Chronic disease IMPACT (chronic disease early detection and improved management in primary care project): An Australian stepped wedge cluster randomised trial.

Background: Interrelated chronic vascular diseases (chronic kidney disease (CKD), type 2 diabetes (T2D) and cardiovascular disease (CVD)) are common with high morbidity and mortality. This study aimed to assess if an electronic-technology-based quality improvement intervention in primary care could improve detection and management of people with and at risk of these diseases. Methods: Stepped-wedge trial with practices randomised to commence intervention in one of five 16-week periods. Intervention included (1) electronic-technology tool extracting data from general practice electronic medical records and generating graphs and lists for audit; (2) education regarding chronic disease and the electronic-technology tool; (3) assistance with quality improvement audit plan development, benchmarking, monitoring and support. De-identified data analysis using R 3.5.1 conducted using Bayesian generalised linear mixed model with practice and time-specific random intercepts. Results: At baseline, eight included practices had 37,946 active patients (attending practice ≥3 times within 2 years) aged ≥18 years. Intervention was associated with increased OR (95% CI) for: kidney health checks (estimated glomerular filtration rate, urine albumin:creatinine ratio (uACR) and blood pressure) in those at risk 1.34 (1.26-1.42); coded diagnosis of CKD 1.18 (1.09-1.27); T2D diagnostic testing (fasting glucose or HbA1c) in those at risk 1.15 (1.08-1.23); uACR in patients with T2D 1.78 (1.56-2.05). Documented eye checks within recommended frequency in patients with T2D decreased 0.85 (0.77-0.96). There were no significant changes in other assessed variables. Conclusions: This electronic-technology-based intervention in primary care has potential to help translate guidelines into practice but requires further refining to achieve widespread improvements across the interrelated chronic vascular diseases.

Using electronic medical record data to assess chronic kidney disease, type 2 diabetes and cardiovascular disease testing, recognition and management as documented in Australian general practice: a cross-sectional analysis.

OBJECTIVES: To evaluate the capacity of general practice (GP) electronic medical record (EMR) data to assess risk factor detection, disease diagnostic testing, diagnosis, monitoring and pharmacotherapy for the interrelated chronic vascular diseases-chronic kidney disease (CKD), type 2 diabetes (T2D) and cardiovascular disease. DESIGN: Cross-sectional analysis of data extracted on a single date for each practice between 12 April 2017 and 18 April 2017 incorporating data from any time on or before data extraction, using baseline data from the Chronic Disease early detection and Improved Management in PrimAry Care ProjecT. Deidentified data were extracted from GP EMRs using the Pen Computer Systems Clinical Audit Tool and descriptive statistics used to describe the study population. SETTING: Eight GPs in Victoria, Australia. PARTICIPANTS: Patients were ≥18 years and attended GP ≥3 times within 24 months. 37 946 patients were included. RESULTS: Risk factor and disease testing/monitoring/treatment were assessed as per Australian guidelines (or US guidelines if none available), with guidelines simplified due to limitations in data availability where required. Risk factor assessment in those requiring it: 30% of patients had body mass index and 46% blood pressure within guideline recommended timeframes. Diagnostic testing in at-risk population: 17% had diagnostic testing as per recommendations for CKD and 37% for T2D. Possible undiagnosed disease: Pathology tests indicating possible disease with no diagnosis already coded were present in 6.7% for CKD, 1.6% for T2D and 0.33% familial hypercholesterolaemia. Overall prevalence: Coded diagnoses were recorded in 3.8% for CKD, 6.6% for T2D, 4.2% for ischaemic heart disease, 1% for heart failure, 1.7% for ischaemic stroke, 0.46% for peripheral vascular disease, 0.06% for familial hypercholesterolaemia and 2% for atrial fibrillation. Pharmaceutical prescriptions: the proportion of patients prescribed guideline-recommended medications ranged from 44% (beta blockers for patients with ischaemic heart disease) to 78% (antiplatelets or anticoagulants for patients with ischaemic stroke). CONCLUSIONS: Using GP EMR data, this study identified recorded diagnoses of chronic vascular diseases generally similar to, or higher than, reported national prevalence. It suggested low levels of extractable documented risk factor assessments, diagnostic testing in those at risk and prescription of guideline-recommended pharmacotherapy for some conditions. These baseline data highlight the utility of GP EMR data for potential use in epidemiological studies and by individual practices to guide targeted quality improvement. It also highlighted some of the challenges of using GP EMR data.

