Search

Background: Thyroid nodules may be detected during the workup of thyroid hormone abnormalities and as incidental findings during unrelated imaging studies. The diagnosis of a thyroid nodule is mainly established by performing fine needle aspiration (FNA) under ultrasound guidance. Thyroid nodules are classified as nondiagnostic, defined in the Bethesda System for Reporting Thyroid Cytopathology as samples with excess blood, cyst fluid only, and lack of thyroid follicular cells. The current study evaluates a series of nondiagnostic FNAs to assess whether repeat sampling improves yield and what patient management, and outcomes are after a nondiagnostic FNA., Methods: Thyroid FNAs from 2016 to 2023 were retrieved from our institution archives. All cases were performed under ultrasound guidance and with rapid on-site evaluation. Cases were assigned the Bethesda System Category. Nondiagnostic FNAs were further reviewed for repeat FNA procedures, potential molecular testing, or diagnostic resections., Results: In total 3104 thyroid FNAs were reviewed, with 153 (4.9%) being nondiagnostic. Of the 154 FNAs, there were 129 patients with an average age of 60 and a male-to-female ratio of 1:3.2. Of the 130 patients, there were 50 patients who underwent 55 repeat FNAs. Thirty-seven (67%) of the repeats were benign, 13 (24%) were nondiagnostic again, and 5 (9%) were atypia of undetermined significance (AUS). Molecular testing was performed on repeat FNAs diagnosed AUS. Four cases showed no mutations and had a high likelihood of being benign. One case did have an NRAS Q61R mutation, and resection revealed a noninvasive follicular thyroid neoplasm with papillary-like nuclear features. Seventeen (13% of all cases) with nondiagnostic FNA were resected. Twelve (71%) thyroidectomies showed benign adenomatous nodules. The remainder showed incidental papillary thyroid microcarcinoma (0.1 cm), an infarcted follicular adenoma, a noninvasive follicular thyroid neoplasm with papillary-like nuclear features, and metastatic renal cell carcinoma (2×)., Conclusion: Thyroid nodules with nondiagnostic cytology are reassuring of being highly likely a benign nodule. Only 5 of the 55 (9%) repeat FNAs yielded abnormalities, with only one of those being truly a follicular neoplasm (confirmed by molecular testing and resection). No primary thyroid malignancies have been identified in follow-up (repeat FNA or surgery). Clinical and ultrasound follow-up may be more appropriate management for nondiagnostic thyroid FNAs., (© 2024 The Author(s). Diagnostic Cytopathology published by Wiley Periodicals LLC.)

Pancreatic cancer, ranking as the third factor in cancer-related deaths, necessitates enhanced diagnostic measures through early detection. In response, SiMoT-Single-molecule with a large Transistor multiplexing array, achieving a Technology Readiness Level of 5, is proposed for a timely identification of pancreatic cancer precursor cysts and is benchmarked against the commercially available chemiluminescent immunoassay SIMOA (Single molecule array) SP-X System. A cohort of 39 samples, comprising 33 cyst fluids and 6 blood plasma specimens, undergoes detailed examination with both technologies. The SiMoT array targets oncoproteins MUC1 and CD55, and oncogene KRAS, while the SIMOA SP-X planar technology exclusively focuses on MUC1 and CD55. Employing Principal Component Analysis (PCA) for multivariate data processing, the SiMoT array demonstrates effective discrimination of malignant/pre-invasive high-grade or potentially malignant low-grade pancreatic cysts from benign non-mucinous cysts. Conversely, PCA analysis applied to SIMOA assay reveals less effective differentiation ability among the three cyst classes. Notably, SiMoT unique capability of concurrently analyzing protein and genetic markers with the threshold of one single molecule in 0.1 mL positions it as a comprehensive and reliable diagnostic tool. The electronic response generated by the SiMoT array facilitates direct digital data communication, suggesting potential applications in the development of field-deployable liquid biopsy., (© 2024 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

Traumatic brain injury (TBI), a significant global health issue, is affecting ∼69 million annually. To better understand TBI's impact on brain function and assess the efficacy of treatments, this study uses a novel temporal-spatial cross-group approach with a porcine model, integrating resting-state functional magnetic resonance imaging (rs-fMRI) for temporal and arterial spin labeling for spatial information. Our research used 18 four-week-old pigs divided into three groups: TBI treated with saline (SLN, n = 6), TBI treated with fecal microbial transplant (FMT, n = 6), and a sham group (sham, n = 6) with only craniectomy surgery as the baseline. By applying machine learning techniques-specifically, independent component analysis and sparse dictionary learning-across seven identified resting-state networks, we assessed the temporal and spatial correlations indicative of treatment efficacy. Both temporal and spatial analyses revealed a consistent increase of correlation between the FMT and sham groups in the executive control and salience networks. Our results are further evidenced by a simulation study designed to mimic the progression of TBI severity through the introduction of variable Gaussian noise to an independent rs-fMRI dataset. The results demonstrate a decreasing temporal correlation between the sham and TBI groups with increasing injury severity, consistent with the experimental results. This study underscores the effectiveness of the methodology in evaluating post-TBI treatments such as the FMT. By presenting comprehensive experimental and simulated data, our research contributes significantly to the field and opens new paths for future investigations into TBI treatment evaluations., (© Wenwu Sun et al., 2024; Published by Mary Ann Liebert, Inc.)

Lactate elevation is a well-characterized biomarker of mitochondrial dysfunction, but its role in diabetic kidney disease (DKD) is not well defined. Urine lactate was measured in patients with type 2 diabetes (T2D) in 3 cohorts (HUNT3, SMART2D, CRIC). Urine and plasma lactate were measured during euglycemic and hyperglycemic clamps in participants with type 1 diabetes (T1D). Patients in the HUNT3 cohort with DKD had elevated urine lactate levels compared with age- and sex-matched controls. In patients in the SMART2D and CRIC cohorts, the third tertile of urine lactate/creatinine was associated with more rapid estimated glomerular filtration rate decline, relative to first tertile. Patients with T1D demonstrated a strong association between glucose and lactate in both plasma and urine. Glucose-stimulated lactate likely derives in part from proximal tubular cells, since lactate production was attenuated with sodium-glucose cotransporter-2 (SGLT2) inhibition in kidney sections and in SGLT2-deficient mice. Several glycolytic genes were elevated in human diabetic proximal tubules. Lactate levels above 2.5 mM potently inhibited mitochondrial oxidative phosphorylation in human proximal tubule (HK2) cells. We conclude that increased lactate production under diabetic conditions can contribute to mitochondrial dysfunction and become a feed-forward component to DKD pathogenesis.

Background: TP53 mutation is present in about 50.8% of lung adenocarcinomas, frequently in combination with other genetic alterations. However, a rare subset harbors the TP53 mutation alone., Methods: Next-generation sequencing was performed in 844 lung adenocarcinomas diagnosed by fine needle aspiration. Fourteen cases (1.7%) showed isolated TP53 alteration and were subjected to a comprehensive analysis., Results: The average age at diagnosis was 65.7 years (range 48-79); 9 males and 5 females. All were smokers with an average pack-year of 40.7 (range 10-70). Ten had metastases, mostly in the brain (n = 4) and pleura (n = 4). After a follow-up period of up to 102 months, 9 died, 3 were alive free of disease, 1 was alive with disease, and 1 was lost to follow-up. The median survival was 12.2 months. Most tumors exhibited poor differentiation, composed of solid sheets with moderate to severe atypia, increased mitotic activity, and necrotic background. Half were positive for TTF-1 and showed p53 overexpression. PD-L1 was positive in 5 cases. Most alterations were missense mutations in exons 5-8, and this mutation type was associated with p53 overexpression. Tumors with combined missense mutation and truncated protein had higher PD-L1 expression along with a trend towards an increase in tumor mutational burden (TMB). CEBPA deletion of undetermined significance was the most common copy number alteration., Conclusion: Isolated TP53 mutation was seen in association with smoking, high-grade cytomorphologic features, adverse prognosis, and recurrent CEBPA deletions. These tumors tend to have strong PD-L1 expression and high TMB, suggesting potential benefit from immune checkpoint inhibitors. Hence, the recognition of this molecular group has prognostic and therapeutic implications., (© 2023 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

