Biomedical subjects
Publications and source records attributed to H Sugihara.
A case of malignant cystosarcoma phyllodes with metastasis in the interpectoral lymph node (Rotter's) is presented in this paper. To the author's best knowledge, this is the first case in Japan on this disease, with lymph node metastasis. Although the surgical management of the disease has not yet been standardized, radical or modified radical mastectomy is thought to be the appropriate procedure.
Two hemorrhagic toxins, HR-2a and HR-2b were originally isolated from Trimeresurus flavoviridis (Habu) venom by Takahashi and Ohsaka (1970). It was reported by the original investigators that no proteolytic activity was detected when casein was used as the substrate. HR-2a and HR-2b were isolated in this laboratory and their proteolytic activities were tested using a variety of different substrates and assay methods. HR-2a and HR-2b were found to contain, respectively, 200 and 219 amino acid residues. Toxicological and biochemical properties of HR-2a and HR-2b were further investigated and are reported in this paper.
Phosphodiesterase was isolated from the venom of Crotalus ruber ruber from the U.S.A. using the gel filtration on a Sephadex G-75 column, followed by anion or cation exchange chromatography. Phosphodiesterase was homogeneous as established by a single band on acrylamide gel electrophoresis and isoelectric focusing electrophoresis. Phosphodiesterase activity was inhibited by ethylenediamine tetraacetic acid (EDTA), o-phenanthroline, thioglycolic acid or p-chloromercuribenzoate (PCMB), but not by soybean trypsin inhibitor (SBTI) or benzamidine. The molecular weight of this enzyme was determined to be approx. 98,000 and the isoelectric point was found to be pH 10.5 by isoelectric focusing with carrier ampholyte. This enzyme contained 1.04 mol zinc per mol. The Michaelis constant (Km) of this enzyme for p-nitrophenyl thymidine-5'-phosphate and inhibition constant (Ki) for PCMB were found to be 8.3 X 10(-3) and 1.2 X 10(-2) M, respectively.
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A radioimmunoassay (RIA) for human pro-LRF(14-69)OH was developed with an antiserum, generated in a rabbit, to [Tyr67]pro-LRF(47-67)NH2 conjugated to BSA. This antiserum bound 28-32% of [125I]pro-LRF(14-69)OH at a final dilution of 1:2500 and the binding was inhibited by pro-LRF(14-69)OH in a dose-dependent manner. The sensitivity of the RIA was 31.2-62.5 pg and the dose that inhibited 50% of the binding to the tracer was 280-320 pg. Intra- and inter-assay coefficients of variation at 50% inhibition were 8 and 12%, respectively. Neither LRF nor pro-LRF(14-37)OH was recognized by the antiserum. The dilution curve generated with human hypothalamic extract was parallel to that of pro-LRF(14-69)OH. In addition the extract yielded a major immunoreactive peak emerging in elution volumes concordant with [125I]pro-LRF(14-69)OH on Sephadex G-50 chromatography.
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To evaluate left ventricular diastolic properties during acute myocardial ischemia, transmitral flow patterns were analyzed by two-dimensional pulsed Doppler echocardiography. In a preliminary study of transient coronary artery occlusion in anesthetized open-chest dogs, the following results were obtained: decreased peak velocity of rapid filling, prolonged rapid filling period, and compensatory increase in the peak velocity of atrial filling. These changes occurred nearly concomitantly with decreased cardiac output, elevated left atrial and left ventricular end-diastolic pressures, and decreased left ventricular systolic pressures. With reperfusion of the coronary artery, these parameters rapidly returned to the base line values. In acute and transient myocardial ischemia induced by intracoronary balloon inflation during therapeutic percutaneous transluminal coronary angioplasty (PTCA) in man, similar changes in these parameters were observed, and the changes were more marked in PTCA for the left anterior descending artery as compared with that of the right coronary artery. In conclusion, noninvasive two-dimensional pulsed Doppler echocardiography is of practical use in evaluating deterioration of the diastolic function of the left ventricle during acute and transient myocardial ischemia.
The feasibility of spatial color mapping vectorcardiography in diagnosing the site and extent of the old myocardial infarct was evaluated in comparison with 201T1 scintigraphy. Vectorcardiographic data made by Frank's technique were entered in the personal computer and the QRS complexes were plotted over 112 points crossing the latitude of each 20 (from 20N to 80S) degrees and longitude of each 20 degrees from 20W to 20E on the spherical body. Eight colors were assigned according to the direction of the QRS vector. To construct a territorial map, the apical, then, the septal, anterior, lateral, inferior and posterior portions were determined from the normal 201Tl scintigrams. Myocardial images obtained in the anterior, left anterior oblique and lateral projections were analyzed by the circumferential profile curve. Diagnostic specificity for infarction was greater than 60%, and especially high (90%) in the anterior, apical and lateral regions, but relatively low (50%) in the septal, inferior and posterior regions. Accuracies exceeded 64%. The infarcted areas assessed by spatial color vectorcardiography agreed with those assessed by 201T1 myocardial scintigraphy. Thus, spatial color vectorcardiography was simple and useful for diagnosing the site and extent of myocardial infarction.
