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Biomedical subjects

M Kunimoto

Publications and source records attributed to M Kunimoto.

At least 109 records · Page 6Linked to original sources
PubMed1987

p-Chloromercuribenzoate-induced dissociation of cytoskeletal proteins in red blood cells of rats.

Effects of p-chloromercuribenzoate (PCMB) on the cytoskeletal organization of rat red blood cells were studied. Upon incubation with 50 microM PCMB in 10 mM Tris-HCl (pH 7.4) at 37 degrees C for 30 min, 80% of actin and 45% of spectrin were released from the ghosts, resulting in the fragmentation of ghost membranes. Addition of 2 mM Mg2+ or 0.1 M KCl, or lowering incubation temperature to 0 degree C substantially inhibited the solubilization of the cytoskeletal proteins and the fragmentation of ghost membranes, which enable to examine the effects of PCMB on the interaction between transmembrane proteins and the peripheral cytoskeletal network. Decreased recoveries of transmembrane proteins, such as band 3 and glycophorin, in Triton shell fraction were observed in the ghosts incubated with PCMB either in the presence of Mg2+ or at 0 degree C. PCMB also inhibited the in vitro association of purified spectrin with spectrin-depleted inside-out vesicles through interaction with proteins in the vesicle, such as bands 2.1 and 3. In the PCMB-treated ghosts, intramembrane particles were highly aggregated, which further supports the PCMB-induced dissociation of the transmembrane proteins from the cytoskeletal network. The decreased recovery of glycophorin in the Triton shell fraction also observed in intact red blood cells upon incubation with PCMB. These results suggest that the main action of PCMB on red cell membranes under physiological condition, at higher ionic strength and in the presence of Mg2+, is to dissociate transmembrane proteins from the peripheral cytoskeletal network, which may modify functions of these proteins.

Actins↗
PubMed1987

Relationship between length-tension relation and 20,000 dalton myosin light chain phosphorylation in guinea-pig taenia caeci.

1. Relationship between length-tension relation and phosphorylation of 20,000 dalton myosin light chain (LC20) in guinea-pig taenia caeci was investigated. 2. At in situ length (Lb), a good linear correlation was obtained between isometric tension and LC20 phosphorylation in high-K+-stimulated muscle. 3. In 100 mM K+-stimulated muscle, the active tension decreased at muscle lengths other than Lb, but no significant decrease in degree of LC20 phosphorylation was observed. 4. These results suggest that in guinea-pig taenia caeci, the major portion of the decrease in active tension at muscle lengths other than Lb is not due to a decrease in degree of activation.

Animals↗
PubMed1986

Detection of metallothionein on nitrocellulose membrane using Western blotting technique and its application to identification of cadmium-binding proteins.

A convenient method for detecting metallothionein (MT) and other cadmium-binding proteins (Cd-BPs) in tissue cytosol was established. A sample that contains MT was separated on a sodium dodecyl sulfate-polyacrylamide gel and then electrophoretically transferred to a nitrocellulose membrane after reduction with 2-mercaptoethanol (2-ME) on the gel. The membrane was incubated in a buffer containing 109Cd and then subjected to autoradiography. MT-I and -II in rat liver cytosol were detected as radioactive bands together with a Cd-BP of 60,000 Da. On the other hand, three Cd-BPs of 40,000, 29,000, and 24,000 Da were detected in the cytosol when the reduction with 2-ME was omitted.

Animals↗
PubMed1986

Efflux of 45Ca from isolated smooth muscle cells of guinea-pig taenia caeci.

Single smooth muscle cells were isolated from guinea-pig taenia caeci by digestion with collagenase. The 45Ca desaturation curve from isolated cells, which were previously washed with Ca2+-free solution containing EGTA in Ca2+-free modified Locke solution, consisted of three components (half-time: 1.0, 3.8 and 12.4 min). The 45Ca efflux from isolated cells in the third component was significantly increased by caffeine. This increase was suppressed by procaine, but was not affected by La3+. These results suggest that, in guinea-pig taenia caeci, there are at least four Ca2+ compartments: superficial low and high affinity bound Ca2+ and cellular low and high affinity bound Ca2+. Caffeine releases Ca2+ from the cellular high affinity binding sites.

Animals↗
PubMed1986

Density increment and decreased survival of rat red blood cells induced by cadmium.