A cohort of 47 patients is screened for pancreatic cancer precursors with a portable 96-well bioelectronic sensing-array for single-molecule assay in cysts fluid and blood plasma, deployable at point-of-care (POC). Pancreatic cancer precursors are mucinous cysts diagnosed with a sensitivity of at most 80% by state-of-the-art cytopathological molecular analyses (e.g., KRAS mut DNA). Adding the simultaneous assay of proteins related to malignant transformation (e.g., MUC1 and CD55) is deemed essential to enhance diagnostic accuracy. The bioelectronic array proposed here, based on single-molecule-with-a-large-transistor (SiMoT) technology, can assay both nucleic acids and proteins at the single-molecule limit-of-identification (LOI) (1% of false-positives and false-negatives). It comprises an enzyme-linked immunosorbent assay (ELISA)-like 8 × 12-array organic-electronics disposable cartridge with an electrolyte-gated organic transistor sensor array, and a reusable reader, integrating a custom Si-IC chip, operating via software installed on a USB-connected smart device. The cartridge is complemented by a 3D-printed sensing gate cover plate. KRAS mut , MUC1, and CD55 biomarkers either in plasma or cysts-fluid from 5 to 6 patients at a time, are multiplexed at single-molecule LOI in 1.5 h. The pancreatic cancer precursors are classified via a machine-learning analysis resulting in at least 96% diagnostic-sensitivity and 100% diagnostic-specificity. This preliminary study opens the way to POC liquid-biopsy-based early diagnosis of pancreatic-cancer precursors in plasma., (© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

Importance: SARS-CoV-2 infection is associated with persistent, relapsing, or new symptoms or other health effects occurring after acute infection, termed postacute sequelae of SARS-CoV-2 infection (PASC), also known as long COVID. Characterizing PASC requires analysis of prospectively and uniformly collected data from diverse uninfected and infected individuals., Objective: To develop a definition of PASC using self-reported symptoms and describe PASC frequencies across cohorts, vaccination status, and number of infections., Design, Setting, and Participants: Prospective observational cohort study of adults with and without SARS-CoV-2 infection at 85 enrolling sites (hospitals, health centers, community organizations) located in 33 states plus Washington, DC, and Puerto Rico. Participants who were enrolled in the RECOVER adult cohort before April 10, 2023, completed a symptom survey 6 months or more after acute symptom onset or test date. Selection included population-based, volunteer, and convenience sampling., Exposure: SARS-CoV-2 infection., Main Outcomes and Measures: PASC and 44 participant-reported symptoms (with severity thresholds)., Results: A total of 9764 participants (89% SARS-CoV-2 infected; 71% female; 16% Hispanic/Latino; 15% non-Hispanic Black; median age, 47 years [IQR, 35-60]) met selection criteria. Adjusted odds ratios were 1.5 or greater (infected vs uninfected participants) for 37 symptoms. Symptoms contributing to PASC score included postexertional malaise, fatigue, brain fog, dizziness, gastrointestinal symptoms, palpitations, changes in sexual desire or capacity, loss of or change in smell or taste, thirst, chronic cough, chest pain, and abnormal movements. Among 2231 participants first infected on or after December 1, 2021, and enrolled within 30 days of infection, 224 (10% [95% CI, 8.8%-11%]) were PASC positive at 6 months., Conclusions and Relevance: A definition of PASC was developed based on symptoms in a prospective cohort study. As a first step to providing a framework for other investigations, iterative refinement that further incorporates other clinical features is needed to support actionable definitions of PASC.

The most abundant cellular divalent cations, Mg 2+ (mM) and Ca 2+ (nM-μM), antagonistically regulate divergent metabolic pathways with several orders of magnitude affinity preference, but the physiological significance of this competition remains elusive. In mice consuming a Western diet, genetic ablation of the mitochondrial Mg 2+ channel Mrs2 prevents weight gain, enhances mitochondrial activity, decreases fat accumulation in the liver, and causes prominent browning of white adipose. Mrs2 deficiency restrains citrate efflux from the mitochondria, making it unavailable to support de novo lipogenesis. As citrate is an endogenous Mg 2+ chelator, this may represent an adaptive response to a perceived deficit of the cation. Transcriptional profiling of liver and white adipose reveals higher expression of genes involved in glycolysis, β-oxidation, thermogenesis, and HIF-1α-targets, in Mrs2 -/- mice that are further enhanced under Western-diet-associated metabolic stress. Thus, lowering m Mg 2+ promotes metabolism and dampens diet-induced obesity and metabolic syndrome., Competing Interests: Declaration of interests M.M. is an inventor on a patent filed by UTHSA on CPACC as a Mrs2 blocker for Mg(2+) in physiology and disease. J.A.B. is a consultant to Pfizer., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

In cytologically indeterminate thyroid nodules undergoing molecular testing, estimated risk of malignancy is variable. Identification of a non-cancer-specific mutation (RAS-like) confirms a neoplastic process but does not differentiate between benign, malignant, and low-risk neoplasms. This study aims to retrospectively evaluate institutional experience of Interpace (ThyGeNEXT ® and ThyraMIR ® ; Pittsburgh, PA) testing and to determine the rate of malignancy in resected nodules, stratified by mutational analysis and microRNA profile. Of 1917 fine need aspirations, 140 (7.3%) underwent Interpace testing: 47 (33.6%) were molecular-not-benign (harbored mutation, fusion, and/or positive miRNA) and 93 (66.4%) were molecular-benign (no mutations or fusions and negative microRNA). Surgery was spared in 79.6% of molecular-benign and 61.4% of all tested patients. Fifty-four (38.6%) underwent resection. Seventeen (89.5%) of the resected molecular-benign were benign and 2 were malignant. Thirteen (37.1%) of the resected molecular-not-benign were benign, 7 (20%) were noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP), and 15 (42.9%) were malignant (p < 0.05, negative predictive value (NPV) 89.4-95.6%, positive predictive value (PPV) 22.3-42.8%). Most molecular-not-benign (72.3%) had RAS-like mutation. Twenty-three were resected: 3 were malignant and 7 were NIFTP. Nodules with non-RAS-like mutations (BRAF V600E-like, others) were more likely to be malignant than RAS-like (H/N/KRAS, BRAF K601E) (p < 0.05, NPV 86.9-96.5%, PPV 100%). Most nodules had RAS-like mutations and most were benign or low-risk neoplasms (NIFTP). This study supports the role of histologic examination in the distinction of malignancy in RAS-like thyroid neoplasms and underscores the role of molecular testing in risk stratification, patient counseling, and operative management., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Introduction: Biomarkers of acute kidney injury (AKI) are often indexed to urine creatinine (UCr) or urine osmolarity (UOsm) to control for urine concentration. We evaluated how these approaches affect the biomarker-outcome association in patients with AKI., Methods: The Assessment, Serial Evaluation, and Subsequent Sequelae in Acute Kidney Injury Study was a cohort of hospitalized patients with and without AKI between 2009 and 2015. Using Cox proportional hazards regression, we assessed the associations and predictions (C-statistics) of urine biomarkers with a composite outcome of incident chronic kidney disease (CKD) and CKD progression. We used 4 approaches to account for urine concentration: indexing and adjusting for UCr and UOsm., Results: Among 1538 participants, 769 (50%) had AKI and 300 (19.5%) developed composite CKD outcome at median follow-up of 4.7 years. UCr and UOsm during hospitalization were inversely associated with the composite CKD outcome. The associations and predictions with CKD were significantly strengthened after indexing or adjusting for UCr or UOsm for urine kidney injury molecule-1 (KIM-1), interleukin-18 (IL-18), and monocyte chemoattractant protein-1 (MCP-1) in patients with AKI. There was no significant improvement with indexing or adjusting UCr or UOsm for albumin, neutrophil gelatinase-associated lipocalin (NGAL), and chitinase 3-like 1 (YKL-40). Uromodulin's (UMOD) inverse association with the outcome was significantly blunted after indexing but not adjusting for UCr or UOsm., Conclusion: UCr and UOsm during hospitalization are inversely associated with development and progression of CKD. Indexing or adjusting for UCr or UOsm strengthened associations and improved predictions for CKD for only some biomarkers. Incorporating urinary concentration should be individualized for each biomarker in research and clinical applications., (© 2022 International Society of Nephrology. Published by Elsevier Inc.)

