Aorta Of Wistar Rats Research Articles

EFFECTS OF HIGH SALT DIET ON ARTERIAL REMODELLING AND THE INTERVENTION OF TELMISARTAN IN WISTAR RATS

ObjectivesTo investigate the effects of dietary salt on aorta and mesenteric artery remodelling in Wistar Rats and explore the possible mechanism of salt-induced arterial remodelling and AngiotensinII receptor blockers telmisartan...

Changes in the expression and function of beta‐adrenergic receptors subtypes In the rat aorta during development

During his life the rat shows changes in the relaxation response mediated by beta adrenergic receptors (βAR). In order to provide additional information on the topic, we studied the changes that may occur in the expression, localization and in vitro function of each subtype βAR in the thoracic aorta of male Wistar rats of two age groups: 3 weeks (prepuberty) and 9 weeks (postpuberty). The results indicate that, after the ninth week, the pretreatment with ICI 118.551 (β2 selective inhibitor, 10‐7 M) but not with CGP 12177 (selective inhibitor of β1, 10‐7 M) inhibited the relaxation response in all groups. However, the inhibition was greater when both antagonists were administered simultaneously. In addition, BRL 37344 (selective β3 agonist) caused relaxation only in prepuberty rats rings, which was blocked by SR 59230A (antagonist β3, 10‐6 M). Consistent with previous data, there was an increase in the expression of β1 and β2 receptors and decreased the β3 from the ninth week rats. Moreover, β1 and β2 subtypes are mainly located in smooth muscle, whereas β3 are located mainly in the endothelium. In conclusion, β1 receptors do not appear to participate functionally by themselves at any age, the β3 only involved in prepuberty rats and β2 are involved in all groups but their affinity is likely to change from the ninth week. Interestingly, the combination of β1 and β2 antagonists produced a synergistic effect which deserves further investigation.

Bee (Apis mellifera) Venom Produced Toxic Effects of Higher Amplitude in Rat Thoracic Aorta than in Skeletal Muscle—An Ultrastructural Study

In this study, changes produced in aorta and triceps surae muscle of Wistar rats as response to bee venom (BV) envenomation were analyzed by transmission electron microscopy and morphometry. A subchronic treatment of 30 days with daily doses of 700 μg BV/kg and an acute-lethal treatment with a single dose of 62 mg BV/kg were performed. The subchronic treatment resulted in endothelial cell retraction, a thicker subendothelial layer, and thinner elastic laminae and musculoelastic layers in aorta, and thicker endothelium and basal laminae in skeletal muscle. In both tissues polymorphous, swollen mitochondria with disrupted cristae were observed. The acute treatment produced extensive endothelial lesions, breakdown of the collagen layer and migration of muscle cells toward the intima in the aorta, and dilatation of endoplasmic reticulum in the skeletal muscle cells. Mitochondria were almost devoid of cristae or with few circular cristae in the smooth muscle cells while most of the mitochondria presented abnormal circular cristae in the skeletal muscle cells. Degenerative alterations in the aorta were of higher intensity in our experiments-both the intima and media strongly responded to BV, in contrast to those found at the level of the skeletal muscle cells where a moderate degenerative myopathy was recorded.

Development of a Growing Rat Model for the In Vivo Assessment of Engineered Aortic Conduits

Numerous limitations of aortic valve grafts currently used in pediatric patients cause the need for alternative prostheses. For the purpose of in vivo evaluation of novel engineered aortic conduit grafts, we aimed at downsizing a previously described model to create a growing rodent model. U-shaped aortic conduits were sutured to the infrarenal aorta of young Wistar rats (70-80 g, n = 10) in an end-to-side manner. Functional assessment was performed by Doppler sonography and high resolution rodent MRI. Histology and immunohistochemistry followed after 8 wk. Postoperative recovery rate was 80%. Conforming to clinical observations, postoperative MRI (d 5) and Doppler sonography (wk 8) revealed unimpaired conduit perfusion. Explanted implants were luminally completely covered by an endothelial cell layer with local hyperplasia and accumulation of α-smooth muscle actin (+) cells. Moreover microcalcification of the decellularized scaffolds was observed. Our downsized model of aortic conduit transplantation enables overall characterization with detailed analysis of maturation of engineered aortic grafts in a growing organism.

