Citations

ABSTRACT Periprosthetic joint infection (PJI) is a devastating complication of total joint arthroplasty that can be challenging to diagnose and treat. Neutrophils are a prominent feature in PJI, with neutrophil extracellular traps (NETs) known to be present; little is known, however, about the physiology of PJI-associated NETs. Here, the proteomes of PJI and non-infectious arthroplasty failure (NIAF), analyzed using liquid chromatography tandem mass-spectrometry (LC-MS/MS) and proximity extension assay (PEA)-based profiling of fluids derived from sonication of arthroplasty components removed from patients who had undergone revision arthroplasty, were compared, with a focus on NET formation. In addition, in vitro effects of synovial fluid components on Staphylococcus aureus- induced NET formation were characterized. LC-MS/MS (PJI n = 85, NIAF n = 26) and PEA (PJI n = 95, NIAF n = 39) showed NET formation and neutrophil degranulation to be associated with PJI, with the complement cascade and platelet activation pathways identified as potential contributors. In vitro studies showed delayed NET formation in artificial synovial fluid compared to standard cell culture media conventionally used to evaluate NETs. Together, these results demonstrate the presence of NETs in PJI, identify potential pathways involved in PJI-associated NETosis, and point to an influence of synovial components on NET formation during PJI. IMPORTANCE Periprosthetic joint infection (PJI) is a major complication of joint replacement, placing a significant burden on patients and the healthcare system. Our research reveals that neutrophils and their antimicrobial “traps” are dominant features of the immune response at the infection joint site. Importantly, we found that the unique environment of the joint delays the formation of these traps, potentially contributing to the pathogenesis of PJI. By establishing a new experimental platform tailored for the infected joint, this work provides a foundational platform for probing these mechanisms and supports the development of improved diagnostic and therapeutic strategies.

Abstract Rationale Idiopathic Pulmonary Fibrosis (IPF) is characterized by chronic progressive pulmonary fibrosis and high mortality. Genetic markers, summarized into a polygenic risk score (PRS), associate with IPF in well-phenotyped research cohorts. Objectives To evaluate the performance of the PRS using real-world data from routinely captured electronic healthcare records. Methods We conducted an observational study evaluating the association of a PRS for IPF with electronic healthcare record diagnosis of IPF as well as lung transplant-free survival in four independent cohorts; the Mass General Brigham Biobank (MGBB), Mayo Clinic Biobank (MCBB), Mayo Clinic Tapestry Cohort (Tapestry), and U.K. Biobank (UKBB). We used multivariable logistic regression and multivariable Cox proportional hazards models adjusting for age, gender, and principal components of ancestry. The cohorts then underwent fixed and random effects meta-analysis. Measurements and Main Results Of 37,709; 44,195; 43,202; and 447,422 participants from MGBB, MCBB, Tapestry and UKBB respectively, 1,015 (2.7%) 2,879 (6.5%), 1,310 (3.0%), and 2,742 (0.6%) participants had an IPF diagnosis. Meta-analysis demonstrated a high-risk PRS associated with IPF diagnosis, OR 2.88 (95%CI 2.41-3.44) compared to all other individuals. A high-risk PRS also associated with the composite endpoint of mortality or lung transplant among those with an IPF diagnosis, HR 1.23(95%CI 1.11-1.35) compared to all other individuals. Conclusions A PRS can identify those at risk for an IPF diagnosis and mortality in biobank-scale data, which may have implications for clinical decisions. Further work is necessary to evaluate the utility of adding genetics in clinical settings.

Summary Long-read nanopore sequencing enables simultaneous detection of germline variation and native DNA base modifications on individual DNA molecules, providing a unique opportunity to investigate allele-specific epigenetic regulation. Here, we performed whole-genome nanopore sequencing on normal and tumor prostate tissues to characterize differential methylation, methylation entropy, and allele-specific methylation (ASM) associated with noncoding genetic variants. Genome-wide analysis identified extensive cancer-associated differentially methylated regions (DMRs), with hypermethylated DMRs significantly enriched near transcription start sites and transcriptional regulatory regions. Integration with transcriptomic datasets revealed strong inverse relationships between promoter methylation and gene expression, while 5-hydroxymethylcytosine (5hmC) levels positively correlated with transcriptional activity across gene bodies. Using fragment-level methylation patterns enabled by long-read sequencing, we further quantified methylation entropy incorporating both 5mCG and 5hmCG states. Cancer-hypermethylated DMRs exhibited markedly reduced entropy, consistent with clonal fixation of methylation states during tumor progression. Entropy profiling across chromatin annotations demonstrated maximal epigenetic heterogeneity at partially modified enhancer-associated regions. To investigate cis-regulatory genetic effects, we developed a simple ASM framework (nanoASM) that can partition sequencing reads by allelic state and identifies allele-specific DMRs directly from long-read data. Compared with conventional population-level mQTL analysis, ASM demonstrated substantially improved statistical efficiency by leveraging within-individual contrasts and reducing sample-level heterogeneity. Although germline single nucleotide polymorphisms (SNPs) were largely shared between normal and tumor tissues, ASM patterns differed substantially, with tumor-associated ASM regions displaying significantly larger genomic span and stronger allelic methylation differences. Comparative analysis with TCGA prostate mQTL and GTEx prostate eQTL datasets demonstrated substantial concordance between ASM directionality and downstream transcriptional effects, particularly for variants located within DMRs and near transcription start sites. At the IRX4 prostate cancer risk locus, ASM identified an androgen-responsive regulatory domain overlapping AR ChIP-seq and H3K27ac peaks, nominating rs6885084 as a candidate functional variant. At the PSCA locus, ASM anchored by rs4736369 was associated with allele-specific methylation, chromatin activation, transcript abundance, and isoform usage. Together, these findings establish nanopore-based ASM analysis as a powerful approach for resolving functional noncoding variants and their regulatory domains they control in prostate cancer.

