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Introduction The kidney is one of the main organs affected by microvascular damage wrought by hypertension. We developed an approach to investigate renal microcirculatory disturbance in live mice by measuring post-occlusive reactive hyperemia (PORH), a reactivity test exploring endothelial and neuro-microvascular functioning. Laser speckle-contrast analysis (LASCA) assesses microvascular blood flow; it provides real-time images of spatial and temporal blood flow dynamics. We compared basal blood flow and PORH test between control and angiotensin-II-treated mice (Ang-II) to validate the model. Objective The study objective was to develop an approach to investigate renal microcirculation, and then to compare microvascular reactivity assessed on LASCA in control versus Ang-II mice. Methods Thirty 7-week-old wild-type C57BL/6J mice were allocated into two groups. One received angiotensin-II via osmotic minipumps (Ang-II; n=15); the other served as control (n=15). Basal blood flow was measured on LASCA. The PORH test was then performed in the two groups. Results Control mice had significantly lower basal renal microcirculatory flow, expressed in perfusion units (PU), than Ang-II-treated mice (1448 ± 96 vs 1703 ± 185 PU, respectively; P < 0.05). Peak flow was lower in controls than in Ang-II mice (1617±104 vs.1724 ± 205 PU, respectively; P=0.21). Control mice had significantly higher kidney PORH than Ang-II mice (8±3 vs 1±4%, respectively; P < 0.05). Conclusion We developed an innovative technique to study renal microcirculation in mice. Ang-II-treated mice showed significantly higher basal blood flow than controls, while PORH was significantly higher in controls than in Ang-II mice.

Immune-mediated, drug-induced hepatitis is a rare complication of halogenated volatile anesthetic administration. IL-4-regulated Th2-polarized reactions initiate this type and other types of hepatitis, while the mechanisms that regulate the severity remain elusive. IL-33 is an innate, IL-4-inducing, Th2-polarizing cytokine that has been detected in patients with liver failure and has been associated with upregulated ST2+Foxp3+CD4+CD25+ T cells; however, roles for IL-33 in drug-induced hepatitis are unclear. We investigated IL-33 in an anesthetic, immune-mediated hepatitis modeled in BALB/c, IL-33-/- and ST2-/- mice, as well as in patients with anesthetic hepatitis. The hepatic IL-33 and ST2 levels were elevated in BALB/c mice (p 0.05) with hepatitis, and anti-IL-33 diminished hepatitis (p 0.05) without reducing IL-33 levels. The complete absence of IL-33 reduced IL-10 (p 0.05) and ST2+Foxp3+CD4+CD25+ T cells (p 0.05), as well as reduced the overall survival (p 0.05), suggesting suppressive roles for IL-33 in anesthetic, immune-mediated hepatitis. All of the mice demonstrated similar levels of CD4+ T-cell proliferation following direct T-cell receptor stimulation, but we detected splenic IL-33 and ST2-negative Foxp3+CD4+CD25+ T cells in ST2-/- mice that developed less hepatitis than BALB/c mice (p 0.05), suggesting that ST2-negative Foxp3+CD4+CD25+ T cells reduced hepatitis. In patients, serum IL-33 and IPEX levels were correlated in controls (r

Introduction Emerging research and clinical evidence suggest that severe COVID‐19 is a microvascular disease, where SARS‐CoV‐2 infection triggers a rapid inflammatory vascular response that leads to endothelial dysfunction and thrombosis. Underlying medical conditions such as obesity, type 2 diabetes and hypertension, have been associated with increased risk for severe illness. SARS‐CoV‐2 enters cells by surface spike protein binding to host cell receptor angiotensin‐converting enzyme 2 (ACE2). Isolated S protein of SARS‐CoV‐1 has been shown to induce an innate immune response. Here we investigate the cytokine response of monocytes with and without comorbidities (obesity and hypertension) exposed to SARS‐CoV‐2 spike protein, and the effects of this response on endothelial barrier function. Methods Human CD14+ monocytes (Lonza, Precision for Medicine) from two Caucasian males in their 50s, one with obesity and hypertension (HTN) and one without, were stimulated with LPS (100 ng/mL [positive control]), recombinant spike receptor‐binding domain (RBD, 10 ug/mL) or serum‐free media (negative control) for 48 hours. Monocyte conditioned media (MCM) was collected and cytokines (IL‐6, IL‐8, IL‐10, IL‐12p70, IL‐1beta, IFN‐gamma, TNF‐alpha, IL‐4) were analyzed by multiplex ELISA. Human lung microvascular endothelial cells (HLMVECs) from a healthy donor were exposed to MCM from healthy or HTN subjects. Endothelial barrier function was measured by electric cell‐substrate impedance sensing (ECIS). Briefly, HLMVECs were seeded on ECIS chambers to proliferate until impedance plateaued. Half of the media was then replaced with MCM. Differences in impedance drops were compared among various conditions. Western blotting was used to investigate Spike RBD internalization. Results Spike RBD and resulted in increased IL‐6, IL‐8, IL‐12p70, and TNF‐alpha compared to negative control (Fig1A). Monocytes from HTN subject showed increased secretion of IL‐6, IL‐8 and IL‐12p70 compared to those from the healthy subject (Fig 1B). HLMVECs showed moderate long‐term barrier disruption, when treated with Spike RBD alone and Spike RBD with MCM from healthy subjects and no difference in transient barrier disruption or recovery. HLMVECs exposed to MCM from HTN subjects showed increased long‐term barrier dysfunction, especially with Spike RBD present, and recovery of barrier function was also hampered (Fig 2). Spike RBD was observed to be internalized by both monocytes and HLMVECs. Discussion Spike RBD alone induces inflammatory cytokine secretion by CD14+ monocytes and is further increased in monocytes from the obese/HTN subject. Exposure to Spike RBD and MCM from both healthy and HTN subjects caused significant long‐term endothelial barrier dysfunction in HLMVECs with increased dysfunction seen for the obese/HTN MCM. These results are consistent with clinical experience with the development of acute respiratory distress syndrome, increased disease severity, and worse outcome in COVID‐19 patients with comorbidities. Further work is being done to characterize the cytokine profile and mechanisms of barrier dysfunction.

Summary Heterozygous gain-of-function (GOF) mutations of hypoxia-inducible factor 2α (HIF2A), a key hypoxia-sensing regulator, are associated with erythrocytosis, thrombosis, and vascular complications that account for morbidity and mortality of patients. We demonstrated that the vascular pathology of HIF2A GOF mutations is independent of erythrocytosis. We generated HIF2A GOF-induced pluripotent stem cells (iPSCs) and differentiated them into endothelial cells (ECs) and smooth muscle cells (SMCs). Unexpectedly, HIF2A-SMCs, but not HIF2A-ECs, were phenotypically aberrant, more contractile, stiffer, and overexpressed endothelin 1 (EDN1), myosin heavy chain, elastin, and fibrillin. EDN1 inhibition and knockdown of EDN1-receptors both reduced HIF2-SMC stiffness. Hif2A GOF heterozygous mice displayed pulmonary hypertension, had SMCs with more disorganized stress fibers and higher stiffness in their pulmonary arterial smooth muscle cells, and had more deformable pulmonary arteries compared with wild-type mice. Our findings suggest that targeting these vascular aberrations could benefit patients with HIF2A GOF and conditions of augmented hypoxia signaling., Graphical abstract, Highlights • HIF2-SMCs are stiffer than WT-SMCs and differ in contractile SMC marker expression • HIF2-SMCs and WT-SMCs differ in EDN1 production and ECM composition • HIF- 2α induces EDN1; EDNI subsequently induces SMC stiffening • Hif2A GOF mouse arterial SMCs have more disorganized stress fibers and are stiffer, Pathophysiology; Biophysics; Biomechanics

