Search

Using a nationally representative panel survey, we examined isolation behaviors among persons in the United States who had positive SARS-CoV-2 test results during January 2021–March 2022. Compared with persons who received provider-administered results, persons with home-based results had 29% (95% CI 5%–47%) lower odds of following isolation recommendations.

Abstract Background Mineworkers in Southern Africa have the highest rates of tuberculosis (TB) among working populations in the world (The World Bank, Benefits and costs associated with reducing tuberculosis among Southern Africa’s mineworkers, 2014), making mineworkers a key population for TB program efforts. The current evaluation aimed to characterize mineworkers and former (ex-) mineworkers, and assess knowledge, attitudes and practices related to TB and HIV care among mineworkers and healthcare workers (HCWs) in Zambia. Methods A mixed-methods evaluation of current and former (ex-) mineworkers and HCWs was conducted in the Copperbelt and North-Western provinces, Zambia. Knowledge, attitudes and practices (KAPs) related to TB care and policies were assessed using a structured survey. Focus Group Discussions (FGDs) were conducted with current and ex-mineworkers to understand perceptions, practices, and barriers related to accessing healthcare for TB. Results Overall, 2,792 mineworkers and 94 HCWs completed the KAP survey, and 206 (171 current, 71 ex-) mineworkers participated in FGDs. Mineworkers and ex-mineworkers were knowledgeable about TB symptoms (cough; 94%), transmission (81.7%) and treatment (99.2%). Yet, barriers to seeking care were evident with 30% of mineworkers experiencing cough, and 19% reporting 2 or more TB symptoms at the time of the survey. The majority of mineworkers (70.9%) were aware of policies barring persons from working after a diagnosis of TB, and themes from FGDs and HCW comments (n = 32/62; 51.6%) recognized fear of job loss as a critical barrier to providing timely screening and appropriate care for TB among mineworkers. The majority (76.9%) of mineworkers indicated they would not disclose their TB status to their supervisor, but would be willing to share their diagnosis with their spouse (73.8%). Conclusion Fear of job loss, driven by governmental policy and mistrust in mining companies, is a major barrier to healthcare access for TB among mineworkers in Zambia. As a result of these findings, the government policy prohibiting persons from working in the mines following TB disease is being repealed. However, major reforms are urgently needed to mitigate TB among mineworkers, including ensuring the rights of mineworkers and their communities to healthy living and working environments, improved social responsibility of mining companies, and facilitating choice and access to affordable, timely, and high-quality healthcare services.

Abstract Mycobacterium tuberculosis transmission dynamics in high-burden settings are poorly understood. Growing evidence suggests transmission may be characterized by extensive individual heterogeneity in secondary cases (i.e., superspreading), yet the degree and influence of such heterogeneity is largely unknown and unmeasured in high burden-settings. We conducted a prospective, population-based molecular epidemiology study of TB transmission in both an urban and rural setting of Botswana, one of the highest TB burden countries in the world. We used these empirical data to fit two mathematical models (urban and rural) that jointly quantified both the effective reproductive number, $$R$$ R , and the propensity for superspreading in each population. We found both urban and rural populations were characterized by a high degree of individual heterogeneity, however such heterogeneity disproportionately impacted the rural population: 99% of secondary transmission was attributed to only 19% of infectious cases in the rural population compared to 60% in the urban population and the median number of incident cases until the first outbreak of 30 cases was only 32 for the rural model compared to 791 in the urban model. These findings suggest individual heterogeneity plays a critical role shaping local TB epidemiology within subpopulations.

Abstract Background Conventional, non-specific preventive migraine treatments often demonstrate low rates of treatment persistence due to poor efficacy or tolerability. Effective, well-tolerated preventive treatments are needed to reduce migraine symptoms, improve function, and enhance quality of life. Atogepant is a migraine-specific oral calcitonin gene–related peptide receptor antagonist that is indicated for the preventive treatment of migraine in adults. This analysis evaluated the safety and tolerability profile of atogepant for the preventive treatment of migraine, including adverse events (AEs) of interest, such as constipation, nausea, hepatic safety, weight changes, and cardiac disorders. Methods This post hoc analysis was performed using data pooled from 2 (12-week) randomized, double-blind, placebo-controlled trials (RCTs) and 2 (40- and 52-week) open-label long-term safety (LTS) trials of oral atogepant for episodic migraine (EM). Results The safety population included 1550 participants from the pooled RCTs (atogepant, n = 1142; placebo, n = 408) and 1424 participants from the pooled LTS trials (atogepant, n = 1228; standard care [SC], n = 196). In total, 643/1142 (56.3%) atogepant participants and 218/408 (53.4%) placebo participants experienced ≥ 1 treatment-emergent AEs (TEAEs) in the RCTs. In the LTS trials, 792/1228 (64.5%) of atogepant participants and 154/196 (78.6%) of SC participants experienced ≥ 1 TEAEs. The most commonly reported TEAEs (≥ 5%) in participants who received atogepant once daily were upper respiratory tract infection (5.3% in RCTs, 7.7% in LTS trials), constipation (6.1% in RCTs, 5.0% in LTS trials), nausea (6.6% in RCTs, 4.6% in LTS trials), and urinary tract infection (3.4% in RCTs, 5.2% in LTS trials). Additionally, weight loss appeared to be dose- and duration-dependent. Most TEAEs were considered unrelated to study drug and few led to discontinuation. Conclusions Overall, atogepant is safe and well tolerated in pooled RCTs and LTS trials for the preventive treatment of EM in adults. Trial registration ClinicalTrials.gov identifiers: NCT02848326 (MD-01), NCT03777059 (ADVANCE), NCT03700320 (study 302), NCT03939312 (study 309). Graphical Abstract

