The example of CaPSURE: lessons learned from a national disease registry

IntroductionAlthough randomized controlled trials (RCTs) remain the gold standard for determining evidence-based clinical practices, large disease registries that enroll large numbers of patients have become paramount as a relatively cost-effective additional tool.MethodsWe highlight the advantages of disease registries focusing on the example of prostate cancer and the Cancer of the Prostate Strategic Urologic Research Endeavor (CaPSURE™) registry.ResultsCaPSURE collects approximately 1,000 clinical and patient-reported variables, in over 13,000 men that are enrolled. Thus far, CaPSURE has yielded over 130 peer-reviewed publications, with several others in press, in key areas of risk migration, practice patterns, outcome prediction, and quality of life outcomes.ConclusionsDisease registries, like CaPSURE complement RCTs and CaPSURE, have provided a means to better understand many aspects of prostate cancer epidemiology, practice patterns, oncologic and HRQOL outcomes, and costs of care across populations. Specialized observational disease registries such as CaPSURE provide insight and have broad implications for disease management and policy

C-Peptide Increases Na,K-ATPase Expression via PKC- and MAP Kinase-Dependent Activation of Transcription Factor ZEB in Human Renal Tubular Cells

Replacement of proinsulin C-peptide in type 1 diabetes ameliorates nerve and kidney dysfunction, conditions which are associated with a decrease in Na,K-ATPase activity. We determined the molecular mechanism by which long term exposure to C-peptide stimulates Na,K-ATPase expression and activity in primary human renal tubular cells (HRTC) in control and hyperglycemic conditions.HRTC were cultured from the outer cortex obtained from patients undergoing elective nephrectomy. Ouabain-sensitive rubidium ((86)Rb(+)) uptake and Na,K-ATPase activity were determined. Abundance of Na,K-ATPase was determined by Western blotting in intact cells or isolated basolateral membranes (BLM). DNA binding activity was determined by electrical mobility shift assay (EMSA). Culturing of HRTCs for 5 days with 1 nM, but not 10 nM of human C-peptide leads to increase in Na,K-ATPase α(1)-subunit protein expression, accompanied with increase in (86)Rb(+) uptake, both in normal- and hyperglycemic conditions. Na,K-ATPase α(1)-subunit expression and Na,K-ATPase activity were reduced in BLM isolated from cells cultured in presence of high glucose. Exposure to1 nM, but not 10 nM of C-peptide increased PKCε phosphorylation as well as phosphorylation and abundance of nuclear ERK1/2 regardless of glucose concentration. Exposure to 1 nM of C-peptide increased DNA binding activity of transcription factor ZEB (AREB6), concomitant with Na,K-ATPase α(1)-subunit mRNA expression. Effects of 1 nM C-peptide on Na,K-ATPase α(1)-subunit expression and/or ZEB DNA binding activity in HRTC were abolished by incubation with PKC or MEK1/2 inhibitors and ZEB siRNA silencing.Despite activation of ERK1/2 and PKC by hyperglycemia, a distinct pool of PKCs and ERK1/2 is involved in regulation of Na,K-ATPase expression and activity by C-peptide. Most likely C-peptide stimulates sodium pump expression via activation of ZEB, a transcription factor that has not been previously implicated in C-peptide-mediated signaling. Importantly, only physiological concentrations of C-peptide elicit this effect

Quantifying the Evolution of Vascular Barrier Disruption in Advanced Atherosclerosis with Semipermeant Nanoparticle Contrast Agents

Acute atherothrombotic occlusion in heart attack and stroke implies disruption of the vascular endothelial barrier that exposes a highly procoagulant intimal milieu. However, the evolution, severity, and pathophysiological consequences of vascular barrier damage in atherosclerotic plaque remain unknown, in part because quantifiable methods and experimental models are lacking for its in vivo assessment.To develop quantitative nondestructive methodologies and models for detecting vascular barrier disruption in advanced plaques.Sustained hypercholesterolemia in New Zealand White (NZW) rabbits for >7-14 months engendered endothelial barrier disruption that was evident from massive and rapid passive penetration and intimal trapping of perfluorocarbon-core nanoparticles (PFC-NP: ∼250 nm diameter) after in vivo circulation for as little as 1 hour. Only older plaques (>7 mo), but not younger plaques (<3 mo) demonstrated the marked enhancement of endothelial permeability to these particles. Electron microscopy revealed a complex of subintimal spongiform channels associated with endothelial apoptosis, superficial erosions, and surface-penetrating cholesterol crystals. Fluorine ((19)F) magnetic resonance imaging and spectroscopy (MRI/MRS) enabled absolute quantification (in nanoMolar) of the passive permeation of PFC-NP into the disrupted vascular lesions by sensing the unique spectral signatures from the fluorine core of plaque-bound PFC-NP.The application of semipermeant nanoparticles reveals the presence of profound barrier disruption in later stage plaques and focuses attention on the disrupted endothelium as a potential contributor to plaque vulnerability. The response to sustained high cholesterol levels yields a progressive deterioration of the vascular barrier that can be quantified with fluorine MRI/MRS of passively permeable nanostructures. The possibility of plaque classification based on the metric of endothelial permeability to nanoparticles is suggested

