mmHg In Vehicle Research Articles

NCX 470 Reduces Intraocular Pressure More Effectively Than Lumigan in Dogs and Enhances Conventional and Uveoscleral Outflow in Non-Human Primates and Human Trabecular Meshwork/Schlemm's Canal Constructs.

Purpose: To determine NCX 470 (0.1%) and Lumigan® (bimatoprost ophthalmic solution, 0.01%-LUM) intraocular pressure (IOP)-lowering activity after single or repeated (5 days) dosing along with changes in aqueous humor (AH) dynamics. Methods: Ocular hypotensive activity of NCX 470 and LUM was compared with vehicle (VEH) in Beagle dogs using TonoVet®. Non-human primates (NHP) and bioengineered three-dimensional (3D) human Trabecular Meshwork/Schlemm's Canal (HTM/HSC™) constructs exposed to transforming growth factor-β2 (TGFβ2) were used to monitor NCX 470 and LUM-induced changes in AH dynamics. Results: NCX 470 (30 μL/eye) showed greater IOP reduction compared with LUM (30 μL/eye) following single AM dosing [maximum change from baseline (CFBmax) = -1.39 ± 0.52, -6.33 ± 0.73, and -3.89 ± 0.66 mmHg (mean ± standard error of the mean) for VEH, NCX 470, and LUM, respectively]. Likewise, repeated 5 days daily dosing of NCX 470 resulted in lower IOP than LUM across the duration of the study (average IOP decrease across tests was -0.45 ± 0.22, -6.06 ± 0.15, and -3.60 ± 0.22 mmHg for VEH, NCX 470, and LUM, respectively). NCX 470 increased outflow facility (Cfl) in vivo in NHP (CflVEH = 0.37 ± 0.09 μL/min/mmHg and CflNCX470 = 0.64 ± 0.17 μL/min/mmHg) as well as in vitro (CHTM/HSC) in HTM/HSC constructs (CHTM/HSC_VEH = 0.47 ± 0.02 μL/min/mm2/mmHg and CHTM/HSC_NCX470 = 0.76 ± 0.03 μL/min/mm2/mmHg). In addition, NCX 470 increased uveoscleral outflow (FuVEH = 0.62 ± 0.26 μL/min and FuNCX470 = 1.53 ± 0.39 μL/min with episcleral venous pressure of 15 mmHg) leaving unaltered aqueous flow (AHFVEH = 2.03 ± 0.22 μL/min and AHFNCX470 = 1.93 ± 0.31 μL/min) in NHP. Conclusions: NCX 470 elicits greater IOP reduction than LUM following single or repeated dosing. Data in NHP and 3D-HTM/HSC constructs suggest that changes in Cfl and Fu account for the robust IOP-lowering effect of NCX 470.

Characterization of Systemic and Regional Hemodynamics and Vascular Dysfunction in Mice with Fecal Induced Peritonitis.

Sepsis is associated with circulatory dysfunction contributing to disturbed blood flow and organ injury. Decreased organ perfusion in sepsis is attributed, in part, to the loss of vasoregulatory mechanisms. Identifying which vascular beds are most susceptible to dysfunction is important for monitoring the recovery of organ function and guiding interventions. This study aimed to investigate the development of vascular dysfunction as sepsis progressed to septic shock. Anesthetized C57Bl/6 mice were instrumented with a fiberoptic pressure sensor in the carotid artery for blood pressure measurements. In subgroups of mice, regional blood flow measurements were taken by positioning a perivascular flow probe around either the left carotid, left renal, or superior mesenteric arteries. Hemodynamic parameters and their responsiveness to bolus doses of vasoactive drugs were recorded prior to and continuously after injection of fecal slurry (1.3 mg/g body weight) for 4 h. Fecal slurry-induced peritonitis reduced mean arterial pressure (62.7 ± 2.4 mmHg vs. 37.5 ± 3.2 mmHg in vehicle and septic mice, respectively), impaired cardiac function, and eventually reduced organ blood flow (71.9%, 66.8%, and 65.1% in the superior mesenteric, renal, and carotid arteries, respectively). The mesenteric vasculature exhibited dysregulation before the renal and carotid arteries, and this underlying dysfunction preceded the blood pressure decline and impaired organ blood flow.