Genome-wide association studies have identified a locus on chromosome 10q22, where many co-inherited single nucleotide polymorphisms (SNPs) are associated with atrial fibrillation (AF). This study seeks to identify the impact of this locus on gene expression at the transcript isoform level in human left atria and to gain insight into potential causal variants. Bulk RNA sequencing was analyzed to identify myozenin 1 (MYOZ1) and synaptopodin 2-like (SYNPO2L) transcript isoforms and the association of common SNPs in this region with transcript isoform expression levels. Chromatin marks were used to suggest candidate regulatory SNPs in this region. Protein amino acid changes were examined for predicted functional consequences. Transfection of MYOZ1 and two SYNPO2L isoforms were performed to localize their encoded proteins in cardiomyocytes derived from stem cells. We identified one MYOZ1 transcript isoform and four SYNPO2L transcript isoforms, two of which encode proteins, while the other two encode long noncoding RNAs (lncRNAs). The risk allele of the strongest AF susceptibility SNP on chromosome 10q22 is associated with decreased MYOZ1 expression and increased expression of the two SNYPO2L lncRNA isoforms. There are many SNPs co-inherited with the top AF-associated SNP due to linkage disequilibrium (LD), including rs11000728, which we propose as the MYOZ1 regulatory SNP, confirmed by reporter gene transfection. In addition, this LD block includes three missense SNPs in the SYNPO2L gene, with the minor protective haplotype predicted to be detrimental to protein function. MYOZ1 and both protein isoforms of SYNPO2L were localized to the sarcomere. This is a complex locus with the potential for several SNPs in a haplotype to alter AF susceptibility by opposing effects on MYOZ1 and SYNPO2L lncRNA expression, along with effects on SYNPO2L protein function.

Recent genome-wide association studies (GWAS) have identified a link between single-nucleotide polymorphisms (SNPs) near the MBOAT7 gene and advanced liver diseases. Specifically, the common MBOAT7 variant (rs641738) associated with reduced MBOAT7 expression is implicated in non-alcoholic fatty liver disease (NAFLD), alcohol-associated liver disease (ALD), and liver fibrosis. However, the precise mechanism underlying MBOAT7-driven liver disease progression remains elusive. Previously, we identified MBOAT7-driven acylation of lysophosphatidylinositol lipids as key mechanism suppressing the progression of NAFLD (Gwag et al., 2019). Here, we show that MBOAT7 loss of function promotes ALD via reorganization of lysosomal lipid homeostasis. Circulating levels of MBOAT7 metabolic products are significantly reduced in heavy drinkers compared to healthy controls. Hepatocyte- (Mboat7-HSKO), but not myeloid-specific (Mboat7-MSKO), deletion of Mboat7 exacerbates ethanol-induced liver injury. Lipidomic profiling reveals a reorganization of the hepatic lipidome in Mboat7-HSKO mice, characterized by increased endosomal/lysosomal lipids. Ethanol-exposed Mboat7-HSKO mice exhibit dysregulated autophagic flux and lysosomal biogenesis, associated with impaired transcription factor EB-mediated lysosomal biogenesis and autophagosome accumulation. This study provides mechanistic insights into how MBOAT7 influences ALD progression through dysregulation of lysosomal biogenesis and autophagic flux, highlighting hepatocyte-specific MBOAT7 loss as a key driver of ethanol-induced liver injury.

Background Pyroptosis executor GsdmD (gasdermin D) promotes atherosclerosis in mice and humans. Disulfiram was recently shown to potently inhibit GsdmD, but the in vivo efficacy and mechanism of disulfiram's antiatherosclerotic activity is yet to be explored. Methods and Results We used human/mouse macrophages, endothelial cells, and smooth muscle cells and a hyperlipidemic mouse model of atherosclerosis to determine disulfiram antiatherosclerotic efficacy and mechanism. The effects of disulfiram on several atheroprotective pathways such as autophagy, efferocytosis, phagocytosis, and gut microbiota were determined. Atomic force microscopy was used to determine the effects of disulfiram on the biophysical properties of the plasma membrane of macrophages. Disulfiram‐fed hyperlipidemic apolipoprotein E−/− mice showed significantly reduced interleukin‐1β release upon in vivo Nlrp3 (NLR family pyrin domain containing 3) inflammasome activation. Disulfiram‐fed mice showed smaller atherosclerotic lesions (~27% and 29% reduction in males and females, respectively) and necrotic core areas (~50% and 46% reduction in males and females, respectively). Disulfiram induced autophagy in macrophages, smooth muscle cells, endothelial cells, hepatocytes/liver, and atherosclerotic plaques. Disulfiram modulated other atheroprotective pathways (eg, efferocytosis, phagocytosis) and gut microbiota. Disulfiram‐treated macrophages showed enhanced phagocytosis/efferocytosis, with the mechanism being a marked increase in cell‐surface expression of efferocytic receptor MerTK. Atomic force microscopy analysis revealed altered biophysical properties of disulfiram‐treated macrophages, showing increased order‐state of plasma membrane and increased adhesion strength. Furthermore, 16sRNA sequencing of disulfiram‐fed hyperlipidemic mice showed highly significant enrichment in atheroprotective gut microbiota Akkermansia and a reduction in atherogenic Romboutsia species. Conclusions Taken together, our data show that disulfiram can simultaneously modulate several atheroprotective pathways in a GsdmD‐dependent as well as GsdmD‐independent manner.

Summary: Continuing emergence of variants of concern resulting in reduced SARS-CoV-2 vaccine efficacy necessitates additional prevention strategies. The structure of VLPCOV-01, a lipid nanoparticle-encapsulated, self-amplifying RNA COVID-19 vaccine with a comparable immune response to BNT162b2, was revised by incorporating a modified base, 5-methylcytosine, to reduce reactogenicity, and an updated receptor-binding domain derived from the Brazil (gamma) variant. Interim analyses of a phase 1 dose-escalation booster vaccination study with the resulting construct, VLPCOV-02, in healthy, previously vaccinated Japanese individuals (N = 96) are reported (jRCT2051230005). A dose-related increase in solicited local and systemic adverse events was observed, which were generally rated mild or moderate. The most commonly occurring events were tenderness, pain, fatigue, and myalgia. Serum SARS-CoV-2 immunoglobulin titers increased during the 4 weeks post-immunization. VLPCOV-02 demonstrated a favorable safety profile compared with VLPCOV-01, with reduced adverse events and fewer fever events at an equivalent dose. These findings support further study of VLPCOV-02.

Abstract Deterministic earthquake prediction remains elusive, but time‐dependent probabilistic seismicity forecasting seems within reach thanks to the development of physics‐based models relating seismicity to stress changes. Difficulties include constraining the earthquake nucleation model and fault initial stress state. Here, we analyze induced earthquakes from the Groningen gas field, where production is strongly seasonal, and seismicity began 3 decades after production started. We use the seismicity response to stress variations to constrain the earthquake nucleation process and calibrate models for time‐dependent forecasting of induced earthquakes. Remarkable agreements of modeled and observed seismicity are obtained when we consider (a) the initial strength excess, (b) the finite duration of earthquake nucleation, and (c) the seasonal variations of gas production. We propose a novel metric to quantify the nucleation model's ability to capture the damped amplitude and the phase of the seismicity response to short‐timescale (seasonal) stress variations which allows further tightening the model's parameters.

