Sources of Variability in Genetic Association Studies: Insights from the Analysis of Hepatic Lipase (LIPC)

Human Mutation. May, 2002  |  Pubmed ID: 11968086

Genetic association studies have been widely used to identify loci that influence plasma lipoprotein concentrations, but few of the associations reported have proved consistently reproducible across different study populations. This lack of consistency is a widely recognized limitation of association studies, and is often ascribed to inadequate statistical power, population substructure, and population-specific linkage disequilibrium. However, few studies have assessed the causes of variability underlying a given genotype-phenotype association. We have examined two possible sources of variability in the association between the -514 polymorphism in hepatic lipase (LIPC) and plasma HDL-C concentrations. First, we compared the association between this polymorphism and hepatic lipase activity in four populations. A single copy of the -514C allele was associated with a 10 mmol.hr(-1).l(-1) increase in hepatic lipase activity in white American and Turkish men but only approximately 5 mmol.hr(-1).l(-1) in Chinese and African-American men. Second, we tested the effects of a stanozolol-induced increase in hepatic lipase activity on plasma HDL-C concentrations in men with normal (< 150mg/dl) or elevated (150-300mg/dl) levels of plasma triglyceride. The increase in hepatic lipase activity was similar in the two groups, but the resulting decline in HDL-C levels was significantly greater in normolipidemic men. These data suggest that the effect of a polymorphism on gene expression can vary among individuals, and that the resulting phenotype may be further modified by interactions with other factors. Three novel LIPC polymorphisms were identified in the study (-1596insC, -2740A>G, and -2880G>C).

Analysis of the Myoglobin Gene in Tibetans Living at High Altitude

High Altitude Medicine & Biology. 2002  |  Pubmed ID: 12006163

Myoglobin, a protein with an important role in muscle oxidative metabolism, is increased in high altitude residents. In the closely related hemoglobins, mutations cause or contribute to human disease. Furthermore, heme-containing proteins may be involved in oxygen sensing. We therefore tested the hypotheses that myoglobin allele frequencies differed in Tibetans, a long-resident human high-altitude population, compared with sea-level residents, and varied in relation to altitude among Tibetans. We obtained the sequence of exon 2 of the myoglobin gene in 146 Tibetans with greater than three generations of stable residence at altitude in rural Tibet. We compared the frequency of known polymorphic sites in this gene among Tibetans living at altitudes of 3000, 3700, and 4500 m and to allele frequencies previously obtained in 525 residents of Dallas, Texas. We also examined the association between different myoglobin genotypes and hemoglobin concentration, used as an index of myoglobin levels. The frequency of the myoglobin 79A allele was higher in the high altitude compared with the sea-level residents, but unchanged with increasing altitude among Tibetans. There was no significant deviation from Hardy-Weinberg equilibrium in any of the Tibetan altitude groups, nor was there any association between myoglobin genotype and hemoglobin concentration. Screening of exon 2 of the myoglobin gene in high altitude Tibetans does not show novel polymorphism or selection for specific myoglobin alleles as a function of altitude of residence or hypoxic challenge.

DNA-loaded Albumin Microbubbles Enhance Ultrasound-mediated Transfection in Vitro

Ultrasound in Medicine & Biology. Jun, 2002  |  Pubmed ID: 12113794

Ultrasound (US), with or without microbubbles, enhances gene transfer in cultured cells, but the effect is modest. We tested if attaching DNA to albumin microbubbles during bubble synthesis could enhance gene expression. Plasmid DNA was loaded on the albumin shell over a range of concentrations (500 to 10,000 microg/mL). Optimal gene expression occurred with loading doses of 4000 microg DNA/mL (4k-loaded bubbles). These microbubbles had diameters of 2.4 +/- 0.7 microm and carried 40 pg DNA/microbubble. DNA-loaded microbubbles had optimal transfection at higher delivered doses of DNA than unloaded bubbles mixed with plasmid. The 4k-loaded bubbles demonstrated a fivefold (p = 0.0003) increase in luciferase reporter expression over that with unloaded bubbles. Similarly, transfection efficiency was better for 4k-loaded microbubbles than unloaded microbubbles (41 +/- 3% vs. 9 +/- 3%, p < 0.0001). DNA loading of microbubbles enhances gene expression and transfection efficiency in US-targeted transfection in vitro and may represent an improved avenue for therapeutic gene delivery in vivo.

