Transcriptional and epigenetic effects of deleting large regions, alone or in combination, from their natural context in the chicken Ig-ß gene.

Previously, we used homologous recombination to delete six groups of cell-type-specific DNase I hypersensitive sites (DHSs), potential transcriptional and epigenetic regulators, scattered in and around the Ig-ß gene from their natural context in B-lymphocyte-derived chicken DT40 cells. Simultaneous deletion of all six groups completely shut down transcription and epigenetic regulation of the Ig-ß gene; therefore, the cooperation of the scattered regulatory regions was essential for transcription and epigenetic regulation. In this study, we regrouped the cell-type-specific DHSs of Ig-ß, those in the original six deletions and three additional ones, into three larger regional groups-the long upstream region, the intron, and the long downstream region-and deleted these groups individually or in combination. Combinatorial deletion of all three regional groups decreased Ig-ß mRNA levels to 0.4% of the control, which was significantly higher than <0.1%, the level resulting from deletion of all six smaller groups. Histone H3 and H4 acetylation and H3K4 dimethylation levels at the Ig-ß promoter were low in cells carrying deletions of all six smaller groups, but intermediate levels of acetylation and enhanced H3K4 dimethylation were observed in cells carrying deletions of all three larger groups. While CG methylation was definitely present at the Ig-ß promoter in cells carrying all six smaller deletions, it was nearly absent from the Ig-ß promoter in cells carrying all three larger deletions. Thus, combinatorial deletion of larger regulatory regions had less effect on transcription and epigenetic regulation at the chicken Ig-ß gene than combinatorial deletion of shorter ones. Analysis of several combinatorial deletions, where combinations included two larger deletions and one smaller deletion, revealed the relative effects of each deletion on transcription of the Ig-ß gene. Investigation of the CG methylation status at the Ig-ß promoter in one combinatorial deletion demonstrated that USI was involved in the maintenance of CG methylation.

[Anesthetic management of two patients with mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS)].

MELAS is a type of the mitochondrial myopathy characterized by elevation of pyruvate and lactate levels in both the blood and cerebrospinal fluid. This syndrome frequently accompanies cerebral infarction like symptoms. Recently, we experienced two patients for anesthesia with MELAS (both 11-year-old girls). V-P shunt construction and IVH reservoir implantation were conducted, respectively. Anesthesia was induced with fentanyl and midazolam, and vecuronium was used to facilitate tracheal intubation. Volatile anesthetic was avoided, and anesthesia was maintained with fentanyl, midazolam, and nitrous oxide. Arterial blood gases and pH were frequently checked, and acetated electrolyte solution was infused mainly during surgery. No complications occurred during anesthesia in both patients. In the anesthetic management for MELAS, measures to prevent malignant hyperthermia must also be considered.

Loxoprofen--another NSAID associated with acute asthmatic death.

Loxoprofen sodium (sodium 2[4-(2-oxocyclopentylmethyl) phenyl] dehydrate; CAS #68767-14-6) is a nonsteroidal, inflammatory drug marketed only in Japan. A case report describes its association with an acute asthmatic death with features resembling those evoked by similar drugs. The analytic methodology is reported. The blood levels of loxoprofen were in the therapeutic range. The tissue concentrations are reported.

Intracellular aluminium inhibits acetylcholine- and caffeine-evoked Ca2+ mobilization.

The effect of intracellular aluminium on Ca2+ signalling in single internally perfused mouse pancreatic acinar cells was investigated by measurement of the Ca2(+)-dependent Cl- current using the patch-clamp whole-cell recording configuration. Acetylcholine (ACh) normally evoked a pulsatile Ca2(+)-dependent Cl- current, but when AlCl3 (1 mM) was present in the internal perfusion solution the ACh responses were virtually absent. When aluminium was acutely infused into the internal perfusion solution, the ACh-evoked Ca2+ signals and also the caffeine-evoked responses quickly disappeared, but the Ca2+ ionophore, ionomycin (100 nM), could still induce a large increase in the Cl- current. It is concluded that intracellular aluminium can abolish receptor-activated intracellular Ca2+ release probably by inhibition of Ca2(+)-induced Ca2+ release.

