Adsorption at the liquid-vapor surface of a binary liquid mixture.

In a binary liquid mixture, the component possessing the lowest surface tension preferentially adsorbs at the liquid-vapor surface. In the past this adsorption behavior has been extensively investigated for critical binary liquid mixtures near the mixture's critical temperature T(c). In this fluctuation-dominated regime the adsorption is described by a universal function of the dimensionless depth zxi where xi is the bulk correlation length. Fewer studies have quantitatively examined adsorption for off-critical mixtures because, in this case, one must carefully account for both the bulk and surface crossover from the fluctuation-dominated regime (close to T(c)) to the mean-field dominated regime (far from T(c)). In this paper we compare extensive liquid-vapor ellipsometric adsorption measurements for the mixture aniline+cyclohexane at a variety of critical and noncritical compositions with the crossover theory of Kiselev and co-workers [J. Chem. Phys. 112, 3370 (2000)].

Ostial renal artery stent placement in patients 75 years of age or older.

Renal artery stent placement has been shown to improve blood pressure (BP) and stabilize renal function in patients with atherosclerotic renovascular disease. However, limited data are available in patients > or = 75 years of age. We analyzed the prestent characteristics and clinical outcomes of patients aged > or = 75 years who underwent renal artery stenting at our institution. We compared these data with those from the remainder of our stent cohort. Nineteen of 89 (21.3%) stent patients were > or = 75 years old. Before intervention, those > or = 75 years were significantly more likely to be women (84.2% v 55%; P = .02), current or former smokers (78.6% v 36.8%; P = .002), and on a greater number of antihypertensive medications (3.68 v 2.80; P = .048). Average clinical follow-up was similar in both groups (23.9 v 23.2 months; P > .05). At last available follow-up, there were more deaths in those > or = 75 years (7/19 v 5/70; P = .038). No significant difference was found in the incidence of dialysis after intervention (3/19 v 7/70). Seventy-four percent of those > or = 75 years had improved BP, 21% were stable, and 5% were worse. Renal function was improved in 26%, stable in 53%, and worse in 21%. Among those > or = 75 years, there was a significant decrease in systolic BP (186.9 to 144.4; P < .01). There was a trend toward decreased diastolic BP and medications. These clinical results did not differ significantly from patients <75 years. Patients > or = 75 years of age with atherosclerotic renovascular disease have a higher incidence of mortality 2 years after renal artery stent placement, but they seem to derive clinical benefit comparable to younger patients.

Chromosomal location of three unconventional myosin heavy chain genes in the mouse.

We have used an interspecific backcross to map the chromosomal locations of the mouse homologs of the genes encoding myr 3 (unconventional myosin from rat), myr 4, and myr 5, members of the myosin super-family of motor proteins. The mouse loci have been named Myo1e, Myo1c, and Myo9b; they map to chromosomes 9 (MGD position 41), 11 (MGD position 46), and 8 (MGD position 31.5), respectively.

The role of the leader sequence coding region in expression and assembly of bacteriorhodopsin.

Bacterio-opsin is made as a precursor in Halobacterium halobium, which has 13 additional residues at the amino terminus. The codons for these residues have been proposed to form a hairpin structure in the mRNA and play a role in ribosome binding; the leader peptide sequence also has been proposed to have a role in membrane insertion of bacteriorhodopsin (BR). We have made mutations in the bop gene region coding for the leader sequence and expressed the mutant genes in an H. halobium mutant lacking wild-type BR. The leader sequence coding region was found to be important for the stability of the mRNA and for its efficient translation. Single base substitutions in this region that did not affect the amino acid sequence caused significant reductions in protein expression. Deletion of the leader region resulted in unstable mRNA and almost no BR production. Introduction of a new ribosome-binding sequence within the coding region of the mature protein restored mRNA stability and some protein expression. Protein made without the leader peptide was properly assembled in the membrane.

