Rat Smooth Muscle Research Articles

Tissue Engineering of Ureteral Grafts: Preparation of Biocompatible Crosslinked Ureteral Scaffolds of Porcine Origin.

The surgical reconstruction of ureteric defects is often associated with post-operative complications and requires additional medical care. Decellularized ureters originating from porcine donors could represent an alternative therapy. Our aim was to investigate the possibility of manufacturing decellularized ureters, the characteristics of the extracellular matrix (ECM) and the biocompatibility of these grafts in vitro/in vivo after treatment with different crosslinking agents. To achieve these goals, native ureters were obtained from pigs and were decellularized. The success of decellularization and the ECM composition were characterized by (immuno)histological staining methods and a DNA-assay. In vitro: scaffolds were crosslinked either with carbodiimide (CDI), genipin (GP), glutaraldehyde, left chemically untreated or were lyophilized. Scaffolds in each group were reseeded with Caco2, LS48, 3T3 cells, or native rat smooth muscle cells (SMC). After 2 weeks, the number of ingrown cells was quantified. In vivo: crosslinked scaffolds were implanted subcutaneously into rats and the type of infiltrating cells were determined after 1, 9, and 30 days. After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced. In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds. In vivo: at day 30, implants were predominantly infiltrated by fibroblasts and M2 anti-inflammatory macrophages. A maximum of MMP3 was observed in the CDI group at day 30. TIMP1 was below the detection limit. In this study, we demonstrated the potential of decellularization to create biocompatible porcine ureteric grafts, whereas a CDI-crosslink may facilitate the remodeling process. The use of decellularized ureteric grafts may represent a novel therapeutic method in reconstruction of ureteric defects.

Class IA Phosphatidylinositol 3-Kinase Isoform p110α Mediates Vascular Remodeling.

Neointima formation after vascular injury remains a significant problem in clinical cardiology, and current preventive strategies are suboptimal. Phosphatidylinositol 3'-kinase is a central downstream mediator of growth factor signaling, but the role of phosphatidylinositol 3'-kinase isoforms in vascular remodeling remains elusive. We sought to systematically characterize the precise role of catalytic class IA phosphatidylinositol 3'-kinase isoforms (p110α, p110β, p110δ), which signal downstream of receptor tyrosine kinases, for vascular remodeling in vivo. Western blot analyses revealed that all 3 isoforms are abundantly expressed in smooth muscle cells. To analyze their significance for receptor tyrosine kinases-dependent cellular responses, we used targeted gene knockdown and isoform-specific small molecule inhibitors of p110α (PIK-75), p110β (TGX-221), and p110δ (IC-87114), respectively. We identified p110α to be crucial for receptor tyrosine kinases signaling, thus affecting proliferation, migration, and survival of rat, murine, and human smooth muscle cells, whereas p110β and p110δ activities were dispensable. Surprisingly, p110δ exerted noncatalytic functions in smooth muscle cell proliferation, but had no effect on migration. Based on these results, we generated a mouse model of smooth muscle cell-specific p110α deficiency (sm-p110α(-/-)). Targeted deletion of p110α in sm-p110α(-/-) mice blunted growth factor-induced cellular responses and abolished neointima formation after balloon injury of the carotid artery in mice. In contrast, p110δ deficiency did not affect vascular remodeling in vivo. Receptor tyrosine kinases-induced phosphatidylinositol 3'-kinase signaling via the p110α isoform plays a central role for vascular remodeling in vivo. Thus, p110α represents a selective target for the prevention of neointima formation after vascular injury, whereas p110β and p110δ expression and activity do not play a significant role.