Effects of acupuncture needle application upon cutaneous microcirculation of rabbit ear lobe.

Microcirculatory effects of the application of an acupuncture needle (32-gauge, silver) to the back, corresponding to Geshu (B17) in human beings, were studied in vivo by microscope, using a transparent ear chamber in conscious rabbits. Although no striking findings were obtained during the needle application for a period of 30 minutes, it was clearly observed that the microvascular blood flow increases gradually in parallel with augmenting spontaneous rhythmic fluctuation of the vessel diameter, namely vasomotion, throughout a continuous observation period longer than 2 hours following release from the needle application. Diameters of arterioles and venules at the full-dilating phases of vasomotion reached levels higher than 200% and 250% of the initial values just before application of the needle, respectively. The clinical efficacy of acupuncture was suggested to be explained at least in part by the increased rhythmic microvascular blood flow in parallel with vasomotion, from the physiological point of view based on the previous investigations.

[Quantitative evaluation of regional myocardial blood flow by digital subtraction angiography: correlations with exercise electrocardiography and Tl-201 myocardial scintigraphy].

We previously reported that the contrast disappearance half-life (T1/2) derived by the computerized washout analysis of digital subtraction coronary arteriograms provides a useful index for quantitatively evaluating regional myocardial blood flow. In the present study, we further evaluated the clinical usefulness of T1/2, comparing it with exercise electrocardiography and exercise thallium-201 myocardial scintigraphy. The study subjects consisted of 25 patients with angina pectoris and 14 patients with normal coronary arteries. Following the manual injection of contrast media into the left anterior descending coronary artery (LAD), a time-density curve was generated in the sectors of the myocardium which were perfused by the LAD and the T1/2 was calculated. T1/2 values correlated closely with double product (r = -0.73). They were significantly greater in patients with exercise-induced ST depression (8.3 +/- 1.0 vs 5.8 +/- 0.7, p less than 0.005). In addition, there was a good correlation between T1/2 values and washout ratio as determined by exercise thallium-201 myocardial scintigraphy, with r = -0.83. Although T1/2 values were within the normal range (mean +/- 2SD of control subjects) in all patients with LAD stenosis of 50 percent or less, these values were abnormally increased, exceeding the normal range, in 11 of the 12 patients with stenosis of 90 percent or more. Compared with exercise electrocardiography, T1/2 values were abnormally prolonged in 11 of the 13 patients with exercise-induced ST depression. Compared with exercise thallium-201 myocardial scintigraphy, T1/2 values were abnormally prolonged in seven of the nine patients with transient perfusion defects.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparable effects of oral diltiazem and verapamil in the treatment of hypertrophic cardiomyopathy. Double-blind crossover study.

The effects of oral diltiazem, 180 mg/day, were compared with those of oral verapamil, 240 mg/day, in 32 patients with hypertrophic cardiomyopathy (HCM), using a double-blind crossover study design. In the first treatment period, diltiazem and verapamil improved subjective complaints in 83% and 71% of those who were symptomatic in the baseline period. Maximal oxygen consumption on exercise stress test increased with verapamil by 2.9 +/- 4.2 ml/Kg/min (p less than 0.05), and tended to increase with diltiazem. Verapamil also reduced the amplitude of negative T wave. In the statistical analysis based on the crossover design, diltiazem and verapamil did not differ in global improvement, overall safety and global utility ratings. In addition, both drugs showed comparable effects on electrocardiographic and echocardiographic variables and exercise tolerance except for minor differences in diastolic blood pressure, T wave amplitude and peak exercise heart rate. On the other hand, verapamil tended to induce more serious side effects, forcing the discontinuation of medication in 3 patients. Therefore, the present study indicates that diltiazem is essentially equally as effective as verapamil and is preferable in the treatment of patients with HCM since it may exhibit fewer serious side effects.

