Evidence of reduced resting blood flow in viable myocardial regions with chronic asynergy.

OBJECTIVES: We tested the hypothesis in patients (n = 24) with ischemic heart disease that chronic contractile dysfunction occurs in myocardial regions with true reduction in rest blood flow. BACKGROUND: Whether viable myocardial regions with chronic contractile dysfunction have true reduction in rest myocardial blood flow is controversial. METHODS: Positron emission tomography (PET) 13N-ammonia was used to measure myocardial blood flow in combination with 18F-fluorodeoxyglucose (18FDG) to assess myocardial viability. Viability also was assessed by dobutamine echo and recovery of function after coronary artery bypass grafting (CABG). Segments (n = 252) were selected based on PET measured reduced resting blood flow and rest asynergy on echo. RESULTS: Regional myocardial viability was present in 20 of 23 patients by PET, 13 of 23 by dobutamine echo and 10 of 11 by postrevascularization criteria. Rest blood flow in normal regions was 1.14+/-0.52 ml/min/g and by definition exceeded (p < 0.005) that in both viable (0.48+/-0.15; n = 8 patients) and nonviable (0.45+/-0.14; n = 8 patients) regions (post-CABG criteria), which did not differ. Correction of rest myocardial blood flow in viable asynergic segments, only, for fibrosis and incomplete tracer recovery raised the level to 0.67+/-0.21 (p < 0.005 vs. normal). Finally, evidence of both stunning (rest asynergy with normal flow) and hibernation was present in 15 of 23 (65%) patients. CONCLUSIONS: Reduced rest blood flow in viable myocardial regions with chronic asynergy is common and cannot be accounted for by partial volume effect. Thus, hypotheses concerning physiologic mechanisms underlying chronic contractile dysfunction should consider the role played by chronic reduction of basal myocardial blood flow.

Noninvasive assessment and necropsy validation of changes in left ventricular mass in ascending aortic banded mice.

Although left ventricular (LV) hypertrophy can be induced by aortic banding, noninvasive assessment of changes in LV mass in mice with a banded ascending aorta by using 2-dimensional (2D) images has not been previously performed. In this study we serially assessed changes in LV mass by 2D echocardiography with a newly available 12-MHz transducer in mice with a banded ascending aorta and validated measurements at necropsy. Estimated by echocardiography, LV mass increased from 74+/- 17 mg before banding to 191.08+/-54 mg at 8 weeks after banding (P <.0001), and excellent correlation was shown with postmortem measurements (r = 0.97). Furthermore, with the use of pulsed Doppler 2-dimensionally guided echocardiography, noninvasive measurement of flow velocities in the ascending aorta before and after the band at the various time points was possible. We propose that 2D echocardiography with a 12-MHz transducer is a powerful tool for serial noninvasive evaluations as an adjunct to the study of cardiac hypertrophy in the murine model.

Platelet-derived growth factor-stimulated expression of the MCP-1 immediate-early gene involves an inhibitory multiprotein complex.

We have demonstrated previously that the seven-nucleotide (nt) motif TTTTGTA (the heptamer) that is present within the proximal 3' untranslated sequences of numerous immediate-early genes is essential for platelet-derived growth factor (PDGF)-stimulated induction of the MCP-1 immediate-early gene. On this basis, the heptamer was suggested to be a conserved regulatory element involved in immediate-early gene expression, although its mechanism of action was unknown. Herein, we demonstrate that the heptamer functions to remove an inhibition of PDGF induction of MCP-1 maintained by two independently acting inhibitory elements present in the MCP-1 5' flanking sequences (designated I* elements). PDGF treatment relieves the I*-mediated inhibition of MCP-1 expression only if the heptamer is also present. One inhibitory element is contained within a 59-nt portion of MCP-1 5' flanking sequences and functions in an orientation-independent and heptamer-regulated manner. Significantly, proteins binding to two DNA sequences contribute to the formation of a single multiprotein complex on the 59-nt I* element. The I*-binding complex contains Sp3, an Sp1-like protein, and a novel DNA-binding protein. Moreover, the complex does not form on two 59-nt sequences containing mutations that reverse the inhibition of PDGF induction maintained by the wild-type I* element. We propose to call the multiprotein I*-binding complex a repressosome and suggest that it acts to repress PDGF-stimulated transcription of MCP-1 in the absence of the heptamer TTTTGTA.

