Awakening Change-Embracing Disability in Medical Training.

This essay describes the author's experience as a medical trainee with narcolepsy with cataplexy, highlighting the toll of self-sacrifice and the need for a cultural shift in medicine to better support trainees with disabilities.

Adult neurogenesis improves spatial information encoding in the mouse hippocampus.

Adult neurogenesis is a unique form of neuronal plasticity in which newly generated neurons are integrated into the adult dentate gyrus in a process that is modulated by environmental stimuli. Adult-born neurons can contribute to spatial memory, but it is unknown whether they alter neural representations of space in the hippocampus. Using in vivo two-photon calcium imaging, we find that male and female mice previously housed in an enriched environment, which triggers an increase in neurogenesis, have increased spatial information encoding in the dentate gyrus. Ablating adult neurogenesis blocks the effect of enrichment and lowers spatial information, as does the chemogenetic silencing of adult-born neurons. Both ablating neurogenesis and silencing adult-born neurons decreases the calcium activity of dentate gyrus neurons, resulting in a decreased amplitude of place-specific responses. These findings are in contrast with previous studies that suggested a predominantly inhibitory action for adult-born neurons. We propose that adult neurogenesis improves representations of space by increasing the gain of dentate gyrus neurons and thereby improving their ability to tune to spatial features. This mechanism may mediate the beneficial effects of environmental enrichment on spatial learning and memory.

Competition shapes the landscape of X-chromosome-linked genetic diversity.

X chromosome inactivation (XCI) generates clonal heterogeneity within XX individuals. Combined with sequence variation between human X chromosomes, XCI gives rise to intra-individual clonal diversity, whereby two sets of clones express mutually exclusive sequence variants present on one or the other X chromosome. Here we ask whether such clones merely co-exist or potentially interact with each other to modulate the contribution of X-linked diversity to organismal development. Focusing on X-linked coding variation in the human STAG2 gene, we show that Stag2variant clones contribute to most tissues at the expected frequencies but fail to form lymphocytes in Stag2WT Stag2variant mouse models. Unexpectedly, the absence of Stag2variant clones from the lymphoid compartment is due not solely to cell-intrinsic defects but requires continuous competition by Stag2WT clones. These findings show that interactions between epigenetically diverse clones can operate in an XX individual to shape the contribution of X-linked genetic diversity in a cell-type-specific manner.

Reelin marks cocaine-activated striatal ensembles, promotes neuronal excitability, and regulates cocaine reward.

Drugs of abuse activate defined neuronal ensembles in brain reward structures such as the nucleus accumbens (NAc), which are thought to promote the enduring synaptic, circuit, and behavioral consequences of drug exposure. While the molecular and cellular effects arising from experience with drugs like cocaine are increasingly well understood, the mechanisms that sculpt NAc ensemble participation are largely unknown. Here, we leveraged unbiased single-nucleus transcriptional profiling to identify expression of the secreted glycoprotein Reelin (encoded by the Reln gene) as a marker of cocaine-activated neuronal ensembles within the rat NAc. Multiplexed in situ detection confirmed selective expression of the immediate early gene Fos in Reln+ neurons after cocaine experience, and also revealed enrichment of Reln mRNA in Drd1 + medium spiny neurons (MSNs) in both the rat and human brain. Using a novel CRISPR interference strategy enabling selective Reln knockdown in the adult NAc, we observed altered expression of genes linked to calcium signaling, emergence of a transcriptional trajectory consistent with loss of cocaine sensitivity, and a striking decrease in MSN intrinsic excitability. At the behavioral level, loss of Reln prevented cocaine locomotor sensitization, abolished cocaine place preference memory, and decreased cocaine self-administration behavior. Together, these results identify Reelin as a critical mechanistic link between ensemble participation and cocaine-induced behavioral adaptations.

A data-driven single-cell and spatial transcriptomic map of the human prefrontal cortex.