Propargylglycine Restores Endothelium-Dependent Relaxation of Aortic Smooth Muscle in Old Rats

Endothelium-dependent relaxation of the vascular smooth muscles (SM) is responsible for the development of basic cardiovascular reactions, and its violation is the basis of aging-associated cardiovascular diseases. We studied effects of inhibiting cystathionine-γ-lyase (CSE), the enzyme for de novo synthesis of hydrogen sulfide, on the endothelium-dependent relaxation of thoracic aorta SM from old rats. It has been shown that an inhibition of CSE by propargylglycine (PAG) results in restoration of decreased ACh-induced relaxation of aortic SM in old rats. That effect of PAG was removed by blocking nitric oxide (NO) synthesis in the endothelial cells. Age-related changes in the levels of H2S2 NO2−and enzyme activity of the constitutive synthesis of NO (cNOS) in the heart were determined. It has been demonstrated that introduced PAG elevates decreased levels of H2S2 NO2−and stimulates suppressed activity of cNOS in old rats. These results suggest that PAG activates alternative ways of H2S synthesis and stimulates the constitutive synthesis of NO to restore endothelial function in old rats.

Vasorelaxant and Antispasmodic Effects of Some Polyphenols and Glycoside in Rat Smooth Muscles

Objective: The present study was undertaken to investigate the effect of Luteolin, chlorogenic acid, amygdalin and tannic acid on vascular smooth muscle contractility. Materials and Methods: The effect of different concentrations of some polyphenols and glycosides on contractile responses of isolated aorta, ileum and trachea to Acetylcholine, KCl and phenylephrine (PE) were evaluated. Results: The results of the current study indicated that luteolin and chlorogenic acid has potent relaxant effects on KCl and PE-induced contractions in rat’s aortic ring. Amygdalin and tannic acid reduced ACh-induced ileum contractions significantly. Conclusions: The inhibitory effect of luteolin and chlorogenic acid on the contraction induced by PE and KCl may be due to their anti-adrenergic and anti-hyperpolarizing effect.

ASSA14-03-37 Study of effects of simvastatin on proliferation, migration and adhension of rat smooth muscle progenitor cells

<h3>Background</h3> To investigate the effects of simvastatin on differentiation and proliferation of rat smooth muscle progenitor cells (SPCs) and identification of compounds that inhibit SPCs differentiation and proliferation for substantial clinical usefulness. <h3>Methods</h3> Total mononuclear cells (MNCs) were isolated from marrow of rats by Ficoll density gradient centrifugation method, and then plated on fibronectin-coated culture dishes. Fresh isolated MNCs were treated with simvastatin (0.01∼10 μmol/l) or vehicle control for 8 d. SPCs were characterised as adherent cells positive for α-SMA by indirect immunofluorescent staining. After 8 days cultured, attached cells were treated with simvastatin (0.01∼10 μmol/l) or vehicle control for 24 h. The proliferation and migration of SPCs were assayed with ³H-TdR incorporation and modified Boyden chamber assay respectively. SPCs adhesion assay was performed by replating those on fibronectin-coated dishes and counting the adherent cells. <h3>Results</h3> Simvastatin potently inhibited SPCs outgrowth. The number of SPCs at 8 days was dramatically decreased by simvatatin. At a concentration as low as 0.01 μmol/l, simvastatin significantly reduced 7.5 ± 5.4% of SPCs (0.01 μmol/l simvastatin vs control: 79 ± 5 vs 85 ± 4, n = 5, p &lt; 0.05). Simvastatin also inhibited SPCs proliferation in a dose-dependent manner, simvastatin significantly reduced 5.8 ± 3.1% of SPCs at a concentration as low as 0.01 μmol/l for 24 h (0.01 μmol/l simvastatin vs control: 3833 ± 126 vs 4070 ± 184, n = 5, p &lt; 0.05). In addition, sirolims also inhibited SPCs migratory and adhesive capacity in a concentration-dependent manner. <h3>Conclusion</h3> Simvastatin could inhibit the differentiation, proliferation and migration of rat smooth muscle progenitor cells.

Does hypercholesterolemia affect the relaxation of the detrusor smooth muscle in rats? In vitro and in vivo studies.

