Electrophysiology and 3D-imaging reveal properties of human intracardiac neurons and increased excitability with atrial fibrillation.

Altered autonomic input to the heart plays a major role in atrial fibrillation (AF). Autonomic neurons termed ganglionated plexi (GP) are clustered on the heart surface to provide the last point of neural control of cardiac function. To date the properties of GP neurons in humans are unknown. Here we have addressed this knowledge gap in human GP neuron structure and physiology in patients with and without AF. Human right atrial GP neurons embedded in epicardial adipose tissue were excised during open heart surgery performed on both non-AF and AF patients and then characterised physiologically by whole cell patch clamp techniques. Structural analysis was also performed after fixation at both the single cell and at the entire GP levels via three-dimensional confocal imaging. Human GP neurons were found to exhibit unique properties and structural complexity with branched neurite outgrowth. Significant differences in excitability were revealed between AF and non-AF GP neurons as measured by lower current to induce action potential firing, a reduced occurrence of low action potential firing rates, decreased accommodation and increased synaptic density. Visualisation of entire GPs showed almost all neurons are cholinergic with a small proportion of noradrenergic and dual phenotype neurons. Phenotypic distribution differences occurred with AF including decreased cholinergic and dual phenotype neurons, and increased noradrenergic neurons. These data show both functional and structural differences occur between GP neurons from patients with and without AF, highlighting that cellular plasticity occurs in neural input to the heart that could alter autonomic influence on atrial function. KEY POINTS: The autonomic nervous system plays a critical role in regulating heart rhythm and the initiation of AF; however, the structural and functional properties of human autonomic neurons in the autonomic ganglionated plexi (GP) remain unknown. Here we perform the first whole cell patch clamp electrophysiological and large tissue confocal imaging analysis of these neurons from patients with and without AF. Our data show human GP neurons are functionally and structurally complex. Measurements of action potential kinetics show higher excitability in GP neurons from AF patients as measured by lower current to induce action potential firing, reduced low firing action potential rates, and decreased action potential accommodation. Confocal imaging shows increased synaptic density and noradrenergic phenotypes in patients with AF. Both functional and structural differences occur in GP neurons from patients with AF that could alter autonomic influence on atrial rhythm.

Polygenic Scores Clarify the Relationship Between Mental Health and Gender Diversity.

Background: Gender-diverse individuals are at increased risk for mental health problems, but it is unclear whether this is due to shared environmental or genetic factors. Methods: In two SPARK samples, we tested for associations of 16 polygenic scores (PGSs) with quantitative measures of gender diversity and mental health. In study 1, 639 independent adults (59% autistic) reported their mental health with the Adult Self-Report and their gender diversity with the Gender Self-Report (GSR). The GSR has 2 dimensions: binary (degree of identification with the gender opposite that implied by sex designated at birth) and nonbinary (degree of identification with a gender that is neither male nor female). In study 2 (N = 5165), we used a categorical measure of gender identity. Results: In study 1, neuropsychiatric PGSs were positively associated with Adult Self-Report scores: externalizing was positively associated with the attention-deficit/hyperactivity disorder PGS (ß = 0.10 [0.03-0.17]), and internalizing was positively associated with the PGSs for depression (ß = 0.07 [0-0.14]) and neuroticism (ß = 0.10 [0.03-0.17]). Interestingly, GSR scores were not significantly associated with any neuropsychiatric PGS. However, GSR nonbinary was positively associated with the cognitive performance PGS (ß = 0.11 [0.05-0.18]), with the effect size comparable in magnitude to the associations of the neuropsychiatric PGSs with the Adult Self-Report. Additionally, GSR binary was positively associated with the nonheterosexual sexual behavior PGS (ß = 0.07 [0-0.14]). In study 2, the cognitive performance PGS effect replicated; transgender and nonbinary individuals had higher PGSs (t316 = 4.16). Conclusions: We showed that while gender diversity is phenotypically positively associated with mental health problems, the strongest PGS associations with gender diversity were with the cognitive performance PGS, not the neuropsychiatric PGSs.