Netarsudil Improves Trabecular Outflow Facility in Patients with Primary Open Angle Glaucoma or Ocular Hypertension: A Phase 2 Study

Intraocular pressure (IOP) reduction is key to controlling primary open angle glaucoma (POAG). Pharmacotherapies for POAG or ocular hypertension (OHT) commonly lower IOP by increasing uveoscleral outflow or decreasing aqueous humor production. Netarsudil (Rhopressa), a Rho kinase inhibitor, reduces IOP by improving trabecular outflow facility, which is reduced in POAG. We investigated the effects of netarsudil on aqueous humor dynamics in patients with POAG or OHT. Double-masked, randomized, vehicle-controlled, Phase 2 trial. Netarsudil 0.02% was instilled in 1 eye and vehicle into the contralateral eye of 20 patients once daily in the morning for 7 days. The primary endpoint was change in mean diurnal outflow facility on day 8 versus that on day 1 (baseline). Outflow facility was measured by using Schiøtz tonography, IOP by pneumotonometry, and episcleral venous pressure (EVP) by automated venomanometry. Eighteen patients (90%) completed the study. Mean diurnal outflow facility increased 0.039 versus 0.007 µL/min/mm Hg from baseline in the netarsudil- and the vehicle-treated groups, respectively (P < .001 vs. baseline for netarsudil), a treatment difference of 0.03 µL/min/mm Hg (P ≤ .001). Mean diurnal IOP change from baseline at day 8 was -4.52 mm Hg for netarsudil versus -0.98 mm Hg for vehicle, a treatment difference of -3.54 mm Hg (P < .0001). Mean diurnal EVP change from baseline was -0.79 mm Hg in the netarsudil-treated group versus 0.10 mm Hg for vehicle, a treatment difference of -0.89 mm Hg (P < .001). All patients reporting an adverse event reported conjunctival hyperemia of mild or moderate severity. Netarsudil acts on the conventional outflow pathway, both proximal and distal, to significantly reduce IOP in POAG and OHT by improving trabecular outflow facility and decreasing EVP.

CCL2 mediates early renal leukocyte infiltration during salt-sensitive hypertension.

Studies examining mechanisms of Dahl salt-sensitive (SS) hypertension have implicated the infiltration of leukocytes in the kidneys, which contribute to renal disease and elevated blood pressure. However, the signaling pathways by which leukocytes traffic to the kidneys remain poorly understood. The present study nominated a signaling pathway by analyzing a kidney RNA sequencing data set from SS rats fed either a low-salt (0.4% NaCl) diet or a high-salt (4.0% NaCl) diet. From this analysis, chemokine (C-C motif) ligand 2 (CCL2) and chemokine (C-C motif) receptor 2 (CCR2) were nominated as a potential pathway modifying renal leukocyte infiltration and contributing to SS hypertension. The functional role of the CCL2/CCR2 pathway was tested by daily administration of CCR2 antagonist (RS-102895 at 5 mg·kg-1·day-1 in DMSO) or DMSO vehicle for 3 or 21 days by intraperitoneal injections during the high salt challenge. Blood pressure, renal leukocyte infiltration, and renal damage were evaluated. The results demonstrated that RS-102895 treatment ameliorated renal damage (urinary albumin excretion; 43.4 ± 5.1 vs. 114.7 ± 15.2 mg/day in vehicle, P < 0.001) and hypertension (144.3 ± 2.2 vs. 158.9 ± 4.8 mmHg in vehicle, P < 0.001) after 21 days of high-salt diet. It was determined that renal leukocyte infiltration was blunted by day 3 of the high-salt diet (1.4 ± 0.1 vs. 1.9 ± 0.2 in vehicle × 106 CD45+ cells/kidney, P = 0.034). An in vitro chemotaxis assay validated the effect of RS-102895 on leukocyte chemotaxis toward CCL2. The results suggest that increased CCL2 in SS kidneys is important in the early recruitment of leukocytes, and blockade of this recruitment by administering RS-102895 subsequently blunted the renal damage and hypertension.

Abstract P149: VEGF Inhibitor-Induced Hypertension and Renal Injury: Prevention by Aspirin?

Vascular endothelial growth factor inhibitor (VEGFi)-induced hypertension and renal injury in cancer patients mimic the clinical phenotype of preeclampsia. ET-1 has been implicated to be involved. Since aspirin can prevent the onset of preeclampsia, we hypothesized that co-treatment of the VEGFi sunitinib (SU) with aspirin might be beneficial and may improve ET-1 status. Male WKY rats were treated with vehicle, 14 mg/kg/day SU, or SU + low or high dose aspirin (5 or 100 mg/kg/day) for 8 days. Mean arterial pressure (MAP) was measured via radiotelemetry. On day 8, 24h urine was collected to determine albuminuria and prostanoid (PGF2α and TxB2) levels. Plasma endothelin-1 (ET-1) was measured via ELISA. Endothelial dysfunction was assessed in iliac vessels and reactive oxygen species (ROS) was measured via aortic superoxide anion production. Renal cyclooxygenase (COX)-1, COX-2 and ET-1 mRNA expression were determined via PCR. SU induced a rapid and sustained increase in MAP (24±2 versus 1±1 mmHg in vehicle on day 6; P&lt;0.001), which was blunted by both low and high aspirin doses (18±3 and 13±4 mmHg respectively on day 6; P&lt;0.05 versus SU). Plasma ET-1 was increased by SU (2.2±0.3 versus 1.4±0.1 pg/ml in vehicle; P&lt;0.05) and this was not affected by aspirin. SU increased albuminuria (1.2±0.2 versus 0.3±0.1 mg/24h in vehicle; P&lt;0.05) which was prevented by high, but not low dose aspirin (0.5±0.1 and 1.3±0.1 mg/24h, respectively), suggesting that this effect is COX-2 dependent. Although renal mRNA expression levels were unchanged, renal COX-2 and ET-1 mRNA expression correlated positively, suggesting that ET-1 upregulates COX-2. SU increased ROS production 2.5-fold (P&lt;0.05 versus vehicle) and this was prevented by both aspirin doses. SU did not induce endothelial dysfunction, nor did it alter prostanoid levels. However, both aspirin doses reduced TxB2 (P&lt;0.05 versus vehicle). In conclusion, high rather than low dose aspirin was more efficacious for the prevention of VEGFi-induced hypertension and renal injury, suggesting that its beneficial effects largely involve COX-2 inhibition. Aspirin may be a novel intervention to allow cancer patients to gain the full benefit of VEGFi without deleterious cardiovascular and renal side effects.