Background: The mechanisms underlying long-term sequelae after AKI remain unclear. Vessel instability, an early response to endothelial injury, may reflect a shared mechanism and early trigger for CKD and heart failure., Methods: To investigate whether plasma angiopoietins, markers of vessel homeostasis, are associated with CKD progression and heart failure admissions after hospitalization in patients with and without AKI, we conducted a prospective cohort study to analyze the balance between angiopoietin-1 (Angpt-1), which maintains vessel stability, and angiopoietin-2 (Angpt-2), which increases vessel destabilization. Three months after discharge, we evaluated the associations between angiopoietins and development of the primary outcomes of CKD progression and heart failure and the secondary outcome of all-cause mortality 3 months after discharge or later., Results: Median age for the 1503 participants was 65.8 years; 746 (50%) had AKI. Compared with the lowest quartile, the highest quartile of the Angpt-1:Angpt-2 ratio was associated with 72% lower risk of CKD progression (adjusted hazard ratio [aHR], 0.28; 95% confidence interval [CI], 0.15 to 0.51), 94% lower risk of heart failure (aHR, 0.06; 95% CI, 0.02 to 0.15), and 82% lower risk of mortality (aHR, 0.18; 95% CI, 0.09 to 0.35) for those with AKI. Among those without AKI, the highest quartile of Angpt-1:Angpt-2 ratio was associated with 71% lower risk of heart failure (aHR, 0.29; 95% CI, 0.12 to 0.69) and 68% less mortality (aHR, 0.32; 95% CI, 0.15 to 0.68). There were no associations with CKD progression., Conclusions: A higher Angpt-1:Angpt-2 ratio was strongly associated with less CKD progression, heart failure, and mortality in the setting of AKI., (Copyright © 2022 by the American Society of Nephrology.)

SARS-CoV-2 is a newly identified coronavirus that causes the respiratory disease called coronavirus disease 2019 (COVID-19). With an urgent need for therapeutics, we lack a full understanding of the molecular basis of SARS-CoV-2-induced cellular damage and disease progression. Here, we conducted transcriptomic analysis of human PBMCs, identified significant changes in mitochondrial, ion channel, and protein quality-control gene products. SARS-CoV-2 proteins selectively target cellular organelle compartments, including the endoplasmic reticulum and mitochondria. M-protein, NSP6, ORF3A, ORF9C, and ORF10 bind to mitochondrial PTP complex components cyclophilin D, SPG-7, ANT, ATP synthase, and a previously undescribed CCDC58 (coiled-coil domain containing protein 58). Knockdown of CCDC58 or mPTP blocker cyclosporin A pretreatment enhances mitochondrial Ca 2+ retention capacity and bioenergetics. SARS-CoV-2 infection exacerbates cardiomyocyte autophagy and promotes cell death that was suppressed by cyclosporin A treatment. Our findings reveal that SARS-CoV-2 viral proteins suppress cardiomyocyte mitochondrial function that disrupts cardiomyocyte Ca 2+ cycling and cell viability., Competing Interests: All authors declare no competing interests., (© 2022 The Authors.)

Introduction: The Milan System for Reporting Salivary Gland Cytopathology (MSRSGC) specifies six categories with estimated risks of malignancy (ROM) and suggested management. The estimated ROM is 25% for Non-Diagnostic (ND) category, and 10% for Non-Neoplastic (NN). This study aimed to investigate histopathologic and clinical outcomes of MSRSGC categories ND and NN at the authors' institution., Materials and Methods: Cytopathology fine needle aspiration reports from 2008-2020 were searched for the word "salivary", "parotid", and "submandibular". Cases fitting Non-Diagnostic (ND) and Non-Neoplastic (NN) categories were identified. Follow-up cyto-/histopathologic and clinical data were extracted., Results: There were 43 ND and 46 NN cases. The average age was 58.3 years. Neoplastic lesions were found in 13 of 43 (30%) ND and 3 of 46 (6.5%) NN. The rate of malignancy in ND category was 14.0% (6/43) and 0% (0/46) in NN category. Four cases in ND (9.3%) and 6 (13.0%) in NN had no neoplasm and instead had an underlying reactive condition (e.g., chronic sialadenitis) or inflammatory lesion (e.g., lymphoepithelial cyst) on histologic follow-up. There was no follow-up pathology in 46.5% NDs (20/43) and 82.6% NNs (38/46); however, no lesions were apparent clinically with a mean follow-up of 3 years and 1.5 years, respectively., Conclusions: MSRSGC categories ND and NN are helpful for reporting salivary gland FNA results. With proper clinical and radiologic correlation, ROM of NN is low; however, ROM of ND remains significant. Repeat FNA after correlation for ND cases seems prudent as neoplasms and malignancies may have gone undetected., (Copyright © 2021 American Society of Cytopathology. Published by Elsevier Inc. All rights reserved.)

Diabetic kidney disease is the leading cause of kidney failure worldwide; in the USA, it accounts for over 50% of individuals entering dialysis or transplant programmes. Unlike other complications of diabetes, the prevalence of diabetic kidney disease has failed to decline over the past 30 years. Hyperglycaemia is the primary aetiological factor responsible for the development of diabetic kidney disease. Once hyperglycaemia becomes established, multiple pathophysiological disturbances, including hypertension, altered tubuloglomerular feedback, renal hypoxia, lipotoxicity, podocyte injury, inflammation, mitochondrial dysfunction, impaired autophagy and increased activity of the sodium-hydrogen exchanger, contribute to progressive glomerular sclerosis and the decline in glomerular filtration rate. The quantitative contribution of each of these abnormalities to the progression of diabetic kidney disease, as well as their role in type 1 and type 2 diabetes mellitus, remains to be determined. Sodium-glucose co-transporter 2 (SGLT2) inhibitors have a beneficial impact on many of these pathophysiological abnormalities; however, as several pathophysiological disturbances contribute to the onset and progression of diabetic kidney disease, multiple agents used in combination will likely be required to slow the progression of disease effectively.

Biological regulatory networks depend upon chemical interactions to process information. Engineering such molecular computing systems is a major challenge for synthetic biology and related fields. The chemical reaction network (CRN) model idealizes chemical interactions, allowing rigorous reasoning about the computational power of chemical kinetics. Here we focus on function computation with CRNs, where we think of the initial concentrations of some species as the input and the equilibrium concentration of another species as the output. Specifically, we are concerned with CRNs that are rate-independent (the computation must be correct independent of the reaction rate law) and composable ( f°g can be computed by concatenating the CRNs computing f and g). Rate independence and composability are important engineering desiderata, permitting implementations that violate mass-action kinetics, or even "well-mixedness", and allowing the systematic construction of complex computation via modular design. We show that to construct composable rate-independent CRNs, it is necessary and sufficient to ensure that the output species of a module is not a reactant in any reaction within the module. We then exactly characterize the functions computable by such CRNs as superadditive, positive-continuous, and piecewise rational linear. Thus composability severely limits rate-independent computation unless more sophisticated input/output encodings are used.

Background: Fine needle aspiration (FNA) of renal masses can distinguish between benign and malignant neoplasms in 73-94% of cases. Previous studies suggested the correct subclassification of renal cell carcinomas (RCCs) by cytomorphology can be achieved in up to 80% of cases. However, as RCCs become increasingly subclassified by molecular signatures, correct subclassification based on cytology alone is increasingly difficult., Design: Two FNA passes (2 stained with Diff-Quik® and 2 with the Papanicolaou method) were performed on all fresh nephrectomy specimens for a 1-year period. There were 30 cases in this study, with 29 primary renal tumors and 1 case of metastatic lung adenocarcinoma. Each case was assigned a random number and came with 2 slides (1 from each staining method). Eight cytopathologists were asked to provide a diagnosis and the World Health Organization/International Society of Urological Pathology (WHO/ISUP) grading if applicable. Fleiss' Kappa and Cohen's Kappa equations were used to look at inter-rater variability., Results: When compared to the surgical pathology diagnosis, the average percent correct diagnosis for all cytopathologist was 35%. Chromophobe RCCs had the best average percent accuracy at 72% followed by clearcell RCC at 48%. Average accuracy for grading RCCs was 40%. Inter-rater variability among the cytopathologists for all RCC diagnoses was fair with a Fleiss' Kappa coefficient of 0.28. For the WHO/ISUP grade, the weighted coefficient for each pathologist ranged from 0.11 to 0.45, ranging from fair to moderate, respectively., Conclusions: Renal tumors are difficult to classify on cytopathology alone. Core needle biopsy and ancillary studies are necessary if diagnosis will change management., (© 2021 S. Karger AG, Basel.)