BackgroundHypertension is a well-established driver of vascular remodeling and stiffening. The goal of this study was to evaluate whether restoring normal blood pressure (BP) fully restores vascular stiffness toward that of normotensive controls.MethodsC57Bl6/J male mice received angiotensin II (angII; 1 μg/kg/min) via infusion pump for 8 weeks (hypertension group: HH), angII for 4 weeks (hypertension group: H4), angII for 4 weeks followed by 4 weeks of recovery (reversal group: HN), or sham treatment (normotensive group: NN). BP, heart rate, and pulse wave velocity (PWV) were measured longitudinally. At the end of the study period, aortas were harvested for testing of vasoreactivity, passive mechanical properties, and vessel structure.ResultsThe HH group exhibited a sustained increase in BP and PWV over the 8-week period (p < 0.01). In the HN group, BP and PWV increased during the 4-week angII infusion, and, though BP was restored during the 4-week recovery, PWV exhibited only partial restoration (p < 0.05). Heart rate was similar in all cohorts. Compared to NN controls, both HH and HN groups had significantly increased wall thickness (p < 0.05 HH vs. NN, p < 0.01 HN vs. NN), mucosal extracellular matrix accumulation (p < 0.0001 HH vs. NN, p < 0.05 HN vs. NN), and intralamellar distance (p < 0.001 HH vs. NN, p < 0.01 HN vs. NN). Both intact and decellularized vessels were noted to have significantly higher passive stiffness in the HH and H4 cohorts than in NN controls (p < 0.0001). However, in the HN cohort, intact vessels were only modestly stiffer than those of NN controls, and decellularized HN vessels were identical to those from the NN controls. Compared to NN controls, the HH and HN cohorts exhibited significantly diminished phenylephrine-induced contraction (p < 0.0001) and endothelium-dependent vasodilation (p < 0.05).ConclusionHypertension causes a significant increase in in vivo aortic stiffness that is only partially reversible after BP normalization. Although hypertension does lead to matrix stiffening, restoration of BP restores matrix mechanics to levels similar to those of normotensive controls. Nevertheless, endothelial and vascular smooth muscle cell dysfunction persist after restoration of normotension. This dysfunction is, in part, responsible for augmented PWV after restoration of BP.

Solar energy is one of the most promising renewable energy that has accelerated and globally about 630 GW has been installed so far. And it projected to reach 8500 GW by 2050 as per IRENA. Solar photovoltaic panels have typically 15-20% efficiency in capturing the incident sunlight. And as the market is maturing, new avenues of research are focused on improving solar PV efficiency at cell level, module level and by adopting solar tracking. Solar photovoltaic panels (PV) are typically mounted at an optimum tilt angle according to the latitude and generally known as Fixed tit deployment. Solar tracking is an alternative methodology to maximize energy captured by a PV panel and is achieved by tracking the sun. This can give an efficiency gain of up to 25% w.r.t fixed tilt but at an additional capital cost of 20%. At Renkube we have developed an innovative PV panel design called MFOT (Motion Free Optical Tracking) that can increase the energy generation of module by the same quantum of 20% as physical trackers but at an additional capital cost of 5-10%. The MFOT panel uses prismatic structures that work on the principal of Total Internal Reflection to increase sunlight incident on solar cell. BDRF (Bi-Directional Reflectance Function) based mirrors is another alternative technology built at Michigan Technology University to boost the illumination on solar cells using mirrors and improve energy generation by up to 20% more. In this paper, we compare the pros and cons of BDRF and MFOT technologies and take a deep dive on various factors such as annual performance gain, geographic constraints, and highlight the seasonal improvements of these systems.

Heterozygous gain-of-function (GOF) mutations of hypoxia-inducible factor 2α (HIF2A), a significant regulator of hypoxia sensing, are associated with erythrocytosis, thrombosis and vascular complications that account for morbidity and mortality of patients with these disorders. We demonstratethat vascular pathology of HIF2A GOF mutations is independent of erythrocytosis. We generated HIF2A GOF induced pluripotent stem cells (iPSCs) and differentiated them into endothelial cells (ECs) and smooth muscle cells (SMCs). Unexpectedly, the HIF2A-SMCs, but not HIF2A-ECs, were phenotypically aberrant, more contractile and stiffer, and overexpressed endothelin 1(EDN1), myosin heavy chain, elastin and fibrillin. EDN1 inhibition or knockdown of EDN1-receptors reduced HIF2-SMC stiffness. Hif2A GOF heterozygous mice displayed early onset of pulmonary hypertension, had SMCs with more disorganized stress fibers and higher stiffness in their pulmonary arterial vascular smooth muscle cells, and more deformable main pulmonary arteries compared with wild type mice. We propose that targeting these vascular aberrations may benefit patients with HIF2A GOF and other conditions ofaugmented hypoxia signaling.

Purpose: To determine whether the absence of transglutaminase 2 enzyme (TG2) in TG2 knockout mice (TG2-/-) protect them against early age-related functional and histological arterial changes. Methods: Pulse wave velocity (PWV) was measured using non-invasive Doppler and mean arterial pressure (MAP) was measured in awake mice using tail-cuff system. Thoracic aortas were excised for evaluation of endothelial dependent vasodilation (EDV) by wire myography, as well as histological analyses. Results: PWV and MAP were similar in TG2-/-mice to age-matched wild type (WT) control mice. Old WT mice exhibited a markedly attenuated EDV as compared to young WT animals. The TG2-/-young and old mice had enhanced EDV responses (p

Background Ischemia-reperfusion injury is characterized by an increase in oxidative stress and leads to significant morbidity and death. The tyrosine kinase c-Abl is activated by oxidative stress and mediates processes that affect endothelial barrier function. We hypothesized treatment with the c-Abl inhibitor imatinib would be protective against ischemia-reperfusion injury in our ex vivo rabbit model. Methods Heart-lung blocs were harvested from rabbits and stored in cold in Perfadex (Vitrolife, Englewood, CO) for 18 hours. Blocs were reperfused for 2 hours in an ex vivo circuit with donor rabbit blood alone (untreated group, n = 7) or donor rabbit blood and 4 mg imatinib (treatment group, n = 10). Serial clinical variables measured every 15 minutes (arterial oxygen and carbon dioxide tension and mean pulmonary artery pressures) and biochemistry of tissue samples before and after reperfusion were assessed. Results Compared with untreated lungs, imatinib treatment improved physiologic parameters, including oxygen, carbon dioxide, and pulmonary artery pressures. Imatinib-treated lungs had less vascular barrier dysfunction as quantified by wet-to-dry weight ratios and bronchoalveolar lavage protein concentrations. Treated lungs showed less inflammation as measured by bronchoalveolar lavage myeloperoxidase assay, less mitochondrial reactive oxygen species production, and increased antioxidant catalase levels. Finally, imatinib protected lungs from DNA damage and p53 upregulation. Conclusions Imatinib treatment significantly improved the physiologic performance of reperfused lungs and biochemical indicators associated with reperfusion injury in this ex vivo model. Further study is necessary to elucidate the mechanism of tyrosine kinase inhibition in lungs exposed to ischemia and reperfusion.

