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Biomedical subjects

K Minakuchi

Publications and source records attributed to K Minakuchi.

At least 37 records · Page 2Linked to original sources
PubMed1998

Endothelin-1 stimulates sodium-dependent calcium efflux from bovine adrenal chromaffin cells in culture.

1. The effect of endothelin (ET)-1 on Ca2+ efflux from cultured bovine adrenal chromaffin cells was examined. ET-1 (10(-7) M) significantly increased intracellular free Ca2+ level ([Ca2+]i), 45Ca2+ uptake and catecholamine secretion in the cells. 2. ET-1 stimulated the efflux of 45Ca2+ from the cells preloaded with 45Ca2+ in a concentration-dependent manner (10(-9)-10(-7) M). This stimulatory effect was inhibited by ET(B) receptor antagonist BQ788, but not by ET(A) receptor antagonist BQ123. Selective ET(B) receptor agonists Suc-[Glu9, Ala11.5]-ET-1 and sarafotoxin S6c (SRTX) also stimulated 45Ca2+ efflux from the cells. 3. ET-1, Suc-[Glu9 Ala11.15]-ET-1 and SRTX increased the level of cyclic GMP in the adrenal chromaffin cells. ET-1 induced an increase in the nitric oxide (NO) level in the cells. The stimulatory effects by which ET- increases NO level and 45Ca2+ efflux were inhibited by NG-monomethyl-L-arginine acetate (L-NMMA), a competitive inhibitor of NO synthase. 4. The 45Ca2+ efflux stimulated by ET-1 was inhibited by deprivation of extracellular Na+, but not by deprivation of Ca2+. 5. These results suggest that ET-1 stimulates an extracellular Na+-dependent Ca2+ efflux through the activation of NO synthase in cultured bovine adrenal chromaffin cells.

Adrenal Glands↗
PubMed1998

Lung cancer: intermittent irradiation synchronized with respiratory motion--results of a pilot study.

PURPOSE: To test the feasibility of a system for intermittent irradiation synchronized with respiratory motion in a clinical setting. MATERIALS AND METHODS: A newly developed gate pulse controller that starts and stops irradiation at a chosen phase of the respiratory cycle by controlling a linear accelerator was used in six patients with lung cancer. A laser displacement sensor was used for the detection of respiratory motion. Three patients underwent radiation therapy during the cycles between 50% expiration and 50% inspiration (step 1), and three patients underwent radiation therapy during the cycles between 70% expiration and 30% inspiration (step 2). RESULTS: The system functioned well; irradiation was verified with portal verification radiography in all six patients. The range of the tumor position during synchronized irradiation was detectable with fast portal localization radiography. The treatment times for steps 1 and 2 were 1.38-1.71 and 2.03-2.18 times longer, respectively, than those for conventional irradiation. CONCLUSION: Synchronized irradiation with the authors' system allowed convenient and reliable reduction of the target volume. Further study is needed to standardize the system for clinical use.

Clinical Protocols↗
PubMed1998

Limited-stage small cell lung cancer: local failure after chemotherapy and radiation therapy.

PURPOSE: To evaluate radiation therapy regimens for improvement in local control in patients with limited-stage small cell lung cancer. MATERIALS AND METHODS: Radical radiation therapy results in 117 patients with limited-stage small cell lung cancer were retrospectively reviewed. The protocols in 90 patients were 40 Gy in 20 fractions (n = 28), 50 Gy in 25 fractions (n = 32), and 45 Gy in 30 fractions (accelerated hyperfractionation, n = 30). The other 27 patients received thoracic irradiation (dose range, 20-60 Gy; median dose, 54 Gy). All patients underwent systemic chemotherapy. RESULTS: The 5-year Kaplan-Meier survival rates in the patients with N0, N1, N2, and N3 disease were 26%, 34%, 18%, and 0%, respectively; the rates of in-field relapse were 25%, 36%, 26%, and 25%, respectively; and the rates of marginal relapse were 0%, 9%, 15%, and 29%, respectively. In 56% of patients with marginal relapse, the relapse site was at the upper margin. The 4-year in-field control rates for the patients who underwent 40, 50, and 45 Gy were 51%, 70%, and 56%, respectively. CONCLUSION: Patients with N3 limited-stage small cell lung cancer should undergo a separate protocol, and the upper margin should be extended in patients with N2 or N3 disease.

