospinal fluid (intra-CSF) administration of serotype 9 adenoassociated viral vectors (AAV9s) encoding sulfamidase corrects both CNS and somatic pathology in MPS IIIA mice. Following vector administration, enzymatic activity increased throughout the brain and in serum, leading to whole body correction of GAG accumulation and lysosomal pathology, normalization of behavioral deficits, and prolonged survival. To test this strategy in a larger animal, we treated beagle dogs using intracisternal or intracerebroventricular delivery. Administration of sulfamidase-encoding AAV9 resulted in transgenic expression throughout the CNS and liver and increased sulfamidase activity in CSF. High-titer serum antibodies against AAV9 only partially blocked CSF-mediated gene transfer to the brains of dogs. Consistently, anti-AAV antibody titers were lower in CSF than in serum collected from healthy and MPS IIIA-affected children. These results support the clinical translation of this approach for the treatment of MPS IIIA and other LSDs with CNS involvement.

Mucopolysaccharidosis type IIIA (MPSIIIA) is a rare lysosomal storage disorder caused by mutations in the sulfamidase gene. Accumulation of glycosaminoglycan (GAG) inside the lysosomes is associated with severe neurodegeneration as well as peripheral organ pathological changes leading to death of affected individuals during adolescence. There is no cure for MPSIIIA. Due to the limitation of the blood-brain barrier, enzyme replacement therapy and gene therapy strategies attempted thus far have not achieved whole-body correction of the disease. After the systemic administration of an adeno-associated virus 9 (AAV9) vector encoding for sulfamidase under the control of a ubiquitous promoter, we were able to obtain widespread expression of the therapeutic transgene in brain and in peripheral organs, and sulfamidase activity in serum of both male and female MPSIIIA mice. This was accompanied by the normalization of GAG storage levels in most peripheral organs. In brain, decrease in GAG tissue content following AAV9 gene transfer of sulfamidase was associated with the resolution of neuroinflammation. Finally, correction of disease phenotype resulted in a remarkable prolongation of survival of both male and female AAV-treated MPSIIIA mice. This proof-of-concept study will be relevant to the future development of therapies for MPSIIIA.

The authors translated from latin to italian the living will of the priest Dateus from Milan, dated 787 a. C., found and published by Ludovico Antonio Muratori in his Antiquitates Italicae Medii Aevi (1740). In the document is specified the request of converting his house into a founding home. Many historians consider this to be the first example of a founding home in the western world. The AA., after review of the history and the literature, noted the lack of any documented activity of the institution, as well as, the unusual model proposed for that time. Therefore, it is the AA. belief that the original document is actually postdated to the 1550 a. C.

Background: To demonstrate that bariatric procedures can be done with natural orifice visualization (NOTES) at 2 institutions (Nucleo Universitario de Estudos de Notes Centro de Cirurgia Experimental Vila do Conde-Junqueira, Vila do Conde, Portugal and Pontificia Universidade Catolica do Rio Grande do Sul, Porto Alegre, RS, Brasil). NOTES is a new surgical approach that is being developed. It consists of the use of a minimally invasive technique in which the surgical procedure is performed through natural orifices, thereby circumventing incisions through the skin., Methods: We performed vertical gastrectomy or laparoscopic sleeve gastrectomy in a porcine model using vaginal route visualization., Results: A laparoscopic vertical sleeve gastrectomy with NOTES visualization in a porcine model was performed with safety., Conclusion: Bariatric procedures can be done with NOTES with results as good as those using laparoscopic techniques.