High intraluminal pressure promotes vascular inflammation via caveolin-1.

The aetiology and progression of hypertension involves various endogenous systems, such as the renin angiotensin system, the sympathetic nervous system, and endothelial dysfunction. Recent data suggest that vascular inflammation may also play a key role in the pathogenesis of hypertension. This study sought to determine whether high intraluminal pressure results in vascular inflammation. Leukocyte adhesion was assessed in rat carotid arteries exposed to 1 h of high intraluminal pressure. The effect of intraluminal pressure on signaling mechanisms including reactive oxygen species production (ROS), arginase expression, and NFĸB translocation was monitored. 1 h exposure to high intraluminal pressure (120 mmHg) resulted in increased leukocyte adhesion and inflammatory gene expression in rat carotid arteries. High intraluminal pressure also resulted in a downstream signaling cascade of ROS production, arginase expression, and NFĸB translocation. This process was found to be angiotensin II-independent and mediated by the mechanosensor caveolae, as caveolin-1 (Cav1)-deficient endothelial cells and mice were protected from pressure-induced vascular inflammatory signaling and leukocyte adhesion. Cav1 deficiency also resulted in a reduction in pressure-induced glomerular macrophage infiltration in vivo. These findings demonstrate Cav1 is an important mechanosensor in pressure-induced vascular and renal inflammation.

Nitroxyl (HNO) reduces endothelial and monocyte activation and promotes M2 macrophage polarization.

Nitroxyl anion (HNO) donors are currently being assessed for their therapeutic utility in several cardiovascular disorders including heart failure. Here, we examine their effect on factors that precede atherosclerosis including endothelial cell and monocyte activation, leucocyte adhesion to the endothelium and macrophage polarization. Similar to the NO donor glyceryl trinitrate (GTN), the HNO donors Angeli's salt (AS) and isopropylamine NONOate (IPA/NO) decreased leucocyte adhesion to activated human umbilical vein endothelial cells (HUVECs) and mouse isolated aorta. This reduction in adhesion was accompanied by a reduction in intercellular adhesion molecule-1 (ICAM-1) and the cytokines monocyte chemoattractant protein 1 (MCP-1) and interleukin 6 (IL-6) which was inhibitor of nuclear factor κB (NFκB) α (IκBα)- and subsequently NFκB-dependent. Intriguingly, the effects of AS on leucocyte adhesion, like those on vasodilation, were found to not be susceptible to pharmacological tolerance, unlike those observed with GTN. As well, HNO reduces monocyte activation and promotes polarization of M2 macrophages. Taken together, our data demonstrate that HNO donors can reduce factors that are associated with and which precede atherosclerosis and may thus be useful therapeutically. Furthermore, since the effects of the HNO donors were not subject to tolerance, this confers an additional advantage over NO donors.

Nitroxyl: a vasodilator of human vessels that is not susceptible to tolerance.

Pre-clinical studies have identified nitroxyl (HNO), the reduced congener of nitric oxide (NO•), as a potent vasodilator which is resistant to tolerance development. The present study explores the efficacy of HNO in human blood vessels and describes, for the first time, a vasodilator for humans that is not susceptible to tolerance. Human radial arteries and saphenous veins were obtained from patients undergoing coronary artery graft surgery and mounted in organ baths. Repeated vasodilator responses to the HNO donor Angeli's salt (AS) and NO• donor glyceryl trinitrate (GTN) were determined. AS- and GTN-induced concentration-dependent vasorelaxation of both human radial arteries (AS pEC50: 6.5 ± 0.2; -log M) and saphenous veins (pEC50: 6.7 ± 0.1) with similar potency. In human radial arteries, GTN-induced relaxation was reduced by the NO• scavenger hydroxocobalamin (HXC; P<0.05) but was unaffected by the HNO scavenger L-cysteine. Alternately, AS was unaffected by HXC but was reduced by L-cysteine (5-fold shift, P<0.05). The sGC (soluble guanylate cyclase) inhibitor ODQ abolished responses to both AS and GTN in arteries and veins (P<0.05). Inhibition of voltage-dependent potassium channels (Kv channels) with 4-AP also significantly reduced responses to AS (pEC50: 5.5) and GTN, suggesting that the relaxation to both redox congeners is cGMP- and Kv channel-dependent. Critically, a concentration-dependent development of tolerance to GTN (1 and 10 µM; P<0.05), but not to AS, was observed in both saphenous veins and radial arteries. Like GTN, the HNO donor AS causes vasorelaxation of human blood vessels via activation of a cGMP-dependent pathway. Unlike GTN, however, it does not develop tolerance in human blood vessels.