A novel ultrasound technique was used to assess the superior segment of the internal jugular vein (IJV) in healthy individuals, evaluating the feasibility of visualizing this vein through the parotid gland and examining positional changes. Eighty-five patients undergoing unrelated carotid ultrasound were included, excluding those with headaches, migraines, or other neurological symptoms. A transducer was placed over the parotid gland, and gray scale images—with and without diameter measurement—and color and spectral Doppler of the IJV were obtained in neutral and head turned contralaterally. Technical feasibility and quantitative measurements with mean values and standard deviations were reported. Significant positional changes were assessed if paired data were available. Successful diameter measurements were obtained in at least one side in 79% to 80% of patients and bilaterally in 75%. Mean diameters were 5.5 cm (right) and 5 cm (left). Velocities were acquired in 82% (unilaterally) and 78% (bilaterally). Mean peak systolic velocities were 42 cm/s (right) and 32 cm/s (left). With head rotation, mean velocities were 46 cm/s (right) and 38 cm/s (left). In patients with paired measurements, velocities increased by over 50% in 19% to 28% of patients ( P <0.05 for both sides). There was no significant change in the diameter when the head was turned. Visualization of the superior IJV using a parotid window and evaluation of its diameter and internal velocities are feasible with the head in both neutral position and turned contralaterally.

Abstract The maintenance of cooperative behaviors within microbial populations remains an evolutionary puzzle, particularly in spatially structured environments like those found in chronic infection. In Pseudomonas aeruginosa , quorum sensing (QS) directs the production of costly public goods that are vulnerable to exploitation by non-producing cheaters. However, the biophysical mechanisms that stabilize this cooperation in complex environments remain poorly understood. Here, we elucidate how the biophysical properties of the bacterial cell surface govern the micron-scale architecture of bacterial aggregates to promote or constrain the stability of cooperation. Using a polymer-structured growth environment, we demonstrate that cell surface hydrophobicity acts as a primary determinant of spatial organization, where hydrophilic wild-type cells assemble into “stacked” aggregates, whereas hydrophobic O-specific antigen-deficient cells form dense “clumps”. In mixed populations, distinct surface properties exhibit phase-separation-like immiscibility, segregating the cells at the micron scale. We show that this segregation directly suppresses cheater fitness by physically sequestering cooperative clusters. Specifically, hydrophobic QS-deficient cells were effectively excluded by hydrophilic QS cooperators, limiting their access to public goods. Furthermore, invasion experiments revealed that hydrophilic cells are inherently fitter, capable of unidirectionally invading populations with a hydrophobic cell surface and independent of social dynamics. Conversely, the need to gain access to nutrient resources can fine-tune these barriers, enabling hydrophilic QS-deficient cells to cluster proximally to hydrophobic cooperators. These findings establish that bacterial cell surface traits impact strongly on microbial social interactions and offer new insights into the maintenance and loss of cooperative traits in chronic infections such as those found in chronic lung infections. Significance Statement Cooperative behaviors are essential for the survival of pathogenic bacteria like Pseudomonas aeruginosa , yet they are continually threatened by “cheater” cells that exploit shared public good resources. We demonstrate that the physical properties of the cell surface, specifically hydrophobicity, act as a key architect of microbial social structure. In polymer-rich environments, differences in surface hydrophobicity drive a distinct “oil-and-water” segregation between cell populations. This micron-scale phase separation physically sequesters cooperative cells, insulating them from exploitation by cheaters. Consequently, spatial structure serves as a biophysical stabilizer of cooperation, limiting the metabolic advantages typically held by cheaters. Our findings demonstrate that microbial social evolution is governed not only by genetic strategies, but by fundamental biophysical constraints that dictate the spatial limits of exploitation.