PDGF-β receptor and PKC have no effect on angiotensin II-induced JAK2 and STAT1 phosphorylation in vascular smooth muscle cells under high glucose condition

Background: The mechanisms responsible for the accelerated cardiovascular disease in diabetes, as well as the increased hypertrophic effects of angiotensin II (Ang II) under hyperglycemic condition, are not very clear. Evidences show that platelet-derived growth factor (PDGF) and protein kinase C (PKC) play a critical role in this effect. In our study, we examined the role of PKC and PDGF receptor on JAK2 and STAT1 phosphorylation under high glucose (HG) condition (25 mmol/L) in response to Ang II in cultured vascular smooth muscle cells (VSMC).Methods: VSMCs were isolated from the thoracic aorta of male Wistar rats and were cultured. Growth-arrested VSMCs were placed in either normal glucose (NG) or HG condition for 48 h and then VSMCs were stimulated with agonists and antagonists. The tyrosine phosphorylation of JAK2 or STAT were determined by immunoblotting using specific antibodies.Results: High glucose markedly increased the phosphorylation of tyrosine residues of JAK2 and serine residues of STAT 1 compared with cells cultured in NG (5.5 mmol/L) with and without Ang II stimulation. Experiments made with specific PDGF-β receptor inhibitor AG1295 and PKC inhibitor GF109203X showed that there were no changes in Ang II-stimulated JAK2 and STAT1 phosphorylation under NG and HG conditions compared with experiments without inhibitors.Conclusion: According to our findings, Ang II-stimulated JAK2 and STAT1 phosphorylation under either NG or HG condition do not proceed via a different pathway rather than PKC and PDGF-β receptor.

Platonin inhibited PDGF-BB-induced proliferation of rat vascular smooth muscle cells via JNK1/2-dependent signaling

To examine the inhibitory actions of the immunoregulator platonin against proliferation of rat vascular smooth muscle cells (VSMCs). VSMCs were prepared from the thoracic aortas of male Wistar rats. Cell proliferation was examined using MTT assays. Cell cycles were analyzed using flow cytometry. c-Jun N-terminal kinase (JNK)1/2, extracellular signal-regulated kinase (ERK)1/2, AKT, and c-Jun phosphorylation or p27 expression were detected using immunoblotting. Pretreatment with platonin (1-5 μmol/L) significantly suppressed VSMC proliferation stimulated by PDGF-BB (10 ng/mL) or 10% fetal bovine serum (FBS), and arrested cell cycle progression in the S and G(2)/M phases. The same concentrations of platonin significantly inhibited the phosphorylation of JNK1/2 but not ERK1/2 or AKT in VSMCs stimulated by PDGF-BB. Furthermore, platonin also attenuated c-Jun phosphorylation and markedly reversed the down-regulation of p27 expression after PDGF-BB stimulation. Platonin inhibited VSMC proliferation, possibly via inhibiting phosphorylation of JNK1/2 and c-Jun, and reversal of p27 down-regulation, thereby leading to cell cycle arrest at the S and G(2)/M phases. Thus, platonin may represent a novel approach for lowering the risk of abnormal VSMC proliferation and related vascular diseases.

Effects induced by inhibitors of the phosphatidylinositol 3‐kinase/Akt and nitric oxide synthase/guanylyl cyclase pathways on the isometric contraction in rat aorta: a comparative study

This work was aimed to determine whether isometric contraction in Wistar rat aorta is related to the phosphatidylinositol 3-kinase (PI3K)/Akt-dependent activation of endothelial nitric oxide synthase (eNOS). Basically, we hypothesized that additional increases in active tone occur after the pharmacological inhibition of a transduction pathway involved in NO synthesis or action. In intact aortic rings contracted with phenylephrine or high K(+), the cumulative administration of the PI3K inhibitor, LY294002, elicited significant decreases--but not supplementary increases--in tone. In endothelium-intact tissues, on the other hand, the Akt1/2 kinase inhibitor did not alter phenylephrine- and K(+)-induced isometric contractions. The PI3K inhibitor wortmannin (1 × 10(-7) m) produced a significant supplementary contraction only in endothelium-intact aortic rings precontracted with phenylephrine. Higher concentrations of this inhibitor produced relaxations of phenylephrine and high K(+)-constricted endothelium-intact and endothelium-denuded aortic rings. LY294002 and wortmannin did not cause any potentiating effect on phenylephrine- and angiotensin II-induced concentration-dependent contractile responses in endothelium-intact tissues. In intact aortic rings contracted with phenylephrine or high K(+), the addition of the NOS inhibitor, L-NAME, or the guanylyl cyclase inhibitor, ODQ, further augmented tone in a concentration-dependent manner, and these supplementary contractions were significantly reduced by endothelium removal. Taken together, our data suggest that the PI3K/Akt pathway is not counteracting aortic isometric contractions by activation of the eNOS. It appears, on the other hand, that the smooth muscle PI3K can stimulate contraction without activation of the protein kinase Akt in response to GPCR agonists and high K(+).