Abstract 105: Therapeutic Suppression of Mtorc2 Signaling Reduces Salt-induced Hypertension and Kidney Injury in SS Rats

The present study explored the protective effect of mTORC2 inhibition in salt-induced hypertension and kidney injury. We have previously reported that enhanced blood pressure salt-sensitivity with renal chronic interstitial infusion of H 2 O 2 (347 nmol/kg/min) in normotensive Sprague Dawley (SD) rats. In the present study, in vivo experiment was performed in which H 2 O 2 (347 nmol/Kg/min) was chronically infused for 3 days into renal interstitium of unilaterally nephrectomized SD rats. A significant increase of mTORC2 activity (pAKT/AKT) was observed in the renal cortex of SD rats infused with H 2 O 2 compared to saline infused rats. We have recently shown that excess production of H 2 O 2 in the SS rat kidneys is the hallmark of salt-induced hypertension. We hypothesized that mTORC2 in the kidney contributes to the development of salt-induced hypertension in SS rats. Rats were treated with PP242 which is an ATP-competitor inhibitor which inhibits the activities of both mTORC1 and mTORC2 whereas rapamycin specifically inhibits mTORC1. PP242 was administrated daily (i.p., 15 mg/Kg/day) for 21 days to SS rats fed a 4.0% NaCl diet. Remarkably, salt-induced hypertension was significantly reduced in SS rats which averaged 119 ± 2 mmHg in PP242 treated rats (n=7) compared to 168 ± 3 mmHg in vehicle treated rats (n=7). Albuminuria was greatly reduced with urine albumin excretion (mg/day) averaging 32.8 ± 3 in PP242 treated rats compared to 256 ± 37 in vehicle treated rats. PP242 treatment notably resulted in reduced infiltration of T lymphocytes in the kidneys of SS rats fed a 4.0% NaCl diet. CD3 + cells/mm 2 averaging 157.0 ± 40.0 compared to 36.0 ± 11.0 in the renal cortex and 218.0 ± 24.0 compared to 24.0 ± 9.0 in the outer medulla in PP242 versus vehicle treated rats. These data show that mTORC2 is required for the intiation of salt-induced hypertension and therapeutic suppression of this pathway virtually abolished salt-sensitivity blood pressure and kidney injury.

Acute and chronic systemic CB1 cannabinoid receptor blockade improves blood pressure regulation and metabolic profile in hypertensive (mRen2)27 rats.

We investigated acute and chronic effects of CB1 cannabinoid receptor blockade in renin‐angiotensin system‐dependent hypertension using rimonabant (SR141716A), an orally active antagonist with central and peripheral actions. In transgenic (mRen2)27 rats, a model of angiotensin II‐dependent hypertension with increased body mass and insulin resistance, acute systemic blockade of CB1 receptors significantly reduced blood pressure within 90 min but had no effect in Sprague‐Dawley rats. No changes in metabolic hormones occurred with the acute treatment. During chronic CB1 receptor blockade, (mRen2)27 rats received daily oral administration of SR141716A (10 mg/kg/day) for 28 days. Systolic blood pressure was significantly reduced within 24 h, and at Day 21 of treatment values were 173 mmHg in vehicle versus 149 mmHg in drug‐treated rats (P < 0.01). This accompanied lower cumulative weight gain (22 vs. 42 g vehicle; P < 0.001), fat mass (2.0 vs. 2.9% of body weight; P < 0.05), and serum leptin (2.8 vs. 6.0 ng/mL; P < 0.05) and insulin (1.0 vs. 1.9 ng/mL; P < 0.01), following an initial transient decrease in food consumption. Conscious hemodynamic recordings indicate twofold increases occurred in spontaneous baroreflex sensitivity (P < 0.05) and heart rate variability (P < 0.01), measures of cardiac vagal tone. The beneficial actions of CB1 receptor blockade in (mRen2)27 rats support the interpretation that an upregulated endocannabinoid system contributes to hypertension and impaired autonomic function in this angiotensin II‐dependent model. We conclude that systemic CB1 receptor blockade may be an effective therapy for angiotensin II‐dependent hypertension and associated metabolic syndrome.