Endothelial C-type natriuretic peptide maintains vascular homeostasis.

The endothelium plays a fundamental role in maintaining vascular homeostasis by releasing factors that regulate local blood flow, systemic blood pressure, and the reactivity of leukocytes and platelets. Accordingly, endothelial dysfunction underpins many cardiovascular diseases, including hypertension, myocardial infarction, and stroke. Herein, we evaluated mice with endothelial-specific deletion of Nppc, which encodes C-type natriuretic peptide (CNP), and determined that this mediator is essential for multiple aspects of vascular regulation. Specifically, disruption of CNP leads to endothelial dysfunction, hypertension, atherogenesis, and aneurysm. Moreover, we identified natriuretic peptide receptor-C (NPR-C) as the cognate receptor that primarily underlies CNP-dependent vasoprotective functions and developed small-molecule NPR-C agonists to target this pathway. Administration of NPR-C agonists promotes a vasorelaxation of isolated resistance arteries and a reduction in blood pressure in wild-type animals that is diminished in mice lacking NPR-C. This work provides a mechanistic explanation for genome-wide association studies that have linked the NPR-C (Npr3) locus with hypertension by demonstrating the importance of CNP/NPR-C signaling in preserving vascular homoeostasis. Furthermore, these results suggest that the CNP/NPR-C pathway has potential as a disease-modifying therapeutic target for cardiovascular disorders.

Raised soluble P-selectin moderately accelerates atherosclerotic plaque progression.

Soluble P-selectin (sP-selectin), a biomarker of inflammatory related pathologies including cardiovascular and peripheral vascular diseases, also has pro-atherosclerotic effects including the ability to increase leukocyte recruitment and modulate thrombotic responses in vivo. The current study explores its role in progressing atherosclerotic plaque disease. Apoe-/- mice placed on a high fat diet (HFD) were given daily injections of recombinant dimeric murine P-selectin (22.5 µg/kg/day) for 8 or 16 weeks. Saline or sE-selectin injections were used as negative controls. In order to assess the role of sP-selectin on atherothrombosis an experimental plaque remodelling murine model, with sm22α-hDTR Apoe-/- mice on a HFD in conjunction with delivery of diphtheria toxin to induce targeted vascular smooth muscle apoptosis, was used. These mice were similarly given daily injections of sP-selectin for 8 or 16 weeks. While plaque mass and aortic lipid content did not change with sP-selectin treatment in Apoe-/- or SM22α-hDTR Apoe-/- mice on HFD, increased plasma MCP-1 and a higher plaque CD45 content in Apoe-/- HFD mice was observed. As well, a significant shift towards a more unstable plaque phenotype in the SM22α-hDTR Apoe-/- HFD mice, with increased macrophage accumulation and lower collagen content, leading to a lower plaque stability index, was observed. These results demonstrate that chronically raised sP-selectin favours progression of an unstable atherosclerotic plaque phenotype.

Endothelial dysfunction in patients with type 2 diabetes post acute coronary syndrome.