Background: Natural orifice translumenal endoscopic surgery (NOTES) is an emerging concept in the recent literature that could lead to potential benefits in clinical applications. Restricted to animal experiments, however, human procedures have not yet been published. Because of the technical and ethical challenges involved in perforation and closure of a healthy organ-as is also seen in operating via the transgastric route-and because of the lack of understanding of the physiopathology and infection risk with these approaches, they have not been applied in the clinical setting. Thus the present study, based on previous animal experiments, describes preliminary clinical application in four cases of transvaginal NOTES cholecystectomy, and discusses safety, feasibility, and potential benefits of the method., Methods: Preliminary acute and survival animal experiments developed by the NOTES Research Group at our institution solved such technical problems for transvaginal NOTES as spatial orientation, insufflation, and instrumentation, making possible the introduction of NOTES as a clinical application. The trials were approved by ethics committee of our institution, and informed consent was obtained from all patients. Since 13 March 2007, four female patients with elective surgical indication for cholecystectomy have undergone transvaginal NOTES cholecystectomy. All intraoperative and postoperative parameters were documented. Vaginal access was achieved under direct vision with conventional instruments, and a 2-channel colonoscope was inserted into the abdominal cavity. After endoscopic insufflation to achieve pneumoperitoneum with CO(2), instruments were inserted through and alongside a colonoscope, allowing successful NOTES cholecystectomy in all patients, with vaginal extraction of the gallbladder. The vaginal wound was closed by direct vision using conventional instruments., Results: The procedure was successful in all patients, with operative time of 45-115 min. Patients experienced low need for postoperative analgesia. Free oral intake was permitted 2 h after the procedure. There were no postoperative complications, and patients were discharged, according to the study protocol, 48 h after the procedure., Conclusions: Preliminary results showed the feasibility and safety of the transvaginal NOTES method in this small initial study population. The technique, developed in our institution, and not transgastric NOTES, may be the preferred approach to serve as the basis for clinical studies.

Background & Aims: Serum cryoglobulins (CGs) are present in patients with chronic hepatitis C virus (HCV) infection, but their long-term clinical importance has not been established. We assessed the development rates, morbidity, and influence on the evolutionary course of hepatitis C of CG., Methods: A cohort of 343 HCV-RNA seropositive outpatients (173 men; age, 58 y; 82 with cirrhosis; 61 treated with interferon) with persistently increased aminotransferase levels and histologically defined liver disease was investigated. Patients initially were investigated for the presence, amount, and type of CG and prospectively followed up with clinical and laboratory examinations every 6 months., Results: At enrollment, CGs were found in 163 (47%) patients at a mean level of 173 +/- 142 mg/L; 80% were type III, and associated to female sex (61% vs 40%, P = .0002) and cirrhosis (29% vs 19%, P = .04). Over the course of 17-130 months (median, 116 mo), de novo CG developed in 25 patients (2.3% per year), including 5 with cryoglobulinemic syndrome (.3% per year). The 10-year rates of progression to cirrhosis and of liver and extrahepatic complications were similar in CG (+) and CG (-) patients (32% vs 34%; 23% vs 16%; 5% vs 3%). The 10-year survival rates were lower for cirrhotic than for noncirrhotic patients (57% vs 91%, P < .00001), independently of CGs., Conclusions: CGs are common in patients with chronic HCV infection, mainly are type III, and do not influence the clinical course of hepatitis C during the first decades, except for the few rare cases of cryoglobulinemic syndrome.

Background: Angioedema without major urticarial flares (hives) is poorly understood. Its causes are diverse, and little is known about its pathogenic mechanisms. We report on our 11 years of experience with this condition and propose a classification of patients affected by angioedema unaccompanied by urticaria., Methods: From 1993 through 2003 at our outpatient clinic, 929 consecutive patients were examined for recurrent angioedema unaccompanied by urticaria. A detailed history was taken to identify known causes of angioedema, with special attention to external allergenic agents. Each patient underwent a complete physical examination, routine laboratory tests (blood cell count, protein electrophoresis, erythrosedimentation rate, examination of stool for ova and parasites, pharyngeal and urine cultures, sinus and dental radiography, and measurements of antitissue autoantibodies and rheumatoid factor in plasma), and complement parameters (C1 inhibitor, C4 and C1q). Further testing was done when pertinent, based on clinical findings. When all results were negative, response to H1-antihistamine was considered., Results: Angioedema could not be classified in 153 patients who were lost to follow-up (16.4%). Among the 776 cases with adequate data, these types of angioedema were identified: 124 (16%) related to external agents such as a drug, insect bite or foodstuff; 85 (11%) related to treatment with angiotensin-converting enzyme inhibitor; 55 (7%) associated with an autoimmune disease or infection; and 197 (25%) caused by C1 inhibitor deficiency. In the other 315 cases (41%), the etiology was undiscovered: 254 cases (33% of the study sample) were idiopathic histaminergic; 40 (5%) were idiopathic nonhistaminergic; and 21 (3%) had other causes of peripheral or generalyzed edema., Interpretation: Our data indicated that angioedema without urticaria could be classified according to specific clinical and pathogenic characteristics, and we have suggested a work-up for patients experiencing this condition.