Abstract 461: Macrophage 12/15-lipoxygenase Controls the Development of Experimental Hypertension through PPAR?

Targeted disruption of the 12/15-lipoxygenase (Alox15) gene in mice prevents the development of angiotensin II-, DOCA-salt-, and L-NAME-induced experimental hypertension. Macrophages are the primary source of Alox15. Using Alox15-/- and wild type (WT) mice, we previously demonstrated that macrophage Alox15 is essential for experimental hypertension; however, the underlying mechanism is not known. Since Alox15-derived metabolites of arachidonic and linoleic acids are potent PPARγ agonists, we hypothesized that activation of macrophage PPARγ is the key step in Alox15 mediation of L-NAME-induced hypertension. Thioglycollate (TG) injection, used to elicit peritoneal macrophages, upregulated PPARγ protein expression and expression of its target gene CD36 in peritoneal macrophages, but not in aorta or kidney, of both WT (6.8±0.3 and 9.3±0.5 fold increase for PPARγ and CD36, respectively) and Alox15-/- mice (4.7±0.4 and 11.1±0.6 fold increase for PPARγ and CD36, respectively). Thus, TG bypassed Alox15 to activate PPARγ selectively in macrophages. Upregulation of PPARγ by TG restored the sensitivity to L-NAME hypertension (100 mg/kg/day L-NAME for 7 days) in Alox15-/- mice (129.4±7.3 mmHg for TG vs. 94.8±5.6 mmHg for non-injected controls, p&lt;0.005). A similar hypertensive effect to L-NAME was observed with the adoptive transfer of TG-elicited Alox15-/- macrophages into Alox15-/- recipient mice (122.4±5.5 mmHg for macrophage vs. 96.2±6.2 mmHg for vehicle, p&lt;0.05). The role of PPARγ was further specified by using the selective PPARγ antagonist, GW9662. When WT mice were treated with GW9662, 50 μg/kg/day for 12 days, L-NAME-induced hypertension did not develop (97.6±4.8 mmHg for GW9662 vs. 131.7±6.5 mmHg for vehicle, p&lt;0.01). The PPARγ antagonist also prevented L-NAME-induced hypertension in TG-injected Alox15-/- mice, indicating a PPARγ-mediated effect in macrophages that results in hypertension. These results demonstrate that TG selectively upregulates macrophage PPARγ, bypassing the missing component in Alox15-/- macrophages, and permitting L-NAME-induced hypertension in Alox15-/- mice. Thus, the macrophage Alox15/PPARγ pathway is an important checkpoint in the development of L-NAME-induced hypertension.

Pharmacological inhibition ofS-nitrosoglutathione reductase improves endothelial vasodilatory function in rats in vivo

Nitric oxide (NO) exerts a wide range of cellular effects in the cardiovascular system. NO is short lived, but S-nitrosoglutathione (GSNO) functions as a stable intracellular bioavailable NO pool. Accordingly, increased levels can facilitate NO-mediated processes, and conversely, catabolism of GSNO by the regulatory enzyme GSNO reductase (GSNOR) can impair these processes. Because dysregulated GSNOR can interfere with processes relevant to cardiovascular health, it follows that inhibition of GSNOR may be beneficial. However, the effect of GSNOR inhibition on vascular activity is unknown. To study the effects of GSNOR inhibition on endothelial function, we treated rats with a small-molecule inhibitor of GSNOR (N6338) that has vasodilatory effects on isolated aortic rings and assessed effects on arterial flow-mediated dilation (FMD), an NO-dependent process. GSNOR inhibition with a single intravenous dose of N6338 preserved FMD (15.3 ± 5.4 vs. 14.2 ± 6.3%, P = nonsignificant) under partial NO synthase inhibition that normally reduces FMD by roughly 50% (14.1 ± 2.9 vs. 7.6 ± 4.4%, P < 0.05). In hypertensive rats, daily oral administration of N6338 for 14 days reduced blood pressure (170.0 ± 5.3/122.7 ± 6.4 vs. 203.8 ± 1.9/143.7 ± 7.5 mmHg for vehicle, P < 0.001) and vascular resistance index (1.5 ± 0.4 vs. 3.2 ± 1.0 mmHg · min · l(-1) for vehicle, P < 0.001), and restored FMD from an initially impaired state (7.4 ± 1.7%, day 0) to a level (13.0 ± 3.1%, day 14, P < 0.001) similar to that observed in normotensive rats. N6338 also reversed the pathological kidney changes exhibited by the hypertensive rats. GSNOR inhibition preserves FMD under conditions of impaired NO production and protects against both microvascular and conduit artery dysfunction in a model of hypertension.