Depressed right ventricular ejection fraction (RVEF) has clear prognostic significance in patients with pulmonary arterial hypertension (PAH). Accordingly, improvements in RVEF represent a desirable end-point in the development of PAH therapies. However, current methods for determination of RVEF require measurement of RV volume and are relatively complex and costly. Here, we validate a novel method for quantitative estimation of RVEF in rats based entirely upon analysis of readily available RV pressure waveforms that eliminates the need for simultaneous volume measurement and can be rapidly applied. Right ventricular pressure and volume (conductance catheter) measurements acquired from 15 rats (7 controls, 8 sugen/hypoxia PAH; 220–250 g) were used for the study. Over the same 10 beat interval, RVEF was directly measured from the volume signal and estimated from the pressure signal. Simultaneous measures were compared by linear regression and Bland-Altman analysis to define bias (accuracy) and precision. Measured RVEF ranged from 0.19 to 0.60 (mean 0.44 ± 0.10) and estimated from 0.19 to 0.52 (mean 0.42 ± 0.09). Across the dataset there was strong correlation (r2 = 0.813), with minimal bias (0.01) and an overall error of 20% consistent with acceptable accuracy and precision. Study results support the potential utility of a method based entirely upon analysis of the RV pressure waveform for assessing drug effects on RVEF in rat models of PAH.

Women who have had preeclampsia have increased cardiovascular disease risk; however, the mechanism(s) responsible for this association remain unclear. Microvascular damage sustained during a preeclamptic pregnancy may persist postpartum. The putative mechanisms mediating this dysfunction include a reduction in NO-dependent dilation and an increased sensitivity to angiotensin II. In this study, we evaluated endothelium-dependent dilation, angiotensin II sensitivity, and the therapeutic effect of angiotensin II receptor blockade (losartan) on endothelium-dependent dilation in vivo in the microvasculature of women with a history of preeclampsia (n=12) and control women who had a healthy pregnancy (n=12). We hypothesized that preeclampsia would have (1) reduced endothelium-dependent dilation, (2) reduced NO-mediated dilation, and (3) increased sensitivity to angiotensin II. We further hypothesized that localized losartan would increase endothelium-dependent vasodilation in preeclampsia. We assessed microvascular endothelium-dependent vasodilator function by measurement of cutaneous vascular conductance responses to graded infusion of acetylcholine (acetylcholine; 10 −7 –102 mmol/L) and a standardized local heating protocol in control sites and sites treated with 15 mmol/L L-NAME ( N G -nitro- l -arginine methyl ester; NO-synthase inhibitor) or 43 µmol/L losartan. Further, we assessed microvascular vasoconstrictor sensitivity to angiotensin II (10 −20 –10 −4 mol/L). Preeclampsia had significantly reduced endothelium-dependent dilation (−0.3±0.5 versus −1.0±0.4 log EC50 ; P P =0.006). Preeclampsia also had augmented vasoconstrictor sensitivity to angiotensin II (−10.2±1.3 versus −8.3±0.5; P =0.006). Angiotensin II type I receptor inhibition augmented endothelium-dependent vasodilation and NO-dependent dilation in preeclampsia but had no effect in healthy pregnancy. These data suggest that women who have had preeclampsia have persistent microvascular dysfunction postpartum, mediated, in part, by increased sensitivity to angiotensin II.

Drug-induced hepatitis is the leading reason that an approved drug is removed from the commercial market. Halogenated anesthetics can induce hepatitis in susceptible persons, and cytochrome p4502E1 (CYP2E1) enzymes responsible for their metabolism induce antibodies in addition to hepatitis. CYP2E1 antibodies detected in anesthetic hepatitis patients have been detected in patients with viral hepatitis, suggesting that these different forms of hepatitis could develop immune reactions to a common segment or epitope of CYP2E1. We have found a common MHC-restricted CYP2E1 epitope in anesthetic and viral hepatitis that is a dominant epitope in anesthetic hepatitis and is significantly associated with fibrosis in patients with viral hepatitis. Along with conformational epitopes, our identification of MHC-restricted CYP2E1 epitopes can be used to develop specific diagnostic tests for drug-induced or viral hepatitis or associated fibrosis or to predict individuals at risk for developing these diseases or their sequelae., Cytochrome p4502E1 (CYP2E1) autoantibodies are biomarkers for drug-induced hepatitis and chronic hepatitis C. However, major histocompatibility-restricted CYP2E1 epitopes associated with these diseases have not been identified. We hypothesized that CYP2E1 epitopes associated with different types of hepatitis may be shared and may impact immune responses and metabolism. SYFPEITHI epitope prediction identified CYP2E1 candidate epitopes that would be recognized by MHC II haplotypes. Candidate epitopes were tested for induction of hepatitis and CYP2E1 autoantibodies in mice and recognition by sera from patients with anesthetic drug-induced and viral hepatitis. Human liver cells treated with epitope hybridoma serum were analyzed for mitochondrial stress. CYP2E1 activity was measured in human microsomes similarly treated. Epitope antibodies in viral hepatitis sera were analyzed using linear regression to uncover associations with liver pathology. A P value of

Background In sickle cell disease (SCD), hemolysis results in the release and activation of arginase, an enzyme that reciprocally regulates nitric oxide (NO) synthase activity and thus, NO production. Simply supplementing the common substrate L-arginine, however, fails to improve NO bioavailability. In this study, we tested the hypothesis that arginase inhibition would improve NO bioavailability and thereby attenuate systemic and pulmonary vascular endothelial dysfunction in transgenic mice with SCD. Methods We studied 5-month-old transgenic sickle cell (SC) mice and age matched wild-type (WT) controls. SC mice were treated with the arginase inhibitor, 2(S)-amino-6-boronohexanoic acid (ABH; approximately 400 μg/d) for 4 weeks or left untreated. Results Vascular arginase activity was significantly higher at baseline in untreated SC mice compared to WT controls (SC versus WT, 346 ± 69.3 vs 69 ± 17.3 pmol urea/mg protein/minute; P = 0.0043; n = 4-5 animals per group). Treatment with ABH may significantly decrease arginase activity to levels near WT controls (SC + ABH 125.2 ± 17.3 pmol urea/mg protein/minute; P = 0.0213). Aortic strips from untreated SC mice showed decreased NO and increased reactive oxygen species (ROS) production (NO: fluorescence rate 0.76 ± 0.14 vs 1.34 ± 0.17 RFU/s; P = 0.0005 and ROS: fluorescence rate 3.96 ± 1.70 vs 1.63 ± 1.20 RFU/s, P = 0.0039; n = 3- animals per group). SC animals treated with ABH for 4 weeks demonstrated NO (fluorescence rate: 1.16 ± 0.16) and ROS (fluorescence rate: 2.02 ± 0.45) levels comparable with age-matched WT controls (n = 3- animals per group). The maximal endothelial-dependent vasorelaxation response to acetylcholine was impaired in aortic rings from SC mice compared with WT (57.7% ± 8.4% vs 80.3% ± 11.0%; P = 0.02; n = 6 animals per group). The endothelial-independent response was not different between groups. In SC mice, the right ventricular cardiac output index and end-systolic elastance were similar (4.60 ± 0.51 vs 2.9 ± 0.85 mL/min/100 g and 0.89 ± 0.48 vs 0.58 ± 0.11 mm Hg/μL), whereas the pulmonary vascular resistance index and right ventricular end-systolic pressure were greater (2.9 ± 0.28 vs 5.5 ± 2.0 mm Hg × min/μL/100 g and 18.9 ± 1.1 vs 23.1 ± 4.0 mm Hg; n = 8 animals per group). Pulse wave velocity (a measure of arterial stiffness) was greater in SC mice compared with WT (3.74 ± 0.54 vs 3.25 ± 0.21 m/s; n = 20 animals per group), arginase inhibition for 4 weeks significantly reduced the vascular SC phenotype to one similar to WT animals (P = 0.0009). Conclusions Arginase inhibition improves NO bioavailability and thereby attenuates systemic and pulmonary vascular endothelial dysfunction in transgenic mice with SCD. Therefore, arginase is a potential therapeutic target in the treatment of cardiovascular dysfunction in SCD.

