Older adults with heart failure treated with carvedilol, bisoprolol, or metoprolol tartrate: risk of mortality.

PURPOSE: The long-term use of ß-blockers has been shown to improve clinical outcomes among patients with heart failure (HF). However, a lack of data persists in assessing whether carvedilol or bisoprolol are superior to metoprolol tartrate in clinical practice. We endeavored to compare the effectiveness of ß-blockers among older adults following a primary hospital admission for HF. METHODS: We conducted a cohort study using Quebec administrative databases to identify patients who were using ß-blockers, carvedilol, bisoprolol, or metoprolol tartrate after the diagnosis of HF. We characterized the patients by the type of ß-blocker prescribed at discharge of their first HF hospitalization. An adjusted multivariate Cox proportional hazards model was used to compare the primary outcome of all-cause mortality. We also conducted analyses by matching for a propensity score for initiation of ß-blocker therapy and assessed the effect on primary outcome. RESULTS: Among 3197 patients with HF with a median follow-up of 2.8 years, the crude annual mortality rates (per 100 person-years) were at 16, 14.9, and 17.7 for metoprolol tartrate, carvedilol, and bisoprolol, respectively. Adjusted hazard ratios of carvedilol (hazard ratio 0.92; 0.78-1.09) and bisoprolol (hazard ratio 1.04; 0.93-1.16) were not significantly different from that of metoprolol tartrate in improving survival. After matching for propensity score, carvedilol and bisoprolol showed no additional benefit with respect to all-cause mortality compared with metoprolol tartrate. CONCLUSIONS: Our evidence suggests no differential effect of ß-blockers on all-cause mortality among older adults with HF. Copyright © 2016 John Wiley & Sons, Ltd.

Induction of anti-anti-idiotype antibodies against sulfated glycosaminoglycans reduces atherosclerosis in apolipoprotein E-deficient mice.

OBJECTIVE: The pathogenesis of atherosclerosis is associated with the early retention of low-density lipoproteins that are trapped in the extracellular matrix of the arterial intima by interaction with glycosaminoglycan side chains of proteoglycans. Mutant mouse/human chimeric antibodies of the murine monoclonal antibody P3, which react with N-glycolyl-containing gangliosides and sulfated glycosaminoglycans, were tested for their potentially antiatherogenic properties through the induction of an idiotypic antibody network that may specifically interfere with the binding of low-density lipoproteins to proteoglycan side chains, low-density lipoprotein modification, and foam cell formation. METHODS AND RESULTS: Apolipoprotein E-deficient mice fed a high-fat, high-cholesterol diet received 5 to 6 doses of chP3R99 or chP3S98 mutant antibodies, showing high and low reactivity, respectively, against their respective antigens. Both chimeric antibodies elicited an immunodominant anti-idiotypic response in the absence of adjuvant. A striking (40%-43%) reduction (P<0.01) in total lesion areas was observed in 18-week-old mice immunized with chP3R99, but not chP3S98, compared with PBS-treated mice. The antiatherosclerotic effect was associated with increased mice sera reactivity against heparin and sulfated glycosaminoglycans, including chondroitin and dermatan sulfate. In addition, purified IgG from chP3R99-immunized mice blocked the retention of apolipoprotein B-containing lipoproteins within the arterial wall of apolipoprotein E(-/-) mice. CONCLUSIONS: The present study supports use of active immunization and the mounting of an idiotypic antibody network response against glycosaminoglycans as a novel approach to target atherosclerosis.

Impact of α-lipoic acid on liver peroxisome proliferator-activated receptor-α, vascular remodeling, and oxidative stress in insulin-resistant rats.