Bioeffects of Myocardial Contrast Microbubble Destruction by Echocardiography

Echocardiography (Mount Kisco, N.Y.). Aug, 2002  |  Pubmed ID: 12356345

Microbubble destruction during contrast echocardiography is known to cause capillary leaks and red blood cell extravasation in skeletal muscle. This study evaluated the biological effects of microbubble destruction on cardiac muscle.

Transcriptional Mapping and Genomic Analysis of the Cardiac Atria and Ventricles

Physiological Genomics. Dec, 2002  |  Pubmed ID: 12502795

The atria and ventricles of the heart have distinct development, structure, and physiology. However, only a few of the genes that underlie the differences between these tissues are known. We used a murine cardiac cDNA microarray to identify genes differentially expressed in the atria and ventricles. The reliability of these findings is supported by highly concordant repetition of hybridization, recognition of previously known atrial and ventricular isoforms of contractile proteins, and confirmation of results by quantitative PCR and in situ hybridization. We examined the most differentially regulated genes for evolutionarily conserved noncoding sequences and found that atrial-expressed genes have more predicted myocyte enhancer factor-2 (MEF2) binding sites than ventricle-predominant genes. We confirmed that messages for MEF2 family members are more abundant in the atria, as are their protein products. Moreover, the activity of a transgenic reporter construct for MEF2 activity is preferentially upregulated in the atria in response to hypertrophic stimuli. This study provides a greater understanding of the molecular differences between atria and ventricles and establishes the framework for an anatomically detailed evaluation of cardiac transcriptional regulation.

Optimization of Ultrasound Parameters for Cardiac Gene Delivery of Adenoviral or Plasmid Deoxyribonucleic Acid by Ultrasound-targeted Microbubble Destruction

Journal of the American College of Cardiology. Jul, 2003  |  Pubmed ID: 12875768

This study was undertaken to optimize echocardiographic parameters for successful gene delivery to the heart and to extend the method from adenoviral to plasmid deoxyribonucleic acid (DNA).

Ultrasound-targeted Microbubble Destruction Can Repeatedly Direct Highly Specific Plasmid Expression to the Heart

Circulation. Aug, 2003  |  Pubmed ID: 12912823

Noninvasive, tissue-specific delivery of therapeutic agents would be a valuable clinical tool. We have previously shown that ultrasound-targeted microbubble destruction can direct expression of an adenoviral reporter to the heart. The present study shows that this method can be applied to selectively deliver plasmid vectors to the heart.

Transcriptional Analysis of Mouse Skeletal Myofiber Diversity and Adaptation to Endurance Exercise

Journal of Muscle Research and Cell Motility. 2003  |  Pubmed ID: 14870974

Vertebrate skeletal myofibers are heterogeneous with respect to their metabolic, contractile and morphological properties. To better understand skeletal myofiber diversity and plasticity at the transcriptional level, we carried out a whole-genome gene expression analysis of skeletal muscles composed primarily of slow/oxidative fibers (soleus) and fast fibers (extensor digitorum longus, EDL). We also followed gene expression changes in plantaris muscles from mice undergoing voluntary wheel running, a protocol that triggers transformation of glycolytic fibers into oxidative ones. Microarray analysis identified 70 genes differentially expressed by 3-fold or greater in soleus vs. EDL muscles and 15 genes up-regulated in exercised vs. sedentary plantaris muscles. A subset of these results were verified by northern blot and/or real-time RT-PCR analyses. Our results expand knowledge of the differences among various types of skeletal myofibers and their adaptation to exercise at the transcriptional level.

Knowledge Discovery by Automated Identification and Ranking of Implicit Relationships

Bioinformatics (Oxford, England). Feb, 2004  |  Pubmed ID: 14960466

MOTIVATION: New relationships are often implicit from existing information, but the amount and growth of published literature limits the scope of analysis an individual can accomplish. Our goal was to develop and test a computational method to identify relationships within scientific reports, such that large sets of relationships between unrelated items could be sought out and statistically ranked for their potential relevance as a set. RESULTS: We first construct a network of tentative relationships between 'objects' of biomedical research interest (e.g. genes, diseases, phenotypes, chemicals) by identifying their co-occurrences within all electronically available MEDLINE records. Relationships shared by two unrelated objects are then ranked against a random network model to estimate the statistical significance of any given grouping. When compared against known relationships, we find that this ranking correlates with both the probability and frequency of object co-occurrence, demonstrating the method is well suited to discover novel relationships based upon existing shared relationships. To test this, we identified compounds whose shared relationships predicted they might affect the development and/or progression of cardiac hypertrophy. When laboratory tests were performed in a rodent model, chlorpromazine was found to reduce the progression of cardiac hypertrophy.