[Apical hypertrophy and its genetic and acquired factors].

Although apical hypertrophy is characterized by a spade-like configuration of the left ventricle and giant negative T waves on electrocardiograms, the identity of apical hypertrophy in the disease spectrum of hypertrophic cardiomyopathy (HCM) is not fully established. The present study compared the demography, familial occurrence, and acquired factors of 43 patients who had apical hypertrophy with those of 104 patients who had asymmetric septal hypertrophy (ASH). Demographically, apical hypertrophy occurred predominantly in middle-aged males (86%). Family surveys showed that 13% of siblings of apical hypertrophy were affected, significantly less than in obstructive (31%) or non-obstructive (29%) HCM with ASH. Thirty-eight percent of siblings of ASH patients less than 35 years of age were affected, with a male/female ratio of 4/5, suggesting an autosomal dominant inheritance. The acquired factors associated with apical hypertrophy were assessed in a case-control study. Relative risk (odds ratio) of the condition was 3.46 (p less than 0.05) in those with histories of hypertension, and increased further to 8.09 (p less than 0.001) in those who were often hypertensive according to their physician's evaluations. Thus a strong association of hypertension with apical hypertrophy was suggested. However, hypertension in this condition was usually mild and labile, the blood pressure reverted to the normal range within several days of hospital admission, implying that transient hypertension during daily activity is associated with apical hypertrophy. Therefore, blood pressure response during exercise stress tests of 25 patients with apical hypertrophy was compared with that of age- and sex-matched controls. Slopes of linear regression between systolic blood pressure and heart rate and oxygen consumption during exercise, were used as indices of blood pressure response. They were significantly greater in apical hypertrophy than in the controls (1.2 +/- 0.4 vs 0.9 +/- 0.3, p less than 0.01 and 4.3 +/- 1.7 vs 2.8 +/- 0.8, p less than 0.001). This trend was observed even in patients without histories of hypertension. These findings suggested that apical hypertrophy has an inheritance pattern different from that of ASH, and has a possible association with acquired factors such as hypertension. Therefore, apical hypertrophy seemed to be a disease entity distinct from HCM with ASH, though it might be included in the disease spectrum of HCM.

Prognosis in hypertrophic cardiomyopathy.

One hundred thirty-six patients with hypertrophic cardiomyopathy were followed up for 1 to 17 years. Twenty-one patients had died, 14 of them suddenly, two from heart failure, two from cerebral embolism, and three from noncardiac causes. Life table analysis revealed that sudden death was significantly associated with young age less than 20 years (relative risk [rr] = 8.63, when compared with those greater than 40 years) and with positive Master's single two-step test (rr = 3.55). Heart failure was more frequent in patients with positive Master's single test (rr = 4.27) and with left ventricular end-diastolic pressure greater than 20 mm Hg (rr = 2.58). Atrial fibrillation, observed in 15 patients, was a poor prognostic sign, resulting in five cardiac deaths and seven heart failures. In contrast, prognosis was favorable in patients with apical hypertrophy with giant negative T wave. Thus Japanese patients with hypertrophic cardiomyopathy showed a prognosis consistent with Western patients, except for excellent outcome of apical hypertrophy.

[Cross-sectional echocardiographic visualization of the infarcted site in myocardial infarction: correlation with electrocardiographic and coronary angiographic findings].

