Role of signal transducers and activators of transcription 1 and -3 in inducible regulation of the human angiotensinogen gene by interleukin-6.

The circulating level of angiotensinogen (AGT) is dynamically regulated as an important determinant of blood pressure and electrolyte homeostasis. Because the mechanisms controlling the regulated expression of human angiotensinogen (hAGT) are unknown, we investigated the inducible regulation of the hAGT gene in well differentiated HepG2 cells. Interleukin-6 (IL-6) stimulation produced a 3.2-fold increase in hAGT mRNA peaking at 96 h after stimulation. Deletional mutagenesis of the hAGT promoter in transient transfection assays identified an IL-6 response domain between nucleotides -350 and -122 containing three reiterated motifs, termed human acute phase response elements (hAPREs). Although mutation of each site individually caused a fall in IL-6-inducible luciferase activity, mutation of all three sites was required to block the IL-6 effect. Electrophoretic mobility shift assay (EMSA), supershift, and microaffinity DNA binding assays indicate IL-6-inducible high-affinity binding of signal transducers and activators of transcription 1 and -3 (STAT1 and -3) to hAPRE1 and -3 but only low-affinity binding to hAPRE2. Expression of a dominant-negative form of STAT3, but not STAT1, produced a concentration-dependent reduction in IL-6-induced hAGT transcription and endogenous mRNA expression. These data indicate that STAT3 plays a major role in hAGT gene induction through three functionally distinct hAPREs in its promoter and suggest a mechanism for its up-regulation during the acute-phase response.

A nalysis of Transcriptional Control Mechanisms I : Techniques for Characterization ofcis-Regulatory Elements.

Transcriptional control, the process controlling when and how much RNA is produced from a DNA template, is a major determinant of gene expression in eukaryotic cells. This process, intensely studied over the last few decades, is under control of specific DNA sequences (cis elements) that function by virtue of their ability to be recognized by sequence-specific DNA-binding proteins (trans-acting elements). Both of these elements function in concert to control the rate and location of RNA transcript formation. Therefore, identification of these cis- and trans- acting elements provides important mechanistic insight into gene expression control. These studies are relevant to understanding aspects of the renin-angiotensin system. For example, a large body of evidence has shown that angiotensinogen (AGT) is a highly inducible gene, regulated by a variety of physiological hormone systems. Because AGT circulates close to its Michaelis-Menten constant (Km) for renin, changes in AGT concentration influence the long-term activity of the RAS [Reviewed in (1)].

Analysis of Transcriptional Control Mechanisms II Techniques for Characterization of trans-Acting Factors.

cis-Acting DNA control elements, enhancers and promoters, function to control gene expression by acting as targets for specific DNA-binding proteins (trans-acting factors). trans-acting factors are sequence-specific DNA-binding proteins that recognize specific signatures in base composition of the DNA, and upon binding, are able to influence transcriptional activity of the core promoter by multiple diverse mechanisms. These mechanisms include direct interaction with the preinitiation complex, recruitment of additional bridging proteins (coactivators), or induction of chromatin remodeling (such as altering nucleosomal phasing). These mechanisms are coordinated to result in changes in gene expression that control critical events in cellular differentiation and responses to extracellular signals or stressors.

Transcriptional regulation of angiotensinogen gene expression.

The renin--angiotensin system (RAS) is an extracellular hormonal system implicated in acute, homeostatic control of peripheral vascular resistance and electrolyte homeostasis. In this tightly regulated system, physiological regulators of blood pressure and fluid balance induce the production of the potent vasoactive angiotensin peptides by sequential proteolysis of the angiotensinogen (AGT) prohormone. AGT is the only known precursor of the angiotensin peptides, whose circulating concentrations influence the tonic activity of the RAS. AGT abundance is regulated at the transcriptional level through hormonal and cell-type specific regulators. In this review, we will discuss the identified mechanisms controlling AGT expression separately for the rodent and human genes. The most intensively investigated gene (rodent AGT) is regulated constitutively by multiple positive- and negative-acting cis factors that function in a cell-type dependent fashion. Inducible rodent AGT expression is mediated through a multihormone-inducible enhancer that integrates signals from steroid and cytokine hormones into AGT transcription. We review recent advances in understanding the mechanism of the nuclear factor-kappa B (NF-kappa B) family in mediating cytokine-induced AGT expression and our recent discoveries on the existence of differentially inducible pools of cytoplasmic NF-kappa B. Constitutive control of the human AGT gene will be discussed; there is surprisingly little information on the cis- and trans-acting regulators controlling inducible expression of human AGT. Finally, we will explore some of the recent developments in gene linkage studies where human AGT alleles have been associated with hypertensive phenotypes through a mechanism that may involve enhanced transcription. These studies have provided a molecular explanation for a subset of heritable hypertensive disorders in humans.

