bluuebunny/crossref_metadata_2025 huggingface dataset:
Retinoic Acid–Induced Tissue Transglutaminase and Apoptosis in Vascular Smooth Muscle Cells
Authors: Ou Hesheng , Haendeler Judith , Aebly Michael R. , Kelly Louise A. , Cholewa Brian C. , Koike George , Kwitek-Black Anne , Jacob Howard J. , Berk Bradford C. , Miano Joseph M.
Published: June 2012

Abstract
—Retinoids exert antiproliferative and prodifferentiating effects in vascular smooth muscle cells (SMCs) and reduce neointimal mass in balloon-injured blood vessels. The mechanisms through which retinoids carry out these effects are unknown but likely involve retinoid receptor-mediated changes in gene expression. Here we report the cloning, chromosomal mapping, and biological activity of the retinoid-response gene rat tissue transglutaminase (tTG). Northern blotting studies showed that tTG is rapidly and dose-dependently induced in a protein synthesis–independent manner after stimulation with the natural retinoid all-
trans
retinoic acid (atRA). The induction of tTG was selective for atRA and its stereoisomers 9-
cis
and 13-
cis
RA, because little or no elevation in mRNA expression was observed with a panel of growth factors. Western blotting and immunofluorescence confocal microscopy showed an accumulation of cytosolic tTG protein after atRA stimulation. Radiolabeled cross-linking studies revealed a corresponding elevation in in vitro tTG activity. The increase in tTG activity was reduced in the presence of 2 distinct inhibitors of tTG (monodansylcadaverine and cystamine). atRA-induced tTG mRNA and protein expression were followed by a significant elevation in SMC apoptosis. Such retinoid-induced programmed cell death could be partially inhibited with each tTG inhibitor and was completely blocked when both inhibitors were used simultaneously. These results establish a role for atRA in the sequential stimulation of tTG and apoptosis in cultured SMCs. atRA-mediated apoptosis in SMCs seems to require the participation of active tTG, suggesting a potential mechanistic link between this retinoid-inducible gene and programmed cell death.
Shear Stress–Dependent Regulation of the Human β-Tubulin Folding Cofactor D Gene
Authors: Schubert Andreas , Cattaruzza Marco , Hecker Markus , Darmer Dorothea , Holtz Juergen , Morawietz Henning
Published: June 2012

Abstract
—The flowing blood generates shear stress at the endothelial cell surface. The endothelial cells modify their phenotype by alterations in gene expression in response to different levels of fluid shear stress. To identify genes involved in this process, human umbilical vein endothelial cells were exposed to laminar shear stress (venous or arterial levels) in a cone-and-plate apparatus for 24 hours. Using the method of RNA arbitrarily primed polymerase chain reaction, we cloned a polymerase chain reaction fragment representing an mRNA species downregulated by arterial compared with venous shear stress (shear stress downregulated gene-1, SSD-1). According to Northern blot analysis, corresponding SSD-1 cDNA clones revealed a similar, time-dependent downregulation after 24 hours of arterial shear stress compared with venous shear stress or static controls. Three SSD-1 mRNA species of 2.8, 4.1, and 4.6 kb were expressed in a tissue-specific manner. The encoded amino acid sequence of the human endothelial SSD-1 isoform (4.1-kb mRNA species) revealed 80.4% identity and 90.9% homology to the bovine β-tubulin folding cofactor D (tfcD) gene. Downregulation of tfcD mRNA expression by shear stress was defined at the level of transcription by nuclear run-on assays. The tfcD protein was downregulated by arterial shear stress. The shear stress–dependent downregulation of tfcD mRNA and protein was attenuated by the NO synthase inhibitor
Nω
-nitro-
l
-arginine methyl ester. Furthermore, the NO donor DETA-NO downregulated tfcD mRNA. Because tfcD was shown to be a microtubule-destabilizing protein, our data suggest a shear stress–dependent regulation of the microtubular dynamics in human endothelial cells.
Plasminogen Is a Critical Determinant of Vascular Remodeling in Mice
Authors: Drew Angela F. , Tucker Heidi L. , Kombrinck Keith W. , Simon Daniel I. , Bugge Thomas H. , Degen Jay L.
Published: June 2012

Abstract
—Extracellular proteolysis is likely to be a feature of vascular remodeling associated with atherosclerotic and restenotic arteries. To investigate the role of plasminogen-mediated proteolysis in remodeling, polyethylene cuffs were placed around the femoral arteries of mice with single and combined deficiencies in plasminogen and fibrinogen. Neointimal development occurred in all mice and was unaffected by genotype. Significant compensatory medial remodeling occurred in the cuffed arteries of control mice but not in plasminogen-deficient mice. Furthermore, focal areas of medial atrophy were frequently observed in plasminogen-deficient mice but not in control animals. A simultaneous deficit of fibrinogen restored the potential of the arteries of plasminogen-deficient mice to enlarge in association with neointimal development but did not eliminate the focal medial atrophy. An intense inflammatory infiltrate occurred in the adventitia of cuffed arteries, which was associated with enhanced matrix deposition. Adventitial collagen deposition was apparent after 28 days in control and fibrinogen-deficient arteries but not in plasminogen-deficient arteries, which contained persistent fibrin. These studies demonstrate that plasmin(ogen) contributes to favorable arterial remodeling and adventitial collagen deposition via a mechanism that is related to fibrinogen, presumably fibrinolysis. In addition, these studies reveal a fibrin-independent role of plasminogen in preventing medial atrophy in challenged vessels. (Circ Res. 2000;87:133-139.)
Evidence for Ca
2+
Activation and Inactivation Sites on the Luminal Side of the Cardiac Ryanodine Receptor Complex
Authors: Ching Li Lien , Williams Alan J. , Sitsapesan Rebecca
Published: June 2012

