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BackgroundMyalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) presents significant challenges for affected children and adolescents, their social environment, and treating physicians, due to its profound impact on quality of life and the lack of causal therapeutic approaches. One crucial aspect of care that has been missing for these patients is comprehensive education for both them and their social circles. ObjectiveThis study protocol aims to outline the goals, study design, execution, and evaluation of the subproject within the BAYNET FOR ME/CFS project. The focus is on developing online education programs for children and adolescents with ME/CFS, as well as for their parents, siblings, and school staff. These programs are designed to improve independent disease management, increase knowledge, and promote interaction with other affected individuals. MethodsIn phase I, the group-based online education programs were developed by a multidisciplinary team based on the ModuS concept created by the Competence Network for Patient Education (KomPaS). These programs were then piloted and finalized. Phase II involved recruiting participants and implementing the finalized programs. Given the restricted physical and cognitive capacities of the affected individuals, the patient education programs were exclusively designed in a digital format to facilitate participation. In phase III, the programs will be evaluated for acceptance, completeness, and participant satisfaction. The qualitative assessment will focus on individual expectations and benefits derived from the training. Phase IV will further assess the programs in terms of improvements in disease knowledge, health-related quality of life, life satisfaction, and family burden. ResultsThe programs were developed, piloted, and finalized during phase I, which ran from December 2022 to May 2023. The pilot phase, from March to May 2023, led to adaptations in the program concept. In total, 8 patients and their parents, 5 siblings, and 59 school staff participated in the piloting. Adjustments were made to the format, content, duration, and schedule to better meet the needs of the affected individuals and their social circles. In phase II, participant recruitment for the patient education program took place from January to July 2023. The study successfully recruited 24 young patients with ME/CFS and their parents, along with 8 siblings and 51 school staff. Two program blocks for patients and parents and 2-3 blocks for siblings and school staff commenced in May 2023 and were completed within the same year. Phase III began after phase II and involves the evaluation of the programs, with the process expected to conclude by the end of 2024. Phase IV, planned for 2025-2026, will involve the rollout of the program to 150 children and their caretakers. This phase will focus on evaluating disease knowledge, health-related quality of life, life satisfaction, and family burden, as well as include longitudinal assessments. ConclusionsThe data aim to support the development of a comprehensive, interprofessional care model for children and adolescents with ME/CFS. International Registered Report Identifier (IRRID)DERR1-10.2196/54679

Introduction: Patients with metastatic NSCLC (mNSCLC) treated with immune checkpoint inhibitors in clinical practice may often not meet the strict inclusion criteria of clinical trials. Our aim was to assess the trial eligibility of patients with mNSCLC treated with pembrolizumab monotherapy in real-world and to compare the outcome of “trial-ineligible” and “potentially trial-eligible” patients. Methods: Data from the prospective, clinical research platform CRISP were used to compare patient characteristics, treatment, and outcome of patients with programmed cell death-ligand 1 tumor proportion score greater than or equal to 50% tumors treated with pembrolizumab monotherapy who are deemed either “potentially trial-eligible” or “trial-ineligible” according to inclusion and exclusion criteria of the registrational studies (KEYNOTE-024 and -042). Results: Of 746 patients included, 343 patients (46.0%) were classified as “trial-ineligible” and had significantly worse outcomes compared with “potentially trial-eligible” patients (n = 403, 54.0%): median progression-free survival: 6.2 (95% confidence interval [CI]: 5.2–8.4) versus 10.3 (95% CI: 8.4–13.8) months, hazard ratio (trial-ineligible versus potentially trial-eligible) of 1.43 (95% CI: 1.19–1.72), p less than 0.001; median overall survival: 15.9 (95% CI: 11.4–20.3) versus 25.3 (95% CI: 19.8–30.4) months, hazard ratio of 1.36 (95% CI: 1.10–1.67), p equals 0.004. Conclusions: Our data reveal that a considerable proportion of patients with mNSCLC are not eligible to participate in a clinical trial and were found to have worse outcomes than potentially trial-eligible patients, whose outcomes were comparable with those obtained from pivotal clinical trials. This is of substantial clinical relevance for physicians discussing outcomes to be expected with their patients and stresses the need for real-world effectiveness analyses.

Abstract Altermagnetism represents an emergent collinear magnetic phase with compensated order and an unconventional alternating even-parity wave spin order in the non-relativistic band structure. We investigate directly this unconventional band splitting near the Fermi energy through spin-integrated soft X-ray angular resolved photoemission spectroscopy. The experimentally obtained angle-dependent photoemission intensity, acquired from epitaxial thin films of the predicted altermagnet CrSb, demonstrates robust agreement with the corresponding band structure calculations. In particular, we observe the distinctive splitting of an electronic band on a low-symmetry path in the Brilliouin zone that connects two points featuring symmetry-induced degeneracy. The measured large magnitude of the spin splitting of approximately 0.6 eV and the position of the band just below the Fermi energy underscores the significance of altermagnets for spintronics based on robust broken time reversal symmetry responses arising from exchange energy scales, akin to ferromagnets, while remaining insensitive to external magnetic fields and possessing THz dynamics, akin to antiferromagnets.

Abstract Static magnetic stimulation (SMS) is a form of non-invasive brain stimulation that alters neural activity and induces neural plasticity that outlasts the period of stimulation. This can modify corticospinal excitability or motor behaviours, suggesting that SMS may alter the intrinsic excitability of neurons. In mammalian neurons, the axon initial segment (AIS) is the site of action potential initiation and undergoes structural plasticity (changes in length and position from the soma) as a homeostatic mechanism to counteract chronic changes in neuronal activity. We investigated whether the chronic application of SMS (6 and 48 h, 0.5 T) induces structural AIS plasticity in postnatally derived primary cortical neurons. Following 6 h of SMS, we observed a shortening in mean AIS length compared to control, that persisted 24 h post stimulation. In contrast, 48 h of SMS induced an immediate distal shift that persisted 24 h post-stimulation. Pharmacological blockade of voltage gated L/T-type calcium channels during stimulation did not prevent SMS-induced AIS structural plasticity. Our findings provide the foundation to expand the use of chronic SMS as a non-invasive method to promote AIS plasticity.

Macroautophagy (referred to as autophagy hereafter) is a major intracellular lysosomal degradation pathway that is responsible for the degradation of misfolded/damaged proteins and organelles. Previous studies showed that autophagy protects against acetaminophen (APAP)-induced injury (AILI) via selective removal of damaged mitochondria and APAP protein adducts. The lysosome is a critical organelle sitting at the end stage of autophagy for autophagic degradation via fusion with autophagosomes. In the present study, we showed that transcription factor EB (TFEB), a master transcription factor for lysosomal biogenesis, was impaired by APAP resulting in decreased lysosomal biogenesis in mouse livers. Genetic loss-of and gain-of function of hepatic TFEB exacerbated or protected against AILI, respectively. Mechanistically, overexpression of TFEB increased clearance of APAP protein adducts and mitochondria biogenesis as well as SQSTM1/p62-dependent non-canonical nuclear factor erythroid 2-related factor 2 (NRF2) activation to protect against AILI. We also performed an unbiased cell-based imaging high-throughput chemical screening on TFEB and identified a group of TFEB agonists. Among these agonists, salinomycin, an anticoccidial and antibacterial agent, activated TFEB and protected against AILI in mice. In conclusion, genetic and pharmacological activating TFEB may be a promising approach for protecting against AILI.

Abstract Assisted reproductive techniques are routinely used in livestock species to increase and enhance productivity. Ovarian hyperstimulation is a process that currently relies on administering pituitary‐derived follicle‐stimulating hormone (FSH) or equine chorionic gonadotropin in combination with other hormones to promote the maturation of multiple follicles and thereby achieve superovulation. The use of partially purified preparations of FSH extracted from natural sources is associated with suboptimal and variable results. Recombinant FSH (rFSH) has been produced in a variety of heterologous organisms. However, attaining a bioactive rFSH of high quality and at low cost for use in livestock remains challenging. Here we report the production and characterization of a single chain bovine rFSH consisting of the β‐ and α‐subunit fused by a polypeptide linker (scbFSH) using Leishmania tarentolae as heterologous expression system. This unicellular eukaryote is non‐pathogenic to mammals, can be grown in bioreactors using simple and inexpensive semisynthetic media at 26°C and does not require CO2 or bovine serum supplementation. Stable cell lines expressing scbFSH in an inducible fashion were generated and characterized for their productivity. Different culture conditions and purification procedures were evaluated, and the recombinant product was biochemically and biologically characterized, including bioassays in an animal model. The results demonstrate that L. tarentolae is a suitable host for producing a homogeneous, glycosylated and biologically active form of scbFSH with a reasonable yield.