Adult↗
PubMed1998

Changes in plasma free and sulfoconjugated catecholamines during the perioperative period of cardiac surgery: effect of continuous infusion of dopamine.

In order to elucidate the pharmacological properties of the formation of sulfoconjugated catecholamines (CAs) in human plasma, we investigated the changes in the plasma levels of free and sulfoconjugated CA during the continuous infusion of dopamine (DA; 4-6 microg/kg/min for 24 h, followed by 3-4 microg/kg/min for 48 h) in patients who had undergone cardiac surgery. The plasma level of free DA increased immediately after the start of the infusion and reached a plateau within 1 h at a level of about 2000 times the basal value. In the control patients who had received non-cardiac surgery without DA infusion, plasma-free DA increased only 5-fold after their operation. The plasma level of DA sulfate increased linearly for 24 h, to 48-fold of the basal value by DA infusion, whereas it showed only a 2-fold increase in the control patients. After 24 h, due to reduction of the infused DA dose, the level of free DA gradually decreased, whereas the level of DA sulfate remained elevated. The plasma levels of free adrenaline (Ad) and noradrenaline (NA) also increased during the DA infusion, but their levels reached a plateau within 1-2 h. Sulfoconjugated Ad and NA increased progressively until the tapering off of DA infusion. In the control patients, both free and conjugated Ad and NA showed transient increases over 12 h after surgery. These results suggest that sulfoconjugation plays a role in regulating the plasma levels of excess free CA, thereby modifying the cardiovascular effects of circulating CA. Measurement of the increase in plasma conjugated CA may be useful as an index of the increase in free CA in plasma due to the administration of an exogenous form or release of endogenous CA from the tissues.

Adult↗
PubMed1998

[Anesthesia for tracheobronchial stent insertion].

We experienced nine cases of general anesthesia for tracheobronchial stent insertion. As far as possible, we anesthetized the patients under spontaneous respiration with inhalation anesthesia, depending on the types of the tracheobronchial stent (Dumon stent or expandable metallic stent), or respiratory status of the patient. Having experienced a case requiring PCPS (percutaneous cardiopulmonary support) to recover from severe ventilatory insufficiency due to tracheal perforation, we, in advance, inserted 18- or 20-gage cannulas into the femoral artery and vein for PCPS standby, and could manage general anesthesia safely for tracheobronchial stent insertion.

Adult↗
PubMed1997

Atrial natriuretic peptide stimulates Na+-dependent Ca2+ efflux from freshly isolated adult rat cardiomyocytes.

In the present study, we examined the effect of atrial natriuretic peptide (ANP) on Ca2+ efflux from freshly isolated adult rat cardiomyocytes. Rat ANP(1-28) stimulated the efflux of 45Ca2+ from the cells in a concentration-dependent manner (10(-8) M to 10(-6) M). The 45Ca2+ efflux was not affected by removal of extracellular Ca2+, but was dependent on the presence of extracellular Na+. In addition, rat ANP(1-28) caused 22Na+ influx into the cells. The 45Ca2+ efflux was also stimulated by C-type natriuretic peptide-22 (CNP-22), but not by rat brain natriuretic peptide-45 (BNP-45). It was also observed that both rat ANP(1-28) and CNP-22 stimulated guanosine 3',5'-cyclic monophosphate production within the cells. These results indicate that ANP stimulates Na+-dependent 45Ca2+ efflux from freshly isolated adult rat cardiomyocytes, probably through Na+/Ca2+ exchange, and that the stimulatory effect of ANP on Ca2+ efflux may be mediated via the natriuretic peptide receptor which has been shown to couple to guanylate cyclase. Since it is reported that Na+/Ca2+ exchange is important in calcium homeostasis within cells, ANP may play a role in the extrusion of intracellular Ca2+ from isolated adult rat cardiomyocytes.

Animals↗
PubMed1997

Difference in interaction of fibronectin with type I collagen and type IV collagen.