The molecular organization of the human neocortex historically has been studied in the context of its histological layers. However, emerging spatial transcriptomic technologies have enabled unbiased identification of transcriptionally defined spatial domains that move beyond classic cytoarchitecture. We used the Visium spatial gene expression platform to generate a data-driven molecular neuroanatomical atlas across the anterior-posterior axis of the human dorsolateral prefrontal cortex. Integration with paired single-nucleus RNA-sequencing data revealed distinct cell type compositions and cell-cell interactions across spatial domains. Using PsychENCODE and publicly available data, we mapped the enrichment of cell types and genes associated with neuropsychiatric disorders to discrete spatial domains.

Could coping with your inner critic be the key to unlocking your scholarly writing?

INTRODUCTION: Though various barriers to productive writing habits exist in academia, scholarship is a critical part of faculty expectations. One barrier that has not been well addressed in the literature is the presence and interference of a negative inner critic, an internal voice or dialogue that criticizes work, halts creativity, and paralyzes writing. COMMENTARY: The purpose of this commentary is to describe the limited evidence-base and anecdotal strategies shown to support increased writing productivity by acknowledging and navigating the inner critic. With strategies such as proper identification, acknowledgment, strong mentor-mentee relationships, personifying the inner critic, embracing a growth mindset, and considering the distinct phases of writing, faculty can cope with their critical inner voice and reclaim control of their scholarly writing productivity. IMPLICATIONS: With such a heavy emphasis on writing productivity for faculty, faculty are encouraged to more formally explore and implement professional development strategies to help navigate their inner critic and bolster writing productivity.

Transcriptomic analysis of the human habenula in schizophrenia.

Pathophysiology of many neuropsychiatric disorders, including schizophrenia (SCZD), is linked to habenula (Hb) function. While pharmacotherapies and deep brain stimulation targeting the Hb are emerging as promising therapeutic treatments, little is known about the cell type-specific transcriptomic organization of the human Hb or how it is altered in SCZD. Here we define the molecular neuroanatomy of the human Hb and identify transcriptomic changes in individuals with SCZD compared to neurotypical controls. Utilizing Hb-enriched postmortem human brain tissue, we performed single nucleus RNA-sequencing (snRNA-seq; n=7 neurotypical donors) and identified 17 molecularly defined Hb cell types across 16,437 nuclei, including 3 medial and 7 lateral Hb populations, several of which were conserved between rodents and humans. Single molecule fluorescent in situ hybridization (smFISH; n=3 neurotypical donors) validated snRNA-seq Hb cell types and mapped their spatial locations. Bulk RNA-sequencing and cell type deconvolution in Hb-enriched tissue from 35 individuals with SCZD and 33 neurotypical controls yielded 45 SCZD-associated differentially expressed genes (DEGs, FDR < 0.05), with 32 (71%) unique to Hb-enriched tissue. eQTL analysis identified 717 independent SNP-gene pairs (FDR < 0.05), where either the SNP is a SCZD risk variant (16 pairs) or the gene is a SCZD DEG (7 pairs). eQTL and SCZD risk colocalization analysis identified 16 colocalized genes. These results identify topographically organized cell types with distinct molecular signatures in the human Hb and demonstrate unique genetic changes associated with SCZD, thereby providing novel molecular insights into the role of Hb in neuropsychiatric disorders. One Sentence Summary: Transcriptomic analysis of the human habenula and identification of molecular changes associated with schizophrenia risk and illness state.

Benchmark of cellular deconvolution methods using a multi-assay reference dataset from postmortem human prefrontal cortex.

Background: Cellular deconvolution of bulk RNA-sequencing (RNA-seq) data using single cell or nuclei RNA-seq (sc/snRNA-seq) reference data is an important strategy for estimating cell type composition in heterogeneous tissues, such as human brain. Computational methods for deconvolution have been developed and benchmarked against simulated data, pseudobulked sc/snRNA-seq data, or immunohistochemistry reference data. A major limitation in developing improved deconvolution algorithms has been the lack of integrated datasets with orthogonal measurements of gene expression and estimates of cell type proportions on the same tissue sample. Deconvolution algorithm performance has not yet been evaluated across different RNA extraction methods (cytosolic, nuclear, or whole cell RNA), different library preparation types (mRNA enrichment vs. ribosomal RNA depletion), or with matched single cell reference datasets. Results: A rich multi-assay dataset was generated in postmortem human dorsolateral prefrontal cortex (DLPFC) from 22 tissue blocks. Assays included spatially-resolved transcriptomics, snRNA-seq, bulk RNA-seq (across six library/extraction RNA-seq combinations), and RNAScope/Immunofluorescence (RNAScope/IF) for six broad cell types. The Mean Ratio method, implemented in the DeconvoBuddies R package, was developed for selecting cell type marker genes. Six computational deconvolution algorithms were evaluated in DLPFC and predicted cell type proportions were compared to orthogonal RNAScope/IF measurements. Conclusions: Bisque and hspe were the most accurate methods, were robust to differences in RNA library types and extractions. This multi-assay dataset showed that cell size differences, marker genes differentially quantified across RNA libraries, and cell composition variability in reference snRNA-seq impact the accuracy of current deconvolution methods.