Relation between coronary "steal" and contractile function at rest in collateral-dependent myocardium of humans with ischemic heart disease.

BACKGROUND: We tested the hypothesis that rest asynergy in collateral-dependent myocardium correlates with coronary steal. METHODS AND RESULTS: PET with [13N]ammonia measured myocardial blood flow and flow reserve in 15 patients with symptomatic chronic ischemic heart disease. Coronary angiography assessed stenosis severity and collateral blood supply. Echocardiography or contrast ventriculography evaluated regional wall motion. Collateral-dependent segments with normal flow at rest and supplied by coronary vessels having /=0.15 mL. min-1. g-1 versus rest. Blood flow at rest in asynergic, collateral-dependent segments with steal (1.15+/-0.35 mL. min-1. g-1) exceeded (P<0.0001) that of asynergic segments without steal (0.81+/-0.24) and those with normal contraction (0.77+/-0.18). Although the flow reserve ratio of segments with normal contraction (1.8+/-0.8) exceeded that of asynergic ones with (0.6+/-0.1) or without (1.3+/-0.4) steal, overlap was great. Correlation between basal contraction and flow reserve ratio in collateral-dependent myocardium was significant but weak (r=0.45, P<0.001). However, segments demonstrating "steal" with adenosine manifested asynergy in 22 of 23 collateral-dependent segments versus 24 of 39 nonsteal segments (chi2=7.10, P<0.01). CONCLUSIONS: Although myocardial flow reserve in collateral-dependent segments with normal contraction exceeded that of asynergic segments, overlap was great. However, in patients with angina or congestive heart failure, left ventricular segments demonstrating steal with adenosine almost always exhibit asynergy at rest. Thus, coronary steal may play an important role in the pathogenesis of chronic contractile impairment at rest, whereas simple reduction of flow reserve may be less important in selected patients.

Effects of dobutamine at maximally tolerated dose on myocardial blood flow in humans with ischemic heart disease.

BACKGROUND: This study tests the hypothesis in humans with ischemic heart disease that myocardial blood flow response to dobutamine is linearly correlated with blood flow response to adenosine. METHODS AND RESULTS: PET with [13N]ammonia was used to measure myocardial blood flow at rest and during adenosine and dobutamine at the maximally tolerated dose. Myocardial segments were defined physiologically on the basis of blood flow response to adenosine: normal, > or = 2 mL x min(-1) x g(-1); abnormal, < 2 mL x min(-1) x g(-1); and "steal," decline versus baseline > or = 0.15 mL x min(-1) x g(-1). The patient population consisted of 11 men and 2 women. Dobutamine increased heart rate (79+/-22 to 115+/-28 bpm) and rate-pressure product (9748+/-2862 to 15,157+/-3433 mm Hg/min) significantly (both P<.01). Myocardial blood flow at rest in abnormal segments (0.50+/-0.23 mL x min(-1) x g(-1)) was reduced (P<.001) versus normal (0.90+/-0.45) and steal (0.92+/-0.60). Nevertheless, in abnormal segments, blood flow increased versus rest (P<.001) with dobutamine (0.83+/-0.43) and adenosine (0.90+/-0.49). In steal segments, myocardial blood flow declined versus baseline (P<.001) with dobutamine (0.68+/-0.46) and adenosine (0.50+/-0.45). In normal segments, myocardial blood flow increased (P<.001) with dobutamine (2.16+/-0.99) and adenosine (3.10+/-0.90). Over the range of flows, the correlation between adenosine and dobutamine was good (r=.78, P<.0001). Although flow with dobutamine in normal segments correlated with rate-pressure product (r=.81, P<.05), the slope of the line was 2.7+/-0.8 (P<.02), and normalized blood flow (3.3+/-2.5 x rest) exceeded normalized rate-pressure product (1.9+/-0.8 x rest; P<.05). CONCLUSIONS: In humans with ischemic heart disease, myocardial blood flow responses to dobutamine and adenosine are linearly correlated over a wide range. The hyperemic response to dobutamine is in excess of that predicted by rate-pressure product and reflects the unmeasured inotropic, oxygen-wasting, and beta2-agonist effects of the drug. Dobutamine induces coronary steal with a frequency approaching that of adenosine.