Mutational analysis of the fingers and palm subdomains of human immunodeficiency virus type-1 (HIV-1) reverse transcriptase.

We have analyzed the human immunodeficiency virus type-1 reverse transcriptase (HIV-1 RT) polymerase domain between amino acids 91 and 157 by site-directed mutagenesis. We have constructed a series of amino acid substitutions using BspMI cassettes, and have assayed the RNA-dependent DNA polymerase, DNA-dependent DNA polymerase, and RNase H activities of the mutant HIV-1 RTs. The regions of HIV-1 RT between amino acids 91 and 119 and between amino acids 151 and 157 lie within the palm subdomain and include part of the polymerase active site. A number of amino acids within these regions have been identified as being directly or indirectly involved with polymerization, since amino acid substitutions at these residues decrease the polymerase activity without affecting RNase H activity. The region of HIV-1 RT between amino acids 120 and 150 lies within the fingers subdomain of the HIV-1 polymerase. We believe that the fingers subdomain plays a role in positioning the template. Many amino acid substitutions in this region decrease or abolish both the polymerase and the RNase H functions.

RNase H activity associated with reverse transcriptase from feline immunodeficiency virus.

Reverse transcription of retroviral genomes requires the action of an RNase H for template switching and primer generation. In this report, we compare enzymatic properties of the RNase H associated with the reverse transcriptase (RT) from feline immunodeficiency virus (FIV) and that from human immunodeficiency virus (HIV). Both enzymes displayed substrate preference for poly[3H](rG) . poly(dC) hybird over poly[3H](rA) . poly(dT) and cation preference for Mg2+ over Mn2+. Activity of the FIV RNase H upon poly(rG) . poly(dC) produced hydrolysis products from 1 to 6 nucleotides in length, similar to that reported for HIV. Dextran sulfates were effective inhibitors of both the FIV and HIV RNase H and RT activities. Nearly identical inhibition constants (0.12 nM) were obtained for all enzyme activities with dextran sulfate 500,000, while different inhibition constants were observed with dextran sulfate 8,000. Our results suggest that FIV and HIV RTs contain a conserved region that is sensitive to the larger dextran sulfate and that dextran sulfate 8,000 may interact at a different site or by a different mechanism.

Hemodialysis clearance of encainide and metabolites.

The disposition of encainide and metabolites O-desmethylencainide (ODE) and 3-methyl-ODE (MODE) was evaluated in a 31-year-old hemodialysis patient following a 25 mg oral dose during an interdialytic period and a second 25 mg oral dose 48 h later, 2 h before a hemodialysis procedure. The inter- and intradialytic elimination half-lives were not different for encainide and its metabolites ODE and MODE. The hemodialysis clearance of encainide, MODE, and ODE are all less than 10% of the creatinine clearance of the dialyzer. Thus, hemodialysis does not result in clinically significant removal of encainide or its metabolites.

Calcium transport properties of cardiac sarcoplasmic reticulum from cardiomyopathic Syrian hamsters (BIO 53.58 and 14.6): evidence for a quantitative defect in dilated myopathic hearts not evident in hypertrophic hearts.

Calcium uptake was measured in homogenates and microsomal preparations enriched in sarcoplasmic reticulum vesicles isolated from hearts of hypertrophic (BIO 14.6) and dilated (BIO 53.58) cardiomyopathic as well as control (F1B) Syrian hamsters at 3, 7, 9, and 11 months of age. Calcium uptake studies were done using the Millipore filtration technique under conditions known to restrict transport to the sarcoplasmic reticulum. Steady-state calcium uptake capacity was used as a measure of the relative amounts of sarcoplasmic reticulum in homogenates prepared from individual hearts. At 3 months of age, there were no differences in calcium uptake in homogenates from control or myopathic hearts. However, by 9 months, although calcium uptake of homogenates from control and hypertrophic hearts was the same, calcium uptake by homogenates from dilated hearts was significantly depressed both in initial rate and capacity. Similar trends were seen in the microsomal vesicle preparations, but the decrease in calcium uptake in the dilated hearts was significantly lower by 3 months of age. The catalytic activity of the sarcoplasmic reticulum transport enzyme was estimated from the ratio of velocity to capacity, which provides a measure of the fractional rate of filling of the sarcoplasmic reticulum with calcium. The velocity-to-capacity ratios were not different at any of the ages in both the homogenate and microsomal preparations. The results of this study demonstrate that a major defect in the dilated cardiomyopathy may be due to a decrease in the volume or number of sarcoplasmic reticulum calcium transport sites rather than a decrease in specific activity of the transport enzyme.