Cardiovascular Gene Therapy: Angiogenesis and Beyond

The American Journal of the Medical Sciences. Mar, 2004  |  Pubmed ID: 15090752

Recent advances in understanding the mechanisms of disease have produced many new targets for gene therapy. However, it has been difficult to convert these new insights into clinically useful applications. In the field of cardiovascular medicine, most clinical studies of gene therapy have focused on angiogenesis as a treatment for ischemia. Initial enthusiasm was supported by small, uncontrolled, phase 1 trials. However, several large efficacy studies have recently been published that have not shown clinically significant improvement, and a few well-publicized complications of gene therapy have cast a pall over the entire field. In this review, we will summarize specific technical aspects of cardiovascular gene therapy, examine the recent series of clinical studies, and explore the direction of future work for the principal cardiovascular diseases.

Effects of Ultrasound-targeted Microbubble Destruction on Cardiac Gene Expression

Ultrasound in Medicine & Biology. Apr, 2004  |  Pubmed ID: 15121256

Ultrasound (US) contrast agents are increasingly used in diagnostic echocardiography. Recent studies have suggested unanticipated effects of microbubble destruction. This study was designed to evaluate gene regulation caused by US-mediated destruction of microbubbles in the heart. During IV infusion of Optison trade mark, triggered US was applied to rat hearts to destroy microbubbles. A control group received only saline and US. RNA was isolated from hearts 24 and 72 h after treatment. Analysis with a deeply representative murine cardiac-specific microarray was used to identify regulated genes. Real-time polymerase chain reaction (PCR) was then applied to verify regulated genes. Microarray analysis revealed only 5 regulated genes in the 24-h group and 4 in the 72-h group. Of these genes, only carbonic anhydrase was significantly upregulated in the 24-h Optison trade mark group (4.3 fold; p = 0.0005) when examined in individual animals by real-time PCR. By this very sensitive technique, the bioeffects of microbubble destruction are negligible.

In Vivo Transcriptional Response of Cardiac Endothelium to Lipopolysaccharide

Arteriosclerosis, Thrombosis, and Vascular Biology. Oct, 2004  |  Pubmed ID: 15319271

Vascular endothelial cells must integrate stimuli from multiple sources, including plasma, leukocytes, and neighboring components of the vessel. These stimuli are difficult to recapitulate in vitro. We have developed a method to examine the in vivo regulation of gene expression in endothelial cells and have applied it to a model of sepsis.

Mice with Cardiomyocyte-specific Disruption of the Endothelin-1 Gene Are Resistant to Hyperthyroid Cardiac Hypertrophy

Proceedings of the National Academy of Sciences of the United States of America. Feb, 2004  |  Pubmed ID: 14764893

Endothelin 1 (ET-1), a potent vasoconstrictor peptide expressed by endothelium, is also produced in the heart in response to a variety of stresses. It induces hypertrophy in cultured cardiac myocytes but only at concentrations far greater than those found in plasma. We tested whether ET-1 generated by cardiac myocytes in vivo is a local signal for cardiac hypertrophy. To avoid the perinatal lethality seen in systemic ET-1-null mice, we used the Cre/loxP system to generate mice with cardiac myocyte-specific disruption of the ET-1 gene. We used the alpha-myosin heavy chain promoter to drive expression of Cre and were able to obtain 75% reduction in ET-1 mRNA in cardiac myocytes isolated from these mice at baseline and after stimulation, in vivo, for 24 h with tri-iodothyronine (T3). Necropsy measurements of cardiac mass indexed for body weight showed a 57% reduction in cardiac hypertrophy in response to 16 days of exogenous T3 in mice homozygous for the disrupted ET-1 allele compared to siblings with an intact ET-1 gene. Moreover, in vivo MRI showed only a 3% increase in left ventricular mass indexed for body weight in mice with the disrupted allele after 3 weeks of T3 treatment versus a 27% increase in mice with an intact ET-1 gene. A reduced hypertrophic response was confirmed by planimetry of cardiac myocytes. We conclude that ET-1, produced locally by cardiac myocytes, and acting in a paracrine/autocrine manner, is an important signal for myocardial hypertrophy that facilitates the response to thyroid hormone.

Use of Ultrasound Contrast Agents for Gene or Drug Delivery in Cardiovascular Medicine

Journal of the American College of Cardiology. Feb, 2005  |  Pubmed ID: 15680708

The clinical utility of ultrasound contrast agents has been established in diagnostic echocardiography. Recently, the use of such agents has been promoted for transport and delivery of various bioactive substances, thus providing a technique for non-invasive gene therapy and organ-specific drug delivery. In this review, we give a critical update of published studies using ultrasound contrast agents for therapeutic use. We discuss the potential applications and limitations of this technique and suggest future applications in cardiovascular medicine.