Transesophageal echocardiography during radiofrequency catheter ablation of ventricular tachycardia.

Radiofrequency lesion formation requires stable catheter tip/endocardial contact. Energy delivery is limited when temperatures are > 100 degrees C due to coagulum formation at the catheter tip. Transesophageal echocardiographic imaging may be useful for monitoring catheter position and detecting boiling. Transesophageal echocardiographic images were recorded during production of 22 radiofrequency lesions in bovine myocardium in a saline bath. Lesion size, tissue temperature and appearance of echo contrast (bubbles) were assessed. In 11 patients, transesophageal echocardiography was used to guide catheter movement and detect boiling during radiofrequency ablation for ventricular tachycardia. In the tissue bath, the appearance of echo bubbles was associated with visual bubbling at the catheter tip, tissue temperatures > 60 degrees C and larger lesions (284 +/- 165 vs 30 +/- 54 mm3; p < 0.001). In humans, transesophageal images easily identified the catheter tip in either ventricle and enabled continuous observation of electrode-tissue contact during radiofrequency application. Transesophageal echocardiographic bubbles appeared in 59 of 217 radiofrequency applications (27%). Continued radiofrequency application after appearance of bubbles was followed by an increase in impedance. Prolonged placement of the probe in heavily sedated patients resulted in a mild sore throat, but no other complications. Transesophageal echocardiographic imaging enables continuous monitoring of catheter position during radiofrequency energy application. The abrupt appearance of echo bubbles indicates boiling and impending coagulum formation at the catheter tip.

Complete heart block complicating retrograde left-heart catheterization of patients with cardiac allografts.

The development of complete heart block during left-sided cardiac catheterization is an uncommon event. We describe three cases of complete heart block complicating left-sided cardiac catheterization of patients with cardiac allografts. Review of the cases performed at our institution suggests that complete heart block during left-sided catheterization may be more common in patients with cardiac allografts.

Ventricular tachycardia after infarction: sources of coronary blood flow to the infarct zone.

The purpose of this study was to determine the sources of coronary blood flow to infarct scars in patients with sustained ventricular tachycardia occurring late after myocardial infarction, which is necessary for transcoronary sclerosis or embolization. Angiograms of 32 consecutive patients (age 63 +/- 8 years, ejection fraction 0.30 +/- 0.10) were reviewed. Sources of blood flow to the infarct zone were identified as coming from a recanalized infarct-related artery, side branch, collateral, or coronary bypass graft. Eighty-four percent of patients in the study had an identifiable blood supply to the area of previous infarction. More than one source of blood flow to anterior infarct locations were observed more often than to inferior infarct locations (53% vs 17%, p = 0.03). Transcoronary mapping for possible chemical ablation should be technically feasible in the majority of patients with ventricular tachycardia. Infarct zone blood flow arises from any of several sources and varies somewhat depending on infarct location.

Metabolic and functional recovery of ischemic human myocardium after coronary angioplasty.

Although revascularization of hypoperfused but metabolically active human myocardium improves segmental function, the temporal relations among restoration of blood flow, normalization of tissue metabolism and recovery of segmental function have not been determined. To examine the effects of coronary angioplasty on 13 asynergic vascular territories in 12 patients, positron emission tomography and two-dimensional echocardiography were performed before and within 72 h of revascularization. Ten patients underwent late echocardiography (67 +/- 19 days) and eight underwent a late positron emission tomographic study (68 +/- 19 days). The extent and severity of abnormalities of wall motion, perfusion and glucose metabolism were expressed as wall motion scores, perfusion defect scores and perfusion-metabolism mismatch scores. Angioplasty significantly increased mean stenosis cross-sectional area (from 0.95 +/- 0.9 to 2.7 +/- 1.4 mm2) and mean cross-sectional luminal diameter (from 0.9 +/- 0.6 to 1.9 +/- 0.5 mm) (both p less than 0.001). Perfusion defect scores in dependent vascular territories improved early after angioplasty (from 116 +/- 166 to 31 +/- 51, p less than 0.002) with no further improvement on the late follow-up study. The mean perfusion-metabolism mismatch score decreased from 159 +/- 175 to 65 +/- 117 early after angioplasty (p less than 0.01) and to 26 +/- 29 at late follow-up (p less than 0.001 vs. before angioplasty; p = NS vs. early after angioplasty). However, absolute rates of glucose utilization remained elevated early after revascularization, normalizing only at late follow-up.(ABSTRACT TRUNCATED AT 250 WORDS)

