Correlation between morning urine estriol concentration and 24-hour estriol excretion

In an attempt to circumvent the need for 24-hour urine collections for estriol analyses in assessment of fetal status, the possibility of using morning urine samples was investigated. Results indicate 1) good correlation between 24-hour estriol excretion and morning estriol concentration, and between corresponding E/C ratios, 2) similar morning and 24-hour estriol concentrations, 3) high dependence of estriol and creatinine excretion on 24-hour urine volume but not on morning volume, 4) larger variations in 24-hour than in morning urine volume, and 5) better consistency of values in morning than in 24-hour samples in pathologic pregnancy. The use of serial morning urine concentrations at least as an outpatient monitoring procedure is suggested.     

Correlation between esophageal motility and 24-hour pH recording in patients with gastroesophageal reflux

The relationship between manometric and pH-metric data was studied in a group of 50 patients with symptoms of gastroesophageal reflux. Using a multiple regression analysis, we found that the total percentage of reflux was significantly correlated to the infradiaphragmatic length and resting pressure of the lower esophageal sphincter and also to the mean amplitude of the contractile waves of the distal esophagus, thus revealing the important role of these factors in the antireflux mechanism. When the patients were divided into groups according to their manometric characteristics and the values of the various pH-metric parameters between these groups compared using a one-way analysis of variance, we found that the amplitude of the contractile waves and the percentage of deglutitions without response were related not only to the total percentage of reflux but also to the number of reflux episodes of greater than 5 min duration and to the duration of the longest episode. This shows that prolonged exposure of the esophageal mucosa to the refluxed material may be due, in part, to an alteration in the capacity for esophageal clearing.     

Correlation between serum-ascites albumin concentration gradient and endoscopic parameters of portal hypertension

OBJECTIVE: We sought to determine the correlation between the level of serum-ascites albumin concentration gradient (SAAG) and the complications of portal hypertension (PHTN), manifested by the presence and grade of esophageal varices (EV). METHODS: Our study included 31 patients with ascites, demonstrated by ultrasonography, who had measurement of the SAAG. All had upper gastrointestinal endoscopy with assessment of the presence and size of EV. High SAAG was considered to be present when the SAAG was > or = 1.1 g/dl and Low SAAG when it measured < 1.1 g/dl. RESULTS: We found that 25 of 31 (80.6%) patients had High SAAG and six of 31 (19.4%) had Low SAAG. Esophageal varices were present in 17 of 25 (68%) patients with High SAAG and in none of six (0%) patients with Low SAAG (p = 0.028). In patients with alcoholic liver disease (ALD), 14 of 14 (100%) had EV. Otherwise, in patients with nonALD, only three of 11 (27.3%) had EV (p < 0.05). The presence of EV was associated with the Child-Pugh Score (p = 0.039). Among patients with High SAAG, EV were present in four of 10 (40%) with SAAG values of 1.10-1.49 g/dl; in four of 6 (66.7%) with SAAG values of 1.50-1.99 g/dl; and in nine of nine (100%) with SAAG values of > or =2.0 g/dl (p = 0.049). The size of the EV had no association with the level of SAAG in patients with High SAAG (p = 0.788), with a Pearson correlation coefficient of R = 0.54 (p = 0.005). Using the Receiver-Operating-Characteristic Curve a SAAG value of > or =1.435 +/- 0.015 g/dl was an accurate indicator of the presence of EV (cutoff point for the higher predictive value: positive = 87.5% and negative = 66.7%). CONCLUSIONS: In patients with ascites the presence of EV is associated only with patients with High SAAG. The presence of EV in patients with ascites and High SAAG is directly related to the degree of SAAG. The size of the EV in patients with ascites and High SAAG is not associated with the degree of SAAG. A SAAG value of > or =1.435 +/- 0.015 g/dl is a useful means to predict the presence of EV in patients with ascites. Finally, in patients with ascites, EV were more prevalent in those with ALD.     

Increased amounts of collagenase and gelatinase in porcine myocardium following ischemia and reperfusion