Tobacco Smoke Dysregulates Endothelial Vasoregulatory Transcripts in Vivo

Physiological Genomics. May, 2005  |  Pubmed ID: 15728332

We hypothesized that human smoking and its deleterious effects on endothelial function can be modeled by exposure of mice to tobacco smoke, and further that these changes would be reflected in gene regulation in vascular endothelium. We used for these studies a mouse strain that expresses green fluorescent protein under the control of an endothelial-specific promoter, Tie-2. Mice were exposed to sidestream smoke from reference cigarettes at 34 mg total suspended particulates/m3. After exposure for 5 days/wk for 1 and 6 wk, aortas were pooled from treatment and control groups. Endothelial cells were rapidly isolated by collagenase treatment followed by fluorescent activated cell sorting to yield populations of >95% purity. RNA isolated from >500 endothelial cells was amplified and analyzed on deeply representative long oligo microarrays. Transcripts dysregulated by >2.5-fold were confirmed by real-time PCR and selected proteins by immunofluorescent localization. In the endothelial cells, the observed more than threefold upregulation of complement factor H (Cfh), calcitonin receptor-like (Calcr1), and soluble epoxide hydrolase (Epxh2) may play a role in hypertensive responses of the vasculature to smoking. We have identified gene regulation in vivo in vascular endothelium that potentially underlies hypertensive responses to tobacco smoke.

Augmentation of Cardiac Protein Delivery Using Ultrasound Targeted Microbubble Destruction

Ultrasound in Medicine & Biology. May, 2005  |  Pubmed ID: 15866418

Gas-filled microbubbles have become an important tool as ultrasonic contrast agents. We have previously shown that ultrasound-targeted microbubble destruction (UTMD) can direct plasmids to the heart. The aim of this study was to evaluate UTMD for protein delivery. Six different groups of rats received 1 microg of luciferase protein with varying protocols: (1) luciferase-loaded microbubbles and ultrasound; (2) luciferase only; (3) luciferase and ultrasound; (4) luciferase-loaded microbubbles; (5) unloaded microbubbles incubated with luciferase and ultrasound; (6) unloaded microbubbles with ultrasound followed by luciferase. Relative luminescence units per mg protein per s were determined in hearts and control organs. The rats that received ultrasound and luciferase-loaded bubbles showed a six-fold higher cardiac luciferase uptake compared with control groups that did not include bubbles. None of the other groups significantly augmented cardiac luciferase activity. We conclude that ultrasound-targeted microbubble destruction can substantially and noninvasively augment organ-specific delivery of proteins.

Targeting Vascular Endothelium with Avidin Microbubbles

Ultrasound in Medicine & Biology. Sep, 2005  |  Pubmed ID: 16176794

Targeting microbubbles (MBs) to specific vascular beds enables contrast ultrasound to be used for molecular imaging. There are several methods for attaching targeting moieties to the surface of MBs. In the present study, we demonstrate that avidin (Av) can be incorporated into the shell of perfluorocarbon-exposed sonicated dextrose albumin (PESDA) MBs (Av-PESDA-MBs) and serve as an anchor that links Av-PESDA-MBs to biotinylated monoclonal antibodies (mAbs). This novel linking strategy was used to conjugate Av-PESDA-MBs to mAbs specific for endoglin (CD105) or a control IgG. MBs targeted to CD105 specifically bound to endothelial cells, but not to fibroblasts, in vitro but Av-PESDA-MBs conjugated with the control IgG did not specifically target either cell type. We conclude that Av-PESDA-MBs represent a novel and attractive tool to conjugate MBs with biotinylated mAbs for the purposes of vascular targeting and molecular imaging.