Mg 2+ is the most abundant divalent cation in metazoans and an essential cofactor for ATP, nucleic acids, and countless metabolic enzymes. To understand how the spatio-temporal dynamics of intracellular Mg 2+ ( i Mg 2+ ) are integrated into cellular signaling, we implemented a comprehensive screen to discover regulators of i Mg 2+ dynamics. Lactate emerged as an activator of rapid release of Mg 2+ from endoplasmic reticulum (ER) stores, which facilitates mitochondrial Mg 2+ ( m Mg 2+ ) uptake in multiple cell types. We demonstrate that this process is remarkably temperature sensitive and mediated through intracellular but not extracellular signals. The ER-mitochondrial Mg 2+ dynamics is selectively stimulated by L-lactate. Further, we show that lactate-mediated m Mg 2+ entry is facilitated by Mrs2, and point mutations in the intermembrane space loop limits m Mg 2+ uptake. Intriguingly, suppression of m Mg 2+ surge alleviates inflammation-induced multi-organ failure. Together, these findings reveal that lactate mobilizes i Mg 2+ and links the m Mg 2+ transport machinery with major metabolic feedback circuits and mitochondrial bioenergetics., Competing Interests: Declaration of Interests M.M. is an inventor on a patent provisionally filed by UTHSA on lactate as a ligand for Mg(2+) dynamics in physiology and disease. All other authors have no financial interests to declare., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Fibrosis is the final common pathway in the pathophysiology of most forms of chronic kidney disease (CKD). As treatment of renal fibrosis still remains largely supportive, a refined understanding of the cellular and molecular mechanisms of kidney fibrosis and the development of novel compounds are urgently needed. Whether arginases play a role in the development of fibrosis in CKD is unclear. We hypothesized that endothelial arginase-2 (Arg2) promotes the development of kidney fibrosis induced by unilateral ureteral obstruction (UUO). Arg2 expression and arginase activity significantly increased following renal fibrosis. Pharmacologic blockade or genetic deficiency of Arg2 conferred kidney protection following renal fibrosis, as reflected by a reduction in kidney interstitial fibrosis and fibrotic markers. Selective deletion of Arg2 in endothelial cells (Tie2Cre/Arg2fl/fl) reduced the level of fibrosis after UUO. In contrast, selective deletion of Arg2 specifically in proximal tubular cells (Ggt1Cre/Arg2fl/fl) failed to reduce renal fibrosis after UUO. Furthermore, arginase inhibition restored kidney nitric oxide (NO) levels, oxidative stress, and mitochondrial function following UUO. These findings indicate that endothelial Arg2 plays a major role in renal fibrosis via its action on NO and mitochondrial function. Blocking Arg2 activity or expression could be a novel therapeutic approach for prevention of CKD.

Interleukin-10 (IL-10), a cytokine with anti-inflammatory effects, is produced by renal parenchymal cells and bone marrow derived cells. Both endogenous and exogenous IL-10 are protective in cisplatin-induced acute kidney injury. However, the source of endogenous IL-10 in cisplatin-induced nephrotoxicity is not clear. Bone marrow chimera experiments in IL10-KO mice indicated that bone marrow derived cells were the primary source of IL-10 in cisplatin nephrotoxicity. Cell specific deletion of IL-10 in T regulatory cells and dendritic cells was accomplished using Foxp3 and CD11c driven cre recombination in IL10flox/flox mice, respectively. Upon treatment with cisplatin, both the IL10flox/flox and the Foxp3YFP-Cre x IL10flox/flox mice developed similar degrees of kidney injury. However, mice with the dendritic cell deletion of IL-10 showed more severe structural and functional changes in the kidney compared to the IL10flox/flox mice. These results indicate that IL-10 from dendritic cells but not from T regulatory cells offers significant endogenous protection against cisplatin induced nephrotoxicity., Competing Interests: The authors have declared that no competing interests exist.

The tricarboxylic acid (TCA) cycle converts the end products of glycolysis and fatty acid β-oxidation into the reducing equivalents NADH and FADH 2 Although mitochondrial matrix uptake of Ca 2+ enhances ATP production, it remains unclear whether deprivation of mitochondrial TCA substrates alters mitochondrial Ca 2+ flux. We investigated the effect of TCA cycle substrates on MCU-mediated mitochondrial matrix uptake of Ca 2+ , mitochondrial bioenergetics, and autophagic flux. Inhibition of glycolysis, mitochondrial pyruvate transport, or mitochondrial fatty acid transport triggered expression of the MCU gatekeeper MICU1 but not the MCU core subunit. Knockdown of mitochondrial pyruvate carrier (MPC) isoforms or expression of the dominant negative mutant MPC1 R97W resulted in increased MICU1 protein abundance and inhibition of MCU-mediated mitochondrial matrix uptake of Ca 2+ We also found that genetic ablation of MPC1 in hepatocytes and mouse embryonic fibroblasts resulted in reduced resting matrix Ca 2+ , likely because of increased MICU1 expression, but resulted in changes in mitochondrial morphology. TCA cycle substrate-dependent MICU1 expression was mediated by the transcription factor early growth response 1 (EGR1). Blocking mitochondrial pyruvate or fatty acid flux was linked to increased autophagy marker abundance. These studies reveal a mechanism that controls the MCU-mediated Ca 2+ flux machinery and that depends on TCA cycle substrate availability. This mechanism generates a metabolic homeostatic circuit that protects cells from bioenergetic crisis and mitochondrial Ca 2+ overload during periods of nutrient stress., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Importance: Among patients who had acute kidney injury (AKI) during hospitalization, there is a need to improve risk prediction such that those at highest risk for subsequent loss of kidney function are identified for appropriate follow-up., Objective: To evaluate the association of post-AKI proteinuria with increased risk of future loss of renal function., Design, Setting, and Participants: The Assessment, Serial Evaluation, and Subsequent Sequelae in Acute Kidney Injury (ASSESS-AKI) Study was a multicenter prospective cohort study including 4 clinical centers in North America included 1538 patients enrolled 3 months after hospital discharge between December 2009 and February 2015., Exposures: Urine albumin-to-creatinine ratio (ACR) quantified 3 months after hospital discharge., Main Outcomes and Measures: Kidney disease progression defined as halving of estimated glomerular filtration rate (eGFR) or end-stage renal disease., Results: Of the 1538 participants, 769 (50%) had AKI durring hospitalization. The baseline study visit took place at a mean (SD) 91 (23) days after discharge. The mean (SD) age was 65 (13) years; the median eGFR was 68 mL/min/1.73 m2; and the median urine ACR was 15 mg/g. Overall, 547 (37%) study participants were women and 195 (13%) were black. After a median follow-up of 4.7 years, 138 (9%) participants had kidney disease progression. Higher post-AKI urine ACR level was associated with increased risk of kidney disease progression (hazard ratio [HR], 1.53 for each doubling; 95% CI, 1.45-1.62), and urine ACR measurement was a strong discriminator for future kidney disease progression (C statistic, 0.82). The performance of urine ACR was stronger in patients who had had AKI than in those who had not (C statistic, 0.70). A comprehensive model of clinical risk factors (eGFR, blood pressure, and demographics) including ACR provided better discrimination for predicting kidney disease progression after hospital discharge among those who had had AKI (C statistic, 0.85) vs those who had not (C statistic, 0.76). In the entire matched cohort, after taking into account urine ACR, eGFR, demographics, and traditional chronic kidney risk factors determined 3 months after discharge, AKI (HR, 1.46; 95% CI, 0.51-4.13 for AKI vs non-AKI) or severity of AKI (HR, 1.54; 95% CI, 0.50-4.72 for AKI stage 1 vs non-AKI; HR, 0.56; 95% CI, 0.07-4.84 for AKI stage 2 vs non-AKI; HR, 2.24; 95% CI, 0.33-15.29 for AKI stage 3 vs non-AKI) was not independently associated with more rapid kidney disease progression., Conclusions and Relevance: Proteinuria level is a valuable risk-stratification tool in the post-AKI period. These results suggest there should be more widespread and routine quantification of proteinuria after hospitalized AKI.

