Maladaptive T-Cell Metabolic Fitness in Autoimmune Diseases.

Immune surveillance and adaptive immune responses, involving continuously circulating and tissue-resident T-lymphocytes, provide host defense against infectious agents and possible malignant transformation while avoiding autoimmune tissue damage. Activation, migration, and deployment of T-cells to affected tissue sites are crucial for mounting an adaptive immune response. An effective adaptive immune defense depends on the ability of T-cells to dynamically reprogram their metabolic requirements in response to environmental cues. Inability of the T-cells to adapt to specific metabolic demands may skew cells to become either hyporesponsive (creating immunocompromised conditions) or hyperactive (causing autoimmune tissue destruction). Here, we review maladaptive T-cell metabolic fitness that can cause autoimmune diseases and discuss how T-cell metabolic programs can potentially be modulated to achieve therapeutic benefits.

Systematic assessment of long-read RNA-seq methods for transcript identification and quantification.

The Long-read RNA-Seq Genome Annotation Assessment Project (LRGASP) Consortium was formed to evaluate the effectiveness of long-read approaches for transcriptome analysis. The consortium generated over 427 million long-read sequences from cDNA and direct RNA datasets, encompassing human, mouse, and manatee species, using different protocols and sequencing platforms. These data were utilized by developers to address challenges in transcript isoform detection and quantification, as well as de novo transcript isoform identification. The study revealed that libraries with longer, more accurate sequences produce more accurate transcripts than those with increased read depth, whereas greater read depth improved quantification accuracy. In well-annotated genomes, tools based on reference sequences demonstrated the best performance. When aiming to detect rare and novel transcripts or when using reference-free approaches, incorporating additional orthogonal data and replicate samples are advised. This collaborative study offers a benchmark for current practices and provides direction for future method development in transcriptome analysis.

Long-term evolution of proliferating cells using the eVOLVER platform.

Experimental evolution using fast-growing unicellular organisms is a unique strategy for deciphering the principles and mechanisms underlying evolutionary processes as well as the architecture and wiring of basic biological functions. Over the past decade, this approach has benefited from the development of powerful systems for the continuous control of the growth of independently evolving cultures. While the first devices compatible with multiplexed experimental evolution remained challenging to implement and required constant user intervention, the recently developed eVOLVER framework represents a fully automated closed-loop system for laboratory evolution assays. However, it remained difficult to maintain and compare parallel evolving cultures in tightly controlled environments over long periods of time using eVOLVER. Furthermore, a number of tools were lacking to cope with the various issues that inevitably occur when conducting such long-term assays. Here we present a significant upgrade of the eVOLVER framework, providing major modifications of the experimental methodology, hardware and software as well as a new stand-alone protocol. Altogether, these adaptations and improvements make the eVOLVER a versatile and unparalleled set-up for long-term experimental evolution.

Long-term evolution of proliferating cells using the eVOLVER platform.

Experimental evolution using fast-growing unicellular organisms is a unique strategy for deciphering the principles and mechanisms underlying evolutionary processes as well as the architecture and wiring of basic biological functions. Over the past decade, this approach has benefited from the development of powerful systems for the continuous control of the growth of independently evolving cultures. While the first devices compatible with multiplexed experimental evolution remained challenging to implement and required constant user intervention, the recently-developed eVOLVER framework represents a fully automated closed-loop system for laboratory evolution assays. However, it remained difficult to maintain and compare parallel evolving cultures in tightly controlled environments over long periods of time using eVOLVER. Furthermore, a number of tools were lacking to cope with the various issues that inevitably occur when conducting such long-term assays. Here we present a significant upgrade of the eVOLVER framework, providing major modifications of the experimental methodology, hardware and software as well as a new standalone protocol. Altogether, these adaptations and improvements make the eVOLVER a versatile and unparalleled setup for long-term experimental evolution.

Effectiveness of Glaucoma Screening and Factors Associated with Follow-up Adherence among Glaucoma Suspects in a Safety-Net Teleretinal Screening Program.