This study sought to determine the impact of α-lipoic acid (LA) on superoxide anion (O(2)(•-)) production and peroxisome proliferator-activated receptor-α (PPARα) expression in liver tissue, plasma free fatty acids (FFA), and aortic remodeling in a rat model of insulin resistance. Sprague-Dawley rats (50-75 g) were given either tap water or a drinking solution containing 10% D-glucose for 14 weeks, combined with a diet with or without LA supplement. O(2)(•-) production was measured by lucigenin chemiluminescence, and PPAR-α expression by Western blotting. Cross-sectional area (CSA) of the aortic media and lumen and number of smooth muscle cells (SMC) were determined histologically. Glucose increased systolic blood pressure (SBP), plasma levels of glucose and insulin, and insulin resistance (HOMA index). All of these effects were attenuated by LA. Whereas glucose had no effect on liver PPAR-α protein level, it decreased plasma FFA. LA decreased the aortic and liver O(2)(•-) production, body weight, and plasma FFA levels in control and glucose-treated rats. Liver PPAR-α protein levels were increased by LA, and negatively correlated with plasma FFA. Medial CSA was reduced in all glucose-treated rats, and positively correlated with plasma FFA but not with SBP or aortic O(2)(•-) production. Glucose also reduced aortic lumen area, so that the media-to-lumen ratio remained unchanged. The ability of LA to lower plasma FFA appears to be mediated, in part, by increased hepatic PPAR-α expression, which may positively affect insulin resistance. Glucose-fed rats may serve as a unique model of aortic atrophic remodeling in hypertension and early metabolic syndrome.

The cardiac neural stem cell phenotype is compromised in streptozotocin-induced diabetic cardiomyopathy.

Neural stem cells were identified in the rat heart and during scar formation and healing participated in sympathetic fiber sprouting and angiogenesis. In the setting of diabetes, impaired wound healing represents a typical pathological feature. These findings provided the impetus to test the hypothesis that experimental diabetes adversely influenced the phenotype of cardiac neural stem cells. Streptozotocin (STZ)-induced diabetic rats were associated with elevated plasma glucose levels, significant loss of body weight and left ventricular contractile dysfunction. In the heart of STZ-diabetic rats, the density of nestin immunoreactive processes emanating from cardiac neural stem cells were reduced. The latter finding was reaffirmed as nestin protein expression was significantly decreased in the heart of STZ-diabetic rats and associated with a concomitant reduction of nestin mRNA. Employing the TUNEL assay, the loss of nestin expression in STZ-diabetic rats was not attributed to widespread cardiac neural stem cell apoptosis. Insulin administration to STZ-diabetic rats with established hyperglycaemia led to a modest recovery of nestin protein expression in cardiac neural stem cells. By contrast, the administration of insulin immediately after STZ injection improved plasma glucose levels and significantly attenuated the loss of nestin protein expression. These data highlight the novel observation that nestin protein expression in cardiac neural stem cells was significantly reduced in STZ-induced type I diabetic rats. The aberrant cardiac neural stem cell phenotype may compromise their biological role and predispose the diabetic heart to maladaptive healing following ischemic injury.

Targeting vascular structure for the treatment of sexual dysfunction.

INTRODUCTION: Erectile dysfunction (ED) and cardiovascular disease often coexist and have many common risk factors. In hypertension, the structure of blood vessels is modified such that there is an increase in medial wall thickness relative to lumen size. Certain antihypertensive agents have been found to induce a regression of vascular structure such that a "hypertensive" vessel appears phenotypically more like that from a normotensive. AIM: To provide an update on the findings to date on the impact of vascular remodeling on erectile function. MAIN OUTCOME MEASURES: Review of peer reviewed literature related to vascular remodeling induced by antihypertensive agents and the potential impact on sexual function. METHODS: A literature review was performed on clinical and experimental evidence regarding the association between cardiovascular disease and ED, the impact of vascular remodeling on these conditions, the impact of antihypertensive therapy on ED, and the mechanisms of antihypertensive drug-induced remodeling. RESULTS: There is increasing evidence that ED may be an early marker for progressing cardiovascular disease. Certain antihypertensive agents have beneficial effects on both vascular structure and erectile function. The major site of resistance in the penile vasculature occurs at the level of the pudendal artery. Although structural remodeling has not yet been investigated in this vessel specifically, antihypertensive drugs have been shown to induce remodeling of the pudendal-penile vasculature and cavernosal arteries. Antihypertensive drug-induced vascular remodeling can be characterized by a decrease in the ratio of wall thickness to lumen diameter, and may result from vascular smooth muscle cell apoptosis, rearrangement of cells around a smaller lumen, and/or changes in the extracellular matrix composition depending on the vessel type. CONCLUSION: Determining the mechanisms involved in antihypertensive drug-induced vascular remodeling in the pudendal vasculature may provide novel targets for the treatment of ED.

Effect of chronic inhibition of nitric oxide on hypertension, insulin resistance, and cardiovascular remodeling in glucose-fed rats.

