Smooth Muscle Cell Metabolism Research Articles

Network Preservation Analysis Reveals Dysregulated Metabolic Pathways in Human Vascular Smooth Muscle Cell Phenotypic Switching.

Vascular smooth muscle cells are key players involved in atherosclerosis, the underlying cause of coronary artery disease. They can play either beneficial or detrimental roles in lesion pathogenesis, depending on the nature of their phenotypic changes. An in-depth characterization of their gene regulatory networks can help better understand how their dysfunction may impact disease progression. We conducted a gene expression network preservation analysis in aortic smooth muscle cells isolated from 151 multiethnic heart transplant donors cultured under quiescent or proliferative conditions. We identified 86 groups of coexpressed genes (modules) across the 2 conditions and focused on the 18 modules that are least preserved between the phenotypic conditions. Three of these modules were significantly enriched for genes belonging to proliferation, migration, cell adhesion, and cell differentiation pathways, characteristic of phenotypically modulated proliferative vascular smooth muscle cells. The majority of the modules, however, were enriched for metabolic pathways consisting of both nitrogen-related and glycolysis-related processes. Therefore, we explored correlations between nitrogen metabolism-related genes and coronary artery disease-associated genes and found significant correlations, suggesting the involvement of the nitrogen metabolism pathway in coronary artery disease pathogenesis. We also created gene regulatory networks enriched for genes in glycolysis and predicted key regulatory genes driving glycolysis dysregulation. Our work suggests that dysregulation of vascular smooth muscle cell metabolism participates in phenotypic transitioning, which may contribute to disease progression, and suggests that AMT (aminomethyltransferase) and MPI (mannose phosphate isomerase) may play an important role in regulating nitrogen and glycolysis-related metabolism in smooth muscle cells.

40-OR: Insulin Stabilized Arterial Plaque by Regulating Vascular Smooth Muscle Cell Metabolism and Differentiation

The main pathology of cardiovascular diseases (CVD) in insulin resistance and diabetes is an acceleration of atherosclerosis and increased risk for the development of unstable plaques. Insulin’s effect on vascular smooth muscle cell (VSMC) has been attributed to be pro-atherogenic due to its actions to increase migration and proliferation. We reported that insulin receptors (IR) are responsible for increasing arterial intimal hyperplasia after wire injury. Surprisingly, mice with VSMC specific deletion of IR (Myh11IRKO/ApoE-/-), atherosclerosis was increased with plaques exhibited unstable pathologies of less VSMC and ECM, but more macrophages and necrosis than ApoE-/- mice. To characterize these unexpected findings in the arterial plaques of Myh11IRKO/ApoE-/- mice, we performed single cell RNA sequencing study of VSMC comparing 4 groups of mice including ApoE-/- and MyH11IRKO/ApoE-/- on normal chow or high fat diet (HFD). Bioinformatics analysis demonstrated that VSMC can be separated into 21 distinctive clusters with those expression enriched for contractile and mitochondrial genes separated from clusters which expressed heat shock protein (HSP) or inflammatory genes. Aorta of HFD insulin resistant WT ApoE-/- mice exhibited significantly increases of multiple clusters of inflammatory VSMCs and, surprisingly, also those clusters enriched for mitochondrial metabolism. However, in Myh11IRKO/ApoE-/- mice on HFD, aortic VMSC clusters with contractile and mitochondrial genes were significantly reduced, but distinct VMSC clusters with expressions of HSP or inflammatory genes were greatly increased. These novel findings, along with metabolic studies in cultured VSMC, indicated that insulin’s actions on VSMC metabolism to reduce the transition of VSMCs subsets from mitochondria enriched VSMCs to stressed and inflammatory VSMCs, which is critical to increase plaque stability and decrease CVD risks in people with insulin resistance and diabetes. Disclosure Q.Li: None. J.Fu: None. K.Park: None. I.Wu: None. G.L.King: Research Support; Janssen Research & Development, LLC.