This research explores the connection between gender diversity, mental health, and genetic factors. It reveals that gender-diverse individuals often experience more mental health issues. Interestingly, rather than finding evidence linking these mental health challenges to genetic risk factors, the study discovered a replicable positive correlation between gender diversity and genetic markers for higher cognitive performance. This suggests that gender-diverse individuals typically have more of these cognitive performance gene variants. Finally, the study presents some early evidence suggesting that interactions between the environment (e.g., stigma) and genetic risk explain some of the elevated risk to mental health in gender-diverse individuals.

Contralateral prophylactic mastectomy in a rural population: A single-institution experience.

Background: The incidence of contralateral prophylactic mastectomy (CPM) for unilateral breast cancer (UBC) has continued to increase, despite an absent survival benefit except in populations at highest risk for developing contralateral breast cancer (CBC). CPM rates may be higher in rural populations but causes remain unclear. A study performed at our institution previously found that 21.8 % of patients with UBC underwent CPM from 2000 to 2009. This study aimed to evaluate the CPM trend at a single institution serving a rural population and identify the CPM rate in average-risk patients. Methods: Retrospective review of patients who underwent mastectomies for UBC at our institution from 2017 to 2021 was performed. Analysis utilized frequencies and percentages, descriptive statistics, chi-square, and independent sample t-tests. Results: A total of 438 patients were included, of whom 64.4 % underwent bilateral mastectomy for UBC (CPM). Patients who underwent CPM were significantly younger, underwent genetic testing, had germline pathogenic variants, had a family history of breast cancer, had smaller tumors, underwent reconstruction, and had more wound infections. Of CPM patients, 50.4 % had no identifiable factors for increased risk of developing CBC. Conclusions: The rate of CPM in a rural population at a single institution increased from 21.8 % to 64.4 % over two decades, with an average-risk CPM rate of 50.4 %. Those that undergo CPM are more likely to undergo reconstruction and have more wound infections. Identifying characteristics of patients undergoing CPM in a rural population and the increased associated risks allows for a better understanding of this trend to guide conversations with patients. Key message: This study demonstrates that the rate of contralateral prophylactic mastectomy for unilateral breast cancers performed at a single institution serving a largely rural population has nearly tripled over the last two decades, with half of these patients having no factors that increase the risk for developing contralateral breast cancers. Contralateral prophylactic mastectomy was significantly associated with smaller tumors, younger age, genetic testing, germline pathogenic variants, family history of breast cancer, breast reconstruction, and increased wound infections.

Differences in Distress Between Rural and Non-rural Appalachian Breast Cancer Patient/Caregiver Dyads During the First Year of Treatment.

Introduction: Breast cancer patients and their caregivers living in rural Appalachia face substantial health disparities compared to their non-rural Appalachian counterparts. However, there is limited research on how these specific health disparities in rural Appalachian communities may impact patient psychological distress and caregiver strain during the first year of breast cancer treatment. Purpose: The purpose of the current study was to assess differences in patient psychological distress (depression and anxiety) and caregiver strain between rural non-rural Appalachian breast-cancer-affected dyads (patients and their caregivers) during the first year of treatment. Methods: A total of 48 Appalachian breast cancer patients (with a Stage I through Stage III diagnosis) and their identified caregiver (together, 'dyads') were identified from The University of Tennessee Medical Center across 2019 to 2020. Dyads completed follow-up surveys throughout the first year of treatment. In this prospective pilot study, measures on anxiety, depression and caregiver strain were self-reported and then analyzed using RM-ANOVA. Results: There was a statistically significant higher number of reports of patient depression and caregiver strain in rural-residing dyads compared to non-rural-residing dyads. However, there was not a statistically significant difference between rural and non-rural Appalachian dyads for patient-reported anxiety during the first year of treatment. Implications: The higher reported patient depression and caregiver strain among rural-residing Appalachian patients may indicate the need for implementing remote (e.g., telehealth) Cognitive Behavioral Therapy (CBT) to address the psychological needs of rural-residing dyads. Additionally, greater education from physicians to rural dyads on what to expect during treatment could alleviate caregiver strain.

Retrospective Analysis of Pelvic Limb Fracture Management in Companion Psittacine Birds (60 Cases).