Abstract
—We have used tryptic digestion to determine whether Ca
2+
can regulate cardiac ryanodine receptor (RyR) channel gating from within the lumen of the sarcoplasmic reticulum (SR) or whether Ca
2+
must first flow through the channel and act via cytosolically located binding sites. Cardiac RyRs were incorporated into bilayers, and trypsin was applied to the luminal side of the bilayer. We found that before exposure to luminal trypsin, the open probability of RyR was increased by raising the luminal [Ca
2+
] from 10 μmol/L to 1 mmol/L, whereas after luminal trypsin exposure, increasing the luminal [Ca
2+
] reduced the open probability. The modification in the response of RyRs to luminal Ca
2+
was not observed with heat-inactivated trypsin, indicating that digestion of luminal sites on the RyR channel complex was responsible. Our results provide strong evidence for the presence of luminally located Ca
2+
activation and inhibition sites and indicate that trypsin digestion leads to selective damage to luminal Ca
2+
activation sites without affecting luminal Ca
2+
inactivation sites. We suggest that changes in luminal [Ca
2+
] will be able to regulate RyR channel gating from within the SR lumen, therefore providing a second Ca
2+
-regulatory effect on RyR channel gating in addition to that of cytosolic Ca
2+
. This luminal Ca
2+
-regulatory mechanism is likely to be an important contributing factor in the potentiation of SR Ca
2+
release that is observed in cardiac cells in response to increases in intra-SR [Ca
2+
].
Contribution of Monocytes/Macrophages to Compensatory Neovascularization
Authors: Moldovan Nicanor I. , Goldschmidt-Clermont Pascal J. , Parker-Thornburg Jan , Shapiro Steven D. , Kolattukudy Pappachan E.
Published: June 2012

Abstract
—In a transgenic model of ischemic cardiomyopathy in which monocytes are attracted to the myocardium by the targeted overexpression of monocyte chemoattractant protein-1 (MCP-1), we have observed the presence of endothelial NO synthase and platelet endothelial cell adhesion molecule-1–negative tunnels, occasionally containing blood-derived cells, that probe the cardiac tissue. Immunohistochemical data show that monocytes/macrophages (MCs/Mphs) drill tunnels using the broad-spectrum mouse macrophage metalloelastase. 5-Bromo-2′-deoxyuridine incorporation and neo-endothelial markers present in the microvasculature of MCP-1 mouse hearts suggest an active angiogenic process. Further studies will be required to establish that the MC-/Mph-drilled tunnels evolve to become capillaries, connected to the existing vessels and colonized by circulating endothelial cell progenitors. This possibility is supported by the availability of these cells, which is demonstrated by cell tagging with β-galactosidase placed under an active endothelial Tie-2 promoter. This phenomenon might represent another mechanism, in addition to the secretion of the angiogenic factors, by which MCs/MPhs may participate in the elaboration of new blood vessels in adult tissues.
Treatment With Dimethylthiourea Prevents Left Ventricular Remodeling and Failure After Experimental Myocardial Infarction in Mice
Authors: Kinugawa Shintaro , Tsutsui Hiroyuki , Hayashidani Shunji , Ide Tomomi , Suematsu Nobuhiro , Satoh Shinji , Utsumi Hideo , Takeshita Akira
Published: June 2012

Abstract
—Oxidative stress might play an important role in the progression of left ventricular (LV) remodeling and failure that occur after myocardial infarction (MI). We determined whether reactive oxygen species (ROS) are increased in the LV remodeling and failure in experimental MI with the use of electron spin resonance spectroscopy and whether the long-term administration of dimethylthiourea (DMTU), hydroxyl radical (·OH) scavenger, could attenuate these changes. We studied 3 groups of mice: sham-operated (sham), MI, and MI animals that received DMTU (MI+DMTU). Drugs were administered to the animals daily via intraperitoneal injection for 4 weeks. ·OH was increased in the noninfarcted myocardium from MI animals, which was abolished in MI+DMTU. Fractional shortening was depressed by 65%, LV chamber diameter was increased by 53%, and the thickness of noninfarcted myocardium was increased by 37% in MI. MI+DMTU animals had significantly better LV contractile function and smaller increases in LV chamber size and hypertrophy than MI animals. Changes in myocyte cross-sectional area determined with LV mid–free wall specimens were concordant with the wall thickness data. Collagen volume fraction of the noninfarcted myocardium showed significant increases in the MI, which were also attenuated with DMTU. Myocardial matrix metalloproteinase-2 activity, measured with gelatin zymography, was increased with MI after 7 and 28 days, which was attenuated in MI+DMTU. Thus, the attenuation of increased myocardial ROS and metalloproteinase activity with DMTU may contribute, at least in part, to its beneficial effects on LV remodeling and failure. Therapies designed to interfere with oxidative stress might be beneficial to prevent myocardial failure.
Dissociation of Sarcoglycans and the Dystrophin Carboxyl Terminus From the Sarcolemma in Enteroviral Cardiomyopathy
Authors: Lee Gil-Hwan , Badorff Cornel , Knowlton Kirk U.
Published: June 2012