Embryonic expression of the Gax homeodomain protein in cardiac, smooth, and skeletal muscle.

Gax is a homeobox-containing gene that has been detected in adult cardiovascular tissues and exhibits a growth arrest-specific pattern of expression in cultured vascular myocytes. To study the regulation of gax during development, we performed immunohistochemistry and in situ hybridization on mouse embryos. Gax was present in mesodermally and, as with other homeobox genes, neuroectodermally derived tissues. Early mesodermal protein expression was limited to the lateral plate and somitic mesoderm. Gax in the cardiac muscle lineage exhibited a biphasic pattern of expression. Expression was prominent in the heart tube of the earliest cardiomyocytes and remained prominent through the looping stage (day 12.5 post coitum [pc]) but fell below the threshold of detection in atria and ventricles by day 13.5 pc. At day 15.5 pc, Gax protein was again detectable but restricted to cells within the compact layer of the ventricular myocardium. Gax expression was also noted in smooth muscle cells as early as day 9.5 pc. In the skeletal muscle lineage, Gax protein was expressed at the onset of somitogenesis before the expression of the myogenic basic helix-loop-helix and MEF2/RSRF family proteins. Subsequently, it was noted at day 9.5 pc in premyogenic cells migrating into head, trunk, and limb buds. Gax was detected in myotomes, premuscle masses, and mature muscle groups. These data suggest an important developmental role for Gax in all muscle lineages.

Quantitative PET measurements of regional myocardial blood flow: observations in humans with ischemic heart disease.

This review focuses on several related issues concerning positron emission tomography measurements of regional myocardial blood flow using 13-N-ammonia in humans. The effect of partial volume correction on estimates of K1, the model parameter describing myocardial blood flow, is considered. In addition a new method for computing K1 images of myocardial flow distribution is briefly described and compared to a standard method. Potential differences between K1 and equilibrium levels of 13-N-ammonia in the myocardium for estimation of myocardial blood flow are discussed also. The issue of heterogeneity of myocardial blood flow and flow reserve in normal volunteers is considered from the clinical point of view in terms of evaluation of patients with ischemic heart disease. Finally, the use of absolute measurement of adenosine-stimulated myocardial blood flow to assess physiological significance of coronary artery stenoses is addressed.

Factors influencing regional myocardial contractile response to inotropic stimulation. Analysis in humans with stable ischemic heart disease.

BACKGROUND: We hypothesized that the response of a myocardial segment to maximal dobutamine reflects not only maximal blood flow but also tethering, metabolic, and beta-blocker status. METHODS AND RESULTS: Patients with stable ischemic heart disease (n = 27) had positron emission tomographic measurement of blood flow at rest and with adenosine, and echocardiography at rest and with dobutamine. Positron emission tomographic measurement of [18F]fluorodeoxyglucose myocardial distribution also was made. Adenosine blood flow in segments that contracted normally at peak dobutamine was similar to that of segments that became hypokinetic (1.06 +/- 0.72 versus 1.02 +/- 0.77 mL.g-1.min-1). Segments that became akinetic failed to augment blood flow (0.68 +/- 0.30 mL.g-1.min-1). Fluorodeoxyglucose-blood flow mismatch was more common in segments with abnormal wall motion at peak dobutamine (24 of 59, 41%) versus those that contracted normally (63 of 269, 23%; chi 2, 7.40; P < .01). In patients off beta-blockers, segments that contracted normally at peak dobutamine increased blood flow with adenosine (0.70 +/- 0.31 to 0.86 +/- 0.46 mL.g-1.min-1; P < .05), whereas those that became abnormal did not (0.63 +/- 0.24 to 0.65 +/- 0.19 mL.g-1.min-1; P = NS). Segments of patients on beta-blockers that contracted normally at peak dobutamine increased blood flow with adenosine (0.78 +/- 0.31 to 1.10 +/- 0.70 mL.g-1.min-1; P < .05), as did segments that became abnormal (0.74 +/- 0.34 to 1.06 +/- 0.82 mL.g-1.min-1; P = NS). However, segments adjacent to ones with abnormal wall motion at rest had higher frequency of abnormal response at peak dobutamine in groups on (48% versus 16%; chi 2, 14.1; P < .001) and off (51% versus 21%; chi 2, 10.9; P < .01) beta-blockers. CONCLUSIONS: Augmented contraction at maximal dobutamine depends not only on increased myocardial blood flow but also on tethering, metabolic, and beta-blocker status. Furthermore, impaired flow reserve does not preclude a normal response to maximal dobutamine, since blood flow need not increase greatly to meet demand.