Asymptomatic myocardial ischemia during percutaneous transluminal coronary angioplasty and importance of prior Q-wave infarction and diabetes mellitus.

Eighty-eight patients undergoing percutaneous transluminal coronary angioplasty (PTCA) of 100 stenoses were studied for the presence of factors deemed significant in the etiology of silent myocardial ischemia. Thirty-two patients were asymptomatic during balloon dilations of 36 arteries, and 56 patients had angina during PTCA of 64 arteries. There were no differences in age, sex, prior anginal history, antianginal regimen, extent of coronary artery disease and number or duration of inflations between the 2 study groups. Previous infarction (33 vs 12%, p less than 0.02), Q waves in the target area (31 vs 7%, p less than 0.005) and diabetes mellitus (36 vs 17%, p less than 0.05) were present more often in the asymptomatic group. Sixty-four% of all asymptomatic patients had either diabetes or previous infarction in the target territory. Collateral circulation was more frequent in asymptomatic patients, probably reflecting the ability of collateral arteries to ameliorate ischemia. During 2-vessel PTCA, patients without angina during dilation of only 1 of the 2 treated arteries (discordant responders) had previous infarction in that artery's territory (5 of 5, 100%), whereas patients without previous infarction were either symptomatic or asymptomatic (concordant responders) during PTCA of both arteries. This study shows that asymptomatic ischemia occurs frequently during PTCA in patients with symptomatic coronary disease. Prior Q-wave infarction and diabetes mellitus are important, independent factors associated with painless ischemia. It is suggested that infarction produces a localized dysfunction of afferent cardiac pain fibers, whereas diabetes can cause a global cardiac sensory neuropathy.

Angioscopy.

Over the last year, research using flexible, ultrathin fiberoptics for vascular imaging (angioscopy) has continued to demonstrate its clinical potential. Vascular surgeons, thus far angioscopy's strongest advocates, have repeatedly demonstrated that the use of this procedure during peripheral vascular bypass surgery can improve graft patency rates. Although it is doubtful that angioscopy could ever replace angiography, the qualitative details of a vessel's surface disclosed by angioscopy are significant and not available by other means, including intravascular ultrasound. Investigators have used angioscopy to evaluate the burgeoning number of new, catheter-based vascular technologies. A combination of angioscopy and laser or atherectomy, however appealing, will require major technologic advances. One such advance has been the balloon-tipped catheter for blood-free imaging, which circumvents the need for potentially hazardous saline flush. Such an imaging system has provided new insights into the diseased myocardium of living patients.

Programmed electrical stimulation at potential ventricular reentry circuit sites. Comparison of observations in humans with predictions from computer simulations.

