Use of ultrasound and MRI to stage the axilla for breast cancer before and after neoadjuvant chemotherapy prior to targeted sentinel lymphadenectomy.

PURPOSE: Prior data from this Center demonstrated that for patients who had biopsy-proven axillary metastases, were ycN0 after neoadjuvant chemotherapy (NAC), and had a wire-directed (targeted) sentinel lymphadenectomy (WD-SLND), 60% were node negative. The hypothesis of this study was that results of axillary imaging either before or after NAC would be predictive of final pathologic status after WD-SLND. METHODS: For patients treated with NAC between 2015 and 2023, ultrasound and MRI images of the axilla were retrospectively reviewed by radiologists specializing in breast imaging, who were blinded to the surgical and pathology results. RESULTS: Of 113 patients who fit the clinical criteria, 66 (58%) were ypN0 at WD-SLND and 34 (30%) had a pathologic complete response to NAC. There was no correlation between the number of abnormal lymph nodes on pre-NAC ultrasound or MRI imaging and the final pathologic status of the lymph nodes. The positive predictive value (PPV) of abnormal post-NAC axillary imaging was 48% for ultrasound and 53% for MRI. The negative predictive value (NPV) for normal post-NAC axillary imaging was 67% for ultrasound and 68% for MRI. CONCLUSION: The results of axillary imaging were not adequate to identify lymph nodes after NAC that were persistently pathologically node positive or those which had become pathologically node negative.

An ALK2 inhibitor, BLU-782, prevents heterotopic ossification in a mouse model of fibrodysplasia ossificans progressiva.

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disease driven by gain-of-function variants in activin receptor-like kinase 2 (ALK2), the most common variant being ALK2R206H. In FOP, ALK2 variants display increased and dysregulated signaling through the bone morphogenetic protein (BMP) pathway resulting in progressive and permanent replacement of skeletal muscle and connective tissues with heterotopic bone, ultimately leading to severe debilitation and premature death. Here, we describe the discovery of BLU-782 (IPN60130), a small-molecule ALK2R206H inhibitor developed for the treatment of FOP. A small-molecule library was screened in a biochemical ALK2 binding assay to identify potent ALK2 binding compounds. Iterative rounds of structure-guided drug design were used to optimize compounds for ALK2R206H binding, ALK2 selectivity, and other desirable pharmacokinetic properties. BLU-782 preferentially bound to ALK2R206H with high affinity, inhibiting signaling from ALK2R206H and other rare FOP variants in cells in vitro without affecting signaling of closely related homologs ALK1, ALK3, and ALK6. In vivo efficacy of BLU-782 was demonstrated using a conditional knock-in ALK2R206H mouse model, where prophylactic oral dosing reduced edema and prevented cartilage and heterotopic ossification (HO) in both muscle and bone injury models. BLU-782 treatment preserved the normal muscle-healing response in ALK2R206H mice. Delayed dosing revealed a short 2-day window after injury when BLU-782 treatment prevented HO in ALK2R206H mice, but dosing delays of 4 days or longer abrogated HO prevention. Together, these data suggest that BLU-782 may be a candidate for prevention of HO in FOP.

Association of body mass index with progression from binge-eating behavior into binge-eating disorder among adolescents in the United States: A prospective analysis of pooled data.

The association between body mass index (BMI) and binge-eating disorder (BED) is well-established. However, data on the extent to which BMI is associated with progression from binge-eating behavior into BED among adolescents are limited, which was the aim of this investigation. Participants were 9964 U.S. adolescents from the Adolescent Brain Cognitive Development (ABCD) Study, aged 9-13 at the time of study enrollment. A computerized parent-reported assessment was used to establish adolescents' binge-eating behaviors and BED. Cox proportional hazards models adjusting for sociodemographic covariates were used to examine prospective associations between BMI and likelihood of BED onset among a) adolescents with binge-eating behavior, and b) adolescents with no binge-eating behavior. Of 975 adolescents who met the study criteria for binge-eating behavior, 89 (9.1%) subsequently met the study criteria for BED. Of 8989 adolescents with no binge-eating behavior, 82 (0.9%) subsequently met the study criteria for BED. BMI percentile was significantly associated with the likelihood of BED onset in participants with (adjusted HR = 1.03, 95% confidence interval [CI] 1.00, 1.06) and participants without (adjusted HR = 1.05, 95% CI 1.03, 1.07) binge-eating behavior. Results were also significant when examining BMI as a dichotomous predictor (above and below 85th percentile) among those with (adjusted HR = 2.60, 95% CI 1.00, 6.68) and those without (adjusted HR = 6.01, 95% CI 3.90, 11.10) binge-eating behavior. Overall, results indicate that elevated BMI is prospectively associated with a greater risk for BED onset among U.S. adolescents with or without binge-eating behavior. Adolescents with a higher BMI may benefit from screening for binge eating, and prevention/early intervention strategies to mitigate the risk for developing BED.

