ROR2/PCP a New Pathway Controlling Endothelial Cell Polarity Under Flow Conditions.

BACKGROUND: Endothelial cells (ECs) are sensitive to physical forces created by blood flow, especially to laminar shear stress. Among the cell responses to laminar flow, EC polarization against the flow direction emerges as a key event, particularly during the development and remodeling of the vascular network. EC adopt an elongated planar cell shape with an asymmetrical distribution of intracellular organelles along the axis of blood flow. This study aimed to investigate the involvement of planar cell polarity via the receptor ROR2 (receptor tyrosine kinase-like orphan receptor 2) in endothelial responses to laminar shear stress. METHODS: We generated a genetic mouse model with EC-specific deletion of Ror2, in combination with in vitro approaches involving loss- and gain-of-function experiments. RESULTS: During the first 2 weeks of life, the endothelium of the mouse aorta undergoes a rapid remodeling associated with a loss of EC polarization against the flow direction. Notably, we found a correlation between ROR2 expression and endothelial polarization levels. Our findings demonstrate that deletion of Ror2 in murine ECs impaired their polarization during the postnatal development of the aorta. In vitro experiments further validated the essential role of ROR2 in both EC collective polarization and directed migration under laminar flow conditions. Exposure to laminar shear stress triggered the relocalization of ROR2 to cell-cell junctions where it formed a complex with VE-Cadherin and ß-catenin, thereby regulating adherens junctions remodeling at the rear and front poles of ECs. Finally, we showed that adherens junctions remodeling and cell polarity induced by ROR2 were dependent on the activation of the small GTPase Cdc42. CONCLUSIONS: This study identified ROR2/planar cell polarity pathway as a new mechanism controlling and coordinating collective polarity patterns of EC during shear stress response.

Impact of Albumin Assays in the Diagnosis of Malnutrition in Hemodialysis Patients: A Cohort Study.

OBJECTIVE: In hemodialysis (HD) patients, malnutrition should be diagnosed by several assessment tools including a plasma albumin concentration of less than 3.8 g/dL or 3.5 g/dL using bromocresol green or immunonephelometry (IN), respectively. However, albumin measurement is not yet standardized and two alternative methods are also commonly used in laboratories: bromocresol purple (BCP) and immunoturbidimetry (IT). This study aimed to revisit the hypoalbuminemia thresholds for BCP and IT, in HD patients. METHODS: Plasma albumin was measured by the four analytical methods during the monthly HD nutritional assessment of 103 prospectively included patients. RESULTS: Significant differences in albumin levels were observed in HD patients depending on the method used. Using BCP or IT with the cut-off at 3.5 g/dL (determined for the general population) we obtained 33% and 9.7% of false hypoalbuminemia in comparison to IN (mean bias of -0.4 g/dL and -0.065 g/dL, respectively). The best hypoalbuminemia threshold for BCP was 3.05 g/dL and 3.4 g/dL for IT. Twenty percent of HD patients were classified as malnourished when albumin was determined by IN. Similar rates were obtained using the new hypoalbuminemia cut-offs for BCP (18.5%) and IT (19.5%). CONCLUSION: To avoid nutritional misclassification of HD patients, we should adjust hypoalbuminemia thresholds when BCP or IT methods are used in laboratories.

Increased Capillary Permeability in Heart Induces Diastolic Dysfunction Independently of Inflammation, Fibrosis, or Cardiomyocyte Dysfunction.