Pelvic limb fractures carry significant morbidity in avian patients, and although management options are well researched, published data on long-term complication rates and mortality outcomes are limited. Here, we present a cross-sectional study evaluating pelvic limb long bone fractures in companion psittacine birds presenting to an exotic-only veterinary hospital in the United Kingdom between 2005 and 2020, focusing on fixation techniques and long-term outcomes. Of the 60 cases that met the inclusion criteria, 22 separate species were represented, with an age range of 8 weeks to 25 years and an even distribution of sexes, among those that had been sexed. The majority of fractures (71.7%) were tibiotarsal; femoral (15%) and tarsometatarsal (13.3%) bones represented the other fracture sites. Several different fracture management methods were used, including external coaptation, surgery, or cage rest. Average time from fracture identification to healing was 33 days, with a median of 31 days and a range of 11-121 days. Satisfactory resolution of fracture repair was achieved in 85.5% (47/55) of cases that were able to be followed to conclusion. Complications were identified in 41.7% (25/60) of fractures of all pelvic long bones. Complications during fracture management were more common in cases treated with external coaptation. The most common complication reported was patient interference with bandages, splints, or both. This study provides an overview of pelvic limb long bone fracture management outcomes, which should prove useful for avian practitioners in clinical practice.

Pedi-R-MAPP | the development, testing, validation, and refinement of a digital nutrition awareness tool.

BACKGROUND & AIMS: The Remote Malnutrition Application (R-MAPP) was developed during the COVID-19 pandemic to provide support for health care professionals (HCPs) working in the community to complete remote nutritional assessments and provide practical guidance for nutritional care. R-MAPP was adapted into Pediatric Remote Malnutrition Application (Pedi-R-MAPP) using a modified Delphi consensus, with the goal of providing a structured approach to completing a nutrition focused assessment as part of a technology enabled care service (TECS) consultation. The aim of this study was to develop and validate a digital version of Pedi-R-MAPP using the IDEAS framework (Integrate, Design, Assess and Share). METHODS: A ten-step process was completed using the IDEAS framework. This involved the four concept processes; Stage-1, Integrate (Step 1-3) identify the problem, specify the goal, and use an evidence-based approach. Stage-2, (Step 4-7) design iteratively and rapidly with user feedback. Stage 3, (Step 8-9) Assess rigorously, and Stage 4 (Step 9-10) publish and launch of the tool. RESULTS: Stage 1:Evidence-based development, Pedi-R-MAPP was developed using Delphi consensus methodology. Stage 2:Iteration & design, HCPs (n = 22) from UK, Europe, South Africa, and North America were involved four workshops to further develop a paper prototype of the tool and complete small-scale testing of a beta version of the tool which resulted in eight iterations. Stage 3:Assess rigorously, Small scale retrospective testing of the tool on children with congenital heart disease (n = 80) was completed by a single researcher, with iterative changes made to improve agreement with summary advice. Large scale testing amongst (n = 745) children in different settings was completed by specialist paediatric dietitians (n = 15) advice who recorded agreement with the summary advice compared with their own clinical assessment. Paediatric dietitians were in overall agreement with the summary advice in the tool 86% (n = 640), compared to their own clinical practice. The main reasons for disagreement were i) frequency of planned review 57.1% (n = 60/105), ii) need for ongoing dietetic review due to chronic condition 20.0% (n = 21/105), iii) disagreement with recommendation for discharge 16.2% (n = 17/105) and iv) concerns with faltering growth and/or need for condition specific growth charts 6.7% (7/105). Iterative changes were made to the algorithm, leading to an improvement in agreement of the summary advice on re-evaluation to 98% (p=<0.0001). CONCLUSION: A digital version of the Pedi-R-MAPP nutrition awareness tool was developed using the IDEAS framework. The summary advice provided by the tool achieved a high level of agreement when compared to paediatric dietetic assessment, by providing a structured approach to completing a remote nutrition focused assessment, along with identifying the frequency of follow-up or an in-person assessment.

Molecular basis of interactions between CaMKII and α-actinin-2 that underlie dendritic spine enlargement.