To determine the contribution of nitric oxide (NO) in cardiovascular remodeling associated to hypertension and insulin resistance, male Sprague-Dawley rats received tap water supplemented or not (control), with 10% D-glucose (G) and/or 50 mg x kg(-1) x d(-1) L-NAME to inhibit NO synthase (G-LN or LN) for 4 weeks. Systolic blood pressure increased by 12%, 26%, and 39% with G, LN, and G-LN treatments, respectively. Hyperinsulinemia and insulin resistance (homeostasis model assessment index) occurred in G-treated rats (P < 0.05) and were further increased in G-LN (P < 0.01). Plasma adrenaline concentrations were markedly increased in all treated groups, especially in G-LN (P < 0.01), whereas noradrenaline was increased in G-treated rats only. Whereas no cardiac hypertrophy or fibrosis was detected, aortic hypertrophy occurred in LN and G-LN rats (P < 0.001) without smooth muscle hyperplasia. Superoxide anion formation was increased in the aorta of all treated groups (P < 0.01) and in the heart of LN (P < 0.05), but reduced nicotinamide adenine dinucleotide phosphate (NAD(P)H) oxidase activity was not affected. In conclusion, the loss of the wide-range protective effects of NO, the increased vascular oxidative stress, and the sympathoadrenal hyperactivity are among the contributing factors leading to the exacerbation of hypertension and insulin resistance in G-LN. These factors were sufficient to cause vascular but not cardiac hypertrophy.

Functional Dosage of Muscarinic Cholinergic Receptor 3 Signalling, Not the Gene Dose, Determines Its Hypertension Pathogenesis.

BACKGROUND: Multiple quantitative trait loci for blood pressure (BP) have been localized throughout human and rodent genomes. Few of them have been functionally identified especially in humans, and little is known about their pathogenic directionality when identified. We focused on Chrm3 encoding the muscarinic cholinergic receptor 3 (M3R) as the causal gene for C17QTL1 in the Dahl salt-sensitive rat model. METHODS AND RESULTS: Congenic knock-ins, gene-specific knockout, and ex vivo and in vivo function studies were applied in the Dahl salt-sensitive rat model of polygenic hypertension. A Chrm3 missense T1667C mutation in the last intracellular domain functionally correlated with a rise in BP increased the M3R signalling and resensitization, and adrenal epinephrogenesis. Gene targeting that abolished the M3R function without affecting any of noncoding Chrm3 variants caused a decrease in BP, indicating that the M3R-mediated signalling promotes hypertension. In contrast, removing 8 amino acids from the M3R first extracellular loop had no effect on BP. CONCLUSIONS: The M3R-specialized signalling constitutes a new pathway of hypertension pathogenesis within the context of a polygenic and quantitative trait. Increased epinephrine in the circulation and secreted from the adrenal glands are suggestive of a molecular mechanism partially mediating M3R to promote hypertension. The structure-function relationships for various M3R domains in their effects on BP pave the way for identifying missense mutations that impact functions on BP as potential diagnostic targets.

RGS5, RGS4, and RGS2 expression and aortic contractibility are dynamically co-regulated during aortic banding-induced hypertrophy.

Overexpression of regulator of G protein signaling 5 (RGS5) in arteries over veins is the most striking difference observed using microarray analysis. The obvious question is what arterial function might require RGS5. Based on functions of homologous proteins in regulating cardiac mass and G-protein-coupled receptor (GPCR) signaling, we proposed that RGS5 and vascular expressed RGS2 and RGS4 could participate in regulating arterial hypertrophy. We used the suprarenal abdominal aorta banding model to induce hypertension and hypertrophy. All 3 RGS messages were expressed in unmanipulated aorta with RGS5 predominating. After 2 days, thoracic aorta lost expression of RGS5, 4, and 2. At 1 week, all three returned to normal, and at 28 days, they increased many fold above normal. Valsartan blockade of angiotensin II (angII)/angII type 1 receptor signaling prevented upregulation of RGS messages but only delayed mass increases, implying wall mass regulation involves both angII-dependent and angII-independent pathways. The abdominal aorta showed less dramatic expression changes in RGS5 and 4, but not 2. Again, those changes were delayed by valsartan treatment with no mass changes. Thoracic aorta contraction to GPCR agonists was examined in aortic explant rings to identify vessel wall physiological changes. In 2-day aorta, the response to Galphaq/i agonists increased above normal, while 28-day aorta had attenuated induced contraction via Galphaq/i agonist, implicating a connection between RGS message levels and changes in GPCR-induced contraction. In vitro overexpression studies showed RGS5 inhibits angII-induced signaling in smooth muscle cells. This study is the first experimental evidence that changes in RGS expression and function correlate with vascular remodeling.