ANISEED (https://www.aniseed.cnrs.fr) is the main model organism database for the worldwide community of scientists working on tunicates, the vertebrate sister-group. Information provided for each species includes functionally-annotated gene and transcript models with orthology relationships within tunicates, and with echinoderms, cephalochordates and vertebrates. Beyond genes the system describes other genetic elements, including repeated elements and cis-regulatory modules. Gene expression profiles for several thousand genes are formalized in both wild-type and experimentally-manipulated conditions, using formal anatomical ontologies. These data can be explored through three complementary types of browsers, each offering a different view-point. A developmental browser summarizes the information in a gene- or territory-centric manner. Advanced genomic browsers integrate the genetic features surrounding genes or gene sets within a species. A Genomicus synteny browser explores the conservation of local gene order across deuterostome. This new release covers an extended taxonomic range of 14 species, including for the first time a non-ascidian species, the appendicularian Oikopleura dioica. Functional annotations, provided for each species, were enhanced through a combination of manual curation of gene models and the development of an improved orthology detection pipeline. Finally, gene expression profiles and anatomical territories can be explored in 4D online through the newly developed Morphonet morphogenetic browser., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: The aim of this study was to explore the effects of irisin on human visceral adipose tissue and adipocytes functions., Methods: Fresh human visceral white adipose tissues derived from 11 donors were used to examine the effects of irisin on browning, adipogenesis and osteogenesis gene expression, and anti-inflammatory properties. Preadipocytes were also used to examine the effects of irisin on mitochondrial respiration, adipogenic differentiation, and osteogenic differentiation., Key Results: Irisin significantly increased cellular mitochondrial energy metabolism in differentiated visceral adipocytes. Irisin also increased mRNA levels of transcriptional regulators of brite/beige adipocytes ( UCP-1, PGC1α, PRDM16, TMEM26, and CD137 ) in subcutaneous white adipose tissue but not in visceral/brown adipose tissue or their derived mature adipocytes. In parallel, irisin increased the protein levels of UCP-1 in subcutaneous white adipose tissue, but had no effect on the expression of this protein in visceral white adipose tissue and perirenal brown adipose tissue. However, irisin inhibited adipogenic differentiation, promoted osteogenic differentiation in visceral adipocytes, down-regulated adipogenesis, and upregulated osteogenesis genes expression in visceral fat tissue. Moreover, administration of irisin reduced the expression of proinflammatory marker mRNAs in both visceral and subcutaneous white adipose tissue., Conclusions: Our data suggest that (1) irisin may increase mitochondrial respiration and glycolysis in visceral adipocytes by a UCP-1 independent pathway; (2) irisin promotes anti-inflammatory activity on fat tissue., Competing Interests: None., (AJTR Copyright © 2019.)

The notochord of the invertebrate chordate Ciona forms a tapered rod at tailbud stages consisting of only 40 cylindrical cells in a single-file column. This tapered shape involves differences in notochord cell volume along the anterior-posterior axis. Here, we quantify sibling cell volume asymmetry throughout the developing notochord and find that there are distinctive patterns of unequal cleavage in all 4 bilateral pairs of A-line primary notochord founder cells and also in the B-line-derived secondary notochord founder cells. A quantitative model confirms that the observed patterns of unequal cleavage are sufficient to explain all the anterior-posterior variation in notochord cell volume. Many examples are known of cells that divide asymmetrically to give daughter cells of different size and fate. Here, by contrast, a series of subtle but iterative and finely patterned asymmetric divisions controls the shape of an entire organ. Quantitative 3D analysis of cell shape and spindle positioning allows us to infer multiple cellular mechanisms driving these unequal cleavages, including polarized displacements of the mitotic spindle, contributions from the shape of the mother cell, and late changes occurring between anaphase and abscission that potentially involve differential cortical contractility. We infer differential use of these mechanisms between different notochord blastomeres and also between different rounds of cell division. These results demonstrate a new role for asymmetric division in directly shaping a developing organ and point toward complex underlying mechanisms., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Fine-needle aspiration (FNA) is frequently utilized in the diagnostic workup of lymphadenopathy. We evaluated the correlation of cytopathology with flow cytometry and tissue biopsy results and assessed the prevalence of specific malignancies in young adults presenting with cervical lymphadenopathy., Methods: Database was searched for cervical lymph node FNA performed by a cytopathologist in patients aged 18-30 years from 2005 to 2017., Results: Cervical lymph node FNA was performed on 48 patients without prior history of malignancy. Nineteen patients had cytology results only, of which all were interpreted as benign reactive lymph node. None developed subsequent malignancies. The remaining 29 patients had cytology with flow cytometry and/or tissue biopsy results. A benign reactive cytology diagnosis was rendered in 18 (62%) cases, of which 11 had concordant diagnosis on flow cytometry, 2 had tissue biopsy, and 3 had both. Eleven (38%) patients had cytology results concerning for a hematologic malignancy, of which 7 were confirmed by flow cytometry and 3 by both flow cytometry and tissue biopsy. Cervical lymph node FNA has 94.1% sensitivity, 83.3% specificity, 88.9% positive predictive value, and 90.9% negative predictive value. The most common hematologic malignancy in our young adult population presenting with cervical lymphadenopathy was Hodgkin lymphoma., Conclusion: FNA is a useful first-line diagnostic procedure for assessing cervical lymphadenopathy in young adults to allow for better triage of specimens for flow cytometry and/or tissue biopsy concerning for a hematologic malignancy and potentially avoid invasive excisional biopsy in a proportion of cases., (© 2019 Wiley Periodicals, Inc.)

The gene regulatory networks underlying Ciona notochord fate specification and differentiation have been extensively investigated, but the regulatory basis for regionalized expression within the notochord is not understood. Here we identify three notochord-expressed genes, C11.331, C12.115 and C8.891, with strongly enriched expression in the secondary notochord cells at the posterior tip of the tail. C11.331 and C12.115 share a distinctive expression pattern that is highly enriched in the secondary notochord lineage but also graded within that lineage with the strongest expression at the posterior tip. Both genes show similar responses to pharmacological perturbations of Wnt and FGF signaling, consistent with an important role for Wnt and FGF ligands expressed at the tail tip. Reporter analysis indicates that the C11.331 cis-regulatory regions are extensively distributed, with multiple non-overlapping regions conferring posterior notochord-enriched expression. Fine-scale analysis of a minimal cis-regulatory module identifies discrete positive and negative elements including a strong silencer. Truncation of the silencer region leads to increased expression in the primary notochord, indicating that C11.331 expression is influenced by putative regulators of primary versus secondary notochord fate. The minimal CRM contains predicted ETS, GATA, LMX and Myb sites, all of which lead to reduced expression in secondary notochord when mutated. These results show that the posterior-enriched notochord expression of C11.331 depends on multiple inputs, including Wnt and FGF signals from the tip of the tail, multiple notochord-specific regulators, and yet-to-be identified regulators of regional identity within the notochord., (Copyright © 2018. Published by Elsevier Inc.)

Acute kidney injury (AKI) is common among hospitalized patients and is associated with high morbidity and mortality. Inflammation is recognized to play an important role in both ischemic and toxic models of AKI. Cisplatin is a widely used and highly effective cancer chemotherapeutic agent but carries the risk of nephrotoxicity. We have used a model of cisplatin-induced AKI to explore the functions of the innate immune response in kidney injury. Several components of innate immunity, such as Toll-like receptor sensing and inflammatory cytokine production, contribute to both ischemic and cisplatin-induced AKI. Importantly, it is the activity of these components in kidney parenchymal cells, rather than immune cells, which mediate AKI. Cellular components of innate immunity, such as neutrophils and dendritic cells, appear to play disparate roles in ischemic vs toxic AKI. Innate immune pathways could be targeted to prevent or treat AKI., Competing Interests: Potential Conflicts of Interest: None disclosed.

Urinary biomarkers are used increasingly for sensitive prediction of kidney injury in preclinical and clinical studies. Given the frequent requirement of anesthesia in various animal models of disease, it is important to define the effects of anesthesia on kidney injury biomarkers to guide the appropriate selection of anesthetic agents and to avoid potential confounders in the interpretation of data. Therefore, we performed a prospective study using male C57BL/6J mice ( n = 45) exposed to a single anesthetic episode to determine the effects several common anesthesia regimens on the urinary excretion of 2 commonly used kidney injury biomarkers: hepatitis A virus cellular receptor 1 (HAVCR1, also known as KIM1) and lipocalin 2 (LCN2, also known as NGAL). We evaluated 3 injectable regimens (ketamine-xylazine, tiletamine-zolazepam, and pentobarbital) and 2 inhalational agents (isoflurane and sevoflurane). Concentrations of HAVCR1 and LCN2 in urine collected at various time points after anesthesia were measured by using ELISA. Administration of ketamine-xylazine resulted in a significant increase in HAVCR1 levels at 6 h after anesthesia but a decrease in LCN2 levels compared with baseline. LCN2 levels steadily increased over the first 24 h after inhalant anesthesia, with a significant increase at 24 h after sevoflurane. These results suggest that injectable anesthesia had early effects on HAVCR1 and LCN2 levels, whereas inhalational agents increased these biomarkers over prolonged time.