Back on the Streets: Examining Emergency Department Return Rates for Unhoused Patients Discharged After Trauma.

BACKGROUND: The unhoused population is at high risk for traumatic injuries and faces unique challenges in accessing follow-up care. However, there is scarce data regarding differences in Emergency Department (ED) return rates and reasons for return between unhoused and housed patients. METHODS: We conducted a 3-year retrospective cohort study at a level-1 trauma center in a large metropolitan area. All patients who presented to the ED with traumatic injuries and were discharged without hospital admission were included in the study. The primary outcome was ED returns for trauma-related complications or new traumatic events <6 months after discharge. Patient characteristics and study outcomes were compared between housed and unhoused groups. RESULTS: A total of 4184 patients were identified, of which 20.3% were unhoused. Compared to housed, unhoused patients were more likely to return to the ED (18.8% vs 13.9%, P < .001), more likely to return for trauma-related complications (4.6% vs 3.1%, P = .045), more likely to return with new trauma (7.1% vs 2.8%, P < .001), and less likely to return for scheduled wound checks (2.5% vs 4.3%, P = .012). Of the patients who returned with trauma-related complications, unhoused patients had a higher proportion of wound infection (20.5% vs 5.7%, P = .008). In the regression analysis, unhoused status was associated with increased odds of ED return with new trauma and decreased odds of return for scheduled wound checks. CONCLUSIONS: This study observed significant disparities between unhoused and housed patients after trauma. Our results suggest that inadequate follow-up in unhoused patients may contribute to further ED return.

Exposure-Response Relationships for Pralsetinib in Patients with RET-Altered Thyroid Cancer or RET Fusion-Positive Nonsmall Cell Lung Cancer.

Pralsetinib is a highly potent oral kinase inhibitor of oncogenic RET (rearranged during transfection) fusions and mutations. Pralsetinib received approval from the United States Food and Drug Administration for the treatment of patients with metastatic RET fusion-positive non-small cell lung cancer (NSCLC), and received accelerated approval for the treatment of patients with RET fusion-positive thyroid cancer. Exposure-response (ER) analyses of efficacy were performed separately in patients with thyroid cancer and in patients with NSCLC, but data for all patients were pooled for the safety analysis. ER models were developed with time-varying exposure; the effect of covariates was also examined. For patients with NSCLC, a higher starting dose was associated with improved progression-free survival (PFS), but this improvement did not correlate with a higher exposure overall. Significant covariates included sex and baseline Eastern Cooperative Oncology Group (ECOG) score. For patients with thyroid cancer, a higher exposure was associated with improved PFS. Significant covariates included prior systemic cancer therapy and ECOG score. For safety, higher exposure was associated with a greater risk of grade ≥3 anemia, pneumonia, and lymphopenia. Patients with an ECOG score of ≥1 had an increased risk of grade ≥3 pneumonia. Non-White patients had a lower risk of grade ≥3 lymphopenia. ER analysis revealed that higher pralsetinib exposure was associated with improved PFS in thyroid cancer, but not in NSCLC. However, a higher starting dose (ie, 400 vs ≤300 mg daily) was correlated with better PFS for all indications. Higher exposure was also associated with an increased risk of grade ≥3 adverse events (AEs); however, the overall incidence of these events was acceptably low (≤20%). This analysis supports the use of a 400 mg starting dose of pralsetinib, allowing for dose reduction in the event of AEs.

Single Ion Pair Is Essential for Stabilizing SHP2's Open Conformation.