A new rat model of diabetic macrovascular complication.

OBJECTIVES: Age-related medial calcification (elastocalcinosis) of large arteries is accelerated in diabetes and appears mainly in distal arteries. The aim was to devise a rat model of elastocalcinosis in association with diabetes to examine the hypothesis that diabetes accelerates vascular calcification experimentally. METHODS: Male Wistar rats received a high fat diet during 2 months followed by a low dose of streptozotocin to induce diabetes (D). Elastocalcinosis was facilitated by 3 weeks of treatment with warfarin and vitamin K (WVK). We started WVK treatment 1 week (D4WVK) and 4 weeks (D7WVK) after the injection of streptozotocin and in age-matched healthy rats. Measurements of hemodynamic and metabolic parameters, aortic and femoral calcium content, and immunohistochemistry for alkaline phosphatase, osteopontin, tumor necrosis factor (TNF)-alpha, and transforming growth factor (TGF)-TGF-beta were performed. RESULTS: Three weeks of WVK treatment alone did not increase the calcium content in the aorta and femoral arteries. However, in the D7WVK group, femoral calcification, but not aortic calcium content, increased significantly as compared to the WVK group. This response was not observed in the D4WVK group. In femoral arteries, strong immunostaining for alkaline phosphatase and osteopontin was observed in the D7WVK group. TNF-alpha and TGF-beta expressions were mainly localized in the adventitia of arteries from diabetic rats. CONCLUSION: We have established a model of accelerated elastocalcinosis in diabetes related to its duration and localized in distal arteries. The modification of local protein expression is also in accordance with clinical data, suggesting that this model could be useful to investigate mechanisms related to this important clinical macrovascular complication of diabetes.

Novel Pathogenesis of Hypertension and Diastolic Dysfunction Caused by M3R (Muscarinic Cholinergic 3 Receptor) Signaling.

Multiple quantitative trait loci for blood pressure (BP) are localized in humans and rodent models. Model studies have not only produced human quantitative trait loci homologues but also provided unforeseen mechanistic insights into the function modality of quantitative trait loci actions. Presently, congenic knockins, gene-specific knockout, and in vitro and in vivo function studies were used in a rat model of polygenic hypertension, DSS (Dahl salt sensitive) rats. One gene previously unknown in regulating BP was detected with 1 structural mutation(s) for each of 2 quantitative trait loci classified into 2 separate epistatic modules 1 and 3. C17QTL1 in epistatic module 2 was identified to be the gene Chrm3 encoding the M3R (muscarinic cholinergic 3 receptor), since a single function-enhancing M3RT556M conversion correlated with elevated BP. To definitively prove that the enhanced M3R function is responsible for BP changes by the DSS alleles of C17QTL1, we generated a Chrm3 gene-specific rat knockout. We observed a reduction in BP without tachycardia in both sexes, regardless of the amount of dietary salt, and an improvement in diastolic and kidney dysfunctions. All occurred in spite of a significant reduction in M3R-dependent vasodilation. The previously seen sexual dimorphism for C17QTL1 on BP disappeared in the absence of M3R. A Chrm3-coding variation increased M3R signaling, correlating with higher BP. Removing the M3R signaling led to a decrease in BP and improvements in cardiac and renal malfunctions. A novel pathogenic pathway accounted for a portion of polygenic hypertension and has implications in applying new diagnostic and therapeutic uses against hypertension and diastolic dysfunction.

Fibroblast apoptosis precedes cardiomyocyte mass reduction during left ventricular remodeling in hypertensive rats treated with amlodipine.