PURPOSE: To evaluate rates and risk factors associated with follow-up adherence to in-person glaucoma evaluations and confirmed glaucoma diagnosis in glaucoma suspects identified through teleretinal diabetic retinopathy screening (TDRS). DESIGN: Retrospective cohort study SUBJECTS: Patients with diabetes identified through teleretinal screening to have large or asymmetric cup-to-disc ratios in a Los Angeles County safety-net primary care-based TDRS program. METHODS: Retrospective chart review was performed to obtain demographic and clinical information for patients with cup-to-disc ratios concerning for glaucoma on TDRS. Patients who completed an in-person follow-up appointment within 1 year of teleretinal screening were adherent. Factors associated with follow-up adherence and diagnosis of glaucoma were analyzed with chi-square and independent t tests along with multivariable logistic regressions. MAIN OUTCOME MEASURES: The proportion of patients with suspected glaucoma who adhered with in-person follow-up examination, proportion of patients with confirmed glaucoma diagnosis, and factors associated with follow-up adherence and glaucoma diagnosis. RESULTS: Eight-hundred seventeen patients with optic discs suspicious for glaucoma were included. Five-hundred thirty-four (65.4%) patients successfully completed an in-person glaucoma evaluation. Among these patients, 62.9% and 24.5% received a diagnosis of glaucoma suspect and glaucomatous optic neuropathy, respectively. Compared with patients aged < 50 years, patients aged 50 to 64 years had 1.57 times higher odds of being adherent with in-person visits (P = 0.036), whereas no difference was seen in those aged ≥ 65 years. For every $10 000 increase in the zip code median income, patients had 11% lower odds of being adherent (P = 0.031). Compared with Latino patients, Black patients had 3.52 times (P < 0.001) higher odds of having confirmed glaucoma. CONCLUSION: The majority of patients referred as glaucoma suspects on TDRS completed a follow-up examination, and nearly a quarter of those examined received a confirmed glaucoma diagnosis. Patients aged ≥ 50 and < 65 years along with those from lower-income neighborhoods were more likely to follow up for an in-person evaluation. Compared with Latino patients, Black patients had a higher risk for a confirmed glaucoma diagnosis. This demonstrates the effectiveness of glaucoma detection in a large-scale TDRS program for a safety-net patient population. FINANCIAL DISCLOSURE(S): The authors have no proprietary or commercial interest in any materials discussed in this article.

High-throughput continuous evolution of compact Cas9 variants targeting single-nucleotide-pyrimidine PAMs.

Despite the availability of Cas9 variants with varied protospacer-adjacent motif (PAM) compatibilities, some genomic loci-especially those with pyrimidine-rich PAM sequences-remain inaccessible by high-activity Cas9 proteins. Moreover, broadening PAM sequence compatibility through engineering can increase off-target activity. With directed evolution, we generated four Cas9 variants that together enable targeting of most pyrimidine-rich PAM sequences in the human genome. Using phage-assisted noncontinuous evolution and eVOLVER-supported phage-assisted continuous evolution, we evolved Nme2Cas9, a compact Cas9 variant, into variants that recognize single-nucleotide pyrimidine-PAM sequences. We developed a general selection strategy that requires functional editing with fully specified target protospacers and PAMs. We applied this selection to evolve high-activity variants eNme2-T.1, eNme2-T.2, eNme2-C and eNme2-C.NR. Variants eNme2-T.1 and eNme2-T.2 offer access to N4TN PAM sequences with comparable editing efficiencies as existing variants, while eNme2-C and eNme2-C.NR offer less restrictive PAM requirements, comparable or higher activity in a variety of human cell types and lower off-target activity at N4CN PAM sequences.

Rates and Patterns of Diagnostic Conversion from Anatomical Narrow Angle to Primary Angle-Closure Glaucoma in the United States.