Right ventricular (RV) systolic and diastolic functions were assessed in patients with previous anteroseptal myocardial infarction to ascertain the influence of interventricular septal ischemia on RV function. Gated right ventriculography with continuous infusion of krypton-81 m was performed in 12 normal subjects and 28 patients with infarction but without significant stenosis of the right coronary artery. Furthermore, RV contractile reserve by postextrasystolic potentiation was evaluated by gated radionuclide ventriculography with 99mTc-HSA. The patients with anteroseptal infarction were divided into two groups by the presence or absence of three hours' redistribution in the septal region on exercise thallium-201 myocardial scintigraphy. Two indices of systolic function (ejection fraction and the peak ejection rate) and three indices of diastolic function (1/3 diastolic filling rate, the peak filling rate and time to the peak filling rate) were derived from the right ventricular time-activity curve and its derivative curve. There was no difference in systolic function among normal subjects and patients with or without redistribution. However, diastolic function was impaired only in the patients without redistribution. The RV contractile reserve in the patients without redistribution was less than in those with it. Thus, RV systolic function was maintained in the patients with anteroseptal infarction, but contractile reserve deteriorated only in severe septal ischemia. Similarly, diastolic function was maintained in mild septal ischemia, but impaired in severe septal ischemia. We concluded that RV systolic and diastolic functions are closely related to interventricular septal ischemia.
To evaluate right ventricular (RV) function in severe left ventricular (LV) failure, we measured RV and LV ejection fractions (EF) in 18 patients with old myocardial infarction (OMI) and in 18 with dilated cardiomyopathy (DCM) using cardiac blood pool scintigraphy. In patients with OMI, RVEF was significantly greater in stage II LV failure than in stage III (functional class of the New York Heart Association, 47 +/- 8% and 28 +/- 12%, respectively: p less than 0.01), and this correlated well with exercise tolerance by bicycle ergometer and mean pulmonary artery pressure (r = 0.83 and r = -0.71, respectively). In patients with DCM, however, there was no correlation between RVEF and these indexes. After the oral administration of denopamine (beta 1 effector), both RVEF and LVEF increased in patients with OMI (35 +/- 13% to 45 +/- 12%, and 27 +/- 9% to 30 +/- 10%: p less than 0.01), but they did not change significantly in patients with DCM. These results indicate that RVEF in patients with OMI correlates well with subjective symptoms, exercise tolerance and RV afterload, but these correlations were not apparent in patients with DCM. We concluded that RV function in cases of severe LV failure has a different meaning between OMI and DCM.
Mature white fat cells (unilocular fat cells) have generally been considered to be in terminal differentiation and, hence, to have no proliferative ability. A new method, referred to as "ceiling culture," has been devised in our laboratory to culture unilocular fat cells in vitro. Under such culture conditions, the fat cells continue to exhibit specific functions of lipid metabolism and proliferate extensively. Intracytoplasmic lipid droplets did not inhibit division of the cells. There were two modes of proliferation of unilocular fat cells: "loculus-dividing" cell division, in which the single loculus of fat in the dividing cell was broken down into multiple droplets and distributed evenly between the daughter cells, and "loculus-preserving" cell division, in which the loculus in the dividing cell was minimally broken down and inherited with its shape preserved by one of the daughter cells with the other getting only a small number of fine lipid droplets. Such findings suggest that unilocular fat cells in mature fat tissue in vivo are probably capable of proliferation in such modes under some conditions.
A kinin-releasing enzyme was isolated from Bitis arietans (puff adder) venom by Sephadex G-100 and DEAE-cellulose column chromatographies. The kinin-releasing enzyme was shown to be homogeneous as demonstrated by a single band on acrylamide gel electrophoresis, isoelectric focusing, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunodiffusion. Its molecular mass is approximately 45 kDa with an isoelectric point of 6.5. Kinin-releasing enzyme possesses proteolytic activity which hydrolyzes the Leu6-Cys7, His10-Leu11 and Ala14-Leu15 bonds of the B chain of oxidized insulin and the A alpha and B beta chain of fibrinogen. Kinin-releasing and benzoyl-L-arginine ethyl ester hydrolytic activities of this enzyme were inhibited by diisopropyl fluorophosphate, suggesting that the serine hydroxyl group is involved in enzymatic activities.