Aging is associated with increased cardiomyocyte loss, left-ventricular hypertrophy, and the accumulation of extracellular matrix, which results in declining cardiac function. The role of the matrix crosslinking enzyme, tissue transglutaminase (TG2), in age-related myocardial stiffness, and contractile function remains incompletely understood. In this study, we examined the role of TG2 in cardiac function, and determined whether TG2 inhibition can prevent age-associated changes in cardiac function. Male Fisher rats (18-month-old) were administered the transglutaminase inhibitor cystamine (study group) or saline (age-matched controls) for 12 weeks via osmotic mini-pumps. Cardiac function was determined by echocardiography and invasive pressure-volume loops. Rat hearts were dissected out, and TG2 expression, activity, and S-nitrosation were determined. Young (6-month-old) males were used as controls. TG2 activity significantly increased in the saline-treated but not in the cystamine-treated aging rat hearts. TG2 expression also increased with age and was unaltered by cystamine treatment. Aged rats showed increased left ventricular (LV) end-systolic dimension and a decrease in fractional shortening compared with young, which was not affected by cystamine. However, cystamine treatment preserved the preload-independent index of LV filling pressure and restored end-diastolic pressure, end-diastolic pressure-volume relationships, and arterial elastance toward young. An increase in TG2 activity contributes to age-associated increase in diastolic stiffness, thereby contributing to age-associated diastolic dysfunction. TG2 may thus represent a novel target for age-associated diastolic heart failure.

Large arteries (especially the aorta) lose elasticity and thicken with aging and as a consequence of other conditions, thus leading to central arterial stiffening and associated adverse effects on blood flow and pressure. Arterial stiffness can be defined and measured in different ways, at a local level or systemically. Increases in either the intrinsic (material) stiffness or net structural (combined geometric and material) arterial stiffness, or both, can increase the velocity at which the pressure pulse travels along the arterial tree and central pulse pressure, which can negatively impact downstream resistance vessels and organs (ie, heart, brain, and kidney). Clarifying temporal and causal relationships between arterial stiffening and hypertension was identified by NHLBI as an important gap of knowledge, with a potential for clinical translation. NIH (National Institutes of Health)-funded studies, more than half of them supported by the NHLBI (Online Figure), have investigated various aspects of arterial stiffening in humans and in experimental models. To enable a more focused research effort on this topic, NHLBI launched a Request for Applications (RFA) HL-10 -027, entitled Cellular and Molecular Mechanisms of Arterial Stiffening and Its Relationship to Development of Hypertension (R01). This initiative supported 11 R01 awards during 2010 to 2015 (Online Table II; cumulative ≈ $20 million dollars in total costs), which represented a significant component of the overall NHLBI investment in this field. Here, we report a summary of important scientific findings that resulted from this NHLBI-initiated research effort, constituting the basis of > 200 original research and review articles (Online Table II), some highlighted here, many conference presentations, and several patents . In humans, increased arterial stiffness, or loss of elastic compliance of large arteries, has been linked to an increased risk of myocardial infarction, heart failure, stroke, and kidney disease, among other conditions. Pulse wave velocity (PWV), …

We utilized innovative textile technology to create tissue-engineered vascular grafts (TEVGs) comprised exclusively of rapidly degrading material poly(glycolic acid). Our new technology led to robust neotissue formation in the TEVGs, especially extracellular matrix formation, such as elastin. In addition, the rapid degradation of the polymer significantly reduced complications, such as stenosis or calcification, as seen with the use of slow degrading polymers in the majority of previous studies for aortic small diameter TEVGs.

Cytochrome p4502E1 (CYP2E1) autoantibodies are biomarkers for drug-induced hepatitis and chronic hepatitis C. However, major histocompatibility-restricted CYP2E1 epitopes associated with these diseases have not been identified. We hypothesized that CYP2E1 epitopes associated with different types of hepatitis may be shared and may impact immune responses and metabolism. SYFPEITHI epitope prediction identified CYP2E1 candidate epitopes that would be recognized by MHC II haplotypes. Candidate epitopes were tested for induction of hepatitis and CYP2E1 autoantibodies in mice and recognition by sera from patients with anesthetic drug-induced and viral hepatitis. Human liver cells treated with epitope hybridoma serum were analyzed for mitochondrial stress. CYP2E1 activity was measured in human microsomes similarly treated. Epitope antibodies in viral hepatitis sera were analyzed using linear regression to uncover associations with liver pathology. A P value of

The lysyl oxidase (LOX) family of enzymes regulates collagen cross-linking. LOX is upregulated in hypertension, increasing vascular stiffness. In vivo human research is sparse, as long-term LOX inhibition in animals causes vascular instability. Our aim was to evaluate the effects of LOX inhibition on cutaneous microvascular function to determine whether LOX function was upregulated in hypertensive humans. Four intradermal microdialysis fibers were placed in the forearm of 10 young [age: 24 ± 1 yr, mean arterial pressure (MAP): 87 ± 2 mmHg], 10 normotensive (age: 50 ± 2 yr, MAP: 84 ± 1 mmHg), and 10 hypertensive (age: 53 ± 2 yr, MAP: 112 ± 2 mmHg) subjects. Two sites were perfused with 10 mM β-aminopropionitrile (BAPN) to inhibit LOX. The remaining two sites were perfused with lactated Ringer solution (control). A norepinephrine dose response (10(−12)−10(−2) M) was performed to examine receptor-mediated vasoconstrictor function. A sodium nitroprusside dose response (10(−8)−10(−1.3) M) was performed to examine vascular smooth muscle vasodilator function. Red blood cell flux was measured via laser-Doppler flowmetry and normalized to cutaneous vascular conductance (flux/MAP). LogEC(50) values were calculated to determine changes in vasosensitivity. Skin tissue samples were analyzed for both extracellular matrix-bound and soluble LOX. LOX inhibition augmented vasoconstrictor sensitivity in young (control: −6.0 and BAPN: −7.1, P = 0.03) and normotensive (control: −4.8 and BAPN: −7.0, P = 0.01) but not hypertensive (control: −6.0 and BAPN: −6.1, P = 0.79) men and women. Relative to young subjects, extracellular matrix-bound LOX expression was higher in hypertensive subjects (young: 100 ± 8 and hypertensive: 162 ± 8, P = 0.002). These results suggest that upregulated LOX may contribute to the vascular stiffness and microvascular dysfunction characteristic in hypertension. NEW & NOTEWORTHY Matrix-bound lysyl oxidase (LOX) and LOX-like 2 expression are upregulated in the microvasculature of hypertensive men and women. Microvascular responsiveness to exogenous stimuli is altered with localized LOX inhibition in healthy men and women but not hypertensive adults. The LOX family differentially affects microvascular function in hypertensive and normotensive men and women.