Regeneration of Three Layers Vascular Wall by using BMP2-Treated MSC Involving HIF-1α and Id1 Expressions Through JAK/STAT Pathways

Engineering living, multilayered blood vessels to form in vivo arteries is a promising alternative to peripheral artery bypass using acellular grafts restricted by thrombosis and occlusion at long term. Bone Morphogenetic Protein 2 (BMP2) is a growth factor determining in the early vascular embryonic development. The aim of the present study was evaluate the collaborative effect of recombinant human--BMP2 and Bone marrow--Mesenchymal stem cells (BM-MSCs) seeded on vascular patch to regenerate a vascular arterial wall in a rat model. BM-MSCs expressing green fluorescent protein (GFP) seeded on vascular patch were cultured in presence of recombinant human-BMP2 [100 ng/mL] during 1 week before their implantation on the abdominal aorta of Wistar rats. We observed after 2 weeks under physiological arterial flow a regeneration of a three layers adult-like arterial wall with a middle layer expressing smooth muscle proteins and a border layer expressing endothelial marker. In vitro study, using Matrigel assay and co-culture of BM-MSCs with endothelial cells demonstrated that rh-BMP2 promoted tube-like formation even at long term (90 days) allowing the organization of thick rails. We demonstrated using inhibitors and siRNAs that rh-BMP2 enhanced the expression of HIF-1α and Id1 through, at least in part, the stimulation of JAK2/STAT3/STAT5 signaling pathways. Rh-BMP2 by mimicking embryological conditions allowed vascular BM-MSCs differentiation.

The Role of Oxidative Stress in Acetylcholine-Induced Relaxation of Deendothelized Arteries

Nitric oxide (NO) produced by endothelium in response to vasorelaxants, such as acetylcholine, induces vasorelaxation of vascular smooth muscle cells (VSMC). It has been found that VSMC express NO-synthase, however, the principal question remained unanswered, if it is physiologically relevant. Because injury of endothelium triggers free-radical production which decreases NO availability we hypothesized that the destruction of arterial anatomical integrity by rubbing off endothelial layer made vessels insensitive to vasodilators as a consequence of oxidative stress. We examined acetylcholine-induced vasorelaxation in deendothelized thoracic aorta (TA), mesenteric artery (MA) and pulmonary artery (PA) of Wistar rats under protection against oxidative stress. Acetylcholine produced vasorelaxation in arteries with intact endothelium, whereas the relaxation in endothelium-denuded ringswas inhibited. Pretreatment of TA,MAandPAdenuded ringswith tempol, free-radical scavenger, improved relaxation to acetylcholine compared to untreated rings. The improved relaxation in all denuded rings was inhibited when ODQ, an inhibitor of guanylate cyclase, or L-NAME, an inhibitor of NOsynthase, were administered contemporary with tempol. Chemiluminiscence method revealed that endothelial denudation of TA and PA increased the production of superoxides. Immunohistochemical staining confirmed expression of NOS3-isoform in both intimal and medial cells in all arteries. Results revealed that deendothelized arteries under protection against oxidative stress exerted relaxation to acetylcholine which was mediated by NO and cGMP. The study suggests that VSMC can release NO in amounts sufficient to account for the vasorelaxation. This finding challenges the concept of the exclusive role of endothelial cells in the relaxation of VSMC.