BACKGROUND: A transient induction of apoptosis accompanies the normalization of left ventricular mass index in spontaneously hypertensive rats (SHR) treated with dihydropyridine calcium-channel blockers. However, the cell type undergoing apoptosis in this model and the temporal correlation with onset cardiac remodeling remain undefined. METHODS AND RESULTS: SHR were treated either with vehicle or amlodipine (20 mg/kg per day) for 4, 7, 10, 14 or 28 days. Amlodipine stably reduced systolic blood pressure by day 2 (-26 +/- 2%) and stably reduced the left ventricular concentration of atrial natriuretic peptide (ANP) mRNA by approximately 50% as early as day 4, suggesting the early reduction of cardiomyocyte stress. Left ventricular mass index was significantly reduced by day 7 (-4.6 +/- 1.5%), in coordination with reduced DNA content (-23 +/- 2%) and non-cardiomyocyte number (-17 +/- 4%). However, the cardiomyocyte cross-sectional area was reduced only starting from day 14. Caspase-3 cleavage was significantly increased at day 7 only. Ultimately, amlodipine for 28 days induced a slight increase in capillary density without affecting total cardiomyocyte number, while reducing the total number of non-cardiomyocytes down to levels seen in untreated normotensive Wistar-Kyoto rats. Bax to Bcl-2 protein ratios were increased from day 7 to day 28. In situ double labeling by the terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) method (apoptosis) combined with rhodamine-labeled lectin binding (endothelial cell marker) revealed a significant increase (> 3-fold) in TUNEL-positive, lectin-negative non-cardiomyocytes in the interstitium between days 7 and 14. CONCLUSIONS: Left ventricular remodeling induced by amlodipine in SHR involves selective deletion of excess fibroblasts via apoptosis prior to cardiomyocyte mass reduction, but after attenuation of ANP gene expression.

Caspase-dependent cell death mediates the early phase of aortic hypertrophy regression in losartan-treated spontaneously hypertensive rats.

Blockade of angiotensin type 1 (AT1) receptors induces smooth muscle cell (SMC) death and regression of aortic hypertrophy in spontaneously hypertensive rats (SHR). We postulated that SMC death and vascular remodeling in this model may be attenuated by z-Val-Ala-Asp(OMe)-CH2F (z-VAD-fmk), a tripeptide inhibitor of caspase enzymes mediating apoptosis. To determine the time course of SMC death and aortic remodeling, SHR were treated with losartan (30 mg/kg per day) for up to 9.5 days. Transient SMC apoptosis occurred in the aortic media with a peak around day 5 of treatment, with increases in the Bax to Bcl-2 protein ratio (>3-fold), in active caspase-3 (5.6-fold), in TUNEL-positive nuclei (19-fold), preceding by 24 hours the peak activation of capase-9 (3.8-fold), and significant reductions in SMC number (46%) and aortic cross-sectional area (8.5%) at 5.5 days. The decrease in total aortic DNA reached significance at 6.5 days (29%). Blood pressure reduction with losartan was progressive and reached significance at day 7 of treatment. Next, we examined the causal link between vascular apoptosis and remodeling. SHR received placebo or losartan (30 mg/kg per day) for 6 days. During the last 24 hours, a subgroup of losartan-treated rats received 3 IV injections of z-VAD-fmk (cumulative dose: 4.4 mg x kg(-1)). All other rats received the vehicle, DMSO. The 24-hour cotreatment with z-VAD-fmk effectively prevented losartan-induced caspase-3 activation and internucleosomal DNA fragmentation, as well as SMC depletion and the reductions in aortic mass and DNA content. Together, these data suggest that caspase-dependent SMC death mediates the early phase of vascular remodeling in response to AT1 receptor blockade in this model of hypertension.

A quantitative trait locus for aortic smooth muscle cell number acting independently of blood pressure: implicating the angiotensin receptor AT1B gene as a candidate.

Vascular hyperplasia may be involved in the remodeling of vasculature. It was unknown whether there were genetic determinants for aortic smooth muscle cell number (SMCN) and, if so, whether they acted independently of those for blood pressure (BP). To unravel this issue, we utilized congenic strains previously constructed for BP studies. These strains were made by replacing various chromosome 2 segments of the Dahl salt-sensitive (S) rat with those of the Milan normotensive rat (MNS). We measured and compared SMCN in aortic cross-sectional areas and BPs of these strains. Consequently, a quantitative trait locus (QTL) for SMCN was localized to a chromosome region not containing a BP QTL, but harboring the locus for the angiotensin II receptor AT1B (Agtr1b). Agtr1b became a candidate for the SMCN QTL because 1) two significant mutations were found in the coding region between S and all congenic strains possessing the MNS alleles, and 2) contractile responses to angiotensin II were significantly and selectively reduced in congenic rats harboring the MNS alleles of the SMCN QTL compared with S rats. The current investigation presents the first line of evidence that a QTL for aortic SMCN exists, and it acts independently of QTLs for BP. The relevant congenic strains developed therein potentially provide novel mammalian models for the studies of vascular remodeling disorders.