PURPOSE: To assess rates of diagnostic conversion from anatomical narrow angle (ANA) to primary angle-closure glaucoma (PACG) in the United States and identify factors associated with diagnostic conversion. DESIGN: Retrospective case-control study. PARTICIPANTS: Patients diagnosed with ANA between the years 2007 and 2019 were identified based on International Classification of Diseases (ICD) codes in the Optum Clinformatics Data Mart Database. Inclusion was limited to newly diagnosed ANA, defined as the following: (1) continuous enrollment during a 2-year look back period and 6-year study period from index (first) date of ANA diagnosis; (2) diagnosis by an ophthalmologist or optometrist and record of gonioscopy; and (3) no history of intraocular pressure (IOP)-lowering drops, laser peripheral iridotomy (LPI), or intraocular surgery. METHODS: Cox proportional hazards models were developed to assess factors associated with diagnostic conversion, defined as a change in ICD code from ANA to PACG. MAIN OUTCOME MEASURES: New diagnosis of PACG within the 6-year study period recorded after an index diagnosis of ANA. RESULTS: Among 3985 patients meeting inclusion criteria, 459 (11.52%) had detected diagnostic conversion to PACG within the study period. The conversion rate was stable at 3.54% per year after the first 6 months of ANA diagnosis. In the Cox proportional hazards model, age > 70 years and early (within 6 months of ANA diagnosis) need for LPI or IOP-lowering drops were positively associated with diagnostic conversion (hazard ratio [HR] > 1.59; P < 0.02). Cataract surgery at any time and late (after 6 months of ANA diagnosis) need for IOP-lowering drops appeared protective against diagnostic conversion (HR < 0.46; P < 0.004). CONCLUSIONS: Annual risk of diagnostic conversion from ANA to PACG is relatively low overall; elderly patients are at higher risk whereas patients receiving cataract surgery are at lower risk. The utility of long-term monitoring seems low for most patients with ANA, highlighting the need for improved clinical methods to identify patients at higher risk for PACG. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Inpatient Care Team Views on Child Life Services: A Scoping Review.

CONTEXT: The utilization of Child Life Services is influenced by interprofessional collaboration and perceptions of other members of the medical team. OBJECTIVES: To summarize studies which address pediatric health care team perspectives on Child Life Services and their utilization in the hospital setting. DATA SOURCES: A comprehensive literature search was conducted with controlled vocabularies and key terms in MEDLINE, Embase, CINAHL, PsycInfo, and Web of Science. STUDY SELECTION: Primary studies published before November 2021 were screened using a predetermined set of inclusion and exclusion criteria. DATA CHARTING: Data charting was performed by 2 independent reviewers. Data extracted include baseline study characteristics, common themes, main outcomes, strengths, and limitations. Because this is not a systematic review, data from included studies was not quantitatively analyzed, but carefully summarized in the manner of a standard scoping review. RESULTS: Nine studies met criteria for inclusion. Common qualitative themes on certified child life specialists include: (1) their broad responsibilities, (2) their positive impact on patients and families, (3) challenges with interprofessional collaboration and integration, and (4) the value of educating others on their roles and responsibilities. CONCLUSIONS: Medical subject headings, controlled vocabulary, or other standardized subject headings that index literature on Child Life Services is limited. However, the existing body of literature supports the positive impact certified child life specialists have on patients and families, despite challenges with complete integration into the interdisciplinary care team. Additional research is required to fully understand and overcome these challenges in continued efforts to further drive patient and family-centered care.

Obstacles for T-lymphocytes in the tumour microenvironment: Therapeutic challenges, advances and opportunities beyond immune checkpoint.

The tumour microenvironment (TME) imposes a major obstacle to infiltrating T-lymphocytes and suppresses their function. Several immune checkpoint proteins that interfere with ligand/receptor interactions and impede T-cell anti-tumour responses have been identified. Immunotherapies that block immune checkpoints have revolutionized the treatment paradigm for many patients with advanced-stage tumours. However, metabolic constraints and soluble factors that exist within the TME exacerbate the functional exhaustion of tumour-infiltrating T-cells. Here we review these multifactorial constraints and mechanisms - elevated immunosuppressive metabolites and enzymes, nutrient insufficiency, hypoxia, increased acidity, immense amounts of extracellular ATP and adenosine, dysregulated bioenergetic and purinergic signalling, and ionic imbalance - that operate in the TME and collectively suppress T-cell function. We discuss how scientific advances could help overcome the complex TME obstacles for tumour-infiltrating T-lymphocytes, aiming to stimulate further research for developing new therapeutic strategies by harnessing the full potential of the immune system in combating cancer.

Improved transcriptome assembly using a hybrid of long and short reads with StringTie.

Short-read RNA sequencing and long-read RNA sequencing each have their strengths and weaknesses for transcriptome assembly. While short reads are highly accurate, they are rarely able to span multiple exons. Long-read technology can capture full-length transcripts, but its relatively high error rate often leads to mis-identified splice sites. Here we present a new release of StringTie that performs hybrid-read assembly. By taking advantage of the strengths of both long and short reads, hybrid-read assembly with StringTie is more accurate than long-read only or short-read only assembly, and on some datasets it can more than double the number of correctly assembled transcripts, while obtaining substantially higher precision than the long-read data assembly alone. Here we demonstrate the improved accuracy on simulated data and real data from Arabidopsis thaliana, Mus musculus, and human. We also show that hybrid-read assembly is more accurate than correcting long reads prior to assembly while also being substantially faster. StringTie is freely available as open source software at https://github.com/gpertea/stringtie.