Resistin contributes to neointimal formation via oxidative stress after vascular injury

Resistin may play a major potential role in vascular remodelling and may contribute to atherogenesis. However, the role of VSMC (vascular smooth muscle cell)-derived resistin in neointimal formation is not well understood. We hypothesize that endogenous resistin derived from VSMCs may contribute to neointimal formation after vascular injury. VSMCs from thoracic aorta of adult Wistar rats were cultured. The carotid artery from adult Wistar rats was injured by balloon catheter. Resistin significantly increased migration and proliferation of VSMCs. Resistin siRNA (small interfering RNA) and resistin antibody significantly inhibited migration and proliferation of VSMCs induced by conditioned medium from stretched VSMCs. Resistin protein and mRNA expression significantly increased at 14days after carotid injury. Resistin siRNA and NAC (N-acetylcysteine) significantly reduced resistin protein and mRNA expression induced by balloon injury. Carotid artery injury increased ROS (reactive oxygen species) production. Treatment with NAC and resistin siRNA decreased ROS production. The neointimal area was significantly increased after carotid injury and was significantly reduced by resistin siRNA and NAC. In conclusion, resistin increases migration and proliferation of VSMCs, and expression of resistin in carotid artery significantly increases after injury. Resistin siRNA attenuates neointimal formation after carotid injury partly through an antioxidative mechanism. Resistin may play a pivotal role in the pathogenesis of neointimal thickening after mechanical injury.

Endothelial regulation of eNOS, PAI-1 and t-PA by testosterone and dihydrotestosterone in vitro and in vivo

The aim of this study is the identification of direct endothelial regulation by the androgens testosterone (T) and dihydrotestosterone (DHT). We tested the effects of T and DHT on nitric oxide (NO) synthesis and on tissue plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) expression in human endothelial cells and in ovariectomized (OVX) rats. The results showed that at physiological concentrations T and DHT increase endothelial synthesis of NO. This depends on a rapid recruitment of the extracellular-related kinase (ERK) 1/2 and of the phosphatidylinositol 3-OH kinase (PI3K)/Akt cascades, resulting in endothelial nitric oxide synthase (eNOS) Ser(1177)-phosphorylation. In addition, a later increase of eNOS expression is found. With supra-physiological amounts of T or DHT the induction of NO synthesis is lost. A concentration-related increase of t-PA expression starting from physiological concentrations of T or DHT is found, whereas PAI-1 is augmented only with higher doses. Although DHT exerts these actions through androgen receptors (AR), T acts in part through aromatase-dependent conversion to 17β-estradiol. Ovariectomy is associated with significant changes in eNOS, t-PA and PAI-1 expression in the aorta of Wistar rats and T and DHT result in modifications on eNOS, PAI-1 and t-PA that are in line with the in vitro experiments. In conclusion, T and DHT act on endothelial cells through AR or via conversion to estradiol. Physiological, but not higher amounts are associated with enhanced NO synthesis and an increased t-PA/PAI-1 ratio. These findings are useful to understand the impact of androgens in ageing individuals.

ChemInform Abstract: NO-Donors. Part 3. Nitrooxyacylated Thiosalicylates and Salicylates - Synthesis and Biological Activities.

Organic nitrates release nitric oxide when incubated with thiosalicylic acid. S-Nitrooxyacylated esters and amides of thiosalicylic acid, together with the corresponding salicylates, were synthesized in order to perform a first in vitro evaluation of these new nitrate-thiol-hybrid prodrugs. These prodrugs might release NO in vivo after biotransformation without the use of endogenous reductives. None of these prodrugs released NO spontaneously when dissolved in buffer solution, but they did activate soluble guanylyl cyclase and induced vasodilatation of phenylephrine-pretreated male Wistar rat aorta in a potency range between that of isosorbiddinitrate and glycerole trinitrate.

Activation of Adventitial Fibroblasts in the Early Stage of the Aortic Transplant Vasculopathy in Rat