L-carnitine treatment improves cardiac performance and restores high-energy phosphate pools in cardiomyopathic Syrian hamster.

Hamsters with either the dilated (BIO 53.58) or hypertrophic (BIO 14.6) form of cardiomyopathy and an inbred control strain of hamster (F1B) were treated for 6 months with high-dose L-carnitine (1 g/kg/day i.p.). After treatment, the animals were killed and their hearts perfused by the isolated working technique. Mechanical performance (as indicated by the double product of heart rate and left ventricular [LV] peak systolic pressure) of both carnitine-treated cardiomyopathic groups was increased significantly above their respective sham-treated groups. Associated with these increases in mechanical performance were significant increases in both peak-positive LV dP/dt (index of contractility) and peak negative dP/dt (index of relaxation) in both carnitine-treated myopathic groups. Serum carnitine levels were increased 10-15 times within 2 hours after injection of L-carnitine in all 3 groups. Myocardial free-carnitine levels were increased twofold in both control and dilated myopathic hearts above their respective sham-treated groups, restoring the level in the dilated hearts comparable to those of controls. Myocardial carnitine levels in the hypertrophic group were not significantly affected by treatment. Total high-energy phosphate stores, i.e., ATP plus creatine phosphate, were restored to control levels by L-carnitine treatment in both cardiomyopathic groups. Levels of the breakdown products of ATP were maintained primarily in the more readily convertible adenosine diphosphate and adenosine monophosphate forms in all three treated groups. These changes resulted in significantly higher ratios of (ATP)/(ADP + AMP + adenosine) and (creatine phosphate)/(creatine) in the treated hearts. This is the first study demonstrating that high-dose L-carnitine treatment results in improved cardiac performance and increased myocardial total high-energy phosphate stores in the Syrian hamster model with one of two distinct forms of cardiomyopathy, i.e., dilated or hypertrophic. The mechanisms for these effects of exogenous L-carnitine treatment cannot be totally explained by changes in oxidative energy metabolism.

Energy metabolism and mechanical function in perfused hearts of Syrian hamsters with dilated or hypertrophic cardiomyopathy.

Energy metabolism was assessed in dilated (congestive) and hypertrophic myopathic hearts from Syrian hamsters after isolated, working heart perfusion with palmitate and/or glucose as substrates. Hearts with these two types of cardiomyopathy were found to be distinctively different from control hearts, and also different from each other. Both cardiomyopathic groups had developed hypertrophy by 3 months but the dilated hearts had a decreased muscle mass by 6 months. In the hypertrophic hearts coronary flow rates per gram of non-collagen protein and, thus, oxygen delivery were markedly increased. With either substrate the hypertrophic hearts maintained more normal levels of adenosine triphosphate in contrast to the dilated hearts whose levels were approximately 50% lower than controls by 6 months of age despite similar heart rates and left ventricular systolic pressure development in all three groups. Lactate to pyruvate ratios in the diseased hearts were comparable to control values. Total coenzyme A levels were statistically lower in the dilated compared to the control group of hearts. Carnitine and its acyl esters, on the other hand, varied markedly with levels of total carnitine decreasing to 50% of control levels in both cardiomyopathic groups by 6 months. In spite of this, the mass action ratios for the carnitine acyl-CoA transferase enzyme complexes were not markedly altered in the control or myopathic hearts regardless of whether palmitate and/or glucose were the perfusate substrates. These results suggest that the decreased carnitine levels are not of sufficient magnitude at this stage in the disease to cause a decrease in cardiac function secondary to restricted energy production. Total carnitine levels were found to be increased in liver and serum of the cardiomyopathic hamsters but unchanged in skeletal muscle. Thus, the deficiency in myocardial carnitine would appear to be due to a specific myocardial problem and not due to a problem of synthesis or supply.