The gene expression landscape of the human locus coeruleus revealed by single-nucleus and spatially-resolved transcriptomics.

Norepinephrine (NE) neurons in the locus coeruleus (LC) make long-range projections throughout the central nervous system, playing critical roles in arousal and mood, as well as various components of cognition including attention, learning, and memory. The LC-NE system is also implicated in multiple neurological and neuropsychiatric disorders. Importantly, LC-NE neurons are highly sensitive to degeneration in both Alzheimer's and Parkinson's disease. Despite the clinical importance of the brain region and the prominent role of LC-NE neurons in a variety of brain and behavioral functions, a detailed molecular characterization of the LC is lacking. Here, we used a combination of spatially-resolved transcriptomics and single-nucleus RNA-sequencing to characterize the molecular landscape of the LC region and the transcriptomic profile of LC-NE neurons in the human brain. We provide a freely accessible resource of these data in web-accessible and downloadable formats.

Left medial liver lobe torsion and postoperative acute gastric rupture in a 2.5-year-old male-castrated Flemish Giant rabbit (Oryctolagus cuniculus).

A 2.5-year-old male-castrated rabbit presented with acute abdominal pain, lethargy, and anorexia. Digital radiography revealed increased left-sided hepatomegaly, gastric dilation, and decreased peritoneal serosal detail. Abdominal ultrasonography identified a torsed left liver lobe, gastric dilation, and peritoneal effusion. Surgery confirmed a left medial liver lobe torsion, with subsequent lobectomy and seven days of hospitalization. The patient re-presented 2 days after discharge and suddenly died while hospitalized, with acute gastric rupture, fulminant peritonitis, and multifocal hepatic infarcts diagnosed on necropsy. We believe this is the first recorded imaging diagnosis of a left medial liver lobe torsion in a rabbit.

Isolated subependymal giant cell astrocytoma (SEGA) in the absence of clinical tuberous sclerosis: two case reports and literature review.

PURPOSE: Subependymal giant cell astrocytoma (SEGA) is a WHO grade I pediatric glioma arising in 5-15% of patients with tuberous sclerosis (TSC). Rare cases of isolated SEGA without TSC have been described. The etiology, genetic mechanisms, natural history, and response to treatment of these lesions are currently unknown. We describe two such cases of isolated SEGA with follow-up. METHODS: Retrospective review was performed at a single institution to describe the clinical course of pathology-confirmed SEGA in patients with germline testing negative for TSC mutations. RESULTS: Two cases of isolated SEGA were identified. Genetic analysis of the tumor specimen was available for one, which revealed an 18 base pair deletion in TSC1. Both cases were managed with surgical resection, one with preoperative embolization. In spite of a gross total resection, one patient experienced recurrence after three years. Treatment with an mTOR inhibitor led to a significant interval reduction of the mass on follow-up MRI. The patient tolerated the medication well for 6 years and is now off of treatment for 2 years with a stable lesion. CONCLUSION: Cases of SEGA outside of the context of TSC are exceedingly rare, with only 48 cases previously described. The genetic mechanisms and treatment response of these lesions are poorly understood. To date, these lesions appear to respond well to mTOR inhibitors and may behave similarly to SEGAs associated with TSC. However, given that experience is extremely limited, these cases should be followed long term to better understand their natural history and treatment response.

Interferons prime the endothelium for toll-like receptor-mediated thrombin generation.

