Integrins, vascular remodeling, and hypertension.
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Animals↗
Biomedical subjects
Anthony M Heagerty
Publications and source records attributed to Anthony M Heagerty.
Identification of Tctex2beta, a novel dynein light chain family member that interacts with different transforming growth factor-beta receptors.
Endoglin is a membrane-inserted protein that is preferentially synthesized in angiogenic vascular endothelial and smooth muscle cells. Endoglin associates with members of the transforming growth factor-beta (TGF-beta) receptor family and has been identified as the gene involved in hereditary hemorrhagic telangiectasia. Although endoglin is known to affect cell responses to TGF-beta, its mode of action is largely unknown. We performed yeast two-hybrid screening of a human placental cDNA library and isolated a new endoglin-binding partner, a novel 221-amino acid member of the Tctex1/2 family of cytoplasmic dynein light chains named Tctex2beta, as the founder of a new Tctex1/2 subfamily. The interaction was localized exclusively to the cytoplasmic domain of endoglin. Reverse transcription-PCR showed expression of Tctex2beta in a wide range of tissues, including vascular endothelial and smooth muscle cells, placenta, and testis, as well as in several tumor cell lines. High expression levels were found in human umbilical vein endothelial cells and the large cell lung cancer cell line. Forced expression of Tctex2beta had a profound inhibitory effect on TGF-beta signaling. Additional Tctex2beta-interacting receptors were identified to be the TGF-beta type II receptor and most likely beta-glycan, but not ALK5, ALK1, or the bone morphogenetic protein type II receptor. Upon fluorescence tagging, co-localization of Tctex2beta and endoglin, as well as Tctex2beta, endoglin, and the TGF-beta type II receptor, was observed by different microscopy techniques. Our findings link endoglin for the first time to microtubule-based minus end-directed transport machinery, suggesting that some endoglin functions might be regulated and directed by its interaction with the cytoplasmic dynein light chain Tctex2beta.
Amino Acid Sequence↗
Beta-adrenoceptor blockade markedly attenuates transgene expression from cytomegalovirus promoters within the cardiovascular system.
OBJECTIVE: The major immediate-early cytomegalovirus enhancer/promoter (MIECMV), widely used in cardiovascular gene therapy, contains several positively regulatory cAMP response elements (CRE). Catecholamine signaling via beta-adrenoceptors might increase transgene expression from MIECMV, and if so, beta-blockers may have a detrimental effect on the efficacy of clinical cardiovascular gene therapy strategies. METHODS AND RESULTS: Cultured smooth muscle cells were exposed to isoprenaline, atenolol, or propranolol, alone and in combination before infection with adenoviruses expressing beta-galactosidase. beta-galactosidase expression was assayed 72 hours later. Isoprenaline increased transgene expression from MIECMV up to 8-fold (P<0.001), but had no effect on a promoter containing no CRE. The effect of isoprenaline was inhibited by beta-blockade and by specific CRE-decoy oligonucleotides. Beta-blockers did not reduce transgene expression below basal levels. After adenovirus-mediated porcine intracoronary gene transfer, however, beta-blockade reduced beta-galactosidase expression by up to 250-fold compared with non-beta-blocked animals (P<0.01). CONCLUSIONS: Enhancement of promoter activity by endogenous catecholamines is essential for high-level transgene expression from MIECMV within the vasculature. Beta-blocker-mediated suppression of transgene expression from MIECMV in vascular tissues has a significant bearing on clinical studies of cardiovascular gene transfer. This is the first described interaction to our knowledge between widely prescribed pharmaceuticals and a commonly used promoter of clinical transgene expression.
Adrenergic beta-Antagonists↗
Adenosine- and hypoxia-induced dilation of human coronary resistance arteries: evidence against the involvement of K(ATP) channels.