Ca2+/calmodulin-dependent protein kinase II (CaMKII) is essential for long-term potentiation (LTP) of excitatory synapses that is linked to learning and memory. In this study, we focused on understanding how interactions between CaMKIIα and the actin-crosslinking protein α-actinin-2 underlie long-lasting changes in dendritic spine architecture. We found that association of the two proteins was unexpectedly elevated within 2 minutes of NMDA receptor stimulation that triggers structural LTP in primary hippocampal neurons. Furthermore, disruption of interactions between the two proteins prevented the accumulation of enlarged mushroom-type dendritic spines following NMDA receptor activation. α-Actinin-2 binds to the regulatory segment of CaMKII. Calorimetry experiments, and a crystal structure of α-actinin-2 EF hands 3 and 4 in complex with the CaMKII regulatory segment, indicate that the regulatory segment of autoinhibited CaMKII is not fully accessible to α-actinin-2. Pull-down experiments show that occupation of the CaMKII substrate-binding groove by GluN2B markedly increases α-actinin-2 access to the CaMKII regulatory segment. Furthermore, in situ labelling experiments are consistent with the notion that recruitment of CaMKII to NMDA receptors contributes to elevated interactions between the kinase and α-actinin-2 during structural LTP. Overall, our study provides new mechanistic insight into the molecular basis of structural LTP and reveals an added layer of sophistication to the function of CaMKII.

Mechanisms of synergistic suppression of ALK-positive lung cancer cell growth by the combination of ALK and SHP2 inhibitors.

Lung cancer is a major cause of cancer-related deaths. Alectinib is the first line of treatment for patients with ALK-positive lung cancer, but the survival rate beyond 2-3 years is low. Co-targeting secondary oncogenic drivers such as SHP2 is a potential strategy for improving drug efficacy. This is because SHP2 is expressed ubiquitously, but ALK expression is largely restricted to cancer cells. Thus, the combination of ALK and SHP2 inhibitors may provide a way to restrict synergistic cytotoxicity to cancer cells only, by reducing the dose of SHP2 inhibitors required for anticancer action and minimising SHP2-dependent systemic toxicity. The objective of this study was to investigate whether the combination of a SHP2 inhibitor (SHP099) with alectinib would synergistically suppress the growth of ALK-positive lung cancer cells. Our results demonstrated that the drug combination significantly and synergistically decreased cell viability at relatively low concentrations in ALK-positive H3122 and H2228 cells, due to G1 cell cycle arrest and increased apoptosis because of suppressed downstream RAS/MAPK signalling. The drug combination also induced the expression of mediators of the intrinsic apoptotic pathway, Bim and cleaved caspase-3, and modulated the expression of cell cycle mediators cyclin D1, cyclin B1, and phosphorylated CDK1.

Heterotopic Ossification Bone Formation in the Frontal Bones of an African Grey Parrot (Psittacus erithacus).

A 5-year-old, male African grey parrot (Psittacus erithacus) was presented with multiple, slow-growing, firm, bilateral masses around the dorsal orbital rims. Computer tomographic imaging revealed mild, incomplete bridging bone formation on the rostrodorsal aspects of the head. A moderate amount of smooth bone formation was identified at the rostrodorsal aspect to the left orbit, with minimal associated soft tissue swelling. Surgical biopsies were collected from the masses and histopathological analysis of the most rostral right mass showed well-differentiated bone, surrounded by dense fibrous connective tissue. Scattered, well-differentiated osteocytes were present within the bone. No evidence of neoplastic changes or infectious agents were identified. The histopathological changes were consistent with metaplastic bone formation. History obtained from the owner revealed recent head trauma, which likely induced the cranial heterotopic ossification in the African grey parrot.

The Development of Breast Cancer-Related Lymphedema After Mastectomy in a Rural Population.

Breast cancer-related lymphedema (BCRL) is a lifelong condition that can impact the quality of life, affecting approximately 20% of breast cancer patients. Risk factors for the development of BCRL after mastectomy in rural populations have not been studied.Retrospective review of mastectomy patients from 2017 to 2021 was performed at a single institution. Statistical analysis included logistic and linear regression models.475 patients were included, and 40 (8.4%) patients were diagnosed with BCRL. Increased odds of developing BCRL were significantly associated with tumor-involved lymph nodes, radiation therapy, axillary lymphadenectomy, adjuvant chemotherapy, and endocrine therapy. Postmastectomy reconstruction significantly reduced the odds of developing BCRL. There was no significant association in our population with age, body mass index, diabetes, tobacco use, cancer type, or complications.This study demonstrates that individuals underrepresented in the literature, such as patients in largely rural communities, have some differences in risk factors for developing BCRL when compared to national studies.