The adenosine A2B receptor (A2BAR) is a member of a family of G-protein coupled receptors (GPCRs), which has a low affinity for adenosine and is now implicated in several pathophysiological conditions. We have demonstrated the beneficial effects of A2BAR activation in enhancing recovery after acute liver injury induced by an acetaminophen (APAP) overdose. While receptor trafficking within the cell is recognized to play a role in GPCR signaling, its role in the mediation of A2BAR effects in the context of APAP-induced liver injury is not well understood. This was investigated here, where C57BL/6J mice were subjected to an APAP overdose (300 mg/kg), and the temporal course of A2BAR intracellular localization was examined. The impact of A2BAR activation or inhibition on trafficking was examined by utilizing the A2BAR agonist BAY 60-6583 or antagonist PSB 603. The modulation of A2BAR trafficking via APAP-induced cell signaling was explored by using 4-methylpyrazole (4MP), an inhibitor of Cyp2E1 and JNK activation. Our results indicate that APAP overdose induced the translocation of A2BAR to mitochondria, which was prevented via 4MP treatment. Furthermore, we demonstrated that A2BAR is localized on the mitochondrial outer membrane and interacts with progesterone receptor membrane component 1 (PGRMC1). While the activation of A2BAR enhanced mitochondrial localization, its inhibition decreased PGRMC1 mitochondria levels and blunted mitochondrial Cyp2E1 activity. Thus, our data reveal a hitherto unrecognized consequence of A2BAR trafficking to mitochondria and its interaction with PGRMC1, which regulates mitochondrial Cyp2E1 activity and modulates APAP-induced liver injury.

As remnants of living organisms, alkenones and isoprenoid glycerol dialkyl glycerol tetraether lipids (isoGDGTs) are widely used biomarkers for determining ocean water temperatures from the past. The organisms that these proxy carriers stem from are influenced by a number of environmental parameters, such as water depth, nutrient availability, light conditions, or seasonality, which all may significantly bias the calibration to ambient water temperatures. Reliable temperature determinations thus remain challenging, especially in higher latitudes and for undersampled regions. We analyzed 33 sediment surface samples from the southern Chilean continental margin and the Drake Passage for alkenones and isoGDGTs and compared the results with gridded instrumental reference data from the World Ocean Atlas 2005 (WOA05) and previously published data from an extended study area covering the central and western South Pacific towards the Aotearoa / New Zealand continental margin. We show that for alkenone-derived sea surface temperatures (SSTs), the widely used global core-top calibration of Müller et al. (1998) yields the smallest deviation of the WOA05-based SSTs. On the contrary, the calibration of Sikes et al. (1997), determined for higher latitudes and summer SSTs, overestimates modern WOA05-based SSTs in both the annual mean and summer. Our alkenone SSTs show a slight seasonal shift of ∼ 1 ∘C at the southern Chilean margin and up to ∼ 2 ∘C in the Drake Passage towards austral summer SSTs. Samples in the central South Pacific, on the other hand, reflect an annual mean signal. We show that for isoGDGT-based temperatures, the subsurface calibration of Kim et al. (2012a) best reflects temperatures from the WOA05 in areas north of the Subantarctic Front (SAF). Temperatures south of the SAF are, in contrast, significantly overestimated by up to 14 ∘C, irrespective of the applied calibration. In addition, we used the GDGT [2]/[3] ratios, which give an indication of the production depth of the isoGDGTs and/or potential influences from land. Our samples reflect a subsurface (0–200 m water depth) rather than a surface (0–50 m water depth) signal in the entire study area and show a correlation with the monthly dust distribution in the South Pacific, indicating terrigenous influences. The overestimation of isoGDGT surface and subsurface temperatures south of the SAF highlights the need for a reassessment of existing calibrations in the polar Southern Ocean. Therefore, we suggest a modified Southern Ocean tetraether index (TEX86)-based calibration for surface and subsurface temperatures, which shows a lower temperature sensitivity and yields principally lower absolute temperatures, which align more closely with WOA05-derived values and also OH–isoGDGT-derived temperatures.

Mitochondria are critical organelles responsible for the maintenance of cellular energy homeostasis. Thus, their dysfunction can have severe consequences in cells responsible for energy-intensive metabolic function, such as hepatocytes. Extensive research over the last decades have identified compromised mitochondrial function as a central feature in the pathophysiology of liver injury induced by an acetaminophen (APAP) overdose, the most common cause of acute liver failure in the United States. While hepatocyte mitochondrial oxidative and nitrosative stress coupled with induction of the mitochondrial permeability transition are well recognized after an APAP overdose, recent studies have revealed additional details about the organelle’s role in APAP pathophysiology. This concise review highlights these new advances, which establish the central role of the mitochondria in APAP pathophysiology, and places them in the context of earlier information in the literature. Adaptive alterations in mitochondrial morphology as well as the role of cellular iron in mitochondrial dysfunction and the organelle’s importance in liver recovery after APAP-induced injury will be discussed.

Abstract This study provides a descriptive characterization of the modern sedimentary processes in Lake Issyk Kul, Kyrgyzstan, important for the selection of a suitable coring or deep‐drilling site, interpretation of future core data and applicability of proxies. The quasi‐equidistant sampling grid of 66 sediment surface samples covers the entirety of the lake basin and is complemented by 10 samples from the major inflows. The methodological approach includes geochemical, granulometric, lipid biomarker, diatom, and statistical analyses. The quantitative and qualitative changes in sediment composition yield information on its generic origin and prevailing transport and depositional environments. The composition of the surface sediments in Issyk Kul is highly heterogenous. Nearshore deposition is mainly controlled by wave action and by fluvial sediment supply with highest quantities of detrital input coming from the high‐energetic, eastern tributaries. Sediments in the deep central basin are mainly produced in situ and dominated by authigenic calcite. Biogenic accumulation is overall low, except for the western extremity of the lake, where the nearshore, shallow‐water, and low‐energetic environment favors aquatic productivity and subsequent preservation of organic material and diatoms. Redeposition of sediments is a dominant process along the slopes across the southern and western basin floor, where run‐out distances of mass movement deposits are up to 5 km. Directional sediment transport by lake currents appears to be less important, except for the transport of very fine‐grained organic matter. Biomarker‐inferred temperature reconstructions suggest lake surface temperatures of ∼15°C in the western littoral zone and in Tyup Bay and a decrease to ∼13°C basinward.

Abstract Background Long periods of uninterrupted sitting, i.e., sedentary bouts, and their relationship with adverse health outcomes have moved into focus of public health recommendations. However, evidence on associations between sedentary bouts and adiposity markers is limited. Our aim was to investigate associations of the daily number of sedentary bouts with waist circumference (WC) and body mass index (BMI) in a sample of middle-aged to older adults. Methods In this cross-sectional study, data were collected from three different studies that took place in the area of Greifswald, Northern Germany, between 2012 and 2018. In total, 460 adults from the general population aged 40 to 75 years and without known cardiovascular disease wore tri-axial accelerometers (ActiGraph Model GT3X+, Pensacola, FL) on the hip for seven consecutive days. A wear time of ≥ 10 h on ≥ 4 days was required for analyses. WC (cm) and BMI (kg m− 2) were measured in a standardized way. Separate multilevel mixed-effects linear regression analyses were used to investigate associations of sedentary bouts (1 to 10 min, >10 to 30 min, and >30 min) with WC and BMI. Models were adjusted for potential confounders including sex, age, school education, employment, current smoking, season of data collection, and composition of accelerometer-based time use. Results Participants (66% females) were on average 57.1 (standard deviation, SD 8.5) years old and 36% had a school education >10 years. The mean number of sedentary bouts per day was 95.1 (SD 25.0) for 1-to-10-minute bouts, 13.3 (SD 3.4) for >10-to-30-minute bouts and 3.5 (SD 1.9) for >30-minute bouts. Mean WC was 91.1 cm (SD 12.3) and mean BMI was 26.9 kg m− 2 (SD 3.8). The daily number of 1-to-10-minute bouts was inversely associated with BMI (b = -0.027; p = 0.047) and the daily number of >30-minute bouts was positively associated with WC (b = 0.330; p = 0.001). All other associations were not statistically significant. Conclusion The findings provide some evidence on favourable associations of short sedentary bouts as well as unfavourable associations of long sedentary bouts with adiposity markers. Our results may contribute to a growing body of literature that can help to define public health recommendations for interrupting prolonged sedentary periods. Trial registration Study 1: German Clinical Trials Register (DRKS00010996); study 2: ClinicalTrials.gov (NCT02990039); study 3: ClinicalTrials.gov (NCT03539237).

