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Biomedical subjects

S Shimizu

Publications and source records attributed to S Shimizu.

At least 595 records · Page 33Linked to original sources
PubMed1993

Detection of high incidence of H-RAS oncogene point mutations in acute myelogenous leukemia.

We have been analyzing RAS p21 proteins and the DNA sequence of leukemic cells. We report here that these cells have high expression of H-RAS p21, which originates from point mutations of RAS oncogenes. The leukemic cells from six patients with acute myelogenous leukemia were separated from heparinized whole blood and bone marrow by a density gradient technique. The expression of RAS oncogenes was analyzed by a fluorescence-activated cell sorting with a panel of monoclonal antibodies. The high expression of DWP, which was reported to recognized activated RAS oncogene, was found in two patients and was associated with high levels of H-RAS expression. These facts prompted us to analyze the DNA sequence of RAS genes with an automated DNA sequencer. Unexpectedly, various kinds of H-RAS point mutations were found in all six cases, including two cases of hot-spot point mutation at codon 12, whereas K-RAS point mutation (no hot-spot point mutations) was found in six cases. The same H-RAS point mutations, at codons 10, 11, and 15, were found in all six cases. To our knowledge, there is no report on H-RAS point mutation in human leukemias. On the basis of these findings, we suggest that H-RAS point mutation together with p53 gene mutation may play an important role in leukemogenesis.

Base Sequence↗
PubMed1993

High-level expression of recombinant human soluble thrombomodulin in serum-free medium by CHO-K1 cells.

Cultivation of gene-engineered Chinese hamster ovary (CHO-K1) cells that produce recombinant human soluble thrombomodulin (rsTM) was investigated to optimize conditions for high-level expression of the protein in a serum-free medium. For economic protein production, oxygenation of cultures with pure O2 permitted sufficient cell growth for high rsTM production with only 1 g/l of microcarriers and a low foetal bovine serum concentration. A longer growth phase (over 5 days) with serum was important to establish sufficient growth of this cell line on the microcarriers for subsequent serum-free culture, and to support a long-term production phase (about 2 months). In the production phase, a high glucose concentration (6.15 g/l) in the serum-free medium was very effective for prolonging the harvest cycle interval. Under these conditions, up to 100 mg/l rsTM was expressed in the conditioned medium. The rates of glucose consumption (G) and lactate production (L) were measured periodically and their ratio (L/G ratio) correlated with rsTM productivity. When the average L/G ratio was lower, reflecting a lower lactate production rate due to appropriate oxygenation of the culture, the specific rsTM production rate increased. Thus it may be possible to estimate protein productivity from L/G ratios calculated from the glucose and lactate measurements.

Animals↗
PubMed1993

Two types of potassium channel regulated by ATP in pancreatic B cells isolated from a type-2 diabetic human.

Two types of K channel regulated by ATP were observed in pancreatic beta cells from a type-2 diabetic man. One type had a conductance of 67 pS at -70 mV in symmetrical 140 mM KCl and was inhibited by intracellular ATP with a half-maximal concentration of 40 microM. ATP inhibition was antagonised by ADP. Tolbutamide inhibited the whole-cell K currents half-maximally at 25 microM. This channel has properties similar to those found for the ATP-sensitive K channel in rodent and normal human beta cells. The second channel type observed was an ATP-activated K channel. It had a conductance of 37 pS at -70 mV in symmetrical 140 mM KCl and was activated half-maximally by 9 microM intracellular ATP. This channel was unaffected by 1 mM tolbutamide. In cell-attached patches, one beta cell out of four tested responded to 20 mM glucose with depolarization. The role of the ATP-activated K channel with respect to the (patho)physiology of the beta cell is uncertain.

Adenosine Diphosphate↗
PubMed1993

Phase IV group study of natural human interferon alpha (HLBI) on chronic myelogeneous leukaemia.

