Fluoxetine effect on aortic nitric oxide-dependent vasorelaxation in the unpredictable chronic mild stress model of depression in mice.

OBJECTIVE: Major depression is an independent risk factor for the development of cardiovascular diseases. However, the exact mechanism by which depression may induce cardiovascular events is unclear. Endothelial dysfunction has been reported as a possible link between depression and subsequent cardiovascular events as described in depressed subjects. The purpose of this study was to investigate endothelial dysfunction and atherosclerosis formation in the aorta of mice exposed to the unpredictable chronic mild stress (UCMS) procedure. METHODS: BALB/c mice were exposed to two 7-week UCMS procedures separated by 6 weeks. Treatments (fluoxetine 10 mg/kg; NaCl 0.9%) started at the third week until the end of the seventh week of each procedure. Endothelial function was evaluated by in vitro assessment of acetylcholine-induced vasorelaxation in aortic rings. By using specific inhibitors for nitric oxide (NO)- and prostacyclin-dependent relaxation, we assessed the part played by NO, prostacyclin, and endothelium-derived hyperpolarizing factor (EDHF)-like mediators in endothelium-dependent relaxation. Atherosclerosis was evaluated by histological examination. RESULTS: Depression-like behavior was increased in the UCMS versus unstressed group and was reversed by chronic fluoxetine treatment. Vascular reactivity study indicated that UCMS induced a decrease in the NO-dependent relaxation that was partially compensated by an EDHF-like dependent relaxation. Because fluoxetine per se increased the NO-dependent relaxation, fluoxetine was able to reverse UCMS effect on the NO component and abolished the EDHF-like component. Atherosclerotic lesion was found in aorta of UCMS and nonstressed animals. CONCLUSIONS: As an independent risk factor, UCMS reproduced the endothelial alterations observed in depression but was not sufficient to provoke morphological alterations.

Investigation of the molecular mechanisms preceding PDE4 inhibitor-induced vasculopathy in rats: tissue inhibitor of metalloproteinase 1, a potential predictive biomarker.

Phosphodiesterase (PDE) 4 inhibitors are a class of drugs that can provide novel therapies for asthma and chronic obstructive pulmonary disease. Their development is frequently hampered by the induction of vascular toxicity in rat mesenteric tissue during preclinical studies. Whereas these vascular lesions in rats have been well characterized histologically, little is known about their pathogenesis and in turn, sensitive and specific biomarkers for preclinical and clinical monitoring do not exist. In order to investigate the early molecular mechanisms underlying vascular injury, time-course studies were performed by treating rats for 2-24 h with high doses of the PDE4 inhibitor CI-1044. Transcriptomics analyses in mesenteric tissue were performed using oligonucleotide microarray and real-time RT-PCR technologies and compared to histopathological observations. In addition, protein measurements were performed in serum samples to identify soluble biomarkers of vascular injury. Our results indicate that molecular alterations preceded the histological observations of inflammatory and necrotic lesions in mesenteric arteries. Some gene expression changes suggest that the development of the lesions could follow a primary modulation of the vascular tone in response to the pharmacological effect of the compound. Activation of genes coding for pro- and antioxidant enzymes, cytokines, adhesion molecules, and tissue inhibitor of metalloproteinase 1 (TIMP-1) indicates that biomechanical stimuli may contribute to vascular oxidant stress, inflammation, and tissue remodeling. TIMP-1 appeared to be an early and sensitive predictive biomarker of the inflammatory and the tissue remodeling components of PDE4 inhibitor-induced vascular injury.

Contractile and relaxant properties of rat-isolated pulmonary veins related to localization and histology.