Acetaminophen (APAP) is a widely used pain reliever that can cause liver injury or liver failure in response to an overdose. Understanding the mechanisms of APAP-induced cell death is critical for identifying new therapeutic targets. In this respect it was hypothesized that hepatocytes die by oncotic necrosis, apoptosis, necroptosis, ferroptosis and more recently pyroptosis. The latter cell death is characterized by caspase-dependent gasdermin cleavage into a C-terminal and an N-terminal fragment, which forms pores in the plasma membrane. The gasdermin pores can release potassium, interleukin-1β (IL-1β), IL-18, and other small molecules in a sublytic phase, which can be the main function of the pores in certain cell types such as inflammatory cells. Alternatively, the process can progress to full lysis of the cell (pyroptosis) with extensive cell contents release. This review discusses the experimental evidence for the involvement of pyroptosis in APAP hepatotoxicity as well as the arguments against pyroptosis as a relevant mechanism of APAP-induced cell death in hepatocytes. Based on the critical evaluation of the currently available literature and understanding of the pathophysiology, it can be concluded that pyroptotic cell death is unlikely to be a relevant contributor to APAP-induced liver injury.

OBJECTIVES:. Proliferation of COVID-19 research underscored the need for improved awareness among investigators, research staff and bedside clinicians of the operational details of clinical studies. The objective was to describe the genesis, goals, participation, procedures, and outcomes of two research operations committees in an academic ICU during the COVID-19 pandemic. DESIGN:. Two-phase, single-center multistudy cohort. SETTING:. University-affiliated ICU in Hamilton, ON, Canada. PATIENTS:. Adult patients in the ICU, medical stepdown unit, or COVID-19 ward. INTERVENTIONS:. None. MEASUREMENTS AND MAIN RESULTS:. An interprofessional COVID Collaborative was convened at the pandemic onset within our department, to proactively coordinate studies, help navigate multiple authentic consent encounters by different research staff, and determine which studies would be suitable for coenrollment. From March 2020 to May 2021, five non-COVID trials continued, two were paused then restarted, and five were launched. Over 15 months, 161 patients were involved in 215 trial enrollments, 110 (51.1%) of which were into a COVID treatment trial. The overall informed consent rate (proportion agreed of those eligible and approached including a priori and deferred consent models) was 83% (215/259). The informed consent rate was lower for COVID-19 trials (110/142, 77.5%) than other trials (105/117, 89.7%; p = 0.01). Patients with COVID-19 were significantly more likely to be coenrolled in two or more studies (29/77, 37.7%) compared with other patients (13/84, 15.5%; p = 0.002). Review items for each new study were collated, refined, and evolved into a modifiable checklist template to set up each study for success. The COVID Collaborative expanded to a more formal Department of Critical Care Research Operations Committee in June 2021, supporting sustainable research operations during and beyond the pandemic. CONCLUSIONS:. Structured coordination and increased communication about research operations among diverse research stakeholders cultivated a sense of shared purpose and enhanced the integrity of clinical research operations.

Abstract Climate change impacts the structure, functioning, and distribution of species and ecosystems. It will shift ecosystem boundaries, potentially affecting vulnerable ecosystems, such as tropical Africa's high mountain ecosystems, i.e., afroalpine ecosystems, and their highly susceptible uniquely adapted species. However, ecosystems along these mountains are not expected to respond similarly to the change. The ericaceous woody vegetation, located between the low-elevation broadleaf forests and high-elevation afroalpine vegetation, are anticipated to be affected differently. We hypothesize that projected climate change will result in an upward expansion and increasing dominance of ericaceous vegetation, which will negatively impact the endemic rich afroalpine ecosystems of the extensive Sanetti plateau. Hence, we modeled the impact of future climate change on the distribution of ericaceous vegetation and discussed its effect on bordering ecosystems in the Bale Mountains. We applied four familiar correlative modeling approaches: bioclim, domain, generalized linear methods, and support vector machines. We used WorldClim’s bioclimatic variables as environmental predictors and two representative concentration pathways (RCPs) of the IPCC Fifth Assessment Report climate change scenarios, namely RCP4.5 and RCP8.5 for future climate projection. The results indicate increased ericaceous vegetation cover on the midaltitude of northwestern and northern parts of the massif, and the Sanetti plateau. We observed upward range expansion and increase of close ericaceous vegetation in midaltitudes, while receding from the lower range across the massif. Moreover, the current ericaceous vegetation range correlates to the temperature and precipitation trends, reaffirming the critical role of temperature and precipitation in determining species distributions along elevational gradients. The results indicate the high likelihood of considerable changes in this biodiversity hotspot in Eastern Africa.

Background & Aims: The liver has a unique capacity to regenerate after injury in a highly orchestrated and regulated manner. Here, we report that O-GlcNAcylation, an intracellular post-translational modification regulated by 2 enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), is a critical termination signal for liver regeneration following partial hepatectomy (PHX). Methods: We studied liver regeneration after PHX on hepatocyte specific OGT and OGA knockout mice (OGT-KO and OGA-KO), which caused a significant decrease (OGT-KO) and increase (OGA-KO) in hepatic O-GlcNAcylation, respectively. Results: OGA-KO mice had normal regeneration, but the OGT-KO mice exhibited substantial defects in termination of liver regeneration with increased liver injury, sustained cell proliferation resulting in significant hepatomegaly, hepatic dysplasia, and appearance of small nodules at 28 days after PHX. This was accompanied by a sustained increase in expression of cyclins along with significant induction in pro-inflammatory and pro-fibrotic gene expression in the OGT-KO livers. RNA-sequencing studies revealed inactivation of hepatocyte nuclear 4 alpha (HNF4α), the master regulator of hepatic differentiation and a known termination signal, in OGT-KO mice at 28 days after PHX, which was confirmed by both Western blot and immunohistochemistry analysis. Furthermore, a significant decrease in HNFα target genes was observed in OGT-KO mice, indicating a lack of hepatocyte differentiation following decreased hepatic O-GlcNAcylation. Immunoprecipitation experiments revealed HNF4α is O-GlcNAcylated in normal differentiated hepatocytes. Conclusions: These studies show that O-GlcNAcylation plays a critical role in the termination of liver regeneration via regulation of HNF4α in hepatocytes.

Acetaminophen (N-acetyl-para-aminophenol (APAP)) is one of the most-studied drugs worldwide [...]

Acetaminophen (APAP) is a widely used analgesic and antipyretic drug, which is safe at therapeutic doses but can cause severe liver injury and even liver failure after overdoses. The mouse model of APAP hepatotoxicity recapitulates closely the human pathophysiology. As a result, this clinically relevant model is frequently used to study mechanisms of drug-induced liver injury and even more so to test potential therapeutic interventions. However, the complexity of the model requires a thorough understanding of the pathophysiology to obtain valid results and mechanistic information that is translatable to the clinic. However, many studies using this model are flawed, which jeopardizes the scientific and clinical relevance. The purpose of this review is to provide a framework of the model where mechanistically sound and clinically relevant data can be obtained. The discussion provides insight into the injury mechanisms and how to study it including the critical roles of drug metabolism, mitochondrial dysfunction, necrotic cell death, autophagy and the sterile inflammatory response. In addition, the most frequently made mistakes when using this model are discussed. Thus, considering these recommendations when studying APAP hepatotoxicity will facilitate the discovery of more clinically relevant interventions.

Acetaminophen (APAP) overdose can induce liver injury and is the most frequent cause of acute liver failure in the United States. We investigated the role of p62/SQSTM1 (referred to as p62) in APAP-induced liver injury (AILI) in mice. We found that the hepatic protein levels of p62 dramatically increased at 24 h after APAP treatment, which was inversely correlated with the hepatic levels of APAP-adducts. APAP also activated mTOR at 24 h, which is associated with increased cell proliferation. In contrast, p62 knockout (KO) mice showed increased hepatic levels of APAP-adducts detected by a specific antibody using Western blot analysis but decreased mTOR activation and cell proliferation with aggravated liver injury at 24 h after APAP treatment. Surprisingly, p62 KO mice recovered from AILI whereas the wild-type mice still sustained liver injury at 48 h. We found increased number of infiltrated macrophages in p62 KO mice that were accompanied with decreased hepatic von Willebrand factor (VWF) and platelet aggregation, which are associated with increased cell proliferation and improved liver injury at 48 h after APAP treatment. Our data indicate that p62 inhibits the late injury phase of AILI by increasing autophagic selective removal of APAP-adducts and mitochondria but impairs the recovery phase of AILI likely by enhancing hepatic blood coagulation.