Src-homology-2-domain-containing PTP-2 (SHP2) is a widely expressed signaling enzyme whose misregulation is associated with multiple human pathologies. SHP2's enzymatic activity is controlled by a conformational equilibrium between its autoinhibited ("closed") state and its activated ("open") state. Although SHP2's closed state has been extensively characterized, the putative structure of its open form has only been revealed in the context of a highly activated mutant (E76K), and no systematic studies of the biochemical determinants of SHP2's open-state stabilization have been reported. To identify amino-acid interactions that are critical for stabilizing SHP2's active state, we carried out a mutagenic study of residues that lie at potentially important interdomain interfaces of the open conformation. The open/closed equilibria of the mutants were evaluated, and we identified several interactions that contribute to the stabilization of SHP2's open state. In particular, our findings establish that an ion pair between glutamate 249 on SHP2's PTP domain and arginine 111 on an interdomain loop is the key determinant of SHP2's open-state stabilization. Mutations that disrupt the R111/E249 ion pair substantially shift SHP2's open/closed equilibrium to the closed state, even compared to wild-type SHP2's basal-state equilibrium, which strongly favors the closed state. To the best of our knowledge, the ion-pair variants uncovered in this study are the first known SHP2 mutants in which autoinhibition is augmented with respect to the wild-type protein. Such "hyperinhibited" mutants may provide useful tools for signaling studies that investigate the connections between SHP2 inhibition and the suppression of human disease progression.

Use of Stroboscopic Goggles in Suture Training Improves Precision and Accuracy.

BACKGROUND: This study aimed to quantify the impact of intermittent visual occlusion via stroboscopic goggles on suture accuracy and precision. METHODS: This crossover study recruited and randomized 72 graduate students to train with stroboscopic goggles early or late in structured suture practice. Participants completed assessments of 10 running sutures with 2 training sessions between baseline and follow-up assessments. The procedure was repeated after crossover. Suture photos were analyzed using ImageJ. Total error among all measurements represented accuracy; standard deviation of error represented precision. Intra- and inter-group trends were identified with Wilcoxon rank-sum tests. RESULTS: Both groups significantly improved in accuracy in the sessions immediately following goggle use, but the group that used goggles later in training continued improving in accuracy and precision while the group that trained with stroboscopic goggles early plateaued. CONCLUSIONS: Using stroboscopic goggles showed quantifiable benefit for augmenting suture training with greatest effect after initial skill acquisition is completed.

Multi-modal skin atlas identifies a multicellular immune-stromal community associated with altered cornification and specific T cell expansion in atopic dermatitis.

In healthy skin, a cutaneous immune system maintains the balance between tolerance towards innocuous environmental antigens and immune responses against pathological agents. In atopic dermatitis (AD), barrier and immune dysfunction result in chronic tissue inflammation. Our understanding of the skin tissue ecosystem in AD remains incomplete with regard to the hallmarks of pathological barrier formation, and cellular state and clonal composition of disease-promoting cells. Here, we generated a multi-modal cell census of 310,691 cells spanning 86 cell subsets from whole skin tissue of 19 adult individuals, including non-lesional and lesional skin from 11 AD patients, and integrated it with 396,321 cells from four studies into a comprehensive human skin cell atlas in health and disease. Reconstruction of human keratinocyte differentiation from basal to cornified layers revealed a disrupted cornification trajectory in AD. This disrupted epithelial differentiation was associated with signals from a unique immune and stromal multicellular community comprised of MMP12 + dendritic cells (DCs), mature migratory DCs, cycling ILCs, NK cells, inflammatory CCL19 + IL4I1 + fibroblasts, and clonally expanded IL13 + IL22 + IL26 + T cells with overlapping type 2 and type 17 characteristics. Cell subsets within this immune and stromal multicellular community were connected by multiple inter-cellular positive feedback loops predicted to impact community assembly and maintenance. AD GWAS gene expression was enriched both in disrupted cornified keratinocytes and in cell subsets from the lesional immune and stromal multicellular community including IL13 + IL22 + IL26 + T cells and ILCs, suggesting that epithelial or immune dysfunction in the context of the observed cellular communication network can initiate and then converge towards AD. Our work highlights specific, disease-associated cell subsets and interactions as potential targets in progression and resolution of chronic inflammation.

Differential Impact of Smoking on Methylome and Transcriptome in Crohn's Disease and Ulcerative Colitis.