The aim of this study was to investigate the in vitro vasomotor properties of rat extra-and intralobar pulmonary veins (PVs) related to their localization and to assess the modulatory role of endothelium on these properties. Segments from PVs were mounted in small vessel myograph and stretched at various diameters (D(10), D(20), D(30)) corresponding to intraluminal pressures of 10, 20 or 30 mmHg. At D(10) or D(20), contractile responses to phenylephrine, U46619 and angiotensin II of distal intralobar part of PVs were smaller compared with those of proximal extralobar part, but no longer different when distal part was stretched at D(30). When submitted to an NO donor, sodium nitroprusside, distal part of PV relaxed more strongly when stretched at D(30) compared with D(10). Acetylcholine and bradykinin were devoid of relaxing effect on distal parts stretched at D(10), but in contrast to acetylcholine, bradykinin slightly relaxed preparations stretched at D(30). Isoprenaline strongly relaxed PVs ( approximately 80% of initial precontraction), with the distal part exhibiting a higher sensitivity to the agonist compared with the proximal part. This relaxation was also observed with salbutamol and suppressed with ICI 118551, which is in favour of the involvement of beta(2)-adrenoceptors in this effect. Preincubation of the preparations with N(G)-nitro-l-arginine methyl ester (10(-4) m) and indomethacin (10(-5) m) did not modify the contractile responses to U46619, nor the relaxing response to isoprenaline, which support that endothelium does not appear to play a significant modulatory role in these responses. Histological and electron microscopical examinations of proximal and distal sections of the same vein show that the layers of smooth muscle cells and cardiomyocytes were thicker in the proximal compared with the distal part. This study shows that, because of morphological heterogeneity of the PVs, the site of dissection and the initial condition of tension can play a significant role upon the sensitivity and the magnitude of the responses to both contractile and relaxing agonists.

Vasorelaxant effects of grape polyphenols in rat isolated aorta. Possible involvement of a purinergic pathway.

The purpose of this study was to investigate the mechanism of the vascular relaxation produced by polyphenolic substances from red wine, with a particular focus on the possible involvement of purinoceptors. With this aim, relaxing responses induced by procyanidin from grape seeds (GSP), anthocyanins, catechin and epicatechin were assessed in rat isolated aortic rings left intact (+E) or endothelium-denuded (-E). In preparations precontracted with noradrenaline, incubation with NG-nitro-L-arginine methyl ester (100 microM, 30 min) fully inhibited the GSP-induced relaxations. Concentration-effect curves to these substances (from 10(-7) to 10(-1) g/L) were determined in depolarized (60 mM KCl) preparations in control condition, after incubation with reactive blue 2 (an antagonist of P2Y purinoceptors, 30 microM), with apyrase (an enzyme which hydrolyses ATP and ADP, 0.8 U/mL) or with alpha,beta-methylene ATP (an inhibitor of ecto ATPases, 10 microM). In (+E) rings, relaxations (expressed as percentage of initial contraction) were 41 +/- 2 and 37 +/- 3 for GSP and anthocyanins, respectively. Only modest relaxations (ca. 10%) were observed in (-E) rings, as it was the case for catechin and epicatechin in (+/- E) rings. Reactive blue 2 or apyrase inhibited the GSP- and anthocyanin-induced relaxations in (+E) rings, while alpha,beta-methylene ATP shifted to the left the relaxation curves obtained with GSP. These data confirm that modest relaxations observed with catechin and epicatechin are not endothelium-dependent but that GSP and anthocyanins induce a relaxing effect, which is related to the integrity of the endothelium and the synthesis and release of nitric oxide (NO). Furthermore, the inhibition by apyrase and the increase by ecto-ATPase inhibition of the GSP- and anthocyanin-induced relaxation suggest that these substances could act via an initial release of nucleotides, which in turn could activate P2Y1 and/or P2Y2 purinoceptors of endothelial cells, trigger the synthesis and release of NO and then lead to relaxation.

Altered aortic vascular reactivity in the unpredictable chronic mild stress model of depression in mice: UCMS causes relaxation impairment to ACh.