To evaluate the effects of hypercholesterolemia on the relaxation function of the urinary bladder, we examined the physiological mechanisms involved in the isoproterenol-induced relaxation in isolated detrusor strips in vitro and voiding behavior in vivo in rats. Adult male Sprague-Dawley rats were fed standard (control, N = 16) or 4% cholesterol diet (hypercholesterolemia, N = 17) for 4weeks. Concentration-response curves for isoproterenol-induced relaxations in carbachol-precontracted detrusor muscle strips were recorded. The contributions of β2- and β3-adrenoceptors and ATP-dependent and Ca(2+)-dependent potassium channels to the relaxation response were investigated by using selective adrenergic agonists salbutamol and BRL 37344 and specific potassium channel inhibitors glibenclamide and charybdotoxin, respectively. Cystometrography was performed to assess bladder function. Hypercholesterolemic rats had higher serum cholesterol and low- and high-density lipoprotein levels than the controls with no sign of atherosclerosis. Isoproterenol-induced relaxation was significantly enhanced in the hypercholesterolemia group. Preincubation with the M2 receptor antagonist attenuated the relaxation response in both groups. The relaxation responses to isoproterenol and salbutamol were similar in both groups, while BRL 37344 appeared to produce a greater relaxant effect in the hypercholesterolemic rats. Also, the inhibitory effects of potassium channel inhibitors on relaxation responses were comparable among the groups. The cystometric findings revealed that threshold and basal pressure values were higher in the hypercholesterolemia group compared with controls. We showed that hypercholesterolemia leads to greater relaxation responses to isoproterenol, appears to impair the braking function of M2 cholinergic receptors on adrenoceptor-induced relaxations in the isolated detrusor muscle, and affects the voiding function in rats.

The interleukin-1β modulator gevokizumab reduces neointimal proliferation and improves reendothelialization in a rat carotid denudation model

ObjectiveExcessive neointima formation often occurs after arterial injury. Interleukin-1β (IL-1β) is a potent pleiotropic cytokine that has been shown to regulate neointimal proliferation. We investigated the effects of the IL-1β modulator gevokizumab in a rat carotid denudation model. MethodsSprague–Dawley rats were subjected to balloon denudation of the right carotid artery and were then randomized to receive a single subcutaneous infusion immediately after balloon injury of saline (control group, n = 13) or gevokizumab (gevokizumab groups, n = 15 in each group: 1, 10 and 50 mg/kg). We evaluated the treatment effects on carotid intima-media thickness (IMT) using ultrasonography, on endothelial regrowth using Evans Blue staining and on inflammatory response using histology. We also assessed the effects of IL-1β and gevokizumab on human umbilical vein endothelial cells (HUVEC) and rat smooth muscle cells. ResultsWe found that carotid IMT, in the proximal part of the denuded artery at day 28, was decreased by gevokizumab 1 mg/kg compared with controls. Neointima area and the intima/media area ratio were both reduced in the gevokizumab 1 mg/kg-treated group. Gevokizumab at the 1 mg/kg dose also improved endothelial regrowth. No effect was observed with gevokizumab 10 or 50 mg/kg. Gevokizumab also decreased the inflammatory effect of IL-1β in in vitro cell experiments and protected HUVECs from IL-1β's deleterious effects on cell migration, apoptosis and proliferation. ConclusionA single administration of gevokizumab 1 mg/kg improves endothelial regrowth and reduces neointima formation in rats following carotid denudation, at least in part through its beneficial effects on endothelial cells.

Nitro-oleic acid decreases transcription of the angiotensin II type I receptor gene in aortic smooth muscle cells

Nitroalkene derivatives of nitro-oleic acid (OA-NO2) regulate pluripotent cell signaling in vivo under physiological and pathological conditions. Angiotensin II type 1 receptor (AT1R) plays an important role in the cardiovascular system. In this study, OA-NO2 reduced the AT1R mRNA level, specifically in primary smooth muscle cells, showing a 70% reduction in rat smooth muscle cells (RASMCs) and a 50% reduction in pig smooth muscle cells (PASMCs). These effects were not observed in CHO cells, which highly express AT1R. The AT1R mRNA decay rate was unchanged after OA-NO2 compared with OA treatment. Nitric oxide (NO) and peroxisome proliferator-activated receptor gamma (PPARγ) did not alter the reduced effects of OA-NO2 on the AT1R mRNA level in SMCs. However, Sp1-mediated activation of the AT1R promoter was reduced in response to OA-NO2 in RASMCs but not 293T cells. In addition, the nuclear factor-kappa B (NF-κB) pathway was involved in the OA-NO2-mediated downregulation of AT1R transcription in SMCs. Taken together, our results demonstrate that OA-NO2 specifically inhibits AT1R mRNA expression in primary smooth muscle cells via the NF-κB pathway.

