PDX proteins from Arabidopsis thaliana as novel substrates of cathepsin B: implications for vitamin B6 biosynthesis regulation.

Vitamin B6 is a critical molecule for metabolism, development, and stress sensitivity in plants. It is a cofactor for numerous biochemical reactions, can serve as an antioxidant, and has the potential to increase tolerance against both biotic and abiotic stressors. Due to the importance of vitamin B6, its biosynthesis is likely tightly regulated. Plants can synthesize vitamin B6 de novo via the concerted activity of Pyridoxine Biosynthesis Protein 1 (PDX1) and PDX2. Previously, PDX proteins have been identified as targets for ubiquitination, indicating they could be marked for degradation by two highly conserved pathways: the Ubiquitin Proteasome Pathway (UPP) and the autophagy pathway. Initial experiments show that PDXs are in fact degraded, but surprisingly, in a ubiquitin-independent manner. Inhibitor studies pointed toward cathepsin B, a conserved lysosomal cysteine protease, which is implicated in both programed cell death and autophagy in humans and plants. In plants, cathepsin Bs are poorly described, and no confirmed substrates have been identified. Here, we present PDX proteins from Arabidopsis thaliana as interactors and substrates of a plant Cathepsin B. These findings not only describe a novel cathepsin B substrate in plants, but also provide new insights into how plants regulate de novo biosynthesis of vitamin B6.

Plant cathepsin B, a versatile protease.

Plant proteases are essential enzymes that play key roles during crucial phases of plant life. Some proteases are mainly involved in general protein turnover and recycle amino acids for protein synthesis. Other proteases are involved in cell signalling, cleave specific substrates and are key players during important genetically controlled molecular processes. Cathepsin B is a cysteine protease that can do both because of its exopeptidase and endopeptidase activities. Animal cathepsin B has been investigated for many years, and much is known about its mode of action and substrate preferences, but much remains to be discovered about this potent protease in plants. Cathepsin B is involved in plant development, germination, senescence, microspore embryogenesis, pathogen defence and responses to abiotic stress, including programmed cell death. This review discusses the structural features, the activity of the enzyme and the differences between the plant and animal forms. We discuss its maturation and subcellular localisation and provide a detailed overview of the involvement of cathepsin B in important plant life processes. A greater understanding of the cell signalling processes involving cathepsin B is needed for applied discoveries in plant biotechnology.

Roadmap for the next decade of plant programmed cell death research.

Programmed cell death (PCD) is fundamentally important for plant development, abiotic stress responses and immunity, but our understanding of its regulation remains fragmented. Building a stronger research community is required to accelerate progress in this area through knowledge exchange and constructive debate. In this Viewpoint, we aim to initiate a collective effort to integrate data across a diverse set of experimental models to facilitate characterisation of the fundamental mechanisms underlying plant PCD and ultimately aid the development of a new plant cell death classification system in the future. We also put forward our vision for the next decade of plant PCD research stemming from discussions held during the 31st New Phytologist workshop, 'The Life and Death Decisions of Plant Cells' that took place at University College Dublin in Ireland (14-15 June 2023). We convey the key areas of significant progress and possible future research directions identified, including resolving the spatiotemporal control of cell death, isolation of its molecular and genetic regulators, and harnessing technical advances for studying PCD events in plants. Further, we review the breadth of potential impacts of plant PCD research and highlight the promising new applications of findings from this dynamically evolving field.

Rhythmic mid-Temporal Theta of Drowsiness Activated by Hyperventilation- Uncommon Trigger of a Rare Benign EEG Variant in Pediatrics. An Educational Review.

Distinguishing abnormal electroencephalogram (EEG) waveforms from benign variants is critical for accurate interpretation of EEG. Hyperventilation (HV) is one of the basic procedures during EEG to enable activation of epileptiform activity. Rarely, HV can activate benign EEG rhythms. Herein, we illustrate two pediatric cases with bursts of rhythmic mid-temporal theta of drowsiness (RMTD), activated by hyperventilation. Continued awareness of this EEG phenomenology and its variations in pediatrics is important in avoiding misdiagnosis of epilepsy.

The challenge of setting restoration targets for macroalgal forests under climate changes.