Repeatability and Reproducibility of 4.5 by 4.5 mm Peripapillary Optical Coherence Tomography Angiography Scans in Glaucoma and Non-Glaucoma Eyes.

PRCIS: Peripapillary vessel parameters from optical coherence tomography angiography (OCTA) 4.5×4.5 mm scans in nonglaucomatous and glaucomatous eyes showed high repeatability and reproducibility, with higher reliability for commercially developed OCTA parameters compared with custom OCTA parameters. PURPOSE: The purpose of this study was to assess intrasession repeatability versus intersession reproducibility of peripapillary vessel parameters from 4.5×4.5 mm OCTA scans in nonglaucomatous eyes and glaucomatous eyes. MATERIALS AND METHODS: In a longitudinal study, peripapillary OCTA scans were quantified using research-oriented custom quantification software that calculated vessel area density (VAD) and flux and clinic-oriented commercially developed software (Cirrus 11.0) that calculated perfusion density (PD) and flux index (FI). Intrasession repeatability and intersession reproducibility were evaluated using within-eye coefficient of variation (CV W ) and intraclass correlation coefficient (ICC). RESULTS: With 127 nonglaucomatous eyes, intrasession CV W for VAD, PD, flux, and FI were 1.900%, 1.174%, 2.787%, and 1.425%, respectively. The intersession CV W were 2.039%, 1.606%, 4.053%, and 2.798%, respectively. Intrasession ICC ranged from 0.903 to 0.956, and intersession ICC ranged from 0.850 to 0.896. Among 144 glaucomatous eyes, intrasession CV W for VAD, PD, flux, and FI were: 3.841%, 1.493%, 5.009%, and 2.432%, respectively. The intersession CV W were 4.991%, 2.155%, 6.360%, and 3.458%, respectively. Intrasession ICC ranged from 0.956 to 0.969, and intersession ICC ranged from 0.918 to 0.964. CONCLUSIONS: Among nonglaucomatous and glaucomatous eyes, the majority of peripapillary OCTA vessel parameters from 4.5×4.5 mm scans had greater intrasession repeatability than intersession reproducibility. There was a greater agreement for the commercially developed quantification parameters than for their custom quantification counterparts.

Intrasession Repeatability and Intersession Reproducibility of Macular Vessel Parameters on Optical Coherence Tomography Angiography in Glaucomatous and Non-Glaucomatous Eyes.

PURPOSE: Using optical coherence tomography angiography (OCTA), this study compared intrasession repeatability versus intersession reproducibility of macular vessel parameters in glaucoma and non-glaucoma subjects. METHODS: 6 × 6 mm2 macular OCTA scans (Cirrus HD-OCT 5000) were acquired from glaucomatous and non-glaucomatous subjects as part of an observational, longitudinal study. Vessel area density (VAD) and vessel skeleton density (VSD) were calculated using research-based quantification software while perfusion density (PDZ) and vessel density (VDZ) were calculated using commercially developed software (Cirrus 11.0, Carl Zeiss Meditec). Intrasession repeatability and intersession reproducibility were determined using within-eye standard deviation (SW), within-eye coefficient of repeatability (CRW), within-eye coefficient of variation (CVW), and intraclass correlation coefficients (ICC). RESULTS: The intrasession repeatability and intersession reproducibility for macular OCTA parameters were similar to one another for both non-glaucomatous and glaucomatous eyes. Intrasession CVW from the non-glaucoma group (n = 73) was 1.097% for VAD, 1.007% for VSD, 2.980% for PDZ, and 2.714% for VDZ. Intersession CVW from the non-glaucoma group (n = 55) was 1.389% for VAD, 1.279% for VSD, 2.935% for PDZ, and 2.695% for VDZ. Intrasession CVW from the glaucoma group (n = 59) was 1.189% for VAD, 0.970% for VSD, 3.827% for PDZ, and 3.542% for VDZ. Intersession CVW from the glaucoma group (n = 45) was 1.412% for VAD, 1.132% for VSD, 3.915% for PDZ, and 3.654% for VDZ. Non-glaucomatous intrasession ICC ranged from 0.711 to 0.824, non-glaucomatous intersession ICC ranged from 0.649 to 0.762, glaucomatous intrasession ICC ranged from 0.710 to 0.853, and glaucomatous intersession ICC ranged from 0.661 to 0.827. CONCLUSIONS: Macular OCTA scans can be a useful tool in monitoring the longitudinal progression of glaucoma due to their high repeatability and reproducibility.