Peranan Intervensi Fisioterapi Metode Senam Kegel Untuk Mengatasi Kegagalan Kerja Sel Otot Polos Pada Kelainan Kandung Kemih Inkontinensia Urin

Urinary incotinensia is a disease that causes sufferers to experience urine without realizing it or is often referred to as bed-wetting. Urinary incotinence causes sufferers to experience difficulty or sleep disturbances, depression, isolation, lack of confidence, irritability, limiting social activities, and spending a lot of money on medication so that they really need a treatment solution. Urinary incontinence results from weakening of the smooth muscle in the bladder organs. One of the treatments that can be done to overcome urinary incontinence is using physiotherapy, namely by doing Kegel exercises. The purpose of this paper is to determine the role of physiotherapy interventions using Kegel exercises to overcome the failure of smooth muscle cells in urinary incontinence bladder disorders. This literature review will discuss four focus studies, namely: 1) the structure and function of the renal organ system, 2) the structure and working mechanism of smooth muscle cells, 3) cellular studies of urinary incontinence, and 4) treatment of urinary incontinence using physiotherapy interventions. Physiotherapy with Kegel exercises has the benefit of strengthening the work of the bladder sphincter and pelvic floor muscles. The mechanism is by increasing smooth muscle cell metabolism. The metabolic rate of the muscles will increase the strength of the smooth muscle and the nervous system to contract, so that the smooth muscle will get stronger. However, Kegel exercises do not completely cure urinary incontinence and only relieve the symptoms it causes.Key words: Physiotherapy; Urinary incontinence; Smooth muscles; Cells; Kegel exercises.

Mechanism of NO Decay in Vascular Smooth Muscle Cells: Role of Cytoglobin and Identification of its Cellular Reducing System

Background In smooth muscle, cytoglobin (Cygb) functions as a potent nitric oxide (NO) dioxygenase and regulates NO metabolism and vascular tone. However, major questions remain regarding which cellular reducing systems regulate Cygb-mediated NO metabolism. Therefore, we performed studies to assess the intracellular concentrations of Cygb and its reducing systems in vascular smooth muscle cells (SMCs), and their role in the process of NO decay. Methods Cygb and its reducing systems, cytochrome B5 reductase (B5R), cytochrome B5 (B5) and cytochrome P450 reductase (CPR) were measured in aortic SMCs. Murine aortic smooth muscle cells (SMCs) were purchased from American Type Culture Collection (ATCC; CRL-2797™) and passaged. For depleting Cygb, B5R or B5, siRNAs were used. SMCs were transfected with Cygb siRNA, B5R siRNA, B5 siRNA or CPR siRNA using Lipofectamine RNAiMAX (Invitrogen). The rate of NO metabolism by normal SMCs of different cell numbers was assessed using a Clark-type NO electrode (NOCHM-4, WPI) utilizing two ports on the side wall of a four-port water-jacketed electrochemical chamber (NOCHM-4, WPI) and additionally another oxygen electrode was placed through second port on the side wall of the chamber. For protein assays, Cygb, B5R and B5 were expressed and purified using reported procedures. Quantitative immunoblotting was used to estimate the cellular concentrations of Cygb and each reducing system protein from comparison to standards of known concentration for each purified protein. Results Intracellular Cygb concentration was estimated as 3.5 µM, while B5R, B5, and CPR were 0.88, 0.38 and 0.15 µM, respectively. NO decay in SMCs was measured following bolus addition of 1 µM NO to air-equilibrated cells. The NO dioxygenation rate (VNO) with 3.5 x 106 cells•ml-1 was 12.7 ± 0.3 nM•s-1. siRNA-mediated knockdown experiments indicated that ~78% of NO metabolism in SMCs was Cygb-dependent. Of this, ~87 % was B5R- and B5-dependent. CPR knockdown resulted in a small decrease in VNO, while depletion of ascorbate had no effect. Kinetic analysis of VNO for the B5R/B5/Cygb system with variation of B5 or B5R concentrations from their SMC levels showed that VNO exhibits apparent Michaelis-Menten behavior for B5 and B5R. In contrast, linear variation was seen with change in Cygb concentration. Conclusions B5R/B5 was demonstrated to be the major reducing system supporting Cygb-mediated NO metabolism in SMCs and studies were performed to experimentally model and predict the alterations of VNO with changes in cellular B5R/B5/Cygb levels.