We studied the presence of collagen degrading enzymes (matrix metalloproteinases, MMPs) in porcine myocardium following ischemia and late reperfusion. In nine pigs, left anterior descending coronary artery was occluded for 6 h followed by reperfusion for 3 h. Six pigs without coronary occlusion served as controls. After the reperfusion period, transmural biopsies from the anterior (ischemic zone) and posterior wall (non-ischemic myocardium) in the left ventricle were obtained and extracted. Heparin-Sepharose isolated components in extracts were analysed for collagenase (triple-helical collagen degradation) and gelatinase activity (zymography). Immunohistochemistry using anti-human (MMP-1, MMP-2, MMP-9, and fibronectin) antibodies was performed on additional biopsies. Collagenase (MMP-1) and gelatinases (MMP-2, MMP-9) could be demonstrated in the extracts of non-ischemic myocardium from ischemic/reperfused as well as control pigs and MMP-1 and MMP-9 activity was found to be increased in ischemic/reperfused myocardium compared with non-ischemic myocardium. In ischemic/reperfused myocardium from live pigs investigated, myocyte necrosis could be confirmed by fibronectin immunoreaction in myocytes and MMP-1 and MMP-9 immunoreactions were increased. MMP-9 was present in cells likely to be infiltrating leukocytes in a patchy distribution throughout the ischemic myocardium. Quite coincident with MMP-9 positive cells, MMP-1 immunoreaction appeared in necrotic myocytes, in addition to reactions observed in vessel walls, endo- and epicardium, and extracellular matrix in non-ischemic myocardium. Thus, the results showed increased amounts of collagenase (MMP-1) and gelatinase (MMP-9) in ischemic/ reperfused myocardium, indicating the appearance of increased amounts of collagen degrading enzymes very early following ischemia and late reperfusion.     

Sarcoplasmic reticulum calcium uptake in human myocardium subjected to ischemia and reperfusion during cardiac surgery

We evaluated the effect of ischemia and reperfusion on sarcoplasmic reticulum Ca uptake in patients subjected to cardiac surgery. Our series included 16 patients (seven female, nine male, age 63 +/- 2 years): five were subjected to aortic valve replacement, five to aortic and mitral valve replacement, six to coronary artery bypass graft. In each case no clinical, electrocardiographic or echocardiographic evidence of perioperative infarction was observed. Biopsies were obtained from the right atrium of each patient before starting extracorporeal circulation, and after the recovery of spontaneous contractile activity, i.e. after cardioplegia-ischemia-reperfusion. The tissue was homogenized, and oxalate-supported Ca uptake, which represents sarcoplasmic reticulum Ca uptake, was measured in the unfractionated homogenate. The assay was performed under basal conditions and in the presence of 900 microM ryanodine, in order to block sarcoplasmic reticulum Ca release channels. Under basal conditions at pCa = 5.85 the rate of sarcoplasmic reticulum Ca uptake averaged 4.76 +/- 0.37 nmol/min per mg of protein in the pre-ischemic samples, and decreased significantly in the post-ischemic samples (3.09 +/- 0.29 nmol/min per mg, P < 0.01). A significant decrease of Ca uptake after ischemia and reperfusion was observed also in the presence of ryanodine (3.53 +/- 0.48 nmol/min per mg) compared to pre-ischemic values (5.98 +/- 0.56 nmol/min per mg, P < 0.01). Additional experiments showed no change in the Ca sensitivity of Ca uptake in the postischemic samples (Kca = 0.48 +/- 0.02 microM, no significant difference after ischemia and reperfusion). In conclusion, active sarcoplasmic reticulum Ca transport was impaired in human atrial myocardium after reversible ischemia and reperfusion.     

Cellular uncoupling during ischemia in hypertrophied and failing rabbit ventricular myocardium: effects of preconditioning

BACKGROUND: Patients with heart failure show a very high incidence of arrhythmias and sudden death that is often preceded by ischemia; however, data on electrophysiological changes during ischemia in failing myocardium are sparse. We studied electrical uncoupling during ischemia in normal and failing myocardium. METHODS AND RESULTS: Tissue resistance, intracellular Ca2+ concentration (Indo-1 fluorescence ratio), and mechanical activity were simultaneously determined in arterially perfused right ventricular papillary muscles from 11 normal and 15 failing rabbits. Heart failure was induced by combined volume and pressure overload. Before sustained ischemia, muscles were subjected to control perfusion (non-PC) or ischemic preconditioning (PC). The onset of uncoupling during ischemia was equal in non-PC normal (13.6+/-0.9 minutes of ischemia) and non-PC failing hearts (13.3+/-0.7 minutes of ischemia). PC postponed uncoupling in normal hearts by 10 minutes. In failing hearts, however, PC caused a large variability in the onset of uncoupling during ischemia (mean, 12.2+/-2.1; range, 5 to 22 minutes of ischemia). The duration of uncoupling process was prolonged in failing hearts (12.9+/-0.9 minutes) compared with normal hearts (7.8+/-0.4 minutes). The degree of heart failure and relative heart weight of the failing hearts significantly correlated with the earlier uncoupling after PC and the duration of uncoupling. In every experiment, the start of Ca2+ rise and contracture preceded uncoupling during ischemia. CONCLUSIONS: The duration of the process of ischemia-induced electrical uncoupling in failing hearts is prolonged compared with that in normal hearts. Ischemic PC has detrimental effects in severely failing papillary muscles because it advances the moment of irreversible ischemic damage.     