Sphenoethmoidal air cell sinusitis: A rare cause of recurrent optic neuritis.

Purpose: To describe the clinical presentation, imaging characteristics and management of a rare case of Onodi cell sinusitis associated with optic neuropathy. Observations: A 46-year-old male presented to the emergency department with progressive left eye vision loss over the course of ten days. The constellation of findings from this patient's history, physical, and fundoscopic exam, as well as CT and MR imaging led to the diagnosis of Onodi air cell sinusitis complicated by optic neuropathy. The patient's symptoms resolved fully and vision returned to baseline with oral antibiotics. Conclusions and Importance: The sphenoethmoidal air cell, also known as the Onodi cell, is an anatomic variant of the paranasal sinuses whose spatial relationship with important neurovascular structures carries significant clinical implications when it becomes inflamed or infected. Our case of Onodi cell sinusitis complicated by optic neuropathy demonstrates how vision loss secondary to sinusitis may resolve with oral antibiotic treatment. We additionally review the relevant anatomy, clinical course and treatment of Onodi cell pathologies.

Ischemic index and distribution of retinal capillary non-perfusion in neovascular glaucoma.

INTRODUCTION: Neovascular Glaucoma (NVG) is a condition normally caused by hypoxic posterior ocular disease, which produces angiogenic factors such as vascular endothelial growth factor (VEGF) that stimulate new vessel proliferation of the anterior segment and angle, eventually leading to angle closure, reduced outflow of aqueous humor and increased intraocular pressure. Without treatment elevated intraocular pressure can rapidly progress to loss of vision. Treatment includes addressing the intraocular pressure and reducing the ischemic drive with panretinal photocoagulation (PRP) of the ischemic retina. Recent imaging advancements allow for ultra-widefield fluorescein angiography (UWFA) which expand the amount of peripheral retina that can be evaluated for non-perfusion. Here we aim to study patterns of non-perfusion in NVG using a group of PRP naïve patients with recent onset NVG. METHODS: This study is a retrospective single-center cross-sectional study of patients seen at LAC + USC Medical Center from January 2015 to April 2020 with new onset NVG, without PRP and with UWFA completed. The percentage of ischemic index of the retina was calculated from the UWFA and evaluated in three distinct zones centered on the fovea: the posterior pole, the mid periphery, and far periphery. To increase sample size, a confirmatory group was included, with PRP allowed prior to UWFA but not before diagnosis. In addition, the time between diagnosis and UWFA was increased to 6 months. RESULTS: A total of 11 eyes met inclusion criteria for the primary group. Ischemic index was found to be 91% in the far periphery, 77% in the mid periphery, and 42% at the posterior pole. The total average ischemic index was 76%. There was a statistically significant difference between the far periphery and posterior pole and mid periphery and posterior pole. A total of 24 eyes met criteria for the confirmatory group. Ischemic index for the confirmatory group was found to be 93% in the far periphery, 75% in the mid periphery, and 35% at the posterior pole. There was a statistically significant difference between the far periphery, posterior pole and mid-periphery. CONCLUSION: This knowledge can be used to further guide treatment and understand risk for NVG.

Wedge Defects on Optical Coherence Tomography Angiography of the Peripapillary Retina in Glaucoma: Prevalence and Associated Clinical Factors.