BACKGROUND: While endothelial dysfunction is suggested to contribute to heart failure with preserved ejection fraction pathophysiology, understanding the importance of the endothelium alone, in the pathogenesis of diastolic abnormalities has not yet been fully elucidated. Here, we investigated the consequences of specific endothelial dysfunction on cardiac function, independently of any comorbidity or risk factor (diabetes or obesity) and their potential effect on cardiomyocyte. METHODS: The ubiquitine ligase Pdzrn3, expressed in endothelial cells (ECs), was shown to destabilize tight junction. A genetic mouse model in which Pdzrn3 is overexpressed in EC (iEC-Pdzrn3) in adults was developed. RESULTS: EC-specific Pdzrn3 expression increased cardiac leakage of IgG and fibrinogen blood-born molecules. The induced edema demonstrated features of diastolic dysfunction, with increased end-diastolic pressure, alteration of dP/dt min, increased natriuretic peptides, in addition to limited exercise capacity, without major signs of cardiac fibrosis and inflammation. Electron microscopic images showed edema with disrupted EC-cardiomyocyte interactions. RNA sequencing analysis of gene expression in cardiac EC demonstrated a decrease in genes coding for endothelial extracellular matrix proteins, which could be related to the fragile blood vessel phenotype. Irregularly shaped capillaries with hemorrhages were found in heart sections of iEC-Pdzrn3 mice. We also found that a high-fat diet was not sufficient to provoke diastolic dysfunction; high-fat diet aggravated cardiac inflammation, associated with an altered cardiac metabolic signature in EC-Pdzrn3 mice, reminiscent of heart failure with preserved ejection fraction features. CONCLUSIONS: An increase of endothelial permeability is responsible for mediating diastolic dysfunction pathophysiology and for aggravating detrimental effects of a high-fat diet on cardiac inflammation and metabolism.

PHACTR-1 (Phosphatase and Actin Regulator 1) Deficiency in Either Endothelial or Smooth Muscle Cells Does Not Predispose Mice to Nonatherosclerotic Arteriopathies in 3 Transgenic Mice.

BACKGROUND: Genome-wide association studies have revealed robust associations of common genetic polymorphisms in an intron of the PHACTR-1 (phosphatase and actin regulator 1) gene (chr6p24), with cervical artery dissection, spontaneous coronary artery dissection, and fibromuscular dysplasia. The aim was to assess its role in the pathogenesis of cervical artery dissection or fibromuscular dysplasia. METHODS: Using various tissue-specific Cre-driver mouse lines, Phactr1 was deleted either in endothelial cells using 2 tissue-specific Cre-driver (PDGFB [platelet-derived growth factor B]-CreERT2 mice and Tie2 [tyrosine kinase with immunoglobulin and EGF homology domains]-Cre) and smooth muscle cells (smooth muscle actin-CreERT2) with a third tissue-specific Cre-driver. RESULTS: To test the efficacy of the Phactr1 deletion after cre-induction, we confirmed first, a decrease in Phactr1 transcription and Phactr1 expression in endothelial cell and smooth muscle cell isolated from Phactr1iPDGFB and Phactr1iSMA mice. Irrespective to the tissue or the duration of the deletion, mice did not spontaneously display pathological phenotype or vascular impairment: mouse survival, growth, blood pressure, large vessel morphology, or actin organization were not different in knockout mice than their comparatives littermates. Challenging vascular function and repair either by angiotensin II-induced hypertension or limb ischemia did not lead to vascular morphology or function impairment in Phactr1-deleted mice. Similarly, there were no more consequences of Phactr1 deletion during embryogenesis in endothelial cells. CONCLUSIONS: Loss of PHACTR-1 function in the cells involved in vascular physiology does not appear to induce a pathological vascular phenotype. The in vivo effect of the intronic variation described in genome-wide association studies is unlikely to involve downregulation in PHACTR-1 expression.

Wnt/frizzled Signaling in Endothelium: A Major Player in Blood-Retinal- and Blood-Brain-Barrier Integrity.