Arterial ketone body ratio and adenosine triphosphate concentration in hepatic ischemia and reperfusion

BACKGROUND/AIMS: The arterial ketone body ratio (AKBR) and the cellular adenosine triphosphate (ATP) concentration have been proposed as indicators of liver function. However, recent studies of the utility of the AKBR as a biochemical marker have been called into question. Furthermore, there is no practical data defining the relationship between ATP concentration and ischemia-reperfusion (IR) changes during liver surgery. METHODOLOGY: The relationship of the AKBR and arterial ATP concentration to IR during hepatectomy was investigated. In 20 patients who underwent hepatectomy, arterial acetoacetate, beta-hydroxybutyrate, and ATP concentrations were measured. The ratio of acetoacetate to beta-hydroxybutyrate (AKBR) was calculated before and after vascular occlusion. RESULTS: The AKBR 15 minutes after clamping was lower than the preclamping values in all of the patients. It increased after unclamping, returning toward the preclamping levels. An AKBR of less than 0.5 prior to clamping did not correlate with preoperative hepatocellular function. An AKBR of less than 0.7 throughout IR was not a consistent risk factor for postoperative complications and liver dysfunction. The arterial ATP concentration did not correlate with the changes during IR or with preoperative hepatocellular function. CONCLUSIONS: Although the AKBR changed during IR as a general indicator of cellular activity, the absolute value of the AKBR was not an accurate predictor of liver function. The arterial ATP concentration also was not a suitable clinical biochemical marker of hepatic function.     

Study of the spectrum of rabbit myocardium nuclear DNAases using models of ischemia and diabetes mellitus

There were alterations in the spectrum of nuclear endo-DNAases in the rabbit myocardium on experimental models of diabetes mellitus and other diseases where the perfused heart was involved i.e. disappearance of the enzyme with 130 kDa from the control extracts was followed by a great increase in the content of low molecular forms in the nuclear preparations obtained from the ischemic heart, as well as that with 145 kDa in all the abnormalities under study.     

Myocardial ischemia and reperfusion are associated with an increased stiffness of remote nonischemic myocardium

During and after an ischemic injury, maintenance and recovery of cardiac function may critically depend on remote nonischemic myocardium. Graded myocardial ischemia is associated with an approximately 50% increase in stiffness of nonischemic myocardium. We determined whether this increase in stiffness is unique to the ischemic period or persists during reperfusion. Ten anesthetized (isoflurane 1.0% vol/vol) open-chest dogs were instrumented to measure left ventricular pressure and dimensions (sonomicrometry) in ischemic and nonischemic myocardium. Regional chamber stiffness and myocardial stiffness were assessed using the end-diastolic pressure-length relationship which was modified by stepwise infusion and withdrawal of 200 mL of the animals' own blood during baseline, 45 min low flow ischemia (systolic bulge), and 60 min after the onset of reperfusion. In remote nonischemic myocardium, regional myocardial ischemia was associated with a significant (P < 0.05) increase in chamber stiffness (+44%) and myocardial stiffness (+48%). Sixty minutes after the onset of reperfusion, chamber stiffness (+54%, P < 0.05 versus baseline) and myocardial stiffness (+55%, P < 0.05 versus baseline) remained increased. Thus, the ischemia-induced increase in stiffness of remote nonischemic myocardium persists for at least 60 min after reperfusion.     

Correlation of EGTA and calcium-blocking agents on the response of the bladder to in vitro ischemia

The effects of repetitive field stimulation (model of hyperrelexia) on the responses of isolated strips of rabbit urinary bladder to FS and carbachol were evaluated under a variety of incubation conditions. Compared to control conditions, 2 h of repetitive FS in normal, oxygenated Tyrode's solution followed by incubation for 1 h with no stimulation resulted in a 50% decrease in contractile response to FS and a 30% decrease in the response to carbachol. Incubation in the absence of O2 and glucose was used as an in vitro model for ischemia. Repetitive stimulation during in vitro ischemia resulted in a significantly greater decrease in the contractile responses to FS and carbachol than did in vitro ischemia without repetitive stimulation. The magnitude of contractile dysfunctions in response to both stimuli were significantly reduced in the presence of EGTA (calcium chelator), diltiazem (calcium channel blocker) or pincidil (potassium channel opener). Incubation with thapsigargin (SR calcium uptake inhibitor) + ryanodine (SR calcium storage inhibitor) had no effect. The results of these studies indicate that inhibition of Ca2+ entry reduces the contractile dysfunctions induced by repetitive stimulation in the presence of in vitro ischemia. Inhibition of Ca2+i storage and release had no significant effect on the magnitude of contractile dysfunctions induced by repetitive stimulation an in vitro ischemia.     