Defining the reducing system of the NO dioxygenase cytoglobin in vascular smooth muscle cells and its critical role in regulating cellular NO decay

In smooth muscle, cytoglobin (Cygb) functions as a potent nitric oxide (NO) dioxygenase and regulates NO metabolism and vascular tone. Major questions remain regarding which cellular reducing systems regulate Cygb-mediated NO metabolism. To better define the Cygb-mediated NO dioxygenation process in vascular smooth muscle cells (SMCs), and the requisite reducing systems that regulate cellular NO decay, we assessed the intracellular concentrations of Cygb and its putative reducing systems and examined their roles in the process of NO decay. Cygb and the reducing systems, cytochrome b5 (B5)/cytochrome b5 reductase (B5R) and cytochrome P450 reductase (CPR) were measured in aortic SMCs. Intracellular Cygb concentration was estimated as 3.5 μM, while B5R, B5, and CPR were 0.88, 0.38, and 0.15 μM, respectively. NO decay in SMCs was measured following bolus addition of NO to air-equilibrated cells. siRNA-mediated knockdown experiments indicated that ∼78% of NO metabolism in SMCs is Cygb-dependent. Of this, ∼87% was B5R- and B5-dependent. CPR knockdown resulted in a small decrease in the NO dioxygenation rate (VNO), while depletion of ascorbate had no effect. Kinetic analysis of VNO for the B5/B5R/Cygb system with variation of B5 or B5R concentrations from their SMC levels showed that VNO exhibits apparent Michaelis–Menten behavior for B5 and B5R. In contrast, linear variation was seen with change in Cygb concentration. Overall, B5/B5R was demonstrated to be the major reducing system supporting Cygb-mediated NO metabolism in SMCs with changes in cellular B5/B5R levels modulating the process of NO decay.

Impact of the Potential Antitumor Agent 2-(4-Hydroxyphenyl) Amino-1,4-Naphthoquinone (Q7) on Vasomotion Is Mediated by the Vascular Endothelium, But Not Vascular Smooth Muscle Cell Metabolism.

Vasomotion is defined as rhythmic oscillations in arterial diameter that regulate the blood flow and blood pressure. Because antitumor treatment may impair vascular functions and increase the blood pressure, we sought to evaluate whether a new naphthoquinone derivative, postulated as an antitumor agent, manifests adverse effects on vascular function. In this article, we evaluated the toxicity of 2-(4-hydroxyphenyl) amino-1,4-naphthoquinone (Q7) and its effects on vascular vasomotion in 3 models of vascular structure: endothelial cells, aortic ring, and smooth muscle cells. Although showing nontoxic effects, Q7 inhibited the formation of capillary-like structures of the EA.hy926 endothelial cell line grown on Matrigel. In exvivo experiments with aortic rings precontracted with phenylephrine (PE, 10-6 M), Q7 (10-5 M) significantly (P < 0.05) reduced vascular rhythmic contractions induced by the acetylcholine (ACh; 10-7-10-5 M), whereas sodium nitroprusside (a nitric oxide donor; 10-8 M) recovered the vasomotion. Furthermore, Q7 (10-5 M) did not decrease KCl-induced vascular rhythmic contractions in the aortic rings precontracted with BaCl2 (a nonselective K+ channel blocker; 10-3 M). Vascular smooth muscle cells (A7r5) preincubated with Q7 (10-5 M) for 3 hours also demonstrated a reduced glucose uptake. However, the Adenosine Triphosphate content was unaffected, suggesting that the rapid reduction in vasomotion observed in vascular reactivity experiments did not involve cellular metabolism but may be due to faster mechanisms involving endothelial nitric oxide and K+ channels leading to oscillations in intracellular Ca2+. In summary, the naphthoquinone derivative Q7 presents low cytotoxicity yet may alter the endothelial cell response and vasomotion in the absence of changes in smooth muscle cell metabolism.

Effect of Natural Cytokine Complex on Metabolism of Smooth Muscle Cells in Myocardial Arteries under Normal Conditions and during Hemodynamic Overload.