BACKGROUND: Smoking is an environmental factor that differentially impacts Crohn's disease (CD) and ulcerative colitis (UC). The mechanism of impact of smoking on disease risk and clinical outcomes remains to be established. METHODS: This study used a prospective cohort of patients with CD or UC. Self-reported smoking status was validated using serum cotinine measurement. We profiled methylation changes in peripheral blood using the Illumina Methylation BeadChip. Transcriptomic profiling was performed on ileal and colonic tissue using an Illumina TruSeq platform. We compared the methylation and transcriptional changes in current, former, and never smokers stratified by disease type. RESULTS: Our cohort included 200 patients with CD or UC with methylation profiles and 160 with transcriptomic data. The mean serum cotinine level was higher in current compared with former or never smokers. Epigenetic changes common to both CD and UC included hypomethylation at AHRR. Smoking-associated MGAT3 hypomethylation was associated with severe disease course only in UC, while IER3 hypomethylation was associated with worse course only in CD. Smoking downregulated several inflammatory pathways in UC. Current smoking in CD but not in UC was associated with upregulation of several genes mediating Paneth cell function. Genes with opposite direction of effects in CD and UC include HSD3B2 and GSTA1. CONCLUSIONS: Our findings suggest both common and differential effects of cigarette smoking on CD and UC. Paneth cell dysfunction may mediate adverse impact of smoking on CD. Bile acid and oxidative stress pathways may be relevant for the differential effect of smoking on CD and UC.

Smoking is a key environmental risk factor for the development of inflammatory bowel disease. Smoking induces changes differential epigenetic changes in the peripheral blood in Crohn's disease and ulcerative colitis. Smoking also induces down regulation of expression of various proinflammatory genes in the colon in ulcerative colitis.

Increased spatial coupling of integrin and collagen IV in the immunoresistant clear cell renal cell carcinoma tumor microenvironment.

Background: Immunotherapy (IO) has improved survival for patients with advanced clear cell renal cell carcinoma (ccRCC), but resistance to therapy develops in most patients. We use cellular-resolution spatial transcriptomics in patients with IO naïve and IO exposed primary ccRCC tumors to better understand IO resistance. Spatial molecular imaging (SMI) was obtained for tumor and adjacent stroma samples. Spatial gene set enrichment analysis (GSEA) and autocorrelation (coupling with high expression) of ligand-receptor transcript pairs were assessed. Multiplex immunofluorescence (mIF) validation was used for significant autocorrelative findings and the cancer genome atlas (TCGA) and the clinical proteomic tumor analysis consortium (CPTAC) databases were queried to assess bulk RNA expression and proteomic correlates. Results: 21 patient samples underwent SMI. Viable tumors following IO harbored more stromal CD8+ T cells and neutrophils than IO naïve tumors. YES1 was significantly upregulated in IO exposed tumor cells. The epithelial-mesenchymal transition pathway was enriched on spatial GSEA and the associated transcript pair COL4A1-ITGAV had significantly higher autocorrelation in the stroma. Fibroblasts, tumor cells, and endothelium had the relative highest expression. More integrin αV+ cells were seen in IO exposed stroma on mIF validation. Compared to other cancers in TCGA, ccRCC tumors have the highest expression of both COL4A1 and ITGAV. In CPTAC, collagen IV protein was more abundant in advanced stages of disease. Conclusions: On spatial transcriptomics, COL4A1 and ITGAV were more autocorrelated in IO-exposed stroma compared to IO-naïve tumors, with high expression amongst fibroblasts, tumor cells, and endothelium. Integrin represents a potential therapeutic target in IO treated ccRCC.

Spatiotemporal cell junction assembly in human iPSC-CM models of arrhythmogenic cardiomyopathy.

Arrhythmogenic cardiomyopathy (ACM) is an inherited cardiac disorder that causes life-threatening arrhythmias and myocardial dysfunction. Pathogenic variants in Plakophilin-2 (PKP2), a desmosome component within specialized cardiac cell junctions, cause the majority of ACM cases. However, the molecular mechanisms by which PKP2 variants induce disease phenotypes remain unclear. Here we built bioengineered platforms using genetically modified human induced pluripotent stem cell-derived cardiomyocytes to model the early spatiotemporal process of cardiomyocyte junction assembly in vitro. Heterozygosity for truncating variant PKP2R413X reduced Wnt/ß-catenin signaling, impaired myofibrillogenesis, delayed mechanical coupling, and reduced calcium wave velocity in engineered tissues. These abnormalities were ameliorated by SB216763, which activated Wnt/ß-catenin signaling, improved cytoskeletal organization, restored cell junction integrity in cell pairs, and improved calcium wave velocity in engineered tissues. Together, these findings highlight the therapeutic potential of modulating Wnt/ß-catenin signaling in a human model of ACM.