Gentamicin inhibits carrier-mediated dipeptide transport in kidney.

The effect of gentamicin on transport of pyroglutamylhistidine (pGlu-His) was examined in rabbit renal brush-border membrane vesicles (BBMV). Gentamicin, an aminoglycoside antibiotic, is limited in its usage because of nephrotoxicity characterized in part by transport defects in the proximal tubule. Since there is no information regarding the effects of gentamicin on renal peptide carriers, uptake of [3H]pGlu-His was measured in BBMV following either in vivo or in vitro exposure to the antibiotic. One hour after in vivo administration, the maximal rate (Vmax) for pGlu-His transport was significantly reduced in isolated membrane vesicles washed free of the drug, but the apparent Michaelis constant (Km) was unaltered. Coincubation of membranes with gentamicin during measurements of pGlu-His uptake had a similar effect, causing a significant decrease in the Vmax but not the Km of transport. The addition of 5 mM magnesium to the uptake medium prevented the in vitro but not the in vivo effect. The data indicate that high doses of gentamicin inhibit the capacity but not the affinity of dipeptide transport in the kidney, prior to morphological changes which typify acute tubular necrosis. The in vitro effect is rapid and involves a direct action of gentamicin on the brush-border membrane. The in vivo experiments show that toxicity may be prolonged and remains following removal of the drug from the renal brush border.

Activation of proximal tubular Na(+)-H+ exchange by angiotensin II.

Stimulation of Na(+)-H+ exchange by angiotensin II (ANG II) was characterized in renal proximal tubular cells. Rabbit proximal nephron segments were incubated in the presence or absence of ANG II (5 x 10(-10) M), after which brush-border membrane vesicles (BBMV) were isolated and assayed for Na(+)-H+ antiporter activity using the acridine orange technique. Both the affinity (for sodium) and capacity of the carrier were elevated significantly (P less than 0.05) within 15 min of incubation with ANG II. To determine whether the stimulation of transport capacity involved a change in Na(+)-H+ antiporter density in the luminal membrane, binding of tritiated 5-(N-methyl-N-isobutyl)amiloride ([3H]MIA) was measured in BBMV derived from control and ANG II-treated nephron segments, following maximal stimulation. This demonstrated a significant (P less than 0.05) increase in the maximal specific binding (Bmax) of [3H]MIA binding in the ANG II-treated group compared with control, of a magnitude sufficient to account for the observed change in maximal velocity (Vmax). The data indicate that the Vmax effect is caused by an apparent increase in the number (density) of active Na(+)-H+ carriers present in the luminal membrane. Finally, to test the possibility that the observed kinetic change involves an exocytic mechanism, the effect of colchicine on ANG II-stimulated antiporter activity was examined. The increase in Vmax due to ANG II was blocked by the addition of 0.5 mM colchicine to the incubation medium, whereas colchicine alone had no significant effect on the Vmax of Na(+)-H+ kinetics.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiple carriers for dipeptide transport: carrier-mediated transport of glycyl-L-proline in renal BBMV.