Abstract
—Enteroviral infection can cause an acquired form of dilated cardiomyopathy. We recently reported that dystrophin is cleaved, functionally impaired, and morphologically disrupted in vitro as well as in vivo during infection with coxsackievirus B3. Genetic dystrophin truncations lead to a marked decrease in dystrophin-associated glycoproteins, whereas expression of only the naturally occurring dystrophin carboxyl terminus, Dp-71, restores the sarcolemmal association of the dystrophin-associated glycoproteins. We sought to determine whether acute cleavage of dystrophin leads to a dissociation of the carboxyl-terminal dystrophin fragment and of the sarcoglycans from the sarcolemma during coxsackievirus B3 infection. We found that in cultured cardiac myocytes and murine hearts infected with coxsackievirus B3, the sarcolemmal localization of the dystrophin carboxyl terminus is lost. The dystrophin-associated glycoproteins α-, β-, γ-, and δ-sarcoglycan and β-dystroglycan were markedly decreased in the membrane fraction of infected cells in culture, and the typical sarcolemmal localization for each of these proteins was lost in coxsackievirus-B3–infected cardiomyocytes in vivo. Furthermore, sucrose gradient ultracentrifugation demonstrated that δ-sarcoglycan was physically dissociated from dystrophin within the membrane fraction. In vivo, the sarcolemmal integrity was functionally impaired with Evans blue dye uptake even though there was no generalized disruption of the sarcolemma of infected myocytes evidenced by intact wheat germ agglutinin staining. In analogy to hereditary sarcoglycanopathies, this disintegration of the sarcoglycan complex may, in addition to the dystrophin cleavage, play an important role in the pathogenesis of enterovirus-induced cardiomyopathy. These results imply a potential role for disruption of the sarcoglycans in an acquired form of heart failure.
Role for Peroxisome Proliferator-Activated Receptor α in Oxidized Phospholipid–Induced Synthesis of Monocyte Chemotactic Protein-1 and Interleukin-8 by Endothelial Cells
Authors: Lee Hans , Shi Weibin , Tontonoz Peter , Wang Shirley , Subbanagounder Ganesamoorthy , Hedrick Catherine C. , Hama Susan , Borromeo Christine , Evans Ronald M. , Berliner Judith A. , Nagy Laszlo
Published: June 2012

Abstract
—The attraction, binding, and entry of monocytes into the vessel wall play an important role in atherogenesis. We have previously shown that minimally oxidized/modified LDL (MM-LDL), a pathogenically relevant lipoprotein, can activate human aortic endothelial cells (HAECs) to produce monocyte chemotactic activators. In the present study, we demonstrate that MM-LDL and oxidation products of 1-palmitoyl-2-arachidonyl-
sn
-glycero-3-phosphocholine (PAPC) activate endothelial cells to synthesize monocyte chemotactic protein-1 (MCP-1) and interleukin-8 (IL-8). Several lines of evidence suggest that this activation is mediated by the lipid-dependent transcription factor peroxisome proliferator-activated receptor α (PPARα), the most abundant member of the PPAR family in HAECs. Treatment of transfected CV-1 cells demonstrated activation of the PPARα ligand-binding domain by MM-LDL, Ox-PAPC, or its component phospholipids, 1-palmitoyl-2-oxovalaroyl-
sn
-glycero-phosphocholine and 1-palmitoyl-2-glutaroyl-
sn
-glycero-phosphocholine; these lipids also activated a consensus peroxisome proliferator-activated receptor response element (PPRE) in transfected HAECs. Furthermore, activation of PPARα with synthetic ligand Wy14,643 stimulates the synthesis of IL-8 and MCP-1 by HAECs. By contrast, troglitazone, a PPARγ agonist, decreased the levels of IL-8 and MCP-1. Finally, we demonstrate that unlike wild-type endothelial cells, endothelial cells derived from PPARα null mice do not produce MCP-1/JE in response to Ox-PAPC and MM-LDL. Together, these data demonstrate a proinflammatory role for PPARα in mediation of the activation of endothelial cells to produce monocyte chemotactic activity in response to oxidized phospholipids and lipoproteins.
Early Activation of the Multicomponent Signaling Complex Associated With Focal Adhesion Kinase Induced by Pressure Overload in the Rat Heart
Authors: Franchini Kleber G. , Torsoni Adriana S. , Soares Paulo H. A. , Saad Mario J. A.
Published: June 2012

Abstract
—Mechanical overload elicits functional and structural adaptive mechanisms in cardiac muscle. Signaling pathways linked to integrin/cytoskeleton complexes may have a function in mediation of the effects of mechanical stimulus in myocardial cells. We investigated the tyrosine phosphorylation and the assembly of the multicomponent signaling complex associated with focal adhesion kinase (Fak) and the actin cytoskeleton in the overloaded myocardium of rats. Pressure overload induced a 3-fold increase in Fak tyrosine phosphorylation within 3 minutes after a 60-mm Hg rise in aortic pressure. A pressure stimulus that lasted for 60 minutes was accompanied by a 5-fold increase in the amount of tyrosine-phosphorylated Fak, and a stimulus as low as 10 mm Hg doubled the amount of tyrosine-phosphorylated Fak in the myocardium within 10 minutes. Pressure overload also induced a time-dependent association of actin with Fak and an increase in the amount of Fak detected in the cytoskeletal fraction of the myocardium. These events were paralleled by c-Src activation and binding to Fak and by an association of Grb2 and p85 subunit of phosphatidylinositol 3-kinase with Fak. Erk1/2 and Akt, two possible downstream effectors of Fak via Grb2 and phosphatidylinositol 3-kinase, were also shown to be activated in parallel with Fak. These findings show that pressure overload induced a rapid activation of the Fak multiple signaling complex in the myocardium of rats, which suggests that this mechanism may have a role in mechanotransduction in the myocardium.
Contractile Reserve and Intracellular Calcium Regulation in Mouse Myocytes From Normal and Hypertrophied Failing Hearts
Authors: Ito Kenta , Yan Xinhua , Tajima Minori , Su Zhi , Barry William H. , Lorell Beverly H.
Published: June 2012