Histoenzymological methods were employed to examine the effects of systemically administered natural cytokine complex including IL-1, IL-2, IL-6, TNFα, MIF, and TGFβ on metabolism of smooth muscle cells in intramural myocardial arteries under physiological conditions and during acute hemodynamic overload of the heart. Natural cytokine complex markedly inhibited metabolism of vascular smooth muscle cells under control conditions and during acute experimental aortal stenosis. In vascular smooth muscle cells, deceleration of tricarboxylic acid cycle, redistribution of the fluxes in glycolytic cascade and its inhibition, down-regulation of oxidation of free fatty acids and their metabolites, and inhibition of the shuttle systems and biosynthetic processes were observed. Inhibition of metabolism in the vascular wall of myocardial arteries correlated with a decrease in their tone and could be partially determined by a decrease in contractile activity of smooth muscle cells. These findings do not exclude the involvement of other factors and mechanisms in down-regulation of metabolism in vascular myocytes in response to increased cytokines levels of in the blood, including their direct effect on biochemical processes in cells.

Histoenzymological Characteristics of Smooth Muscle Cells in Myocardial Vessels: A Comparative Study under Conditions of Increased Left or Right Ventricular Afterload.

Histoenzymological methods were used to study metabolism of smooth muscle cells of intramural myocardial arteries during experimental aortic or pulmonary artery stenosis. Aortic stenosis was accompanied by changes in smooth muscles of the left ventricle manifested by deceleration of tricarboxylic acid cycle, inhibition of oxidation of free fatty acids and their metabolites, flux redistribution in the glycolytic cascade, and inhibition of shuttle systems and biosynthetic processes. Similar metabolic alterations were observed in vessels of the ventricular septum, but they were not revealed in vessels of the right ventricle (except glycolysis stimulation). Under conditions of pulmonary artery stenosis, histoenzymological alterations in vascular smooth muscle of both ventricles and ventricular septum were similar, which attested to acceleration of tricarboxylic acid cycle, stimulation of oxidation of the free fatty acids with their metabolites, acceleration of glycolysis, and activation of the shuttle systems and biosynthetic processes. Comparative analysis of histoenzymological alterations revealed substantial differences in the character of metabolic changes under conditions of increased left and right ventricular afterload, which can be caused by peculiarities in myocardial blood flow, severity of circulatory disorders, severity of hypoxia, and intensity of processes maintaining ionic homeostasis in vascular smooth muscles and transport across the histohematic barriers. The data attest to important metabolic role of glycolysis in vascular smooth muscles of the myocardium, especially under conditions of enhanced afterload of the right ventricle.

Complex I dysfunction underlies the glycolytic switch in pulmonary hypertensive smooth muscle cells

ATP is essential for cellular function and is usually produced through oxidative phosphorylation. However, mitochondrial dysfunction is now being recognized as an important contributing factor in the development cardiovascular diseases, such as pulmonary hypertension (PH). In PH there is a metabolic change from oxidative phosphorylation to mainly glycolysis for energy production. However, the mechanisms underlying this glycolytic switch are only poorly understood. In particular the role of the respiratory Complexes in the mitochondrial dysfunction associated with PH is unresolved and was the focus of our investigations. We report that smooth muscle cells isolated from the pulmonary vessels of rats with PH (PH-PASMC), induced by a single injection of monocrotaline, have attenuated mitochondrial function and enhanced glycolysis. Further, utilizing a novel live cell assay, we were able to demonstrate that the mitochondrial dysfunction in PH-PASMC correlates with deficiencies in the activities of Complexes I–III. Further, we observed that there was an increase in mitochondrial reactive oxygen species generation and mitochondrial membrane potential in the PASMC isolated from rats with PH. We further found that the defect in Complex I activity was due to a loss of Complex I assembly, although the assembly of Complexes II and III were both maintained. Thus, we conclude that loss of Complex I assembly may be involved in the switch of energy metabolism in smooth muscle cells to glycolysis and that maintaining Complex I activity may be a potential therapeutic target for the treatment of PH.

LIM-only protein FHL2 is a positive regulator of liver X receptors in smooth muscle cells involved in lipid homeostasis.

