The cyclic GMP modulators YC-1 and zaprinast reduce vessel remodeling through anti-proliferative and pro-apoptotic effects

Guanosine-specific cyclic nucleotide signaling is suggested to serve protective actions in the vasculature; however, the influence of selective pharmacologic modulation of cyclic guanosine monophosphate (GMP)-synthesizing soluble guanylate cyclase (sGC) or cyclic GMP-degrading phosphodiesterase (PDE) on vessel remodeling has not been thoroughly examined. In this study, rat carotid artery balloon injury was performed and the growth-modulating effects of the sGC stimulator YC-1 or the cGMP-dependent PDE-V inhibitor zaprinast were examined. YC-1 or zaprinast elevated vessel cyclic GMP content, reduced medial wall and neointimal cell proliferation, stimulated medial and neointimal cellular apoptosis, and markedly attenuated neointimal remodeling in comparable fashion. Interestingly, sGC inhibition by ODQ failed to noticeably alter neointimal growth, and concomitant zaprinast with YC-1 did not modify any parameter compared to individual treatments. These results provide novel in vivo evidence that YC-1 and zaprinast inhibit injury-induced vascular remodeling through anti-mitogenic and pro-apoptotic actions and may offer promising therapeutic approaches against vasoproliferative disorders. Originally published J Cardiovasc Pharmacol Ther, Vol. 14, No. 2, June 200

The Dopamine D3 Receptor Knockout Mouse Mimics Aging-Related Changes in Autonomic Function and Cardiac Fibrosis

Blood pressure increases with age, and dysfunction of the dopamine D3 receptor has been implicated in the pathogenesis of hypertension. To evaluate the role of the D3 receptor in aging-related hypertension, we assessed cardiac structure and function in differently aged (2 mo, 1 yr, 2 yr) wild type (WT) and young (2 mo) D3 receptor knockout mice (D3KO). In WT, systolic and diastolic blood pressures and rate-pressure product (RPP) significantly increased with age, while heart rate significantly decreased. Blood pressure values, heart rate and RPP of young D3KO were significantly elevated over age-matched WT, but similar to those of the 2 yr old WT. Echocardiography revealed that the functional measurements of ejection fraction and fractional shortening decreased significantly with age in WT and that they were significantly smaller in D3KO compared to young WT. Despite this functional change however, cardiac morphology remained similar between the age-matched WT and D3KO. Additional morphometric analyses confirmed an aging-related increase in left ventricle (LV) and myocyte cross-sectional areas in WT, but found no difference between age-matched young WT and D3KO. In contrast, interstitial fibrosis, which increased with age in WT, was significantly elevated in the D3KO over age-matched WT, and similar to 2 yr old WT. Western analyses of myocardial homogenates revealed significantly increased levels of pro- and mature collagen type I in young D3KO. Column zymography revealed that activities of myocardial MMP-2 and MMP-9 increased with age in WTs, but in D3KO, only MMP-9 activity was significantly increased over age-matched WTs. Our data provide evidence that the dopamine D3 receptor has a critical role in the emergence of aging-related cardiac fibrosis, remodeling, and dysfunction

Assessment of HIF-1α expression and release following endothelial injury in-vitro and in-vivo

Background: Endothelial injury is an early and enduring feature of cardiovascular disease. Inflammation and hypoxia may be responsible for this, and are often associated with the up-regulation of several transcriptional factors that include Hypoxia Inducible Factor-1 (HIF-1). Although it has been reported that HIF-1α is detectable in plasma, it is known to be unstable. Our aim was to optimize an assay for HIF-1α to be applied to in vitro and in vivo applications, and to use this assay to assess the release kinetics of HIF-1 following endothelial injury. Methods: An ELISA for the measurement of HIF in cell-culture medium and plasma was optimized, and the assay used to determine the best conditions for sample collection and storage. The results of the ELISA were validated using Western blotting and immunohistochemistry (IHC). In vitro, a standardized injury was produced in a monolayer of rat aortic endothelial cells (RAECs) and intracellular HIF-1α was measured at intervals over 24 hours. In vivo, a rat angioplasty model was used. The right carotid artery was injured using a 2F Fogarty balloon catheter. HIF-1α was measured in the plasma and in the arterial tissue (0, 1, 2, 3 and 5 days post injury). Results: The HIF-1α ELISA had a limit of detection of 2.7 pg/ mL and was linear up to 1000 pg/ mL. Between and within-assay coefficient of variation values were less than 15%. HIF-1α was unstable in cell lysates and plasma, and it was necessary to add a protease inhibitor immediately after collection, and to store samples at -800C prior to analysis. The dynamics of HIF-1α release were different for the in vitro and in vivo models. In vitro, HIF-1α reached maximum concentrations approximately 2h post injury, whereas peak values in plasma and tissues occurred approximately 2 days post injury, in the balloon injury model. Conclusion: HIF-1α can be measured in plasma, but this requires careful sample collection and storage. The carotid artery balloon injury model is associated with the transient release of HIF-1α into the circulation that probably reflects the hypoxia induced in the artery wall