Synthetic polyamine BPA-C8 inhibits TGF-β1-mediated conversion of human dermal fibroblast to myofibroblasts and establishment of galectin-1-rich extracellular matrix in vitro.

Cancer-associated fibroblasts (CAFs) play a role in the progression of malignant tumors. They are formed by conversion of fibroblasts to smooth muscle α-actin-positive (SMA-positive) myofibroblasts. Polyamines are known to change the arrangement of the actin cytoskeleton by binding to the anionic actin. We tested the effect of the synthetic polyamine BPA-C8 on the transition of human dermal fibroblasts to myofibroblasts induced either by TGF-β1 alone or by TGF-β1 together with adhesion/growth-regulatory galectin-1. Pre-existing CAFs, myofibroblasts from pancreatitis, and rat smooth muscle cells were also exposed to BPA-C8. BPA-C8 impaired myofibroblast formation from activated fibroblasts, but it had no effect on cells already expressing SMA. BPA-C8 also reduced the occurrence of an extracellular matrix around the activated fibroblasts. The reported data thus extend current insights into polyamine activity, adding interference with tumor progression to the tumor-promoting processes warranting study.

P23 Pharmacological induction of CBS-mediated H2S production attenuates renal damage following deep hypothermia and rewarming

Hypothermic static preservation is commonly applied to preserve organ grafts during transplantation. However, cold ischemia is associated with the induction of renal injury, in which reactive oxygen species (ROS) play a major role. We recently reported that cellular uptake of catecholamines prevents hypothermia/rewarming-induced cell death by inhibiting ROS production. We also showed in cultured rat smooth muscle aortic cells that administration of catecholamines such as dopamine boosts endogenous H 2 S production leading to cell survival under hypothermic condition. Therefore in this study, we investigated whether these mechanisms are operational in vivo in the rat kidney, by investigating the effect of dopamine on renal injury induced by hypothermia and rewarming under ketamine anesthesia. Our results show that forced hypothermia followed by rewarming of rats induces glomerular and interstitial injury in animals anesthetized with ketamine, as indicated by histopathological changes, higher KIM-1 expression, influx of neutrophils and macrophages, and high ROS generation. Addition of dopamine on the other hand, reduced renal hypothermia/rewarming injury in animals anesthetized with ketamine and upregulated CBS expression and subsequent H 2 S production. These demonstrate that dopamine has renoprotection via CBS-induced H 2 S production against organ damage induced by hypothermia/rewarming.

Role of mitochondrial KATP channels in PKCβ‐dependent constriction of small pulmonary arteries (1090.10)

PKCβ mediates vasoconstriction through a unique signaling pathway in the pulmonary circulation that involves mitochondrial reactive oxygen species (mitoROS). We hypothesized that PKCβ elicits these responses through activation of vascular smooth muscle mitochondrial KATP channels (mitoKATP). To test this hypothesis, we examined constriction in endothelium‐disrupted, pressurized rat pulmonary arteries (~150 μm diameter) and mitoROS production (by MitoSOX fluorescence) in cultured rat and human pulmonary arterial smooth muscle cells (PASMC) in response to the PKC agonist phorbol 12‐myristate 13‐acetate (PMA). PMA caused vasoconstriction and mitoROS production that were significantly attenuated (p&lt;0.05) by both the PKCβ inhibitor LY‐333,531 (10 nM) and PKCβ siRNA. Consistent with our hypothesis, the specific mitoKATP channel antagonist 5‐hydroxydecanoate (5‐HD; 500 μM) inhibited vasoconstriction and mitoROS production to both PMA and the mitoKATP activator diazoxide (10‐5‐ 10‐3 M). Consistent with a role for mitoKATP in PKCβ‐dependent mitoROS generation, PMA caused mitochondrial membrane potential depolarization (detected by tetramethylrhodamine methyl ester fluorescence), which was prevented by 5‐HD (p&lt;0.05 vs. vehicle) in both pulmonary arteries and human PASMC. We conclude that PKCβ enhances mitoROS production via mitoKATP activation in PASMC to mediate vasoconstriction.Grant Funding Source: Supported by NIH R01 HL088192 and American Heart Association grants 1POST5260026 and 13GRNT14