The LIM-only protein FHL2 is expressed in smooth muscle cells (SMCs) and inhibits SMC-rich-lesion formation. To further elucidate the role of FHL2 in SMCs, we compared the transcriptomes of SMCs derived from wild-type (WT) and FHL2 knockout (KO) mice. This revealed that in addition to the previously recognized involvement of FHL2 in SMC proliferation, the cholesterol synthesis and liver X receptor (LXR) pathways are altered in the absence of FHL2. Using coimmunoprecipitation experiments, we found that FHL2 interacts with the two LXR isoforms, LXRα and LXRβ. Furthermore, FHL2 strongly enhances transcriptional activity of LXR element (LXRE)-containing reporter constructs. Chromatin immunoprecipitation (ChIP) experiments on the ABCG1 promoter revealed that FHL2 enhances the association of LXRβ with DNA. In line with these observations, we observed reduced basal transcriptional LXR activity in FHL2-KO SMCs compared to WT SMCs. This was also reflected in reduced expression of LXR target genes in intact aorta and aortic SMCs of FHL2-KO mice. Functionally, the absence of FHL2 resulted in attenuated cholesterol efflux to both ApoA-1 and high-density lipoprotein (HDL), in agreement with reduced LXR signaling. Collectively, our findings demonstrate that FHL2 is a transcriptional coactivator of LXRs and points toward FHL2 being an important determinant of cholesterol metabolism in SMCs.

Penile rehabilitation after pelvic cancer surgery.

Erectile dysfunction is the most common complication after pelvic radical surgery. Rehabilitation programs are increasingly being used in clinical practice but there is no high level of evidence supporting its efficacy. The principle of early penile rehabilitation stems from animal studies showing early histological and molecular changes associated with penile corporal hypoxia after cavernous nerve injury. The concept of early penile rehabilitation was developed in late nineties with a subsequent number of clinical studies supporting early pharmacologic penile rehabilitation. These studies included all available phosphodiesterase type 5 inhibitors, intracavernosal injection and intraurethral use of prostaglandin E1 and to lesser extent vacuum erectile devices. However, these studies are of small number, difficult to interpret, and often with no control group. Furthermore, no studies have proven an in vivo derangement of endothelial or smooth muscle cell metabolism secondary to a prolonged flaccid state. The purpose of the present report is a synthetic overview of the literature in order to analyze the concept and the rationale of rehabilitation program of erectile dysfunction following radical pelvic surgery and the evidence of such programs in clinical practice. Emphasis will be placed on penile rehabilitation programs after radical cystoprostatectomy, radical prostatectomy, and rectal cancer treatment. Future perspectives are also analyzed.

Mammalian Target of Rapamycin Complex 2 (mTORC2) Coordinates Pulmonary Artery Smooth Muscle Cell Metabolism, Proliferation, and Survival in Pulmonary Arterial Hypertension

Enhanced proliferation, resistance to apoptosis, and metabolic shift to glycolysis of pulmonary arterial vascular smooth muscle cells (PAVSMCs) are key pathophysiological components of pulmonary vascular remodeling in idiopathic pulmonary arterial hypertension (PAH). The role of the distinct mammalian target of rapamycin (mTOR) complexes mTORC1 (mTOR-Raptor) and mTORC2 (mTOR-Rictor) in PAVSMC proliferation and survival in PAH and their therapeutic relevance are unknown. Immunohistochemical and immunoblot analyses revealed that mTORC1 and mTORC2 pathways are markedly upregulated in small remodeled pulmonary arteries and isolated distal PAVSMCs from subjects with idiopathic PAH that have increased ATP levels, proliferation, and survival that depend on glycolytic metabolism. Small interfering RNA- and pharmacology-based analysis showed that although both mTORC1 and mTORC2 contribute to proliferation, only mTORC2 is required for ATP generation and survival of idiopathic PAH PAVSMCs. mTORC2 downregulated the energy sensor AMP-activated protein kinase, which led to activation of mTORC1-S6 and increased proliferation, as well as a deficiency of the proapoptotic protein Bim and idiopathic PAH PAVSMC survival. NADPH oxidase 4 (Nox4) protein levels were increased in idiopathic PAH PAVSMCs, which was necessary for mTORC2 activation, proliferation, and survival. Nox4 levels and mTORC2 signaling were significantly upregulated in small pulmonary arteries from hypoxia-exposed rats at days 2 to 28 of hypoxia. Treatment with the mTOR kinase inhibitor PP242 at days 15 to 28 suppressed mTORC2 but not Nox4, induced smooth muscle-specific apoptosis in small pulmonary arteries, and reversed hypoxia-induced pulmonary vascular remodeling in rats. These data provide a novel mechanistic link of Nox4-dependent activation of mTORC2 via the energy sensor AMP-activated protein kinase to increased proliferation and survival of PAVSMCs in PAH, which suggests a new potential pathway for therapeutic interventions.