UNLABELLED: This single visit study examined whether endothelial function, in addition to cardiovascular (CV) risk factors and plasma microparticle content, was normalised in 15 patients with type 2 diabetes + acute coronary syndrome (ACS) (6 weeks-6 months post cardiac event) undergoing standard clinical care compared to 16 sex- and age-matched healthy controls. RESULTS: While total and low-density lipoprotein (LDL) cholesterol levels were well controlled in the patients with type 2 diabetes + ACS, residual CV risk profiles such as increased body mass index (BMI), systolic blood pressure, glucose levels and triglycerides and lower high-density lipoprotein (HDL) levels were still apparent. Endothelium-dependent responses to acetylcholine (ACh) were significantly lower in type 2 diabetes + ACS patients compared to controls. Correspondingly, the reactive hyperaemic index (RHI) was lower in the patient cohort. Endothelial microparticle (EMP) levels (CD31(+), CD41(-)) were 40% lower in the patient cohort. Simultaneous analysis of platelet microparticle (PMP) levels (CD41(+)) showed no difference between cohorts. CONCLUSIONS: Patients with type 2 diabetes suffering from recent ACS exhibit residual CV risk factors despite being on standard clinical care. In addition, these patients continue to present with endothelial dysfunction despite having lower levels of EMPs.

C-type natriuretic peptide (CNP): cardiovascular roles and potential as a therapeutic target.

Natriuretic peptides play a fundamental role in cardiovascular homeostasis by modulation of fluid and electrolyte balance and vascular tone. C-type natriuretic peptide (CNP) represents the paracrine element of the natriuretic peptide axis which complements the endocrine actions of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). CNP is produced by the endothelium and the heart and appears to play a prominent role in vascular and cardiac function, both physiologically and pathologically. This provides a rationale for the therapeutic potential of pharmacological interventions targeted to CNP signalling. This article provides an overview of the biology and pharmacology of CNP, with emphasis on the cardiovascular system, and discusses pathologies in which drugs designed to manipulate CNP signalling maybe of clinical benefit.

Omega-conotoxin GVIA mimetics based on an anthranilamide core: effect of variation in ammonium side chain lengths and incorporation of fluorine.

A number of omega-conotoxin GVIA mimetics based on an anthranilamide core were prepared and tested for their affinity for rat brain Ca(v)2.2 channels. Features such as the presence of hydroxyl and fluoro substituents on the tyrosine side chain mimic, the length of the chains on the lysine/arginine side chain mimics and the use of diguanidino and diamino substituents rather than mono-guanidine/mono-amine substitution were examined. The diguanidinylated compounds proved to be the most active and deletion of the hydroxyl substituent had a limited influence on activity. The SAR associated with variation in the lysine/arginine side chain mimics was not strong. The introduction of a fluoro substituent into the tyrosine mimic produced the most active compound prepared in this study (2g), with an EC(50) at rat brain Ca(v)2.2 channels of 6 microM.

Low molecular weight non-peptide mimics of omega-conotoxin GVIA.

We report the synthesis and biological activity of a low molecular weight non-peptidic mimic of the analgesic peptide omega-conotoxin GVIA. The molecular weight of this compound presents a reduction by 193g/mol compared to a previously reported lead. This compound exhibits an EC(50) of 5.8microM and is accessible in only six synthetic steps compared to the original lead (13 steps). We also report several improvements to the original synthetic route.

Conopressin-T from Conus tulipa reveals an antagonist switch in vasopressin-like peptides.

We report the discovery of conopressin-T, a novel bioactive peptide isolated from Conus tulipa venom. Conopressin-T belongs to the vasopressin-like peptide family and displays high sequence homology to the mammalian hormone oxytocin (OT) and to vasotocin, the endogenous vasopressin analogue found in teleost fish, the cone snail's prey. Conopressin-T was found to act as a selective antagonist at the human V 1a receptor. All peptides in this family contain two conserved amino acids within the exocyclic tripeptide (Pro7 and Gly9), which are replaced with Leu7 and Val9 in conopressin-T. Whereas conopressin-T binds only to OT and V 1a receptors, an L7P analogue had increased affinity for the V 1a receptor and weak V2 receptor binding. Surprisingly, replacing Gly9 with Val9 in OT and vasopressin revealed that this position can function as an agonist/antagonist switch at the V 1a receptor. NMR structures of both conopressin-T and L7P analogue revealed a marked difference in the orientation of the exocyclic tripeptide that may serve as templates for the design of novel ligands with enhanced affinity for the V 1a receptor.