Abstract Background Physical fitness is an essential marker of health. The literature regarding the question of whether individuals with asthma have reduced physical fitness compared to their non-asthmatic peers is inconsistent and focuses on the cardiorespiratory endurance dimension. This study provides a comparison of different dimensions of physical fitness in individuals with and without asthma on the basis of the German population-based study “KiGGS” (German Health Interview and Examination Survey for Children and Adolescents) and its in-depth study “MoMo” (2009–2012: wave 1 and 2014–2017: wave 2). Methods In total, 7731 individuals aged 6–30 years were included in this cross-sectional analysis at two measurement waves, including 353 individuals with and 7378 without asthma. The 12-month prevalence of physician-diagnosed asthma was assessed by interview. Physical fitness was measured by six test items of the MoMo test profile. “Cardiorespiratory endurance” was measured by an ergometric test, “muscular strength” by standing long jump, push-ups and sit-ups and “coordination” by jumping sideways and balancing backwards. Because of the broad age range of the sample, age- and sex-specific percentiles were used. Physical activity, age, gender and general state of health were assessed by questionnaire. Results The individuals with asthma reported a poorer general state of health at both measurement waves. However, the results of the fitness tests indicated that they were as physically fit as their peers without asthma in relation to cardiorespiratory endurance and muscular strength. The mean percentiles were all within the same range. The results of the comparisons of coordination performance were inconsistent. At wave 1 they were within the same range, at wave 2 individuals with asthma showed a poorer coordination performance (p = 0.041; HL = 4.125, CI of HL 0.155–8.125). Conclusions To the best of our knowledge, this is the first study to compare the physical fitness of individuals with and without asthma by considering several dimensions of physical fitness. The study demonstrates that cardiorespiratory endurance and muscular strength are not reduced in individuals with asthma. The results of the comparisons at the two measurement waves were remarkably stable.

Na przełomie XIX i XX wieku w parlamencie westminsterskim rozegrał się jeden z najważniejszych, trwający wiele lat akt konkurencji między obydwoma izbami parlamentu o wpływ na realizację funkcji ustawodawczej w procedurze parlamentarnej. W niniejszym artykule, będącym wynikiem znacznie szerszych studiów nad przekształceniami wewnętrznymi parlamentaryzmu brytyjskiego, przedstawiono społeczne i polityczne tło tej walki toczącej się na forum obydwu izb między dwiema siłami politycznymi: Partią Konserwatywną i Partią Liberalną, wzmocnioną przez taktycznych sojuszników. W tekście starano się przedstawić prawny wynik tego sporu w postaci słynnego Parliament Act 1911, ale także – na podstawie dzisiejszego stanu wiedzy opartego również na zachowanych stenogramach wystąpień parlamentarnych – klimat tego sporu oraz jego społeczne i polityczne tło.

Fixed-timed artificial insemination (FTAI) protocols for beef cattle in South America are primarily based on estradiol esters and intravaginal progesterone-releasing devices (IVPD). The objective of this study was to determine the optimal gonadotropin-releasing hormone (GnRH)-based protocol as an alternative to the use of estrogen-based protocols in grazing Bos taurus suckling beef cows. All cows received an IVPD on the day of protocol initiation and prostaglandin F2α (PG) plus equine chorionic gonadotropin (eCG) treatments at the time of IVPD removal. In Experiment 1, cows (n = 235) were randomly assigned to one of four treatments: (i) 7-day estradiol = 2 mg of estradiol benzoate (EB) at IVPD insertion on Day 9 and 1 mg of estradiol cypionate (ECP) at IVPD removal on Day 2; (ii) 7-day GnRH = 10 µg of GnRH at IVPD insertion on Day 10, IVPD removal on Day 3 and GnRH at FTAI; (iii) 7 & 7 estradiol = PG at IVPD insertion on Day 16, EB on Day 9 and ECP at IVPD removal on Day 2; (iv) 7 & 7 GnRH = PG at IVPD insertion on Day 17, GnRH on Day 10, IVPD removal on Day 3 and GnRH at FTAI. In Experiment 2, cows (n = 462) were randomly assigned to one of four treatments: (i) 6-day estradiol = EB at IVPD insertion on Day 9, IVPD removal on Day 3 and GnRH at FTAI; (ii) 7-day estradiol; (iii) 7-day GnRH; (iv) 7 & 7 GnRH. In Experiment 1, plasma progesterone concentrations and percentage of cows with a corpus luteum (CL) at IVPD removal, and pregnancy per AI (P/AI) were greater for cows subjected to GnRH-based protocols compared with cows subjected to estrogen-based protocols (p < 0.01). In Experiment 2, cows subjected to the 7 & 7 GnRH protocol had the greatest P/AI (p < 0.01). In summary, GnRH-based FTAI protocols resulted in similar or greater P/AI compared to estrogen-based FTAI protocols in grazing postpartum Bos taurus suckled beef cows. The greatest P/AI was attained with the 7 & 7 GnRH protocol.

Acute hepatic failure is associated with high morbidity and mortality for which the only definitive therapy is liver transplantation. Some fraction of those who undergo emergency transplantation have been shown to recover native liver function when transplanted with an auxiliary hepatic graft that leaves part of the native liver intact. Thus, transplantation could have been averted with the development and use of some form of hepatic support. The costs of developing and testing liver support systems could be dramatically reduced by the availability of a reliable large animal model of hepatic failure with a large therapeutic window that allows the assessment of efficacy and timing of intervention. Non-lethal forms of hepatic injury were examined in combination with liver-directed radiation in non-human primates (NHPs) to develop a model of acute hepatic failure that mimics the human condition. Porcine hepatocyte transplantation was then tested as a potential therapy for acute hepatic failure. After liver-directed radiation therapy, delivery of a non-lethal hepatic ischemia-reperfusion injury reliably and rapidly generated liver failure providing conditions that can enable pre-clinical testing of liver support or replacement therapies. Unfortunately, in preliminary studies, low hepatocyte engraftment and over-immune suppression interfered with the ability to assess the efficacy of transplanted porcine hepatocytes in the model. A model of acute liver failure in NHPs was created that recapitulates the pathophysiology and pathology of the clinical condition, does so with reasonably predictable kinetics, and results in 100% mortality. The model allowed preliminary testing of xenogeneic hepatocyte transplantation as a potential therapy.

Currently approved repetitive transcranial magnetic stimulation (rTMS) protocols for the treatment of major depressive disorder (MDD) involve once-daily (weekday) stimulation sessions, with 10 Hz or intermittent theta burst stimulation (iTBS) frequencies, over 4–6 weeks. Recently, accelerated treatment protocols (multiple daily stimulation sessions for 1–2 weeks) have been increasingly studied to optimize rTMS treatments. Accelerated protocols might confer unique advantages for adolescents and young adults but there are many knowledge gaps related to dosing in this age group. Off-label, clinical practice frequently outpaces solid evidence as rigorous clinical trials require substantial time and resources. Murine models present an opportunity for high throughput dose finding studies to focus subsequent clinical trials in humans. This project investigated the brain and behavioural effects of an accelerated low-intensity rTMS (LI-rTMS) protocol in a young adult rodent model of chronic restraint stress (CRS). Depression and anxiety-related behaviours were induced in young adult male Sprague Dawley rats using the CRS model, followed by the 3-times-daily delivery of 10 Hz LI-rTMS, for two weeks. Behaviour was assessed using the Elevated Plus Maze and Forced Swim Test, and functional, chemical, and structural brain changes measured using magnetic resonance imaging techniques. CRS induced an agitated depression-like phenotype but therapeutic effects from the accelerated protocol were not detected. Our findings suggest that the age of rodents may impact response to CRS and LI-rTMS. Future studies should also examine higher intensities of rTMS and accelerated theta burst protocols.