The process of site selection and spatial planning has received scarce attention in the scientific literature dealing with marine restoration, suggesting the need to better address how spatial planning tools could guide restoration interventions. In this study, for the first time, the consequences of adopting different restoration targets and criteria on spatial restoration prioritization have been assessed at a regional scale, including the consideration of climate changes. We applied the decision-support tool Marxan, widely used in systematic conservation planning on Mediterranean macroalgal forests. The loss of this habitat has been largely documented, with limited evidences of natural recovery. Spatial priorities were identified under six planning scenarios, considering three main restoration targets to reflect the objectives of the EU Biodiversity Strategy for 2030. Results show that the number of suitable sites for restoration is very limited at basin scale, and targets are only achieved when the recovery of 10% of regressing and extinct macroalgal forests is planned. Increasing targets translates into including unsuitable areas for restoration in Marxan solutions, amplifying the risk of ineffective interventions. Our analysis supports macroalgal forests restoration and provides guiding principles and criteria to strengthen the effectiveness of restoration actions across habitats. The constraints in finding suitable areas for restoration are discussed, and recommendations to guide planning to support future restoration interventions are also included.

Potential and limitations of digital ethnographic research: A case study on a web community.

Introduction: This work aims at transposing ethnographic research into digital contexts to probe its potential and limitations in a specific field of study: that of sexuality, particularly suited to ethnographic exploration. We chose as our case study a web community of Italian asexual people. As we shall see, this allowed us to simultaneously explore both the various techniques called into play in digital ethnography and the digital as a specific sphere within which sexuality takes on a very peculiar meaning. Digital sociality is paramount for the definition of imaginaries, meanings, and practices that could not be explored elsewhere. This is due to the implicit characteristics of the population studied, which does not find corresponding physical spaces of aggregation. Methods: The paper will present the research design using this specific case study to address some of the typical dilemmas that researchers face when following the digital ethnographic approach and will explore the research results as an example of the kind of analysis available with the information and data collected through this method. Results and discussion: The conclusions will attempt to briefly outline the shortfalls and advantages of this method, considering its application to this specific field of study.

Adjuvant chemoradiotherapy (gemcitabine-based) in pancreatic adenocarcinoma: the Pisa University experience.

INTRODUCTION: The role of adjuvant chemoradiotherapy in patients with pancreatic adenocarcinoma (PA) is controversial. In this study we aimed to assess the feasibility, disease-free survival (DFS) and overall survival (OS) of adjuvant chemoradiotherapy (gemcitabine based) in patients with resected PA and their correlation with prognostic factors. METHODS: 122 resected patients (stage ≥IIa) treated between February 1999 and December 2013 were analyzed. Two cycles of gemcitabine (1,000 mg/m2 on days 1, 8 and 15 every 28 days) were administered before concomitant radiotherapy (45 Gy/25 fractions) and chemotherapy (gemcitabine 300 mg/m2 weekly). RESULTS: Median follow-up was 22.7 months (range 4-109). Gastrointestinal toxicity (G3), neutropenia (G3-G4) and cardiac toxicity (G2-G3) were observed in 2.4%, 10.6% and 1.6% of patients, respectively. OS at 12, 24 and 60 months was 79%, 55% and 31%, respectively (median 25 months). Two-year OS in patients with postoperative Karnofsky performance status (KPS) ≤70 and ≥80 was 37.1% and 62.3%, respectively (p<0.0001). OS was better in the group of patients with a postoperative CA 19-9 level ≤100 U/mL (p = 0.014). Median DFS was 17 months. CONCLUSIONS: The combination of concomitant gemcitabine and radiotherapy in patients with radically resected PA was well tolerated and associated with a low incidence of local recurrences. Five-year OS was significantly influenced by postoperative KPS and CA 19-9 values.

Dose tracking assessment for image-guided radiotherapy of the prostate bed and the impact on clinical workflow.