Transplant vasculopathy (TV) is the most significant obstacle to long-term success of organ transplantation. Increasing attention has been paid to the role of adventitia in vascular diseases. We evaluated the role of adventitial fibroblasts in the development of TV. Thoracic aortas from Sprague-Dawley (SD) rats transplanted into the abdominal aortas of Wistar rats worked as allografts, and isografts (SD to SD) were control. Grafts were removed on days 3, 7, and 14 for histologic, morphometric, and immunohistochemical detection of vimentin, alpha-smooth muscle actin, Ki-67, CD3, transforming growth factor-beta1 (TGF-beta1), monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinase-7 (MMP-7), and quantitative real-time reverse transcriptase polymerase chain reaction for TGF-beta1, MCP-1, MMP-7, tumor necrosis factor-alpha, and interleukin-1beta. In the allografts, neointima thickness and neointima/media thickness ratios were slightly increased at 7 days and significantly increased at 14 days. Immunostaining of vimentin and alpha-smooth muscle actin showed adventitial fibroblasts activation and differentiation into myofibroblasts. Ki-67-positive nuclei were observed in the adventitia 3 days after allografting and subsequently in the neointima. No more than 4% CD3-positive cells were found in adventitia in all the groups. Compared with isografts, TGF-beta1, MMP-7, and MCP-1 were expressed in the adventitia before neointima formation and were significantly increased in allografts at all time points. Tumor necrosis factor-alpha and interleukin-1beta were also significantly increased in adventitia in allografts. These results demonstrated that adventitial fibroblasts are activated and can produce cytokines and chemokines before the neointimal hyperplasia. They may exert a potential effect on the development of neointimal hyperplasia in TV.

Matrix metalloproteinases cleave the β2-adrenergic receptor in spontaneously hypertensive rats

We recently observed the enhanced serine and matrix metalloproteinase (MMP) activity in the spontaneously hypertensive rat (SHR) compared with its normotensive Wistar-Kyoto (WKY) rat and the cleavage of membrane receptors in the SHR by MMPs. We demonstrate in vivo that MMP-7 and MMP-9 injection leads to a vasoconstrictor response in microvessels of rats that is blocked by a specific MMP inhibitor (GM-6001, 1 microM). Multiple pathways may be responsible. Since the beta(2)-adrenergic receptor (beta(2)-AR) is susceptible to the action of endogenous MMPs, we hypothesize that MMPs in the plasma of SHRs are able to cleave the extracellular domain of the beta(2)-AR. SHR arterioles respond in an attenuated fashion to beta(2)-AR agonists and antagonists. Aorta and heart muscle of control Wistar rats were exposed for 24 h (37 degrees C) to fresh plasma of male Wistar and WKY rats and SHRs with and without doxycycline (30 microM) and EDTA (10 mM) to reduce MMP activity. The density of extracellular and intracellular domains of beta(2)-AR was determined by immunohistochemistry. The density of the extracellular domain of beta(2)-AR is reduced in aortic endothelial cells and cardiac microvessels of SHRs compared with that of WKY or Wistar rats. Treatment of the aorta and the heart of control Wistar rats with plasma from SHRs, but not from WKY rats, reduced the number of extracellular domains, but not intracellular domains, of beta(2)-AR in aortic endothelial cells and cardiac microvessels. MMP inhibitors (EDTA and doxycycline) prevented the cleavage of the extracellular domain. Thus MMPs may contribute to the reduced density of the extracellular domain of beta(2)-AR in blood vessels and to the increased arteriolar tone of SHRs compared with normotensive rats.

A novel solid state telemetry sensor for the chronic measurement of pressure in rats

Current telemetry devices for the measurement of pressure in rats utilise a catheter with a biocompatible gel and fluid which transmit the pressure waveform to a sensor. The reuse of gel/fluid based catheters is problematic with the need to regel and remove blood clots, furthermore the catheter is often damaged after repeated uses and requires refurbishment. These gel filled catheters have typically a very low frequency response; not allowing for performing meaningful dynamic analysis of cardiac pressure waveforms. These gel filled catheters also struggle with air bubbles during the regel operation which may also affect the signal integrity. We have developed a fully implantable telemeter incorporating a novel micro‐tip 2F catheter via a MEMs based sensor. The transmitter incorporates high frequency sampling of the pressure waveform and digital transmission to a remote receiver. The total implant weight is 14 gm and suitable for implantation in animals over 200 gm. Digital transmission of the signals permits co‐housing of animals without crosstalk. The ability to recharge the batteries inductively whilst monitoring provides the platform for long term and/or repeated use in animals. We report on the performance of telemeters when implanted in the aorta of Wistar rats for up to 1 month. The frequency response of the system was 500Hz allowing high fidelity measurement of arterial pressures. This high fidelity system for making direct measurements of internal pressures could provide a valuable research tool for expanding the scope of chronic studies while improving the ease of use of telemetry systems.