Abstract 55: Involvement of Matrix Metalloproteinase-2 During the Progression of Renal Disease in Diabetic Dahl Salt-sensitive Rats

Recently, our laboratory reported that Dahl salt-sensitive (SS) rats develop a form of renal disease following induction of diabetes with streptozotocin (STZ) that is similar to patients with diabetic nephropathy (DN). The progression of renal injury in this model was associated with increased levels of matrix metalloproteinase-2 (MMP-2) in the renal cortex. The present study examined the role of MMP-2 during the progression of diabetes-induced renal injury by comparing the development of proteinuria and renal injury following the induction of type I diabetes in SS rats and in a MMP-2 Zinc finger KO strain of SS rats with an 8 base pair frame-shift deletion (1433-1440) in exon 7 (MMP-2 ZN KO strain). The glomerular expression of MMP-2 protein was non-detectable in the MMP-2 ZN KO strain compared to SS rats fed a low salt (LS) diet. We next performed studies using 9 week-old SS and MMP-2 ZN KO rats treated with either vehicle or (2) STZ, 50 mg/kg (i.p.) to induce diabetes and fed the rats a low salt (LS) diet containing 0.4% NaCl to minimize the development of hypertension. At 18 weeks of age, protein excretion increased to 303±39 mg/day in STZ-treated SS rats (n=7) versus 112±12 mg/day in vehicle treated rats. Protein excretion only increased to 150±23 mg/day in the STZ-treated MMP-2 ZN KO strain (n=7). Blood pressure was not significantly altered and averaged 109±4 mmHg in vehicle and STZ-treated SS rats versus 108±4 mmHg in the STZ-treated MMP2-ZN KO animals. STZ-treated SS rats exhibited marked glomerular injury and extensive renal fibrosis. The degree of glomerulosclerosis and renal interstitial fibrosis was significantly reduced in the kidneys of the MMP-2 ZN KO strain. These data indicate that the progression of diabetes-induced renal injury in STZ-SS rats is associated with upregulation of the expression and activity of MMP-2 and that KO of MMP-2 gene function markedly reduces the development of proteinuria and renal injury in this model of type I diabetic nephropathy. These results also suggest that selective MMP inhibitors may be useful to prevent the development and/or progression of chronic kidney disease in the millions of patients suffering from diabetes.

Renal Inhibition of Heme Oxygenase-1 Increases Blood Pressure in Angiotensin II-Dependent Hypertension

The goal of this study was to test the hypothesis that renal medullary heme oxygenase (HO) acts as a buffer against Ang-II dependent hypertension. To test this hypothesis, renal medullary HO activity was blocked using QC-13, an imidazole-dioxolane HO-1 inhibitor, or SnMP, a classical porphyrin based HO inhibitor. HO inhibitors were infused via IRMI catheters throughout the study starting 3 days prior to implantation of an osmotic minipump which delivered Ang II or saline vehicle. MAP was increased by Ang II infusion and further increased by IRMI infusion of QC-13 or SnMP. MAP averaged 113 ± 3, 120 ± 7, 141 ± 2, 153 ± 2, and 154 ± 3 mmHg in vehicle, vehicle + IRMI QC-13, Ang II, Ang II + IRMI QC-13, and Ang II + IRMI SnMP treated mice, respectively (n = 6). Inhibition of renal medullary HO activity with QC-13 in Ang II infused mice was also associated with a significant increase in superoxide production as well as significant decreases in antioxidant enzymes catalase and MnSOD. These results demonstrate that renal inhibition of HO exacerbates Ang II dependent hypertension through a mechanism which is associated with increases in superoxide production and decreases in antioxidant enzymes.

The Soluble Epoxide Hydrolase Inhibitor AR9281 Decreases Blood Pressure, Ameliorates Renal Injury and Improves Vascular Function in Hypertension.

Soluble epoxide hydrolase inhibitors (sEHIs) are demonstrating promise as potential pharmaceutical agents for the treatment of cardiovascular disease, diabetes, inflammation, and kidney disease. The present study determined the ability of a first-in-class sEHI, AR9281, to decrease blood pressure, improve vascular function, and decrease renal inflammation and injury in angiotensin hypertension. Rats were infused with angiotensin and AR9281 was given orally during the 14-day infusion period. Systolic blood pressure averaged 180 ± 5 mmHg in vehicle treated and AR9281 treatment significantly lowered blood pressure to 142 ± 7 mmHg in angiotensin hypertension. Histological analysis demonstrated decreased injury to the juxtamedullary glomeruli. Renal expression of inflammatory genes was increased in angiotensin hypertension and two weeks of AR9281 treatment decreased this index of renal inflammation. Vascular function in angiotensin hypertension was also improved by AR9281 treatment. Decreased afferent arteriolar and mesenteric resistance endothelial dependent dilator responses were ameliorated by AR9281 treatment of angiotensin hypertensive rats. These data demonstrate that the first-in-class sEHI, AR9281, lowers blood pressure, improves vascular function and reduces renal damage in angiotensin hypertension.