PRCIS: Among subjects with glaucoma, wedge-shaped defects on optical coherence tomography angiography (OCTA) were associated with disc hemorrhages (DH), paracentral visual field (VF) defects, increased cup-to-disc ratio (CDR), and thinner retinal nerve fiber layer (RNFL). PURPOSE: To examine determinants of wedge defects on peripapillary OCTA in glaucoma. MATERIALS AND METHODS: A total of 278 eyes of 186 subjects with mild to severe primary open-angle glaucoma underwent 6×6 spectral-domain OCTA imaging of the superficial peripapillary retina from 2016 to 2020 at an academic practice. Wedge defects were defined as focal microvasculature loss that extends outward from the optic nerve in an arcuate, wedge shape. Logistic regression models controlling for intereye correlation identified variables significantly associated with wedge defects. Eyes with profound microvasculature loss in both hemispheres were excluded. Candidate variables included: age, sex, race or ethnicity, diabetes, hypertension, follow-up duration, baseline untreated intraocular pressure, intraocular pressure at time of imaging, DH history, paracentral VF defects, CDR, central corneal thickness, spherical equivalent, VF mean deviation, RNFL thickness, and glaucoma stage. RESULTS: Of 278 eyes, 126 (45.3%) had wedge defects in at least 1 hemisphere. In our multivariable logistic regression model, wedge defects were associated with DH history [odds ratio (OR): 3.19, 95% confidence interval (CI): 1.05-9.69, P=0.041], paracentral VF defects [OR: 4.38 (95% CI: 2.11-9.11), P<0.0001], larger CDR [OR: 1.27 (95% CI: 1.03-1.56), P=0.024, per 0.1 increase], and thinner RNFL [OR: 1.71 (95% CI: 1.25-2.34), P=0.0009, per 10 µm decrease]. CONCLUSION: DH history and paracentral VF defects were independently associated with wedge defects on OCTA, which was present in 45.3% of primary open-angle glaucoma patients. These findings may provide insight into glaucoma pathogenesis.

Racial and Sociodemographic Disparities in the Detection of Narrow Angles before Detection of Primary Angle-Closure Glaucoma in the United States.

PURPOSE: To assess the proportion of newly diagnosed cases of primary angle-closure glaucoma (PACG) with and without prior diagnosis of anatomical narrow angle (ANA) and to identify sociodemographic risk factors for late detection (PACG without prior ANA diagnosis). DESIGN: Retrospective cohort study. METHODS: One hundred two thousand six hundred seventeen patients with PACG were identified from the Optum Clinformatics Data Mart Database (2007-2019). Patients with newly diagnosed PACG met the following criteria: (1) diagnosis made by an ophthalmologist, (2) disease observable for at least 12 months before diagnosis, and (3) no history of treatment before diagnosis unless preceded by a diagnosis of ANA. Multivariate logistic regression modeling was performed to identify sociodemographic risk factors for late detection. MAIN OUTCOME MEASURES: Proportion of patients with newly diagnosed PACG without prior ANA diagnosis and sociodemographic factors associated with late detection. RESULTS: Thirty-one thousand forty-four patients were eligible. More than 70% of PACG cases were detected without prior ANA diagnosis, regardless of patient age, sex, or race. The odds of late detection were significantly higher (P < 0.001) among men (odds ratio [OR], 1.32; 95% confidence interval [CI], 1.25-1.40), Black patients (OR, 1.25; 95% CI, 1.15-1.37), and patients 80 years of age or older (OR, 1.28; 95% CI, 1.11-1.47) or living in Southern (OR, 1.30; 95% CI, 1.22-1.40) or Pacific (OR, 1.27; 95% CI, 1.16-1.36) regions. Findings were similar for patients with PACG with a record of gonioscopy and treatment or with a 24-month lookback period. CONCLUSIONS: Most patients who receive a new diagnosis of PACG in the United States do not have a prior diagnosis of ANA. The elderly, men, and Black patients are at higher risk of late detection. A need exists for increased disease awareness among providers and more accessible tools to detect patients at risk of developing PACG.

The steroidal lactone withaferin A impedes T-cell motility by inhibiting the kinase ZAP70 and subsequent kinome signaling.

The steroidal lactone withaferin A (WFA) is a dietary phytochemical, derived from Withania somnifera. It exhibits a wide range of biological properties, including immunomodulatory, anti-inflammatory, antistress, and anticancer activities. Here we investigated the effect of WFA on T-cell motility, which is crucial for adaptive immune responses as well as autoimmune reactions. We found that WFA dose-dependently (within the concentration range of 0.3-1.25 µM) inhibited the ability of human T-cells to migrate via cross-linking of the lymphocyte function-associated antigen-1 (LFA-1) integrin with its ligand, intercellular adhesion molecule 1 (ICAM-1). Coimmunoprecipitation of WFA interacting proteins and subsequent tandem mass spectrometry identified a WFA-interactome consisting of 273 proteins in motile T-cells. In particular, our data revealed significant enrichment of the zeta-chain-associated protein kinase 70 (ZAP70) and cytoskeletal actin protein interaction networks upon stimulation. Phospho-peptide mapping and kinome analysis substantiated kinase signaling downstream of ZAP70 as a key WFA target, which was further confirmed by bait-pulldown and Western immunoblotting assays. The WFA-ZAP70 interaction was disrupted by a disulfide reducing agent dithiothreitol, suggesting an involvement of cysteine covalent binding interface. In silico docking predicted WFA binding to ZAP70 at cystine 560 and 564 residues. These findings provide a mechanistic insight whereby WFA binds to and inhibits the ZAP70 kinase and impedes T-cell motility. We therefore conclude that WFA may be exploited to pharmacologically control host immune responses and potentially prevent autoimmune-mediated pathologies.