Abstract
—Mouse myocyte contractility and the changes induced by pressure overload are not fully understood. We studied contractile reserve in isolated left ventricular myocytes from mice with ascending aortic stenosis (AS) during compensatory hypertrophy (4-week AS) and the later stage of early failure (7-week AS) and from control mice. Myocyte contraction and [Ca
2+
]
i
transients with fluo-3 were measured simultaneously. At baseline (0.5 Hz, 1.5 mmol/L [Ca
2+
]
o
, 25°C), the amplitude of myocyte shortening and peak-systolic [Ca
2+
]
i
in 7-week AS were not different from those of controls, whereas contraction, relaxation, and the decline of [Ca
2+
]
i
transients were slower. In response to the challenge of high [Ca
2+
]
o
, fractional cell shortening was severely depressed with reduced peak-systolic [Ca
2+
]
i
in 7-week AS compared with controls. In response to rapid pacing stimulation, cell shortening and peak-systolic [Ca
2+
]
i
increased in controls, but this response was depressed in 7-week AS. In contrast, the responses to both challenge with high [Ca
2+
]
o
and rapid pacing in 4-week AS were similar to those of controls. Although protein levels of Na
+
-Ca
2+
exchanger were increased in both 4-week and 7-week AS, the ratio of SR Ca
2+
-ATPase to phospholamban protein levels was depressed in 7-week AS compared with controls but not in 4-week AS. This was associated with an impaired capacity to increase sarcoplasmic reticulum Ca
2+
load during high work states in 7-week AS myocytes. In hypertrophied failing mouse myocytes, depressed contractile reserve is related to an impaired augmentation of systolic [Ca
2+
]
i
and SR Ca
2+
load and simulates findings in human failing myocytes.
Enhanced Na
+
Channel Intermediate Inactivation in Brugada Syndrome
Authors: Wang Dao W. , Makita Naomasa , Kitabatake Akira , Balser Jeffrey R. , George Alfred L.
Published: June 2012

Abstract
—Brugada syndrome is an inherited cardiac disease that causes sudden death related to idiopathic ventricular fibrillation in a structurally normal heart. The disease is characterized by ST-segment elevation in the right precordial ECG leads and is frequently accompanied by an apparent right bundle-branch block. The biophysical properties of the
SCN5A
mutation T1620M associated with Brugada syndrome were examined for defects in intermediate inactivation (
I
M
), a gating process in Na
+
channels with kinetic features intermediate between fast and slow inactivation. Cultured mammalian cells expressing T1620M Na
+
channels in the presence of the human β
1
subunit exhibit enhanced intermediate inactivation at both 22°C and 32°C compared with wild-type recombinant human heart Na
+
channels (WT-hH1). Our findings support the hypothesis that Brugada syndrome is caused, in part, by functionally reduced Na
+
current in the myocardium due to an increased proportion of Na
+
channels that enter the
I
M
state. This phenomenon may contribute significantly to arrhythmogenesis in patients with Brugada syndrome. The full text of this article is available at http://www.circresaha.org.
Mechanisms of NO/cGMP-Dependent Vasorelaxation
Authors: Sausbier Matthias , Schubert Rudolf , Voigt Viktor , Hirneiss Christoph , Pfeifer Alexander , Korth Michael , Kleppisch Thomas , Ruth Peter , Hofmann Franz
Published: June 2012

Abstract
—Both cGMP-dependent and -independent mechanisms have been implicated in the regulation of vascular tone by NO. We analyzed acetylcholine (ACh)- and NO-induced relaxation in pressurized small arteries and aortic rings from wild-type (wt) and cGMP kinase I–deficient (cGKI
–/–
) mice. Low concentrations of NO and ACh decreased the spontaneous myogenic tone in wt but not in cGKI
–/–
arteries. However, contractions of cGKI
–/–
arteries and aortic rings were reduced by high concentrations (10 μmol/L) of 2-(
N
,
N
-diethylamino)-diazenolate-2-oxide (DEA-NO). Iberiotoxin, a specific blocker of Ca
2+
-activated K
+
(BK
Ca
) channels, only partially prevented the relaxation induced by DEA-NO or ACh in pressurized vessels and aortic rings. DEA-NO increased the activity of BK
Ca
channels only in vascular smooth muscle cells isolated from wt cGKI
+/+
mice. These results suggest that low physiological concentrations of NO decrease vascular tone through activation of cGKI, whereas high concentrations of DEA-NO relax vascular smooth muscle independent of cGKI and BK
Ca
. NO-stimulated, cGKI-independent relaxation was antagonized by the inhibition of soluble guanylyl cyclase or cAMP kinase (cAK). DEA-NO increased cGMP to levels that are sufficient to activate cAK. cAMP-dependent relaxation was unperturbed in cGKI
–/–
vessels. In conclusion, low concentrations of NO relax vessels by activation of cGKI, whereas in the absence of cGKI, NO can relax small and large vessels by cGMP-dependent activation of cAK.
Upregulation of Immunoreactive Angiotensin II Release and Angiotensinogen mRNA Expression by High-Frequency Preganglionic Stimulation at the Canine Cardiac Sympathetic Ganglia
Authors: Kushiku Kazushi , Yamada Hiromi , Shibata Kazuhiko , Tokunaga Ryoko , Katsuragi Takeshi , Furukawa Tatsuo
Published: June 2012