In our studies on fibronectin, difference in binding to type I collagen and type IV collagen was observed and analysed. Four different fragments, which consist of I6-II1-II2-I7-I8-I9, I6-II1-II2-I7, I6-II1-II2, and I8-I9 within the collagen binding domain, have been isolated from proteolytic digests of fibronectin. The N-terminal fragments of the collagen binding domain showed the binding affinity to both of type I and type IV collagens. On the other hand, the C-terminal portion of the domain, I8-I9, only bound to type I collagen. The newly developed monoclonal antibody FN 40, which recognizes type I9 homology repeat of the collagen binding domain, inhibited the binding of fibronectin to type I collagen in a dose-dependent manner. Three overlapping fragments, I6-II1-II2-I7, I6-II1-II2-I7-I8, and I7-I8-I9, which have been expressed in Escherichia coli, showed the similar binding affinity to type IV collagen, whereas the fragment containing type I9 repeat (I7-I8-I9) showed the significantly stronger binding activity to type I collagen. In addition, the expressed fragment lacking I9 competitively inhibited the binding of fibronectin to type IV collagen. In contrast, the interference of this fragment to type I collagen binding activity of fibronectin was not significant. These data indicate that the collagen binding domain contains at least 2 sites for interaction with each type of collagen and that type I9 repeat is important for type I collagen binding, whereas I7 is for type IV collagen binding. Chemical modification studies revealed that the three-dimensional structure of fibronectin is more important for type I collagen binding.

Antibodies, Monoclonal↗
PubMed1997

Structure of the rice glutaredoxin (thioltransferase) gene.

We have isolated the gene encoding a glutaredoxin in rice (Oryza sativa L.) and determined the nucleotide (nt) sequence of about a 4.2 kb long. The cloned gene (gRASC8) was found to contain four exons interrupted by three introns. The first exon begins the ATG translation start codon and the four exons code for a protein composed of 112 amino acids. The tetrapeptide -Cys-Pro-Phe-Cys- [-Cys-Pro-Phe(Tyr)-Cys-] which constitutes an active site of Escherichia coli and mammalian glutaredoxins, was conserved. The nt sequence contained consensus TATA and CAAT boxes, and two polyadenylation signals. Southern blot analysis of rice genomic DNA suggests that there are two copies of the glutaredoxin genes in rice.

Amino Acid Sequence↗
PubMed1997

Serotonin increases Na(+)-dependent Ca2+ efflux from bovine adrenal chromaffin cells in culture.

The effect of serotonin (5-hydroxytryptamine, 5-HT) on Ca2+ mobilization in bovine adrenal chromaffin cells in culture was examined. 5-HT (10(-5) M) did not increase secretion of catecholamine, uptake of 45Ca2+ and levels of intracellular free Ca2+ ([Ca/+]i). However, 5-HT (10(-8)-10(-5) M) stimulated the efflux of 45Ca2+ from cultured bovine adrenal chromaffin cells in a concentration-dependent manner. Its stimulatory effect on 45Ca2+ efflux was inhibited by cyproheptadine (a 5-HT1A and 5-HT2 receptor antagonist) or mianserin (a 5-HT2 receptor antagonist). The increase in 5-HT-stimulated 45Ca2+ efflux was dependent on extracellular Na+ concentration, but not extracellular Ca2+ concentration. These results indicate that stimulation of the 5-HT receptors induces extracellular Na(+)-dependent Ca2+ efflux from bovine adrenal chromaffin cells in culture, probably by acceleration of Na+/Ca2+ exchange.

Adrenal Glands↗
PubMed1997

Potentiation of histamine-induced catecholamine secretion by ouabain in cultured bovine adrenal chromaffin cells is dependent on calcium and sodium influx.

The effects of histamine on catecholamine secretion from cultured bovine adrenal chromaffin cells were studied in the presence of ouabain, an inhibitor of Na+-K+ ATPase. The purpose of this study was to determine whether Na+, as well as Ca2+, was involved in histamine receptor-mediated catecholamine secretion. Histamine (10(-8)-10(-5) M)-induced catecholamine secretion was markedly potentiated by addition of ouabain (10(-5) M) and was inhibited by a histamine-H1 receptor antagonist or incubation in a Ca2+-free medium. Histamine-induced 45Ca2+ influx was also potentiated by addition of ouabain. Ouabain alone or in the presence of histamine increased 22Na+ influx into the cells. In an additional set of experiments, cells were preincubated in the presence or absence of Na+ for 30 min (+/- histamine and ouabain), washed and then catecholamine secretion was measured following exposure to 2.2 mM Ca2+ for 15 min. Preincubation with histamine alone with or without Na+ had no effect of Ca2+-induced secretion of catecholamine. Preincubation with ouabain alone or with ouabain plus histamine produced a slight stimulation of catecholamine secretion in Na+-free medium and a large stimulation in Na+-containing medium. These results suggested that stimulation of the histamine-H1 receptor and inhibition of the Na+ pump both increase intracellular Na+ levels, resulting in increases in Ca2+ influx and catecholamine secretion.