A pro-inflammatory and fibrous cap thinning transcriptome profile accompanies carotid plaque rupture leading to stroke.

Atherosclerotic plaque rupture is the etiology of ischemic stroke and myocardial infarction. The molecular mechanisms responsible for rupture remain unclear, in part, due to the lack of data from plaques at the time of rupture. Ribosome-depleted total RNA was sequenced from carotid plaques obtained from patients undergoing carotid endarterectomy with high-grade stenosis and either (1) a carotid-related ischemic cerebrovascular event within the previous 5 days ('recently ruptured,' n = 6) or (2) an absence of a cerebrovascular event ('asymptomatic,' n = 5). Principal component analysis confirmed plaque rupture was responsible for the greatest percentage of the variability between samples (23.2%), and recently ruptured plaques were enriched for transcripts associated with inflammation and extracellular matrix degradation. Hierarchical clustering achieved differentiation of the asymptomatic from the recently ruptured plaques. This analysis also found co-expression of transcripts for immunoglobulins and B lymphocyte function, matrix metalloproteinases, and interferon response genes. Examination of the differentially expressed genes supported the importance of inflammation and inhibition of proliferation and migration coupled with an increase in apoptosis. Thus, the transcriptome of recently ruptured plaques is enriched with transcripts associated with inflammation and fibrous cap thinning and support further examination of the role of B lymphocytes and interferons in atherosclerotic plaque rupture.

Clinical autism subscales have common genetic liabilities that are heritable, pleiotropic, and generalizable to the general population.

The complexity of autism's phenotypic spectra is well-known, yet most genetic research uses case-control status as the target trait. It is undetermined if autistic symptom domain severity underlying this heterogeneity is heritable and pleiotropic with other psychiatric and behavior traits in the same manner as autism case-control status. In N = 6064 autistic children in the SPARK cohort, we investigated the common genetic properties of twelve subscales from three clinical autism instruments measuring autistic traits: the Social Communication Questionnaire (SCQ), the Repetitive Behavior Scale-Revised (RBS-R), and the Developmental Coordination Disorder Questionnaire (DCDQ). Educational attainment polygenic scores (PGS) were significantly negatively correlated with eleven subscales, while ADHD and major depression PGS were positively correlated with ten and eight of the autism subscales, respectively. Loneliness and neuroticism PGS were also positively correlated with many subscales. Significant PGS by sex interactions were found-surprisingly, the autism case-control PGS was negatively correlated in females and had no strong correlation in males. SNP-heritability of the DCDQ subscales ranged from 0.04 to 0.08, RBS-R subscales ranged from 0.09 to 0.24, and SCQ subscales ranged from 0 to 0.12. GWAS in SPARK followed by estimation of polygenic scores (PGS) in the typically-developing ABCD cohort (N = 5285), revealed significant associations of RBS-R subscale PGS with autism-related behavioral traits, with several subscale PGS more strongly correlated than the autism case-control PGS. Overall, our analyses suggest that the clinical autism subscale traits show variability in SNP-heritability, PGS associations, and significant PGS by sex interactions, underscoring the heterogeneity in autistic traits at a genetic level. Furthermore, of the three instruments investigated, the RBS-R shows the greatest evidence of genetic signal in both (1) autistic samples (greater heritability) and (2) general population samples (strongest PGS associations).

Assaying Protein Kinase A Activity Using a FRET-Based Sensor Purified from Mammalian Cells.