FcgammaRIIB Mediates C-reactive Protein Inhibition of Endothelial NO Synthase

Circulation Research. Nov, 2005  |  Pubmed ID: 16269657

C-reactive protein (CRP) is an acute-phase reactant that is positively correlated with cardiovascular disease risk and endothelial dysfunction. Whether CRP has direct actions on endothelium and the mechanisms underlying such actions are unknown. Here we show in cultured endothelium that CRP prevents endothelial NO synthase (eNOS) activation by diverse agonists, resulting in the promotion of monocyte adhesion. CRP antagonism of eNOS occurs nongenomically and is attributable to blunted eNOS phosphorylation at Ser1179. Okadaic acid or knockdown of PP2A by short-interference RNA reverses CRP antagonism of eNOS, indicating a key role for the phosphatase. Aggregated IgG, the known ligand for Fcgamma receptors, causes parallel okadaic acid-sensitive loss of eNOS function, FcgammaRIIB expression is demonstrable in endothelium, and heterologous expression studies reveal that CRP antagonism of eNOS requires FcgammaRIIB. In FcgammaRIIB(+/+) mice, CRP blunts acetylcholine-induced increases in carotid artery vascular conductance; in contrast, CRP enhances acetylcholine responses in FcgammaRIIB(-/-) mice. Thus FcgammaRIIB mediates CRP inhibition of eNOS via PP2A, providing a mechanistic link between CRP and endothelial dysfunction.

Potential Bioeffects of Ultrasonic Destruction of Microbubble Contrast Agents

Journal of the American College of Cardiology. Apr, 2006  |  Pubmed ID: 16580538

Efficient Gene Delivery to Pancreatic Islets with Ultrasonic Microbubble Destruction Technology

Proceedings of the National Academy of Sciences of the United States of America. May, 2006  |  Pubmed ID: 16709667

This study describes a method of gene delivery to pancreatic islets of adult, living animals by ultrasound targeted microbubble destruction (UTMD). The technique involves incorporation of plasmids into the phospholipid shell of gas-filled microbubbles, which are then infused into rats and destroyed within the pancreatic microcirculation with ultrasound. Specific delivery of genes to islet beta cells by UTMD was achieved by using a plasmid containing a rat insulin 1 promoter (RIP), and reporter gene expression was regulated appropriately by glucose in animals that received a RIP-luciferase plasmid. To demonstrate biological efficacy, we used UTMD to deliver RIP-human insulin and RIP-hexokinase I plasmids to islets of adult rats. Delivery of the former plasmid resulted in clear increases in circulating human C-peptide and decreased blood glucose levels, whereas delivery of the latter plasmid resulted in a clear increase in hexokinase I protein expression in islets, increased insulin levels in blood, and decreased circulating glucose levels. We conclude that UTMD allows relatively noninvasive delivery of genes to pancreatic islets with an efficiency sufficient to modulate beta cell function in adult animals.

Melatonin Administration Does Not Affect Isoproterenol-induced LVH

IEEE Engineering in Medicine and Biology Magazine : the Quarterly Magazine of the Engineering in Medicine & Biology Society. May-Jun, 2006  |  Pubmed ID: 16764435

Endogenous Endothelin-1 is Required for Cardiomyocyte Survival in Vivo

Circulation. Aug, 2006  |  Pubmed ID: 16908774

Endothelin-1 (ET-1) has potent vasoconstrictor and hypertrophic actions. Pharmacological antagonists of endothelin receptors attenuate cardiac hypertrophy, have been approved for treatment of pulmonary hypertension, and are under investigation for treatment of heart failure. To investigate the role of ET-1 in the heart, we created mice with cardiomyocyte deletion of ET-1.

Transcriptional Analysis of Doxorubicin-induced Cardiotoxicity

American Journal of Physiology. Heart and Circulatory Physiology. Mar, 2006  |  Pubmed ID: 16243910

Doxorubicin is an effective chemotherapeutic agent against a broad range of tumors. However, a threshold dose of doxorubicin causes an unacceptably high incidence of heart failure and limits its clinical utility. We have established two models of doxorubicin cardiotoxicity in mice: 1) in an acute model, mice are treated with 15 mg/kg of doxorubicin once; and 2) in a chronic model, they receive 3 mg/kg weekly for 12 wk. Using echocardiography, we have monitored left ventricular function during treatment in the chronic model and seen the expected development of dilated cardiomyopathy. Treated mice showed histological abnormalities similar to those seen in patients with doxorubicin cardiomyopathy. To investigate transcriptional regulation in these models, we used a muscle-specific cDNA microarray. We have identified genes that respond to doxorubicin exposure in both models and confirmed these results using real-time PCR. In the acute model, a set of genes is regulated early and rapidly returns to baseline levels, consistent with the half-life of doxorubicin. In the chronic model, which mimics the clinical situation much more closely, we identified dysregulated genes that implicate specific mechanisms of cardiac toxicity. These include STARS, a hypertrophy-responsive gene; SNF1-kinase, a potential modulator of ATP levels; and AXUD1, a downstream target of the proapoptotic regulator AXIN1.