Isolation and characterization of rufoxin, a novel protein exhibiting neurotoxicity from venom of the psammophiine, Rhamphiophis oxyrhynchus (Rufous beaked snake).

Colubrid snake venoms potentially represent a vast source of novel biological actives and structural motifs owing to their diverse phylogeny. The present study describes the identification of rufoxin, a neurotoxin from the venom of Rhamphiophis oxyrhynchus (Rufous beaked snake) which is a member of the African colubrid lineage, the psammophiines. Rufoxin (1 microM) displayed reversible post-synaptic neurotoxic activity as evidenced by significant inhibition of indirect twitches and responses to exogenous nicotinic agonists in the chick biventer cervicis nerve-muscle preparation. Rufoxin (0.1-1.0 microM) also caused a rightward parallel shift of cumulative concentration-response curves to carbachol (CCh; 0.6-80 microM) without a significant depression of the maximum response, suggestive of classical competitive antagonism at the skeletal muscle nicotinic receptor. Rufoxin lacks NH(2)-terminal sequence homology to previously identified snake venom toxins. This work indicates a wider distribution of neurotoxins across the advanced snake superfamily than previously described.

Isolation and characterisation of conomap-Vt, a D-amino acid containing excitatory peptide from the venom of a vermivorous cone snail.

Cone snail venom is a rich source of bioactives, in particular small disulfide rich peptides that disrupt synaptic transmission. Here, we report the discovery of conomap-Vt (Conp-Vt), an unusual linear tetradecapeptide isolated from Conus vitulinus venom. The sequence displays no homology to known conopeptides, but displays significant homology to peptides of the MATP (myoactive tetradecapeptide) family, which are important endogenous neuromodulators in molluscs, annelids and insects. Conp-Vt showed potent excitatory activity in several snail isolated tissue preparations. Similar to ACh, repeated doses of Conp-Vt were tachyphylactic. Since nicotinic and muscarinic antagonists failed to block its effect and Conp-Vt desensitised tissue remained responsive to ACh, it appears that Conp-Vt contractions were non-cholinergic in origin. Finally, biochemical studies revealed that Conp-Vt is the first member of the MATP family with a d-amino acid. Interestingly, the isomerization of L-Phe to D-Phe enhanced biological activity, suggesting that this post-translational modified conopeptide may have evolved for prey capture.

Pharmacological characterisation of a neurotoxin from the venom of Boiga dendrophila (mangrove catsnake).

In this study, we have pharmacologically characterised boigatoxin-A, a three finger toxin isolated from the venom of the colubrid, Boiga dendrophila (Mangrove catsnake). In the chick biventer cervicis nerve-muscle preparation boigatoxin-A (1 microM) displayed poorly reversible postsynaptic blockade as evidenced by the inhibition of indirect (0.1 Hz, 0.2 ms, supramaximal V) twitches and responses to exogenous acetylcholine (1 mM) and carbachol (20 microM). Boigatoxin-A (0.3-0.5 microM) caused a concentration-dependent depression of the maximum response of cumulative concentration response curves to CCh (0.6-80 microM). Boigatoxin-A (1 microM) induced readily reversible inhibition of electrically evoked (0.2 Hz, 0.3 ms, 70-100 V) twitches of the prostatic segment of the rat vas deferens. This inhibition was not significantly attenuated by 8-phenyltheophylline (20 microM) or idazoxan (1 microM). Boigatoxin-A (1 microM) did not affect alpha,beta-mATP (10 microM) or noradrenaline (25 microM) responses in unstimulated epididymal segments of the rat vas deferens. Our data suggests that this toxin has weak postsynaptic neurotoxicity in skeletal muscle and also prejunctional neurotoxic activity in the smooth muscle of the rat vas deferens to inhibit the release of neurotransmitter(s), but not via prejunctional purinergic or adrenergic receptors. This is the first report of such activity for a toxin isolated from snake venom and reinforces the largely untapped potential of colubrid venoms.

A biochemical and pharmacological examination of Rhamphiophis oxyrhynchus (Rufous beaked snake) venom.