Prevalence of Low-Frequency, Antiviral Resistance Variants in SARS-CoV-2 Isolates in Ontario, Canada, 2020-2023.

Importance: Nirmatrelvir-ritonavir is an oral antiviral medication that improves outcomes in SARS-CoV-2 infections. However, there is concern that antiviral resistance will develop and that these viruses could be selected for after treatment. Objective: To determine the prevalence of low-frequency SARS-CoV-2 variants in patient samples that could be selected for by nirmatrelvir-ritonavir. Design, Setting, and Participants: This retrospective cohort study was conducted at 4 laboratories that serve community hospitals, academic tertiary care centers, and COVID-19 assessment centers in Ontario, Canada. Participants included symptomatic or asymptomatic patients who tested positive for SARS-CoV-2 virus and submitted virus samples for diagnostic testing between March 2020 and January 2023. Exposure: SARS-CoV-2 infection. Main Outcomes and Measures: Samples with sufficient viral load underwent next-generation genome sequencing to identify low-frequency antiviral resistance variants that could not be identified through conventional sequencing. Results: This study included 78 866 clinical samples with next-generation whole-genome sequencing data for SARS-CoV-2. Low-frequency variants in the viral nsp5 gene were identified in 128 isolates (0.16%), and no single variant associated with antiviral resistance was predominate. Conclusions and Relevance: This cohort study of low-frequency variants resistant to nirmatrelvir-ritonavir found that these variants were very rare in samples from patients with SARS-CoV-2, suggesting that selection of these variants by nirmatrelvir-ritonavir following the initiation of treatment may also be rare. Surveillance efforts that involve sequencing of viral isolates should continue to monitor for novel resistance variants as nirmatrelvir-ritonavir is used more broadly.

School Nurses' Experiences and Needs in COVID-19 Pandemic Response: A Qualitative Study.

The coronavirus disease 2019 pandemic drastically shifted the public health burden onto the school health system. We, therefore, explored the school nurses' experiences and needs for confronting the pandemic in school settings across Korea. This qualitative investigation enrolled 30 participants after their written consent. A written interview conducted through email and focus group interviews using an online meeting application verified participants' opinions and experiences. Collected data were subjected to text-mining and content analysis. The school nurse was found to be responsible for many tasks, including education on preventive measures with unclear quarantine guidelines. Content analysis identified three key themes: "aggravated difficulties owing to lack of support resulted in burnout," "reflection on supportive resources; direct provisions were helpful," and "needs and suggestions to safeguard students' health in the event of another pandemic." Recommendations should all be considered to keep schools and students safe from future pandemics.

Fibre-infused gel scaffolds guide cardiomyocyte alignment in 3D-printed ventricles.

Hydrogels are attractive materials for tissue engineering, but efforts to date have shown limited ability to produce the microstructural features necessary to promote cellular self-organization into hierarchical three-dimensional (3D) organ models. Here we develop a hydrogel ink containing prefabricated gelatin fibres to print 3D organ-level scaffolds that recapitulate the intra- and intercellular organization of the heart. The addition of prefabricated gelatin fibres to hydrogels enables the tailoring of the ink rheology, allowing for a controlled sol-gel transition to achieve precise printing of free-standing 3D structures without additional supporting materials. Shear-induced alignment of fibres during ink extrusion provides microscale geometric cues that promote the self-organization of cultured human cardiomyocytes into anisotropic muscular tissues in vitro. The resulting 3D-printed ventricle in vitro model exhibited biomimetic anisotropic electrophysiological and contractile properties.

[68 Ga]-DOTATATE PET/MR-based evaluation of physiologic somatostatin receptor 2 expression in the adult pituitary gland as a function of age and sex in a prospective cohort.