Microvascular dysfunction originating during a preeclamptic pregnancy persists postpartum and probably contributes to increased CVD risk in these women. One putative mechanism contributing to this dysfunction is increased vasoconstrictor sensitivity to endothelin-1 (ET-1), mediated by alterations in ET-1 receptor type-B (ETBR). We evaluated ET-1 sensitivity, ETAR, and ETBR contributions to ET-1-mediated constriction, and the mechanistic role of ETBR in endothelium-dependent dilation in vivo in the microvasculature of postpartum women who had preeclampsia (PrEC, n=12) and control women who had a healthy pregnancy (HC, n=12). We hypothesized that (1) PrEC would have a greater vasoconstrictor response to ET-1, and (2) reduced ETBR-mediated dilation. We further hypothesized that ETBR-blockade would attenuate endothelium-dependent vasodilation in HC, but not PrEC. Microvascular reactivity was assessed by measurement of cutaneous vascular conductance responses to graded infusion of ET-1 (10−20–10−8 mol/l), ET-1 + 500 nmol/l BQ-123 (ETAR-blockade), and ET-1 + 300 nmol/l BQ-788 (ETBR-blockade), and during graded infusion of acetylcholine (ACh, 10−7–102 mmol/l) and a standardized local heating protocol with and without ETBR-inhibition. PrEC had an increased vasoconstriction response to ET-1 (P=0.02). PrEC demonstrated reduced dilation responses to selective ETBR stimulation with ET-1 (P=0.01). ETBR-inhibition augmented ET-1-mediated constriction in HC (P=0.01) but attenuated ET-1-mediated constriction in PrEC (P=0.003). ETBR-inhibition attenuated endothelium-dependent vasodilation responses to 100mmol/l ACh (P=0.04) and local heat (P=0.003) in HC but increased vasodilation (ACh: P=0.01; local heat: P=0.03) in PrEC. Women who have had preeclampsia demonstrate augmented vasoconstrictor sensitivity to ET-1, mediated by altered ETBR signaling. Furthermore, altered ETBR function contributes to diminished endothelium-dependent dilation in previously preeclamptic women.

Background The structural elements of the vascular wall, namely, extracellular matrix and smooth muscle cells ( SMC s), contribute to the overall stiffness of the vessel. In this study, we examined the crosslinking‐dependent and crosslinking‐independent roles of tissue transglutaminase ( TG 2) in vascular function and stiffness. Methods and Results SMC s were isolated from the aortae of TG 2−/− and wild‐type ( WT ) mice. Cell adhesion was examined by using electrical cell–substrate impedance sensing and PicoGreen assay. Cell motility was examined using a Boyden chamber assay. Cell proliferation was examined by electrical cell–substrate impedance sensing and EdU incorporation assays. Cell micromechanics were studied using magnetic torsion cytometry and spontaneous nanobead tracer motions. Aortic mechanics were examined by tensile testing. Vasoreactivity was studied by wire myography. SMC s from TG 2−/− mice had delayed adhesion, reduced motility, and accelerated de‐adhesion and proliferation rates compared with those from WT . TG 2−/− SMC s were stiffer and displayed fewer cytoskeletal remodeling events than WT . Collagen assembly was delayed in TG 2−/− SMC s and recovered with adenoviral transduction of TG 2. Aortic rings from TG 2−/− mice were less stiff than those from WT ; stiffness was partly recovered by incubation with guinea pig liver TG 2 independent of crosslinking function. TG 2−/− rings showed augmented response to phenylephrine‐mediated vasoconstriction when compared with WT . In human coronary arteries, vascular media and plaque, high abundance of fibronectin expression, and colocalization with TG 2 were observed. Conclusions TG 2 modulates vascular function/tone by altering SMC contractility independent of its crosslinking function and contributes to vascular stiffness by regulating SMC proliferation and matrix remodeling.

Stiffening of conduit arteries is a risk factor for cardiovascular morbidity. Aortic wall stiffening increases pulsatile hemodynamic forces that are detrimental to the microcirculation in highly perfused organs, such as the heart, brain, and kidney. Arterial stiffness is associated with hypertension but presumed to be due to an adaptive response to increased hemodynamic load. In contrast, a recent clinical study found that stiffness precedes and may contribute to the development of hypertension although the mechanisms underlying hypertension are unknown. Here, we report that in a diet-induced model of obesity, arterial stiffness, measured in vivo, develops within 1 month of the initiation of the diet and precedes the development of hypertension by 5 months. Diet-induced obese mice recapitulate the metabolic syndrome and are characterized by inflammation in visceral fat and aorta. Normalization of the metabolic state by weight loss resulted in return of arterial stiffness and blood pressure to normal. Our findings support the hypothesis that arterial stiffness is a cause rather than a consequence of hypertension.

Objectives Ischemia/reperfusion injury (IRI) is a common complication of lung transplantation (LTx). Hydrogen sulfide (H2S) is a novel agent previously shown to slow metabolism and scavenge reactive oxygen species, potentially mitigating IRI. We hypothesized that pretreatment with inhaled H2S would improve graft function in an ex vivo model of LTx. Methods Rabbits (n = 10) were ventilated for 2 h prior to heart-lung bloc procurement. The treatment group (n = 5) inhaled room air (21% O2) supplemented with 150 ppm H2S while the control group (n = 5) inhaled room air alone. Both groups were gradually cooled to 34°C. All heart-lung blocs were then recovered and cold-stored in low-potassium dextran solution for 18 h. Following storage, the blocs were reperfused with donor rabbit blood in an ex vivo apparatus. Serial clinical parameters were assessed and serial tissue biochemistry was examined. Results Prior to heart-lung bloc procurement, rabbits pretreated with H2S exhibited similar oxygenation (P = 0.1), ventilation (P = 0.7), and heart rate (P = 0.5); however, treated rabbits exhibited consistently higher mean arterial blood pressures (P = 0.01). During reperfusion, lungs pretreated with H2S had better oxygenation (P

Background Endothelial dysfunction and arterial stiffening play major roles in cardiovascular diseases. The critical role for the miR-181 family in vascular inflammation has been documented. Here we tested whether the miR-181 family can influence the pathogenesis of hypertension and vascular stiffening. Methods and results qPCR data showed a significant decrease in miR-181b expression in the aorta of the older mice. Eight miR-181a1/b1-/- mice and wild types (C57BL6J:WT) were followed weekly for pulse wave velocity (PWV) and blood pressure measurements. After 20 weeks, the mice were tested for endothelial function and aortic modulus. There was a progressive increase in PWV and higher systolic blood pressure in miR-181a1/b1-/- mice compared with WTs. At 21 weeks, aortic modulus was significantly greater in the miR-181a1/b1-/- group, and serum TGF-β was found to be elevated at this time. A luciferase reporter assay confirmed miR-181b targets TGF-βi (TGF-β induced) in the aortic VSMCs. In contrast, wire myography revealed unaltered endothelial function along with higher nitric oxide production in the miR-181a1/b1-/- group. Cultured VECs and VSMCs from the mouse aorta showed more secreted TGF-β in VSMCs of the miR-181a1/b1-/- group; whereas, no change was observed from VECs. Circulating levels of angiotensin II were similar in both groups. Treatment with losartan (0.6 g/L) prevented the increase in PWV, blood pressure, and vascular stiffness in miR-181a1/b1-/- mice. Immunohistochemistry and western blot for p-SMAD2/3 validated the inhibitory effect of losartan on TGF-β signaling in miR-181a1/b1-/- mice. Conclusions Decreased miR-181b with aging plays a critical role in ECM remodeling by removing the brake on the TGF-β, pSMAD2/3 pathway.