The venom of R. oxyrhynchus, a member of the psammophiine subfamily of the colubrid assemblage, was examined for biological activity using biochemical and pharmacological techniques. Venom displayed a high protein content, a complex electrophorectic profile and PLA2 activity but no detectable proteolytic or haematological activities. In the chick biventer cervicis nerve muscle preparation, venom (1-10 microg/ml) displayed postsynaptic neurotoxic activity as evidenced by inhibition of indirect (0.1 Hz, 0.2 ms, supramaximal V) twitches and responses to exogenous acetylcholine (1 mM) and carbachol (20 microM). This inhibitory effect was poorly reversible by washing. Venom (30-50 microg/ml) caused a rapid and readily reversible inhibition of direct (0.1 Hz, 2 ms, supramaximal V) twitches of the chick biventer cervicis nerve muscle preparation without morphological changes to the muscle fibers. Venom (30-100 microg/ml) inhibited electrically-evoked (0.2 Hz, 0.3 ms, 70-100 V) twitches of the prostatic segment of the rat vas deferens. This inhibitory effect was not significantly attenuated by 8-phenyltheophylline (8-PT; 20 microM), idazoxan (1 microM), a combination of ranitidine (0.2 microM) and thioperamide (10 microM) or capsazepine (10 microM). Venom (5 mg/kg) induced hypotension with subsequent cardiovascular collapse in the anaesthetised rat. The cardiovascular collapse was prevented by artificial respiration of the animals prior to venom administration. The biological activities demonstrated by R. oxyrhynchus venom may aid in prey envenomation strategies such as prey immobilisation. This study provides further evidence that colubrid venoms are comprised of multiple components which can display a variety of actions, some of which may be novel, therefore reinforcing the largely untapped potential of colubrid venoms.

In vitro neuromuscular activity of 'colubrid' venoms: clinical and evolutionary implications.

In this study, venoms from species in the Colubrinae, Homalopsinae, Natricinae, Pseudoxyrhophiinae and Psammophiinae snake families were assayed for activity in the chick biventer cervicis skeletal nerve muscle preparation. Boiga dendrophila, Boiga cynodon, Boiga dendrophila gemincincta, Boiga drapiezii, Boiga irregularis, Boiga nigriceps and Telescopus dhara venoms (10 microg/ml) displayed postsynaptic neuromuscular activity as evidenced by inhibition of indirect (0.1 Hz, 0.2 ms, supramaximal V) twitches. Neostigmine (5 microM) reversed the inhibition caused by B. cynodon venom (10 microg/ml) while the inhibitory effects of Psammophis mossambicus venom (10 microg/ml) spontaneously reversed, indicating a reversible mode of action for both venoms. Trimorphodon biscutatus (10 microg/ml) displayed irreversible presynaptic neurotoxic activity. Detectable levels of phospholipase A2 activity were found only in T. biscutatus, T. dhara and P. mossambicus venoms. The results demonstrate a hitherto unsuspected diversity of pharmacological actions in all lineages which may have implications ranging from clinical management of envenomings to venom evolution.

Isolation of a neurotoxin (alpha-colubritoxin) from a nonvenomous colubrid: evidence for early origin of venom in snakes.

The evolution of venom in advanced snakes has been a focus of long-standing interest. Here we provide the first complete amino acid sequence of a colubrid toxin, which we have called alpha-colubritoxin, isolated from the Asian ratsnake Coelognathus radiatus (formerly known as Elaphe radiata), an archetypal nonvenomous snake as sold in pet stores. This potent postsynaptic neurotoxin displays readily reversible, competitive antagonism at the nicotinic receptor. The toxin is homologous with, and phylogenetically rooted within, the three-finger toxins, previously thought unique to elapids, suggesting that this toxin family was recruited into the chemical arsenal of advanced snakes early in their evolutionary history. LC-MS analysis of venoms from most other advanced snake lineages revealed the widespread presence of components of the same molecular weight class, suggesting the ubiquity of three-finger toxins across advanced snakes, with the exclusion of Viperidae. These results support the role of venom as a key evolutionary innovation in the early diversification of advanced snakes and provide evidence that forces a fundamental rethink of the very concept of nonvenomous snake.