Keeping the Engine Primed: HIF Factors As Key Regulators of Cardiac Metabolism and Angiogenesis During Ischemia

Journal of Molecular Medicine (Berlin, Germany). Dec, 2007  |  Pubmed ID: 18026917

Myocardial ischemia, the most common cause of cardiac hypoxia in clinical medicine, occurs when oxygen delivery cannot meet myocardial metabolic requirements in the heart. This deficiency can result from either a reduced supply of oxygen (decreased coronary bloodflow) or an increased myocardial demand for oxygen (increased wall stress or afterload). Patients with stable coronary artery disease as well as patients experiencing acute myocardial infarction can experience episodes of severe ischemia. Although hypoxia is an obligatory component, it is not the sole environmental stress experienced by the ischemic heart. Reperfusion after ischemia is associated with increased oxidative stress as the heart reverts to aerobic respiration and thereby generates toxic levels of reactive oxygen species (ROS). During mild ischemia, mitochondrial function is partially compromised and substrate preferences adapt to sustain adequate ATP generation. With severe ischemia, mitochondrial function is markedly compromised and anaerobic metabolism must provide energy no matter what the cost in generation of toxic ROS adducts. Ischemia produces a variety of environmental stresses that impair cardiovascular function. As a result, multiple signaling pathways are activated in mammalian cells during ischemia/reperfusion injury in an attempt to minimize cellular injury and maintain cardiac output. Amongst the transcriptional regulators activated are members of the hypoxia inducible factor (HIF) transcription factor family. HIF factors regulate a variety of genes that affect a myriad of cellular processes including metabolism, angiogenesis, cell survival, and oxygen delivery, all of which are important in the heart. In this review, we will focus on the metabolic and angiogenic aspects of HIF biology as they relate to the heart during ischemia. We will review the metabolic requirements of the heart under normal as well as hypoxic conditions, the effects of preconditioning and its regulation as it pertains to HIF biology, the apparent roles of HIF-1 and HIF-2 in intermediary metabolism, and translational applications of HIF-1 and HIF-2 biology to cardiac angiogenesis. Increased understanding of the role of HIFs in cardiac ischemia will ultimately influence clinical cardiovascular practice.

Broken Heart Syndrome--a Critical Review of Transient Left Ventricular Apical Ballooning

Hawaii Medical Journal. Oct, 2007  |  Pubmed ID: 18065123

Transient left ventricular apical ballooning (TLVAB) is a recently proposed clinical entity that mimics an acute coronary syndrome in its clinical presentation. Retrospective studies suggest that it may account for 1% of myocardial infarction diagnoses. We provide a review of TLVAB including its clinical presentation, diagnostic studies, etiology, pathogenesis, criticism, complications, and management.

In Vivo Endothelial Gene Regulation in Diabetes

Cardiovascular Diabetology. 2008  |  Pubmed ID: 18423039

An authentic survey of the transcript-level response of the diabetic endothelium in vivo is key to understanding diabetic cardiovascular complications such as accelerated atherosclerosis and endothelial dysfunction.

Down-regulation of Caveolin-1, an Inhibitor of Transforming Growth Factor-beta Signaling, in Acute Allergen-induced Airway Remodeling

The Journal of Biological Chemistry. Feb, 2008  |  Pubmed ID: 18056268

Asthma can progress to subepithelial airway fibrosis, mediated in large part by transforming growth factor-beta (TGF-beta). The scaffolding protein caveolin-1 (cav1) can inhibit the activity of TGF-beta, perhaps by forming membrane invaginations that enfold TGF-beta receptors. The study goals were 1) to evaluate how allergen challenge affects lung expression of cav1 and the density of caveolae in vivo 2) to determine whether reduced cav1 expression is mediated by interleukin (IL)-4 and 3) to measure the effects of decreased expression of cav1 on TGF-beta signaling. C57BL/6J, IL-4-deficient mice, and cav1-deficient mice, sensitized by intraperitoneal injections of phosphate-buffered saline or ovalbumin (OVA) at days 0 and 12, received intranasal phosphate-buffered saline or OVA challenges at days 24, 26, and 28. Additionally, another group of C57BL/6J mice received IL-4 by intratracheal instillation for 7 days. We confirmed that the OVA-allergen challenge increased eosinophilia and T-helper type 2-related cytokine levels (IL-4, IL-5, and IL-13) in bronchoalveolar lavage. Allergen challenge reduced lung cav1 mRNA abundance by 40%, cav1 protein by 30%, and the number of lung fibroblast caveolae by 50%. Administration of IL-4 in vivo also substantially decreased cav1 expression. In contrast, the allergen challenge did not decrease cav1 expression in IL-4-deficient mice. The reduced expression of cav1 was associated with activation of TGF-beta signaling that was further enhanced in OVA-sensitized and challenged cav1-deficient mice. This study demonstrates a previously unknown modulation of TGF-beta signaling by IL-4, via cav1, suggesting novel therapeutic targets for controlling the effects of TGF-beta and thereby ameliorating pathological airway remodeling.