BACKGROUND: Respiratory infection is associated with microvascular thrombus formation and marked elevation in cytokine levels. The role of cytokines elaborated by the pulmonary epithelium in thrombotic responses is poorly understood. OBJECTIVES: Our goal was to identify cytokines of pulmonary epithelial cell origin that enhance thrombin generation in the endothelium at concentrations equal to or less than those found in the circulation during infection. METHODS: We screened multiple cytokines produced by the pulmonary epithelium for the ability to enhance toll-like receptor (TLR)-mediated endothelial thrombin generation. Effects of cytokines on tissue factor and thrombomodulin expression, cytokine selectivity for different TLRs, and prothrombotic activity of endogenous cytokines in conditioned medium from pulmonary human epithelial cells were evaluated. RESULTS: MIP-1ß, MCP-1, IL-10, IL-6, IL-1ß, TNFα, IFNα, IFNß, and IFNγ were tested for their ability to enhance TLR3-mediated thrombin generation on endothelial cells. Only interferons (IFNs) and TNFα promoted TLR3-mediated thrombin generation at levels that circulate during infection. IFNs robustly enhanced tissue factor expression when used in conjunction with TLR agonists and reduced thrombomodulin expression in the endothelium independently of TLRs. IFNα, which is typically elevated with viral infection, only synergized with TLR3 agonists mimicking viral pathogen-associated molecular patterns. In contrast, IFNγ, which is typically observed in bacterial infection, synergized more effectively with TLR4 agonists released by bacteria. Conditioned media from inflamed pulmonary epithelial cells primed the endothelium for TLR-mediated thrombin generation. Anti-IFN type I antibodies blocked this effect, indicating that endogenous IFNs prime the endothelium for TLR-mediated thrombin generation. CONCLUSION: IFNs elaborated by the pulmonary epithelium are necessary and sufficient to enhance TLR-mediated thrombin generation.

Implications of outgroup selection in the phylogenetic inference of hominoids and fossil hominins.

Understanding the phylogenetic relationships among hominins and other hominoid species is critical to the study of human origins. However, phylogenetic inferences are dependent on both the character data and taxon sampling used. Previous studies of hominin phylogenetics have used Papio and Colobus as outgroups in their analyses; however, these extant monkeys possess many derived traits that may confound the polarities of morphological changes among living apes and hominins. Here, we consider Victoriapithecus and Ekembo as more suitable outgroups. Both Victoriapithecus and Ekembo are anatomically well known and are widely accepted as morphologically primitive stem cercopithecoid and hominoid taxa, respectively, making them more appropriate for inferring polarity for later-occurring hominoid- and hominin-focused analyses. Craniodental characters for both taxa were scored and then added to a previously published matrix of fossil hominin and extant hominoid taxa, replacing outgroups Papio and Colobus over a series of iterative analyses using both parsimony and Bayesian inference methods. Neither the addition nor replacement of outgroup taxa changed tree topology in any analysis. Importantly, however, bootstrap support values and posterior probabilities for nodes supporting their relationships generally increased compared to previous analyses. These increases were the highest at extant hominoid and basal hominin nodes, recovering the molecular ape phylogeny with considerably higher support and strengthening the inferred relationships among basal hominins. Interestingly, however, the inclusion of both extant and fossil outgroups reduced support for the crown hominid node. Our findings suggest that, in addition to improving character polarity estimation, including fossil outgroups generally strengthens confidence in relationships among extant hominoid and basal hominins.

Data-driven identification of total RNA expression genes for estimation of RNA abundance in heterogeneous cell types highlighted in brain tissue.

We define and identify a new class of control genes for next-generation sequencing called total RNA expression genes (TREGs), which correlate with total RNA abundance in cell types of different sizes and transcriptional activity. We provide a data-driven method to identify TREGs from single-cell RNA sequencing data, allowing the estimation of total amount of RNA when restricted to quantifying a limited number of genes. We demonstrate our method in postmortem human brain using multiplex single-molecule fluorescent in situ hybridization and compare candidate TREGs against classic housekeeping genes. We identify AKT3 as a top TREG across five brain regions.

The reconstructed cranium of Pierolapithecus and the evolution of the great ape face.