1. The ATP-sensitive potassium (K(ATP)) channel may be an important mediator of metabolic dilation in the human coronary circulation. As adenosine and hypoxia are considered to be major mediators of metabolic dilation in the coronary circulation, we investigated the effect of glibenclamide (a K(ATP) channel blocker) on adenosine and hypoxic dilation of human coronary resistance arteries, with myogenic tone, in vitro. 2. Vessels were dissected from the atrial appendage from consenting patients and studied in vitro using a pressure arteriograph system. Segments of coronary resistance artery were pressurized to 60 mmHg and the vessels studied developed spontaneous myogenic tone. 3. The K(ATP) opener pinacidil (final conc. 5 x 10(-6) M) resulted in dilation, which was completely reversed by 5 x 10(-6) glibenclamide (84+/-14 vs -10+/-9%, pinacidil and pinacidil plus glibenclamide, respectively, P=0.009, n=5). 4. Adenosine (final conc. 10(-5) M) resulted in dilation, glibenclamide (5 x 10(-6) and 10(-5) M) was without effect (118+/-12 vs 104+/-16% adenosine and adenosine plus 10(-5) glibenclamide, respectively, n.s., n=4). 5. Hypoxia (8+/-3 mmHg O2) resulted in a dilation that reversed when normoxic conditions were restored (60+/-9 vs 3+/-11% hypoxia and post-hypoxia, respectively, P=0.014, n=3). The hypoxic dilation was not affected by glibenclamide (63+/-14 vs 55+/-6% hypoxia and hypoxia plus glibenclamide, respectively, n.s., n=4). In a further series of experiments, vessels were incubated with glibenclamide (5 x 10(-6)) prior to a hypoxic challenge; again, glibenclamide was without effect on the hypoxic dilation (-0.008+/-2 vs 95+/-3% glibenclamide and glibenclamide plus hypoxia, respectively, P=0.0005, n=3). 6. These data demonstrate that glibenclamide is without effect on both adenosine and hypoxic dilation of human coronary resistance arteries with myogenic tone, from the right atrial appendage in vitro. Our findings suggest that the K(ATP) channel is unlikely to be a major mediator of metabolic dilation in these arteries.
Adenosine↗
Investigation of the role of TASK-2 channels in rat pulmonary arteries; pharmacological and functional studies following RNA interference procedures.
In the present study, we investigated the ability of RNA interference technology to suppress TASK-2 potassium channel expression in human embryonic kidney (HEK293) cells stably transfected with TASK-2 cDNA and in rat isolated intact pulmonary arteries. Lipofectamine-induced transfection of a specific siRNA sequence targeted against TASK-2 resulted in a dose- and time-dependent decrease in TASK-2 channel protein expression. In siRNA-transfected cells the TASK-2 peak currents were significantly smaller than in control cells at every investigated pH, while the pH sensitivity was not altered. Using scrambled siRNA as a negative control, there were no significant changes in TASK-2 protein expression or current compared to mock-transfected cells. In TASK-2 siRNA-transfected small pulmonary arteries, but not in scrambled siRNA-treated vessels, myocyte resting membrane potential at pH 7.4 was significantly less negative and the hyperpolarisations in response to increasing pH from 6.4 to 8.4 were significantly smaller compared with control. The application of levcromakalim (10 microM), NS1619 (33 microM) and a potassium channel inhibitor cocktail (5 mM 4-aminopyridine, 10 mM tetraethylammonium chloride, 30 microM Ba2+ and 10 microM glibenclamide) had similar effects in control and in siRNA-transfected vessels. The TASK-1 (anandamide-sensitive) contribution to resting membrane potential was comparable in each group. Clofilium (100 microM) generated significantly smaller responses in transfected artery segments. These results suggest that RNA interference techniques are effective at inhibiting TASK-2 channel expression in cultured cells and in intact vessels and that TASK-2 channels have a functional role in setting the membrane potential of pulmonary artery myocytes.
Animals↗
Middle cerebral artery structure and distensibility during developing and established phases of hypertension in the spontaneously hypertensive rat.