Under the control of Kanagawa CML/HLBI phase IV study group in Japan, 18 cases out of registered 30 cases of chronic myelogeneous leukaemia consisting of 17 chronic phase and 1 accelerated phase, during July, 1991 to January, 1992, were analyzed for their hematological responses and cytogentic responses preliminarily. Hematological response rate (PR + CR) was 83.3% including 50.0% of CR, as judged by Kimura's criteria after treatment with HLBI alone (16 cases) or/and with other chemotherapy (2 cases). The dosage and duration of HLBI therapy required to get into the complete remission ranged from 212 to 1272 millions IU and between 6 to 42 weeks (mean value was 20 weeks), respectively. The clonally proliferated leukocytes and decreased physiological hematopoiesis started to recover from 2 to 4 weeks and reached their normal ranges from 16 weeks after 6 millions IU of HLBI were administered every day. In the 4 cases examined, 3 cases showed minimal cytogenetic responses and a case showed no cytogenetic response. Slight and temporary adverse effects were observed in 15 out of 18 cases (83.3%) including fever, general malaise, appetite loss, eruption, diarrhea, glossitis, hypogustation, weight loss and local muscle pain.

Adult↗
PubMed1993

Inhibition of cell growth by bafilomycin A1, a selective inhibitor of vacuolar H(+)-ATPase.

Bafilomycin A1, a potent selective inhibitor of vacuolar H(+)-ATPase, inhibited the growth of a variety of cultured cells dose-dependently, including golden hamster embryo and NIH-3T3 fibroblasts, whether or not they were transformed, and PC12 and HeLa cells. The concentration of bafilomycin A1 for 50% inhibition of cell growth ranged from 10 to 50 nM. The dose response was nearly parallel with that of the bafilomycin A1-induced lysosomal pH increase. The degree of pH increase for growth inhibition produced by bafilomycin A1 was similar to that produced by NH4Cl in which little difference was recognized in effect among cell types.

3T3 Cells↗
PubMed1993

Comparative study of QRST values from body surface potential mapping, 12-lead ECGs, VCGs in detecting inferior myocardial infarction, and evaluating the severity of left ventricular wall motion abnormalities in simulated left bundle branch block.

The authors compared the ability of QRST time-integral values (QRST values) from body surface potential maps (BSPM), 12-lead electrocardiograms (ECGs), and Frank lead vectorcardiograms (VCGs) in diagnosing a prior inferior myocardial infarction (MI) in simulated left bundle branch block (LBBB). The study included 32 patients whose digitized ECGs were recorded simultaneously for BSPM, ECGs, and VCGs during normal sinus rhythm and during right ventricular pacing simulating LBBB (18 with and 14 without an inferior MI). QRST values were calculated in each lead point of ECGs. Data on 608 normal subjects were used as controls; mean +/- 2 SD was regarded as the normal range. The following parameters were derived: sigma DM, sigma DE, sigma DV, the sum of the differences between the normal mean QRST value, and the QRST value of a given patient in leads where the QRST value was less than the normal range ("-2 SD area") in BSPM, ECGs, and VCGs (Y lead). The correlation coefficients for sigma DM, sigma DE, and sigma DV between the two activation sequences were highly significant. Sensitivity and specificity were as follows: 89% and 93% for sigma DM > 100 mV.ms, 89% and 93% for sigma DE > 50 mV.ms, and 56% and 100% for sigma DV > 10 mV.ms, respectively. Although sigma DM, sigma DE, and sigma DV were significantly (P < .01) correlated with the asynergy index calculated from left ventriculograms, sigma DM showed the best correlation. QRST values from BSPM, ECGs, and VCGs provide information that is useful in detecting an inferior MI and in estimating the severity of left ventricular wall motion abnormalities in the setting of LBBB. Of the three parameters, BSPM showed the best correlation with the severity of left ventricular wall motion abnormalities.

Aged↗
PubMed1993

High blood O2 affinity and relationship of O2 uptake and delivery in resting muscle.