Inhibition of vascular smooth muscle cell proliferation and neointimal formation in injured arteries by a novel, oral mitogen-activated protein kinase/extracellular signal-regulated kinase inhibitor.

BACKGROUND: Mitogen-activated protein kinases (MAPKs) are rapidly induced after arterial injury in different animal models. However, their precise role in vascular smooth muscle cell (VSMC) proliferation and neointimal formation in vivo remains to be determined. METHODS AND RESULTS: We investigated the properties of a novel, selective inhibitor of the upstream kinase, MAPK/extracellular signal-regulated kinase, that is orally active (PD0185625). In vitro, PD0185625 was shown to abrogate p44/p42 MAPK activation in VSMCs after serum stimulation. This was associated with a dose-dependent inhibition of VSMC proliferation. In vivo, PD0185625 was administered orally to rats (200 mg x kg(-1) x d(-1)) beginning 2 days before balloon injury of the left carotid artery and for 2 weeks thereafter. Treatment with PD0185625 led to nearly complete inhibition of p44/p42 MAPK activation after balloon injury. This resulted in a significant decrease in VSMC proliferation (BrdU incorporation) at day 7 after injury. Moreover, neointimal formation was significantly reduced in PD0185625-treated animals at 14 and 28 days after arterial injury. We found that PD0185625 did not increase the rate of apoptotic cell death but prevented cell cycle progression and induced a G1 block. CONCLUSIONS: PD0185625 reduced neointimal formation after arterial injury. The mechanism involved inhibition of VSMC proliferation via a G1 block of the cell cycle. Orally active selective MAPK inhibitors could represent a novel therapeutic approach for vascular diseases.

Reversal of interstitial fibroblast hyperplasia via apoptosis in hypertensive rat heart with valsartan or enalapril.

OBJECTIVE: Renin-angiotensin system inhibitors transiently induce apoptosis at the onset of cardiac hypertrophy regression in spontaneously hypertensive rats (SHRs). The focus of this study is to evaluate the cell selectivity of this response. METHODS: SHRs were treated with valsartan or enalapril (30 mg kg(-1) day(-1)) or placebo for 1 to 4 weeks. Stereological and morphological data were obtained from immunohistological analyses. Apoptosis was quantified by DEVDase (caspase-3-like) activity assay and immunoblot analysis of apoptosis-regulatory proteins (Bax and Bcl-2). Identification of the apoptotic cell type was conducted using in situ TUNEL labeling, in conjunction with alpha-sarcomeric actin or lectin immunoreactivity as markers for cardiomyocytes and endothelial cells, respectively. RESULTS: Stereological analysis of the left ventricle revealed significant non-cardiomyocyte hyperplasia in placebo-treated SHRs (239+/-29x10(6) nuclei) as compared to untreated age-matched normotensive Wistar-Kyoto (WKY) rats (107+/-12x10(6)). In contrast, the number of cardiomyocyte nuclei was comparable between untreated SHRs (48+/-4x10(6)) and WKY rats. After 4 weeks of valsartan or enalapril treatment, SHRs showed significant reductions in systolic blood pressure (>28%), left ventricular hypertrophy (>9%) and cardiomyocyte cross-sectional area (>17%). Moreover, these treatments abolished non-cardiomyocyte hyperplasia in SHR left ventricle without affecting cardiomyocyte number, capillary density or number of capillary per cardiomyocyte nucleus. As a mechanism of cell deletion consistent with apoptosis induction, ventricles showed increased caspase-3 activation (>4.5-fold) as well as Bax to Bcl-2 protein ratio (>3.2-fold) within 2 weeks of valsartan or enalapril treatment. Immunohistological analysis revealed a significant increase in TUNEL-positive, lectin-negative non-cardiomyocytes, suggesting a rise in apoptotic interstitial fibroblasts in the left ventricle within 2 weeks of treatment with valsartan or enalapril (>63%), with a return to baseline (0.033+/-0.003%) at 4 weeks. Treatments did not affect right ventricular mass, apoptosis or cellularity. CONCLUSION: Cardiac apoptosis induction during regression of left ventricular hypertrophy reverses interstitial fibroblast hyperplasia in SHRs treated with inhibitors of the renin-angiotensin system.