A spectrophotometric method for inorganic phosphate determination in the presence of thiol compounds is described. Thiol compounds, which interfere with the measurement of inorganic phosphate by a modification of the method of Gomori, are removed by carboxymethylation by iodoacetate prior to the formation and reduction of phosphomolybdate complex. A linear standard curve is obtained by this method, and the method is suitable for the assay of a phosphate-releasing enzyme when the measurement must be performed in the presence of thiol compounds.
Two sets of cDNA clones were isolated from cDNA libraries prepared from poly(A+) RNA of rabbit lung and spleen by screening with the cDNA probe for the large subunit (80-kDa subunit) of chicken calcium-dependent protease (Ca2+-protease; Ohno, S., Emori, Y., Imajoh, S., Kawasaki, H., Kisaragi, M., and Suzuki, K. (1984) Nature 312, 566-570). The two sets of clones were identified as cDNA clones for two Ca2+-protease isozymes with high (mu-type) and low (m-type) calcium sensitivities from a comparison of the primary structures deduced from the nucleotide sequences with partial amino acid sequences from the two isozymes. The cDNA clones for the 80-kDa subunits of the mu- and m-type Ca2+-proteases contained, in total, about 1.5- and 2.2-kilobase cDNA inserts, respectively, which correspond roughly to the C-terminal halves of the coding regions and the entire 3'-noncoding regions. The two isozymes are encoded by two distinct mRNA species present in all the tissues examined, although the amount of mRNA significantly differs among the various tissues. Four E-F hand structures, typical calcium-binding structures in various calcium-binding proteins such as calmodulin, were detected in the C-terminal regions of both isozymes, as in the case of chicken Ca2+-protease. Comparison of the amino acid sequences of the two rabbit isozymes and the corresponding region of the chicken enzyme revealed marked homology, which indicates that these three enzymes have the same evolutionary origin. Furthermore, we suggest that the mu-type rabbit Ca2+-protease, rather than the m-type, is similar to chicken Ca2+-protease, which is regarded as an m-type enzyme in the C-terminal region. The evolution and molecular basis of the differences in calcium sensitivities of the Ca2+-proteases are discussed.
Female and male rats were treated with monosodium glutamate (MSG; 4 mg/g b. wt.) as neonates and the capability of the pituitary gland to secrete growth hormone (GH) in response to an intravenous injection of human GH-releasing factor (GRF) was evaluated under pentobarbital anesthesia on 109 days of life. Immunoreactive GRF content in the pituitary stalk-median eminence tissue in MSG-treated rats was less than 20% of that of control. A significant dose-dependent plasma GH response was observed after the administration of two doses of human GRF (0.25 and 1 microgram/kg b. wt., i.v.) in both control and MSG-treated rats. The responses between MSG-treated and control rats were comparable in female rats, but they were significantly reduced in male MSG-treated rats. These results show that the pituitary's responsiveness to exogenous GRF is well preserved in MSG-treated rats despite prolonged and severe depletion of endogenous GRF and there exists a sex difference in the effect of MSG on GH secretion elicited by GRF.
Ehrlich cancer cells and inflammatory cells in mouse ascitic fluid were hydrolyzed and stained with acridine orange (AO). The AO hydrolysis curves for G1/G2 + M phase cancer cells and inflammatory cells were differentially determined using flow cytometry by monitoring the metachromatic red-shifted fluorescence of the fluorochrome bound to the single-stranded DNA produced by acid hydrolysis. By computer fitting of the Bateman function to the hydrolysis curves, the kinetic parameters k1 (rate constant for the production of single-stranded DNA), k2 (rate constant for the degradation of the produced single-stranded DNA), and y0 (theoretical value of the single-stranded DNA present initially) were determined. It was found that the k2 value, which reflects the degree of DNA instability, was much higher for cancer cells in both the G1 and G2 + M phases than for inflammatory cells. This finding led us to develop a method for the differential AO staining of cancer cells and non-cancerous cells utilizing the different degree of DNA instability at acid hydrolysis. AO staining after hydrolysis with 2N HCl at 30 degrees C for 8.5 min was found to be the optimal method. In the 60 cases of human malignant epithelial and nonepithelial tumors tested, all of the malignant tumor cells emitted metachromatic red fluorescence, while all of the nonmalignant tumor cells (5 cases of benign tumor) and normal cells emitted orthochromatic green fluorescence when observed with a violet excitation light under a fluorescence microscope. This new technique can be a useful tool for the screening of malignancy in exfoliative cytology and also for basic cancer research.