Trajectory of Pain, Functional Limitation, and Parental Coping Resources Following Pediatric Short-stay Surgery: Factors Impacting Rate of Recovery.

OBJECTIVES: Although there are many benefits of short-stay hospital admissions for high volume, pediatric surgical procedures, this model of care places greater responsibility on parents for the management of children's pain. This study aimed to document the trajectory of child pain outcomes and a range of parent-reported functional outcomes following discharge from a short-stay surgical admission. Moreover, we aimed to document the trajectory of parental perceived personal coping resources. Second, we assessed whether parental dispositional factors, assessed before hospital discharge, predicted the child's pain intensity and parent-reported functional recovery. METHODS: Participants included children (aged 4 to 14 y) admitted for a short-stay tonsillectomy or appendectomy, and their parents. Parents completed a questionnaire before discharge from hospital. Demographic and surgical information was recorded from medical records. Following discharge, daily assessments of pain and functioning were carried out over a 10-day period using iPods or mobile phones. Predischarge and postdischarge data were obtained for 55 child and parent dyads. RESULTS: Pain intensity scores returned to low levels (2/10 or less) by day 5 for appendectomy and day 10 for tonsillectomy. Parents' perceived personal coping resources increased more slowly following tonsillectomy than appendectomy. Controlling for time since surgery and parental coping resources, parental pain-related catastrophizing was a significant predictor of child pain and functional recovery. DISCUSSION: Short-stay surgery results in parents facing considerable burden in managing their child's pain and functional impairment over a 10-day period. The potential value of screening for parental pain-related catastrophizing before discharge from hospital warrants further consideration and may enable identification of children likely to experience poorer recovery.

Hemiretinal Asymmetry in Peripapillary Vessel Density in Healthy, Glaucoma Suspect, and Glaucoma Eyes.

PURPOSE: To investigate hemiretinal asymmetry in radial peripapillary capillary vessel area density (VAD) of healthy, glaucoma suspect, and glaucoma eyes of varying severity and its diagnostic utility for glaucoma. DESIGN: Population-based, cross-sectional study. METHODS: Optic disc scans (6 × 6 mm) were collected on optical coherence tomography angiography (OCTA) to obtain VAD and on optical coherence tomography (OCT) to measure circumpapillary retinal nerve fiber layer (RNFL) thickness. Hemiretinal difference in VAD (hdVAD) was defined as the absolute difference between superior and inferior hemiretinal VAD. Age-adjusted multivariable linear regression of hdVAD on glaucoma severity was performed. Areas under curves (AUCs) were calculated from predicted probabilities generated by multiple logistic regression of glaucoma severity on age-adjusted single and combined parameters. RESULTS: A total of 1,043 eyes of 1,043 participants (587 healthy, 270 suspect, 67 mild, 54 moderate, 65 severe glaucoma) were included. After age adjustment, mean hdVAD was similar between healthy and suspect (P = .225), higher in mild vs suspect (P < .001), and higher in moderate vs mild (P = .018), but lower in severe vs moderate (P = .001). AUCs of hdVAD were highest for discriminating mild (0.685) and moderate (0.681) glaucoma from healthy. Combining hdVAD and global RNFL (gRNFL) yielded the highest AUCs of all parameters for mild (0.818) and any POAG (0.859) and resulted in significantly better diagnostic accuracy than either hdVAD or gRNFL alone (P < .05 for all comparisons). CONCLUSIONS: hdVAD is higher in early glaucoma and may help with early detection when damage is focal, but its diagnostic ability appears less robust in advanced glaucoma when damage is diffuse.