PURPOSE: The pituitary gland has the fourth highest physiologic avidity of [68 Ga]-DOTATATE. In order to guide our understanding of [68 Ga]-DOTATATE PET in clinical contexts, accurate characterization of the normal pituitary gland is first required. This study aimed to characterize the normal pituitary gland using dedicated brain [68 Ga]-DOTATATE PET/MRI as a function of age and sex. METHODS: A total of 95 patients with a normal pituitary gland underwent brain [68 Ga]-DOTATATE PET examinations for the purpose of diagnosing CNS SSTR2 positive tumors (mean age: 58.9, 73% female). Maximum SUV of the pituitary gland was obtained in each patient. SUV of superior sagittal sinus was obtained to calculate normalized SUV score (SUVR) of the gland. The anatomic size of the gland was collected as maximum sagittal height (MSH). Correlations with age and sex were analyzed. RESULTS: The mean SUV and SUVR of the pituitary gland were 17.6 (range: 7-59.5, SD = 7.1) and 13.8 (range: 3.3-52.6, SD = 7.2), respectively. Older females had significantly higher SUV of the pituitary gland compared to younger females. When stratified by age and sex, both older and younger females had significantly higher pituitary SUV than older males. SUVR did not differ significantly by age or sex. MSH of the pituitary gland in younger females was significantly greater than in younger males at all age cutoffs. CONCLUSION: This study provides an empiric profiling of the physiological [68 Ga]-DOTATATE avidity of the pituitary gland. The findings suggest that SUV may vary by age and sex and can help guide the use of [68 Ga]-DOTATATE PET/MRI in clinical and research settings. Future studies can build on these findings to investigate further the relationship between pituitary biology and demographic factors.

Substance misuse training in pharmacy education: Results from a pilot study on the integration of an interprofessional experience.

Background: Studies show that pharmacists are unsure in their ability to screen patients for substance abuse. This study evaluates the efficacy of incorporating interprofessional education (IPE) into a substance misuse training program on pharmacy students' learning outcomes in providing screening and counseling for substance misuse. Methods: Pharmacy students from 2019 to 2020 completed 3 substance misuse training modules. Students from the class of 2020 completed an additional IPE event. Both cohorts completed pre- and post-surveys that evaluated knowledge of content and comfort level with patient screening and counseling regarding substance misuse. Paired student t-tests and difference-in-difference analyses were used to evaluate the impact of the IPE event. Findings: Both cohorts (n = 127) showed statistically significant improvement in learning outcomes in providing substance misuse screening and counseling. IPE yielded extremely positive feedback from all students, but its addition to the overall training did not improve learning outcomes. This may be attributed to the differences in baseline knowledge of each class cohort. Conclusion: Substance misuse training successfully improved pharmacy student knowledge and comfort level with providing patient screening and counseling services. Though the IPE event did not improve learning outcomes, qualitative student feedback was overwhelmingly positive and supports the continued incorporation of IPE.

Increases in hydraulic conductance and solute permeability in a mouse model of ascending thoracic aortic aneurysm.

Large elastic arteries, such as the aorta, contain concentric layers of elastic laminae composed mainly of the extracellular matrix protein elastin. The structure of the elastic laminae could affect transmural mass transport and contribute to aortic disease progression. We studied the effects of a genetic mutation (LoxM292R/+, referred to as MU) in mice associated with ascending thoracic aortic aneurysm (TAA) on the mass transport and elastic laminae structure. Solute absent fluid flux and hydraulic conductance through the ascending aortic wall were not significantly different between groups, however solute present fluid flux, hydraulic conductance, solute flux, and solute permeability of 4 kDa FITC-dextran were significantly increased in the MU group, indicating that movement of small molecules into the aortic wall is facilitated in MU mice. Quantification from light microscopy images of the ascending aorta showed no significant differences in wall thickness, or inner elastic lamina fenestration size and density, but an increase in the number of elastic laminae breaks in the MU group. Ultrastructural comparisons from transmission electron micrographs suggest less dense and disorganized elastic laminae in MU aorta that may also contribute to the transport differences. Our results provide an initial investigation into the connections between mass transport and elastic laminae structure, specifically in a genetic mouse aneurysm model, which can be further used to understand TAA pathology and develop treatment strategies.

Destabilization of the SHP2 and SHP1 protein tyrosine phosphatase domains by a non-conserved "backdoor" cysteine.