Administration of a substituted adamantyl urea inhibitor of soluble epoxide hydrolase protects the kidney from damage in hypertensive Goto–Kakizaki rats

Hypertension and Type 2 diabetes are co-morbid diseases that lead to the development of nephropathy. sEH (soluble epoxide hydrolase) inhibitors are reported to provide protection from renal injury. We hypothesized that the sEH inhibitor AUDA [12-(3-adamantan-1-yl-ureido)-dodecanoic acid] protects the kidney from the development of nephropathy associated with hypertension and Type 2 diabetes. Hypertension was induced in spontaneously diabetic GK (Goto-Kakizaki) rats using AngII (angiotensin II) and a high-salt diet. Hypertensive GK rats were treated for 2 weeks with either AUDA or its vehicle added to drinking water. MAP (mean arterial pressure) increased from 118+/-2 mmHg to 182+/-20 and 187+/-6 mmHg for vehicle and AUDA-treated hypertensive GK rats respectively. AUDA treatment did not alter blood glucose. Hypertension in GK rats resulted in a 17-fold increase in urinary albumin excretion, which was decreased with AUDA treatment. Renal histological evaluation determined that AUDA treatment decreased glomerular and tubular damage. In addition, AUDA treatment attenuated macrophage infiltration and inhibited urinary excretion of MCP-1 (monocyte chemoattractant protein-1) and kidney cortex MCP-1 gene expression. Taken together, these results provide evidence that sEH inhibition with AUDA attenuates the progression of renal damage associated with hypertension and Type 2 diabetes.

Kidney-Specific Induction of Heme Oxygenase-1 Prevents Angiotensin II Hypertension

The main goal of this study was to determine whether kidney-specific induction of heme oxygenase-1 (HO-1) can prevent the development of angiotensin (Ang) II-dependent hypertension. To test this hypothesis, intrarenal medullary interstitial catheters were implanted into the left kidney of uninephrectomized mice. Infusion of cobalt protoporphyrin (CoPP; 250 microg/mL; at 50 microL/h for 48 hours) resulted in significant induction of HO-1 in the renal medulla when examined 2 weeks after the infusion with no induction observed in other organs, such as the heart or liver. Next, we examined the effect of renal-specific induction of HO-1 on the development of Ang II-dependent hypertension. CoPP or vehicle (0.1 mol/L NaOH [pH 8.3]) was infused as indicated above 2 days before implantation of an osmotic minipump, which delivered Ang II or saline vehicle at a rate of 1 microg/kg per minute. Mean arterial pressure was measured in conscious, unrestrained mice for 3 consecutive days starting on day 7 after implantation of the minipumps. Mean arterial pressure averaged 114+/-5, 122+/-4, 162+/-2, and 125+/-6 mm Hg in vehicle-, intrarenal medullary interstitial CoPP-, Ang II-, and Ang II + intrarenal medullary interstitial CoPP-treated mice, respectively (n=6 or 7). These results demonstrate that kidney-specific induction of HO-1 prevents the development of Ang II-dependent hypertension and that induction of HO-1 in the kidney may be the mechanism by which systemic delivery of CoPP lowers blood pressure in Ang II-dependent hypertension.

Renal Medullary Infusion of CoPP Prevents Angiotensin‐II Dependent Hypertension in Mice

Several studies have established systemic induction of Heme Oxygenase-1 (HO-1) as a viable means of attenuating the development of hypertension in several experimental models. However, the precise mechanism behind the anti-hypertensive actions of HO-1 induction is still unclear. In this study, we tested the hypothesis that kidney specific induction of HO-1 can prevent the development of Ang-II dependent hypertension. To test this hypothesis, renal medullary interstitial catheters were implanted into the left kidney of uninephrectomized mice. Infusion of a known inducer of HO-1, cobalt protoporphyrin (CoPP; 250 μg/ml; at 50 μl/hr for 48hrs), resulted in significant induction of HO-1 mainly in the renal medulla when examined 2 weeks after the infusion with no induction observed in other organs such as the brain, heart, and liver. Next, we examined the effect of this approach of inducing of HO-1 on the development of Ang II dependent hypertension. CoPP or Vehicle (0.1 M NaOH, pH 8.3) was infused as indicated above 2 days prior to implantation of an osmotic minipump which delivered Ang II at a rate of 1 μg/kg/min. Mean arterial pressure (MAP) was measured in conscious, unrestrained mice for 3 consecutive days starting 7 days after the implantation of the Ang II minipumps. MAP averaged 112 ± 4, 121 ± 4, 162 ± 2 and 126 ± 6 mmHg in Vehicle + Vehicle, CoPP + Vehicle, Vehicle + Ang II and CoPP + Ang II treated mice respectively (n=6). These results suggest that renal medullary infusion of CoPP to induce HO-1 specifically in the kidneys can prevent the development of Ang II dependent hypertension in mice. Induction of HO-1 in the renal medulla may be the mechanism by which systemic delivery of CoPP lowers blood pressure in Ang-II dependent hypertension.