Background: External beam irradiation is an accepted treatment for skeletal malignancies. Radiation acts on both cancerous and normal cells and, depending on the balance of these effects, may promote or impair bone healing after pathologic fracture. Previous studies suggest an adverse effect of radiation on endochondral ossification, but the existence of differential effects of radiation on the two distinct bone healing pathways is unknown., Questions/purposes: The purpose of this study was to investigate the differential effects of external beam irradiation on endochondral compared with intramembranous ossification with intramedullary nail and plate fixation of fractures inducing the two respective osseous healing pathways through assessment of (1) bone biology by histomorphometric analysis of cartilage area and micro-CT volumetric assessment of the calcified callus; and (2) mechanical properties of the healing fracture by four-point bending failure analysis of bending stiffness and strength., Methods: Thirty-six male Sprague-Dawley rats underwent bilateral iatrogenic femur fracture: one side was repaired with an intramedullary nail and the other with compression plating. Three days postoperatively, half (n = 18) received 8-Gray external beam irradiation to each fracture. Rodents were euthanized at 1, 2, and 4 weeks postoperatively (n = 3/group) for quantitative histomorphometry of cartilage area and micro-CT assessment of callus volume. The remaining rodents were euthanized at 3 months (n = 9/group) and subjected to four-point bending tests to assess stiffness and maximum strength., Results: Nailed femurs that were irradiated exhibited a reduction in cartilage area at both 2 weeks (1.08 ± 1.13 mm versus 37.32 ± 19.88 mm; 95% confidence interval [CI] of the difference, 4.32-68.16 mm; p = 0.034) and 4 weeks (4.60 ± 3.97 mm versus 39.10 ± 16.28 mm; 95% CI of the difference, 7.64-61.36 mm; p = 0.023) compared with nonirradiated fractures. There was also a decrease in the volume ratio of calcified callus at 4 weeks (0.35 ± 0.08 versus 0.51 ± 0.05; 95% CI of the difference, 0.01-0.31; p = 0.042) compared with nonirradiated fractures. By contrast, there was no difference in cartilage area or calcified callus between irradiated and nonirradiated plated femurs. The stiffness (128.84 ± 76.60 N/mm versus 26.99 ± 26.07 N/mm; 95% CI of the difference, 44.67-159.03 N/mm; p = 0.012) and maximum strength (41.44 ± 22.06 N versus 23.75 ± 11.00 N; 95% CI of the difference, 0.27-35.11 N; p = 0.047) of irradiated plated femurs was greater than the irradiated nailed femurs. However, for nonirradiated femurs, the maximum strength of nailed fractures (36.05 ± 17.34 N versus 15.63 ± 5.19 N; 95% CI of the difference, 3.96-36.88 N; p = 0.022) was greater than plated fractures, and there was no difference in stiffness between the nailed and plated fractures., Conclusions: In this model, external beam irradiation was found to preferentially inhibit endochondral over intramembranous ossification with the greatest impairment in healing of radiated fractures repaired with intramedullary nails compared with those fixed with plates. Future work with larger sample sizes might focus on further elucidating the observed differences in mechanical properties., Clinical Relevance: This work suggests that there may be a rationale for compression plating rather than intramedullary nailing of long bone fractures in select circumstances where bony union is desirable, adjunctive radiation treatment is required, and bone stock is sufficient for plate and screw fixation.

The Myxomavirus-derived protein Serp-1 has potent anti-inflammatory activity in models of vasculitis, lupus, viral sepsis, and transplant. Serp-1 has also been tested successfully in a Phase IIa clinical trial in unstable angina, representing a "first-in-class" therapeutic. Recently, peptides derived from the reactive center loop (RCL) have been developed as stand-alone therapeutics for reducing vasculitis and improving survival in MHV68-infected mice. However, both Serp-1 and the RCL peptides lose activity in MHV68-infected mice after antibiotic suppression of intestinal microbiota. Here, we utilize a structure-guided approach to design and test a series of next-generation RCL peptides with improved therapeutic potential that is not reduced when the peptides are combined with antibiotic treatments. The crystal structure of cleaved Serp-1 was determined to 2.5 Å resolution and reveals a classical serpin structure with potential for serpin-derived RCL peptides to bind and inhibit mammalian serpins, plasminogen activator inhibitor 1 (PAI-1), anti-thrombin III (ATIII), and α-1 antitrypsin (A1AT), and target proteases. Using in silico modeling of the Serp-1 RCL peptide, S-7, we designed several modified RCL peptides that were predicted to have stronger interactions with human serpins because of the larger number of stabilizing hydrogen bonds. Two of these peptides (MPS7-8 and -9) displayed extended activity, improving survival where activity was previously lost in antibiotic-treated MHV68-infected mice (P < 0.0001). Mass spectrometry and kinetic assays suggest interaction of the peptides with ATIII, A1AT, and target proteases in mouse and human plasma. In summary, we present the next step toward the development of a promising new class of anti-inflammatory serpin-based therapeutics.

Ischemia/reperfusion is a common cause of acute kidney injury (AKI). However, mechanisms underlying the sudden loss in kidney function and tissue injury remain to be fully elucidated. Here, we investigated the role of peptidyl arginine deiminase-4 (PAD4), which converts arginine to citrulline and plays a role in epigenetic regulation and inflammation, in renal ischemia/reperfusion injury. PAD4 expression was highly induced in infiltrating leukocytes 24 hours following renal ischemia and reperfusion. This induction was accompanied by citrullination of histone H3 and formation of neutrophil extracellular traps in kidneys of wild-type mice. By contrast, PAD4-deficient mice did not form neutrophil extracellular traps, expressed lower levels of pro-inflammatory cytokines and were partially protected from renal ischemia/reperfusion-induced AKI. Furthermore, PAD4-deficient mice recovered kidney function 48 hours after ischemia/reperfusion, whereas kidney function in the wild-type mice progressively worsened. Administration of DNase I, which degrades neutrophil extracellular traps or the PAD-specific inhibitor YW3-56 before ischemia, partially prevented renal ischemia/reperfusion-induced AKI. Notably, transfer of neutrophils from wild-type, but not from PAD4-deficient mice, was sufficient to restore renal neutrophil extracellular trap formation and impair kidney function following renal ischemia/reperfusion. Thus, neutrophil PAD4 plays a pivotal role in renal ischemia/reperfusion-induced AKI., (Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

ANISEED (www.aniseed.cnrs.fr) is the main model organism database for tunicates, the sister-group of vertebrates. This release gives access to annotated genomes, gene expression patterns, and anatomical descriptions for nine ascidian species. It provides increased integration with external molecular and taxonomy databases, better support for epigenomics datasets, in particular RNA-seq, ChIP-seq and SELEX-seq, and features novel interactive interfaces for existing and novel datatypes. In particular, the cross-species navigation and comparison is enhanced through a novel taxonomy section describing each represented species and through the implementation of interactive phylogenetic gene trees for 60% of tunicate genes. The gene expression section displays the results of RNA-seq experiments for the three major model species of solitary ascidians. Gene expression is controlled by the binding of transcription factors to cis-regulatory sequences. A high-resolution description of the DNA-binding specificity for 131 Ciona robusta (formerly C. intestinalis type A) transcription factors by SELEX-seq is provided and used to map candidate binding sites across the Ciona robusta and Phallusia mammillata genomes. Finally, use of a WashU Epigenome browser enhances genome navigation, while a Genomicus server was set up to explore microsynteny relationships within tunicates and with vertebrates, Amphioxus, echinoderms and hemichordates., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The U.S. Air Force conducts vector and arbovirus surveillance worldwide. We report on two Rhabdoviridae detected at Holloman Air Force Base in Otero County, New Mexico including the nearly complete 11-kb genome of Merida virus, which was not previously known from the USA, and a novel virus tentatively named Dillard's Draw virus. Merida virus was previously only known from Mexico. Dillard's Draw virus represents a novel arbovirus most closely related to an avian pathogen from Australia and shares some genetic similarity to Durham virus from the eastern United States.

Background: Acute kidney injury (AKI) is a complex disease burdened by uncertainties of definition, management strategies, and prognosis. This study explores the relationship between demographic characteristics of nephrologists and their perceptions about the definition, management, and follow-up of AKI., Methods: We developed a Web-based survey, the International Survey on Acute Kidney Injury (ISAKI), consisting of 29 items in 4 categories: (1) demographic and practice characteristics, (2) definition of AKI, (3) management of renal replacement therapy (RRT) in AKI, and (4) sequelae of AKI. A multivariable stepwise logistic regression model was used to examine relationships between the dependent variables and the demographic characteristics of the respondents., Results: Responses from 743 nephrologists from 90 countries were analyzed. The majority (60%) of respondents reported using RIFLE and/or AKIN criteria regularly to define AKI, although US nephrologists were less likely to do so (OR: 0.58; 95% CI: 0.42-0.85). The most common initial RRT modality was intermittent hemodialysis (63.5%), followed by continuous RRT (23.8%). Faculty affiliation was associated with a higher likelihood of using a dialysis schedule of ≥4 times a week (OR: 1.75; 95% CI: 1.20-2.55). The respondents believed that a single episode of AKI increases the likelihood of development of chronic kidney disease (CKD) (55%), subsequent AKI (36%), and rapid progression of preexisting CKD (87%). US nephrologists were less likely to recommend follow-up after resolution of AKI (OR: 0.15; 95% CI: 0.07-0.33)., Conclusions: Our findings highlight the need for a widely accepted consensus definition of AKI, a uniform approach to management, and improved follow-up after resolution of AKI episodes.