To determine whether multiple carriers are responsible for luminal uptake of glycyl-L-proline (Gly-Pro) in the renal proximal tubule, transport of Gly-[3H]Pro was measured in brush-border membrane vesicles (BBMV). A Line-weaver-Burk analysis of Michaelis-Menten kinetics revealed the presence of two carriers: a lower affinity, higher capacity carrier (Km = 1.3 x 10(-2) M; Vmax = 4.6 x 10(-8) mol.mg-1.min-1) and a higher affinity, lower capacity carrier (Km = 2.7 x 10(-7) M; Vmax = 7.8 x 10(-13) mol.mg-1.min-1). The dipeptides Gly-Sar, beta Ala-His, and pyroGlu-His competitively inhibited the low-affinity carrier. No effect on the Km or Vmax of Gly-Pro transport in this range was seen in the presence of the dipeptides Gly-Gly or cycloHis-Pro. The high-affinity carrier exhibited a different inhibition spectrum. Competitive inhibition of Gly-Pro transport was demonstrated for the dipeptides Gly-Gly and Gly-Sar. However, none of the other peptides tested above altered Gly-Pro transport in the high-affinity range, including pyroGlu-His, which is transported by a high-affinity carrier. At both low (4 x 10(-8) M) and high (4 x 10(-3) M) concentrations, uptake of Gly-Pro was stimulated in the presence of an inwardly directed H+ gradient but was unaffected by the presence of an inward Na+ gradient. In addition, measurements in the presence of valinomycin and an outwardly directed K+ gradient strongly suggest that H(+)-stimulated uptake at both concentrations is electrogenic.(ABSTRACT TRUNCATED AT 250 WORDS)

Long-term neurologic outcome in psychogenic water drinkers with severe symptomatic hyponatremia: the effect of rapid correction.

PURPOSE: The purpose of our study was to ascertain the safety of rapidly correcting acute symptomatic hyponatremia in psychogenic water drinkers, particularly in regard to any delayed adverse neurologic sequelae. PATIENTS AND METHODS: We reviewed the medical records of all known psychogenic water drinkers (34) in our hospital from 1977 to 1989. Using seizure as a marker of severity, we identified 13 patients having a total of 27 episodes associated with severe hyponatremia. We evaluated the charts of those patients in detail to assess the mode of treatment, rate of correction, and long-term neurologic outcome. None of the patients experienced respiratory arrest before treatment, which was initiated within 2 hours of seizure. RESULTS: For all 27 episodes, the initial serum sodium level (mean +/- SE) was 110.9 +/- 1.2 mmol/L, and the rate of correction (mean +/- SE) was 1.65 +/- 0.2 mmol/L/hour. All but one episode were corrected "rapidly" (initial correction rate of 0.7 or more mmol/L/hour) to 120 to 130 mmol/L within 12 hours. The absolute change in the serum sodium level was 15.1 +/- 1.2 mmol/L in 12 hours, 21.6 +/- 1.4 mmol/L in 24 hours, and 25.9 +/- 1.4 mmol/L in 48 hours. In no instance did therapy induce hypernatremia. All patients recovered immediately after treatment. There was no clinical or radiologic evidence of adverse neurologic sequelae immediately after treatment or after 6 years of follow-up. CONCLUSION: In this series of male psychogenic water drinkers, early "rapid" correction of acute symptomatic hyponatremia by raising the serum sodium level 15 mmol/L in 12 hours while maintaining an absolute change in the serum sodium level of 26 mmol/L within 48 hours produced no long-term neurologic sequelae.

Low-affinity transport of pyroglutamyl-histidine in renal brush-border membrane vesicles.

These studies were performed to determine if a low-affinity carrier is present in the luminal membrane of proximal tubular cells for the transport of the dipeptide, pyroglutamyl-histidine (pGlu-His). We have previously described the existence of a specific, high-affinity, low-capacity [transport constant (Kt) = 9.3 X 10(-8) M, Vmax = 6.1 X 10(-12) mol.mg-1.min-1] carrier for pGlu-His in renal brush-border membrane vesicles. In the present study, we sought to demonstrate that multiple carriers exist for the transport of a single dipeptide by determining whether a low-affinity carrier also exists for the uptake of pGlu-His. Transport of pGlu-His into brush-border membrane vesicles was saturable over the concentration range of 10(-5)-10(-3) M, yielding a Kt of 6.3 X 10(-5) M and a Vmax of 2.2 X 10(-10) mol.mg-1.min-1. Uptake was inhibited by the dipeptides glycyl-proline, glycyl-sarcosine, and carnosine but not by the tripeptide pyroglutamyl-histidyl-prolinamide. We conclude that 1) pGlu-His is transported across the luminal membrane of the proximal tubule by multiple carriers and 2) the lower affinity carrier, unlike the higher affinity carrier, is nonspecific with respect to other dipeptides.