The Wnt/frizzled signaling pathway is one of the major regulators of endothelial biology, controlling key cellular activities. Many secreted Wnt ligands have been identified and can initiate diverse signaling via binding to a complex set of Frizzled (Fzd) transmembrane receptors and coreceptors. Roughly, Wnt signaling is subdivided into two pathways: the canonical Wnt/ß-catenin signaling pathway whose main downstream effector is the transcriptional coactivator ß-catenin, and the noncanonical Wnt signaling pathway, which is subdivided into the Wnt/Ca2+ pathway and the planar cell polarity pathway. Here, we will focus on its cross talk with other angiogenic pathways and on its role in blood-retinal- and blood-brain-barrier formation and its maintenance in a differentiated state. We will unravel how retinal vascular pathologies and neurovascular degenerative diseases result from disruption of the Wnt pathway related to vascular instability, and highlight current research into therapeutic options.

Plasma creatinine below limit of quantification in a patient with acute kidney injury.

BACKGROUND: Acute kidney injury (AKI) is an infrequent complication of inflammatory bowel disease and can be exceptionally linked to interstitial nephritis secondary to anti-inflammatory drugs, such as Pentasa® (5-ASA). CASE PRESENTATION: We present an case of an 80-year-old man who presented chronic diarrheas treated by Pentasa®. He developed AKI, evidenced by high plasma creatinine dosed in his local laboratory. At the hospital admission, plasma creatinine was exceptionally undetectable by the enzymatic method while Jaffe's method successfully determined it. Creatinine measurement by the enzymatic method was gradually restored during hospital stay, concomitant with the discontinuation of 5-ASA administration, suggesting that this drug could interfere with creatinine enzymatic assay. Creatinine enzymatic assays combine serial reactions. The last one called Trinder reaction, catalyzed by a peroxidase, uses H2O2 to convert uncolored dye in a colored compound, proportionally to creatinine concentration. We showed that AKI related-plasma accumulation of 5-ASA, could participate in the negative interference observed on creatinine measurement, by scavenging H2O2. Interestingly, all Trinder reaction-based measurements (uric acid, lipase, lactate, triglycerides and cholesterol) were affected. Negative interference of 5-ASA was confirmed by interferogram experiments on all Trinder reaction-dependent assays. CONCLUSION: All Trinder-dependent parameters should be interpreted with the patient's treatment knowledge, in particular salicylate derivatives.

Covichem: A biochemical severity risk score of COVID-19 upon hospital admission.

Clinical and laboratory predictors of COVID-19 severity are now well described and combined to propose mortality or severity scores. However, they all necessitate saturable equipment such as scanners, or procedures difficult to implement such as blood gas measures. To provide an easy and fast COVID-19 severity risk score upon hospital admission, and keeping in mind the above limits, we sought for a scoring system needing limited invasive data such as a simple blood test and co-morbidity assessment by anamnesis. A retrospective study of 303 patients (203 from Bordeaux University hospital and an external independent cohort of 100 patients from Paris Pitié-Salpêtrière hospital) collected clinical and biochemical parameters at admission. Using stepwise model selection by Akaike Information Criterion (AIC), we built the severity score Covichem. Among 26 tested variables, 7: obesity, cardiovascular conditions, plasma sodium, albumin, ferritin, LDH and CK were the independent predictors of severity used in Covichem (accuracy 0.87, AUROC 0.91). Accuracy was 0.92 in the external validation cohort (89% sensitivity and 95% specificity). Covichem score could be useful as a rapid, costless and easy to implement severity assessment tool during acute COVID-19 pandemic waves.

Mast Cells Are the Trigger of Small Vessel Disease and Diastolic Dysfunction in Diabetic Obese Mice.

[Figure: see text].

Characterization of acute kidney injury in critically ill patients with severe coronavirus disease 2019.