Novel therapeutic interventions for preventing or attenuating kidney injury following ischemia-reperfusion injury (IRI) remain a focus of significant interest. Currently, there are no definitive therapeutic or preventive approaches available for ischemic acute kidney injury (AKI). Our objective is to determine 1 ) whether renal arginase activity or expression is increased in renal IRI, and 2 ) whether arginase plays a role in development of renal IRI. The impact of arginase activity and expression on renal damage was evaluated in male C57BL/6J (wild type) and arginase-2 (ARG2)-deficient ( Arg2 -/- ) mice subjected to bilateral renal ischemia for 28 min, followed by reperfusion for 24 h. ARG2 expression and arginase activity significantly increased following renal IRI, paralleling the increase in kidney injury. Pharmacological blockade or genetic deficiency of Arg2 conferred kidney protection in renal IRI. Arg2 -/- mice had significantly attenuated kidney injury and lower plasma creatinine and blood urea nitrogen levels after renal IRI. Blocking arginases using S -(2-boronoethyl)-l-cysteine (BEC) 18 h before ischemia mimicked arginase deficiency by reducing kidney injury, histopathological changes and kidney injury marker-1 expression, renal apoptosis, kidney inflammatory cell recruitment and inflammatory cytokines, and kidney oxidative stress; increasing kidney nitric oxide (NO) production and endothelial NO synthase (eNOS) phosphorylation, kidney peroxisome proliferator-activated receptor-γ coactivator-1α expression, and mitochondrial ATP; and preserving kidney mitochondrial ultrastructure compared with vehicle-treated IRI mice. Importantly, BEC-treated eNOS-knockout mice failed to reduce blood urea nitrogen and creatinine following renal IRI. These findings indicate that ARG2 plays a major role in renal IRI, via an eNOS-dependent mechanism, and that blocking ARG2 activity or expression could be a novel therapeutic approach for prevention of AKI., (Copyright © 2017 the American Physiological Society.)

Inflammation is an important mediator of most forms of acute kidney injury (AKI). Although neutrophils are prominent components of the inflammatory cascade, the precise role of neutrophils in AKI and the mechanisms by which they contribute to AKI remain controversial. In this issue, Deng et al. identify an important cross talk between renal epithelial cells and neutrophils involving the production and action of leukotriene B4 in mediating cisplatin AKI. We discuss the possible explanations for the discrepant findings that have been reported for neutrophils in cisplatin AKI., (Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Fructose consumption has been linked to hypertension in animal models and human studies, and endogenous fructose metabolism has been shown to promote acute and chronic kidney injury in mice. A recent study published in Nature Communications demonstrates a reduction in ischemic acute kidney injury with genetic knockout or inhibition of fructokinase, which catalyzes the first step in fructose metabolism. Although the role of this pathway in human kidney disease remains unclear, the recent description of several candidate fructokinase inhibitors may allow for clinical studies in the future., (Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Inflammation is a central pathophysiologic mechanism that contributes to diabetes mellitus and diabetic nephropathy. Recently, we showed that macrophages directly contribute to diabetic renal injury and that pharmacological blockade or genetic deficiency of chemokine (C-C motif) receptor 2 (CCR2) confers kidney protection in diabetic nephropathy. However, the direct role of CCR2 in kidney-derived cells such as podocytes in diabetic nephropathy remains unclear. To study this, we developed a transgenic mouse model expressing CCR2 specifically in podocytes (Tg[NPHS2-Ccr2]) on a nephropathy-prone (DBA/2J) and CCR2-deficient (Ccr2 -/- ) background with heterozygous Ccr2 +/- littermate controls. Diabetes was induced by streptozotocin. As expected, absence of CCR2 conferred kidney protection after nine weeks of diabetes. In contrast, transgenic CCR2 overexpression in the podocytes of Ccr2 -/- mice resulted in significantly increased albuminuria, blood urea nitrogen, histopathologic changes, kidney fibronectin and type 1 collagen expression, podocyte loss, and glomerular apoptosis after nine weeks of streptozotocin-induced diabetes. Interestingly, there was no concurrent increase in kidney macrophage recruitment or inflammatory cytokine levels in the mice. These findings support a direct role for CCR2 expression in podocytes to mediate diabetic renal injury, independent of monocyte/macrophage recruitment. Thus, targeting the CCR2 signaling cascade in podocytes could be a novel therapeutic approach for treatment of diabetic nephropathy., (Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Protein biomarkers, especially cytokines, play a pivotal role in the diagnosis and treatment of a wide spectrum of diseases. Therefore, a critical need for advanced cytokine sensors has been rapidly growing and will continue to expand to promote clinical testing, new biomarker development, and disease studies. In particular, sensors employing transduction principles of various optical modalities have emerged as the most common means of detection. In typical cytokine assays which are based on the binding affinities between the analytes of cytokines and their specific antibodies, optical schemes represent the most widely used mechanisms, with some serving as the gold standard against which all existing and new sensors are benchmarked. With recent advancements in nanoscience and nanotechnology, many of the recently emerging technologies for cytokine detection exploit various forms of nanomaterials for improved sensing capabilities. Nanomaterials have been demonstrated to exhibit exceptional optical properties unique to their reduced dimensionality. Novel sensing approaches based on the newly identified properties of nanomaterials have shown drastically improved performances in both the qualitative and quantitative analyses of cytokines. This article brings together the fundamentals in the literature that are central to different optical modalities developed for cytokine detection. Recent advancements in the applications of novel technologies are also discussed in terms of those that enable highly sensitive and multiplexed cytokine quantification spanning a wide dynamic range. For each highlighted optical technique, its current detection capabilities as well as associated challenges are discussed. Lastly, an outlook for nanomaterial-based cytokine sensors is provided from the perspective of optimizing the technologies for sensitivity and multiplexity as well as promoting widespread adaptations of the emerging optical techniques by lowering high thresholds currently present in the new approaches.

Human noroviruses are a leading cause of gastroenteritis, and so, vaccine development is desperately needed. Elucidating viral mechanisms of immune antagonism can provide key insight into designing effective immunization platforms. We recently revealed that B cells are targets of norovirus infection. Because noroviruses can regulate antigen presentation by infected macrophages and B cells can function as antigen-presenting cells, we tested whether noroviruses regulate B-cell-mediated antigen presentation and the biological consequence of such regulation. Indeed, murine noroviruses could prevent B-cell expression of antigen presentation molecules and this directly correlated with impaired control of acute infection. In addition to B cells, acute control required MHC class I molecules, CD8 + T cells, and granzymes, supporting a model whereby B cells act as antigen presenting cells to activate cytotoxic CD8 + T cells. This immune pathway was active prior to the induction of antiviral antibody responses. As in macrophages, the minor structural protein VP2 regulated B-cell antigen presentation in a virus-specific manner. Commensal bacteria were not required for the activation of this pathway and ultimately only B cells were required for the clearance of viral infection. These findings provide new insight into the role of B cells in stimulating antiviral CD8 + T-cell responses.

Background: Although stored urine samples are often used in biomarker studies focused on acute and chronic kidney disease, how storage time impacts biomarker levels is not well understood., Methods: 866 subjects enrolled in the NIDDK-sponsored ASsessment, Serial Evaluation, and Subsequent Sequelae in Acute Kidney Injury (ASSESS-AKI) Study were included. Samples were processed under standard conditions and stored at -70°C until analyzed. Kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), interleukin-18 (IL-18), and liver fatty acid binding protein (L-FABP) were measured in urine samples collected during the index hospitalization or an outpatient visit 3 months later. Mixed effects models were used to determine the effect of storage time on biomarker levels and stratified by visit., Results: Median storage was 17.8 months (25-75% IQR 10.6-23.7) for samples from the index hospitalization and 14.6 months (IQR 7.3-20.4) for outpatient samples. In the mixed effects models, the only significant association between storage time and biomarker concentration was for KIM-1 in outpatient samples, where each month of storage was associated with a 1.7% decrease (95% CI -3% to -0.3%). There was no relationship between storage time and KIM-1 levels in samples from the index hospitalization., Conclusion: There was no significant impact of storage time over a median of 18 months on urine KIM-1, NGAL, IL-18 or L-FABP in hospitalized samples; a statistically significant effect towards a decrease over time was noted for KIM-1 in outpatient samples. Additional studies are needed to determine whether longer periods of storage at -70°C systematically impact levels of these analytes., Competing Interests: W. Brian Reeves is an Associate Editor of Kidney International, a journal of the International Society of Nephrology. Kathleen D. Liu and Chi-yuan Hsu had reagents donated for assays by Abbott Laboratories. Prasad Devarajan has patents submitted for the use of NGAL as a biomarker of kidney injury. T. Alp Ikizler is the Chair of the Medical Advisory Board for DSI, Inc, the Chair of the Scientific Advisory Board for Satellite Health, has received honoraria from Hospira for an advisory board, from Fresenius-Kabi for lecturing and for advisory boards, and from the American Board of Internal Medicine as a member of the Nephrology subspecialty committee. Jonathan Himmelfarb is the chair of the steering committee for a clinical trial for Thrasos, Inc. James Kaufman is the study director for a large randomized clinical trial for the Department of Veterans Affairs Cooperative Study Program. Because this is an ongoing study of human subjects with confidential data, the data cannot be shared at present because of patient confidentiality. At the end of the parent ASSESS-AKI study, a de-identified data set will be prepared in compliance with NIH guidelines and made publicly available. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