Acetaminophen (N‐acetyl‐para‐aminophenol [APAP]) overdose is the most common cause of drug‐induced liver injury in the Western world and has limited therapeutic options. As an important dietary component intake, fructose is mainly metabolized in liver, but its impact on APAP‐induced liver injury is not well established. We aimed to examine whether fructose supplementation could protect against APAP‐induced hepatotoxicity and to determine potential fructose‐sensitive intracellular mediators. We found that both high‐fructose diet feeding before APAP injection and fructose gavage after APAP injection reduced APAP‐induced liver injury with a concomitant induction of the hepatic carbohydrate‐response element‐binding protein α (ChREBPα)–fibroblast growth factor 21 (FGF21) pathway. In contrast, Chrebpα liver‐specific‐knockout (Chrebpα‐LKO) mice failed to respond to fructose following APAP overdose, suggesting that ChREBPα is the essential intracellular mediator of fructose‐induced hepatoprotective action. Primary mouse hepatocytes with deletion of Fgf21 also failed to show fructose protection against APAP hepatotoxicity. Furthermore, overexpression of FGF21 in the liver was sufficient to reverse liver toxicity in APAP‐injected Chrebpα‐LKO mice. Conclusion: Fructose protects against APAP‐induced hepatotoxicity likely through its ability to activate the hepatocyte ChREBPα–FGF21 axis.

Background: Acetaminophen (APAP) overdose can cause liver injury and liver failure, which is one of the most common causes of drug-induced liver injury in the United States. Pharmacological activation of autophagy by inhibiting mechanistic target of rapamycin (mTOR) protects against APAP-induced liver injury likely via autophagic removal of APAP-adducts and damaged mitochondria. In the present study, we aimed to investigate the role of genetic ablation of mTOR pathways in mouse liver in APAP-induced liver injury and liver repair/regeneration. Methods: Albumin-Cre (Alb-Cre) mice, mTORf/f and Raptorf/f mice (C57BL/6J background) were crossbred to produce liver-specific mTOR knockout (L-mTOR KO, Alb Cre+/-, mTORf/f) and liver-specific Raptor KO (L-Raptor, Alb Cre+/-, Raptor f/f) mice. Alb-Cre littermates were used as wild-type (WT) mice. These mice were treated with APAP for various time points for up to 48 h. Liver injury, cell proliferation, autophagy and mTOR activation were determined. Results: We found that genetic deletion of neither Raptor, an important adaptor protein in mTOR complex 1, nor mTOR, in the mouse liver significantly protected against APAP-induced liver injury despite increased hepatic autophagic flux. Genetic deletion of Raptor or mTOR in mouse livers did not affect APAP metabolism and APAP-induced c-Jun N-terminal kinase (JNK) activation, but slightly improved mouse survival likely due to increased hepatocyte proliferation. Conclusions: Our results indicate that genetic ablation of mTOR in mouse livers does not protect against APAP-induced liver injury but may slightly improve liver regeneration and mouse survival after APAP overdose.

Mitochondria have been studied for decades from the standpoint of metabolism and ATP generation. However, in recent years mitochondrial dynamics and its influence on bioenergetics and cellular homeostasis is also being appreciated. Mitochondria undergo regular cycles of fusion and fission regulated by various cues including cellular energy requirements and pathophysiological stimuli, and the network of critical proteins and membrane lipids involved in mitochondrial dynamics is being revealed. Hepatocytes are highly metabolic cells which have abundant mitochondria suggesting a biologically relevant role for mitochondrial dynamics in hepatocyte injury and recovery. Here we review information on molecular mediators of mitochondrial dynamics and their alteration in drug-induced liver injury. Based on current information, it is evident that changes in mitochondrial fusion and fission are hallmarks of liver pathophysiology ranging from acetaminophen-induced or cholestatic liver injury to chronic liver diseases. These alterations in mitochondrial dynamics influence multiple related mitochondrial responses such as mitophagy and mitochondrial biogenesis, which are important adaptive responses facilitating liver recovery in several contexts, including drug-induced liver injury. The current focus on characterization of molecular mechanisms of mitochondrial dynamics is of immense relevance to liver pathophysiology and have the potential to provide significant insight into mechanisms of liver recovery and regeneration after injury.

Background: The purpose of this proof-of-concept feasibility study was to determine if spike-triggered intraspinal microstimulation (ISMS), a form of activity dependent stimulation (ADS), results in improved motor performance in an ambulatory rat model of spinal cord injury (SCI). Methods: Experiments were carried out in adult male Sprague Dawley rats with moderate thoracic contusion injury. Rats were assigned to one of two groups: Control or ADS therapy. Four weeks post-SCI, all rats were implanted with a recording microelectrode in the left hindlimb motor cortex and a fine-wire stimulating electrode in the contralateral lumbar spinal cord. ADS was administered for 4 hours/day, 4 days/week, for 4 weeks. During therapy sessions, single-unit spikes were discriminated in real time in the hindlimb motor cortex and used to trigger stimulation in the spinal cord ventral horn. Control rats were similarly implanted with electrodes but did not receive stimulation therapy. Results: Motor performances of each rat were evaluated before SCI contusion, once a week post-SCI for four weeks (prior to electrode implantation), and once a week post-conditioning for four weeks. Basso, Beattie, and Bresnahan (BBB) locomotor scores were significantly improved in ADS rats compared to Control rats at 1 and 2 weeks after initiation of therapy. Foot fault scores on the Horizontal Ladder were significantly improved in ADS rats compared to pre-therapy ADS and Control rats after 1 week of therapy and recovered to near pre-injury scores after 3 weeks of therapy. The Ledged Beam test showed deficits after SCI in both ADS and Control rats but there were no significant differences between groups after 4 weeks of ADS therapy. Conclusions: These results show that chronic stimulation after spinal cord injury using a methodology of spike-triggered ISMS enhances behavioral recovery of locomotor function as measured by the BBB score and the Horizontal Ladder task. However, it is still uncertain if the behavioral improvements seen were dependent on spike-triggered ISMS.

Abstract Background Scaling-up and sustaining healthcare interventions can be challenging. Our objective was to describe how the 3 Wishes Project (3WP), a personalized end-of-life intervention, was scaled-up and sustained in an intensive care unit (ICU). Methods In a longitudinal mixed-methods study from January 12,013 - December 31, 2018, dying patients and families were invited to participate if the probability of patient death was > 95% or after a decision to withdraw life support. A research team member or bedside clinician learned more about each of the patients and their family, then elicited and implemented at least 3 personalized wishes for patients and/or family members. We used a qualitative descriptive approach to analyze interviews and focus groups conducted with 25 clinicians who cared for the enrolled patients. We used descriptive statistics to summarize patient, wish, and clinician characteristics, and analyzed outcome data in quarters using Statistical Process Control charts. The primary outcome was enrollment of terminally ill patients and respective families; the secondary outcome was the number of wishes per patient; tertiary outcomes included wish features and stakeholder involvement. Results Both qualitative and quantitative analyses suggested a three-phase approach to the scale-up of this intervention during which 369 dying patients were enrolled, having 2039 terminal wishes implemented. From a research project to clinical program to an approach to practice, we documented a three-fold increase in enrolment with a five-fold increase in total wishes implemented, without a change in cost. Beginning as a study, the protocol provided structure; starting gradually enabled frontline staff to experience and recognize the value of acts of compassion for patients, families, and clinicians. The transition to a clinical program was marked by handover from the research staff to bedside staff, whereby project catalysts mentored project champions to create staff partnerships, and family engagement became more intentional. The final transition involved empowering staff to integrate the program as an approach to care, expanding it within and beyond the organization. Conclusions The 3WP is an end-of-life intervention which was implemented as a study, scaled-up into a clinical program, and sustained by becoming integrated into practice as an approach to care.

Background & Aims: A coordinated stress and regenerative response is important after hepatocyte damage. Here, we investigate the phenotypes that result from genetic abrogation of individual components of the checkpoint kinase 2/transformation-related protein 53 (p53)/cyclin-dependent kinase inhibitor 1A (p21) pathway in a murine model of metabolic liver injury. Methods: Nitisinone was reduced or withdrawn in Fah-/- mice lacking Chk2, p53, or p21, and survival, tumor development, liver injury, and regeneration were analyzed. Partial hepatectomies were performed and mice were challenged with the Fas antibody Jo2. Results: In a model of metabolic liver injury, loss of p53, but not Chk2, impairs the oxidative stress response and aggravates liver damage, indicative of a direct p53-dependent protective effect on hepatocytes. Cell-cycle control during chronic liver injury critically depends on the presence of both p53 and its downstream effector p21. In p53-deficient hepatocytes, unchecked proliferation occurs despite a strong induction of p21, showing a complex interdependency between p21 and p53. The increased regenerative potential in the absence of p53 cannot fully compensate the surplus injury and is not sufficient to promote survival. Despite the distinct phenotypes associated with the loss of individual components of the DNA damage response, gene expression patterns are dominated by the severity of liver injury, but reflect distinct effects of p53 on proliferation and the anti-oxidative stress response. Conclusions: Characteristic phenotypes result from the genetic abrogation of individual components of the DNA damage-response cascade in a liver injury model. The extent to which loss of gene function can be compensated, or affects injury and proliferation, is related to the level at which the cascade is interrupted. Accession numbers of repository for expression microarray data: GSE156983, GSE156263, GSE156852, and GSE156252.