Abstract
—The possible involvement of the local angiotensin system in ganglionic functions was investigated in the canine cardiac sympathetic ganglia. Positive chronotropic responses to preganglionic stellate stimulation at high frequencies, after intravenous administration of pentolinium plus atropine, were inhibited by the nonpeptide angiotensin AT
1
receptor antagonist forasartan or the angiotensin I–converting enzyme inhibitor captopril, whereas the rate increases elicited by the postganglionic stellate stimulation and norepinephrine given intravenously failed to be inhibited by these antagonists. The levels of endogenous immunoreactive angiotensin II, as determined by radioimmunoassay in the incubation medium of the stellate and inferior cervical ganglia, were increased after the high-frequency preganglionic stimulation of the isolated ganglia. The increment of the peptide was also antagonized by the pretreatment with captopril but not by a chymase inhibitor, chymostatin. The expression of angiotensinogen mRNA was observed in the stellate ganglion, adrenal, liver, and lung but not in the ovary and spleen. The expression of the mRNA in the stellate and inferior cervical ganglia increased after high-frequency preganglionic stimulation of the in vivo dogs for a period of 1 hour. These results indicate that an intrinsic angiotensin I–converting enzyme–dependent angiotensin system exists in the cardiac sympathetic ganglia, which is activated by high-frequency preganglionic stimulation.
Homeobox Protein Hex Induces SMemb/Nonmuscle Myosin Heavy Chain-B Gene Expression Through the cAMP-Responsive Element
Authors: Sekiguchi Kenichi , Kurabayashi Masahiko , Oyama Yuko , Aihara Yasushi , Tanaka Toru , Sakamoto Hironosuke , Hoshino Yoichi , Kanda Tsugiyasu , Yokoyama Tomoyuki , Shimomura Yukio , Iijima Hirokazu , Ohyama Yoshio , Nagai Ryozo
Published: June 2012

Abstract
—Recent studies have shown that the homeobox gene Hex plays an important role in inducing differentiation of vascular endothelial cells. In this study, we examined the expression of Hex in vascular smooth muscle cells (VSMCs) in vitro and in vivo. Immunohistochemistry showed a marked induction of Hex protein in neointimal VSMCs after balloon injury in rat aorta. Western and reverse transcriptase–polymerase chain reaction analyses demonstrated that Hex was abundantly expressed in cultured VSMCs, whereas it was undetectable in other cell types or in normal aorta. The expression pattern of Hex was similar to that of SMemb/NMHC-B, a nonmuscle isoform of myosin heavy chain that we have previously reported to be a molecular marker of dedifferentiated VSMCs. We next examined the role of Hex in SMemb gene transcription. Promoter analysis demonstrated that the sequence identical to consensus cAMP-responsive element (CRE) located at −481 of the SMemb promoter was critical for Hex responsiveness. Mutant Hex expression vector, which lacks the homeodomain, failed to stimulate SMemb gene transcription, suggesting the requirement of the homeodomain for its transactivation. Elecrophoretic mobility shift assay showed that Hex binds to a consensus binding sequence for homeobox proteins, but not to CRE. Cotransfection of protein kinase A expression vector increased the ability of Hex to stimulate SMemb promoter activity in a CRE-dependent manner. Overexpression of CRE binding protein (CREB), but not Mut-CREB which contains mutation at Ser133, strongly activated Hex-induced SMemb promoter activity. These results suggest that Hex mediates transcriptional induction of the SMemb/NMHC-B gene via its homeodomain, and Hex can function as a transcriptional modulator of CRE-dependent transcription in VSMCs.
Reduction of [Ca
2+
]
i
Restores Uncoupled β-Adrenergic Signaling in Isolated Perfused Transgenic Mouse Hearts
Authors: Serikov Vladimir B. , Petrashevskaya Natalia N. , Canning Amy M. , Schwartz Arnold
Published: June 2012

The effects of alterations in calcium in the perfusion media were studied on β-adrenergic coupling in isolated hearts from 3 different transgenic mice: cardiac-specific overexpressed α

1

subunit of L-type calcium channel, overexpressed Gα

q

, and phospholamban knockout. Isolated hearts from all 3 models, when studied at [Ca

2+

] of 2 mmol/L in the perfusate, showed the usual blunted or no response to β-adrenergic stimulation. Lowering [Ca

2+

] to 0.75 to 1.5 mmol/L unloaded the hearts of calcium and restored to nearly normal the responsiveness to β-agonist stimulation.
Gene Transfer as an Approach to Treating Hemophilia
Authors: High Katherine A.
Published: June 2012

Abstract
—Hemophilia is an X-linked bleeding diathesis caused by a deficiency of either factor VIII or factor IX. Present treatment for hemophilia involves intravenous infusion of either recombinant or plasma-derived clotting factor concentrates. Problems with this treatment method, including the expense, need for intravenous access, and risks of blood-borne disease transmission, have fueled an interest in developing a gene-transfer approach to treatment. On the basis of experience with protein concentrate therapy, it seems likely that even modest elevations in circulating levels of factor VIII or factor IX can prevent most of the mortality and much of the morbidity associated with the disease. Hemophilia has a number of advantages as a model system for working out strategies for gene transfer as an approach to the treatment of genetic diseases; these include wide latitude in choice of target tissue, a wide therapeutic window for levels of circulating factor, ease of determining therapeutic endpoints, and existence of excellent animal models of the disease. Preclinical studies over the last decade have recently culminated in the initiation of clinical trials of gene transfer for hemophilia A and B. Three trials, each using different vectors and target tissues, are presently underway, and two additional trials are in late planning stages. This report reviews the preclinical data underlying these strategies and the design of the ongoing and proposed clinical trials.
Regional Differences in Integrin Expression
Authors: Davenpeck Kelly L. , Marcinkiewicz Cezary , Wang Dian , Niculescu Rodica , Shi Yi , Martin Jack L. , Zalewski Andrew
Published: June 2012