Renal ischemia-reperfusion injury in the rat is prevented by a novel immune modulation therapy.

BACKGROUND: Vasogen Inc.'s (Mississauga, Ontario, Canada) immune modulation therapy (IMT) is a therapy in which cells from the patient's own blood are modified by ex vivo exposure to specific physicochemical stressors, including oxidation, ultraviolet (UV) light, and an elevated temperature. The therapy has been shown to have a beneficial effect in models of inflammation and vascular diseases. This study tested the hypothesis that IMT can prevent renal ischemia-reperfusion (I/R) injury in rats. METHODS: Whole blood was collected from syngeneic age-matched donors by cardiac puncture. It was treated with a combination of controlled physiochemical stressors consisting of elevated temperature, a gas mixture of medical oxygen containing ozone, and UV light. The treated blood (150 microL) was injected in the gluteal muscle. Control animals received the same volume of untreated blood or physiological saline. Transient (45 or 60 minutes) left-renal ischemia was produced with simultaneous contralateral nephrectomy in treated and control spontaneously hypertensive rats (SHR). Young and old male and female rats were studied. Plasma creatinine, diuresis, and the survival rates of each group were compared. Renal apoptosis-necrosis was estimated by DNA laddering, histology, and in situ terminal deoxynucleotidyl transferase assay. mRNA levels of several regulators of apoptosis-regeneration were determined in control and postischemic kidneys by Northern blotting. RESULTS: IMT pretreatment of SHR significantly reduced renal I/R injury compared with equivalent placebo treatments consisting of untreated blood- or saline-injected SHR, as evidenced by a significant increase of the survival rate curves in young and old male SHR, which correlated with 24-hour postischemic diuresis. The increases in plasma creatinine following renal I/R were significantly lower in IMT-treated young male and old female SHR compared with saline or untreated blood-injected controls. Dilution analysis showed that the protective effect of treated blood was lost by dilution. Loss of epithelial cells was reduced in IMT-treated rats, with a significant decline in the peak of apoptosis 12 hours after acute ischemic renal injury. IMT did not modify the pattern of mRNA levels of several genes involved in the inflammation and regeneration processes. CONCLUSION: Our data demonstrate that IMT prevents the destruction of kidney tissue and the resulting animal death caused by renal I/R injury.

Synergistic interaction between enalapril, L-arginine and tetrahydrobiopterin in smooth muscle cell apoptosis and aortic remodeling induction in SHR.

Smooth muscle cell (SMC) apoptosis occurs at the onset of enalapril-induced regression of aortic hypertrophy in SHR. A potential mechanism is the correction of endothelial dysfunction (ED) leading to reduced production of reactive oxygen species and enhanced bioavailability of nitric oxide (NO), a potent apoptosis inducer. Stimulants of NO include the precursor L-arginine and the NO synthase cofactor tetrahydrobiopterin (BH(4)), which correct ED in several models. The objective was to examine the relationships between ED and the cell growth/death balance during vascular remodeling induced by enalapril in SHR. SHR, 10-week-old, received enalapril (ENA: 30 mg x kg(-1) x day(-1) p.o.) for 1 or 2 weeks, or a co-treatment of L-arginine (2.0 g x kg(-1) x day(-1) p.o.) and BH(4) (5.4 mg x kg(-1) x day(-1) i.p. twice daily) administered alone (group: LB) or in combination with enalapril (ENA+LB) for 1 week. Controls received vehicle. After 1 week, ED was completely corrected with LB but not affected significantly by ENA, whereas both treatments failed to induce SMC apoptosis or aortic remodeling. The correction of ED and the induction of SMC apoptosis (3.3-fold increase in TUNEL labeling) required 2 weeks of ENA treatment. The combination of LB with ENA for 1 week, however, was additive for the reduction of SMC proliferation, and synergistic for the induction of apoptosis and regression of vascular hypertrophy. These interactions were independent of blood pressure regulation. Our results suggest that the correction of ED is not sufficient to induce SMC apoptosis and vascular remodeling, although it facilitates these responses during enalapril treatment.