Cardioprotective Signaling by Endothelin

Trends in Cardiovascular Medicine. Oct, 2008  |  Pubmed ID: 19232951

The endothelin axis promotes vasoconstriction, suggesting that antagonists of endothelin signaling might be useful in treatment of heart failure. However, promising results from animal trials have not been recapitulated in heart failure patients. Here we review the role of major signaling pathways in the heart that are involved in cell survival initiated by ET-1. These pathways include mitogen-activated protein kinase, phosphatidyl inositol-1,4,5-triphosphate kinase (PI3K-AKT), nuclear factor-kappaB (NF-kappaB), and calcineurin signaling. A better understanding of endothelin-mediated signaling in cardiac cell survival may allow a reevaluation of endothelin receptor antagonists (ETRAs) in the treatment of heart failure.

Tiny Bubbles and Endocytosis?

Circulation Research. Mar, 2009  |  Pubmed ID: 19286610

Conditional HIF-1alpha Expression Produces a Reversible Cardiomyopathy

PloS One. 2010  |  Pubmed ID: 20657781

The response to hypoxia in tissues is regulated by the heterodimeric transcription factor Hypoxia Inducible Factor-1 (HIF-1).

Transcriptional Analysis of the Endothelial Response to Diabetes Reveals a Role for Galectin-3

Physiological Genomics. Oct, 2011  |  Pubmed ID: 21791638

To characterize the endothelial dysfunction associated with Type II diabetes, we surveyed transcriptional responses in the vascular endothelia of mice receiving a diabetogenic, high-fat diet. Tie2-GFP mice were fed a diet containing 60% fat calories (HFD); controls were littermates fed normal chow. Following 4, 6, and 8 wk, aortic and leg muscle tissues were enzymatically dispersed, and endothelial cells were obtained by fluorescence-activated cell sorting. Relative mRNA abundance in HFD vs. control endothelia was measured with long-oligo microarrays; highly dysregulated genes were confirmed by real-time PCR and protein quantification. HFD mice were hyperglycemic by 2 wk and displayed vascular insulin resistance and decreased glucose tolerance by 5 and 6 wk, respectively. Endothelial transcripts upregulated by HFD included galectin-3 (Lgals3), 5-lipoxygenase-activating protein, and chemokine ligands 8 and 9. Increased LGALS3 protein was detected in muscle endothelium by immunohistology accompanied by elevated LGALS3 in the serum of HFD mice. Our comprehensive analysis of the endothelial transcriptional response in a model of Type II diabetes reveals novel regulation of transcripts with roles in inflammation, insulin sensitivity, oxidative stress, and atherosclerosis. Increased endothelial expression and elevated humoral levels of LGALS3 supports a role for this molecule in the vascular response to diabetes, and its potential as a direct biomarker for the inflammatory state in diabetes.

Conditional HIF-1 Induction Produces Multistage Neovascularization with Stage-specific Sensitivity to VEGFR Inhibitors and Myeloid Cell Independence

Blood. Apr, 2011  |  Pubmed ID: 21307392

Neovascularization is a crucial component of tumor growth and ischemia. Although prior work primarily used disease models, delineation of neovascularization in the absence of disease can reveal intrinsic mechanisms of microvessel regulation amenable to manipulation in illness. We created a conditional model of epithelial HIF-1 induction in adult mice (TetON-HIF-1 mice). Longitudinal photoacoustic microscopy (L-PAM) was coincidentally developed for noninvasive, label-free serial imaging of red blood cell-perfused vasculature in the same mouse for weeks to months. TetON-HIF-1 mice evidenced 3 stages of neovascularization: development, maintenance, and transgene-dependent regression. Regression occurred despite extensive and tight pericyte coverage. L-PAM mapped microvascular architecture and quantified volumetric changes in neocapillary morphogenesis, arteriovenous remodeling, and microvessel regression. Developmental stage endothelial proliferation down-regulation was associated with a DNA damage checkpoint consisting of p53, p21, and endothelial γ-H2AX induction. The neovasculature was temporally responsive to VEGFR2 immuno-blockade, with the developmental stage sensitive, and the maintenance stage resistant, to DC101 treatment. L-PAM analysis also pinpointed microvessels ablated or resistant to VEGFR2 immuno-blockade. HIF-1-recruited myeloid cells did not mediate VEGFR2 inhibitor resistance. Thus, HIF-1 neovascularization in the absence of disease is self-regulated via cell autonomous endothelial checkpoints, and resistant to angiogenesis inhibitors independent of myeloid cells.