Effect of pH and calcium concentration on calcium diffusion in streptococcal model-plaque biofilms

Demineralization during a cariogenic episode is affected by storage and transport in dental plaque of ions released from enamel, and by the effect on both of plaque fluid pH and ion concentrations. To investigate this, 45Ca effusion from a condensed film of streptococci was measured at pH 7, 6 and 5, and 0-20 mmol/l calcium. Cells were loaded into effusion chambers and the appearance of 45Ca and [3H]-inulin in carrier-containing but initially tracer-free buffer was measured. Ratios of 45Ca and [3H]-inulin activity in the initial suspending solution and at equilibrium in the clearance solution, permitted calculation of extracellular volume and bound calcium. The rate of Ca appearance was proportional to the retarded diffusion coefficient (rDe), which was related to the effective diffusion coefficient (De) by: rDe = De/(1 + R) in which R is the ratio of bound to free Ca2+. The rate of Ca2+ effusion increased with calcium concentration, converging on a value of 2.8 x 10(-10) m2/sec. At low pH it reached convergence at a lower [Ca]. This demonstrates that calcium effusion is dependent on binding, so a high proportion of binding sites in plaque will reduce mineral loss in vivo. Loss of binding sites at low pH will increase mineral loss.     

A subunit interaction in chloroplast ATP synthase determined by genetic complementation between chloroplast and bacterial ATP synthase genes

F1F0-ATP synthases utilize protein conformational changes induced by a transmembrane proton gradient to synthesize ATP. The allosteric cooperativity of these multisubunit enzymes presumably requires numerous protein-protein interactions within the enzyme complex. To correlate known in vitro changes in subunit structure with in vivo allosteric interactions, we introduced the beta subunit of spinach chloroplast coupling factor 1 ATP into a bacterial F1 ATP synthase. A cloned atpB gene, encoding the complete chloroplast beta subunit, complemented a chromosomal deletion of the cognate uncD gene in Escherichia coli and was incorporated into a functional hybrid F1 ATP synthase. The cysteine residue at position 63 in chloroplast beta is known to be located at the interface between alpha and beta subunits and to be conformationally coupled, in vitro, to the nucleotide binding site > 40 A away. Enlarging the side chain of chloroplast coupling factor 1 beta residue 63 from Cys to Trp blocked ATP synthesis in vivo without significantly impairing ATPase activity or ADP binding in vitro. The in vivo coupling of nucleotide binding at catalytic sites to transmembrane proton movement may thus involve an interaction, via conformational changes, between the amino-terminal domains of the alpha and beta subunits.     

Time-resolved charge translocation by the Ca-ATPase from sarcoplasmic reticulum after an ATP concentration jump

Time-resolved measurements of currents generated by Ca-ATPase from fragmented sarcoplasmic reticulum (SR) are described. SR vesicles spontaneously adsorb to a black lipid membrane acting as a capacitive electrode. Charge translocation by the enzyme is initiated by an ATP concentration jump performed by the light-induced conversion of an inactive precursor (caged ATP) to ATP with a time constant of 2.0 ms at pH 6.2 and 24 degrees C. The shape of the current signal is triphasic, an initial current flow into the vesicle lumen is followed by an outward current and a second slow inward current. The time course of the current signal can be described by five relaxation rate constants, lambda1 to lambda5 plus a fixed delay D approximately 1-3 ms. The electrical signal shows that 1) the reaction cycle of the Ca-ATPase contains two electrogenic steps; 2) positive charge is moved toward the luminal side in the first rapid step and toward the cytoplasmic side in the second slow step; 3) at least one electroneutral reaction precedes the electrogenic steps. Relaxation rate constant lambda3 reflects ATP binding, with lambda(3,max) approximately 100 s(-1). This step is electroneutral. Comparison with the kinetics of the reaction cycle shows that the first electrogenic step (inward current) occurs before the decay of E2P. Candidates are the formation of phosphoenzyme from E1ATP (lambda2 approximately 200 s[-1]) and the E1P --> E2P transition (D approximately 1 ms or lambda1 approximately 300 s[-1]). The second electrogenic transition (outward current) follows the formation of E2P (lambda4 approximately 3 s[-1]) and is tentatively assigned to H+ countertransport after the dissociation of Ca2+. Quenched flow experiments performed under the conditions of the electrical measurements 1) demonstrate competition by caged ATP for ATP-dependent phosphoenzyme formation and 2) yield a rate constant for phosphoenzyme formation of 200 s(-1). These results indicate that ATP and caged ATP compete for the substrate binding site, as suggested by the ATP dependence of lambda3 and favor correlation of lambda2 with phosphoenzyme formation.