Increased migratory properties of aortal smooth muscle cells exposed to calcitriol in culture

The aim of the present study was to examine the effect of 1,25(OH) 2D 3 (calcitriol) on SMC (smooth muscle cell) migration, especially in the context to atherogenesis. SMCs were obtained from the aortas of newborn Wistar rats by enzymatic digestion. Different aspects of cell behavior during migration in culture were examined by phase contrast, fluorescence and electron microscopy (TEM, SEM) and supported by flow cytometric and biochemical analyses. Morphological studies revealed that supra-physiological (1–100 nmol/l) concentrations of calcitriol inhibit SMC differentiation, therefore these cells display several hallmarks of the synthetic state. Dynamic changes in actin cytoskeleton organization were a critical event in SMC shape, adhesion and spreading. Calcitriol diminished stress fibers assembly and focal adhesions formation. Reduced expression of β 1-integrin receptors on SMC surface after exposition to calcitriol coincided with increased proliferative and migratory activities of these cells. Moreover, after calcitriol stimulation, the ability of SMCs to the production of proinflamatory cytokines IFN-γ, TNF-α and IL-6 was inhibited. The results from these comparative investigations indicate that 1,25(OH) 2D 3 inhibit differentiation and facilitate SMC migration in culture. It has been also suggested that such responses of SMCs to calcitriol play a beneficial role in fibrous cap formation during atherosclerotic process.

Mechanisms underlying the biphasic effect of vitamin K1 (phylloquinone) on arterial blood pressure

Phylloquinone (vitamin K(1), VK(1)) is widely used therapeutically and intravenous administration of this quinone can induce hypotension. We aimed to investigate the mechanisms underlying the effects induced by VK(1) on arterial blood pressure. With this purpose a catheter was inserted into the abdominal aorta of male Wistar rats for blood pressure and heart rate recording. Bolus intravenous injection of VK(1) (0.5-20 mgkg(-1)) produced a transient increase in blood pressure followed by a fall. Both the pressor and depressor response induced by VK(1) were dose-dependent. On the other hand, intravenous injection of VK(1) did not alter heart rate. The nitric oxide synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 10 and 20 mgkg(-1)) reduced both the increase and decrease in blood pressure induced by VK(1) (5 mgkg(-1)). On the other hand, indometacin (10 mg kg(-1)), a non-selective cyclooxygenase inhibitor, did not alter the increase in mean arterial pressure (MAP) induced by VK(1). However, VK(1)-induced fall in MAP was significantly attenuated by indometacin. We concluded that VK(1) induces a dose-dependent effect on blood pressure that consists of an acute increase followed by a more sustained decrease in MAP. The hypotension induced by VK(1) involves the activation of the nitric oxide (NO) pathway and the release of vasodilator prostanoid(s).

Regulation of resistin by cyclic mechanical stretch in cultured rat vascular smooth muscle cells

Resistin has a potential role in atherosclerosis; however, the molecular mechanism underlying the increase in resistin expression in atherosclerosis remains unclear. As mechanical stretch plays an important role in atherosclerosis, in the present study we sought to investigate the cellular and molecular mechanisms underlying the regulation of resistin by cyclic mechanical stretch in VSMCs (vascular smooth muscle cells). VSMCs from thoracic aorta of adult Wistar rats were cultured and subjected to cyclic stretch. Cyclic mechanical stretch significantly increased resistin protein and mRNA expression as compared with control cells without stretch. The specific p38 MAPK (mitogen-activated protein kinase) inhibitor SB203580, the antioxidant N-acetylcysteine and p38 MAPK siRNA (small interfering RNA) attenuated the induction of resistin protein by cyclic stretch. Cyclic stretch significantly increased the phosphorylation of p38 MAPK, whereas pre-treatment with SB203580 and N-acetylcysteine significantly inhibited this effect. Cyclic stretch significantly increased ROS (reactive oxygen species) production, and pre-treatment with N-acetylcysteine significantly inhibited stretch-induced ROS production. Cyclic stretch also increased STAT3 (signal transducer and activator of transcription 3)-binding activity and resistin promoter activity, and resistin promoter activity was abolished when STAT3 in the promoter area was mutated. Pre-treatment with SB203580 and N-acetylcysteine significantly attenuated resistin promoter activity induced by cyclic stretch. Cyclic stretch increased the secretion of AngII (angiotensin II) and resistin from cultured VSMCs. Exogenous AngII increased resistin expression, and AngII receptor inhibition attenuated this effect. In conclusion, cyclic mechanical stretch increases resistin expression in cultured rat VSMCs. Stretch-induced resistin expression is mediated through ROS, and the p38 MAPK and STAT3 pathways. Therefore resistin induced by cyclic stretch may contribute to the pathogenesis of atherosclerosis under haemodynamic overload.Key