Adrenal Medulla↗
PubMed1997

Purification and characterization of glutaredoxin (thioltransferase) from rice (Oryza sativa L.).

We purified and characterized glutaredoxin (thioltransferase), which catalyzes thiol/disulfide exchange reaction, for the first time in plants. The purification procedure employed an immunoabsorbent, antiglutaredoxin-Sepharose. Glutaredoxin was purified about 2,200-fold from rice bran and it appeared to be homogeneous on SDS-PAGE. MALDI-TOF mass spectrometry revealed that the protein has a molecular mass of 11,097.9 Da. Rice glutaredoxin consists of 105 amino acid residues, containing the tetrapeptide -Cys-Phe-Pro (Tyr)-Cys-, which constitutes the active site of Escherichia coli and mammalian glutaredoxins. Inactivation assay also indicated that cysteine residues are responsible for enzyme activity. Kinetic analyses revealed that the enzyme did not exhibit normal Michaelis-Menten kinetics. The enzyme has an optimum pH of about 8.7 with 2-hydroxyethyl disulfide as a substrate. In addition, rice glutaredoxin has dehydroascorbate reductase activity, like mammalian glutaredoxin.

Amino Acid Sequence↗
PubMed1997

Chondrocyte migration to fibronectin, type I collagen, and type II collagen.

It is well known that cellular interactions, such as cell adhesion, migration, invasion, between cells and the extracellular matrix are mediated by the integrin family of cell surface receptors. Chondrocytes are surrounded by an abundant extracellular matrix, but there is less information on the cellular receptors which interact with this matrix. In our studies, fibronectin, type I collagen, and type II collagen promoted haptotactic and chemotactic migration of chondrocytes, as determined using a modified Boyden chamber system. Treatment of chondrocytes with tyrosine kinase inhibitor, herbimycin or genistein, resulted in a dose dependent inhibition of migration toward these matrix proteins, whereas adhesion of chondrocytes was not influenced. This indicated the existence of functional relationships between protein tyrosine phosphorylation and chondrocyte migration following the adhesion of chondrocytes to matrix proteins. Further study showed that the peptide GRGDSP inhibited chondrocyte migration to fibronectin but not to collagens. On the other hand, chondrocytes migrated toward the tetra-RGD containing peptide, but not the peptide GRGDSP, in a dose dependent fashion. These observations suggest that cross-linking or clustering of integrins is essential to induce transmembrane signaling related to tyrosine phosphorylation for chondrocyte migration toward fibronectin.

Animals↗
PubMed1997

Establishment of a standardized assay system of fibronectin activity using fibronectin-mediated cell adhesion.

An in vitro assay of fibronectin (FN) was established based on the adhesion of baby hamster kidney (BHK) cells through the cell-binding domain of FN. Each well of a microtiter plate was coated with samples or various concentrations of standard FN. Bovine serum albumin was further coated to prevent the non-specific adhesion of the cells. Various numbers of BHK cells were plated and incubated. After washing out the non-attached cells, the number of attached cells was measured using neutral red (NR)-staining. The conditions for the assay were optimal when 1 x 10(5) cells/well were plated and incubated for 90 min. The linear relationship between the concentration of FN coated and the absorbance of NR was observed in the range of 0.1-1.0 micro/ml of FN. The inhibition of cell binding by the peptides containing an Arg-Gly-Asp (RGD) sequence demonstrated that this assay system depended on FN-mediated cell adhesion through the major cell-binding domain.

Animals↗
PubMed1997

Effect of K+ channel openers on K+ channel in cultured human dermal papilla cells.

Minoxidil sulfate and pinacidil, well-known activators of the ATP-sensitive K+ (KATP) channel, induce hair growth in clinical studies. The opening of K+ channels is thought to be an important mechanism in the regulation of hair follicles. In the present study, we used the patch clamp technique to characterize the K+ channels and tested the effect of K+ channel openers on K+ channels in cultured human dermal papilla cells. In dermal papilla cells, the Ca(2+)-activated K+ (KCa) channel with large conductance (179.3 +/- 13.1 pS in symmetrical 150 mM K+ solutions, n = 9) was dominant and we could not observe KATP channels in cell-attached and inside-out patches. In addition, minoxidil and pinacidil failed to activate KATP or KCa channels. In inside-out membrane patches, the channel was blocked by 10 mM tetraethylammonium ion, 2 mM 4-aminopyridine to the cytosolic face of the membrane or by lowering Ca2+ using 10 mM EGTA, but not by glibenclamide. In the cell-attached patch configurations, extracellular application of 1 mM sodium nitroprusside, a nitrovasodilator, activated the KCa channel. Methylene blue (2 mM) inhibited channel activation by sodium nitroprusside. Extracellular application of 20 mM dibutyryl cGMP activated the KCa channel, suggesting that channel activation is mediated by cGMP. Nitrovasodilators, which have no effect on hair growth, now appear to activate KCa channels in dermal papilla cells. These results suggest that increased K+ permeability itself in dermal papilla cells may not be sufficient for promotion of hair growth.