BACKGROUND: Coronavirus disease 2019 (COVID-19)-associated acute kidney injury (AKI) frequency, severity and characterization in critically ill patients has not been reported. METHODS: Single-centre cohort performed from 3 March 2020 to 14 April 2020 in four intensive care units in Bordeaux University Hospital, France. All patients with COVID-19 and pulmonary severity criteria were included. AKI was defined using Kidney Disease: Improving Global Outcomes (KDIGO) criteria. A systematic urinary analysis was performed. The incidence, severity, clinical presentation, biological characterization (transient versus persistent AKI; proteinuria, haematuria and glycosuria) and short-term outcomes were evaluated. RESULTS: Seventy-one patients were included, with basal serum creatinine (SCr) of 69 ± 21 µmol/L. At admission, AKI was present in 8/71 (11%) patients. Median [interquartile range (IQR)] follow-up was 17 (12-23) days. AKI developed in a total of 57/71 (80%) patients, with 35% Stage 1, 35% Stage 2 and 30% Stage 3 AKI; 10/57 (18%) required renal replacement therapy (RRT). Transient AKI was present in only 4/55 (7%) patients and persistent AKI was observed in 51/55 (93%). Patients with persistent AKI developed a median (IQR) urine protein/creatinine of 82 (54-140) (mg/mmol) with an albuminuria/proteinuria ratio of 0.23 ± 20, indicating predominant tubulointerstitial injury. Only two (4%) patients had glycosuria. At Day 7 after onset of AKI, six (11%) patients remained dependent on RRT, nine (16%) had SCr >200 µmol/L and four (7%) had died. Day 7 and Day 14 renal recovery occurred in 28% and 52%, respectively. CONCLUSION: Severe COVID-19-associated AKI is frequent, persistent, severe and characterized by an almost exclusive tubulointerstitial injury without glycosuria.

Pigments test strips: A rapid companion test to exclude sub-arachnoid haemorrhage.

OBJECTIVES: Subarachnoid haemorrhage (SAH) is characterised by 25% of mortality or induces long-term care. It needs immediate diagnosis with computed tomography (CT) scan. For the inconclusive CT scans, the detection of haem pigments can be performed in the cerebrospinal fluid (CSF). The reference method is spectrophotometry but it requires a large volume of CSF, and specific equipment. Sometimes, urine test strips are used as an alternative method for haem pigments detection. However, this method needs validation in SAH context. The aim of the study was to compare the performance of Multistix® urine test strips for haem pigments detection to the reference spectrophotometry and the final clinical SAH diagnosis. METHODS: We collected 136 CSFs sampled for suspected SAH. We detected haem pigments with urine test strips and spectrophotometry and compared performances for 100 samples. RESULTS: Urine tests strips displayed a high sensitivity (0.97) as compared to the reference spectrophotometry for haem pigments detection. Interestingly, absence of haem pigments fully correlated with absence of SAH. CONCLUSIONS: Negative Multistix® urine test strips could help to exclude SAH diagnosis in combination with clinical data when a spectrophotometer is not available, or as a bedside diagnosis test.

Therapies targeting Frizzled-7/ß-catenin pathway prevent the development of pathological angiogenesis in an ischemic retinopathy model.

Retinopathies remain major causes of visual impairment in diabetic patients and premature infants. Introduction of anti-angiogenic drugs targeting vascular endothelial growth factor (VEGF) has transformed therapy for these proliferative retinopathies. However, limitations associated with anti-VEGF medications require to unravel new pathways of vessel growth to identify potential drug targets. Here, we investigated the role of Wnt/Frizzled-7 (Fzd7) pathway in a mouse model of oxygen-induced retinopathy (OIR). Using transgenic mice, which enabled endothelium-specific and time-specific Fzd7 deletion, we demonstrated that Fzd7 controls both vaso-obliteration and neovascular phases (NV). Deletion of Fzd7 at P12, after the ischemic phase of OIR, prevented formation of aberrant neovessels into the vitreous by suppressing proliferation of endothelial cells (EC) in tufts. Next we validated in vitro two Frd7 blocking strategies: a monoclonal antibody (mAbFzd7) against Fzd7 and a soluble Fzd7 receptor (CRD). In vivo a single intravitreal microinjection of mAbFzd7 or CRD significantly attenuated retinal neovascularization (NV) in mice with OIR. Molecular analysis revealed that Fzd7 may act through the activation of Wnt/ß-catenin and Jagged1 expression to control EC proliferation in extra-retinal neovessels. We identified Fzd7/ß-catenin signaling as new regulator of pathological retinal NV. Fzd7 appears to be a potent pharmacological target to prevent or treat aberrant angiogenesis of ischemic retinopathies.