Evaluation of travoprost as adjunctive therapy in patients with uncontrolled intraocular pressure while using timolol 0.5%

PURPOSE: To evaluate the intraocular pressure-lowering efficacy and safety of travoprost 0.0015% and 0.004%, dosed daily in the evening compared with vehicle, in patients with open-angle glaucoma or ocular hypertension, whose intraocular pressure was not adequately controlled on timolol 0.5% twice daily (twice daily). METHODS: Subjects who qualified at screening began a run-in period dosing timolol twice daily for 3 weeks. If the subjects had an intraocular pressure of 24 to 36 mm Hg at 8 am and 21 to 36 mm Hg at 10 am and 4 pm in at least one eye on timolol, they were randomized to one of two concentrations of travoprost (0.0015% or 0.004%) or vehicle solution every day and were followed for 6 months. Four hundred twenty-six subjects were randomized. The mean intraocular pressure at 8 am, 10 am, and 4 pm in the patient’s eye with the higher intraocular pressure was used for the analysis. RESULTS: Mean baseline values (25 mm Hg) for subjects at eligibility (while maintained on timolol) were not significantly different ( P < .0001) among the treatment groups. The intraocular pressure was lowered an additional −5.7 to −7.2 mm Hg and −5.1 to −6.7 mm Hg in the travoprost 0.004% and 0.0015% concentrations, respectively. These changes were significantly ( P ≤ .0001) different from the vehicle group (−1.3 to −2.8 mm Hg). The intraocular pressure range on treatment at all visit times over the 6-month treatment period ranged from 17.9 to 19.2 mm Hg for travoprost 0.004% and 18.3 to 20.1 mm Hg for travoprost 0.0015% compared with 22.4 to 24.1 mm Hg for vehicle. Average hyperemia scores ranged from trace to mild (mean 0.5 on a scale of 0 = none/trace; 1= mild; 2 = moderate; 3 = severe) for all treatment groups. No iris pigmentation changes were observed in any patient during this study. There were no clinically or statistically significant changes from baseline in visual acuity, ocular cells and flare, fundus parameter, cup-to-disk ratio and visual field between the treatment groups. There were no serious adverse events reported for any treatment group. CONCLUSIONS: Travoprost produced clinically relevant and statistically significant additional intraocular pressure reductions from baseline when used adjunctively with timolol in subjects with open-angle glaucoma or ocular hypertension.

Melatonin increases pial artery tone and decreases the lower limit of cerebral blood flow autoregulation.

We studied a possible link between the melatonin-induced increase in cerebral arteriolar tone and the melatonin-induced shift in cerebral blood flow (CBF) autoregulation to a lower pressure level. Using the cranial window technique, we showed that intravenous infusion of melatonin constricted pial arterioles (-5.1 +/- 1.3 and -5.4 +/- 0.7 microm at 60 and 600 ng/kg/h, respectively). Perivascular application of luzindole alone had no significant effect but abolished vasoconstriction induced by melatonin (-0.5 +/- 0.7 and + 3.0 +/- 1.2 microm at 60 and 600 ng/kg/h respectively). Using a combination of the hydrogen clearance and cranial window techniques, we showed that intravenous infusion of melatonin had no effect on baseline CBF but shifted the lower limit (LL) of CBF autoregulation (stepwise hypotension) to a lower pressure (90 +/- 2 mmHg in vehicle vs. 71 +/- 3 and 51 +/- 5 mmHg, both P < 0.05, after melatonin at 60 and 600 ng/kg/h, respectively). As melatonin had no effect on systemic blood pressure yet shifted the LL of CBF autoregulation, the security margin increased (28 +/- 5 in controls vs. 38 +/- 3 and 55 +/- 5% after melatonin at 60 and 600 ng/kg/h, respectively, both P < 0.05). The higher i.v. infusion rate of melatonin increased the relative arteriolar dilatory response to hypotension but did not increase absolute diameter at any given pressure level. Our results show that melatonin shifts the LL of CBF autoregulation to a lower systemic pressure level. This effect does not appear to be explained by the effect of melatonin on cerebral arteriolar diameter.

Mechanism of decreased adenosine protection in reperfusion injury of aging rats.

