Caveolar peroxynitrite formation impairs endothelial TRPV4 channels and elevates pulmonary arterial pressure in pulmonary hypertension.

Recent studies have focused on the contribution of capillary endothelial TRPV4 channels to pulmonary pathologies, including lung edema and lung injury. However, in pulmonary hypertension (PH), small pulmonary arteries are the focus of the pathology, and endothelial TRPV4 channels in this crucial anatomy remain unexplored in PH. Here, we provide evidence that TRPV4 channels in endothelial cell caveolae maintain a low pulmonary arterial pressure under normal conditions. Moreover, the activity of caveolar TRPV4 channels is impaired in pulmonary arteries from mouse models of PH and PH patients. In PH, up-regulation of iNOS and NOX1 enzymes at endothelial cell caveolae results in the formation of the oxidant molecule peroxynitrite. Peroxynitrite, in turn, targets the structural protein caveolin-1 to reduce the activity of TRPV4 channels. These results suggest that endothelial caveolin-1-TRPV4 channel signaling lowers pulmonary arterial pressure, and impairment of endothelial caveolin-1-TRPV4 channel signaling contributes to elevated pulmonary arterial pressure in PH. Thus, inhibiting NOX1 or iNOS activity, or lowering endothelial peroxynitrite levels, may represent strategies for restoring vasodilation and pulmonary arterial pressure in PH.

Novel Smooth Muscle Ca2+-Signaling Nanodomains in Blood Pressure Regulation.

BACKGROUND: Ca2+ signals in smooth muscle cells (SMCs) contribute to vascular resistance and control blood pressure. Increased vascular resistance in hypertension has been attributed to impaired SMC Ca2+ signaling mechanisms. In this regard, transient receptor potential vanilloid 4 (TRPV4SMC) ion channels are a crucial Ca2+ entry pathway in SMCs. However, their role in blood pressure regulation has not been identified. METHODS: We used SMC-specific TRPV4-/- (TRPV4SMC-/-) mice to assess the role of TRPV4SMC channels in blood pressure regulation. We determined the contribution of TRPV4SMC channels to the constrictor effect of α1 adrenergic receptor (α1AR) stimulation and elevated intraluminal pressure: 2 main physiologic stimuli that constrict resistance-sized arteries. The contribution of spatially separated TRPV4SMC channel subpopulations to elevated blood pressure in hypertension was evaluated in angiotensin II-infused mice and patients with hypertension. RESULTS: We provide first evidence that TRPV4SMC channel activity elevates resting blood pressure in normal mice. α1AR stimulation activated TRPV4SMC channels through PKCα (protein kinase Cα) signaling, which contributed significantly to vasoconstriction and blood pressure elevation. Intraluminal pressure-induced TRPV4SMC channel activity opposed vasoconstriction through activation of Ca2+-sensitive K+ (BK) channels, indicating functionally opposite pools of TRPV4SMC channels. Superresolution imaging of SMCs revealed spatially separated α1AR:TRPV4 and TRPV4:BK nanodomains in SMCs. These data suggest that spatially separated α1AR-TRPV4SMC and intraluminal pressure-TRPV4SMC-BK channel signaling have opposite effects on blood pressure, with α1AR-TRPV4SMC signaling dominating under resting conditions. Furthermore, in patients with hypertension and a mouse model of hypertension, constrictor α1AR-PKCα-TRPV4 signaling was upregulated, whereas dilator pressure-TRPV4-BK channel signaling was disrupted, thereby increasing vasoconstriction and elevating blood pressure. CONCLUSIONS: Our data identify novel smooth muscle Ca2+-signaling nanodomains that regulate blood pressure and demonstrate their impairment in hypertension.

Calcium Signal Profiles in Vascular Endothelium from Cdh5-GCaMP8 and Cx40-GCaMP2 Mice.