Pierolapithecus catalaunicus (~12 million years ago, northeastern Spain) is key to understanding the mosaic nature of hominid (great ape and human) evolution. Notably, its skeleton indicates that an orthograde (upright) body plan preceded suspensory adaptations in hominid evolution. However, there is ongoing debate about this species, partly because the sole known cranium, preserving a nearly complete face, suffers from taphonomic damage. We 1) carried out a micro computerized tomography (CT) based virtual reconstruction of the Pierolapithecus cranium, 2) assessed its morphological affinities using a series of two-dimensional (2D) and three-dimensional (3D) morphometric analyses, and 3) modeled the evolution of key aspects of ape face form. The reconstruction clarifies many aspects of the facial morphology of Pierolapithecus. Our results indicate that it is most similar to great apes (fossil and extant) in overall face shape and size and is morphologically distinct from other Middle Miocene apes. Crown great apes can be distinguished from other taxa in several facial metrics (e.g., low midfacial prognathism, relatively tall faces) and only some of these features are found in Pierolapithecus, which is most consistent with a stem (basal) hominid position. The inferred morphology at all ancestral nodes within the hominoid (ape and human) tree is closer to great apes than to hylobatids (gibbons and siamangs), which are convergent with other smaller anthropoids. Our analyses support a hominid ancestor that was distinct from all extant and fossil hominids in overall facial shape and shared many features with Pierolapithecus. This reconstructed ancestral morphotype represents a testable hypothesis that can be reevaluated as new fossils are discovered.

Variation in access and prescription of vedolizumab and ustekinumab in paediatric patients with inflammatory bowel disease: a UK-wide study.

BACKGROUND: Therapeutic options for paediatric inflammatory bowel disease (IBD) are limited, especially for younger children. Unlike in adults, vedolizumab and ustekinumab are not licensed for paediatric use in the UK. We aimed to understand the real-world access to, and use of, these therapies in the paediatric population. METHODS: We surveyed UK IBD centres to assess the incident use of vedolizumab and ustekinumab from 1 January 2021 to 31 December 2021. We collected information on funding, dose escalations and therapeutic drug monitoring. RESULTS: 18 of 21 centres responded, covering an estimated 5260 patients. One hundred and thirteen were started on vedolizumab, prescription incidence 2.2%, median prescriptions per centre was 4 (range 1-20). Considering ustekinumab, 73 patients were commenced, prescription incidence 1.4%. Median prescription per centre was 3.5 (range 1-13). Prescription rates at each centre were not predicted by patient number cared for at that centre (p=0.2). Dose escalation was common in vedolizumab (66.7% centres) and ustekinumab (55.5%).Funding strategies varied substantially, and multiple funding sources were used; 12 of 18 centres (66.7%) reported funding through routine National Health Service (NHS) England/Scottish arrangements. There was local NHS trust funding in 8 of 18 centres (44.4%). Individual funding requests (IFRs) were used in 5 of 18 (27.8%), although IFRs are reserved for patients with unique additional characteristics. Four centres were unable to achieve funding in pre-pubescent children. CONCLUSIONS: There is widespread use of vedolizumab and ustekinumab across the UK, although practice is highly variable. Access to therapy appeared to differ substantially. There is a growing disparity between international guidelines and real-world practice. Establishing early and effective therapy in all patients remains a priority.

Delayed formation of neural representations of space in aged mice.

Aging is associated with cognitive deficits, with spatial memory being very susceptible to decline. The hippocampal dentate gyrus (DG) is important for processing spatial information in the brain and is particularly vulnerable to aging, yet its sparse activity has led to difficulties in assessing changes in this area. Using in vivo two-photon calcium imaging, we compared DG neuronal activity and representations of space in young and aged mice walking on an unfamiliar treadmill. We found that calcium activity was significantly higher and less tuned to location in aged mice, resulting in decreased spatial information encoded in the DG. However, with repeated exposure to the same treadmill, both spatial tuning and information levels in aged mice became similar to young mice, while activity remained elevated. Our results show that spatial representations of novel environments are impaired in the aged hippocampus and gradually improve with increased familiarity. Moreover, while the aged DG is hyperexcitable, this does not disrupt neural representations of familiar environments.