Comparison of endothelial promoter efficiency and specificity in mice reveals a subset of Pdgfb-positive hematopoietic cells.

Essentials To reliably study the respective roles of blood and endothelial cells in hemostasis, mouse models with a strong and specific endothelial expression of the Cre recombinase are needed. Using mT/mG reporter mice and conditional JAK2V617F/WT mice, we compared Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 with well-characterized Tie2-Cre mice. Comparison of recombination efficiency and specificity towards blood lineage reveals major differences between endothelial transgenic mice. Cre-mediated recombination occurs in a small number of adult hematopoietic stem cells in Pdgfb-iCreERT2;JAK2V617F/WT transgenic mice. SUMMARY: Background The vessel wall, and particularly blood endothelial cells (BECs), are intensively studied to better understand hemostasis and target thrombosis. To understand the specific role of BECs, it is important to have mouse models that allow specific and homogeneous expression of genes of interest in all BEC beds without concomitant expression in blood cells. Inducible Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 transgenic mice are widely used for BEC targeting. However, issues remain in terms of recombination efficiency and specificity regarding hematopoietic cells. Objectives To determine which mouse model to choose when strong expression of a transgene is required in adult BECs from various organs, without concomitant expression in hematopoietic cells. Methods Using mT/mG reporter mice to measure recombination efficiency and conditional JAK2V617F/WT mice to assess specificity regarding hematopoietic cells, we compared Pdgfb-iCreERT2 and Cdh5(PAC)-CreERT2 with well-characterized Tie2-Cre mice. Results Adult Cdh5(PAC)-CreERT2 mice are endothelial specific but require a dose of 10 mg of tamoxifen to allow constant Cre expression. Pdgfb-iCreERT2 mice injected with 5 mg of tamoxifen are appropriate for most endothelial research fields except liver studies, as hepatic sinusoid ECs are not recombined. Surprisingly, 2 months after induction of Cre-mediated recombination, all Pdgfb-iCreERT2;JAK2V617F/WT mice developed a myeloproliferative neoplasm that is related to the presence of JAK2V617F in hematopoietic cells, showing for the first time that Cre-mediated recombination occurs in a small number of adult hematopoietic stem cells in Pdgfb-iCreERT2 transgenic mice. Conclusion This study provides useful guidelines for choosing the best mouse line to study the role of BECs in hemostasis and thrombosis.

Targeting the Sphingosine 1-Phosphate Axis Exerts Potent Antitumor Activity in BRAFi-Resistant Melanomas.

BRAF inhibitors (BRAFi) are used to treat patients with melanoma harboring the V600E mutation. However, resistance to BRAFi is inevitable. Here, we identified sphingosine 1-phosphate (S1P) receptors as regulators of BRAFV600E-mutant melanoma cell-autonomous resistance to BRAFi. Moreover, our results reveal a distinct sphingolipid profile, that is, a tendency for increased very long-chain ceramide species, in the plasma of patients with melanoma who achieve a response to BRAFi therapy as compared with patients with progressive disease. Treatment with BRAFi resulted in a strong decrease in S1PR1/3 expression in sensitive but not in resistant cells. Genetic and pharmacologic interventions, that increase ceramide/S1P ratio, downregulated S1PR expression and blocked BRAFi-resistant melanoma cell growth. This effect was associated with a decreased expression of MITF and Bcl-2. Moreover, the BH3 mimetic ABT-737 improved the antitumor activity of approaches targeting S1P-metabolizing enzymes in BRAFi-resistant melanoma cells. Collectively, our findings indicate that targeting the S1P/S1PR axis could provide effective therapeutic options for patients with melanoma who relapse after BRAFi therapy.