Exercise testing in children before and after surgical treatment of aortic stenosis.

Twenty-three children with valvar or discrete subvalvar aortic stenosis underwent a controlled, progressive bicycle exercise test within 6 months before and 3-30 months after surgery for left ventricular outflow tract obstruction. The patients were divided into three groups according to the preoperative resting gradient of left ventricular to aortic peak systolic pressure: 30-69 mm Hg (group A), 70-99 mm Hg (group B), and greater than or equal to 100 mm Hg (group C). Preoperatively, 19 of 23 patients (83%) developed significant ST depression (greater than or equal to 1.0 mm) during exercise, whereas only seven (30%) had abnormal ST depression at rest. Postoperatively, mean exercise-induced ST depression regressed to less than 1 mm in all three groups. In the total population the frequency of ST depression greater than 1 mm was significantly reduced after surgical treatment and mean total work and peak exercise systolic blood pressure were significantly increased within 12 months after surgery. Total work increased significantly in group B within 12 months and in group C within 13-24 months after surgery, but remained unchanged on group A. Peak exercise heart rates were similar before and after surgery in each group. Peak exercise systolic pressures increased after surgery in all three groups, but the mean differences were statistically significant only in group C patients tested 13-24 months after surgery. The results of this study show that exercise testing is useful for quantifying the severity of aortic stenosis and documenting the clinical improvement (or lack thereof) after surgical treatment, and that properly supervised exercise testing can be performed at minimal risk to children with significant aortic stenosis.

Control of fatty acid metabolism in ischemic and hypoxic hearts.

The effects of whole heart ischemia on fatty acid metabolism were studied in the isolated, perfused rat heart. A reduction in coronary flow and oxygen consumption resulted in lower rates of palmitate uptake and oxidation to CO2. This decrease in metabolic rate was associated with increased tissue levels of long chain acyl coenzyme A and long chain acylcarnitine. Cellular levels of acetyl-CoA, acetylcarnitine, free CoA, and free carnitine decreased. These changes in CoA and its acyl derivatives indicate that beta oxidation became the limiting step in fatty acid metabolism. The rate of beta oxidation was probably limited by high levels of NADH and FADH2 secondary to a reduced supply of oxygen. Tissue levels of neutral lipids showed a slight increase durning ischemia, but incorporation of [U-14C]palmitate into lipid was not altered significantly. Although both substrates for lipid synthesis were present in higher concentrations during ischemia, compartmentalization of long chain acyl-CoA in the mitochondrial matrix and alpha-glycerol phosphate in the cytosol may have accounted for the relatively low rate of lipid synthesis.

Fatty acid oxidation by isolated perfused working hearts of aged rats.

It has been reported that mitochondria isolated from hearts of old rats have lower respiratory activity than mitochondria from young rats. In order to determine the physiological correlates of these changes, the metabolism of hearts from young and old rats has been compared in a perfused working heart preparation. The oxidation of [14C]palmitate to 14CO2, oxygen consumption, and nucleotide levels were measured under different cardiac workloads. The hearts from old animals performed less cardiac work and utilized less oxygen and palmitate in proportion to tissue mass, but the ratio of oxygen consumed to pressure developed was unaltered. There was a small but significant decrease in cardiac efficiency expressed as the ratio between the rate of oxygen consumed and ventricular pressure development. Tissue levels of total carnitine and long-chain acylcarnitine derivatives were greatly reduced in the older heart without significant change in free CoA, acetyl-CoA, or long-chain acyl-CoA. The adenine nucleotide levels were not significantly different in the two groups. The results appear consistent with the in vitro studies on isolated mitochondria.