Specific Antioxidant Selenoproteins Are Induced in the Heart During Hypertrophy

Archives of Biochemistry and Biophysics. Aug, 2011  |  Pubmed ID: 21621505

Selenium (Se) is thought to confer cardioprotective effects through the actions of antioxidant selenoprotein enzymes that directly limit levels of ROS such as hydrogen peroxide (H(2)O(2)) or that reverse oxidative damage to lipids and proteins. To determine how the selenoproteome responds to myocardial hypertrophy, two mouse models were employed: triidothyronine (T3)- or isoproterenol (ISO)-treatment. After 7days of T3- and ISO-treatment, cardiac stress was demonstrated by increased H(2)O(2) and caspase-3 activity. Neither treatment produced significant increases in phospholipid peroxidation or TUNEL-positive cells, suggesting that antioxidant systems were protecting the cardiomyocytes from damage. Many selenoprotein mRNAs were induced by T3- and ISO-treatment, with levels of methionine sulfoxide reductase 1 (MsrB1, also called SelR) mRNA showing the largest increases. MsrB enzymatic activity was also elevated in both models of cardiac stress, while glutathione peroxidase (GPx) activity and thioredoxin reductase (Trxrd) activity were moderately and nonsignificantly increased, respectively. Western blot assays revealed a marked increase in MsrB1 and moderate increases in GPx3, GPx4, and Trxrd1, particularly in T3-treated hearts. Thus, the main response of the selenoproteome during hypertrophy does not involve increased GPx1, but increased GPx3 for reducing extracellular H(2)O(2) and increased GPx4, Trxrd1, and MsrB1 for minimizing intracellular oxidative damage.

Genetic Determinants of Lipid Traits in Diverse Populations from the Population Architecture Using Genomics and Epidemiology (PAGE) Study

PLoS Genetics. Jun, 2011  |  Pubmed ID: 21738485

For the past five years, genome-wide association studies (GWAS) have identified hundreds of common variants associated with human diseases and traits, including high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and triglyceride (TG) levels. Approximately 95 loci associated with lipid levels have been identified primarily among populations of European ancestry. The Population Architecture using Genomics and Epidemiology (PAGE) study was established in 2008 to characterize GWAS-identified variants in diverse population-based studies. We genotyped 49 GWAS-identified SNPs associated with one or more lipid traits in at least two PAGE studies and across six racial/ethnic groups. We performed a meta-analysis testing for SNP associations with fasting HDL-C, LDL-C, and ln(TG) levels in self-identified European American (~20,000), African American (~9,000), American Indian (~6,000), Mexican American/Hispanic (~2,500), Japanese/East Asian (~690), and Pacific Islander/Native Hawaiian (~175) adults, regardless of lipid-lowering medication use. We replicated 55 of 60 (92%) SNP associations tested in European Americans at p<0.05. Despite sufficient power, we were unable to replicate ABCA1 rs4149268 and rs1883025, CETP rs1864163, and TTC39B rs471364 previously associated with HDL-C and MAFB rs6102059 previously associated with LDL-C. Based on significance (p<0.05) and consistent direction of effect, a majority of replicated genotype-phentoype associations for HDL-C, LDL-C, and ln(TG) in European Americans generalized to African Americans (48%, 61%, and 57%), American Indians (45%, 64%, and 77%), and Mexican Americans/Hispanics (57%, 56%, and 86%). Overall, 16 associations generalized across all three populations. For the associations that did not generalize, differences in effect sizes, allele frequencies, and linkage disequilibrium offer clues to the next generation of association studies for these traits.

Hypercholesterolemia Potentiates Aortic Endothelial Response to Inhaled Diesel Exhaust

Inhalation Toxicology. Jan, 2011  |  Pubmed ID: 21222557

Inhalation of diesel exhaust induces vascular effects including impaired endothelial function and increased atherosclerosis.

Toward a Holistic View of Transcriptional Regulation

Circulation. Cardiovascular Genetics. Feb, 2011  |  Pubmed ID: 21325161

Pancreatic Adenocarcinoma Associated With Werner's Syndrome (Adult-Onset Progeria)

Gastrointestinal Cancer Research : GCR. Jan, 2011  |  Pubmed ID: 21464868