INTRODUCTION: Studies in Cx40-GCaMP2 mice, which express calcium biosensor GCaMP2 in the endothelium under connexin 40 promoter, have identified the unique properties of endothelial calcium signals. However, Cx40-GCaMP2 mouse is associated with a narrow dynamic range and lack of signal in the venous endothelium. Recent studies have proposed many GCaMPs (GCaMP5/6/7/8) with improved properties although their performance in endothelium-specific calcium studies is not known. METHODS: We characterized a newly developed mouse line that constitutively expresses GCaMP8 in the endothelium under the VE-cadherin (Cdh5-GCaMP8) promoter. Calcium signals through endothelial IP3 receptors and TRP vanilloid 4 (TRPV4) ion channels were recorded in mesenteric arteries (MAs) and veins from Cdh5-GCaMP8 and Cx40-GCaMP2 mice. RESULTS: Cdh5-GCaMP8 mice showed lower baseline fluorescence intensity, higher dynamic range, and higher amplitudes of individual calcium signals than Cx40-GCaMP2 mice. Importantly, Cdh5-GCaMP8 mice enabled the first recordings of discrete calcium signals in the intact venous endothelium and revealed striking differences in IP3 receptor and TRPV4 channel calcium signals between MAs and mesenteric veins. CONCLUSION: Our findings suggest that Cdh5-GCaMP8 mice represent significant improvements in dynamic range, sensitivity for low-intensity signals, and the ability to record calcium signals in venous endothelium.

Colonization With Levofloxacin-resistant Extended-spectrum ß-Lactamase-producing Enterobacteriaceae and Risk of Bacteremia in Hematopoietic Stem Cell Transplant Recipients.

Background: Bacteremia caused by extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae (ESBL-E) is associated with inadequate empirical therapy and substantial mortality in neutropenic patients. Strategies are needed to identify neutropenic patients at high risk of these infections. Methods: From April 2014 to September 2016, we collected perianal swabs, both at admission and weekly thereafter, from patients undergoing hematopoietic stem cell transplantation (HSCT). Patients received prophylactic levofloxacin while neutropenic. Swabs were plated onto selective agar, colonies were identified and underwent antimicrobial susceptibility testing, and phenotypic ESBL testing and polymerase chain reaction for ß-lactamase genes were performed on ceftriaxone-resistant Enterobacteriaceae. We then determined the prevalence of pre-transplant ESBL-E colonization and risk of ESBL-E bacteremia. Colonizing and bloodstream isolates from patients with ESBL-E bacteremia underwent multilocus sequence typing and pulsed-field gel electrophoresis. Results: We analyzed 312 patients, including 212 allogeneic and 100 autologous HSCT recipients. Ten percent (31/312) of patients had pre-transplant ESBL-E colonization. Susceptibility rates of colonizing ESBL-E were: levofloxacin, 25%; cefepime, 9%; piperacillin-tazobactam, 84%; and meropenem, 97%. Of 31 patients colonized with ESBL-E pre-transplant, 10 (32%) developed ESBL-E bacteremia during their transplant admission, compared to 1 (0.4%) of 281 patients not colonized with ESBL-E (P < .001). All bloodstream ESBL-E were levofloxacin-resistant and colonizing and bloodstream isolates from individual patients had identical genotypic profiles. Conclusions: HSCT recipients who are colonized with levofloxacin-resistant ESBL-E pre-transplant and receive levofloxacin prophylaxis have high rates of bacteremia from their colonizing strain during neutropenia. Assessing for ESBL-E colonization in neutropenic patients could lead to optimization of empirical antibacterial therapy.

Synthesis of the WXYZA' domain of maitotoxin.

A synthesis of the WXYZA' domain (7) of the marine neurotoxin maitotoxin (1) is reported. The convergent synthetic strategy involves construction of key building blocks 11 and 12, their coupling, and the elaboration of the resulting ester (10) to the target molecule through a ring-closing metathesis and a hydroxy dithioketal cyclization as the key steps. For the construction of fragment 11, the Noyori reduction/Achmatowicz rearrangement and hydroxy epoxide opening technologies were applied (starting from furfuryl alcohol (13)), whereas for the synthesis of fragment 12, a carbohydrate-based approach was adopted (starting from 2-deoxy-D-ribose (14)). The synthesized WXYZA' domain (7) of maitotoxin (1) exhibited the expected (13)C NMR chemical shifts, supporting the originally assigned structure of the corresponding region of the natural product.

Epidemiology of Bloodstream Infections Caused by Escherichia coli and Klebsiella pneumoniae That Are Piperacillin-Tazobactam-Nonsusceptible but Ceftriaxone-Susceptible.