Abstract The RD‐X19 is an investigational, handheld medical device precisely engineered to emit blue light through the oral cavity to target the oropharynx and surrounding tissues. At doses shown to be noncytotoxic in an in vitro three‐dimensional human epithelial tissue model, the monochromatic visible light delivered by RD‐X19 results in light‐initiated expression of immune stimulating cytokines IL‐1α and IL‐1β, with corresponding inhibition of severe acute respiratory syndrome‐coronavirus 2 (SARS‐CoV‐2) replication. A single exposure of 425 nm blue light at 60 J/cm2 led to greater than 99% reductions against all SARS‐CoV‐2 strains tested in vitro, including the more transmissible (Alpha) and immune evasive (Beta) variants. These preclinical findings along with other studies led to a randomized, double‐blind, sham‐controlled early feasibility study using the investigational device as a treatment for outpatients with mild to moderate coronavirus disease 2019 (COVID‐19). The study enrolled 31 subjects with a positive SARS‐CoV‐2 antigen test and at least two moderate COVID‐19 signs and symptoms at baseline. Subjects were randomized 2:1 (RD‐X19: sham) and treated twice daily for 4 days. Efficacy outcome measures included assessments of SARS‐CoV‐2 saliva viral load and clinical assessments of COVID‐19. There were no local application site reactions and no device‐related adverse events. At the end of the study (day 8), the mean change in log10 viral load was −3.29 for RD‐X19 and −1.81 for sham, demonstrating a treatment benefit of −1.48 logs (95% confidence internal, −2.88 to −0.071, nominal p = 0.040). Among the clinical outcome measures, differences between RD‐X19 and sham were also observed, with a 57‐h reduction of median time to sustained resolution of COVID‐19 signs and symptoms (log rank test, nominal p = 0.044).

Abstract In-stent restenosis (ISR) complicates revascularization in the coronary and peripheral arteries. Apolipoprotein A1 (apoA1), the principal protein component of HDL possesses inherent anti-atherosclerotic and anti-restenotic properties. These beneficial traits are lost when wild type apoA1(WT) is subjected to oxidative modifications. We investigated whether local delivery of adeno-associated viral (AAV) vectors expressing oxidation-resistant apoA1(4WF) preserves apoA1 functionality. The efflux of 3H-cholesterol from macrophages to the media conditioned by endogenously produced apoA1(4WF) was 2.1-fold higher than for apoA1(WT) conditioned media in the presence of hypochlorous acid emulating conditions of oxidative stress. The proliferation of apoA1(WT)- and apoA1(4FW)-transduced rat aortic smooth muscle cells (SMC) was inhibited by 66% ± 10% and 65% ± 11%, respectively, in comparison with non-transduced SMC (p

Objective: To identify radiomic and clinical features associated with post-ablation recurrence of AF, given that cardiac morphologic changes are associated with persistent atrial fibrillation (AF), and initiating triggers of AF often arise from the pulmonary veins which are targeted in ablation. Methods: Subjects with pre-ablation contrast CT scans prior to first-time catheter ablation for AF between 2014–2016 were retrospectively identified. A training dataset (D1) was constructed from left atrial and pulmonary vein morphometric features extracted from equal numbers of consecutively included subjects with and without AF recurrence determined at 1 year. The top-performing combination of feature selection and classifier methods based on C-statistic was evaluated on a validation dataset (D2), composed of subjects retrospectively identified between 2005–2010. Clinical models ( $\text{M}_{\mathrm {C}}$ ) were similarly evaluated and compared to radiomic ( $\text{M}_{\mathrm {R}}$ ) and radiomic-clinical models ( $\text{M}_{\mathrm {RC}}$ ), each independently validated on D2. Results: Of 150 subjects in D1, 108 received radiofrequency ablation and 42 received cryoballoon. Radiomic features of recurrence included greater right carina angle, reduced anterior-posterior atrial diameter, greater atrial volume normalized to height, and steeper right inferior pulmonary vein angle. Clinical features predicting recurrence included older age, greater BMI, hypertension, and warfarin use; apixaban use was associated with reduced recurrence. AF recurrence was predicted with radio-frequency ablation models on D2 subjects with C-statistics of 0.68, 0.63, and 0.70 for radiomic, clinical, and combined feature models, though these were not prognostic in patients treated with cryoballoon. Conclusions: Pulmonary vein morphology associated with increased likelihood of AF recurrence within 1 year of catheter ablation was identified on cardiac CT. Significance: Radiomic and clinical features-based predictive models may assist in identifying atrial fibrillation ablation candidates with greatest likelihood of successful outcome.

Introduction Glioblastoma is the most common malignant primary brain tumour with a median overall survival of 12–15 months (range 6–17 months), even with maximal treatment involving debulking neurosurgery and adjuvant concomitant chemoradiotherapy. The use of postoperative imaging to detect progression is of high importance to clinicians and patients, but currently, the optimal follow-up schedule is yet to be defined. It is also unclear how adhering to National Institute for Health and Care Excellence (NICE) guidelines—which are based on general consensus rather than evidence—affects patient outcomes such as progression-free and overall survival. The primary aim of this study is to assess MRI monitoring practice after surgery for glioblastoma, and to evaluate its association with patient outcomes.Methods and analysis ImagiNg Timing aftER surgery for glioblastoma: an eVALuation of practice in Great Britain and Ireland is a retrospective multicentre study that will include 450 patients with an operated glioblastoma, treated with any adjuvant therapy regimen in the UK and Ireland. Adult patients ≥18 years diagnosed with glioblastoma and undergoing surgery between 1 August 2018 and 1 February 2019 will be included. Clinical and radiological scanning data will be collected until the date of death or date of last known follow-up. Anonymised data will be uploaded to an online Castor database. Adherence to NICE guidelines and the effect of being concordant with NICE guidelines will be identified using descriptive statistics and Kaplan-Meier survival analysis.Ethics and dissemination Each participating centre is required to gain local institutional approval for data collection and sharing. Formal ethical approval is not required since this is a service evaluation. Results of the study will be reported through peer-reviewed presentations and articles, and will be disseminated to participating centres, patients and the public.