Background Ischemia–reperfusion injury is a common complication after lung transplantation. Ischemia–reperfusion injury is thought to be mediated by reactive oxygen species (ROS). Hydrogen sulfide (H2S) is a novel agent that has been previously shown to scavenge ROS and slow metabolism. We evaluated the effect of infused H2S on the presence of ROS after reperfusion in an ex vivo model of lung transplantation. Methods Heart–Lung blocks were recovered from New Zealand white rabbits (n = 12) and cold stored in Perfadex solution for 18 h. After storage, the heart–lung blocks were reperfused ex vivo with donor rabbit blood. In the treatment group (n = 7), a bolus of sodium H2S was added at the beginning of reperfusion (100 μg/kg) and continuously infused throughout the 2-h experiment (1 mg/kg/h). The vehicle group (n = 5) received an equivalent volume of saline. Serial airway and pulmonary artery pressures and arterial and venous blood gases were measured. Results Oxygenation and pulmonary artery pressures were similar between the 2 groups. However, treatment with H2S resulted in a dramatic reduction in the presence of ROS after 2 h of reperfusion (4,851 ± 2,139 versus 235 ± 462 related fluorescence units/mg protein; P = 0.003). A trend was seen toward increased levels of cyclic guanosine monophosphate in the H2S-treated group (3.08 ± 1.69 versus 1.73 ± 1.41 fmol/mg tissue; P = .23). Conclusions After prolonged ischemia, infusion of H2S during reperfusion was associated with a significant decrease in the presence of ROS, a suspected mediator of ischemia–reperfusion injury. To our knowledge, the present study represents the first reported therapeutic use of H2S in an experimental model of lung transplantation.

Introduction Reduced nitric oxide (NO) bioavailability and increased generation of reactive oxygen species and inflammation with advancing age play a major role in the development of cardiovascular disease (CVD) risk factors of hypertension, endothelial dysfunction and arterial stiffness. Therefore, increase in NO bioavailability by supplementing inorganic nitrite and improve in antioxidant defense system by supplementing N-acetylcysteine (NAC) could protect against the CVD. Hypertension is highly prevalent in a population and cause endothelial dysfunction and arterial stiffness. Hence spontaneously hypertensive rats (SHR) were used in this study as a model for hypertension-mediated endothelial dysfunction and arterial stiffness. Hypothesis Treatment of SHR with nitrite or NAC will reduce blood pressure, arterial stiffness, and endothelial dysfunction. Methods SHR (18 weeks) and normotensive Wistar Kyoto (WKY) rats were divided into 4 groups: 1) WKY, 2) SHR-cont, 3) SHR-nitrite and 4) SHR-NAC. Group 3 rats received nitrite (60 mg/L) in drinking water and group 4 rats received NAC pH 7.2 (1.2 g/L) for 3 months. Mean arterial pressure (MAP; tail-cuff method) and pulse wave velocity (PWV, a measure of arterial stiffness) were measured. Aortic endothelial function was determined with a wire myograph. Results MAP & PWV of SHR-cont were 1.8-fold and 1.6-fold higher compared with WKY respectively. MAP was modestly decreased in nitrite and NAC groups compared with that of SHR-cont. PWV did not decrease significantly with NAC or nitrite treatment. Endothelium dependent vasorelaxation was significantly decreased in SHR-cont compared with WKY. This decreased vasorelaxation was restored to WKY level in both nitrite and NAC groups. Conclusion Nitrite and NAC treatments completely protected against the hypertension induced endothelium dysfunction in SHR. Nitrite and NAC appears to be very promising dietary components to protect against age-associated endothelial dysfunction.

Essential hypertension is a proinflammatory, proconstrictor disease coinciding with endothelial dysfunction and inward vessel remodeling. Using the skin circulation, our aim was to determine whether inducible NO synthase (iNOS) upregulation attenuates NO-dependent cutaneous vasodilation in hypertensive humans. We hypothesized that, with hypertension, localized iNOS inhibition would restore vasodilation in response to NO-dependent stimuli, and iNOS expression would be increased and phosphorylated vasodilator-stimulated phosphoprotein would be decreased. For, in vivo protocols, 4 intradermal microdialysis fibers were placed in 9 hypertensive and 10 normotensive men and women (systolic blood pressure: 146±4 versus 113±2 mm Hg; P N ω - propyl - l -arginine), and nonselective NOS inhibited ( N G -nitro- l -arginine methyl ester). Cutaneous vascular conductance was calculated (percentage of sodium nitroprusside) during standardized local heating (42°C) and acetylcholine dose-response protocols (0.01, 0.10, 1.00, 5.00, 10.00, 50.00, 100.00 mmol/L). The NO-dependent local heating response was attenuated at control (95±2% versus 76±2% cutaneous vascular conductance; P P P P P P =0.006) and phosphorylated vasodilator-stimulated phosphoprotein was decreased in skin from hypertensive humans ( P =0.04). These data suggest that iNOS is upregulated in essential hypertensive humans and contributes to reduced NO-dependent cutaneous vasodilation.

Elevated low-density lipoproteins (LDLs) are associated with vascular dysfunction evident in the cutaneous microvasculature. We hypothesized that uncoupled endothelial nitric oxide synthase (NOS3) through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic (HC) humans and that a statin intervention would decrease arginase activity. Five microdialysis fibres were placed in the skin of nine normocholesterolaemic (NC: LDL level 95±4 mg dl⁻¹) and nine hypercholesterolaemic (HC: LDL: 177±6 mg dl⁻¹) men and women before and after 3 months of systemic atrovastatin. Sites served as control, NOS inhibited, arginase inhibited, L-arginine supplemented and arginase inhibited plus L-arginine supplemented. Skin blood flow was measured while local skin heating (42°C) induced NO-dependent vasodilatation. L-NAME was infused after the established plateau in all sites to quantify NO-dependent vasodilatation. Data were normalized to maximum cutaneous vascular conductance (CVC(max)). Skin samples were obtained to measure total arginase activity and arginase I and arginase II protein. Vasodilatation was reduced in hyperocholesterolaemic subjects (HC: 76±2 vs. NC: 94±3%CVC(max), P 0.05). Arginase activity and protein were increased in HC skin (P < 0.05 vs. NC) and activity decreased with atrovastatin treatment (P < 0.05). Reduced NOS3 substrate availability through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic humans, which is corrected with atorvastatin therapy.

A Wireless Mesh Network (WMN) is a promising way of providing low-cost broadband Internet access. The underlying routing protocol naively assumes that all the nodes in the network are non-malicious. The open architecture of WMN, multi-hop nature of communication, different management styles, and wireless communication paves way to malicious attackers. The attackers can exploit hidden loopholes in the multipath mesh routing protocol to have a suction attack called the blackhole attack. The attacker can falsify routing metrics such as the shortest transmission time to reach any destination and thereby suck the network traffic. We propose a novel strategy by employing mobile honeypot agents that utilize their topological knowledge and detect such spurious route advertisements. They are deployed as roaming software agents that tour the network and lure attackers by sending route request advertisements. We collect valuable information on attacker's strategy from the intrusion logs gathered at a given honeypot. We finally evaluate the effectiveness of the proposed architecture using simulation in ns-2.

Extracorporeal circulation provides critical life support in the face of cardiopulmonary or renal failure, but it also introduces a host of unique morbidities characterized by edema formation, cardiac insufficiency, autonomic dysfunction, and altered vasomotor function. We tested the hypothesis that cyclohexanone (CHX), a solvent used in production of extracorporeal circuits and intravenous (IV) bags, leaches into the contained fluids and can replicate these clinical morbidities. Crystalloid fluid samples from circuits and IV bags were analyzed by gas chromatography-mass spectrometry to provide a range of clinical CHX exposure levels, revealing CHX contamination of sampled fluids (9.63–3,694 μg/l). In vivo rat studies were conducted ( n = 49) to investigate the effects of a bolus IV infusion of CHX vs. saline alone on cardiovascular function, baroreflex responsiveness, and edema formation. Cardiovascular function was evaluated by cardiac output, heart rate, stroke volume, vascular resistance, arterial pressure, and ventricular contractility. Baroreflex function was assessed by mean femoral arterial pressure responses to bilateral carotid occlusion. Edema formation was assessed by the ratio of wet to dry organ weights for lungs, liver, kidneys, and skin. CHX infusion led to systemic hypotension; pulmonary hypertension; depressed contractility, heart rate, stroke volume, and cardiac output; and elevated vascular resistance ( P < 0.05). Mean arterial pressure responsiveness to carotid occlusion was dampened after CHX infusion (from +17.25 ± 1.8 to +5.61 ± 3.2 mmHg; P < 0.05). CHX infusion led to significantly higher wet-to-dry weight ratios vs. saline only (3.8 ± 0.06 vs. 3.5 ± 0.05; P < 0.05). CHX can reproduce clinical cardiovascular, neurological, and edema morbidities associated with extracorporeal circulatory treatment.