Use of intraoperative topical antibiotics to reduce intrathecal baclofen pump surgical site infections: a single institution's experience over 24 years.

OBJECTIVE: Patients requiring intrathecal baclofen (ITB) therapy are at high risk for surgical site infections (SSIs) given their poor functional status. After years of a nominal infection rate, there was an inexplicable increase in ITB pump infections at the authors' institution and multiple investigations offered no solution. Use of intraoperative topical antibiotics is well-documented in the orthopedic literature and was considered for ITB pump insertion. In this study, the authors investigated whether intraoperative vancomycin and tobramycin powder at the ITB pump site could reduce SSIs. METHODS: Operative and infection data were collected and analyzed retrospectively to determine the efficacy of this change. Patients were stratified into three cohorts (1998-2009, 2010-2012, and 2013-2021) to better understand the trends before and after implementation of intraoperative topical antibiotics. Each cohort had similar demographics. RESULTS: One hundred fifty-four patients underwent 272 ITB pump procedures between 1998 and 2021 (131 in 1998-2009, 49 in 2010-2012, and 92 in 2013-2021) for cerebral palsy (69.5%), spastic quadriparesis due to traumatic brain injury (7.1%), anoxic brain injury (6.5%), and other causes (16.9%). Infection rates were reduced from a high of 32% in 2010-2011 to 3.8% over the last 2.5 years (p = 0.0094). There were no adverse effects from the use of topical antibiotics. CONCLUSIONS: In the setting of an intractable rise in ITB pump infections, the addition of intraoperative topical antibiotics significantly reduced postoperative infections in a high-risk population. One could appreciate a significant drop each year in the rate of infections after the institution of intraoperative topical antibiotics. The reduction in SSIs significantly improved the long-term outcomes for these patients.

Delayed formation of neural representations of space in aged mice.

Aging is associated with cognitive deficits, with spatial memory being very susceptible to decline. The hippocampal dentate gyrus (DG) is important for processing spatial information in the brain and is particularly vulnerable to aging, yet its sparse activity has led to difficulties in assessing changes in this area. Using in vivo two-photon calcium imaging, we compared DG neuronal activity and representations of space in young and aged mice walking on an unfamiliar treadmill. We found that calcium activity was significantly higher and less tuned to location in aged mice, resulting in decreased spatial information encoded in the DG. However, with repeated exposure to the same treadmill, both spatial tuning and information levels in aged mice became similar to young mice, while activity remained elevated. Our results show that spatial representations of novel environments are impaired in the aged hippocampus and gradually improve with increased familiarity. Moreover, while the aged DG is hyperexcitable, this does not disrupt neural representations of familiar environments.

Promoting continuing professional development (CPD) through a novel CPD advanced pharmacy practice experience.

BACKGROUND AND PURPOSE: The Accreditation Council for Pharmacy Education Standards 2016 and the Entrustable Professional Activities highlight the importance of continuing professional development (CPD) in pharmacy education. Furthermore, pharmacy graduates must self-direct their learning to sustain professional knowledge, skills, and practice. An advanced pharmacy practice experience (APPE) dedicated to CPD can help meet standards of pharmacy education and prepare students for a career of lifelong learning. EDUCATIONAL ACTIVITY AND SETTING: An innovative CPD APPE centered on the CPD framework and student self-directed learning was developed and implemented by three colleges of pharmacy. Students enrolled in the new CPD APPE were introduced to the CPD framework, engaged in reflection, developed personalized learning objectives, and participated in self-directed learning activities to meet identified educational needs. FINDINGS: Student performance outcomes were assessed via written reflections, portfolio documentation, and attendance records. The novel CPD rotation showed positive findings regarding student-perceived satisfaction, achievement of learning outcomes, and foundational lifelong learning habits. As soon-to-be graduates and practicing pharmacists, final-year pharmacy students are well poised to learn and apply the CPD framework and develop the skills needed to become lifelong learners. SUMMARY: Experiences across three colleges of pharmacy demonstrated that a CPD APPE is feasible, valuable, and effective to integrate comprehensive CPD training within pharmacy education. Other programs within the academy may utilize this scalable model to prepare APPE students to engage in self-directed CPD and lifelong learning as health professionals.