Three dimensional assessment of the site of myocardial infarct was performed using cross-sectional echocardiography in 68 patients with old myocardial infarction. Patients with a history or electrocardiographic findings suggestive of double or multiple infarctions were excluded from the study. In patients with abnormal Q waves in V1 to V3, a regional wall motion abnormality (asynergy) was observed in the anterior portion of the interventricular septum (IVS) and the anterior free wall of the left ventricle (LV) which was extended from the base to apex. Most of them had a significant stenosis in the left anterior descending artery (LAD) distal to the first diagonal branch. Patients with Q waves in V1 to V5 or V6 showed extensive asynergy in the anterior IVS, anterior and lateral free walls of the LV extended from the base to apex. LAD stenosis proximal to the first diagonal branch seemed to be the corresponding coronary lesion. In patients with Q waves in V3 to V5 or V6, asynergy was limited to the apical half of the anterior wall of the LV. In patients with Q waves in II, III and a VF, asynergy was observed in the basal half of the posterior wall and the posterior portion of the IVS.

Insulin-like activity of photochemically obtained monovalent monomeric concanavalin A in the presence of anti-concanavalin A antibodies: dependence on multivalency for stimulation of glucose oxidation of rat fat cells.

Insulin-like action of monovalent monomeric concanavalin A (m-Con A) was examined in rat adipocytes in the presence of anti-m-Con A antiserum. The antisera from rabbits injected with m-Con A reacted with not only monovalent monomeric but also tetravalent tetrameric concanavalin A (alpha-Con A) in Ouchterlony double diffusion analysis. m-Con A alone did not show any appreciable effect on glucose oxidation of adipocytes while it slightly inhibited glycerol release stimulated by epinephrine. In contrast, exposure of adipocytes to m-Con A in the presence of antibodies to m-Con A resulted in stimulation of glucose oxidation and inhibition of epinephrine-stimulated lipolysis. The stimulation and the inhibition with m-Con A in the presence of the antibodies were of the same degree as those with alpha-Con A. Both alpha- and m-Con A were slightly active in inhibiting 125I-labeled insulin binding. These results demonstrate that the ability of anti-m-Con A antiserum to aggregate m-Con A bound to receptors on the isolated-adipocyte plasma membrane allowed m-Con A to mimic the biological activity of insulin and that the aggregation of receptors for ligands other than insulin can induce insulin-like action in rat fat cells.

[Exercise two-dimensional echocardiography: correlation between exercise induced asynergy and coronary artery lesions].

Exercise two-dimensional (2-D) echocardiography was performed in patients with suspected coronary artery disease, and exercise induced left ventricular asynergy was evaluated qualitatively and was compared with the coronary artery stenosis and electrocardiographic ST changes. Subjects were 12 patients with angina of effort, 8 patients with spontaneous angina, 8 patients with chest pain syndrome with the normal coronary artery, and 7 patients with hypertrophic cardiomyopathy (HCM). Cases with myocardial infarction were excluded from this study. 1) Left ventricular asynergy during exercise was observed in 10 and ST depression in 11 of 12 patients with angina of effort. In patients with spontaneous angina, left ventricular asynergy and ST depression during exercise were observed in 2 of 8 patients without anginal pain, and both patients had coronary artery stenosis of 90% or more. 2) Exercise induced asynergy was also observed in 4 of 7 patients with HCM without coronary artery stenosis. It seemed likely that the markedly hypertrophied myocardium and impairment of left ventricular compliance and relaxation may induce relative myocardial ischemia.

Effect of reserpinization on insulin secretion in the rat.

Reserpine-induced supersensitivity to the insulin-releasing action of a beta-adrenergic agonist, isoprenaline, and of glucose was studied in vivo and in vitro. The subcutaneous injection of rats with reserpine (0.05 to 10 mg kg-1) enhanced the action of isoprenaline on insulin secretion. ED50 of isoprenaline for insulin secretion was changed little after reserpinization, whereas maximum effect of the beta-agonist was enhanced by pretreating rats with reserpine. Glucose-stimulated insulin secretion was also enhanced in the reserpinized rats. Pancreases isolated from the reserpinized rats secreted more insulin in response to phentolamine in the presence of glucose and isoprenaline. These results suggest that the supersensitivity in insulin secretion induced by reserpine may be non-specific.