Male Wistar rats were injected with CdCl2 subcutaneously to examine in vivo effects of Cd on density and survival of red blood cells. During the 7 days after administration of 1.0 mg Cd/kg, the following sequence of events occurred: (1) a progressive increase in the amount of more dense red blood cells concomitant with a decrease in that of light red blood cells from the first to the third day; (2) an increase in the spleen weight at the third day; (3) a decrease in the hematocrit value and an increase in the amount of light red blood cells at the fifth day; and (4) a recovery of the hematocrit value at the seventh day. Five days after administration, the hematocrit value decreased in a dose-dependent mode and the decrease was significant at the 1% level at 1.0 and 1.5 mg Cd/kg. A highly significant splenomegaly was also observed at 0.5 to 1.5 mg Cd/kg. In order to label red blood cells in vivo, [3H]diisopropylfluorophosphate ([3H]DFP) was injected into rats. At Day 11, Cd at either 0.5 or 1.0 mg/kg was administered to [3H]DFP-prelabeled animals. Cd administration accelerated 3H-labeled red cell clearance from the blood. Six days after Cd administration, the radioactivity of red blood cells was 76 and 68% (P less than 0.05) of the control at 0.5 and 1.0 mg Cd/kg, respectively. In vitro treatment of rat red blood cells with either 0.5 or 1.0 mM CdCl2 also increased red cell density and accelerated in vivo clearance of red blood cells from the recipient circulation. These results show that Cd at low dose can cause anemia by increasing red cell density and by accelerating red cell sequestration, presumably in the spleen.

Animals↗
PubMed1986

Changes in membrane properties of rat red blood cells induced by cadmium accumulating in the membrane fraction.

When rat red blood cells were incubated in a cadmium (Cd)-free medium following 1-h pretreatment with 0.5 mM CdCl2, incorporated Cd was retained in the cell during 14-h incubation and progressively accumulated in the membrane fraction, especially in the cytoskeleton fraction. In parallel to this accumulation, red cell filterability decreased and echinocytic cells increased, although intracellular ATP was maintained at the control level. The echinocytic shape was maintained in ghosts and cytoskeletons prepared from the Cd-loaded cells. In addition, the association of bands 2.1, 3, 4.2, and 4.5 with cytoskeletons increased and dissociation of cytoskeletal networks at low ionic strength decreased as the incubation time increased. Pretreatment of red blood cells with Cd also induced a release of small vesicles. These vesicles contained hemoglobin but were depleted of spectrin and actin, showing a phospholipid composition similar to that of red cell ghosts. These results suggest that the organization of cell membranes, especially cytoskeletal networks, is altered by Cd accumulation in the cytoskeleton fraction, which results in acceleration of age-related changes of red blood cells such as shape change and decreased filterability.

Adenosine Triphosphate↗
PubMed1985

Vesicle release from rat red cell ghosts and increased association of cell membrane proteins with cytoskeletons induced by cadmium.

When rat red cell ghosts were incubated with 0.1-0.5 mM CdCl2 in 10 mM Tris-HCl (pH 7.4) at 37 degrees C for 30 min, they became irregular in shape and released small vesicles. The release of vesicles was dependent on the incubation temperature and Cd2+ concentration. The maximum release occurred at 37 degrees C in the presence of 0.2 mM Cd2+. The protein composition of Cd2+-induced vesicles was similar to that of the vesicles released from ATP-depleted red cells. Upon incubation with 0.1-0.2 mM Cd2+, more than 90% of the Cd2+ added to the incubation buffer was recovered in ghosts and 15-20% of the ghost Cd2+ was located on the cytoskeletons prepared by washing ghosts with 0.5% Triton X-100 solution containing 0.1 M KCl and 10 mM Tris-HCl (pH 7.4). Moreover, the cytoskeletons prepared from Cd2+-treated ghosts markedly contained cell membrane proteins, bands 2.1, 3, 4.2 and 4.5, and glycophorins. The association of bands 3 and 4.2 with cytoskeletons increased with increasing concentrations of Cd2+ added to the incubation buffer and saturated at 0.2 mM Cd2+. The solubilization of cytoskeletal proteins, bands 1, 2 and 5, from ghosts at low ionic strength was almost completely suppressed by preincubation of ghosts with 0.1 mM Cd2+. HgCl2, PbCl2 and ZnCl2 at 0.2 mM each also produced an increased association of cell membrane proteins with cytoskeletons, whereas CaCl2 and MgCl2 did not.

Actins↗
PubMed1985

An apparent acceleration of age-related changes of rat red blood cells by cadmium.