Carrier-mediated transport of pyroglutamyl-histidine in renal brush border membrane vesicles.

These studies were performed to determine if a transmembrane carrier for pyroglutamyl-histidine (pGlu-His) is present in the luminal membrane of renal proximal tubular cells. Previous studies have suggested the intact transepithelial transport of pGlu-His, a dipeptide formed by the hydrolysis of luteinizing hormone-releasing hormone by enzymes associated with the brush border in the proximal nephron. With the use of a renal brush border membrane vesicle preparation, pGlu-His showed H+-stimulated, Na-independent, saturable transport into an osmotically active space. High-pressure liquid chromatographic analysis of both the intravesicular and extravesicular fluids indicated intact uptake of the dipeptide. The transport constant (Kt) and Vmax for pGlu-His transport were 9.3 X 10(-8) M and 6.1 X 10(-12) mol.mg-1.min-1, respectively. Transport of pGlu-His was not inhibited by the dipeptides glycyl-proline, glycyl-sarcosine, and N-beta-alanyl-L-histidine, which have been previously shown to be transported into renal brush border vesicles via a single, low-affinity, high-capacity, Na-independent, and H+-stimulated peptide carrier. In addition, the gamma-glutamyl-containing peptides gamma-glutamyl-histidine and N(N-L-gamma-glutamyl-L-cysteinyl)glycine and the tripeptide pyroglutamyl-histidyl-prolinamide were without an inhibitory effect. In contrast, transport of pGlu-His was inhibited by the dipeptide pyroglutamyl-alanine. This study demonstrates the existence of a high-affinity, low-capacity H+ cotransport system for pGlu-His in the proximal tubular luminal plasmalemma, which appears to be specific for pyroglutamyl-containing dipeptides. The data indicate that multiple dipeptide carriers are present in the proximal nephron.

Renal handling of luteinizing hormone releasing hormone: a model for peptide transport and hydrolysis.

This study provides evidence that: 1) LHRH is degraded by renal brush border hydrolases, followed by reabsorption of oligopeptide metabolites in the proximal kidney tubule. 2) Peptide carriers are present in the luminal membrane of the proximal nephron, which apparently function to reabsorb oligopeptide metabolites resulting from hydrolysis of filtered peptides, including LHRH. 3) Renal brush border hydrolysis of LHRH involves cleavage at multiple sites by endopeptidases like angiotensin I-converting enzyme and endopeptidase 24.11; D-amino acid substituents at these sites may alter the expected cleavage pattern of the analogs. 4) A transcytotic pathway is present in the proximal nephron which is facilitated by endocytosis of cationic macromolecules; such a pathway may function to reabsorb hydrolytically resistant peptides, but the issue of potential toxicity must be clarified.

Identification and characterization of a monoclonal antibody recognizing a galactose-binding domain of the toxin ricin.

A monoclonal antibody raised against purified ricin B chain, 75/3B12, blocked ricin toxicity 30- to 100-fold in vitro. The 75/3B12 IgG and F(ab')2 blocked ricin binding to cell surface galactose-containing receptors. The 75/3B12 Fab bound ricin D with a Ka of 10(7) M-1, and this binding was blocked by asialofetuin, lactose, and N-acetylgalactosamine--molecules which interact with the ricin galactose-binding site--but not by fetuin, sucrose, or glucose. The 75/3B12 Fab contained no detectable carbohydrate and, according to several lines of evidence, did not bind ricin via Ig carbohydrate determinants. The monoclonal antibody appears to recognize a galactose-binding site on ricin D via the variable region of the antibody. The 75/3B12 Fab bound ricin E only 1/50 as well as ricin D and bound the Ricinus agglutinin only 1/80 as well as ricin D. The antibody specificity indicates that structural differences exist in the galactose-binding sites of the Ricinus communis lectins. Abrin and other lectins which bind galactose or N-acetylgalactosamine were not significantly bound by the monoclonal antibody. In vitro, the antibody blocked the nontarget toxicity of immunotoxins similarly to lactose. However, in vivo, unlike lactose, the 75/3B12 antibody protected mice from ricin toxicity.