Abstract
—There is increasing evidence to suggest that coronary smooth muscle cells (SMCs) differ from noncoronary SMCs. As integrin adhesion molecules regulate many SMC functions, we hypothesized that differences in integrin expression on coronary and noncoronary SMCs may account for cellular differences. Analysis of integrin expression on freshly isolated porcine coronary and noncoronary SMCs revealed that coronary SMCs express significantly less α
5
β
1
than noncoronary SMCs, whereas the expression of total β
1
and that of α
v
β
3
are similar. Consistent with these findings, coronary SMCs demonstrated significantly less adhesion to fibronectin, compared with carotid artery SMCs. As α
5
β
1
-mediated signaling has been associated with cellular proliferation, the effects of differential α
5
β
1
expression on cell proliferation were examined by comparing primary coronary and carotid artery SMC proliferation. Coronary SMC growth was significantly lower than that of carotid artery SMCs when plated on fibronectin or type I collagen. Blocking α
5
β
1
function on carotid artery SMCs produced a significant decrease in cellular proliferation, resulting in growth similar to that of coronary SMCs. Furthermore, blocking α
5
β
1
, but not α
v
β
3
, inhibited loss of α-smooth muscle actin in proliferating SMCs. Proliferating coronary SMCs were found to upregulate α
5
β
1
expression, further indicating a role for α
5
β
1
in SMC growth. These results suggest that dissimilar α
5
β
1
integrin expression may mediate regional differences in phenotype of vascular SMCs.
Cyclin A/cdk2 Activation Is Involved in Hypoxia-Induced Apoptosis in Cardiomyocytes
Authors: Adachi Susumu , Ito Hiroshi , Tamamori-Adachi Mimi , Ono Yuichi , Nozato Toshihiro , Abe Shinji , Ikeda Masa-aki , Marumo Fumiaki , Hiroe Michiaki
Published: June 2012

Abstract
—Cardiomyocytes are terminally differentiated cells characterized as withdrawal cell-cycle machinery, but nonetheless they are known to express cell-cycle regulators. Because many proteins related to the cell cycle induce apoptosis in proliferating cells, we examined the involvement of these proteins in the apoptosis pathway in cardiomyocytes. Primary rat cardiomyocytes were exposed to a severe hypoxic condition to induce apoptosis. The apoptosis rate of cardiomyocytes increased to ≈40% under 24 hours of hypoxia as evaluated by the TUNEL method. The cyclin A protein level assessed by immunoblot analysis accumulated in a time-dependent manner in cardiomyocytes, but there was no increase in nonmyocytes. Hypoxia increased the activity of cyclin A–associated kinase but not the activity of cyclin E–associated kinase, and the apoptosis was inhibited by infection of dominant-negative cdk2 adenovirus, suggesting that cyclin A and its associated kinase play significant roles in the apoptosis of cardiomyocytes. To investigate the cyclin A–mediated apoptosis, we infected cultured cells with cyclin A adenovirus. Apoptosis was induced in 63±12% of the infected cardiomyocytes in contrast to only 12±3% of the LacZ-infected control cells. In addition, the cells in the hypoxic condition showed an increase in caspase-3 activity and a subsequent decrease in p21
cip1/waf1
protein, which is partly cleaved by caspase-3. These findings confirm that cyclin A–associated kinase mediates hypoxia-induced apoptosis in cardiomyocytes, and they also suggest that additional elements of the cell-cycle–dependent machinery participate in this mechanism.
Retinoids Induce Fibroblast Growth Factor-2 Production in Endothelial Cells via Retinoic Acid Receptor α Activation and Stimulate Angiogenesis In Vitro and In Vivo
Authors: Gaetano Carlo , Catalano Alfonso , Illi Barbara , Felici Angelina , Minucci Saverio , Palumbo Roberta , Facchiano Francesco , Mangoni Antonella , Mancarella Salvatore , Mühlhauser Judith , Capogrossi Maurizio C.
Published: June 2012

Abstract
—The effect of retinoic acid (RA) on endothelial cells is still controversial and was examined in the present study. In bovine aortic endothelial cells (BAECs),
all-trans
RA (ATRA) and 9-
cis
RA (9CRA), but not 13-
cis
RA (13CRA), induced fibroblast growth factor-2 (FGF-2) production and exhibited a biphasic dose-dependent effect to enhance BAEC proliferation and differentiation into tubular structures on reconstituted basement membrane proteins (Matrigel); both processes were inhibited by FGF-2–neutralizing antibody. The pan RA receptor (RAR)-selective ligand (E)-4-[2-(5,5,8,8,-tetramethyl-5,6,7,8-tetrahydro-2-naphtalenyl)-1-propenyl] benzoic acid and the RARα-selective ligand 4-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphtyl)-ethenyl] benzoic acid stimulated the production of FGF-2, whereas the addition of the RARα-antagonist RO 41-5253 inhibited this effect. In BAECs, the forced expression of RARα, but not RARβ or RARγ, enhanced FGF-2 production, whereas the RARα-dominant negative, Δ403, blocked this effect. Furthermore, RARα overexpression directly stimulated BAEC differentiation on Matrigel and potentiated the effects of ATRA in this assay. Finally, ATRA-treated BAECs coinjected with Matrigel subcutaneously in mice induced neovascularization within the Matrigel plug, and ATRA also enhanced angiogenesis in the chicken chorioallantoic membrane assay. In conclusion, RA can stimulate endothelial cell proliferation and differentiation in vitro via enhanced RARα-dependent FGF-2 production, and it can also induce angiogenesis in vivo. The full text of this article is available at http://www.circresaha.org.
I
f
Current and Spontaneous Activity in Mouse Embryonic Ventricular Myocytes
Authors: Yasui Kenji , Liu Weiran , Opthof Tobias , Kada Kenji , Lee Jong-Kook , Kamiya Kaichiro , Kodama Itsuo
Published: June 2012