BACKGROUND: The cumulative dose was compared with the planned dose among fourteen patients undergoing image-guided, intensity-modulated radiotherapy of the prostate bed. Moreover, we investigated the feasibility of adding dose tracking to the routine workflow for radiotherapy. METHODS: Daily cone beam computed tomography was conducted for image-guided radiotherapy, and weekly cumulative delivered doses were calculated for dose tracking. Deformable image registration was applied to map weekly dose distributions to the original treatment plan and to create a cumulative dose distribution. The dose-volume histogram (DVH) cut-off points for the rectum and bladder and the planning target volume (PTV), were used to compare the planned and cumulative delivered doses. The additional time required by the departmental staff to complete these duties was recorded. RESULTS: The PTV coverage of the delivered treatment did not satisfy the expected goal for three patients (V98% >98%). In another three patients, the DVH cut-off point for the bladder was higher than the limits, while for the rectum, treatment was as expected in all cases (two patients failed both their bladder constraints and the PTV coverage). Overall, four patients did not satisfy one or more criteria at the end of their treatment. CONCLUSIONS: A well-defined strategy for dose tracking assessment is feasible, would have minimal impact on the workload of a radiotherapy department, and may offer objective information to support radiation oncologists in making decisions about adaptive procedures.

Chronological Evolution of Magnetic Resonance Imaging Findings in Children With Febrile Infection-Related Epilepsy Syndrome.

OBJECTIVE: To describe and analyze the chronological evolution of the radiological findings in seven children with febrile infection-related epilepsy syndrome. METHODS: This is a retrospective study describing the radiological findings and evolution in seven children with febrile infection-related epilepsy syndrome who presented from 2009 to 2013. The children all fit the defined clinical criteria for febrile infection-related epilepsy syndrome; all had a history of normal psychomotor development who presented with acute-onset catastrophic partial status epilepticus associated with a febrile illness or unspecific infectious process. The children were identified from the author's weekly review of the pediatric inpatient service, and then the data were collected and analyzed retrospectively. RESULTS: Six males and one female ranging from 3 months to 9 years of age presented with status epilepticus preceded by a febrile illness. Extensive investigations for infectious, autoimmune, and metabolic etiologies were unremarkable. Multiple antiepileptic medications were attempted, including drug-induced coma in all of them, with poor response. Immunotherapy with intravenous steroids or intravenous immunoglobulin (three patients had both) was tried in six of seven patients with a poor response. Ketogenic diet was initiated in four of seven patients with limited response. Serial magnetic resonance imaging studies, done from the initial presentation through 18 months of follow-up, showed evolution from normal imaging to severe cerebral atrophy. Progressive cytotoxic edema involving mostly bilateral hippocampi and temporal lobes was appreciated in one to three weeks. At one month from seizure onset, mild to moderate cerebral atrophy and hippocampal sclerosis was appreciated that continued to progress over the next year. After six to twelve months, most of the patients showed moderate to severe cerebral atrophy and by one year, cerebellar atrophy was also appreciated. CONCLUSION: Febrile infection-related epilepsy syndrome is a devastating epilepsy syndrome of childhood without a diagnostic biologic marker. The magnetic resonance imaging findings appear to be progressive and typical. Thus, combined with the clinical course, imaging findings can help to confirm the diagnosis (until a biologic marker is found). This hopefully will allow multicentered treatment protocols in the future.

Dosimetric impact of different multileaf collimators on prostate intensity modulated treatment planning.

AIM: The main purpose of this study is to perform a dosimetric comparison on target volumes and organs at risks (OARs) between prostate intensity modulated treatment plans (IMRT) optimized with different multileaf collimators (MLCs). BACKGROUND: The use of MLCs with a small leaf width in the IMRT optimization may improve conformity around the tumor target whilst reducing the dose to normal tissues. MATERIALS AND METHODS: Two linacs mounting MLCs with 5 and 10 mm leaf-width, respectively, implemented in Pinnacle(3) treatment planning system were used for this work. Nineteen patients with prostate carcinoma undergoing a radiotherapy treatment were enrolled. Treatment planning with different setup arrangements (7 and 5 beams) were performed for each patient and each machine. Dose volume histograms (DVHs) cut-off points were used in the treatment planning comparison. RESULTS: Comparable planning target volume (PTV) coverage was obtained with 7- and 5-beam configuration (both with 5 and 10 mm MLC leaf-width). The comparison of bladder and rectum DVH cut-off points for the 5-beam arrangement shows that 52.6% of the plans optimized with a larger leaf-width did not satisfy at least one of the OARs' constraints. This percentage is reduced to 10.5% for the smaller leaf-width. If a 7-beam arrangement is used the value of 52.6% decreases to 21.1% while the value of 10.5% remains unchanged. CONCLUSION: MLCs collimators with different widths and number of leaves lead to a comparable prostate treatment planning if a proper adjustment is made of the number of gantry angles.