Various mechanisms were proposed as substantial drivers of (sub)tropical South American hydroclimate changes during the last deglaciation. However, the interpretation of past precipitation records from the regions affected by the South American Summer Monsoon, the dominant hydroclimatic system in (sub)tropical South America, still insufficiently consider feedbacks between oceanic and atmospheric processes evident in modern observational data. Here, we evaluate ocean-atmosphere feedbacks active in the region from 19 to 4 ka based on a multi-proxy record comprising lipid biomarker, bulk sediment elemental composition and foraminiferal geochemistry from a sediment core retrieved from the tropical western South Atlantic offshore eastern Brazil at ~22°S. Our proxy data together with existing paleoclimate records show that the consideration of large scale synoptic climatic features across South America is crucial for understanding the past spatio-temporal rainfall variability, especially during the last deglaciation. While the paleohydrological data from our study site show relatively stable precipitation across the deglaciation in the core region of the South Atlantic Convergence Zone, distinct hydroclimatic gradients developed across the continent during Heinrich Stadial 1, which climaxed at ~16 ka. By then, the prevalent atmospheric and oceanic configuration caused more frequent extreme climatic events associated with positive rainfall in the northern portion of eastern South America and in the southeastern portion of the continent. These climatic extremes resulted from substantial warming of the sub(tropical) western South Atlantic sea surface that fostered oceanic moisture transport towards the continent and the reconfiguration of quasi-stationary atmospheric patterns. We further find that enhanced continental precipitation in combination with low glacial sea level strongly impacted marine ecosystems via enhanced terrigenous organic matter input in line with augmented nutrient release to the ocean. Extreme rainfall events similar to those that occurred during Heinrich Stadial 1 are likely to recur in South America as a consequence of global warming, because the projected reduction of the intra-hemispheric temperature gradient may lead to the development of atmospheric patterns similar to those in force during Heinrich Stadial 1.

Anthropogenic climate change is challenging biodiversity conservation worldwide. Climate change metrics derived from future climate predictions help to assess potential impacts of climate change on biodiversity. Here we calculated future climate change velocities across biogeographical regions of terrestrial Europe and the Natura 2000 protected area network, the largest protected area network on Earth. We applied climate projections for the year 2070, considering two emission scenarios, six global climate models and a fine spatial resolution. Areas with very high climate change velocity were identified as climate change hotspots, while areas with very low velocity were recognized as coldspots. We further revealed where and to what extent climate change hotspots and coldspots coincide with Natura 2000 sites. We found that climate change velocities are projected highest in the Continental and Boreal regions, and lowest in the Mediterranean and Anatolian regions. However, the Alpine region will likely contain largest areal proportions of climate change hotspots, while areal proportions of coldspots are projected largest in the Mediterranean region. High mountain regions such as the Alps show a high proportion of Natura 2000 sites that coincide with climate change hotspots. Both, hotspots and coldspots, are geographically associated with areas of topographic diversity. Low topographical diversity indicates high climate change exposure. The impact of hotspots increases with spatial isolation. Oceanic climate buffers climate change exposure in contrast to continental climate. However, continental regions of Europe tend to exhibit less spatial isolation. We recommend conservation action in climate change hotspots and coldspots to simultaneously protect the most climate-exposed biodiversity as well as climate change refugia. Climate change hotspots and coldspots overlapping with Natura 2000 sites should be considered priority conservation sites because new protected areas are hard to realize in densely populated landscapes of Europe. This study directs European conservation management and policy towards meeting international conservation goals in a climate-smart way.

Abstract To investigate abdominal volume determined by a new body scanner algorithm as anthropometric marker for Metabolic Syndrome (MetS) and its parameters compared to manually measured waist circumference (WC), we performed body scans in 411 participants (38% men, 20-81 years). WC and triglyceride, HDL-cholesterol, and fasting glucose concentrations, and blood pressure were assessed as MetS parameters. We used Spearman correlations and linear regression to investigate associations and goodness-of-fit (R², BIC) of abdominal volume and WC with MetS parameters, and logistic regression to analyse the discriminative power of WC and abdominal volume to assess likelihoods of MetS components and MetS. Correlations with triglyceride, HDL-cholesterol, and glucose concentration were slightly stronger for abdominal volume (r; 0.32, −0.32, and 0.34, respectively) than for WC (0.28, −0.28, and 0.29, respectively). Explained variances in MetS parameters were slightly higher and goodness-of-fit slightly better for abdominal volume than for WC, but differences were small. Exemplarily, glucose levels were 0.28 mmol/L higher (R² = 0.25; BIC = 945.5) per 1-SD higher WC, and 0.35 mmol/L higher (R² = 0.28; BIC = 929.1) per 1-SD higher abdominal volume. The discriminative power to estimate MetS components was similar for WC and abdominal volume. Our data show that abdominal volume allows metabolic characterization comparable to established WC.

Abstract Background Protein microarrays are a versatile and widely used tool for analyzing complex protein mixtures. Membrane arrays utilize antibodies which are captured on a membrane to specifically immobilize several proteins of interest at once. Using detection antibodies, the bound protein-antibody-complex is converted into visual signals, which can be quantified using densitometry. The reliability of such densitometric assessments depends on a variety of factors, not only sample preparation and the choice of acquisition device but also the selected analysis software and the algorithms used for readout and processing data. Currently available software packages use a single image of a membrane at an optimal exposure time selected for that specific experimental framework. This selection is based on a user’s best guess and is subject to inter-user variability or the acquisition device algorithm. With modern image acquisition systems proving the capacity to collect signal development over time, this information can be used to improve densitometric measurements. Here we introduce proMAD, a toolkit for protein microarray analysis providing a novel systemic approach for the quantification of membrane arrays based on the kinetics of the analytical reaction. Results Briefly, our toolkit ensures an exact membrane alignment, utilizing basic computer vision techniques. It also provides a stable method to estimate the background light level. Finally, we model the light production over time, utilizing the knowledge about the reaction kinetics of the underlying horseradish peroxidase-based signal detection method. Conclusion proMAD incorporates the reaction kinetics of the enzyme to model the signal development over time for each membrane creating an individual, self-referencing concept. Variations of membranes within a given experimental set up can be accounted for, allowing for a better comparison of such. While the open-source library can be implemented in existing workflows and used for highly user-tailored analytic setups, the web application, on the other hand, provides easy platform-independent access to the core algorithm to a wide range of researchers. proMAD’s inherent flexibility has the potential to cover a wide range of use-cases and enables the automation of data analytic tasks.

The zebrafish is a powerful model organism to study the mechanisms governing transition metal ions within whole brain tissue. Zinc is one of the most abundant metal ions in the brain, playing a critical pathophysiological role in neurodegenerative diseases. The homeostasis of free, ionic zinc (Zn2+) is a key intersection point in many of these diseases, including Alzheimer’s disease and Parkinson’s disease. A Zn2+ imbalance can eventuate several disturbances that may lead to the development of neurodegenerative changes. Therefore, compact, reliable approaches that allow the optical detection of Zn2+ across the whole brain would contribute to our current understanding of the mechanisms that underlie neurological disease pathology. We developed an engineered fluorescence protein-based nanoprobe that can spatially and temporally resolve Zn2+ in living zebrafish brain tissue. The self-assembled engineered fluorescence protein on gold nanoparticles was shown to be confined to defined locations within the brain tissue, enabling site specific studies, compared to fluorescent protein-based molecular tools, which diffuse throughout the brain tissue. Two-photon excitation microscopy confirmed the physical and photometrical stability of these nanoprobes in living zebrafish (Danio rerio) brain tissue, while the addition of Zn2+ quenched the nanoprobe fluorescence. Combining orthogonal sensing methods with our engineered nanoprobes will enable the study of imbalances in homeostatic Zn2+ regulation. The proposed bionanoprobe system offers a versatile platform to couple metal ion specific linkers and contribute to the understanding of neurological diseases.