To clarify the effects of high oxygen (O2) affinity of blood under stagnant hypoxic conditions, we studied the relationship between VO2 and O2 delivery (CaO2.flow) in isolated dog gracilis muscles perfused with low and normal P50 blood. The high affinity blood was prepared by either short-term incubation (P50 = 24.8 +/- 1.0 Torr) or long-term refrigerated storage (P50 = 19.5 +/- 1.5 Torr) with sodium cyanate. VO2 of the resting muscle was independent of O2 delivery above a critical level (0.45 ml.min-1.100 g-1), but was dependent on it below this level. The critical O2 delivery (0.45 ml.min-1.100 g-1), the maximal O2 extraction ratio (0.62), and the plateau VO2 (0.28 ml.min-1.100 g-1) in the present perfusions with low P50 blood were similar to those with normal blood. "Critical PvO2" (i.e. PvO2 at the critical O2 delivery) was 33 Torr in the perfusions with normal blood but only 19 Torr in the low P50 perfusions. The perfusion pressure-blood flow relationship was not influenced by the affinity change. These results suggest that the true critical PvO2 in resting muscle is lower than 19 Torr and hence that the decrease in VO2 in resting skeletal muscle under stagnant flow conditions is not caused by a decrease in PO2 driving pressure, but by a decrease in the surface area available for diffusion due to a closure of capillaries induced by the low perfusion pressure.

Animals↗
PubMed1993

Induction of copper-zinc superoxide dismutase in gerbil hippocampus after ischemia.

To assess the role of Cu-Zn superoxide dismutase (CuZnSOD) in regulating cellular antioxidant defenses, we studied the induction of CuZnSOD mRNA by an in situ hybridization technique and of CuZnSOD protein by an immunocytochemical method in the gerbil hippocampus following 5 min of transient global ischemia. For hybridization, we synthesized 48-mer oligonucleotide (base 465-512) complementary to rat CuZnSOD mRNA. Northern blot analysis showed hybridization to a single band of molecular weight 0.65 kb. After 5 min of ischemia, the signal became stronger at 3 and 24 h and returned to the control level 3 days later. In situ hybridization histochemistry revealed an increase in labeling throughout the hippocampus, especially in the granular layer 3 h following ischemia. The increase was prolonged only in the CA1 pyramidal layer after 24 h and was eliminated within 3 days or later. Conversely, analysis by Western blotting revealed that the insult produced few effects on the induction of CuZnSOD protein. Immunocytochemistry for CuZnSOD revealed a reduced immunostaining in the CA1 pyramidal layer at 24 h of recirculation when the persistent expression of CuZnSOD mRNA was shown in the same area. Our findings suggest that the expression of endogenous CuZnSOD is temporarily stimulated by an ischemic insult without increasing the protein level. The prolonged increase in mRNA and the decrease in the protein of CuZnSOD in the CA1 neurons seem to imply an important role of the endogenous antioxidant enzyme that protects against the detrimental effects of superoxide radicals on delayed neuronal death.

Animals↗
PubMed1993

Exposure of human spermatozoa to the cumulus oophorus results in increased relative force as measured by a 760 nm laser optical trap.

Spermatozoa change their movement characteristics in response to different environmental conditions. To investigate the relative force of spermatozoa exhibiting different motility patterns, a laser optical trap was used. A laser beam at 760 nm was directed through a microscope objective and focused above the spermatozoa to create a three-dimensional optical trap. Spermatozoa were trapped at 300 mW, and laser power was reduced until spermatozoa could escape. The force generated by the flagellar movement was proportional to the laser power at which the spermatozoa escaped from the trap. Three motility patterns were studied: linear, hyperactivated, and cumulus-related. Mean escape power for spermatozoa displaying linear motility was 59.5 +/- 43 mW, for hyperactivated motility 122.3 +/- 67 mW (P < 0.0001) and for cumulus-related motility 200.6 +/- 44.2 mW (P < 0.0001). In this study, we showed that human spermatozoa generated more relative force upon exposure to the cumulus mass. The combination of small-amplitude lateral head displacement and higher relative force may produce a 'drilling' effect which is synergistic with the enzymatic digestion of the cumulus matrix during the fertilization process.