Protein tyrosine phosphatases (PTPs) are critical regulators of cellular signal transduction that catalyze the hydrolytic dephosphorylation of phosphotyrosine in substrate proteins. Among several conserved features in classical PTP domains are an active-site cysteine residue that is necessary for catalysis and a "backdoor" cysteine residue that can serve to protect the active-site cysteine from irreversible oxidation. Curiously, two biologically important phosphatases, Src homology domain-containing PTPs 2 and 1 (SHP2 and SHP1), each contain an additional backdoor cysteine residue at a position of the PTP domain that is occupied by proline in almost all other classical PTPs (position 333 in human SHP2 numbering). Here we show that the presence of cysteine 333 significantly destabilizes the fold of the PTP domains in the SHPs. We find that replacement of cysteine 333 with proline confers increased thermal stability on the SHP2 and SHP1 PTP domains, as measured by temperature-dependent activity assays and differential scanning fluorimetry. Conversely, we show that substantial destabilization of the PTP-domain fold is conferred by introduction of a non-natural cysteine residue in a non-SHP PTP that contains proline at the 333 position. It has previously been suggested that the extra backdoor cysteine of the SHP PTPs may work in tandem with the conserved backdoor cysteine to provide protection from irreversible oxidative enzyme inactivation. If so, our current results suggest that, during the course of mammalian evolution, the SHP proteins have developed extra protection from oxidation at the cost of the thermal instability that is conferred by the presence of their PTP domains' second backdoor cysteine.

Recreating the heart's helical structure-function relationship with focused rotary jet spinning.

Helical alignments within the heart's musculature have been speculated to be important in achieving physiological pumping efficiencies. Testing this possibility is difficult, however, because it is challenging to reproduce the fine spatial features and complex structures of the heart's musculature using current techniques. Here we report focused rotary jet spinning (FRJS), an additive manufacturing approach that enables rapid fabrication of micro/nanofiber scaffolds with programmable alignments in three-dimensional geometries. Seeding these scaffolds with cardiomyocytes enabled the biofabrication of tissue-engineered ventricles, with helically aligned models displaying more uniform deformations, greater apical shortening, and increased ejection fractions compared with circumferential alignments. The ability of FRJS to control fiber arrangements in three dimensions offers a streamlined approach to fabricating tissues and organs, with this work demonstrating how helical architectures contribute to cardiac performance.

Evaluating diagnostic accuracy and determining optimal diagnostic thresholds of different approaches to [68Ga]-DOTATATE PET/MRI analysis in patients with meningioma.

Multiple approaches with [68Ga]-DOTATATE, a somatostatin analog PET radiotracer, have demonstrated clinical utility in evaluation of meningioma but have not been compared directly. Our purpose was to compare diagnostic performance of different approaches to quantitative brain [68Ga]-DOTATATE PET/MRI analysis in patients with suspected meningioma recurrence and to establish the optimal diagnostic threshold for each method. Patients with suspected meningioma were imaged prospectively with [68Ga]-DOTATATE brain PET/MRI. Lesions were classified as meningiomas and post-treatment change (PTC), using follow-up pathology and MRI as reference standard. Lesions were reclassified using the following methods: absolute maximum SUV threshold (SUV), SUV ratio (SUVR) to superior sagittal sinus (SSS) (SUVRsss), SUVR to the pituitary gland (SUVRpit), and SUVR to the normal brain parenchyma (SUVRnorm). Diagnostic performance of the four methods was compared using contingency tables and McNemar's test. Previously published pre-determined thresholds were assessed where applicable. The optimal thresholds for each method were identified using Youden's J statistics. 166 meningiomas and 41 PTC lesions were identified across 62 patients. SUV, SUVRsss, SUVRpit, and SUVRnorm of meningioma were significantly higher than those of PTC (P < 0.0001). The optimal thresholds for SUV, SUVRsss, SUVRpit, and SUVRnorm were 4.7, 3.2, 0.3, and 62.6, respectively. At the optimal thresholds, SUV had the highest specificity (97.6%) and SUVRsss had the highest sensitivity (86.1%). An ROC analysis of SUV, SUVRsss, SUVRpit, and SUVRnorm revealed AUC of 0.932, 0.910, 0.915, and 0.800, respectively (P < 0.0001). Developing a diagnostic threshold is key to wider clinical translation of [68Ga]-DOTATATE PET/MRI in meningioma evaluation. We found that the SUVRsss method may have the most robust combination of sensitivity and specificity in the diagnosis of meningioma in the post-treatment setting, with the optimal threshold of 3.2. Future studies validating our findings in different patient populations are needed to continue optimizing the diagnostic performance of [68Ga]-DOTATATE PET/MRI in meningioma patients.Trial registration: ClinicalTrials.gov Identifier: NCT04081701. Registered 9 September 2019. https://clinicaltrials.gov/ct2/show/NCT04081701 .