A wireless mesh network is a wired extension of a multihop ad hoc network that defines a new paradigm for broadband wireless Internet access. A packet originating from a mesh client is relayed collaboratively in a multihop fashion by the intermediate mesh routers toward an Internet gateway. All existing mesh routing protocols assume that each MR honestly participates in packet forwarding. This is valid only in a network managed by a single trusted authority. However, a community-based WMN can be formed by a group of independent MRs operated by different service providers. It is a real challenge to establish a priori trust in a multi-operator WMN. In such a situation, a selfish MR might be motivated to monopolize the wireless channel for itself by intentionally dropping others' packets. This results in severe performance degradation. Thus, enforcing collaboration is a determinant aspect in designing a secure and reliable WMN. In this article we analyze selfishness of MRs in a multi-operator WMN and explore its overall negative impact on network performance. We finally present a summary of various existing schemes with respect to detecting selfishness, analyze their usefulness in WMNs, and highlight their relative advantages and deficiencies.

We present a protocol for measuring in vivo aortic stiffness in mice using high-resolution ultrasound imaging. Aortic diameter is measured by ultrasound and aortic blood pressure is measured invasively with a solid-state pressure catheter. Blood pressure is raised then lowered incrementally by intravenous infusion of vasoactive drugs phenylephrine and sodium nitroprusside. Aortic diameter is measured for each pressure step to characterize the pressure-diameter relationship of the ascending aorta. Stiffness indices derived from the pressure-diameter relationship can be calculated from the data collected. Calculation of arterial compliance is described in this protocol. This technique can be used to investigate mechanisms underlying increased aortic stiffness associated with cardiovascular disease and aging. The technique produces a physiologically relevant measure of stiffness compared to ex vivo approaches because physiological influences on aortic stiffness are incorporated in the measurement. The primary limitation of this technique is the measurement error introduced from the movement of the aorta during the cardiac cycle. This motion can be compensated by adjusting the location of the probe with the aortic movement as well as making multiple measurements of the aortic pressure-diameter relationship and expanding the experimental group size.

Hydrogen sulfide (H2S) has emerged as an important gasotransmitter in the vasculature. In this study, we tested the hypothesis that H2S contributes to coronary vasoregulation and evaluated the physiological relevance of two sources of H2S, namely, cystathionine-γ-lyase (CSE) and 3-mercaptypyruvate sulfertransferase (MPST). MPST was detected in human coronary artery endothelial cells as well as rat and mouse coronary artery; CSE was not detected in the coronary vasculature. Rat coronary artery homogenates produced H2S through the MPST pathway but not the CSE pathway in vitro. In vivo coronary vasorelaxation response was similar in CSE knockout mice, wild-type mice (WT), and WT mice treated with the CSE inhibitor propargylglycine, suggesting that CSE-produced H2S does not have a significant role in coronary vasoregulation in vivo. Ex vivo, the MPST substrate 3-mercaptopyruvate (3-MP) and H2S donor sodium hydrosulfide (NaHS) elicited similar coronary vasoreactivity responses. Pyruvate did not have any effects on vasoreactivity. The vasoactive effect of H2S appeared to be nitric oxide (NO) dependent: H2S induced coronary vasoconstriction in the presence of NO and vasorelaxation in its absence. Maximal endothelial-dependent relaxation was intact after 3-MP and NaHS induced an increase in preconstriction tone, suggesting that endothelial NO synthase activity was not significantly inhibited. In vitro, H2S reacted with NO, which may, in part explain the vasoconstrictive effects of 3-MP and NaHS. Taken together, these data show that MPST rather than CSE generates H2S in coronary artery, mediating its effects through direct modulation of NO. This has important implications for H2S-based therapy in healthy and diseased coronary arteries.

Wireless Mesh Networking (WMN), the static mesh routers (MRs) cooperatively relay each other packets to the Internet Gateway (IGW). The routing protocols assume all the nodes in the network to be non-malicious. However, the open architecture of WMNs paves way to malicious attackers who can exploit hidden loopholes in the routing protocol. In this paper, we mainly focus on the vulnerability of the network to a suction attack called blackhole attack. In detecting such attacks, we explore the use of intelligent agents called Honeypots which are roaming virtual software agents that generate a dummy Route Request (RREQ) packets to lure and trap blackhole attackers. We illustrate the performance of our proposed detection approach by extensive simulation results using the ns-2 simulator.

In the recent years, wireless mesh network (WMN) has evolved as a new paradigm for broadband wireless Internet access. The self-configurability, open wireless infrastructure, and different management styles of WMN make them vulnerable to malicious attackers. As a first step to secure WMNs, it is critical to incorporate an authentication mechanism for mesh clients. The existing proposals are primarily based on public key certificates, which incur considerable overhead in signature verification. We propose a network layer authentication mechanism called Merkle Tree based Mesh Authentication Protocol (MT-MAP) for WMNs. It incorporates inexpensive hash operations using Merkle tree to authenticate single/multihop mesh clients. We also show how the use of hash tree facilitates fast and periodic refresh of authentication certificates. Finally, we present a security analysis of MT-MAP against impersonation and replay attacks.

In the last few years, Wireless Sensor Networks (WSNs) have emerged as a disrupting technology for myriad military and civilian applications. They demand an accurate location of the event detected and is done by using a mobile beacon node to provide accurate location and assume a benign environment. However, in a hostile military environment, such a node can be easily tampered by an adversary. In this paper, we propose a distributed cluster based anomaly detection scheme by assigning few randomly chosen cluster heads a critical task of estimating the reliability of the mobile beacon node. As localization of remaining nodes is cautiously performed only after verifying the authenticity of the mobile beacon node, a considerable overhead is saved in the incorrect localization of the entire network. We perform extensive simulation for different attacks and observe our scheme to have a high detection rate of 99% and a very less false positive rate of 20%.

Wireless mesh network (WMN) is evolving to be a new paradigm for wireless Internet connectivity as it obviates the need for wired infrastructure at every access point (AP) a.k.a mesh router (MR). The MRs collaboratively forward the traffic towards the Internet gateway (IGW). The self-configurable architecture of MRs paves way for malicious intruders to conduct a denial-of-service attack (DoS) on the MRs by flooding the network with a large volume of traffic; thus rendering the system inaccessible to the real users. In this paper, we present a cache based defense at the MRs to identify flooding style DoS attacks. We use a most frequently used cache mechanism to identify such flows and raise an early alert to curb them. We effectively avert any performance degradation by dropping the identified attack flows along the forwarding routers. Simulation results indicate that our scheme offers an active line of defense against DoS attacks

Wireless Mesh Networks (WMNs) are evolving to be the key technology of the future. As the WMNs are envisioned to provide high bandwidth broadband service to a large community of users, the Internet Gateway (IGW) which acts as a central point of internet attachment for the mesh routers, it is likely to be a potential bottleneck because of its limited wireless link capacity. We propose a novel technique that elegantly balances the load among the different IGWs in a WMN. We switch the point of attachment of an active source serviced gateway depending on the average queue length at the IGW. The proposed load balancing scheme includes: an initial gateway discovery module, which determines a primary gateway for a mesh router and a load balancing module that rebalances the load among the gateways. We use ns-2 [9] for evaluating our proposed scheme and we observe that the proposed scheme is able to balance the traffic efficiently.