Female↗
PubMed1993

Inorganic pyrophosphatase of clofibrate-induced rat liver peroxisomes.

Clofibrate increased inorganic pyrophosphatase (PPase) activity in peroxisomes more than 12-fold (740 milliunits/head, 15.9 +/- 5.0 milliunits/mg protein) in rat liver. The distribution of cytochrome c oxidase and that of the PPase in a Nycodenz gradient suggested that the PPase is an original peroxisomal enzyme and not a mitochondrial contaminant: This was confirmed by second Nycodenz gradient centrifugation. The optimum pH of the peroxisomal PPase was about 8.5. The activity was specific to inorganic pyrophosphate (PPi), the Km value for PPi being 34.1 +/- 3.3 microM. It was strictly dependent on Mg2+ and showed a sigmoidal dose-response for Mg2+ with an S0.5 value of 100 microM. The activity was inhibited by Ca2+, p-chloromercuriphenylsulfonic acid, Hg2+, N-ethylmaleimide, and NaF. The functions of peroxisomal PPase are discussed.

Animals↗
PubMed1993

Structural study of the N-linked oligosaccharides of hepatocyte growth factor by two-dimensional sugar mapping.

The structures of the N-linked oligosaccharides on recombinant human hepatocyte growth factor (rh-HGF) expressed by Chinese hamster ovary (CHO) cells were studied by two-dimensional sugar mapping. The oligosaccharides released from the glycopeptides by peptide: N-glycosidase F (PNGase F) treatment were tagged with 2-aminopyridine at the reducing ends. The alpha-chain was linked by biantennary, triantennary, and tetraantennary oligosaccharides, but the dominant oligosaccharides linking the beta-chain were biantennary (> 85%). There was no significant difference in oligosaccharide structures between the two glycosylation sites on each chain, that is, Asn263 and Asn371 on the alpha-chain, and Asn535 and Asn622 on the beta-chain. The linkage of sialic acid to the non-reducing terminal galactose was identified as NeuAc alpha(2-3) by 1H-NMR spectrometry. The structures of the N-linked oligosaccharides from rat HGF were also studied. Triantennary oligosaccharides were obtained from the alpha-chain and a biantennary oligosaccharide was obtained from the beta-chain. This result indicates that the alpha-chain is also linked by higher branched oligosaccharides than the beta-chain in rat HGF.

Aminopyridines↗
PubMed1993

Malignant epithelioid schwannoma of the skin showing partial HMB-45 positivity.

A malignant epithelioid schwannoma occurred on the right second toe of a 30-year-old Japanese man. It was a firm, flesh-colored, benign-appearing nodule and measured 13 x 9 mm in diameter and 6 mm in height. To our knowledge, this is the first case of malignant epithelioid schwannoma occurring on the toe. Histopathology was characterized by a circumscribed nodule in the dermis that predominantly consisted of atypical large epithelioid cells with some spindle cells whose proliferation was similar to that of the Verocay bodies seen in ordinary schwannoma. Fontana-Masson staining demonstrated no melanin pigment in the tumor at the light microscopic level. The eosinophilic cytoplasm contained abundant glycogen and was positive for S-100 protein and HMB-45, as usually seen in melanomas. Electron microscopy revealed that there was an abundance of long-spacing collagen in the extracellular matrix, and the cells contained numerous dense-cored granules. But no definite melanosomes were observed in any stage. As far as we are aware, this is the first case of a malignant epithelioid schwannoma showing HMB-45 positivity.

Adult↗
PubMed1993

Further study of experimental gastritis in rats: endocrine and neurocrine profile.