Protein Kinase A (PKA) is the major intracellular receptor for cAMP. Research into this prototype kinase is supported by kinase assays that are typically performed in vitro using radio-labeled ATP. For in vivo studies, genetically encoded FRET-based sensors have become popular for monitoring PKA activity. Here, we show that it is also possible to apply such reporters in vitro. We describe how to express and purify milligram quantities of a FRET-based PKA activity reporter using cultured human embryonic kidney cells. We demonstrate how to utilize the purified reporter in a plate reader to determine the IC50 for the widely utilized PKA inhibitor H89 in the presence of a physiologically relevant concentration of ATP. The protocol takes advantage of the economical transfection reagent polyethylenimine and can be performed in a standard cell culture facility. Whereas assays based on radiolabelling are more sensitive, the approach presented here has several advantages: It enables continuous measurement of changes in substrate phosphorylation; a single preparation produces enough reporter for thousands of recordings; the reporter has a long shelf life; and it avoids the safety considerations that arise when working with radioactive material.

Pedi-R-MAPP: The development of a nutritional awareness tool for use in remote paediatric consultations using a modified Delphi consensus.

BACKGROUND & AIMS: The Remote Malnutrition Application (R-MAPP) was developed during the COVID-19 pandemic to provide support for health care professionals (HCPs) working in the community to complete remote nutritional assessments, and provide practical guidance for nutritional care. The aim of this study was to modify the R-MAPP into a version suitable for children, Pediatric Remote Malnutrition Application (Pedi-R-MAPP), and provide a structured approach to completing a nutrition focused assessment as part of a technology enabled care service (TECS) consultation. METHODS: A ten-step process was completed: 1) permission to modify adult R-MAPP, 2) literature search to inform the Pedi-R-MAPP content, 3) Pedi-R-MAPP draft, 4) international survey of HCP practice using TECS, 5) nutrition experts invited to participate in a modified Delphi process, 6) first stakeholder meeting to agree purpose/draft of the tool, 7) round-one online survey, 8) statements with consensus removed from survey, 9) round-two online survey for statements with no consensus and 10) second stakeholder meeting with finalisation of the Pedi-R-MAPP nutrition awareness tool. RESULTS: The international survey completed by 463 HCPs, 55% paediatricians, 38% dietitians, 7% nurses/others. When HCPs were asked to look back over the last 12 months, dietitians (n = 110) reported that 5.7 ± 10.6 out of every 10 appointments were completed in person; compared to paediatricians (n = 182) who reported 7.5 ± 7.0 out of every 10 appointments to be in person (p < 0.0001), with the remainder completed as TECS consultations. Overall, 74 articles were identified and used to develop the Pedi-R-MAPP which included colour-coded advice using a traffic light system; green, amber, red and purple. Eighteen participants agreed to participate in the Delphi consensus and completed both rounds of the modified Delphi survey. Agreement was reached at the first meeting on the purpose and draft sections of the proposed tool. In round-one of the online survey, 86% (n = 89/104) of statements reached consensus, whereas in round-two 12.5% (n = 13/104) of statements reached no consensus. At the second expert meeting, contested statements were discussed until agreement was reached and the Pedi-R-MAPP could be finalised. CONCLUSION: The Pedi-R-MAPP nutrition awareness tool was developed using a modified Delphi consensus. This tool aims to support the technological transformation fast-tracked by the COVID-19 pandemic by providing a structured approach to completing a remote nutrition focused assessment, as well as identifying the frequency of follow up along with those children who may require in-person assessment.

Detection of individual spin species via frequency-modulated charge pumping.

We utilize a frequency-modulated charge pumping methodology to measure quickly and conveniently single "charge per cycle" in highly scaled Si/SiO2 metal-oxide-semiconductor field effect transistors. This is indicative of detection and manipulation of a single interface trap spin species located at the boundary between the SiO2 gate dielectric and Si substrate (almost certainly a Pb type center). This demonstration in sub-micrometer devices in which Dennard scaling of the gate oxide has yielded extremely large gate oxide leakage currents eliminates interference between the charge pumping current and the leakage phenomenon. The result is the ability to measure single trap charge pumping reliably and easily, which would otherwise be completely inaccessible due to oxide leakage. This work provides a unique and readily available avenue for single spin species detection and manipulation, which can be applied as a quantized standard of electrical current as well as to serve as a potentially useful platform for developing quantum engineering technologies. Finally, we discuss potential underlying physical mechanisms that are involved in producing a seemingly contradictory measure of both odd and even integer values for charge per cycle.

Osseous Migration of a Perforating Gastrointestinal Foreign Body in an Indian Runner Duck (Anas platyrhynchos domesticus).