A combined approach for the early recognition of acute kidney injury after adult cardiac surgery.

BACKGROUND: Cardiac surgery-associated acute kidney injury (CSA-AKI) is a frequent complication. The current criteria to detect CSA-AKI rise only when organic dysfunction has occurred. The Doppler Renal Resistive Index (RRI) and the urinary G1 cell cycle arrest proteins (TIMP-2 and IGFBP7) have been advocated to predict CSA-AKI at an early stage after cardiac surgery. The primary objective was to determine the predictive value of these new markers to detect CSA-AKI after elective heart surgery in patients at risk to develop AKI. METHODS: In a prospective observational trial, we studied 50 patients scheduled for elective on-pump heart surgery at high risk for CSA-AKI. The primary outcome was the incidence of AKI according to the KDIGO criteria recording the urine output every hour until ICU discharge and measuring the serum creatinine levels on each postoperative day until the post-procedure peak values were reached or until the 7th postoperative day. The RRI and the urinary proteins [TIMP-2]*[IGFBP7] were measured concomitantly: before surgery, 1hour (H1), 4-hour (H4), 12-hour (H12), and 24-hour (H24) after surgery. RESULTS: Thirty-seven patients (74%) developed CSA-AKI. Urinary [TIMP-2]*[IGFBP7] at H12 were significantly higher in patients that developed AKI (0.62, [interquartile] [0.20-1.18] vs. 0.30 [0.07-0.47] P=0.044) with an area under the receiver-operating characteristic curve of 0.69 [0.53-0.84]. The best sensitivity (65%) and specificity (62%) was achieved for a cutoff value of 0.3 (ng.mL-1)2.1000-1. The H12 time-point was the only in which the RRI values measured showed a trend toward statistical significance in patients that developed AKI (0.72 (Standard deviation)±(0.06) vs. 0.68±(0.07) P=0.065). The combination of the two markers ([TIMP-2]*[IGFBP7]+RRI) at H12 showed an increased performance of the accuracy with an area under the receiver-operating characteristic curve of 0.78 [0.62-0.93]. CONCLUSIONS: In a population at risk of developing CSA-AKI, neither the RRI nor urinary [TIMP-2]*[IGFBP7] detect CSA-AKI occurring in the first post-operative week within the first 24 postoperative hours.

New markers for early detection of acute kidney injury after transcatheter aortic valve implantation.

BACKGROUND: Acute kidney injury (AKI) is a frequent complication after a transcatheter aortic valve implantation (TAVI). Biomarkers such as urinary G1 cell cycle arrest proteins (TIMP-2 and IGFBP7) and sonographic evaluation (Doppler Renal Resistive Index [RRI]) have been advocated to predict AKI at an early stage after a TAVI-procedure. The primary aim was to determine the predictive value of these markers to detect AKI after a TAVI-procedure at an early phase. PATIENTS AND METHODS: In a prospective observational study, 62 consecutive patients were scheduled for a TAVI. AKI was assessed based on the KDIGO criteria. Biomarkers and RRI were measured concomitantly before TAVI, at the first micturition post-implantation and the first micturition on the morning after the procedure. RESULTS: Twenty-two patients (35%) developed AKI. On the first day after the TAVI-procedure, urinary TIMP-2 and IGFBP7 concentrations increased significantly in patients who developed AKI (0.1, [interquartile] [0.1-0.35] to 0.40 [0.10-1.00] vs. 0.2 [0.1-0.5] to 0.10 [0.10-0.20], P=0.012) with an area under the receiver-operating characteristic curve of 0.71 [0.55-0.83]. Sensitivity was 0.57 and specificity was 0.83 for a cut-off value of 0.35. No significant increases in RRI were found in patients who developed AKI. CONCLUSIONS: Based on the current guidelines for the diagnosis of AKI, the urinary proteins TIMP-2 and IGFBP7 do not detect AKI at an early stage accurately in patients undergoing a TAVI-procedure.