Inhibition of Interleukin-6 Expression by Curcumin in Rat Vascular Smooth Muscle Explants &lt;I&gt;In Vitro&lt;/I&gt;

In atherosclerotic plaques, infectious agents from microbes may release Lipopolysaccharide (LPS) and heat shock proteins that can stimulate the production of mediators, i.e., pro-inflammatory cytokines, by vascular endothelial cells and Smooth Muscle Cells (SMCs). The elevated level of Interleukin-6 (IL-6) is strongly associated with the development of atherosclerosis. Here, we investigated whether curcumin isolated from the rhizome of Curcuma longa affected the expression of IL-6 at protein and gene levels in rat smooth muscle cells treated with LPS in vitro by conducting ELISA and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) assays. LPS at 2 µg mL-1 activated the expression of IL-6 protein and mRNA in SMCs. Curcumin (1-15 µM) caused the decreased levels of IL-6 protein and mRNA in the dose-dependent manner in LPS-induced SMCs, indicating its potential antiatherosclerotic effect for cardiovascular risk management. In addition, curcumin also partially blocked the activation of LPS-induced phosphorylation of MAP kinases, i.e., ERK1/2, p38 and JNK, in SMCs, suggesting it may inhibit IL-6 expression via attenuating MAP kinase signaling pathways in LPS-induced SMCs. These data may in part explain the molecular action of antiatherosclerotic effects of curcumin.

Contribution of ATP-Sensitive K+ Channels to Contraction of Rat Tail Artery is Significantly Reduced in Males but Not Females Rats in Streptozotocin-Induced Diabetes: Pharmacological Analysis

The aim of this article is to find out possible gender differences of participation of ATP-sensitive potassium channels in regulation of contractility in vascular smooth muscles of rats with diabetes. We used isolated tail artery of both genders rats with streptazotocin-induced diabetes (SID). After 3 weeks, we compared concentration-effects curves obtained with phenylephrine (0.1 to 300 μM) in the presence of 100 μM of diazoxid in control and SID rats of both genders. Diazoxide at 100 μM concentration significantly antagonizes concentration-dependent contraction of tail artery induced by phenylephrine in both male and female control rats. However, this antagonism disappeared in male but not female rats treated by streptozotocin. Our results indicate that SID leads to downregulation or functional malformation (lack of reaction to the opener) of ATP-sensitive potassium channels in males, but not in females rats. This observation has to be confirmed by immunochemistry analysis and taking into account in humans.

Direct writing of bio-functional coatings for cardiovascular applications

The surface modification of metallic biomaterials is of critical importance to enhance the biocompatibility of surgical implant materials and devices. This article investigates the use of a direct-write inkjet technique for multilayer coatings of a biodegradable polymer (polyester urethane urea (PEUU)) embedded with an anti-proliferation drug paclitaxel (Taxol). The direct-write inkjet technique provides selective patterning capability for depositing multimaterial coatings on three-dimensional implant devices such as pins, screws, and stents for orthopedic and vascular applications. Drug release profiles were studied to observe the influence of drug loading and coating thickness for obtaining tunable release kinetics. Platelet deposition studies were conducted following ovine blood contact and significant reduction in platelet deposition was observed on the Taxol loaded PEUU substrate compared with the unloaded control. Rat smooth muscle cells were used for cell proliferation studies. Significant reduction in cell growth was observed following the release of anti-proliferative drug from the biopolymer thin film. This research provides a basis for developing anti-proliferative biocompatible coatings for different biomedical device applications.