OBJECTIVE: The aims of the current study were to examine the structural properties of middle cerebral arteries (MCA) from young (5-7 weeks) and adult (20-24 weeks) spontaneously hypertensive rats (SHR), compared with age-matched Wistar-Kyoto (WKY) control rats. DESIGN: MCA segments (8-10 per group) were secured onto glass pipettes in a small vessel chamber and studied using a pressure arteriograph system. Vessels were perfused in Ca2+-free physiological salt solution to ensure the absence of tone. The wall thickness and lumen diameter were recorded at intraluminal pressures ranging from 3 to 180 mmHg using a video dimension analyser. RESULTS: There was a borderline increase in systolic pressure of the young SHR, compared with WKY controls, but the systolic pressure of the older SHR was significantly raised. The MCA lumen diameter from young SHR was reduced across the entire pressure range and arterial distensibility was not reduced, compared with WKY vessels. The MCA lumen diameter from adult SHR was reduced at high pressure, but converged with the lumen diameter of the WKY vessels at 3 mmHg, and the stress-strain relation was shifted to the left, compared with the WKY vessels; nevertheless, the slope of the tangential elastic modulus-stress relation was not significantly increased. The pressure-wall cross-sectional area relationship did not differ between strains at either time point. CONCLUSIONS: These data demonstrate eutrophic inward remodelling of the MCA from young SHR, compared with WKY controls. In the adult SHR the structural changes are probably a consequence of a reduced arterial distensibility.
Aging↗
alphaV integrins are necessary for eutrophic inward remodeling of small arteries in hypertension.
Human essential hypertension is characterized by eutrophic remodeling of small arteries, with little evidence of hypertrophy. Likewise, vessels of young hypertensive TGR(mRen2)27 animals have undergone similar structural alterations. The role of integrins in resistance arteries of TGR(mRen2)27 during the eutrophic-remodeling process was examined as blood pressure rose. Initially, 8 alpha and 3 beta integrins were identified and levels of expression investigated using RT-PCR. As pressure increased and remodeling advanced, integrin expression profiles revealed that only alphaV was significantly raised. In conjunction, we confirmed elevated integrin alphaV protein levels in TGR(mRen2)27 rat arteries and localization to the media using immunofluorescence. beta1 and beta3, but not beta5 integrin subunits were coprecipitated with integrin alphaV and are implicated in the eutrophic remodeling process. Administration of a peptide antagonist of alphaVbeta3 abolished remodeling but enhanced growth, indicating that hypertrophy supervened as a response to hypertension-induced increases in wall stress. We have established that the only upregulated integrin, the alphaV subunit of integrin alphaVbeta3, has a crucial role in the hypertensive remodeling process of TGR(mRen2)27 rat resistance arteries. During hypertensive remodeling, functions of specific alphaVbeta3-extracellular matrix interactions are likely to allow vascular smooth muscle cell-length autoregulation, which includes a migratory process, to maintain a narrowed lumen after a prolonged constricted state.
Animals↗
Identification and characterization of vascular calcification-associated factor, a novel gene upregulated during vascular calcification in vitro and in vivo.
OBJECTIVE: Vascular calcification, with its increasing clinical sequelae, presents an important and unresolved dilemma in cardiac and vascular practice. We aimed to identify molecules involved in this process to develop strategies for treatment or prevention. METHODS AND RESULTS: Using subtractive hybridization, a novel cDNA, designated vascular calcification-associated factor (VCAF), has been isolated from a bovine retinal pericyte cDNA library generated during the differentiation and mineralization of these cells in vitro. RNA ligase-mediated rapid amplification of cDNA ends was used to compile the 740-bp bovine cDNA sequence. Database searching reveals that VCAF has novel nucleotide/amino acid sequences. RNA analysis confirms that VCAF is upregulated in mineralized pericytes and is present in human calcified arteries but not noncalcified arteries. Protein analysis using a VCAF antibody confirms the presence of an 18-kDa protein in calcified nodules but not in confluent pericytes. Adenoviral antisense VCAF gene delivery reduces VCAF protein levels and accelerates pericyte differentiation compared with controls. CONCLUSIONS: We demonstrate the isolation of a novel gene, VCAF, which is upregulated during vascular calcification in vitro and in vivo. Antisense VCAF gene delivery accelerates pericyte differentiation, implicating a role for VCAF in this clinically significant pathological process.
Adenoviridae↗
Influence of changes of blood pressure on vascular angiotensin II receptor subtype expression.
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Animals↗
Effects of angiotensin type-1 receptor antagonism on small artery function in patients with type 2 diabetes mellitus.