Abstract We have previously shown that the DBA/2J versus AKR/J mouse strain is associated with decreased autophagy-mediated lysosomal hydrolysis of cholesterol esters. Our objective was to determine differences in lysosome function in AKR/J and DBA/2J macrophages, and identify the responsible genes. Using a novel dual-labeled indicator of lysosome function, DBA/2J versus AKR/J bone marrow derived macrophages had significantly decreased lysosome function. We performed quantitative trait loci mapping of lysosome function in bone marrow macrophages from an AKR/J × DBA/2J strain intercross. Four distinct lysosome function loci were identified, which we named macrophage lysosome function modifier (Mlfm) Mlfm1 through Mlfm4. The strongest locus Mlfm1 harbors the Gpnmb gene, which has been shown to recruit autophagy protein light chain 3 to autophagosomes for lysosome fusion. The parental DBA/2J strain has a nonsense variant in Gpnmb. siRNA knockdown of Gpnmb in AKR/J macrophages decreased lysosome function, and Gpnmb deletion through CRISP/Cas9 editing in RAW 264.7 mouse macrophages also demonstrated a similar result. Furthermore, a DBA/2 substrain, called DBA/2J-Gpnmb+/SjJ, contains the wildtype Gpnmb gene, and macrophages from this Gpnmb-preserved DBA/2 substrain exhibited recovered lysosome function. In conclusion, we identified Gpnmb as a causal modifier gene of lysosome function in this strain pair.

Objective: Hemifacial atrophy (HFA) is a rare disorder characterized by progressive unilateral wasting facial soft tissue, muscle, and/or bone. Trigeminal nerve abnormalities may contribute to or result from disease pathophysiology. We aimed to gain further insights into the role of trigeminal pathophysiology along the HFA severity spectrum. Methods: A systematic literature review was performed according to PRISMA standards. Retrospective cases of HFA from the literature and Mayo Clinic EMG database were pooled for descriptive and semi-quantitative analysis. Results: Overall, 13 total HFA patients were identified through literature and database reviews. Trigeminal nerve testing was abnormal in 9/13 (69%), exclusively in moderate-severe cases. Abnormalities suggested a peripheral (7/9, 78%) or mixed central/peripheral (2/9, 22%) localization. Trigeminal nerve abnormalities were not identified in any of the 4 cases with mild disease severity. Conclusion: Moderate to severe cases of HFA were associated with electrophysiological trigeminal abnormalities. No abnormalities were seen in mild cases of HFA. Significance: Trigeminal nerve electrophysiology may serve as a biomarker of moderate-severe disease progression, likely reflecting the consequences of progressive soft tissue atrophy.

Here the authors show that a murine norovirus causes self-resolving diarrhea in the absence of systemic disease in wild-type neonatal mice and show that lymphocytes affect disease in a dual manner. The data suggests that neonatal mice are a promising model for further studies.

Objective: The most common kidney cancer, clear cell renal cell carcinoma (ccRCC), is closely associated with obesity. The “clear cell” variant of RCC gets its name from the large lipid droplets that accumulate in the tumor cells. Although renal lipid metabolism is altered in ccRCC, the mechanisms and lipids driving this are not well understood. Methods: We used shotgun lipidomics in human ccRCC tumors and matched normal adjacent renal tissue. To assess MBOAT7s gene expression across tumor severity, we examined histologically graded human ccRCC samples. We then utilized genome editing in ccRCC cell lines to understand the role of MBOAT7 in ccRCC progression. Results: We identified a lipid signature for ccRCC that includes an increase in arachidonic acid-enriched phosphatidylinositols (AA-PI). In parallel, we found that ccRCC tumors have increased expression of acyltransferase enzyme membrane bound O-acyltransferase domain containing 7 (MBOAT7) that contributes to AA-PI synthesis. In ccRCC patients, MBOAT7 expression increases with tumor grade, and increased MBOAT7 expression correlates with poor survival. Genetic deletion of MBOAT7 in ccRCC cells decreases proliferation and induces cell cycle arrest, and MBOAT7−/− cells fail to form tumors in vivo. RNAseq of MBOAT7−/− cells identified alterations in cell migration and extracellular matrix organization that were functionally validated in migration assays. Conclusions: This study highlights the accumulation of AA-PI in ccRCC and demonstrates a novel way to decrease the AA-PI pool in ccRCC by limiting MBOAT7. Our data reveal that metastatic ccRCC is associated with altered AA-PI metabolism and identify MBOAT7 as a novel target in advanced ccRCC. Keywords: Clear cell renal carcinoma, Lipid, Phosphatidylinositol, Metabolism

Background Transgenic overexpression of apolipoprotein A-I (apoA1) has been shown to delay atherosclerosis lesion progression and promote lesion regression in mouse models; however, apoA1 is subject to oxidation by myeloperoxidase (MPO) and loss of function. The activity of oxidant resistant human apoA1 was compared to unmodified human apoA1 in mouse models of atherosclerosis progression and regression. Methods and results Human apoA1 and the MPO oxidant resistant 4WF isoform transgenic mice were bred to LDL receptor deficient (LDLr KO) mice and fed a western-type diet. High level expression of these human apoA1 isoforms did not lead to increased HDL-cholesterol levels on the LDLr KO background. In males and females, lesion progression was studied over time, and both apoA1 and 4WF transgenic mice vs. LDLr KO mice had significant and similar delayed lesion progression and reduced non-HDL cholesterol. Using time points with equivalent lesion areas, lesion regression was initiated by feeding the mice a low-fat control diet containing a microsomal triglyceride transfer protein inhibitor for 7 weeks. Lesions regressed more in the male apoA1 and 4WF transgenics vs. the LDLr KO, but the 4WF isoform was not superior to the unmodified isoform in promoting lesion regression. Conclusions Both human apoA1 and the 4WF MPO oxidant resistant apoA1 isoform delayed lesion progression and promoted lesion regression in LDLr KO mice, with more pronounced effects in males than females; moreover, the 4WF isoform functioned similarly to the unmodified human apoA1 isoform.