Bupropion Overdose Presenting as Status Epilepticus in an Infant.

BACKGROUND: Bupropion is a monocyclic antidepressant in the aminoketone class, structurally related to amphetamines. The Food and Drug Administration withdrew this product from the market in 1986 after seizures were reported in bulimic patients. It was later reintroduced in 1989 when the incidence of seizures was shown to be dose-related in the immediate release preparation. Massive bupropion ingestion has been associated with status epilepticus and cardiogenic shock in adults. Seizures have been reported in children, but not status epilepticus. This report highlights a patient who presented with status epilepticus and developed cardiopulmonary arrest after bupropion ingestion. False-positive amphetamine diagnosis from urine drug screen on presentation was reported. METHOD: We review the presentation, clinical course, diagnostic studies, and outcome of this patient. We then review the literature regarding bupropion overdose in children. RESULT: Symptoms of bupropion toxicity and risk for seizures are dose-dependent and fatalities have been reported. Our patient developed status epilepticus and cardiopulmonary arrest and then progressed to have a hypoxic ischemic encephalopathy and refractory symptomatic partial seizures. CONCLUSION: Our report highlights the need to keep this medication away from children in order to prevent accidental overdose.

No benefit of adjuvant Fluorouracil Leucovorin chemotherapy after neoadjuvant chemoradiotherapy in locally advanced cancer of the rectum (LARC): Long term results of a randomized trial (I-CNR-RT).

BACKGROUND AND PURPOSE: To evaluate the effect of adjuvant chemotherapy (ACT) in locally advanced rectal cancer (LARC) after neoadjuvant chemoradiation (NACT-RT). The study was funded by the Italian National Research Council (CNR). METHODS: From September 1992 to January 2001, 655 patients with LARC (clinically T3-4, any N) treated with NACT-RT and surgery, were randomized in two arms: follow-up (Arm A) or 6 cycles of ACT with 5 fluorouracil (5FU)-Folinic Acid (Arm B). NACT-RT consisted of 45Gy/28/ff concurrent with 5FU (350mg/sqm) and Folinic Acid (20mg/sqm) on days 1-5 and 29-33; surgery was performed after 4-6weeks. Median follow up was 63·7months. Primary end point was overall survival (OS). RESULTS: 634/655 patients were evaluable (Arm A 310, Arm B 324); 92·5% of Arm A and 91% of Arm B patients received the preoperative treatment as in the protocol; 294 patients of Arm A (94·8%) and 296 of Arm B (91·3%) underwent a radical resection; complete pathologic response and overall downstaging rates did not show any significant difference in the two arms. 83/297 (28%) patients in Arm B, never started ACT. Five year OS and DFS did not show any significant difference in the two treatment arms. Distant metastases occurred in 62 patients (21%) in Arm A and in 58 (19·6%) in Arm B. CONCLUSIONS: In patients with LARC treated with NACT-RT, the addition of ACT did not improve 5year OS and DFS and had no impact on the distant metastasis rate.

Adenosina induce arritmias ventriculares en corazones con miocardiopatía chagásica crónica / Adenosine induces ventricular arrythmias in hearts with chronic chagas cardiomyopathy