An Indian runner duck (Anas platyrhynchos domesticus) was presented for a second opinion after a linear, metallic foreign body was identified on radiographic images. The primary veterinarian performed diagnostic imaging while investigating the presenting complaint of the duck's left pelvic limb lameness. The images obtained from a computed tomography scan performed during the second-opinion visit revealed a linear, metallic foreign body with an associated migration tract originating from the ventriculus and terminating in the proximal left femur. Significant osteomyelitis was noted at the proximal left femur associated with the presence of the linear, metallic object. The foreign body and the adhesions associated with its migration were removed in 2, staged, surgical procedures. Although penetrating ventricular foreign bodies have been previously reported, migration through the cortex of a long bone is an unusual presentation. This case demonstrates that perforating, migrating, gastrointestinal foreign bodies can result in lameness refractory to analgesia and ancillary supportive care.

Interactive effects of (±)-trans-U50488 and its stereoisomers with cannabinoids.

The adverse effects of mu opioid agonists have spurred a renewed interest in using kappa opioid receptor (KOR) agonists as analgesics. KOR agonists also have potential for development as diuretics for the treatment of edema and hypertension. Here, we evaluated the discriminative stimulus, antinociceptive, and diuretic effects of the kappa agonist (±)-trans-U-50488 and its stereoisomers (-)-(1S,2S)-U-50488 or (+)-(1R,2R)-U-50488) alone and in combination with the cannabinoid agonist (-)-CP 55,940. To establish (±)-U-50488 as a discriminative stimulus, rats (n = 12) were trained to discriminate intraperitoneal (i.p.) administration of 5.6 mg/kg of (±)-trans-U-50488 from saline under a fixed-ratio 20 (FR-20) schedule of food reinforcement. Then, antinociception was assessed using two procedures: warm water tail withdrawal and von Frey paw withdrawal. Diuretic effects were assessed in separate rats (n = 6/group). Doses of (±)-U-50488 and (-)-U-50488 that served as discriminative stimuli produced significant increases in urine output, but at lower doses than those that produced antinociception. In contrast, (+)-U-50488 alone had no discriminative stimulus or diuretic effects at the doses tested, but did produce antinociception in the von Frey assay. When three cannabinoids and morphine were tested in the (±)-U-50488 discrimination procedure to determine the similarity of these drugs' discriminative stimulus effects to those for (±)-U-50488, the rank order similarity was (-)-CP 55,940 > (-)-trans-THC > (+)-WIN 55,212-2 ≥ morphine. (-)-CP 55,940 alone (0.056 mg/kg) partially substituted for the discriminative stimulus effects of (±)-U-50488 and produced significant diuretic and antinociceptive effects. (-)-CP 55,940 in combination with (±)-U-50488 also produced a two-fold leftward shift in the discriminative stimulus curve for (±)-U-50488, and near-additive antinociception with (±)-U-50488 and (+)-U-50488. Further, the diuretic effect of (-)-CP 55,940 was enhanced by a dose of (+)-U50488, which itself did not alter urine output. These data together indicate that a combination of cannabinoid and kappa opioid agonists can enhance diuresis, but may have limited potential for serving as opioid-sparing pharmacotherapeutics for treatment of pain.

The effect of metformin in EML4-ALK+ lung cancer alone and in combination with crizotinib in cell and rodent models.