Endothelial dysfunction has been demonstrated to occur in small arteries from patients with type 2 diabetes and hypertension. The effects of angiotensin II receptor blockade on vessel function were examined using pressure myography in a randomized 12-week double-blind placebo-controlled parallel group study using candesartan cilexitil. The maximal vascular response to acetylcholine (Ach) was impaired at baseline and improved with candesartan. This improvement was primarily caused by an effect in the nitric oxide component of Ach-mediated dilatation. The degree of endothelial dysfunction directly correlated with serum low-density lipoprotein cholesterol levels. Sodium nitroprusside-induced endothelium-independent dilatation was reduced in diabetic patients and intervention with candesartan lead to an improvement in EC50 with no change in maximal response. Vasoconstriction to norepinephrine was normal and did not change with intervention, but responses to angiotensin II were reduced after candesartan in diabetic patients. These results demonstrate that even brief treatment with angiotensin II receptor blockade is associated with a significant improvement in resistance vessel endothelial function.
Acetylcholine↗
Roles of large and small arteries in vascular disease: pitfalls in interpreting ethnic variations.
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Arteries↗
Small artery structure and hypertension: adaptive changes and target organ damage.
Hypertension is known to be associated with an increase in the wall/lumen ratio (W/L) of the resistance arteries. Growth, eutrophic remodelling and changes in arterial distensibility can all contribute to an increase W/L. Wall stress may stimulate growth, whereas remodelling and/or reduced distensibility may be the result of prolonged contraction. Impaired or overwhelmed pressure-induced myogenic constriction, causing increased wall stress, may explain the small artery growth seen in diabetes and secondary hypertension, respectively. We find that small artery growth is a greater predictor of cardiovascular events than an increased W/L.
Adaptation, Physiological↗
Adenovirus-mediated gene transfer of transforming growth factor-beta3, but not transforming growth factor-beta1, inhibits constrictive remodeling and reduces luminal loss after coronary angioplasty.
BACKGROUND: Extracellular matrix (ECM) remodeling is central to the development of restenosis after PTCA. Substantial evidence implicates transforming growth factor-beta1 (TGF-beta1), a regulator of ECM deposition by vascular cells, in its pathogenesis. TGF-beta3 reduces TGF-beta1-induced ECM deposition in cutaneous wounds. We therefore investigated the effects of intracoronary expression of TGF-beta3 and TGF-beta1 on luminal loss after angioplasty. METHODS AND RESULTS: Porcine coronary arteries received an adenovirus expressing TGF-beta3, TGF-beta1, or lacZ (beta-galactosidase), or PBS only, at the site of angioplasty. Morphometric analysis 28 days after angioplasty confirmed reduced luminal loss in TGF-beta3 vessels (-0.65+/-0.10 mm2) compared with lacZ (-1.18+/-0.19 mm2) or PBS only (-1.19+/-0.17 mm2; P=0.003). Luminal loss was not reduced in TGF-beta1 vessels (-1.02+/-0.19 mm2; P=0.48). An increase in the external elastic lamina area in TGF-beta3-treated vessels (+0.73+/-0.32 mm2) contrasted with decreases in control vessels (mean, -0.53+/-0.17 mm2; P=0.001) and TGF-beta1 vessels (-0.87+/-0.34 mm2; P=0.003). Collagen content increased at the site of injury in TGF-beta3-treated vessels (26.1+/-14.2%) but decreased in the lacZ (-22.8+/-6.6%) and PBS-only (-23.4+/-7.0%; P=0.002) groups and was not significantly changed in TGF-beta1-treated vessels. CONCLUSIONS: Expression of TGF-beta3 inhibits constrictive remodeling after PTCA and reduces luminal loss. This is accompanied by increased adventitial collagen, which may act as an external "scaffold" preventing vessel constriction. These findings confirm the potential of gene therapies that modify ECM remodeling for prophylaxis of restenosis.
Adenoviridae↗
Myogenic and structural properties of cerebral arteries from the stroke-prone spontaneously hypertensive rat.