Abstract
—Knowledge of the initiation of electrical and contractile activity in the embryonic heart relies to a large extent on data obtained in chicken. In recent years, molecular biological techniques have raised an interest in mouse physiology, including early embryonic development. We studied action potentials and the occurrence of one of the pacemaker currents,
I
f
, by the whole-cell voltage and current-clamp technique at the earliest stage at which a regular heartbeat is established (9.5 days postcoitum) and at 1 day before birth. We show, first, that at the early stage there is a prominent
I
f
in mouse embryonic ventricles, which decreases by 82% before birth in concert with the loss of regular spontaneous activity of ventricular cells. Second, the decrease in
I
f
current is associated with a slight change in channel gating kinetics and a decrease in total mRNA expression of the genes encoding for
I
f
current. Third, the most prevalent mRNA subtype is switched from HCN4 to HCN2 during the second half of embryonic development. Fourth, the
I
f
current may be modulated by the β-adrenergic cascade, although the coupling to the β-adrenoceptor in the sarcolemma itself is not yet mature. We conclude that
I
f
current of the sinus node type is present in early embryonic mouse ventricular cells. In association with a loss of
I
f
current, the ventricle tends to lose pacemaker potency during the second half of embryonic development.
Effects of Cholesterol-Fat Diets on Pigeons Susceptible and Resistant to Atherosclerosis
Authors: CLARKSON THOMAS B. , LOFLAND HUGH B.
Published: June 2012

Cholesterol-fat diets are markedly more atherogenic when fed to a strain of pigeons susceptible to spontaneous atherosclerosis, as compared to a strain resistant to the disease. A breed of pigeons intermediate in susceptibility was found to be intermediate in their response to a high cholesterol-high fat diet. Serum cholesterol levels were similar among the three breeds.

The White Carneau pigeons developed lesions complicated by connective tissue proliferation while the Autosexing Kings and Racing Homers developed merely "pure atheroma" or "fatty streaks."
Cardiovascular Dynamics of Vasovagal Reactions in Man
Authors: GREENE MURRAY A. , BOLTAX ADOLPH J. , ULBERG ROBERT J.
Published: June 2012

The dynamics of vasovagal reactions which occurred in 2 patients during right heart catheterization were studied. Both subjects initially had systemic hypertension with corresponding increases in peripheral resistances. Subject B had considerably decreased blood flow, moderately elevated pulmonary artery and right ventricular end-diastolic pressures, and elevated central blood volume. These parameters were normal in subject A.

The characteristic features of the reactions were appreciable declines in cardiac outputs coincident with decreased systemic arterial systolic, diastolic, and pulse pressures, decreased heart rates, and slight increases in calculated peripheral resistances. Stroke volumes were moderately reduced. A slight decrease in pulmonary artery systolic pressure occurred in subject A. In subject B, pulmonary artery and right ventricular enddiastolic pressures declined to normal and central blood volume decreased moderately. The return of heart rate to control values had no significant effects on the other physiologic parameters.

The data demonstrate that reductions in cardiac output, due primarily to decreased stroke outputs, were predominantly responsible for the arterial hypotensions during the vasovagal reactions in these 2 subjects. The ineffectiveness of atropine in restoring hemodynamics in one subject suggested the importance of inhibition of efferent sympathetic activity in the etiology of this reaction. The failure of peripheral resistance to compensate for decreased blood flow suggested inhibition of arteriolar tone.
Potentiation of Smooth Muscle Contraction by Adrenal Steroids
Authors: FOWLER NOBLE O. , CHOU NORA H.F.
Published: June 2012

The rabbit aortic strip preparation of Furchgott was used to test the effect of adrenal steroids upon the muscle contraction produced by 0.1 to 0.2
µ
g. of levarterenol. The steroids tested were corticosterone, hydrocortisone, aldosterone acetate, androsterone, and dehydroisoandrosterone. Amounts of 1 to 300
µ
g. were added to the 20-ml. muscle bath. Potentiation was produced by each steroid; the percentage increase of amplitude of contraction was from 36.1 to 74.2 percent. Corticosterone produced consistent potentiation in amounts as small as 5
µ
g.; 200
µ
g. of aldosterone acetate was required to produce potentiation. Maximum potentiation was produced by hydrocortisone. The observed potentiation may be due to an effect of adrenal steroids upon transcellular sodium and potassium gradients.
Effect of Changes in Frequency of Stimulation Upon Rabbit Ventricular Action Potential
Authors: GIBBS COLIN L. , JOHNSON EDWARD A.
Published: June 2012

It has been demonstrated that in rabbit ventricular muscle, the area of the action potential is at a maximum at a stimulus frequency ranging from 1 to 3 sec.
-1
. At frequencies on either side of this range the action potential decreases in area. The changes in action-potential shape at high rates of stimulation are caused by an earlier "take off" of the fast phase of repolarization, and at low rates of stimulation the changes in shape are predominantly caused by a drop in the height of the plateau. It has also been shown that the ratio of the area of the extra action potential to the area of the normal action potential depends upon the stimulus frequency and the interval between the normal and extra action potentials. Upon any chauge in the stimulus frequency, the area of the ventricular action potential overshoots its final value before stabilizing. The shape of the action potential is dependent upon the previous history of the muscle. By altering the frequency of stimulation, it has also been shown that the changes in action-potential shape at low frequencies of stimulation, in contrast to those at high frequencies, are very rapid and take place immediately when the basic stimulus frequency is altered.
Pharmacological Evidence for the Importance of Catecholamines in Cardiac Rhythmicity
Authors: ROBERTS JAY , MODELL WALTER
Published: June 2012