BACKGROUND: Piperacillin-tazobactam-nonsusceptible (TZP-NS) Enterobacteriaceae are typically also resistant to ceftriaxone. We recently encountered bacteremias due to Escherichia coli (Ec) and Klebsiella pneumoniae (Kp) that were TZP-NS but ceftriaxone-susceptible (CRO-S). METHODS: We reviewed all Ec and Kp bacteremias from 2011 to 2015 at our center and assessed the prevalence, antimicrobial susceptibilities, genetic profiles, patient characteristics, treatments, and outcomes of TZP-NS/CRO-S infections. We identified risk factors for TZP-NS/CRO-S infections compared with Ec and Kp bacteremias that were TZP-S and CRO-S (Control Group 1) and compared outcomes of patients with TZP-NS/CRO-S bacteremias, Control Group 1, and patients bacteremic with extended-spectrum ß-lactamase (ESBL)-producing Ec and Kp. RESULTS: There were 1857 Ec and Kp bacteremia episodes, of which 78 (4.2%) were TZP-NS/CRO-S (Ec: 50/1227 [4.1%]; Kp: 28/630 [4.4%]). All TZP-NS/CRO-S isolates were also ampicillin-sulbactam-NS. Of 32 TZP-NS/CRO-S isolates that were sequenced, 28 (88%) harbored bla TEM-1 or bla SHV-1, none had an ESBL or AmpC ß-lactamase gene, and many sequence types were represented. Independent risk factors for TZP-NS/CRO-S bacteremia were exposure to ß-lactam/ß-lactamase inhibitors (BL/BLIs; adjusted odds ratio [aOR], 5.5; P < .001) and cephalosporins (aOR, 3.0; P = .04). Thirty-day mortality after TZP-NS/CRO-S bacteremia was 25%, which was similar to control groups and was similar in patients treated empirically with BL/BLIs compared with those treated with cephalosporins or carbapenems. Targeted therapy with cephalosporins did not yield a higher 30-day mortality rate than carbapenem therapy. CONCLUSIONS: TZP-NS/CRO-S Ec and Kp are emerging causes of bacteremia, and further research is needed to better understand the epidemiology, resistance mechanisms, and clinical impact of these strains.

Solid-phase approach to tetrahydroquinolones using a sulfur linker cleaved by SmI2.

[reaction: see text] A sulfur alpha-heteroatom-substituted carbonyl (HASC) linker has been utilized in a solid-phase approach to tetrahydroquinolones. The route illustrates the compatibility of the linker system with palladium-catalyzed transformations and its utility for library synthesis. The linker is cleaved by electron transfer from samarium(II) iodide.

The growing threat of multidrug-resistant Gram-negative infections in patients with hematologic malignancies.

Prolonged neutropenia and chemotherapy-induced mucositis render patients with hematologic malignancies highly vulnerable to Gram-negative bacteremia. Unfortunately, multidrug-resistant (MDR) Gram-negative bacteria are increasingly encountered globally, and current guidelines for empirical antibiotic coverage in these patients may not adequately treat these bacteria. This expansion of resistance, coupled with traditional culturing techniques requiring 2-4 days for bacterial identification and antimicrobial susceptibility results, have grave implications for these immunocompromised hosts. This review characterizes the epidemiology, risk factors, resistance mechanisms, recommended treatments, and outcomes of the MDR Gram-negative bacteria that commonly cause infections in patients with hematologic malignancies. We also examine the infection prevention strategies in hematology patients, such as infection control practices, antimicrobial stewardship, and targeted decolonization. Finally, we assess the strategies to improve outcomes of the infected patients, including gastrointestinal screening to guide empirical antibiotic therapy, new rapid diagnostic tools for expeditious identification of MDR pathogens, and use of two new antimicrobial agents, ceftolozane/tazobactam and ceftazidime/avibactam.

A review of antithrombotic therapy for transcatheter aortic valve replacement.

Aortic stenosis (AS) is a common valvular pathological finding in older adults. A latent period followed by rapid progression after the onset of symptoms can result in a high rate of death if left untreated. Aortic valve replacement (AVR) remains the standard of care for patients with severe symptomatic AS; however, due to comorbidities and age, patients may be ineligible for surgical AVR. Transcatheter AVR (TAVR) is an alternative treatment for patients with severe symptomatic AS for whom surgery is not an option. The most recent guidelines and consensus statement discussing TAVR support the use of antithrombotic therapy during and after TAVR procedures to prevent thrombotic complications. Intravenous unfractionated heparin titrated to target activated clotting times during the procedure and dual antiplatelet therapy before and after TAVR with aspirin and clopidogrel may be used. However, it is unclear which antithrombotic regimen may provide optimal protection for early and late thrombotic events in patients who undergo TAVR. This review evaluates the current guidelines, trials, and registry data discussing antithrombotic regimens for TAVR.