Abstract Aim Due to their longevity and structure, forest ecosystems are particularly affected by climate change with consequences for their biodiversity, functioning, and services to mankind. In the European Union (EU), natural and seminatural forests are protected by the Habitats Directive and the Natura 2000 network. This study aimed to assess the exposure of three legally defined forest habitat types to climate change, namely (a) Tilio‐Acerion forests of slopes, screes, and ravines (9180*), (b) bog woodlands (91D0*), and (c) alluvial forests with Alnus glutinosa and Fraxinus excelsior (91E0*). We analyzed possible changes in their Bavarian distribution, including their potential future coverage by Natura 2000 sites. We hypothesized that protected areas (PAs) with larger elevational ranges will remain suitable for the forests as they allow for altitudinal distribution shifts. Methods To estimate changes in range size and coverage by PAs, we combined correlative species distribution models (SDMs) with spatial analyses. Ensembles of SDM‐algorithms were applied to two climate change scenarios (RCP4.5 and RCP8.5) of the HadGEM2‐ES model for the period 2061–2080. Results Our results revealed that bog woodlands experience the highest range losses (>2/3) and lowest PA coverage (max. 15% of sites with suitable conditions). Tilio‐Acerion forests exhibit opposing trends depending on the scenario, while alluvial forests are less exposed to climatic changes. As expected, the impacts of climate change are more pronounced under the “business as usual” scenario (RCP8.5). Additionally, PAs in flat landscapes are more likely to lose environmental suitability for currently established forest habitat types. Main conclusions Based on these findings, we advocate the expansion of the Natura 2000 network particularly in consideration of elevational gradients, connectivity, and projected climatic suitability. Nonclimatic stressors on forest ecosystems, especially bog woodlands, should be decreased and climate change mitigation efforts enhanced. We recommend transferring the approach to other habitat types and regions.

Liver injury and acute liver failure caused by acetaminophen (APAP) overdose is the clinically most important drug toxicity in Western countries. Mechanistic investigations have revealed a central role of mitochondria in the pathophysiology. Excess formation of the reactive metabolite N-acetyl-p-benzoquinone imine (NAPQI) after an overdose leads to hepatic glutathione depletion, mitochondrial protein adducts formation and an initial oxidant stress, which triggers the activation of mitogen activated protein (MAP) kinase cascade ultimately leading to c-jun N-terminal kinase (JNK) phosphorylation. Phospho-JNK translocates to the mitochondria and amplifies the oxidative and nitrosative stress eventually causing the mitochondrial membrane permeability transition pore opening and cessation of adenosine triphosphate (ATP) synthesis. In addition, mitochondrial matrix swelling ruptures the outer membrane and releases endonucleases, which cause nuclear deoxyribonucleic acid (DNA) fragmentation. Together, the nuclear DNA damage and the extensive mitochondrial dysfunction result in necrotic cell death. However, the pro-cell death signaling events are counteracted by adaptive responses such as autophagy and mitochondrial biogenesis. The improved mechanistic insight into the pathophysiology leads to better understanding of the mechanisms of action of the existing antidote N-acetylcysteine and justifies the clinical testing of novel therapeutics such as 4-methylpyrazole and calmangafodipir. Keywords: Acetaminophen (APAP) hepatotoxicity, Mitochondrial dysfunction, Biogenesis, Oxidant stress, Peroxynitrite, c-Jun N-terminal kinase (JNK)

Abstract Background Protected areas (PAs) are aimed to hold the environmental conditions that facilitate species and ecosystems to persist. PAs can become climatically unsuitable and unable to sustain their current number of species under climate change. The Natura 2000 (N2K) is the largest coordinated conservation tool assigned to maintain the long-term survival of Europe’s most significant species and habitats. In attempting to understand the effectiveness of PAs in the face of climate change scenarios, we tested two hypotheses: (1) PAs in the Alpine and the Boreal biogeographical regions will experience more newly emerged climate conditions (hotter and drier) compared to the climate representation of other biogeographical regions under future climate in Europe and (2) PAs in the Mediterranean and the Continental biogeographical regions will face more consistency in climate conditions due to less area of disappearing and novel climate in future. Methods Current climate data (1960–1990) and projections for 2050 and 2070 of PAs of N2K were extracted from WorldClim global climate data. Principal components analysis (PCA) was performed to construct climate space for the PAs across the biogeographical regions based on 19 climatic variables assessed at 5-km resolution. ArcMap 10.1 was used to map the location of the novel and disappearing climates. Results PAs in the Alpine region will experience more novel climate conditions in the future compared to other biogeographical regions. The future projections showed that 17.70% of the PAs in the Alpine region will experience novel climate by 2070. Considerable climate consistency was observed in the PAs in the Continental region compared to the other biogeographical regions. Our results showed that about 176 km2 of the selected PAs in the Continental region will face new emerging climate, while about 110 km2 will disappear under RCP 8.5 scenario. The prediction also revealed that in the Mediterranean region 08 PAs will experience novel climate and 786 km2 areas in these PAs will face disappearing climate by 2070. We found that fewer areas of PAs in the Boreal regions will experience disappearing climate in both the scenarios. Conclusions The portion of novel climate conditions can be seen as a future opportunity to assign new reserves for the species. Our study highlights the importance of conservation planning to increase the connectivity between PAs, identifying novel conservation zones to maximize representation of habitats during the emerging climatic changes as well as designing strategies, management, and monitoring of the individual PAs.

Background and aim: Overdose of acetaminophen (APAP) leads to liver injury, which is one of the most common causes of liver failure in the United States. We previously demonstrated that pharmacological activation of autophagy protects against APAP-induced liver injury in mice via removal of damaged mitochondria and APAP-adducts (APAP-ADs). Using an image-based high-throughput screening for autophagy modulators, we recently identified that chlorpromazine (CPZ), a dopamine inhibitor used for anti-schizophrenia, is a potent autophagy inducer in vitro. Therefore, the aim of the present study is to determine whether CPZ may protect against APAP-induced liver injury via inducing autophagy. Methods: Wild type C57BL/6J mice were injected with APAP to induce liver injury. CPZ was administrated either at the same time with APAP (co-treatment) or 2 h later after APAP administration (post-treatment). Hemotoxyline and eosin (H&E) staining of liver histology, terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling (TUNEL) staining of necrotic cell death as well as serum levels of alanine aminotransferase (ALT) were used to monitor liver injury. Results: We found that CPZ markedly protected against APAP-induced liver injury as demonstrated by decreased serum levels of ALT, liver necrotic areas as well as TUNEL-positive cells in mice that were either co-treated or post-treated with CPZ. Mechanistically, we observed that CPZ increased the number of autolysosomes and decreased APAP-induced c-Jun N-terminal kinase activation without affecting the metabolic activation of APAP. Pharmacological inhibition of autophagy by chloroquine partially weakened the protective effects of CPZ against APAP-induced liver injury. Conclusions: Our results indicate that CPZ ameliorates APAP-induced liver injury partially via activating hepatic autophagy and inhibiting JNK activation. Keywords: Chlorpromazine (CPZ), Acetaminophen (APAP), Drug-induced liver injury, Necrosis, Autolysosome, Hepatotoxicity, Glutathione (GSH), Oxidant stress

Abstract Background Despite the large number of studies and reviews available, the evidence regarding the biological determinants of physical activity (PA) is inconclusive. In this umbrella review, we summarized the current evidence on the biological determinants of PA across the life course, by pooling the results of the available systematic literature reviews (SLRs) and meta-analyses (MAs). Methods We conducted an online search on MEDLINE, ISI Web of Science, Scopus, and SPORTDiscus databases up to January 2018. SLRs and MAs of observational studies that investigated the association between biological determinants of PA and having PA as outcome were considered eligible. The extracted data were assessed based on the importance of the determinants, the strength of evidence, and the methodological quality. Results We identified 19 reviews of which most were of moderate methodological quality. Determinants that were studied most frequently among all ages and demonstrated evidence suggesting a positive association to PA were younger age, being male, higher health status, and higher physical fitness levels. Among adults, normal birth weight was found to be positively associated to PA with convincing strength of evidence, while findings among adolescents were inconsistent and with limited strength of evidence. Conclusions Different social or behavioral factors may contribute to the decrease of PA with age and among females versus males, and creating programmes targeted at diverse ages, female population, and adults with abnormal birth weight is recommended. Future studies should use prospective study designs, standardized definitions of PA, and objective measurement methods of PA assessment.