Major depression is an independent risk factor for the development of cardiovascular disease. This impact of depression on vascular function seems to be mediated by the endothelial dysfunction, defined as an impairment of endothelium-dependent vasorelaxation, which represents a reliable predictor of atherosclerosis and has been regularly found to be associated with depression. This study aimed at investigating aortic vascular reactivity in mice submitted to the unpredictable chronic mild stress (UCMS) procedure, a reliable model of depression. The results confirm the effectiveness of the UCMS procedure to induce neuroendocrine, physical and behavioral depression-like alterations as well as a significant decrease of acetylcholine-induced vasorelaxation without any effect on phenylephrine-induced vasoconstriction. In this study, we reveal an altered vascular reactivity in an animal model of depression, demonstrating an endothelial dysfunction reminiscent to the one found in depressed patients.

An increased regional blood flow precedes mesenteric inflammation in rats treated by a phosphodiesterase 4 inhibitor.

The study was undertaken to assess the hemodynamic effects induced by a single dose of the phosphodiesterase 4 (PDE4) inhibitor, CI-1044, which is known to cause mesenteric vascular alterations in rats. In the present study, an administration of 160 mg/kg of CI-1044 caused perivascular and interstitial inflammation, with infiltrates of admixed neutrophils and macrophages but without evidence of vascular necrosis (ileum, 15/20 rats; duodenum + jejunum, 7/20 rats). Four hours after administration, blood pressure was decreased (- 13%). A fluorescent microsphere technique demonstrated that, in these conditions, cardiac output was doubled (+ 100%) and total peripheral resistance was decreased (- 54%). The largest increases in blood flow were measured in the duodenum (+ 101%), in the jejunum (+ 110%), and in the ileum (+ 192%). Therefore, the mesentery was the most sensitive organ affected by the drug and, within this area, parts with the highest incidence of vascular alteration were those which had shown the highest increase in flow. In addition, isolated precontracted mesenteric resistance arteries dissected from untreated animals were fully relaxed when incubated with increasing concentrations of CI-1044 up to 2.5 x 10(-5)M. At this latter concentration, contractile abilities and sensitivities to the physiological agonist noradrenaline (NA) and to the thromboxane analogue U46619 were significantly attenuated (- 28 and - 27%, respectively). This effect could lead to a decreased response to NA and possibly to other agonists in vivo consistent with the vasodilation observed with the microsphere technique. These data provide evidence that the PDE4 inhibitor CI-1044 induces changes of vascular tone that could lead to histological alterations in the mesenteric area.

Altered gene expression in rat mesenteric tissue following in vivo exposure to a phosphodiesterase 4 inhibitor.

Vascular injury is a relatively common finding during the pre-clinical toxicity testing of drugs. The mechanisms of the injury are poorly understood and in turn, sensitive and specific biomarkers for pre-clinical and clinical monitoring do not exist. The present study was undertaken to investigate the molecular mechanisms of drug-induced vascular injury in mesenteric tissue of rats treated with the selective phosphodiesterase 4 (PDE4) inhibitor CI-1044. In a time-course study, male Sprague Dawley rats were given daily doses of 40 or 80 mg/kg for 1, 2 or 3 successive days and were euthanized the following day. Gene expression profiles in mesenteric tissue were determined using Affymetrix RG_U34A microarrays and fibrinogen and cytokine measurements were performed in blood samples. Hierarchical clustering analysis produced a clear pattern separation of the animals with inflammation, animal with inflammation and necrosis and animals without any lesion. Genes associated with inflammation, procoagulation, extracellular matrix remodeling were up-regulated. An altered expression of genes involved in vascular tone regulation, lipid and glucose metabolism was also observed. Selected genes expression changes were confirmed by TaqMan real-time RT-PCR. The inflammatory process was also detected in the bloodstream at the protein level since fibrinogen, IL6 and IL1beta concentrations were increased in treated animals. Overall, the present study reveals several molecular changes supporting the hypothesis by which PDE4 inhibitor-induced vascular lesions in rats are triggered by an inflammatory mechanism and/or a vascular tone dysregulation.