In rodent models, female sex has been shown to be protective against ischemia-reperfusion injury. A recent publication suggests that this sex-dependent response to injury may have clinical implications for delayed graft function after kidney transplantation., (Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Determining ultratrace amounts of protein biomarkers in patient samples in a straightforward and quantitative manner is extremely important for early disease diagnosis and treatment. Here, we successfully demonstrate the novel use of zinc oxide nanorods (ZnO NRs) in the ultrasensitive and quantitative detection of two acute kidney injury (AKI)-related protein biomarkers, tumor necrosis factor (TNF)-α and interleukin (IL)-8, directly from patient samples. We first validate the ZnO NRs-based IL-8 results via comparison with those obtained from using a conventional enzyme-linked immunosorbent method in samples from 38 individuals. We further assess the full detection capability of the ZnO NRs-based technique by quantifying TNF-α, whose levels in human urine are often below the detection limits of conventional methods. Using the ZnO NR platforms, we determine the TNF-α concentrations of all 46 patient samples tested, down to the fg per mL level. Subsequently, we screen for TNF-α levels in approximately 50 additional samples collected from different patient groups in order to demonstrate a potential use of the ZnO NRs-based assay in assessing cytokine levels useful for further clinical monitoring. Our research efforts demonstrate that ZnO NRs can be straightforwardly employed in the rapid, ultrasensitive, quantitative, and simultaneous detection of multiple AKI-related biomarkers directly in patient urine samples, providing an unparalleled detection capability beyond those of conventional analysis methods. Additional key advantages of the ZnO NRs-based approach include a fast detection speed, low-volume assay condition, multiplexing ability, and easy automation/integration capability to existing fluorescence instrumentation. Therefore, we anticipate that our ZnO NRs-based detection method will be highly beneficial for overcoming the frequent challenges in early biomarker development and treatment assessment, pertaining to the facile and ultrasensitive quantification of hard-to-trace biomolecules.

The design and development of a calorimetric biosensing system enabling relatively high throughput sample analysis are reported. The calorimetric biosensor system consists of a thin (∼20 μm) micromachined Y-cut quartz crystal resonator (QCR) as a temperature sensor placed in close proximity to a fluidic chamber packed with an immobilized enzyme. Layer by layer enzyme immobilization of urease is demonstrated and its activity as a function of the number of layers, pH, and time has been evaluated. This configuration enables a sensing system where a transducer element is physically separated from the analyte solution of interest and is thereby free from fouling effects typically associated with biochemical reactions occuring on the sensor surface. The performance of this biosensing system is demonstrated by detection of 1-200 mM urea in phosphate buffer via a flow injection analysis (FIA) technique. Miniaturized fluidic systems were used to provide continuous flow through a reaction column. Under this configuration the biosensor has an ultimate resolution of less than 1 mM urea and showed a linear response between 0-50 mM. This work demonstrates a sensing modality in which the sensor itself is not fouled or contaminated by the solution of interest and the enzyme immobilized Kapton® fluidic reaction column can be used as a disposable cartridge. Such a system enables reuse and reliability for long term sampling measurements. Based on this concept a biosensing system is envisioned which can perform rapid measurements to detect biomarkers such as glucose, creatinine, cholesterol, urea and lactate in urine and blood continuously over extended periods of time.

Monocyte/macrophage recruitment correlates strongly with the progression of diabetic nephropathy. Tumor necrosis factor-α (TNF-α) is produced by monocytes/macrophages but the direct role of TNF-α and/or macrophage-derived TNF-α in the progression of diabetic nephropathy remains unclear. Here we tested whether inhibition of TNF-α confers kidney protection in diabetic nephropathy via a macrophage-derived TNF-α-dependent pathway. Compared to vehicle-treated mice, blockade of TNF-α with a murine anti-TNF-α antibody conferred kidney protection in Ins2(Akita) mice as indicated by reductions in albuminuria, plasma creatinine, histopathologic changes, kidney macrophage recruitment, and plasma inflammatory cytokine levels at 18 weeks of age. To assess the direct role of macrophage-derived TNF-α in diabetic nephropathy, we generated macrophage-specific TNF-α-deficient mice (CD11b(Cre)/TNF-α(Flox/Flox)). Conditional ablation of TNF-α in macrophages significantly reduced albuminuria, the increase in plasma creatinine and blood urea nitrogen, histopathologic changes, and kidney macrophage recruitment compared to diabetic TNF-α(Flox/Flox) control mice after 12 weeks of streptozotocin-induced diabetes. Thus, production of TNF-α by macrophages plays a major role in diabetic renal injury. Hence, blocking TNF-α could be a novel therapeutic approach for treatment of diabetic nephropathy.

Cisplatin is a very effective chemotherapeutic agent used against a wide range of solid tumors. A major adverse effect of cisplatin therapy is acute kidney injury (AKI). Neutrophils are reported to infiltrate and exacerbate injury in a wide range of sterile inflammatory models of tissue injury. Here, we studied the kinetics of neutrophil infiltration into kidneys and their role in cisplatin-mediated AKI. Mice treated with cisplatin showed an increase in circulating neutrophils 24 and 48 h after cisplatin administration. Cisplatin treatment caused an increase in kidney leukocytes with neutrophils accounting for the majority of the infiltrating leukocytes. The extent of neutrophil infiltration coincided with the severity of kidney injury and renal dysfunction. To examine the functional relevance of infiltrating neutrophils in cisplatin nephrotoxicity, we depleted neutrophils using a neutrophil-specific antibody (anti-Ly-6G). This antibody resulted in greater than 90% depletion of neutrophils in both the blood and kidney. Of note, depletion of neutrophils had no impact on the extent of cisplatin-induced AKI as compared to non-depleted mice. Earlier, we reported that dendritic cell depletion in CD11c-DTRtg mice causes exacerbation of AKI and a dramatic increase in renal neutrophils. Thus, we also examined the role of neutrophils in dendritic cell-depleted mice treated with cisplatin. Dendritic cell depletion exacerbated AKI in spite of neutrophil depletion. These data demonstrate that cisplatin nephrotoxicity is not mediated by neutrophils and that dendritic cells protect kidneys via neutrophil-independent mechanisms.

Zhou et al. identify progranulin (PGRN) as a protective mediator that limits inflammation in murine models of acute kidney injury (AKI) and reduces its severity. Deficiency of PGRN was associated with increased inflammation and increased injury in ischemic and nephrotoxic models of AKI. Exogenous PGRN reduced AKI even when administered after AKI was established. Interference in NOD2 pathways is suggested as a possible mechanism for protection. PGRN-based therapeutics might have application in the treatment or prevention of AKI.

The Ciona notochord displays planar cell polarity (PCP), with anterior localization of Prickle (Pk) and Strabismus (Stbm). We report that a myosin is polarized anteriorly in these cells and strongly colocalizes with Stbm. Disruption of the actin/myosin machinery with cytochalasin or blebbistatin disrupts polarization of Pk and Stbm, but not of myosin complexes, suggesting a PCP-independent aspect of myosin localization. Wash out of cytochalasin restored Pk polarization, but not if done in the presence of blebbistatin, suggesting an active role for myosin in core PCP protein localization. On the other hand, in the pk mutant line, aimless, myosin polarization is disrupted in approximately one third of the cells, indicating a reciprocal action of core PCP signaling on myosin localization. Our results indicate a complex relationship between the actomyosin cytoskeleton and core PCP components in which myosin is not simply a downstream target of PCP signaling, but also required for PCP protein localization.