Quantitative trait locus mapping for interleukin-1β release after inflammasome priming and activation was performed on bone-marrow-derived macrophages (BMDM) from an AKRxDBA/2 mouse strain intercross. The strongest associated locus mapped very close to the Pycard gene on chromosome 7, which codes for the inflammasome adaptor protein apoptosis-associated speck-like protein containing a CARD (ASC). The DBA/2 and AKR Pycard genes only differ at a single-nucleotide polymorphism (SNP) in their 3’ untranslated region (UTR). DBA/2 vs. AKR BMDM had increased levels of Pycard mRNA expression and ASC protein, and increased inflammasome speck formation, which was associated with increased Pycard mRNA stability without an increased transcription rate. CRISPR/Cas9 gene editing was performed on DBA/2 embryonic stem cells to change the Pycard 3’UTR SNP from the DBA/2 to the AKR allele. This single base change significantly reduced Pycard expression and inflammasome activity after cells were differentiated into macrophages due to reduced Pycard mRNA stability.

Abstract Background The Royal College of Radiologists (RCR) and General Medical Council (GMC) encourage learning from mistakes. But negative feedback can be a demoralising process with adverse implications for staff morale, clinical engagement, team working and perhaps even patient outcomes. We first reviewed the literature regarding positive feedback and teamworking. We wanted to see if we could reconcile our guidance to review and learn from mistakes with evidence that positive interactions had a better effect on teamworking and outcomes than negative interactions. We then aimed to review and categorise the over 600 (mainly discrepancy) cases discussed in our educational cases meeting into educational ‘themes’. Finally, we explored whether we could use these educational themes to deliver the same teaching points in a more positive way. Methods and results The attendance records, programmes and educational cases from 30 consecutive bimonthly meetings between 2011 and 2017 were prospectively collated and retrospectively analysed. Six hundred and thirty-two cases were collated over the study period where 76% of the cases submitted were discrepancies, or perceived errors. Eight percent were ‘good spots’ where examples of good calls, excellent reporting, exemplary practice or subtle findings that were successfully reported. Eight percent were educational cases in which no mistake had been made. The remaining 7% included procedural complications or system errors. Conclusion By analysing the pattern of discrepancies in a department and delivering the teaching in a less negative way, the ‘lead’ of clinical errors can be turned in to the ‘gold’ of useful educational tools. Interrogating the whole database periodically can enable a more constructive, wider view of the meeting itself, highlight recurrent deficiencies in practice, and point to where the need for continuing medical training is greatest. Three ways in which our department have utilised this material are outlined: the use of ‘good spots’, arrangement of targeted teaching and production of specialist educational material. These techniques can all contribute to a more positive learning experience with the emphasis on acknowledging and celebrating excellence (ACE).

Pericardial effusions (PEFs) are often missed on Computed Tomography (CT), which particularly affects the outcome of patients presenting with hemodynamic compromise. An automatic PEF detection, segmentation, and classification tool would expedite and improve CT based PEF diagnosis; 258 CTs with (206 with simple PEF, 52 with hemopericardium) and without PEF (each 134 with contrast, 124 non-enhanced) were identified using the radiology report (01/2016–01/2021). PEF were manually 3D-segmented. A deep convolutional neural network (nnU-Net) was trained on 316 cases and separately tested on the remaining 200 and 22 external post-mortem CTs. Inter-reader variability was tested on 40 CTs. PEF classification utilized the median Hounsfield unit from each prediction. The sensitivity and specificity for PEF detection was 97% (95% CI 91.48–99.38%) and 100.00% (95% CI 96.38–100.00%) and 89.74% and 83.61% for diagnosing hemopericardium (AUC 0.944, 95% CI 0.904–0.984). Model performance (Dice coefficient: 0.75 ± 0.01) was non-inferior to inter-reader (0.69 ± 0.02) and was unaffected by contrast administration nor alternative chest pathology (p > 0.05). External dataset testing yielded similar results. Our model reliably detects, segments, and classifies PEF on CT in a complex dataset, potentially serving as an alert tool whilst enhancing report quality. The model and corresponding datasets are publicly available.

Background: SARS-CoV-2 enters cells by binding of its spike protein to angiotensin-converting enzyme 2 (ACE2). Angiotensin-converting enzyme inhibitors (ACEIs) or angiotensin II receptor blockers (ARBs) have been reported to increase ACE2 expression in animal models, and worse outcomes are reported in patients with co-morbidities commonly treated with these agents, leading to controversy during the COVID-19 pandemic over whether these drugs might be helpful or harmful. Methods: : Animal, in vitro and clinical data relevant to the biology of the renin-angiotensin system (RAS), its interaction with the kallikrein-kinin system (KKS) and SARS-CoV-2, and clinical studies were reviewed. Findings and Interpretation: SARS-CoV-2 hijacks ACE2to invade and damage cells, downregulating ACE2, reducing its protective effects and exacerbating injurious Ang II effects. However, retrospective observational studies do not show higher risk of infection with ACEI or ARB use. Nevertheless, study of the RAS and KKS in the setting of coronaviral infection may yield therapeutic targets.

Background: The Systemic Therapy for At Risk Teens (START) trial is a randomised controlled trial of multisystemic therapy (MST) compared with management as usual (MAU). The present study reports on long-term follow-up of the trial (to 60 months). Objectives: The primary objective was to compare MST and MAU for the proportion of young people in each group with criminal convictions up to 60 months post baseline. Secondary outcomes included group comparisons of psychological and behavioural factors. An economic analysis was carried out to determine the cost-effectiveness of MST compared with MAU. Two qualitative studies were conducted to better understand the subjective experiences of the participants. Design: Primary outcomes (collected up to 60 months) were collected using a centralised police database. Secondary outcomes were evaluated using self-report questionnaires completed by both young people and parents or carers at the 24-, 36- and 48-month follow-ups. Research assistants were blind to treatment allocation. Setting: Participants were recruited from participating MST sites in nine areas of England. Secondary outcomes were typically collected within the family home. Participants: A total of 684 families were recruited into the START trial and allocated randomly to a treatment group. Of these, 487 remained in the second phase of the trial. Young people were aged, on average, 13.8 years at baseline, with 63% male and 37% female. Interventions: MST is a manualised programme for young people exhibiting antisocial behaviour and their families that uses principles from cognitive–behavioural and family therapy to provide an individualised approach. MAU content was not prespecified, but consisted of the standard care offered to young people who met eligibility for the trial. Main outcome measures: Young people’s offending was evaluated using the Police National Computer. Secondary measures included validated self-report measures completed by both the young person and their parent or carer. The economic evaluation took a broad perspective and outcomes were assessed in terms of quality-adjusted life-years and offending. Results: No significant differences were found in the proportion of offending between the groups (hazard ratio 1.03, 95% confidence interval 0.84 to 1.26; p = 0.78). No differences were found between the groups on secondary outcome measures, with a few exceptions that did not hold up consistently across the follow-up period. The economic analysis did not find evidence to support the cost-effectiveness of MST compared with MAU. Outcomes from the qualitative studies suggest that families mostly felt positive about MST, and that MST was associated with greater maturity in young men. Limitations: Some intended evaluations were not possible to deliver. Selective attrition may have influenced the nature of the sample size. It is also unclear how representative the MAU services were of reality. Future research: Recommendations are made for the evaluation of MST in populations with more severe behavioural problems, as well as for identifying and testing new moderators. Conclusions: The results of the second phase of the START trial do not support the long-term superiority of MST to MAU, but elements of the intervention may be adapted successfully. Trial registration: Current Controlled Trials ISRCTN77132214 and London South-East REC registration number 09/H1102/55. Funding: This project was funded by the National Institute for Health Research (NIHR) Health Services and Delivery Research programme and will be published in full in Health Services and Delivery Research; Vol. 8, No. 23. See the NIHR Journals Library website for further project information.