Cell based studies have suggested that the diabetes drug metformin may combine with the anaplastic lymphoma kinase receptor (ALK) inhibitor crizotinib to increase ALK positive lung cancer cell killing and overcome crizotinib resistance. We therefore tested metformin alone and in combination with crizotinib in vivo, by employing a xenograft mouse model of ALK positive lung cancer. We found that 14 days of daily oral metformin (100 mg/kg) alone had a moderate but statistically significant effect on tumour growth suppression, but in combination with crizotinib, produced no greater tumour suppression than crizotinib (25 mg/kg) alone. We also reassessed the effect of metformin on EML4-ALK positive lung cancer (H3122) cell viability. Although metformin alone did have a moderate effect on cell viability (30% suppression) this was only at a clinically irrelevant concentration (5 mM) and there was no additive effect with cytotoxic concentrations of crizotinib. Moreover, metformin did not overcome crizotinib resistance in our resistant cells. Nevertheless, we were able to show that metformin induces a G1-cell cycle arrest and apoptosis alone and in combination with crizotinib. Also, consistent with earlier work, the addition of insulin-like growth factor-1 (IGF-1) to EML4-ALK positive cancer cells reduced cell killing by crizotinib. We therefore hypothesised that the effect of metformin in vivo was not due to direct cytotoxicity on cancer cells, but by modulation of IGF-1 expression. We therefore measured levels of IGF-1 in plasma taken from mice treated with metformin, but found no difference between the drug treatment and control groups. We further hypothesised that the effect of metformin could be due to modulation of thrombospondin 1 (TSP-1), which metformin has been proposed to regulatein vivo, but again we found no difference between the experimental groups. Finally, we investigated the potential for liver and kidney toxicity, as well as CYP3A based interactions, from the combination of metformin with crizotinib.

High Energy Side and Rear American Football Head Impacts Cause Obvious Performance Decrement on Video.

The objective of this study was to compare head impact data acquired with an impact monitoring mouthguard (IMM) to the video-observed behavior of athletes' post-collision relative to their pre-collision behaviors. A total of n = 83 college and high school American football players wore the IMM and were video-recorded over 260 athlete-exposures. Ex-athletes and clinicians reviewed the video in a two-step process and categorized abnormal post-collision behaviors according to previously published Obvious Performance Decrement (OPD) definitions. Engineers qualitatively reviewed datasets to check head impact and non-head impact signal frequency and magnitude. The ex-athlete reviewers identified 2305 head impacts and 16 potential OPD impacts, 13 of which were separately categorized as Likely-OPD impacts by the clinical reviewers. All 13 Likely-OPD impacts were in the top 1% of impacts measured by the IMM (ranges 40-100 g, 3.3-7.0 m/s and 35-118 J) and 12 of the 13 impacts (92%) were to the side or rear of the head. These findings require confirmation in a larger data set before proposing any type of OPD impact magnitude or direction threshold exists. However, OPD cases in this study compare favorably with previously published impact monitoring studies in high school and college American football players that looked for OPD signs, impact magnitude and direction. Our OPD findings also compare well with NFL reconstruction studies for ranges of concussion and sub-concussive impact magnitudes in side/rear collisions, as well as prior theory, analytical models and empirical research that suggest a directional sensitivity to brain injury exists for single high-energy impacts.

Inhibition of Mitogen-Activated Protein Kinase Kinase Alone and in Combination with Anaplastic Lymphoma Kinase (ALK) Inhibition Suppresses Tumor Growth in a Mouse Model of ALK-Positive Lung Cancer.

Anaplastic lymphoma kinase (ALK)-positive non-small-cell lung cancer most commonly arises through EML4 (Echinoderm Microtuble Like 4)-ALK chromosomal fusion. We have previously demonstrated that combination of the ALK inhibitor crizotinib with the MEK inhibitor selumetinib was highly effective at reducing cell viability of ALK-positive non-small-cell lung cancer (H3122) cells. In this study, we further investigated the efficacy of crizotinib and selumetinib combination therapy in an in vivo xenograft model of ALK-positive lung cancer. Crizotinib decreased tumor volume by 52% compared with control, and the drug combination reduced tumor growth compared with crizotinib. In addition, MEK inhibition alone reduced tumor growth by 59% compared with control. Crizotinib and selumetinib alone and in combination were nontoxic at the dose of 25 mg/kg, with values for ALT (<80 U/l) and creatinine (<2 mg/dl) within the normal range. Our results support the combined use of crizotinib with selumetinib in ALK-positive lung cancer but raise the possibility that a sufficient dose of an MEK inhibitor alone may be as effective as adding an MEK inhibitor to an ALK inhibitor. SIGNIFICANCE STATEMENT: This study contains in vivo evidence supporting the use of combination MEK inhibitors in ALK+ lung cancer research, both singularly and in combination with ALK inhibitors. Contrary to previously published reports, our results suggest that it is possible to gain much of the benefit from combination treatment with an MEK inhibitor alone, at a tolerable dose.