Endothelial cells use dynamic actin to facilitate lymphocyte transendothelial migration and maintain the monolayer barrier

The vascular endothelium is a highly dynamic structure, and the integrity of its barrier function is tightly regulated. Normally impenetrable to cells, the endothelium actively assists lymphocytes to exit the bloodstream during inflammation. The actin cytoskeleton of the endothelial cell (EC) is known to facilitate transmigration, but the cellular and molecular mechanisms are not well understood. Here we report that actin assembly in the EC, induced by Arp2/3 complex under control of WAVE2, is important for several steps in the process of transmigration. To begin transmigration, ECs deploy actin-based membrane protrusions that create a cup-shaped docking structure for the lymphocyte. We found that docking structure formation involves the localization and activation of Arp2/3 complex by WAVE2. The next step in transmigration is creation of a migratory pore, and we found that endothelial WAVE2 is needed for lymphocytes to follow a transcellular route through an EC. Later, ECs use actin-based protrusions to close the gap behind the lymphocyte, which we discovered is also driven by WAVE2. Finally, we found that ECs in resting endothelial monolayers use lamellipodial protrusions dependent on WAVE2 to form and maintain contacts and junctions between cells

Vascular Dysfunction in Horses with Endocrinopathic Laminitis

Endocrinopathic laminitis (EL) is a vascular condition of the equine hoof resulting in severe lameness with both welfare and economic implications. EL occurs in association with equine metabolic syndrome and equine Cushing's disease. Vascular dysfunction, most commonly due to endothelial dysfunction, is associated with cardiovascular risk in people with metabolic syndrome and Cushing's syndrome. We tested the hypothesis that horses with EL have vascular, specifically endothelial, dysfunction. Healthy horses (n = 6) and horses with EL (n = 6) destined for euthanasia were recruited. We studied vessels from the hooves (laminar artery, laminar vein) and the facial skin (facial skin arteries) by small vessel wire myography. The response to vasoconstrictors phenylephrine (10-9-10-5M) and 5-hydroxytryptamine (5HT; 10-9-10-5M) and the vasodilator acetylcholine (10-9-10-5M) was determined. In comparison with healthy controls, acetylcholine-induced relaxation was dramatically reduced in all intact vessels from horses with EL (% relaxation of healthy laminar arteries 323.5 ± 94.1% v EL 90.8 ± 4.4%, P = 0.01, laminar veins 129.4 ± 14.8% v EL 71.2 ± 4.1%, P = 0.005 and facial skin arteries 182.0 ± 40.7% v EL 91.4 ± 4.5%, P = 0.01). In addition, contractile responses to phenylephrine and 5HT were increased in intact laminar veins from horses with EL compared with healthy horses; these differences were endothelium-independent. Sensitivity to phenylephrine was reduced in intact laminar arteries (P = 0.006) and veins (P = 0.009) from horses with EL. Horses with EL exhibit significant vascular dysfunction in laminar vessels and in facial skin arteries. The systemic nature of the abnormalities suggest this dysfunction is associated with the underlying endocrinopathy and not local changes to the hoof

A new class of glycomimetic drugs to prevent free fatty acid-induced endothelial dysfunction

Background: Carbohydrates play a major role in cell signaling in many biological processes. We have developed a set of glycomimetic drugs that mimic the structure of carbohydrates and represent a novel source of therapeutics for endothelial dysfunction, a key initiating factor in cardiovascular complications. Purpose: Our objective was to determine the protective effects of small molecule glycomimetics against free fatty acid­induced endothelial dysfunction, focusing on nitric oxide (NO) and oxidative stress pathways. Methods: Four glycomimetics were synthesized by the stepwise transformation of 2,5­dihydroxybenzoic acid to a range of 2,5­substituted benzoic acid derivatives, incorporating the key sulfate groups to mimic the interactions of heparan sulfate. Endothelial function was assessed using acetylcholine­induced, endotheliumdependent relaxation in mouse thoracic aortic rings using wire myography. Human umbilical vein endothelial cell (HUVEC) behavior was evaluated in the presence or absence of the free fatty acid, palmitate, with or without glycomimetics (1µM). DAF­2 and H2DCF­DA assays were used to determine nitric oxide (NO) and reactive oxygen species (ROS) production, respectively. Lipid peroxidation colorimetric and antioxidant enzyme activity assays were also carried out. RT­PCR and western blotting were utilized to measure Akt, eNOS, Nrf­2, NQO­1 and HO­1 expression. Results: Ex vivo endothelium­dependent relaxation was significantly improved by the glycomimetics under palmitate­induced oxidative stress. In vitro studies showed that the glycomimetics protected HUVECs against the palmitate­induced oxidative stress and enhanced NO production. We demonstrate that the protective effects of pre­incubation with glycomimetics occurred via upregulation of Akt/eNOS signaling, activation of the Nrf2/ARE pathway, and suppression of ROS­induced lipid peroxidation. Conclusion: We have developed a novel set of small molecule glycomimetics that protect against free fatty acidinduced endothelial dysfunction and thus, represent a new category of therapeutic drugs to target endothelial damage, the first line of defense against cardiovascular disease