Wireless Mesh Networks (WMNs) are evolving to be the key technology of the future. The self-configuring nature of WMNs and the ease, with which a mesh router/mesh point can be added, makes it pertinent to ensure their secure operation. All the routing protocols in WMNs naively assume the nodes to be co-operative in forwarding each other's packets. However, a node can behave selfishly by discretely dropping other's packets, in an attempt to maximize its throughput. In this paper, we present a distributed scheme called, Distributed Self-policing Architecture for Fostering Node Cooperation (D-SAFNC), for enforcing cooperation among the nodes in a WMN. We use a distributed approach in isolating any selfish node with the help of localized detection agents called sink nodes. We study the effectiveness of our scheme through simulations using ns-2 which reaffirm that D-SAFNC can successfully prevent any performance degradation due to the presence of selfish nodes.

The online version of the original article can be found underdoi:10.1007/s00421-011-2257-9.M.Khan J.Steppan S.Ryoo L.Santhanam T.Berkowitz D. Nyhan D. E. Berkowitz (&)Department of Anesthesiology and Critical Care Medicine,Johns Hopkins University Medicine, Tower 711, 600 N Wolfe St.,Baltimore, MD 21287, USAe-mail: dberkowi@bme.jhu.eduK. H. SchuleriDepartment of Medicine, Johns Hopkins University Medicine,600 N Wolfe St., Baltimore, MD 21287, USAE. Tuday L. Bugaj A. A. Shoukas D. E. BerkowitzDepartment of Biomedical Engineering,Johns Hopkins University Medicine, 600 N Wolfe St.,Baltimore, MD 21287, USA

Hypertension (HTN) increases the risk for cardiac disease and stroke. Endogenous hydrogen sulfide (H 2 S), which plays a prominent role in a multitude of pathologies like inflammation/sepsis, hypertension, peripheral and cerebro-vascular, and coronary artery disease, is now well characterized as a physiologic vasodilator [1] . H 2 S is produced by cystathionine- γ -lyase (CSE) in vascular endothelium. It caters to relaxation through sulfhydration (posttranslational modification) of IK, SK and K ATP channels, resulting in vascular smooth muscle cell hyperpolarization. This effect is independent of the Nitric Oxide (NO)/cGMP/PKG axis [2] . Tang et al. studied H 2 S in spontaneously hypertensive (SHR) rats and demonstrated a loss of CSE/H 2 S in HTN and a decrease in BP (blood pressure) on substitution of H 2 S [3] . Aim To elicit if loss of endogenous H 2 S contributes to the pathogenesis of hypertension. Results Tail cuff was used to measure BP in SHR and control Wistar Kyoto (WKY) rats starting at age 4 weeks (w) up until SHRs became hypertensive. Infusion of glybenclamide (20 mg/kg) a K ATP channel blocker, revealed a significant increase in systolic BP (SBP) in 4-week-old SHR (ΔSBP = 43.16 ± 24.24%, n = 5) and 4 w WKY (ΔSBP = 89.97 ± 41.40%, n = 4), while little change was demonstrated in hypertensive 90 w SHR (ΔSBP = 24.07 ± 21.99%, n = 4), measured invasively via aortic catheterization.In a separate experiments, aortic rings isolated from age matched SHRs ( n = 5) and WKYs ( n = 5), were mounted on a myograph, prepared in physiologic buffer at 37 °C, and constricted with phenylephrine (1 μM), for assessment of endothelial function. Acetylcholine (ACH) dependent maximum relaxation on treatment with L-Name (eNOS inhibitor) was modestly attenuated in aortas of 4 w WKY (92.79 ± 2.64–83.26 ± 3.35%), 4 w SHR (83.50 ± 4.47–59.48 ± 8.75%) and 90-week-old WKYs (71.39 ± 15.06–15.77 ± 5.16%). Contrary to that, this response was completely blocked in 90 w SHRs (35.32 ± 5.16–0.24 ± 3.17%). Treatment with propargylglycine (CSE blocker) did not affect ACH mediated maximum relaxation in 90 w SHR (35.32 ± 5.16–29.40 ± 5.86%), but it significantly attenuated the responses in 4 w WKYs (92.79 ± 2.64–56.30 ± 4.55), 4 w SHRs (83.50 ± 4.47–71.63 ± 12.25) and 90 w SHRs (71.39 ± 15.06–8.13 ± 4.61%). Strikingly, rings from normotensive 16 w SHR had a complete L-Name sensitive attenuation of endothelial relaxation (63.25 ± 5.15–2.74 ± 1.75%). Conclusion There appears to be a NO independent component of endothelial relaxation in normotensive blood vessels, which is sensitive to CSE inhibitors and blockage of K ATP channels. This NO independent component of relaxation is absent in HTN and pre-HTN vessels; hence implying that loss of the CSE/H 2 S/K ATP axis could be a mechanism preceding HTN. We aims to measure CSE activity and sulfhydration of Kir 6.1-Cysteine 43 and GAPDH in normotensive/pre and post HTN blood vessels in different animal models of HTN to elicit if CSE/H 2 S/K ATP axis is a potential target for the treatment of hypertension.

Novel Nebulized Arginase Inhibitor Increases Human Lung Compliance during Normothermic Ex Vivo Lung Perfusion T.J. George, G.J. Arnaoutakis, C.A. Beaty, S.K. Jandu, L. Santhanam, D.E. Berkowitz, A.S. Shah. The Division of Cardiac Surgery, The Johns Hopkins Medical Institutions, Baltimore, MD; The Department of Anesthesiology and Critical Care Medicine, The Johns Hopkins Medical Institutions, Baltimore, MD.

Background Vascular aging is closely associated with increased vascular stiffness. It has recently been demonstrated that decreased nitric oxide ( NO )‐induced S‐nitrosylation of tissue transglutaminase ( TG 2) contributes to age‐related vascular stiffness. In the current study, we tested the hypothesis that exercise restores NO signaling and attenuates vascular stiffness by decreasing TG 2 activity and cross‐linking in an aging rat model. Methods and Results Rats were subjected to 12 weeks of moderate aerobic exercise. Aging was associated with diminished phosphorylated endothelial nitric oxide synthase and phosphorylated vasodilator‐stimulated phosphoprotein abundance, suggesting reduced NO signaling. TG 2 cross‐linking activity was significantly increased in old animals, whereas TG 2 abundance remained unchanged. These alterations were attenuated in the exercise cohort. Simultaneous measurement of blood pressure and pulse wave velocity ( PWV ) demonstrated increased aortic stiffness in old rats, compared to young, at all values of mean arterial pressure ( MAP ). The PWV ‐ MAP correlation in the old sedentary and old exercise cohorts was similar. Tensile testing of the vessels showed increased stiffness of the aorta in the old phenotype with a modest restoration of mechanical properties toward the young phenotype with exercise. Conclusions Increased vascular stiffness during aging is associated with decreased TG 2 S‐nitrosylation, increased TG 2 cross‐linking activity, and increased vascular stiffness likely the result of decreased NO bioavailability. In this study, a brief period of moderate aerobic exercise enhanced NO signaling, attenuated TG cross‐linking activity, and reduced ex vivo tensile properties, but failed to reverse functional vascular stiffness in vivo, as measured by PWV .