Role of microangiopathy in diabetic cardiomyopathy

Although heart disease due to diabetes is mainly associated with complications of the large vessels, microvascular abnormalities are also considered to be involved in altering cardiac structure and function. Three major defects, such as endothelial dysfunction, alteration in the production/release of hormones, and shift in metabolism of smooth muscle cells, have been suggested to produce damage to the small arteries and capillaries (microangiopathy) due to hyperglycemia, and promote the development of diabetic cardiomyopathy. These factors may either act alone or in combination to produce oxidative stress as well as changes in cellular signaling and gene transcription, which in turn cause vasoconstriction and structural remodeling of the coronary vessels. Such alterations in microvasculature produce hypoperfusion of the myocardium and thereby lower the energy status resulting in changes in Ca(2+)-handling, apoptosis, and decreased cardiac contractile force. This article discusses diabetes-induced mechanisms of microvascular damage leading to cardiac dysfunction that is characterized by myocardial dilatation, cardiac hypertrophy as well as early diastolic and late systolic defects. Metabolic defects and changes in neurohumoral system due to diabetes, which promote disturbances in vascular homeostasis, are highlighted. In addition, increase in the vulnerability of the diabetic heart to the development of heart failure and the signaling pathways integrating nuclear factor κB and protein kinase C in diabetic cardiomyopathy are also described for comparison.

Influence of human cytomegalovirus infection on cholesterol metabolism in smooth muscle cells of umbilical arteries and the related mechanisms

Objective To investigate the effect of human cytomegalovirus(HCMV) infection on cholesterol metabolism in smooth muscle cells of the umbilical arteries and the related mechanisms.Methods Vascular smooth muscle cells were isolated from umbilical arteries and were challenged with 1 MOI HCMV,and the intercellular content of cholesterol was determined.Cells treated with DMEM/F12 solution containing 3% fetal bovine serum were taken as mock infection controls.The differentially expressed genes were screened by cDNA microarray in each group,and the abnormally expressed genes associated with cholesterol metabolism were identified by fluorescence quantitative RT-PCR.Results The intracellular content of cholesterol([0.71±0.06] μmol/10~6 cells) was significantly higher in HCMV infection group at 72 h after infection compared with that in the control group(t=7.950,P=0.000 2).cDNA microarray and fluorescence quantitative RT-PCR showed that the expressions of HMG-CoA synthetase and HMG-CoA reductase were greatly up-regulated compared with those in the mock infection group 48 and 72 h after infection(P0.01).Conclusion Human cytomegalovirus infection can increase the synthesis of cholesterol through up-regulating cholesterol metabolism-associated genes in vascular smooth muscle cells,leading to unbalancd cholesterol metabolism.

The pathomechanism and treatment of CADASIL

CADASIL has been reported notably in Europe, has recently been found to occur in Japan and, with the increase in the number of reported cases, has been attracting attention. Currently, the diagnosis of CADASIL is established clinically if the patient: (1) develops the condition at a relatively young age (40-50 years), (2) is not at risk for stroke, (3) has repeated attacks of lacunar infarction with gradual progression to pseudobulbar paralysis and dementia, and (4) has other family members with similar symptoms. The diagnosis is also established by imaging and laboratory studies if: (1) MRI reveals leukoaraiosis and multiple small infarcts in the deep white matter, basal ganglia, thalamus, and pons, with hyperintensities of the temporal pole and external capsule bilaterally; (2) electron microscopy demonstrates granular osmiophilic material (GOM) around vascular smooth muscles in the brain, skeletal muscle, and skin; and (3) DNA analysis shows notch3 mutations. The mechanism of development of CADASIL due to notch3 mutations is still unknown. However, a recent study revealed that Notch3 ectodomain is a major component of GOM. On binding to the ligand, Notch3 normally undergoes proteolytic cleavage, with the resulting clearance of the extracellular domain. However, in CADASIL, it accumulates as GOM and potentially inhibits normal metabolism in smooth muscle cells. It is necessary to suppress the production of GOM for fundamental therapy of CADASIL.