The purpose of this study was to determine whether the protective effects of adenosine on myocardial ischemia-reperfusion injury are altered with age, and if so, to clarify the mechanisms that underlie this change related to nitric oxide (NO) derived from the vascular endothelium. Isolated perfused rat hearts were exposed to 30 min of ischemia and 60 min of reperfusion. In the adult hearts, administration of adenosine (5 micromol/l) stimulated NO release (1. 06 +/- 0.19 nmol. min(-1). g(-1), P < 0.01 vs. vehicle), increased coronary flow, improved cardiac functional recovery (left ventricular developed pressure 79 +/- 3.8 vs. 57 +/- 3.1 mmHg in vehicle, P < 0.001; maximal rate of left ventricular pressure development 2,385 +/- 103 vs. 1,780 +/- 96 in vehicle, P < 0.001), and reduced myocardial creatine kinase loss (95 +/- 3.9 vs. 159 +/- 4.6 U/100 mg protein, P < 0.01). In aged hearts, adenosine-stimulated NO release was markedly reduced (+0.42 +/- 0.12 nmol. min(-1). g(-1) vs. vehicle), and the cardioprotective effects of adenosine were also attenuated. Inhibition of NO production in the adult hearts significantly decreased the cardioprotective effects of adenosine, whereas supplementation of NO in the aged hearts significantly enhanced the cardioprotective effects of adenosine. The results show that the protective effects of adenosine on myocardial ischemia-reperfusion injury are markedly diminished in aged animals, and that the loss in NO release in response to adenosine may be at least partially responsible for this age-related alteration.

Delayed myocardial preconditioning by α1-adrenoceptors involves inhibition of apoptosis

Objective: Previous studies have demonstrated that α1-adrenoceptor activation increases myocardial resistance to ischemic injury 24 hours later. Here we tested the hypothesis that delayed protection is associated with limited infarction and involves altered expression of pro-apoptotic and/or anti-apoptotic proteins. Methods: Rabbits were treated with phenylephrine or an equivalent volume of vehicle (n = 6 per group). Twenty-four hours after pretreatment, isolated hearts were perfused with a bovine erythrocyte suspension in modified Krebs solution, subjected to 45 minutes of global ischemia (37°C), and reperfused for 120 minutes. Infarct size was determined by triphenyltetrazolium chloride staining. Apoptosis was quantified by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. Left ventricular tissue from separate groups of animals (n = 5 per group), 24 hours after pretreatment with phenylephrine or vehicle but without ischemia and reperfusion, was analyzed by Western blotting for content of the anti-apoptotic protein, bcl x, and pro-apoptotic protein, bax. Results: Isolated hearts after phenylephrine pretreatment had increased end-reperfusion developed pressures (56.8 ± 4.9 vs 36.2 ± 3.9 mm Hg for vehicle, P = .008) and decreased elevated end-diastolic pressures (26.7 ± 4.5 vs 42.3 ± 5.0 mm Hg for vehicle, P = .04). Phenylephrine pretreatment abrogated infarction (9.9 ± 2.4% vs 42.6 ± 6.3% for vehicle, P = .002) and reduced the number of apoptotic nuclei (24 ± 4.8 vs 51 ± 4.6 for vehicle, P = .038). Analysis by Western blotting showed that the ratio of bcl x to bax protein increased in phenylephrine-pretreated hearts (2.65 ± 0.5 vs 1.0 ± 0.1 for vehicle, P = .008). Conclusion: Delayed myocardial protection to infarction mediated by α1-adrenoceptor activation involves an increased bcl x/bax ratio, thereby limiting apoptotic cell death. (J Thorac Cardiovasc Surg 1999;117:980-6)

Thromboxane is required for full expression of angiotensin hypertension in rats.

Recent studies suggest that thromboxane (TX) mediates a significant component of angiotensin II (ANG II)-induced hypertension. However, there is little information to support the hypothesis that this relationship is important during chronic, physiological increases in ANG II, particularly while controlling for variation in endogenous ANG II levels induced by TX inhibition. This study tested that hypothesis in 27 chronically instrumented rats. After baseline measurements, suppression of endogenous TX was induced and maintained throughout the study in 13 rats by i.v. infusion of the TX synthesis inhibitor (TSI) U63557A: the other 14 rats received vehicle. Baseline mean arterial pressure (MAP) was not different between groups and was unchanged by TSI or vehicle. Continuous inhibition of ANG II production was then initiated in both groups of rats by i.v. infusion of the angiotensin-converting enzyme inhibitor (ACEI) benazepril. ACEI reduced blood pressure similarly in vehicle and TSI rats, from 105 +/- 2 to 91 +/- 2 mm Hg and 103 +/- 1 to 89 +/- 1 mm Hg, respectively. ANG II was then infused at 5 ng.kg-1.min-1 i.v. for 7 days in six rats from each group to restore ANG II activity to baseline levels. This dose increased MAP to 103 +/- 2 and 101 +/- 1 mm Hg in vehicle and TSI rats, respectively, values not different from pre-ACEI levels. Seven TSI rats and eight vehicle rats received a higher dose of ANG II (20 ng.kg-1.min-1 i.v.). After 7 days, MAP was higher in vehicle than in TSI rats (143 +/- 5 versus 120 +/- 4 mm Hg). These results suggest that endogenous TX is an important determinant of MAP in ANG II hypertension but may have a diminished role in blood pressure regulation when ANG II is at normal and subnormal levels.