Dogs with complete heart block were used to explore the dependence of the intrinsic rhythmicity of the heart on catecholamine activity. By the use of reserpine, hexamethonium, and surgical sympathectomy, it was demonstrated that, while reduction in catecholamine activity slows both the atrial and the ventricular rates, the ventricular pace maker was always affected to a greater degree. The effect of a combination of reserpine and hexamethonium on the ventricle was greater than when either agent was administered alone, suggesting that the greater the sympathetic blockade, the greater the depression in pacemaker rhythmicity. Since the course of its action on the heart paralleled that of catecholamine depletion, it seems probable that the effect of reserpine on rhythmicity was due to its catecholamine depleting action. It was thought unlikely that prolonged hypotension contributed to the reserpine action on the heart. It is indicated that disturbances in electrolyte balance and blood pH did not play a role in the effects observed.
Quantitative Adsorption of Antibody by the Isolated Heart and the Intensity of Cardiac Anaphylaxis
Authors: FEIGEN GEORGE A. , VUREK GERALD G. , IRVIN WILLIAM S. , PETERSON JANICE K.
Published: June 2012

The present experiments show that the adsorption of antibodies to the guinea pig heart can be achieved by perfusing the heart with rabbit antiovalbumin solutions. The amount of radioactive antibody remaining on the tissue is a definite function of the concentration of antibody in the bulk phase instilled into the coronary circulation. When the antibody-loaded organ is challenged with ovalbumin, the efflux rate of antibody is increased; it is also increased in response to such nonspecific agents as histamine and epinephrine. Since the output rate is not affected when the heart fails to react to rechallenge with antigen, the release of antibody may be entirely dependent upon the mechanical response.

The maximal amount.it of histamine-like material that can be produced by the heart undergoing anaphylaxis was found to be 1 x 10
-s
moles/Gm./min., estimated as histamine; the minimal amount of antibody necessary to release this quantity was calculated to be 0.343 µg. of specific antiovalbumin/100 mg. of heart. This load can be achieved by perfusing the heart with an antibody solution containing 30 µg. of specific γ-globulin. Perfusion with an antibody solution containing 50 µg./ml. showed the amount of histamine subsequently liberated by the same dose of ovalbumin to be reduced by 50 per cent.

The rate and amplitude of contraction were increased to the same absolute values in all cases in which either ovalbumin or histamine was employed to set off the reaction, suggesting that the release of active material at even the low sensitizing doses of antibody used exceeded the ceiling of the physiological response. The decrease in coronary flow was inversely related to the dose of sensitizing antibody and bore little relation to the changes in this parameter induced by the subsequent administration of histamine.
Concealed Conduction
Authors: HOFFMAN BRIAN F. , CRANEFIELD PAUL F. , STUCKEY JACKSON H.
Published: June 2012

The existence of concealed conduction of atrial beats has been demonstrated. Concealed conduction within the atrioventricular node has been studied by recording the transmembrane potential of single nodal fibers. Conduction block in this part of the specialized conducting system depends more upon decremental conduction within certain nodal fibers than upon differences in action-potential duration. Also, the extreme delay of activation that results from such decremental conduction explains certain changes in atrioventricular delay associated with premature atrial beats as well as a prolonged refractoriness of atrioventricular transmission after concealed nodal conduction of an atrial extrasystole.

In the in situ dog heart, local bipolar electrograms recorded from various parts of the bundle of His, bundle branches, and peripheral Purkinje fibers have shown that con cealed conduction may extend through the atrioventricular node and the bundle of His and that block may occur between common bundle and bundle branches or between the latter and the peripheral Purkinje fiber. Block in these locations, which results from local differences in the action-potential duration, has been compared to decremental conduction within the atrioventricular node.
Testicular Action and Structural Relationships of Compounds with Antiatherogenic Activity
Authors: MALINOW MANUEL RENÉ , MOGUILEVSKY JAIME A. , LEMA BALTAZAR , BUR GRATO E. , ERENFRYD AARÓN
Published: June 2012

Cholesterol-fed chickens have been injected subcutaneously for eight weeks with the following drugs: estradiol, diethylstilbestrol, estriol, androstanolone, δ5-androstenediol and androstanediol. Aortic and coronary atherosclerosis, blood and aortic cholesterol, as well as several endocrine structures have been studied. Coronary atherosclerosis has been prevented by estradiol and by diethylstilbestrol, while the other steroids have been ineffective at the dosage used here. When birds were considered individually, no correlation was found between antiatherogenic activity and endocrine changes, thus confirming that testicular depression is not necessary for arterial protection. An attempt to correlate molecular structure and arterial changes has been carried out.
Disposal of Intravenously Administered Fat in Subjects with Atherosclerosis and in Normal Controls
Authors: MASHFORD M. L. , NESTEL P. J.
Published: June 2012

A. series of experiments has been performed in which a standard dose of an artificial oil emulsion (Lipomul I.V.) was given to 5 groups of subjects and the disappearance of fat from their blood was observed by plasma optical density measurements. The groups were age-matched and consisted of a control group; 3 groups who had suffered a myocardial infarction and were respectively untreated, receiving heparin and receiving phenoprocoumon after recovery from the acute stage of the illness; and the last group which consisted of patients who suffered from intermittent claudication. The control, untreated infarct and peripheral vascular disease groups exhibited similar curves of optical density on time. The mean peak optical density was significantly lower in the heparin-treated than in the untreated group. Conversely, the mean peak optical density of the phenoprocoumontreated group was significantly higher than that of the untreated or control groups.
Localization and Function of the Medullary Vasomotor Center in Infracollicularly Decerebrated Cats
Authors: LINDGREN PERCY
Published: June 2012

The medullary vasomotor centers were studied in three cats, decerebrated below the mesencephalon four to nine days before the $$$cute experiment. Exploration of the dorsal regions of the medulla oblongata revealed that the vasomotor active regions had about the same extension as in intact animals. Medullary pressor and depressor reflexes, elicited by electrical stimulation of a peripheral nerve and a sinus nerve, were also of a normal type in the decerebrated cats. The results indicate that the medullary vasomotor center constitutes a region of nuclear character with cell bodies and synapses functioning as a tonic and reflex center. In this respect, it seems to function normally even after the degeneration of vasomotor pathways from higher centers.