A multi-hit endocrine model of intrinsic adult-onset asthma

Epidemiological studies indicate that adult-onset asthma is initiated by stress (anxiety and depression), obesity and menopause. Ironically, despite our understanding of the various stressors that promote chronic adult-onset asthma, most of which are known to elevate cortisol production via the hypothalamic–pituitary–adrenal (HPA) axis, inhaled and systemic corticosteroids are the mainstay for the treatment of chronic asthma. This implicates other endocrine or cellular changes independent of cortisol synthesis in non-allergic adult-onset asthma. The mechanism by which corticosteroids are thought to modulate bronchial tone in relieving asthma is via corticosteroid-responsive genes that increase PGE 2 and cAMP production which promote muscle relaxation. Therefore, any physiological condition that suppresses intracellular PGE 2 and cAMP production would counter cortisol-induced muscle relaxation and potentially trigger non-allergic adult-onset asthma. Stress, obesity and menopause act on three interrelated endocrine pathways, the serotonergic, leptinergic and hypothalamic pathways, all of which operate through receptors to modulate cAMP and Ca 2+ metabolism in smooth muscle cells (SMCs). We propose that the level of SMC cAMP, as determined by overall signaling through corticosteroid receptors, leptin receptors and the GPCRs of the HPG and serotonergic pathways, will regulate bronchial tone (i.e. the ‘Multi-Hit Endocrine Model of Adult-Onset Asthma’). Thus, decreases in HPG (menopause) and serotonergic (depression) signaling and increases in leptinergic (obesity) signaling relative to HPA signaling would decrease cellular SMC cAMP and promote muscle contraction. This model can explain the discrepant epidemiological data associating stress, obesity, depression and menopause with adult-onset asthma and is supported by basic and clinical data. Treatment of depressed or menopausal asthmatics with selective serotonin reuptake inhibitors or hormone replacement therapy, respectively, alleviates bronchoconstriction. Future therapeutic strategies might therefore target the serotonergic, leptinergic and hypothalamic pathways in regulating cellular cAMP production and bronchoconstriction for the treatment of adult-onset asthma.

ATP stimulates MMP-2 release from human aortic smooth muscle cells via JNK signaling pathway

Aortic smooth muscle cell release of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2) has been implicated in aortic aneurysm pathogenesis, but proximal modulation of release is poorly understood. Extracellular nucleotides regulate vascular smooth muscle cell metabolism in response to physiochemical stresses, but nucleotide modulation of MMP and/or TIMP release has not been reported. We hypothesized that nucleotides modulate MMP-2 and TIMP-2 release from human aortic smooth muscle cells (HASMCs) via distinct purinergic receptors and signaling pathways. We exposed HASMCs to exogenous ATP and other nucleotides with and without interleukin-1beta (IL-1beta). HASMCs were pretreated in some experiments with apyrase, which degrades ATP, and inhibitors of ERK1/2, JNK, and p38 MAPK. MMP-2 and TIMP-2 released into supernatant were assessed using ELISA and Western blotting. ATP, adenosine, and UTP significantly stimulated MMP-2 release in the presence of IL-1beta (300 nM ATP: 181 +/- 22%, P = 0.003; 30 microm adenosine: 244 +/- 150%, P = 0.001; and 200 microm UTP: 153 +/- 40%, P = 0.015; vs. 100% constitutive). ATP also stimulated MMP-2 release in the absence of IL-1beta (100 microm ATP: 148 +/- 38% vs. 100% constitutive). Apyrase significantly reduced ATP-stimulated MMP-2 release (apyrase + 500 nM ATP: 59 +/- 3% vs. 124 +/- 7% with 500 nM ATP). Rank-order agonist potency for MMP-2 release was consistent with ATP activation of PAY and PAY receptors. ATP induced phosphorylation of intracellular JNK, and inhibition of the JNK pathway blocked ATP-stimulated MMP-2 release, indicating signaling via this pathway. Nucleotides are thus novel stimulants of MMP-2 release from HASMCs and may provide a mechanistic link between physiochemical stress in the aorta and aneurysms, especially in the context of inflammation.