Metastasis suppressor NM23 limits oxidative stress in mammals by preventing activation of stress-activated protein kinases/JNKs through its nucleoside diphosphate kinase activity.

NME1 (nonmetastatic expressed 1) gene, which encodes nucleoside diphosphate kinase (NDPK) A [also known as nonmetastatic clone 23 (NM23)-H1 in humans and NM23-M1 in mice], is a suppressor of metastasis, but several lines of evidence-mostly from plants-also implicate it in the regulation of the oxidative stress response. Here, our aim was to investigate the physiologic relevance of NDPK A with respect to the oxidative stress response in mammals and to study its molecular basis. NME1-knockout mice died sooner, suffered greater hepatocyte injury, and had lower superoxide dismutase activity than did wild-type (WT) mice in response to paraquat-induced acute oxidative stress. Deletion of NME1 reduced total NDPK activity and exacerbated activation of the stress-related MAPK, JNK, in the liver in response to paraquat. In a mouse transformed hepatocyte cell line and in primary cultures of normal human keratinocytes, MAPK activation in response to H2O2 and UVB, respectively, was dampened by expression of NM23-M1/NM23-H1, dependent on its NDPK catalytic activity. Furthermore, excess or depletion of NM23-M1/NM23-H1 NDPK activity did not affect the intracellular bulk concentration of nucleoside di- and triphosphates. NME1-deficient mouse embryo fibroblasts grew poorly in culture, were more sensitive to stress than WT fibroblasts, and did not immortalize, which suggested that they senesce earlier than do WT fibroblasts. Collectively, these results indicate that the NDPK activity of NM23-M1/NM23-H1 protects cells from acute oxidative stress by inhibiting activation of JNK in mammal models.-Peuchant, E., Bats, M.-L., Moranvillier, I., Lepoivre, M., Guitton, J., Wendum, D., Lacombe, M.-L., Moreau-Gaudry, F., Boissan, M., Dabernat, S. Metastasis suppressor NM23 limits oxidative stress in mammals by preventing activation of stress-activated protein kinases/JNKs through its nucleoside diphosphate kinase activity.

Fzd7 (Frizzled-7) Expressed by Endothelial Cells Controls Blood Vessel Formation Through Wnt/ß-Catenin Canonical Signaling.

OBJECTIVE: Vessel formation requires precise orchestration of a series of morphometric and molecular events controlled by a multitude of angiogenic factors and morphogens. Wnt/frizzled signaling is required for proper vascular formation. In this study, we investigated the role of the Fzd7 (frizzled-7) receptor in retinal vascular development and its relationship with the Wnt/ß-catenin canonical pathway and Notch signaling. APPROACH AND RESULTS: Using transgenic mice, we demonstrated that Fzd7 is required for postnatal vascular formation. Endothelial cell (EC) deletion of fzd7 (fzd7ECKO) delayed retinal plexus formation because of an impairment in tip cell phenotype and a decrease in stalk cell proliferation. Dvl (dishevelled) proteins are a main component of Wnt signaling and play a functionally redundant role. We found that Dvl3 depletion in dvl1-/- mice mimicked the fzd7ECKO vascular phenotype and demonstrated that Fzd7 acted via ß-catenin activation by showing that LiCl treatment rescued impairment in tip and stalk cell phenotypes induced in fzd7 mutants. Deletion of fzd7 or Dvl1/3 induced a strong decrease in Wnt canonical genes and Notch partners' expression. Genetic and pharmacological rescue strategies demonstrated that Fzd7 acted via ß-catenin activation, upstream of Notch signaling to control Dll4 and Jagged1 EC expression. CONCLUSIONS: Fzd7 expressed by EC drives postnatal angiogenesis via activation of Dvl/ß-catenin signaling and can control the integrative interaction of Wnt and Notch signaling during postnatal angiogenesis.

Kif26b controls endothelial cell polarity through the Dishevelled/Daam1-dependent planar cell polarity-signaling pathway.