ABSTRACT Enzalutamide and abiraterone are hormonal treatments that improve survival in metastatic castration‐resistant prostate cancer. Identifying genetic variants associated with the clearance of these drugs may aid in improved dosing and outcomes. We performed genetic association studies of enzalutamide and abiraterone oral clearance in the Alliance A031201 clinical trial. Genome‐wide genotyping was performed with the primary analysis limited to European‐descent participants. Pharmacogene metabolic phenotypes were estimated using PyPGx and Stargazer. Associations of metabolic activity groups for CYP3A4 , CYP3A5 , CYP2C19 and SLCO1B1 with enzalutamide clearance ( N = 706) and CYP3A4 , SLCO2B1 and UGT1A4 with abiraterone clearance ( N = 323) were tested by linear regression. Targeted SNP associations were assessed for abiraterone clearance at loci proximal to major metabolizing genes. Full genome‐wide association studies were performed for both sets of clearance values. No significant associations were identified between metabolic phenotypes and enzalutamide or abiraterone oral clearance SNPs in the SULT2A1 5′ flanking region were significantly associated with lower abiraterone clearance, (rs296373, minor allele frequency = 0.15, β = −0.457, p = 3.2E‐06). Liver protein and liver and adrenal gland gene expression QTL databases indicated significantly lower SULT2A1 expression patterns for individuals carrying associated alleles, likely explaining the lower abiraterone oral clearance. CYP2C8*3 was associated with higher enzalutamide clearance ( p = 0.012), but this was not significant after correction for multiple testing. This study is the first to identify the genetic association of SULT2A1, known to be involved in the metabolism of steroids in the liver and adrenal glands, with abiraterone clearance. Genetic variation in SULT2A1 may be useful to inform personalized dosing of abiraterone. ClinicalTrials.gov Identifier: NCT01949337

BACKGROUND: Familial hypercholesterolemia (FH) is an autosomal dominant genetic disorder that increases risk for premature coronary artery disease and has accessible and effective interventions. The Dutch lipid clinic network is currently the most used diagnostic criterion; however, genetic sequencing provides a definitive diagnosis of FH. The goals of this study were to determine whether germline genetic screening using exome sequencing could be used to efficiently identify individuals who were genotype positive for FH. METHODS: Participants were recruited from 3 geographically and racially diverse sites in the United States (Rochester, MN; Phoenix, AZ; and Jacksonville, FL). Participants underwent Exome+ sequencing (dba Helix, San Mateo, CA) and return of results for specific genetic findings in APOB , LDLR , or PCSK9 . A chart review was performed to collect demographics, personal, and family cardiovascular history. RESULTS: At the time of the study, 84 413 participants were enrolled in the Tapestry study. Annotation and interpretation of all variants in genes for FH resulted in the identification of 419 likely pathogenic and pathogenic variants (prevalence, 0.50%), which included 116 APOB , 298 LDLR , and 5 PCSK9 . Sixty-six percent were female, the mean body mass index was 27.3, with 12.3% reporting a history of diabetes. Hypertriglyceridemia (≥150 mg/dL) was present in 39.5% and reduced HDL (<50 mg/dL) was present in 56.7% of patients. 27.5% of patients were not on cholesterol-lowering medications, and only 10% of FH carriers were at goal low-density lipoprotein cholesterol levels. A history of coronary artery disease was reported in 22.4% of the cohort. Nearly 90% of these participants were newly diagnosed carriers of FH. Only 30.8% of confirmed genetic diagnoses of FH satisfied clinical (Dutch lipid clinic network) criteria for a diagnosis. CONCLUSIONS: Our results emphasize the need for wider utilization of germline genetic sequencing for enhanced screening and detection of individuals who have familial hypercholesterolemia. REGISTRATION: URL: https://www.clinicaltrials.gov ; Unique identifier: NCT05212428.

Objectives: To determine the feasibility of reduced-dose chest computed tomographic angiography (CTA) with convolutional neural network (CNN) denoising for detecting pulmonary arteriovenous malformations (pAVMs) in children with hereditary hemorrhagic telangiectasia (cwHHT). Methods: Fifteen cwHHT underwent a chest CTA (ie, a controlled “study” dose). Noise was inserted to simulate a quarter dose (QD) exam. Images were reconstructed using iterative reconstruction (IR) and our self-trained CNN denoising model. For each case, 3 sets of images were created: study dose (SD)+IR, QD+IR, and QD+CNN. Two thoracic radiologists independently scored each set to assess quality, spatial resolution, artifacts, and the presence of pAVMs using 4-level ordinal scales. Quantitative assessments of image quality were performed using contrast-to-noise ratios (CNRs) with comparisons made between the experimental conditions. Results: Thirteen of the 15 patients recruited with hereditary hemorrhagic telangiectasia (mean age: 9.3±4.5 y) were positive for pAVM by transthoracic contrast echocardiography. The sensitivities using QD+CNN were 0.85 and 1.00 for readers 1 and 2, respectively. This was compared with 0.69 and 0.84 using QD+IR versus 0.85 and 0.92 for SD+IR. Inter-reader agreement for pAVM detection utilizing QD+CNN was moderate and resulted in kappa=0.59 ( P =0.012). The subjective assessments for QD+CNN were comparable to the SD technique. Regression analysis of reader scores revealed improved quality in QD+CNN versus QD+IR ( P =0.001). Similarly, the QD+CNN condition demonstrated the highest CNRs. Conclusions: Reduced-dose chest CTA with CNN denoising provides a level of sensitivity comparable to standard dose CTA and high CNRs for the detection of pAVMs in cwHHT.