Rate-limiting steps of carbohydrate and fatty acid metabolism in ischemic hearts.

Control of glycolysis and fatty acid oxidation in ischemic myocardium was studied in isolated working rat hearts. Coronary flow was reduced to the whole heart. In ischemic tissue, oxygen consumption, glycolysis and fatty acid oxidation all decreased in proportion to the restriction in coronary flow. Inhibition of glycolysis developed at the level of glyceraldehyde-3-phosphate dehydrogenase. Restricted flux through this step appeared to result from accumulation of lactate, H+ and NADH. The rate of glycolysis was inversely related to accumulation of lactate. Additions of high levels of lactate to the perfusate inhibited glycolysis in aerobic, anoxic and ischemic hearts. The mechanism of this effect of lactate in anaerobic hearts is unknown, but does not appear to be related to pH changes. Oxidation of fatty acids was restricted at the level of beta-oxidation and high levels of both long-chain acyl CoA and carnitine derivatives accumulated.

Effect of coronary blood flow on glycolytic flux and intracellular pH in isolated rat hearts.

The rate of coronary blood flow was varied in isolated working rat heart preparations to determine its influence on the rate of glocose utilization, tissue high-energy phosphates, and intracellular pH. A 60% reduction in coronary blood flow resulted in a 30% reduction in oxygen consumption, an accelerated rate of glusoe utilization, lower tissue levels of high-energy phosphate, and higher tissue levels of lactate and H+. Ventricular performance deteriorated as reflected by a decrease in heart rate and peak systolic pressure. Further reductions in coronary blood flow resulted in inhibition of glycolysis, a greater decrease in tissue levels of high-energy phosphates, and higher tissue levels of both lactate and H+. These changes in glycolytic flux, tissue metabolites, and ventricular performance were proportional to the degree of restriction in coronary blood flow. The importance of coronary blood flow and washout of the interstitial space in the maintenance of accelerated glycolytic flux in oxygen-deficient hearts is emphasized. It is concluded that acceleration of ATP production from glycolysis can occur only in the marginally ischemic tissue in the peripheral area of tissue supplied by an occluded artery. The central area of tissue which receives a low rate of coronary blood flow will have a reduced rate of ATP production due to both a lack of oxygen and an inhibition of glycolysis.

Relationship between coronary flow and adenosine triphosphate production from glycolysis and oxidative metabolism.

The relationship between coronary flow and adenosine triphosphate ATP production was determined in isolated rat hearts and in situ pigs hearts. The major source of ATP in ischemic hearts was oxidative phosphorylation. Oxidation of glucose accounted for most of the residual oxygen consumption in ischemic hearts when the concentration of fatty acids was low, but at 1.2 mM palmitate fatty acids were oxidized in preference to carbohydrate, as in aerobic hearts. The rates of ATP production from both glycolysis and oxidative metabolism were decreased in proportion to the reduction in coronary flow in oxygen-deficient hearts. Glycolysis was reduced to below aerobic rates when coronary flow was about 0.5 ml/min/g tissue in both rat hearts perfused with bicarbonate buffer and blood-perfused pig hearts. Tissue level of high energy phosphates reflected the rates of ATP production and declined in proportion to the reduction in coronary flow. In addition, tissue lactate and H+ accumulated in proportion to the restriction in flow.

Inhibition of glycolysis in hearts during ischemic perfusion.

Rates of glycolysis were determined in the isolated perfused rat heart under aerobic, anoxic, and ischemic conditions. The rate was accelerated in anoxic and was inhibited in ischemic tissue. Glycolytic inhibition developed at the level of glyceraldehyde-3-P dehydrogenase and was associated with accumulation of high levels of tissue lactate and H+.