We conducted this study to identify the endocrine and neurocrine mechanisms of gastric mucosal protection in rats with experimental atrophic gastritis (erosive atrophic antritis) induced by prolonged exposure to taurocholate. This resulting gastritis was characterized by a significant reduction of parietal cell mass, a decrease in mucosal thickness, decreased numbers of pyloric glands, infiltration by inflammatory cells, and fibrotic proliferation in the gastric mucosa. Mucosal erosions were also prominent. These morphologic and morphometric findings indicate the presence of erosive atrophic gastritis, as previously described. Fasting levels of serum gastrin increased significantly in the rats with gastritis versus controls, whereas the mucosal gastrin levels did not differ significantly from those of controls. Mucosal levels of somatostatin decreased significantly, and vasoactive intestinal peptide (VIP) increased significantly in the pyloric sphincter region. These findings suggest that these peptides and neuropeptides are involved in the induction of this form of gastritis. The peptides may play an important role in the mechanisms of gastric mucosal protection (i.e., gastrin is an aggressive and somatostatin is a defensive factor, and VIP promotes the reflux of bile into the stomach by relaxing the pyloric sphincter).

Animals↗
PubMed1993

A comparison of four solutions for cold storage of pancreatic islets.

Clinical pancreatic islet transplantation requires cold storage of islets for some hours. The best solution for cold storage will be that which maintains islet viability for the longest time. We have examined the viability of rat and human islet after storage at 4 degrees C for up to 6 days in Hanks' balanced salt solution, University of Wisconsin solution (UW), and 2 modified versions of this last solution, Sumimoto D (SD) and histidine-lactobionate (HL). The integrity of cold-stored rat and human islets of Langerhans has been examined using supravital stains and electron microscopy. In addition, the viability of cold-stored rat islets was tested by intraportal transplantation into syngeneic streptozotocin-induced diabetic recipients. The in vitro studies showed good preservation of islets stored in UW, SD, and HL for up to 72 hr. In comparison, storage for periods as short as 24 hr in HBSS markedly reduced islet integrity. The transplantation studies showed that rat islets cold stored in HBSS solution for 24 hr were not able to reverse experimental diabetes, whereas those stored in the other 3 solutions for 24 hr successfully reversed diabetes within 1 week of transplantation. After 48-hr cold storage, only islets preserved in HL solution were uniformly capable of producing functioning islet grafts. None of the tested solutions was able to maintain islet viability sufficiently to allow successful transplantation after more than 48 hr of cold storage. These experiments demonstrate that good islet viability is maintained for up to 24 hr of storage in UW, SD, and HL, and even after 48-hr cold storage in HL solution, whereas preservation in HBSS solution was deleterious to islet viability within 24 hr.

Adenosine↗
PubMed1993

Effects of right ventricular pacing on QRST isointegral maps in patients with and without myocardial infarction: body surface distribution of significant changes in QRST area compared to supraventricular complex.

To assess the effects of right ventricular (RV) pacing on body surface QRST distributions, we recorded QRST isointegral maps (I-maps) during sinus rhythm and RV pacing in 25 patients with anterior myocardial infarction (MI), 19 with inferior MI, and 14 without MI. The QRST values at each lead point recorded during sinus rhythm and RV pacing with an 87-lead system were analyzed with a paired t-test in each patient. An abnormal decrease in the QRST value of the I-map was assessed by the difference map, which indicated a "-2SD area," where the QRST integral value was less than the normal range (mean - 2SD) calculated from 608 normal individuals. The I-maps were similar during the two activation sequences in patients with and without MI. However, during RV pacing, QRST values significantly decreased over the upper right anterior chest and increased over the lower left anterior chest and back. The sigma DMs (sum of QRST integral values below the normal range) for both activation sequences were strongly correlated in patients with anterior MI and with inferior MI (r = 0.91 and r = 0.92, respectively; P < 0.001). Although small but significant changes in QRST values were detected, the distribution of the "-2SD area" and the sigma DM were similar during both activation sequences in patients with prior MI. Thus, these findings demonstrate that an altered activation sequence produces small but significant changes in QRST values but that I-maps still provide information that is useful for the diagnosis of MI during RV pacing.

Adult↗