The effect of cadmium at nonhemolytic concentrations on rat red blood cells was examined in vitro and compared with that of mercury. When rat red blood cells were incubated at 37 degrees C with 0.5 to 2.0 mM CdCl2 for 60 min, age-related changes of red blood cells occurred. These changes developed in a dose-dependent manner and included increases in cell density and in the number of echinocytes and a decrease in filterability. The incubation of red blood cells with 0.5 and 1.0 mM CdCl2 resulted in their shortened in vivo survival. Neither metabolic blockade, such as a fall in the adenosine triphosphate and glutathione contents, nor potassium leakage occurred at nonhemolytic concentrations of Cd. The effect of HgCl2 on age-related changes of red blood cells occurred at 5- to 20-fold lower concentrations than those of CdCl2. These results show that age-related changes were apparently accelerated by nonhemolytic concentrations of Cd and may be, at least in part, due to the interaction of Cd with thiol residues of cell membranes.

Adenosine Triphosphate↗
PubMed1985

In vivo effects of nitrogen dioxide on the blood nitrate level and the Na+,K+-ATPase activity of red blood cells of rats.

Male Wistar rats were exposed to 0.4, 1.2 and 4.0 ppm NO2 for 13 weeks to examine the effects of NO2 on the blood nitrate concentration and the Na+,K+-ATPase activity of red blood cells. Exposures to 1.2 and 4.0 ppm NO2 caused an elevation of the blood nitrate level at the first, third and eighth week. The maximum concentration attained was 148% (P less than 0.001, at the third week) and 201% (P less than 0.001, at the eighth week) of the controls at 1.2 and 4.0 ppm NO2, respectively. On the other hand, the nitrate concentration was decreased to the control level at the second, fourth and thirteenth weeks. 0.4 ppm NO2 caused a progressive but slight increase in the blood nitrate concentration from the third week and reached the maximum (122% of the control, P less than 0.001) at the eighth week. The Na+,K+-ATPase activity decreased slightly from the first week upon exposure to 4.0 ppm NO2 and reached the minimum (72% of the control, P less than 0.05) at the third week. Subsequently, the activity was increased to 159% (P less than 0.001) of the control at the eighth week. Exposure to 0.4 and 1.2 ppm NO2 caused fundamentally similar but less significant alterations of the Na+,K+-ATPase activity.

Animals↗
PubMed1984

Effects of nitrate and nitrite, chemical intermediates of inhaled nitrogen dioxide, on membrane components of red blood cells of rats.

Rat blood was incubated at 37 degrees C for 60 min with either NaNO3 or NaNO2 to examine the relationship between the decrease in the hexose content and Ca2+,Mg2+-ATPase activity of red cell membranes, and NO3- and NO2-. The hexose content decreased depending on the NaNO2 concentration up to 100 microM reaching 76% (p less than 0.05) of the control value. NaNO3 had little effect on the hexose content. On the other hand, the Ca2+,Mg2+-ATPase activity decreased depending on the NaNO3 concentration up to 200 microM, where the activity reached 75% (p less than 0.01) of the control value. The effect of NaNO2 on this activity was smaller than that of NaNO3. The sialic acid content and the Na+,K+-ATPase activity did not show significant alterations by incubation with NaNO2 and NaNO3 at below 100 microM. To examine the in vivo effects of NO2- and NO3-, 50 mM NaNO3 was intravenously injected into rats five times at hourly intervals (dose: 1.0 ml/kg body weight), and blood was collected 1 hr after the last injection. The activities of Ca2+,Mg2+- and Na+,K+-ATPases of red cell membranes were decreased to 68% (p less than 0.05) and 80% of the control value, respectively. Reduction by injection of 50 mM NaNO2 was smaller than that by 50 mM NaNO3. The results show that the hexose content and the Ca2+,Mg2+-ATPase activity of red cell membranes were decreased by NO-x that increased in the blood during short-term exposure of rats to NO2.

Animals↗
PubMed1984

Composition and metabolism of fatty acids in phospholipids of density-separated red cells of rats.

Fatty acid compositions of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) and the rates of fatty acid esterification to these phospholipids (PL) were measured in intact rat red cell populations of different ages separated by density gradient centrifugation in order to clarify changes in membrane lipids of red blood cells during in vivo aging. Fatty acid compositions of PC and PE altered progressively as red cells became denser. Changes in unsaturated fatty acids occurred predominantly at the 2-position of PC and PE and those in saturated fatty acids at both positions. The esterification rates of 5 major fatty acids decreased as red cells became denser and those of oleic acid, linoleic acid and arachidonic acid to both PC and PE of fraction I cells (oldest cells) were 37-51% those of fraction IV cells (youngest cells). Reduction in the rates of fatty acid esterification appeared to occur in the course of red cell maturation because reticulocyte-enriched cell fractions showed 4.5-14.5 times higher rates of linoleic acid and arachidonic acid esterifications to PC and PE.

Animals↗