Insulin and Blood Pressure

Hypertension has long been known to be more prevalent among obese subjects or in patients with diabetes, ie, in states of insulin resistance.1 After the demonstration that essential hypertension is an insulin-resistant state in its own right,2 it was logical to include hypertension in the insulin resistance syndrome,3 later transmuted into the metabolic syndrome.4 A large number of studies, both epidemiological and physiological, have explored this association and, more in general, the interrelationships between blood pressure and insulin action. Summarizing the available evidence—and critically analyzing its merits and pitfalls—is beyond the scope of this brief commentary. It may be nevertheless useful to recall a few general points. First, the association between insulin resistance and hypertension extends into the normal state as an association between insulin action on glucose metabolism and blood pressure levels5; this clearly speaks for the existence of multiple cross-talk between the 2 homeostatic systems. Second, longitudinal studies have confirmed that insulin resistance (as measured by its surrogate, fasting hyperinsulinemia) may precede the development of frank hypertension.6 The reverse temporal sequence, ie, hypertension antedating insulin resistance, has not been documented. Third, associations, even if fairly consistent, do …

Cadmium accumulation in aortas of smokers.

Abdominal aortic aneurysm is a smoking-related disorder. Cadmium, inhaled from cigarettes, may accumulate in the aorta and facilitate weakening of the aorta through adverse effects on smooth muscle cell metabolism. Cadmium was measured by atomic absorption spectrometry in infrarenal aortas from 13 patients with abdominal aortic aneurysm and from 17 age- and sex-matched patients with normal-diameter abdominal aorta. Total cadmium content was associated with smoking, assessed as pack-years (r=0.54, P=0.004), but was similar in aneurysmal and undilated aortas. The cadmium content (mean+/-SE) was higher in the media (3.25+/-0.53 ng/mg dry wt, 7+/-1.2 micromol/L) than in the intima or adventitia (1.14+/-0.24 and 1.87+/-0.38 ng/mg dry wt, respectively; ANOVA, P<0.005). There was a strong correlation between medial cadmium content and pack-years of smoking (r=0.87, P<0.001). In aortic smooth muscle cells cultured on fibrillar collagen, cadmium inhibited DNA synthesis and collagen synthesis and diminished cell numbers (IC(50) 2 micromol/L, 6 micromol/L, and 6 micromol/L, respectively), but higher concentrations of cadmium were required for upregulation of metallothionein (EC(50) 23 micromol/L). The cadmium content of the aorta increases in direct proportion to the pack-years of cigarettes smoked, with selective accumulation in the medial layer. However, the cadmium content of aneurysmal aortas was not higher than that of nondilated aortas for patients with matched smoking history. In smokers, the level of cadmium accumulation is probably sufficient to impair the viability of cultured smooth muscle cells. Similar mechanisms could underlie the development of degenerative aortic disease in smokers.

Oxidative stress leads to cholesterol accumulation in vascular smooth muscle cells

The transformation of macrophages and smooth muscle cells into foam cells by modified low-density lipoproteins (LDL) is one of the key events of atherogenesis. Effects of free radicals have mainly been studied in LDL, and other than toxicity, data dealing with direct action of free radicals on cells are scarce. This study focused on the direct effects of free radicals on cholesterol metabolism of smooth muscle cells. A free radical generator, azobis-amidinopropane dihydrochloride, was used, and conditions for a standardized oxidative stress were set up in vascular smooth muscle cells. After free radical action, the cells presented an accumulation of cholesterol that appeared to be the result of: (i) an increase in cholesterol biosynthesis and esterification; (ii) a decrease in cell cholesteryl ester hydrolysis; and (iii) a reduced cholesterol efflux. All these parameters were opposed by antioxidants. In addition, oxidant stress induced an increased degradation of acetyl-LDL, whereas no change was noted for native LDL. From this data, it was concluded that cholesterol metabolism of vascular smooth muscle cells was markedly altered by in vitro treatment with free radicals, although cell viability was unaffected. The resulting disturbance in cholesterol metabolism favors accumulation of cholesterol and cholesteryl esters in vascular cells, and thus may contribute to the formation of smooth muscle foam cells.