Cells, Cultured↗
PubMed1997

Effect of evodiamine on catecholamine secretion from bovine adrenal medulla.

The effect of evodiamine on catecholamine secretion from bovine adrenal medulla was investigated. Evodiamine, a bioactive component isolated from dry unripened fruit of Evodia rutaecarpa Bentham, was found to stimulate the secretion of catecholamine from perfused bovine adrenal medulla at a concentration of 10 microM and its effect persisted for at least 30 min. This stimulatory effect of evodiamine was abolished by omission of Ca2+ from the perfusion fluid. Evodiamine (0.1-10 microM) markedly enhanced the secretion of catecholamine from the adrenal medulla induced by acetylcholine (100 microM or high K+(56 mM). The secretion of catecholamine was promptly enhanced by acetylcholine or high K+, but returned to the control level on treatment for 20 min. However, when evodiamine was added to the perfusion fluid after acetylcholine or high K+ stimulation for 10 min, the secretion of catecholamine again increased greatly. These results indicate that evodiamine not only stimulated the secretion of catecholamine from bovine adrenal medulla but also reversed insensitivity of these cells to acetylcholine or high K+ stimulation.

Adrenal Medulla↗
PubMed1996

Sensitive quantitation of nitric oxide by EPR spectroscopy.

A recent method for NO detection is electron paramagnetic resonance (EPR) with ferrous and mononitrosyl dithiocarbamate (Fe2+ (DETC)2) for spin trapping [Menon, N.K., et al., J.Mol. Cell Cardiol., 23:389; 1991]. However, by this technique, we failed to detect the spectrum of the NOFe2+ (DETC)2 complex in biological systems because of the low solubility of Fe2+ (DETC)2 and rapid oxidation of NOFe2+ (DETC)2 complex. To overcome these problems, we modified this method by adding albumin to solubilize Fe2+ (DETC)2 and Na2S2O4 as a strong reductant to increase the sensitivity and stability of the EPR spectrum of the NOFe2+ (DETC)2 complex. The optimal concentrations of these reagents were 3.3 mM of Fe2+ and DETC, 33 mg/ml albumin and 2 M Na2S2O4. The detection limit was less than 10 pmol/ml under these conditions. By this modified method, we succeeded in quantifying NO production from porcine aorta induced by forskolin.

Animals↗
PubMed1996

Potentiation by ouabain of catecholamine secretion from bovine adrenal chromaffin cells in culture induced by pituitary adenylate cyclase-activating polypeptide: evidence for involvements of Na+ and Ca2+ movements.

The effect of pituitary adenylate cyclase-activating polypeptide (PACAP) on catecholamine secretion with ouabain, an inhibitor of Na(+)-K+ ATPase, in cultured bovine adrenal chromaffin cells was examined, to determine whether movement of Na+, as well as Ca2+, is involved in the secretory process. PACAP (10(-10)-10(-6)M)-induced catecholamine secretion was markedly potentiated by addition of ouabain (10(-5)M). When cultured cells were preincubated with PACAP for 30 min in Ca(2+)-free medium in the presence of ouabain and then stimulated for 15 min with Ca(2+)-containing medium without PACAP or ouabain, their catecholamine secretion was dependent on the external Ca2+ concentration, and 45Ca2+ influx into the cells was increased. When the cells had been preincubated with PACAP and ouabain in Na(+)-free sucrose medium, their Ca(2+)-induced catecholamine secretion was greatly reduced. PACAP increased 22Na+ influx into cells treated with ouabain. These results suggest that stimulation by PACAP and inhibition of the Na(+)-pump both increase the intracellular Na+ level, resulting in increase in Ca2+ influx and catecholamine secretion.

Adenylyl Cyclases↗