Three-dimensional (3D) cell culture models that provide a biologically relevant microenvironment are imperative to investigate cell–cell and cell–matrix interactions in vitro. Semi-synthetic star-shaped poly(ethylene glycol) (starPEG)–heparin hydrogels are widely used for 3D cell culture due to their highly tuneable biochemical and biomechanical properties. Changes in gene expression levels are commonly used as a measure of cellular responses. However, the isolation of high-quality RNA presents a challenge as contamination of the RNA with hydrogel residue, such as polymer or glycosaminoglycan fragments, can impact template quality and quantity, limiting effective gene expression analyses. Here, we compare two protocols for the extraction of high-quality RNA from starPEG–heparin hydrogels and assess three subsequent purification techniques. Removal of hydrogel residue by centrifugation was found to be essential for obtaining high-quality RNA in both isolation methods. However, purification of the RNA did not result in further improvements in RNA quality. Furthermore, we show the suitability of the extracted RNA for cDNA synthesis of three endogenous control genes confirmed via quantitative polymerase chain reaction (qPCR). The methods and techniques shown can be tailored for other hydrogel models based on natural or semi-synthetic materials to provide robust templates for all gene expression analyses.

BackgroundVisceral leishmaniasis (VL) is re-emerging in Armenia since 1999 with 167 cases recorded until 2019. The objectives of this study were (i) to determine for the first time the genetic diversity and population structure of the causative agent of VL in Armenia; (ii) to compare these genotypes with those from most endemic regions worldwide; (iii) to monitor the diversity of vectors in Armenia; (iv) to predict the distribution of the vectors and VL in time and space by ecological niche modeling.Methodology/principal findingsHuman samples from different parts of Armenia previously identified by ITS-1-RFLP as L. infantum were studied by Multilocus Microsatellite Typing (MLMT). These data were combined with previously typed L. infantum strains from the main global endemic regions for population structure analysis. Within the 23 Armenian L. infantum strains 22 different genotypes were identified. The combined analysis revealed that all strains belong to the worldwide predominating MON1-population, however most closely related to a subpopulation from Southeastern Europe, Maghreb, Middle East and Central Asia. The three observed Armenian clusters grouped within this subpopulation with strains from Greece/Turkey, and from Central Asia, respectively. Ecological niche modeling based on VL cases and collected proven vectors (P. balcanicus, P. kandelakii) identified Yerevan and districts Lori, Tavush, Syunik, Armavir, Ararat bordering Georgia, Turkey, Iran and Azerbaijan as most suitable for the vectors and with the highest risk for VL transmission. Due to climate change the suitable habitat for VL transmission will expand in future all over Armenia.ConclusionsGenetic diversity and population structure of the causative agent of VL in Armenia were addressed for the first time. Further genotyping studies should be performed with samples from infected humans, animals and sand flies from all active foci including the neighboring countries to understand transmission cycles, re-emergence, spread, and epidemiology of VL in Armenia and the entire Transcaucasus enabling epidemiological monitoring.

Secondary education around the world has been significantly disrupted by covid-19. Students have been forced into new ways of independent learning, often using remote technologies, but without the social nuances and direct teacher interactions of a normal classroom environment. Using data from the School Attitudes Survey—which surveys students regarding the perceived level of difficulty, anxiety level, self-efficacy, enjoyability, subject relevance, and opportunities for creativity with regards to each of their school subjects—this study examines students' responses to this disruption from two very different schools with two very different experiences of the pandemic. This paper reports on the composite attitudinal profiles of students in the senior secondary levels at each school (Years 10–12, n = 834). The findings challenged our expectation that the increased difficulty and anxiety caused by the disruption would reduce perceived opportunities for creativity. Indeed, our analyses showed that the students at both schools demonstrated generally positive attitudes toward their learning and strongly associated opportunities for creativity with other attitudinal constructs including enjoyability, subject relevance, and self-efficacy. These complex associations made by the students appear to have buffered the impacts of the disruption, and they may even have supported creative resilience.

Abstract Background The study estimated service coverage for severe mental disorders (psychosis, bipolar disorder and moderate-severe depression), globally and regionally, using data collected from the Mental Health Atlas 2017. Methods Service coverage was defined as the proportion of people with a disorder contacting a mental health service among those estimated to have the disorder during a 12-month period. We drew upon 12-month service utilisation data from the Mental Health Atlas 2017. Expected prevalent cases of individual disorders were estimated using the disorder-specific prevalence rate estimates of the Global Burden of Disease Study 2016 and total population sizes. Methods for assessing the validity of country-reported service utilisation data were developed and applied. Outcomes From 177 countries, 50 countries provided reliable service coverage estimates for psychosis, along with 56 countries for bipolar disorder, and 65 countries for depression. The mean service coverage for psychosis was lowest in low- [10.9% (95% confidence interval (CI) 3.3–30.4)] and lower middle-income countries [21.5% (95% CI 11.9–35.7)] and highest in high-income countries [59.5% (95% CI 42.9–74.1)]. Service coverage for bipolar disorder ranged between 3.1% (95% CI 0.8–11.5) and 10.4% (95% CI 6.7–15.8). Mean service coverage for moderate-severe depression ranged between 2.9% (95% CI 1.3–6.3) for low-income countries and 31.1% (95% CI 18.3–47.6) for high-income countries. Interpretation The reporting method utilised by the Mental Health Atlas appears to be reliable for psychosis but not for depression. This method of estimating service coverage provides progress in tracking an important indicator for mental health; however, it highlights that considerable work is needed to further develop global mental health information systems.

The impairment of LDL receptor-related protein-1 (LRP1) in numerous cell types is associated with obesity, diabetes, and fatty liver disease. Here, we compared the metabolic phenotype of C57BL/6J wild-type and LRP1 knock-in mice carrying an inactivating mutation in the distal NPxY motif after feeding a low-fat diet or high-fat (HF) diet with cholesterol supplementation (HFHC) or HF diet without cholesterol supplementation. In response to HF feeding, both groups developed hyperglycemia, hyperinsulinemia, hyperlipidemia, increased adiposity, and adipose tissue inflammation and liver steatosis. However, LRP1 NPxY mutation prevents HFHC diet-induced hypercholesterolemia, reduces adipose tissue and brain inflammation, and limits liver progression to steatohepatitis. Nevertheless, this mutation does not protect against HFHC diet-induced insulin resistance. The selective metabolic improvement observed in HFHC diet-fed LRP1 NPxY mutant mice is due to an apparent increase of hepatic LDL receptor levels, leading to an elevated rate of plasma lipoprotein clearance and lower hepatic cholesterol levels. The unique metabolic phenotypes displayed by LRP1 NPxY mutant mice indicate an LRP1-cholesterol axis in modulating tissue inflammation. The LRP1 NPxY mutant mouse phenotype differs from phenotypes observed in mice with tissue-specific LRP1 inactivation, thus highlighting the importance of an integrative approach to evaluate how global LRP1 dysfunction contributes to metabolic disease development.

Northern Africa’s past climate is characterized by a prolonged humid period known as the African Humid Period (AHP), giving origin to the “Green Sahara” and supporting human settlements into areas that are now desert. The spatial and temporal extent of climate change associated with the AHP is, however, subject to ongoing debate. Uncertainties arise from the complex nature of African climate, which is controlled by the strength and interactions of different monsoonal systems, resulting in meridional shifts in rainfall belts and zonal movements of the Congo Air Boundary. Here, we examine a ∼12,500-years record of hydroclimate variability from Lake Dendi located in the Ethiopian highlands based on a combination of plant-wax-specific hydrogen (δD) and carbon (δ13C) isotopes. In addition, pollen data from the same sediment core are used to investigate the response of the regional vegetation to changing climate. Our δD record indicates high precipitation during peak AHP (ca. 10 to 8 ka BP) followed by a gradual transition toward a drier late Holocene climate. Likewise, vegetation cover changed from predominant grassland toward an arid montane forest dominated by Juniperus and Podocarpus accompanied by a general reduction of understory grasses. This trend is corroborated by δ13C values pointing to an increased contribution of C3 plants during the mid-to late Holocene. Peak aridity occurred around 2 ka BP, followed by a return to a generally wetter climate possibly linked to enhanced Indian Ocean Monsoon strength. During the last millennium, increased anthropogenic activity, i.e., deforestation and agriculture is indicated by the pollen data, in agreement with intensified human impact recorded for the region. The magnitude of δD change (40‰) between peak wet conditions and late Holocene aridity is in line with other regional δD records of East Africa influenced by the CAB. The timing and pace of aridification parallels those of African and Indian monsoon records indicating a gradual response to local insolation change. Our new record combining plant-wax δD and δ13C values with pollen highlights the sensitive responses of the regional vegetation to precipitation changes in the Ethiopian highlands.