The purpose of this study was to define specific types of resetting responses to programmed electrical stimulation during human ventricular tachycardia and to use computer simulations of reentry circuits to assess the possible mechanisms and pacing site location relative to the reentry circuit for each type of response. The effects of scanning single stimuli at 35 left ventricular endocardial sites during sustained monomorphic ventricular tachycardia in 12 patients were studied. In considering alterations in QRS configuration and the delay between the stimulus and the advanced QRS, we identified three types of resetting responses to scanning stimuli consistent with stimulation at sites in or near the reentry circuit at 12 abnormal endocardial sites in eight patients. Type 1: all capturing stimuli were followed after a delay by early QRS complexes that had the same configuration as the tachycardia complexes. Type 2: late stimuli reset tachycardia as in type 1 but early stimuli reset the tachycardia after altering the QRS configuration. Type 3: late stimuli reset tachycardia as in type 1, but early stimuli advanced tachycardia with a short stimulus to QRS delay without altering the QRS configuration. In the simulations, premature depolarization of sites in the circuit produced orthodromic and antidromic wavefronts. The orthodromic wavefront propagated through the circuit and exited the circuit at the same site as did the previous tachycardia wavefronts and advanced the tachycardia without altering the configuration of the advanced QRS. The antidromic wavefront of relatively late stimuli was confined within or near the circuit by collision with the orthodromic wavefront of the preceding tachycardia beat and failed to alter ventricular activation distant from the circuit. Therefore, the QRS configuration after the stimulus was unchanged. A type 1 response occurred when all capturing stimuli produced this effect. However, with increasing stimulus prematurity, the antidromic wavefront propagated farther before colliding with an orthodromic wavefront, and under some conditions, it exited the circuit from a site other than the original circuit "exit," and altered the ventricular activation sequence distant from the circuit and, therefore, the QRS configuration, producing a type 2 pattern. The type 3 pattern occurred when the antidromic wavefront of early premature beats captured the original circuit exit. The effect of a stimulus was dependent on the stimulus prematurity, the relative conduction times from the stimulation site to the potential sites of "exit" from the circuit, and the timing of the excitable gap at the stimulation site.(ABSTRACT TRUNCATED AT 400 WORDS)

Simultaneous cannulation: a technique for percutaneous transluminal coronary angioplasty of chronic total occlusions.

Three patients with indications for percutaneous transluminal angioplasty of a totally occluded coronary artery also had distal opacification by a collateral vessel. The balloon catheter and guide wire were manipulated proximally through the guide catheter. With a second catheter, angiography of the collateral vessel opacified the occluded artery distal to the obstruction. In all three cases, simultaneous catheterization was safe and aided our assessment of distal vessel contour, length of occlusion, and the intraluminal position of the wire beyond the complete obstruction. We conclude that this technique may improve success rates for dilating chronic total obstructions by allowing safer manipulation of less flexible wires.

Coronary angioscopy in patients with unstable angina pectoris.

To visualize intracoronary lesions in patients with different clinical expressions of coronary disease, we performed coronary angioscopy during coronary-artery bypass surgery in 10 patients with unstable angina and 10 patients with stable coronary disease. We examined a total of 32 vessels, using flexible fiberoptic angioscopes. Twenty-two vessels had no acute intimal lesion; three had complex plaques, six had thrombi, and one had both. Coronary angiography correctly identified the absence of complex plaque and thrombus in 22 vessels, but it detected only one of four complex plaques and one of seven thrombi. On angioscopy, none of the 17 arteries in the patients with stable coronary disease had either a complex plaque or thrombus. In the "offending" arteries of the patients with unstable angina, all three patients with accelerated angina had complex plaques and all seven with angina at rest had thrombi. We conclude that angioscopy frequently reveals complex plaques or thrombi not detected by coronary angiography. Our observations suggest that anginal syndromes that are refractory to medical treatment can be caused by unstable pathologic processes in the intima. Ulceration of plaques may increase the frequency and severity of effort angina, and the subsequent development of partially occlusive thrombi may cause unstable rest angina.

Cardiac morphology in rats with growth hormone-producing tumours.

Cardiac enlargement and dysfunction are common in patients with acromegaly. Whether these changes are a direct consequence of growth hormone excess is obscured by the high frequency of hypertension, diabetes mellitus, or atherosclerosis in acromegalic patients. In this study, the effects of chronic elevations of growth hormone (GH) upon the heart were studied in rats with GH-producing tumours implanted subcutaneously for 4 weeks. Geometric measurements and histology were employed to detect the presence of cardiac changes. Increased mass was observed in the tumour-bearing animals. When compared with controls, in tumour-bearing rats there were significantly greater (P less than 0.05) right (0.17 +/- 0.03 v. 0.13 +/- 0.01 g) and left (0.62 +/- 0.05 v. 0.50 +/- 0.04 g) ventricular weights, external cardiac dimensions, and myocardial fibre diameters (9.4 +/- 0.6 v. 8.3 +/- 0.4 micron). However, these increases were linearly-related to increased body mass in the tumour-bearing group so that the ratios of ventricular weights to body weight were similar in both groups. Furthermore, no pathologic changes such as myocardial fibrosis or asymmetric septal hypertrophy were present in the tumour-bearing rats. Thus, under the conditions of this study, growth hormone excess induced cardiac growth, which appeared to represent a manifestation of generalized body growth rather than a distinct pathologic process.