High-density lipoprotein cholesterol (HDL-C) levels are inversely correlated with coronary heart disease (CHD) in multiple epidemiological studies, but whether HDL is causal or merely associated with CHD is unclear. Recent trials for HDL-raising drugs were either not effective in reducing CHD events or, if beneficial in reducing CHD events, were not conclusive as the findings could be attributed to the drugs’ LDL-reducing activity. Furthermore, the first large Mendelian randomization study did not causally relate HDL-C levels to decreased CHD. Thus, the hypothesis that HDL is protective against CHD has been rightfully challenged. However, subsequent Mendelian randomization studies found HDL characteristics that are causally related to decreased CHD. Many aspects of HDL structure and function, especially in reverse cholesterol transport, may be better indicators of HDL’s protective activity than simply measuring HDL-C. Cholesterol efflux capacity is associated with lower levels of prevalent and incident CHD, even after adjustment for HDL-C and apolipoprotein A-1 levels. Also, subjects with very high levels of HDL-C, including those with rare mutations that disrupt hepatic HDL uptake and reverse cholesterol transport, may be at higher risk for CHD than those with moderate levels. We describe here several cell-based and cell-free in vitro assays of HDL structure and function that may be used in clinical studies to determine which of HDL’s functions are best associated with protection against CHD. We conclude that the HDL hypothesis may need revision based on studies of HDL structure and function, but that the HDL hypothesis is not dead yet.

Exertional rhabdomyolysis is a metabolic event characterized by the release of muscle content into the circulation due to exercise-driven breakdown of skeletal muscle. Recurrent exertional rhabdomyolysis has been associated with metabolic myopathies and mitochondrial disorders, a clinically and genetically heterogeneous group of predominantly autosomal recessive, monogenic conditions. Although genetics factors are well recognized in recurrent rhabdomyolysis, the underlying causes and mechanisms of exercise-driven muscle breakdown remain unknown in a substantial number of cases. We present clinical and genetic study results from seven adult male subjects with recurrent exertional rhabdomyolysis. In all subject, whole exome sequencing identified multiple heterozygous variants in genes associated with monogenic metabolic and/or mitochondrial disorders. These variants consisted of known pathogenic and/or new likely pathogenic variants in combination with other rare deleterious alleles. The presence of heterozygous pathogenic and rare deleterious variants in multiple genes suggests an oligogenic inheritance for exertional rhabdomyolysis etiology. Our data imply that exertional rhabdomyolysis can reflect cumulative effects or synergistic interactions of deleterious variants in multiple genes that are likely to compromise muscle metabolism under the stress of exercise. Keywords: Exertional rhabdomyolysis, Metabolic disorders, Mitochondrial diseases, Multiple genetic variants, Synergistic heterozygosity, Oligogenic inheritance

Abstract Background Suprachoroidal haemorrhage is a rare complication of either medical anticoagulation treatment or intraocular surgical procedures. Suprachoroidal haemorrhages often have devastating visual outcome despite conservative and/or surgical intervention. Case presentation A patient with known Open Angle Glaucoma and Atrial Fibrillation on warfarin presents symptoms and signs suggestive acute angle closure. Examination reveals the underlying cause is a large, macula involving, spontaneous suprachoroidal haemorrhage secondary to loss of anti-coagulation control. Following aggressive medical treatment and surgical intervention, including drainage combined cataract extraction with intraocular lens implant, pars-plana vitrectomy, and external drainage of suprachoroidal haematoma, we managed to preserve the patient’s eye and some of its function. Conclusion Spontaneous suprachoroidal haemorrhages are rare complications of loss of anticoagulation control. Our case shows that aggressive treatment in selected cases can offer a relatively good outcome.

Abstract Background Frailty is a state of vulnerability to poor resolution of homeostasis after a stressor event and is strongly associated with adverse outcomes. Therefore, the assessment of frailty may be an essential part of evaluation in any healthcare encounter that might result in an escalation of care. The purpose of the study was to assess the frequency and association of frailty with clinical outcomes in patients subject to rapid response team (RRT) review. Methods In this multi-national prospective observational cohort study, centres with existing RRTs collected data over a 7-day period, with follow up of all patients at 24 h following their RRT call and at hospital discharge or 30 days following the event trigger (whichever came sooner). Investigators also collected data on the triggers and interventions provided and a bedside assessment on the level of patients’ frailty using a clinical frailty scale. Results Amongst 1133 patients, 40% were screened as frail, which was associated with older age (p

Abnormal PR interval duration is associated with risk for atrial fibrillation and heart block. Here, van Setten et al. identify 44 PR interval loci in a genome-wide association study of over 92,000 individuals and find genetic overlap with QRS duration, heart rate and atrial fibrillation.

This article discusses connections between transnational multi-faith social movements addressing climate change on a global scale with local expressions of religion and ecology in Indonesia. It connects two trends in literature on religion and ecology: 1) spatial analysis of religion and the natural environment and 2) studies of religious environmental social movements. Many studies of these movements put religious activists at the center, which suggests that they operate in a separate zone, somewhat disconnected from the local communities they aim to represent and reform. This articles argues that religious environmental movements can be better understood by placing them in the middle, as actors embedded in and shaped by overlapping global and local spaces. The article begins with a brief review of literature on religion and ecology relevant to a spatial analysis of religion and environmental social movements. It then argues that studies about religious environmental movements can be enriched by studying how movements are embedded in global and local contexts. It provides examples of how the Indonesian context (among others) has helped to shape global religious environmental movements. Next, the article presents case studies in Indonesia demonstrating how environmental activists share a dynamic relationship with their contexts, and how religious environmental discourses are co-created by local communities and religious activists. The article concludes with suggestions for further study about creative adaption to climate change at the local and global level.