Angiogenesis involves the coordinated growth and migration of endothelial cells (ECs) toward a proangiogenic signal. The Wnt planar cell polarity (PCP) pathway, through the recruitment of Dishevelled (Dvl) and Dvl-associated activator of morphogenesis (Daam1), has been proposed to regulate cell actin cytoskeleton and microtubule (MT) reorganization for oriented cell migration. Here we report that Kif26b--a kinesin--and Daam1 cooperatively regulate initiation of EC sprouting and directional migration via MT reorganization. First, we find that Kif26b is recruited within the Dvl3/Daam1 complex. Using a three-dimensional in vitro angiogenesis assay, we show that Kif26b and Daam1 depletion impairs tip cell polarization and destabilizes extended vascular processes. Kif26b depletion specifically alters EC directional migration and mislocalized MT organizing center (MTOC)/Golgi and myosin IIB cell rear enrichment. Therefore the cell fails to establish a proper front-rear polarity. Of interest, Kif26b ectopic expression rescues the siDaam1 polarization defect phenotype. Finally, we show that Kif26b functions in MT stabilization, which is indispensable for asymmetrical cell structure reorganization. These data demonstrate that Kif26b, together with Dvl3/Daam1, initiates cell polarity through the control of PCP signaling pathway-dependent activation.

Kinetic eGFR and Novel AKI Biomarkers to Predict Renal Recovery.

BACKGROUND AND OBJECTIVES: Prompt recognition of severe renal impairment could improve the early management of critically ill patients. We compared the value of kinetic eGFR, plasma neutrophil gelatinase-associated lipocalin (NGAL), and urine tissue inhibitor of metalloproteinase-2 and urine insulin-like growth factor-binding protein 7 ([TIMP-2]*[IGFBP7]) in predicting short-term recovery from AKI and major adverse kidney events. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: During the 6-month study period, 245 patients were admitted to our intensive care unit. This study included 57 consecutive patients presenting with AKI within the first 24 hours after admission. AKI markers were evaluated at inclusion (day 0) and 24 hours later (day 1). Kinetic eGFR was calculated on day 1 according to serum creatinine evolution. Renal recovery was defined as normalization of serum creatinine with reversal of oliguria within 48 hours. Major adverse kidney events included death, need for RRT, or persistence of renal dysfunction at hospital discharge. RESULTS: Plasma NGAL and [TIMP-2]*[IGFBP7] predicted renal recovery, with area under the receiver-operating characteristic curve (AUC-ROC) values between 0.70 and 0.79 at inclusion. Although plasma NGAL values frequently reached the maximal measurement range, their decrease on day 1 predicted recovery. The kinetic eGFR calculation after initial resuscitation provided the best AUC-ROC value for renal recovery, at 0.87. The best predictions for major adverse kidney events were provided by [TIMP-2]*[IGFBP7] and kinetic eGFR (equal AUC-ROCs of 0.81). Combining AKI markers in addition to clinical prediction models improved the discrimination and reclassification of patients who will recover from AKI or suffer from major adverse kidney events. CONCLUSIONS: Biomarkers of kidney damage predicted short-term renal recovery and major adverse kidney events for an unselected cohort of critically ill patients. Calculating the kinetic eGFR imposed a delay after initial resuscitation but provided a good diagnostic and prognostic approach. The utility of functional and damage AKI marker combinations in addition to clinical information requires validation in larger prospective studies.

Genetic disorders of simple sphingolipid metabolism.

A better understanding of the functions sphingolipids play in living organisms can be achieved by analyzing the biochemical and physiological changes that result from genetic alterations of sphingolipid metabolism. This review summarizes the current knowledge gained from studies both on human patients and mutant animals (mice, cats, dogs, and cattle) with genetic disorders of sphingolipid metabolism. Genetic alterations affecting the biosynthesis, transport, or degradation of simple sphingolipids are discussed.