Kinin B2 receptor is not involved in enalapril-induced apoptosis and regression of hypertrophy in spontaneously hypertensive rat aorta: possible role of B1 receptor.

1. Treatment with enalapril induces smooth muscle cell apoptosis and regression of aortic hypertrophy in spontaneously hypertensive rats (SHRs), whereas combined blockade of angiotensin II AT(1) and AT(2) receptors does not. We postulated that vascular apoptosis with enalapril involves enhanced half-life of bradykinin (BK) and kinin B(2) receptor stimulation. 2. SHR, 11-weeks old, were treated for 4 weeks with enalapril (30 mg kg(-1) day(-1)), Hoe 140 (500 microg kg(-1) day(-1); B(2) receptor antagonist), alone or in combination. Controls received vehicle. 3. The half-life of hypotensive responses to intra-arterial bolus injections of BK were significantly increased in SHR anesthetized after 4 weeks of enalapril, an effect prevented by Hoe 140. The magnitude of BK-induced hypotension was significantly attenuated in all rats treated with Hoe 140. 4. As compared to placebo, enalapril treatment significantly reduced blood pressure (-34+/-2%), aortic hypertrophy (-20+/-3%), hyperplasia (-37+/-5%) and DNA synthesis (-61+/-8%), while it increased aortic DNA fragmentation by two-fold. Hoe 140 given alone or in combination with enalapril affected none of these parameters. 5. As a possible alternative mechanism, aortae isolated during the second week of enalapril treatment showed a transient upregulation of contractile responses to des-Arg(9)BK (EC(50)<1 nM), which were significantly reduced by [Leu(8)]des-Arg(9)BK (10 microM). Moreover, in vitro receptor autoradiography revealed an increase in expression of B(1) and B(2) receptor binding sites by 8-11 days of enalapril treatment. 6. Aortic apoptosis induction and hypertrophy regression with enalapril do not involve kinin B(2) receptors in SHR. Kinins acting via B(1) receptors remains a candidate mechanism.

Genetics of programmed cell death and proliferation.

Vascular remodeling and hypertrophy-hyperplasia of the heart and kidney are the hallmarks of hypertensions, being involved in the long-term maintenance of elevated blood pressure and the development of cardiovascular and renal hypertension. Both augmented proliferation and unscheduled programmed cell death (apoptosis) contribute to this phenomenon. In the present article, we summarize the results of these studies with an emphasis on the relative impact of genetic determinants and the environment.

Reduction of advanced-glycation end products levels and inhibition of RAGE signaling decreases rat vascular calcification induced by diabetes.

Advanced-glycation end products (AGEs) were recently implicated in vascular calcification, through a process mediated by RAGE (receptor for AGEs). Although a correlation between AGEs levels and vascular calcification was established, there is no evidence that reducing in vivo AGEs deposition or inhibiting AGEs-RAGE signaling pathways can decrease medial calcification. We evaluated the impact of inhibiting AGEs formation by pyridoxamine or elimination of AGEs by alagebrium on diabetic medial calcification. We also evaluated if the inhibition of AGEs-RAGE signaling pathways can prevent calcification. Rats were fed a high fat diet during 2 months before receiving a low dose of streptozotocin. Then, calcification was induced with warfarin. Pyridoxamine was administered at the beginning of warfarin treatment while alagebrium was administered 3 weeks after the beginning of warfarin treatment. Results demonstrate that AGEs inhibitors prevent the time-dependent accumulation of AGEs in femoral arteries of diabetic rats. This effect was accompanied by a reduced diabetes-accelerated calcification. Ex vivo experiments showed that N-methylpyridinium, an agonist of RAGE, induced calcification of diabetic femoral arteries, a process inhibited by antioxidants and different inhibitors of signaling pathways associated to RAGE activation. The physiological importance of oxidative stress was demonstrated by the reduction of femoral artery calcification in diabetic rats treated with apocynin, an inhibitor of reactive oxygen species production. We demonstrated that AGE inhibitors prevent or limit medial calcification. We also showed that diabetes-accelerated calcification is prevented by antioxidants. Thus, inhibiting the association of AGE-RAGE or the downstream signaling reduced medial calcification in diabetes.