The aims of the study were to compare the myogenic and structural properties of middle cerebral arteries (MCAs) from the stroke-prone spontaneously hypertensive rat (SHRSP) with MCAs from the spontaneously hypertensive rat (SHR) before stroke development in SHRSP. Rats were fed a "Japanese" diet (low-protein rat chow and 1% NaCl in drinking water) for 8 wk, and cerebral arteries were studied in vitro at 12 wk using a pressure arteriograph. Systolic pressure was significantly increased in SHRSP compared with SHR at 12 wk. Between 60 and 180 mmHg, MCAs from SHR maintained an essentially constant diameter, i.e., displayed a "myogenic range," whereas the diameter of MCAs from SHRSP progressively increased as a function of pressure. Passive lumen diameter of MCAs from SHRSP was reduced at high pressure, and wall thickness and wall/lumen were increased, compared with SHR. Wall cross-sectional area was also increased in MCAs from SHRSP compared with the SHR, indicating growth. The stress-strain relationship was shifted to the left in MCAs from SHRSP, indicating decreased MCA distensibility compared with SHR. However, collagen staining with picrosirius red revealed a redistribution of collagen to the outer half of the MCA wall in SHRSP compared with SHR. These data demonstrate impaired myogenic properties in prestroke SHRSP compared with SHR, which may explain stroke development. The structural differences in MCAs from SHRSP compared with SHR were a consequence of both growth and a reduced distensibility.
Animals↗
Ongoing trials: what should we expect after ALLHAT?
The publication of the Antihypertensive and Lipid-lowering Treatment to Prevent Heart Attack Trial (ALLHAT) results, when used in conjunction with a new meta-analysis, provides hypertension research with a watershed. Data demonstrate clearly that lowering blood pressure is the most important aspect of hypertension management, and that all agents are similarly effective. There is little sustainable evidence for pressure-independent advantages for any class of drug at this time. Therefore, by combining these findings with additional information on target levels of blood pressure and safety, plus the use of aspirin and statins, we can formulate health care policies that attack hypertension in a holistic risk-based way. The future will focus on gaining extra benefit for patients by testing the efficacy and superiority of drug combinations using novel agents that favorably influence intermediate markers such as arterial stiffness, and arresting the ever-increasing burden of cognitive impairment and dementia.
Antihypertensive Agents↗
Expression of latent TGF-beta binding proteins and association with TGF-beta 1 and fibrillin-1 following arterial injury.
OBJECTIVES: Transforming growth factor-beta (TGF-beta), a potent regulator of wound healing and scar formation, is thought to have a key role in the response to arterial injury. Latent TGF-beta binding proteins (LTBPs), members of the fibrillin superfamily, govern TGF-beta1 release, targeting and activation in vitro and also play a role as structural components of fibrillin-rich microfibrils. Despite the potential of LTBPs to modulate the response to arterial injury through either or both of these mechanisms, as yet their expression and function in the injured vasculature remain poorly defined. METHODS: In this study, a porcine model of coronary angioplasty was used to investigate LTBP-1 and LTBP-2 synthesis and their association with TGF-beta 1 and fibrillin-1. RESULTS: After angioplasty, increased LTBP-1 and LTBP-2 immunostaining was detected in a similar distribution to increased TGF-beta 1 expression in the neointima and in the neoadventitia. Overnight organ cultures revealed the formation of large latent TGF-beta 1 complexes containing LTBP-1. Increased LTBP-1 proteolysis after arterial injury correlated with increased active and latent TGF-beta levels. LTBP-2 synthesis increased in response to arterial injury but was neither present in large latent complexes nor proteolytically processed. LTBP-1 and LTBP-2 both co-localised to fibrillin-rich fibrillar structures in the neointima and adventitia. CONCLUSIONS: These data suggest that LTBP-1 may have a TGF-beta 1 binding role in the arterial response to injury, and that LTBP-1 and LTBP-2 may have a structural role in association with microfibrils within the developing neointimal lesion. LTBP-1 proteolysis is potentially an important regulatory step for TGF-beta activation in the vasculature and inhibition of proteolysis could represent a novel therapeutic modality for controlling the arterial injury response.
Angioplasty, Balloon, Coronary↗
The amplifier hypothesis: persisting dissent.
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Animals↗