Impaired flow-induced arterial remodeling in DOCA-salt hypertensive rats

Arteries from young healthy animals respond to chronic changes in blood flow and blood pressure by structural remodeling. We tested whether the ability to respond to decreased (-90%) or increased (+100%) blood flow is impaired during the development of deoxycorticosterone acetate (DOCA)-salt hypertension in rats, a model for an upregulated endothelin-1 system. Mesenteric small arteries (MrA) were exposed to low blood flow (LF) or high blood flow (HF) for 4 or 7 weeks. The bioavailability of vasoactive peptides was modified by chronic treatment of the rats with the dual neutral endopeptidase (NEP)/endothelin-converting enzyme (ECE) inhibitor SOL1. After 3 or 6 weeks of hypertension, the MrA showed hypertrophic arterial remodeling (3 weeks: media cross-sectional area (mCSA): 10 +/- 1 x 10(3) to 17 +/- 2 x 10(3) mu m(2); 6 weeks: 13 +/- 2 x 10(3) to 24 +/- 3 x 10(3) mu m(2)). After 3, but not 6, weeks of hypertension, the arterial diameter was increased (empty set: 385 +/- 13 to 463 +/- 14 mu m). SOL1 reduced hypertrophy after 3 weeks of hypertension (mCSA: 6 x 10(3) +/- 1 x 10(3) mu m(2)). The diameter of the HF arteries of normotensive rats increased (empty set: 463 +/- 22 mu m) but no expansion occurred in the HF arteries of hypertensive rats (empty set: 471 +/- 16 mu m). MrA from SOL1-treated hypertensive rats did show a significant diameter increase (empty set: 419 +/- 13 to 475 +/- 16 mu m). Arteries exposed to LF showed inward remodeling in normotensive and hypertensive rats (mean empty set between 235 and 290 mu m), and infiltration of monocyte/ macrophages. SOL1 treatment did not affect the arterial diameter of LF arteries but reduced the infiltration of monocyte/ macrophages. We show for the first time that flow-induced remodeling is impaired during the development of DOCA-salt hypertension and that this can be prevented by chronic NEP/ECE inhibition. Hypertension Research (2012) 35, 1093-1101; doi:10.1038/hr.2012.94; published online 12 July 201

Mesenteric Resistance Arteries in Type 2 Diabetic db/db Mice Undergo Outward Remodeling

Resistance vessel remodeling is controlled by myriad of hemodynamic and neurohormonal factors. This study characterized structural and molecular remodeling in mesenteric resistance arteries (MRAs) in diabetic (db/db) and control (Db/db) mice.Structural properties were assessed in isolated MRAs from 12 and 16 wk-old db/db and Db/db mice by pressure myography. Matrix regulatory proteins were measured by Western blot analysis. Mean arterial pressure and superior mesenteric blood flow were measured in 12 wk-old mice by telemetry and a Doppler flow nanoprobe, respectively.Blood pressure was similar between groups. Lumen diameter and medial cross-sectional area were significantly increased in 16 wk-old db/db MRA compared to control, indicating outward hypertrophic remodeling. Moreover, wall stress and cross-sectional compliance were significantly larger in diabetic arteries. These remodeling indices were associated with increased expression of matrix regulatory proteins matrix metalloproteinase (MMP)-9, MMP-12, tissue inhibitors of matrix metalloproteinase (TIMP)-1, TIMP-2, and plasminogen activator inhibitor-1 (PAI-1) in db/db arteries. Finally, superior mesenteric artery blood flow was increased by 46% in 12 wk-old db/db mice, a finding that preceded mesenteric resistance artery remodeling.These data suggest that flow-induced hemodynamic changes may supersede the local neurohormonal and metabolic milieu to culminate in hypertrophic outward remodeling of type 2 DM mesenteric resistance arteries

A method of inducing global cerebral ischemia.

The four-vessel occlusion (4-VO) method of global forebrain cerebral ischemia mimics the human clinical condition of cardiac arrest. It results in selective neuronal damage and is a useful experimental system to dissect underlying mechanisms behind ischemic phenomena such as the differential susceptibility of CA1 compared to the CA3 region of the hippocampus. It also provides a "proof-of-principle" system for testing out potential agents for neuroprotection