Intramyocardial Injection of a Synthetic Hydrogel with Delivery of bFGF and IGF1 in a Rat Model of Ischemic Cardiomyopathy

It is increasingly appreciated that the properties of a biomaterial used in intramyocardial injection therapy influence the outcomes of infarcted hearts that are treated. In this report the extended in vivo efficacy of a thermally responsive material that can deliver dual growth factors while providing a slow degradation time and high mechanical stiffness is examined. Copolymers consisting of N-isopropylacrylamide, 2-hydroxyethyl methacrylate, and degradable methacrylate polylactide were synthesized. The release of bioactive basic fibroblast growth factor (bFGF) and insulin-like growth factor 1 (IGF1) from the gel and loaded poly(lactide-co-glycolide) microparticles was assessed. Hydrogel with or without loaded growth factors was injected into 2 week-old infarcts in Lewis rats and animals were followed for 16 weeks. The hydrogel released bioactive bFGF and IGF1 as shown by mitogenic effects on rat smooth muscle cells in vitro. Cardiac function and geometry were improved for 16 weeks after hydrogel injection compared to saline injection. Despite demonstrating that left ventricular levels of bFGF and IGF1 were elevated for two weeks after injection of growth factor loaded gels, both functional and histological assessment showed no added benefit to inclusion of these proteins. This result points to the complexity of designing appropriate materials for this application and suggests that the nature of the material alone, without exogenous growth factors, has a direct ability to influence cardiac remodeling.

In vitro and in vivo degradation evaluation of novel iron-bioceramic composites for bone implant applications

Biodegradable metals such as magnesium, iron and their alloys have been known as potential materials for temporary medical implants. However, most of the studies on biodegradable metals have been focusing on optimizing their mechanical properties and degradation behavior with no emphasis on improving their bioactivity behavior. We therefore investigated the possibility of improving iron biodegradation rate and bioactivity by incorporating various bioactive bioceramics. The iron-based bioceramic (hydroxyapatite, tricalcium phosphate and biphasic calcium phosphate) composites were prepared by mechanical mixing and sintering process. Degradation studies indicated that the addition of bioceramics lowered the corrosion potential of the composites and slightly increased their corrosion rate compared to that of pure iron. In vitro cytotoxicity results showed an increase of cellular activity when rat smooth muscle cells interacted with the degrading composites compared to pure iron. X-ray radiogram analysis showed a consistent degradation progress with that found in vivo and positive tissue response up to 70days implantation in sheep animal model. Therefore, the iron-based bioceramic composites have the potential to be used for biodegradable bone implant applications.

Controlled Oxygen Release from Pyridone Endoperoxides Promotes Cell Survival under Anoxic Conditions

In tissue engineering, survival of larger constructs remains challenging due to limited supply of oxygen caused by a lack of early vascularization. Controlled release of oxygen from small organic molecules represents a possible strategy to prevent cell death under anoxic conditions. A comprehensive study of methylated pyridone-derived endoperoxides has led to the development of water-soluble molecules that undergo retro-Diels-Alder reactions in aqueous environment releasing oxygen in high yield and with half-lives of up to 13 h. These molecules in combination with vitamin C as singlet oxygen quencher significantly improved survival of 3T3 fibroblasts and rat smooth muscle cells challenged with oxygen-depleted conditions.