Abstract Objective To determine if deficiency of CD6, a cell surface protein on lymphocytes that alters natural antibody production, increases atherosclerosis in ApoE-deficient mice fed a chow or a western-type diet. Results We compared cholesterol levels, IgM, B1a cells, and aortic root lesion areas in ApoE-deficient vs. CD6/ApoE double deficient mice. Feeding the high-fat western type diet increased all parameters, except for B1a cell numbers decreased. Sex also had an effect on many parameters with males having increased body weights, higher high density lipoprotein cholesterol, higher B1a cells, but smaller atherosclerotic lesions if chow fed mice; however, this sex effect on atherosclerosis was absent in mice fed the western-type diet. CD6 deficiency had no effect on atherosclerosis in both male and female mice on either diet. Thus, loss of CD6 on lymphocytes did not lead to expected reductions in B1a cells and protective IgM levels, and in turn did not alter atherosclerosis in mice.

Deep brain stimulation of the subthalamic nucleus (STN-DBS) is an effective treatment for Parkinson’s disease (PD). In this study, we used an interpretative phenomenological analysis to explore how 10 male people with PD experienced life after STN-DBS surgery. Two themes emerged. The first, “Healed and relieved: all that glitters is not gold,” highlights the benefits and the personal “costs” of surgery. The second, “The change within: new interpretations of the present and future unfold,” explores how patients reinterpreted their lives as individuals and members of society in the present and as they face their future. Relief, gratitude, disappointment, and the need for social support are expressed as well as a new appraisal of values and the future. STN-DBS alters the life course of people with PD, and this study provides new insight into psychological and social issues that surgery raises for the patient and their family system. These psychosocial issues should be taken into account when preparing the patient and their family for surgery or supporting them postoperatively.

Episodic status migrainosus is a previously described migraine variant where the exclusive phenotype is that of relapsing status migrainosus. The majority of cases have been reported based on retrospective chart review. A prospectively identified case fulfilling previously published criteria for episodic status migrainosus is reported, with an enduring pattern of attacks meeting criteria for status migrainosus over a 35-year period. A family history in a maternal aunt of a similar phenotype was noted. This prospectively identified case further validates the existence of episodic status migrainosus as a novel nosologic entity. The family history in our case suggests the possibility of genetic susceptibility. Perspective regarding a potential temporal continuum between acute migraine, status migrainosus, and new daily persistent headache is described. Status migrainosus may occur both as a complication of migraine and in the pure form of episodic status migrainosus.

Bariatric surgery improves glycemic control better than medical therapy; however, the effect of bariatric surgery on HDL function is not well characterized. Serum samples were available at baseline, 1-, and 5-years post procedures, for 90 patients with obesity and type 2 diabetes who were randomized to intensive medical therapy (n = 20), Roux-en-Y gastric bypass (RYGB, n = 37), or sleeve gastrectomy (SG, n = 33) as part of the STAMPEDE clinical trial. We examined serum HDL function by two independent assays, apolipoprotein A-1 exchange rate (AER) and cholesterol efflux capacity (CEC). Compared with baseline, AER was significantly higher at 5 years for participants in all treatment groups, but only increased significantly at 1 year in the RYGB and SG groups. CEC was divided into ABCA1-dependent and independent fractions, and the later was correlated with AER. ABCA1-independent CEC increased significantly only at 5 years in both surgical groups, but did not significantly change in the medical therapy group. There was no significant change in ABCA1-dependent CEC in any group. The increase in AER, but not ABCA1-independent CEC, was correlated with the reduction in body mass index and glycated hemoglobin levels among all subjects at 5 years, indicating that AER as a measure of HDL function would be a better reflection of therapy versus CEC.

Lipoproteins are supramolecular assemblies of proteins and lipids with dynamic characteristics critically linked to their biological functions as plasma lipid transporters and lipid exchangers. Among them, spherical high-density lipoproteins are the most abundant forms of high-density lipoprotein (HDL) in human plasma, active participants in reverse cholesterol transport, and associated with reduced development of atherosclerosis. Here, we employed elastic incoherent neutron scattering (EINS) and hydrogen-deuterium exchange mass spectrometry (HDX-MS) to determine the average particle dynamics and protein backbone local mobility of physiologically competent discoidal and spherical HDL particles reconstituted with human apolipoprotein A-I (apoA-I). Our EINS measurements indicated that discoidal HDL was more dynamic than spherical HDL at ambient temperatures, in agreement with their lipid-protein composition. Combining small-angle neutron scattering (SANS) with contrast variation and MS cross-linking, we showed earlier that the most likely organization of the three apolipoprotein A-I (apoA-I) chains in spherical HDL is a combination of a hairpin monomer and a helical antiparallel dimer. Here, we corroborated those findings with kinetic studies, employing hydrogen-deuterium exchange mass spectrometry (HDX-MS). Many overlapping apoA-I digested peptides exhibited bimodal HDX kinetics behavior, suggesting that apoA-I regions with the same amino acid composition located on different apoA-I chains had different conformations and/or interaction environments.

Introduction The Office for National Statistics (ONS) is transforming the way it produces statistics and how it facilitates policy and academic research. Legislative change will enable access to administrative and commercial data to underpin this transformation but only through an innovative record linkage will the full benefits of transformation be realised. Objectives and Approach Greater access to non-survey data brings with it huge opportunities for both social and economic statistics in the UK - significant improvements in timeliness and geographic fidelity. There are also risks in working with large, complex datasets which have not been compiled for statistical analysis. ONS are developing the Integrated Data Enabling Analysis and Statistics (IDEAS) initiative to provide ONS staff to linked datasets, without constructing a large database of everything. The approach involves routinely linking data to three indices (person, address, business) and assigning unique reference numbers. Data are then de-identified but can be re-joined for defined projects. Results The session will set out case studies of linkage work that either have already been taken forward using IDEAS or those which are planned. Particular emphasis will be given to how IDEAS can facilitate research across a range of statistical research topics by linking on person, address and business. By making anonymised linked data available in a safe setting the IDEAS framework also has the potential to facilitate research across government and across academic research. ONS have ambitious plans to use this approach to draw together expertise from across both sectors to be able to realise the benefits of linked where knowledge is shared on data quality, methods and previous research undertaken. Conclusion/Implications The IDEAS framework is as step change in the production of official statistics. With appropriate legal, ethical and security controls, linked data within IDEAS can be repeated and refined for different applications. The framework enables collaboration across disciplines and has the potential to be using across government and academia.