Potentiation of insulin secretory response of rats by BCG injection.

Effects of BCG cells on blood insulin, glucose, glycerol and free fatty acid (FFA), and liver glycogen were examined in rats in vivo. Freund's complete adjuvant (FCA) was used as a solution containing BCG cells. Insulin secretion and lactate production by isoproterenol were enhanced by FCA injection. Hyperglycemia induced by epinephrine was somewhat attenuated by treating rats with FCA. The increment of plasma FFA by isoproterenol was smaller in FAC-injected rats than that in control rats. The amount of glycogen in liver was increased by FCA treatment Anti-insulin antiserum (AIS) did not modify the lactate producing action of isoproterenol. FFA release stimulated by isoproterenol was enhanced by AIS. Liver glycogen content in FCA-injected rats was decreased by AIS injection. These results suggest that some component(s) of BCG cells may enhance insulin release stimulated by secretagogues, and insulin thus released attenuates some metabolic effects of the agonist.

[Detection of exercise induced left ventricular asynergy by two-dimensional echocardiography (author's transl)].

In order to detect an exercise induced asynergy, cross-sectional echocardiography was performed during multistage maximal bicycle ergometer stress test in the supine position. 1) Left ventricular (LV) asynergy occurred earlier than or simultaneously with the appreciable ST segment change. 2) In patients with angina, LV asynergy appeared in the area of dominant coronary stenosis, while ST depression was seen in V3-6 as well as II, III and aVF, despite of the localized area of asynergy. 3) In patients with myocardial infarction, LV asynergy increased or extended over or around the infarcted area except one case, ST segment elevated in the leads over the infarction with abnormal Q waves and depressed in the reciprocal leads. These observations revealed that ST depression does not necessarily mean an occurrence of new ischemia over the corresponding area in myocardial infarction. Thus exercise cross-sectional echocardiography was demonstrated to be a good method to detect an exercise induced ischemia and would be particularly valuable in view of the coronary artery bypass.

[Secondary atypical hypertrophy: hypertrophic cardiomyopathy with acquired risk factors (author's transl)].

New clinical concept of "secondary atypical hypertrophy" was proposed for hypertrophic cardiomyopathy (HCM) associated with an acquired risk factors such as hypertension, strenuous exercise, etc, based on the following findings. 1) Case-control study suggested that history of hypertension, physical labor and weight gain were thought to be risk factors of apical hypertrophy. History of hypertension was also demonstrated in 55% of older cases of HCM (greater than 35 yrs) with asymmetric septal hypertrophy, and this figure was appreciably higher than that in general population (around 26% in our population survey). Thus hypertension was suggested to have a causal relationship to HCM as an important risk factor in older cases. 2) Even in cases without hypertension, 29% of non-obstructive HCM exhibited a marked increase in systolic blood pressure on bicycle ergometer stress test, suggesting an important of transient hypertension during exercise as a risk factor of unusual hypertrophy. 3) The cases of HCM with definite family history and the cases of HCM with secondary atypical hypertrophy, with acquired risk factors such as hypertension, presented different clinical features. The latter is older and predominant in males. In the former, QRS pattern was les distorted, suggesting milder congenital defect of the myocardium. Left ventricular function was less impaired in cases with secondary atypical hypertrophy if judged from diastolic descent rate of the mitral valve, left ventricular enddiastolic pressure and functional aerobic impairment (FAI). On the other hand, no differences were observed between cases with ventricular septal hypertrophy and left ventricular outflow obstruction. 4) Follow-up study demonstrated rather favorable prognosis in cases with secondary atypical hypertrophy because of the absence of sudden death despite of their older age. These distinctive difference in clinical features, prognosis and then management in cases with HCM with acquired risk factors suggested a validity of the concept of secondary atypical hypertrophy in clinical practice.