BRCA1 shields vascular smooth muscle cells from oxidative stress

Excessive production of reactive oxygen species (ROS), in part via upregulation of DNA damage pathways, is a central mechanism governing pathologic activation of vascular smooth muscle cells (VSMCs). We hypothesized that the breast cancer 1, early onset (BRCA1) gene that is involved in cellular resistance to DNA damage limits ROS production and oxidative stress in VSMCs. We evaluated basal and H2O2-stimulated expression of BRCA1 in human aortic smooth muscle cells (HASMCs). In vitro gain-of-function experiments were performed in BRCA1 adenovirus (Ad-BRCA1)-transfected HASMCs. ROS production and expression of Nox1 and its key regulatory subunit p47phox, key components of the ROS-generating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system, were evaluated. In vivo gain-of-function experiments were performed in spontaneously hypertensive (SHR) rats treated with Ad-BRCA1 (5 × 10(10) IU/rat). Blood pressure, vascular ROS generation, Nox1, and p47phox expression were measured. BRCA1 was constitutively expressed in murine, rat, and human smooth muscle cells (SMCs). H2O2 significantly reduced BRCA1 expression with a resultant increase in ROS generation. BRCA1-overexpressing HASMCs were protected against H2O2-induced ROS generation, in part, via downregulation of the ROS-producing NADPH oxidase subunits Nox1 and p47phox. Ad-BRCA1 treatment in SHR rats was associated with a sustained increase in aortic BRCA1 expression, lower aortic ROS production, reduced γH2A.X levels, greater RAD51 foci, and decreases in blood pressure. BRCA1 is a novel and previously unrecognized target that may shield VSMCs from oxidative stress by inhibiting NADPH Nox1-dependent ROS production. Gene- and/or cell-based approaches that improve BRCA1 bioavailability may represent a new approach in the treatment of diverse vascular diseases associated with an aberrant VSMC phenotype.

Sexually dimorphic urethral activity in response to pharmacological activation of 5-HT1A receptors in the rat

In this study, we examined the possibility that 5-HT1A receptors may underlie sexually dimorphic mechanisms affecting the regulation of urethral functions in anesthetized rats. Simultaneous recordings of intravesical pressure under isovolumetric conditions, external urethral sphincter-electromyography, and urethral perfusion pressure were used to examine the effects of a 5-HT1A receptor agonist [8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT)] and antagonist (WAY-100635) on bladder and urethral functions. This research also evaluated the effects of 8-OH-DPAT and α-bungarotoxin (a neuromuscular blockade agent) on urethral continence using leak point pressure testing, and the distribution of 5-HT1A receptors in the lower urinary tract was assessed by immunohistochemistry. The serotonergic mechanism that controls the urinary bladder and external urethral sphincter-electromyography activity showed no significant sexual differences, but urethral activity in urethral perfusion pressure and leak point pressure values exhibited some sexual differences. 8-OH-DPAT enhanced urethral pressure during continence in rats of both sexes, but the drug elevated the pressure during voiding in male rats and reduced it in female rats. The distribution of 5-HT1A receptors in the spinal cord also showed some sexual differences. The present study contributes to our understanding of the role of 5-HT1A receptors in physiological and immunohistochemical properties of urethral smooth muscle in rats of different sexes. These findings may be a basis for the future development of pharmacotherapies for stress urinary incontinence in men.

Altered expression of caveolin 2 and 3 in smooth muscle of rat urinary bladder by 17β-estradiol

BackgroundThe purpose of this study was to investigate the effect of estrogen alteration on the expression of caveolin 2 and 3 in rat smooth muscle of urinary bladder.MethodsFemale Sprague–Dawley rats were divided into three groups: control, bilateral ovariectomy (Ovx), and bilateral ovariectomy followed by subcutaneous injections of 17β-estradiol (Ovx?+?Est). After 4 weeks, urodynamic measurements were taken to ascertain the contraction interval and contraction pressure. The expression and cellular localization of caveolin 2 and 3 were determined by Western blot and immunohistochemistry in rat urinary bladder smooth muscle.ResultsIn cystometrograms, the contraction interval (min) was significantly lower in the Ovx group (3.1?±?1.5) than in the control group (5.6?±?1.2), but was increased after estrogen treatment (9.3?±?1.0). Conversely, the average contraction pressure (mmHg) was higher in the Ovx group (26.2?±?2.3) than in the control group (21.9?±?3.1), and was decreased after estrogen treatment (23.8?±?3.5). Caveolin 2 and 3 expression was localized in the cell membrane of the smooth muscle. The protein expression of both caveolin 2 and 3 was significantly lower after ovariectomy and was restored to the control levels after 17β-estradiol treatment.ConclusionsHormonal alteration causes a significant change in the expression of caveolin 2 and 3 in smooth muscle of rat urinary bladder. These findings suggest that these molecules might have functional roles in the detrusor overactivity that occurs in association with hormonal alteration.