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation.

This study investigated the use of Acceptance and Commitment Therapy (ACT) as a treatment for music performance anxiety (MPA) in an uncontrolled pilot design. ACT is a newer, “third-wave” therapy that differs from previous MPA treatments, because its goal is not to reduce symptoms of MPA. Rather, ACT aims to enhance psychological flexibility in the presence of unwanted symptoms through the promotion of six core processes collectively known as the ACT “Hexaflex.” A small group of student vocalists (N = 7) from an elite choral college were recruited using objective criteria for evaluating MPA. Participants received 12 ACT sessions, and their baseline functioning served as a pre-treatment control. Treatment consisted of an orientation to ACT, identifying experientially avoidant behaviors, facilitation of Hexaflex processes, group performances in which valued behaviors were practiced in front of one another, meditations, homework, and completion of self-report measures before, during, and after treatment (at a 1- and 3-month follow-up). Improvements were observed in participants' cognitive defusion, acceptance of MPA symptoms, and psychological flexibility at post-treatment and follow-ups. Students also appeared to improve their performance quality and reduce their shame over having MPA. These results add to existing research suggesting ACT is a promising intervention for MPA, while also highlighting how vocal students may be less impaired by physical MPA symptoms.

The aim of this study was to identify structural and functional brain changes that accompanied the transition from chronic (CM; ≥ 15 headache days/month) to episodic (EM; < 15 headache days/month) migraine following prophylactic treatment with onabotulinumtoxinA. Specifically, we examined whether CM patients responsive to prophylaxis (responders; n = 11), as evidenced by a reversal in disease status (defined by at least a 50% reduction in migraine frequency and < 15 headache days/month), compared to CM patients whose migraine frequency remained unchanged (non-responders; n = 12), showed differences in cortical thickness using surface-based morphometry. We also investigated whether areas showing group differences in cortical thickness displayed altered resting-state functional connectivity (RS-FC) using seed-to-voxel analyses. Migraine characteristics measured across groups included disease duration, pain intensity and headache frequency. Patient reports’ of headache frequency over the four weeks prior to (pre-treatment) and following (post-treatment) prophylaxis were compared (post minus pre) and this measure served as the clinical endpoint that determined group assignment. All patients were scanned within two weeks of the post-treatment visit. Results revealed that responders showed significant cortical thickening in the right primary somatosensory cortex (SI) and anterior insula, and left superior temporal gyrus and pars opercularis compared to non-responders. In addition, disease duration was negatively correlated with cortical thickness in fronto-parietal and temporo-occipital regions in responders but not non-responders, with the exception of the primary motor cortex (MI) that showed the opposite pattern; disease duration was positively associated with MI cortical thickness in responders versus non-responders. Our seed-based RS-FC analyses revealed anti-correlations between the SI seed and lateral occipital (LOC) and dorsomedial prefrontal cortices in responders, whereas non-responders showed increased connectivity between the pars opercularis seed and LOC. Overall, our findings revealed distinct morphometric and functional brain changes in CM patients that reverted to episodic migraine following prophylactic treatment compared to CM patients that showed no change in disease status. Elucidating the CNS changes involved in disease reversal may be critical to discovering interventions that prevent or slow the progression of CM. Such changes may aid in the evaluation of treatments as well as provide markers for disease ‘de-chronification’.

Objective and Background: We hypothesized that evaluation scores for attending neurologists by patients and residents would parallel one another. Additionally, we hypothesized that provider productivity would be also be associated with performance evaluations by patients and residents. Methods: In a university neurology department, we collected individual Clinician and Group Consumer Assessment of Healthcare Providers and Systems patient satisfaction scores and standardized resident evaluation scores (n = 22 faculty members). We performed bivariate analysis of doctor–patient satisfaction versus resident evaluation scores. Results: Attending neurologists with higher patient satisfaction received lower resident evaluation scores ( P < .05). There seem to be disproportionate neurologists with low evaluations not meeting clinical productivity targets. Conclusion: Finding a significant inverse correlation was surprising. Perhaps what is valued by patients in their physician is not what residents value in teachers. That deserves further study. Maybe attending physicians who spend their energy on the patient experience do not have sufficient time to devote to teaching and vice versa. That neurologists with low evaluation scores appear more likely to not meet productivity targets supports this idea.

Inactivation of the von Hippel-Lindau (VHL) gene and activation of the hypoxia-inducible factor (HIF) in susceptible cells precedes formation of tumorlets and frank tumor in the epididymis of male VHL patients. We performed detailed histologic and molecular pathologic analysis of tumor-free epididymal tissues from VHL patients to obtain further insight into early epididymal tumorigenesis. Four epididymides from two VHL patients were serially sectioned to allow for three-dimensional visualization of morphologic changes. Areas of interest were genetically analyzed by tissue microdissection, immunohistochemistry for HIF and markers for mesonephric differentiation, and in situ hybridization for HIF downstream target vascular endothelial growth factor. Structural analysis of the epididymides revealed marked deviations from the regular anatomic structure resulting from impaired organogenesis. Selected efferent ductules were represented by disorganized mesonephric cells, and the maldeveloped mesonephric material was VHL-deficient by allelic deletion analysis. Furthermore, we observed maldeveloped mesonephric material near cystic structures, which were also VHL-deficient and were apparent derivatives of maldeveloped material. Finally, a subset of VHL-deficient cells was structurally integrated in regular efferent ductules; proliferation of intraductular VHL-deficient cells manifests itself as papillary growth into the ductular lumen. Furthermore, we clarify that that there is a pathogenetic continuum between microscopic tumorlets and formation of tumor. In multiple locations, three-dimensional reconstruction revealed papillary growth to extend deeply into ductular lumina, indicative of progression into early hamartoma-like neoplasia. We conclude epididymal tumorigenesis in VHL disease to occur in two distinct sequential steps: maldevelopment of VHL-deficient mesonephric cells, followed by neoplastic papillary proliferation.