Dextran sulfate modulates MAP kinase signaling and reduces endothelial injury in a rat aortic clamping model

Mitogen-activated protein kinases (MAPKs), including JNK, p38, and ERK1/2, noticeably influence ischemia/reperfusion injury (IRI). The complement inhibitor dextran sulfate (DXS) associates with damaged endothelium denudated of its heparan sulfate proteoglycan (HSPG) layer. Other glycosaminoglycan analogs are known to influence MAPK signaling. Hypothetically therefore, targeted intravascular cytoprotection by DXS may function in part through influencing MAPK activation to reduce IRI-induced damage of the vasculature. IRI of the infrarenal aorta of male Wistar rats was induced by 90 minutes clamping followed by 120 minutes reperfusion. DXS (5 mg/mL) or physiologic saline (NaCl controls) was infused locally into the ischemic aortic segment immediately prior to reperfusion. Ninety minutes ischemia-only and heparinase infusion (maximal damage) experiments, as well as native rat aorta, served as controls. Aortas were excised following termination of the experiments for further analysis. DXS significantly inhibited IRI-induced JNK and ERK1/2 activation (P = .043; P =.005) without influencing the p38 pathway (P =.110). Reduced aortic injury, with significant inhibition of apoptosis (P = .032 for DXS vs NaCl), correlated with decreased nuclear factor kappaB translocation within the aortic wall. DXS treatment clearly reduced C1q, C4b/c, C3b/c, and C9 complement deposition, whilst preserving endothelial cell integrity and reducing reperfusion-induced HSPG shedding. Protection was associated with binding of fluorescein labeled DXS to ischemically damaged tissue. Local application of DXS into ischemic vasculature immediately prior to reperfusion reduces complement deposition and preserves endothelial integrity, partially through modulating activation of MAPKs and may offer a new approach to tackle IRI in vascular surgical procedures. The purpose of the present study was to determine the role of dextran sulfate (DXS), a glycosaminoglycan analog and complement inhibitor, in modulating intracellular MAPK signaling pathways, reducing complement activation and ultimately attenuating ischemia/reperfusion injury (IRI) in a rat aortic-clamping model, in part a surrogate model to study the microvasculature. The study shows a role for DXS in ameliorating endothelial injury by reducing IRI-mediated damage and intravascular, local inflammation in the affected aortic segment. DXS may be envisaged as an endothelial protectant in vascular injury, such as occurs during vascular surgical procedures.

Endothelial dysfunction in the early- and late-stage type-2 diabetic Goto-Kakizaki rat aorta

As there are increasing evidences that human diabetes induces cardiovascular dysfunction, we investigated the type-2 diabetes-induced endothelial dysfunction in the early and late-stage Goto-Kakizaki (GK) rat aorta. We performed organ bath studies, and examined the changes in expression levels of muscarinic M(3) receptor, endothelial, inducible, and neuronal nitric oxide synthase (eNOS, iNOS, and nNOS, respectively) mRNAs in the rat aorta utilizing real-time polymerase chain reaction in 12-week-old and 70-week-old GK rats as well as in age-matched Wistar rats. In the 12-week-old GK rat aorta, a significant increase in norepinephrine-induced contraction and a significant decrease in acetylcholine-induced relaxation as well as significant increases in expression levels of muscarinic M(3) receptor and eNOS and a significant decrease in nNOS mRNAs were observed compared to age-matched controls. In the older GK rat aorta, significant decreases in acetylcholine- and nitroglycerine-induced relaxations as well as significant decreases in the expression levels of muscarinic M(3) receptor, eNOS, iNOS, and nNOS mRNAs were observed compared to those in the younger GK rats. In contrast, although significant decreases in acetylcholine and nitroglycerine-induced relaxations were observed, the expression levels of muscarinic M(3) receptor, eNOS, iNOS, and nNOS mRNAs in the older Wistar rats aorta were unchanged, increased, increased and decreased, respectively, compared to the younger Wistar rat aorta. These results indicate that endothelial dysfunction in the rat aorta progresses with age and development of diabetes condition, and that decreased relaxations in the late-stage rat aorta may be due to these alterations.