La adenosina liberada durante la isquemia y la hipoxia es capaz de inducir arritmias ventriculares, fenómenos también observados en la enfermedad de Chagas. En este trabajo caracterizamos farmacológicamente las propiedades arritmogénicas de la adenosina en ratas Sprague Dawley sanas (n = 14) y con miocarditis chagásica crónica (n = 14), que fueron sometidas a estudios electrocardiográficos y farmacológicos en preparaciones de corazón aislado tipo Langendorff. La adenosina incrementó las arritmias ventriculares en ambos grupos en forma dosis-dependiente, y en un 50% de las ratas chagásicas indujo fibrilación ventricular respecto a un 7,14% de las ratas sanas (p < 0,05). La fibrilación fue prevenida por antagonistas A1 (DPCPX) y A2a (8-CSC). Las arritmias se asociaron a un QT prolongado, a depolarizaciones tempranas, a fenómenos R sobre T y torsade de pointes. En conclusión, la adenosina es un fármaco proarrítmico capaz de inducir fibrilación ventricular en ratas chagásicas (AU)

Adenosine released during ischemia and hypoxia can induce ventricular arrhythmias. This phenomenon is also observed in Chagas disease. This study involved pharmacologic analysis of the arrhythmogenic properties of adenosine in healthy Sprague-Dawley rats (n=14) and in rats with chronic Chagas cardiomyopathy (n=14). Electrocardiographic and pharmacologic studies were performed on isolated hearts prepared using the Langendorff method. Adenosine increased ventricular arrhythmias in both groups of animals in a dose-dependent manner: 50% of chagasic rats developed ventricular fibrillation compared with 7.14% of healthy rats (P < .05). Fibrillation was prevented by A1 (i.e., DPCPX) and A2a (i.e., 8-CSC) receptor antagonists. Arrhythmia was associated with a prolonged QT interval, early depolarization, and the R-on-T and torsade de pointes phenomena. In conclusion, adenosine is a proarrythmic drug that is able to induce ventricular fibrillation in chagasic rat hearts (AU)

[Adenosine induces ventricular arrythmias in hearts with chronic chagas cardiomyopathy]. / Adenosina induce arritmias ventriculares en corazones con miocardiopatía chagásica crónica.

Adenosine released during ischemia and hypoxia can induce ventricular arrhythmias. This phenomenon is also observed in Chagas disease. This study involved pharmacologic analysis of the arrhythmogenic properties of adenosine in healthy Sprague-Dawley rats (n=14) and in rats with chronic Chagas cardiomyopathy (n=14). Electrocardiographic and pharmacologic studies were performed on isolated hearts prepared using the Langendorff method. Adenosine increased ventricular arrhythmias in both groups of animals in a dose-dependent manner: 50% of chagasic rats developed ventricular fibrillation compared with 7.14% of healthy rats (P< .05). Fibrillation was prevented by A1 (i.e., DPCPX) and A2a (i.e., 8-CSC) receptor antagonists. Arrhythmia was associated with a prolonged QT interval, early depolarization, and the R-on-T and torsade de pointes phenomena. In conclusion, adenosine is a proarrythmic drug that is able to induce ventricular fibrillation in chagasic rat hearts.

A single-institutional brachytherapy experience in the management of esophageal cancer.

PURPOSE: The aim of this work was to analyze the efficacy of high-dose-rate brachytherapy in the management of patients with esophageal cancer. METHODS AND MATERIALS: Between 2001 and 2008, 104 patients (88 males and 16 females) received a brachytherapy treatment as a part of a personalized multimodal approach. The median age was 72 years, and the median Karnofsky performance status was 60. Brachytherapy was used in different situations: to complete a primary treatment with radical intent in patients not suitable for surgery; to control local recurrences; or to obtain a rapid and durable palliation of dysphagia in patients with metastases or in poor general condition. In selected cases, endoscopic ultrasound images were integrated in treatment planning procedure to obtain a more accurate volume definition. Efficacy in controlling dysphagia was assessed 1 month after brachytherapy. RESULTS: Fifty-one patients received a radical treatment, and 53 patients were treated to relieve the symptoms. The median overall survival was 20.8 months. Lymph node involvement was found to be a significant prognostic factor for overall survival and dysphagia. Dysphagia was controlled in 88 patients (84.6%), and the median dysphagia-free interval was 17.5 months. Early and late adverse events were generally mild to moderate. The most severe effects were esophagotracheal fistulas, observed in 3 patients. CONCLUSIONS: Brachytherapy has an important role in the management of esophageal cancer with a low rate of complications. endoscopic ultrasound-assisted treatment planning can be useful for a better individualization of curative treatments.