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D T Yamaguchi

Publications and source records attributed to D T Yamaguchi.

At least 37 records · Page 2Linked to original sources
PubMed1991

Prostaglandins enhance parathyroid hormone-evoked increase in free cytosolic calcium concentration in osteoblast-like cells.

Prostaglandins (PGs) are autocrine or paracrine hormones that may interact with circulating hormones such as parathyroid hormone (PTH) in bone. We examined the interaction of the PGs, PGF2 alpha, PGE2, and 6-keto-PGF1 alpha with PTH to enhance the rapid, initial transient rise in free cytosolic calcium ([Ca2+]i) and cAMP levels stimulated by PTH. Pretreatment of UMR-106, MC3T3-E1, and neonatal rat calvarial osteoblast-like cells by PGs resulted in an enhancement of the early transient rise in [Ca2+]i stimulated by PTH. PGF2 alpha was approximately 100 times more potent than PGE2. PGE2 itself was more potent than 6-keto-PGF1 alpha in enhancing PTH-stimulated rise in [Ca2+]i. Near-maximal augmentation was achieved at PGF2 alpha doses of 10 nM and PGE2 of 1 microM. The degree of augmentation in [Ca2+]i by PGF2 alpha was independent of preincubation time. PGF2 alpha pretreatment did not alter the EC50 for the PTH-induced [Ca2+]i increase but only the extent of rise in [Ca2+]i at each dose of PTH. The augmented increase in [Ca2+]i was mostly due to enhanced PTH-mediated release of Ca2+ from intracellular stores. PGF2 alpha did not stimulate an increase in PTH receptor number as assessed by [125I]-PTH-related peptide binding. PG pretreatment partially reversed PTH inhibition of cell proliferation, suggesting that an increase in [Ca2+]i may play a role in tempering the anti-proliferative effect of PTH mediated by cAMP. These studies suggest a new mode by which PGs can affect cellular activity.

6-Ketoprostaglandin F1 alpha↗
PubMed1991

Cellular immunosenescence: an overview.

Recent studies on space flights suggest that certain T cell immunologic activities are vulnerable to microgravitation. It would be desirable to know the extent to which these changes can be prevented or reversed. Since the changes observed are analogous to the effects of aging on immunity, a brief overview is presented of our current knowledge of age-related changes in immune cells and of the various interventional methods which have been used successfully in preventing the decline with age and in elevating the levels of immune functions of old individuals.

Aging↗
PubMed1990

Cytosolic pH regulation in osteoblasts. Regulation of anion exchange by intracellular pH and Ca2+ ions.

Measurements of cytosolic pH (pHi) 36Cl fluxes and free cytosolic Ca2+ concentration ([Ca2+]i) were performed in the clonal osteosarcoma cell line UMR-106 to characterize the kinetic properties of Cl-/HCO3- (OH-) exchange and its regulation by pHi and [Ca2+]i. Suspending cells in Cl(-)-free medium resulted in rapid cytosolic alkalinization from pHi 7.05 to approximately 7.42. Subsequently, the cytosol acidified to pHi 7.31. Extracellular HCO3- increased the rate and extent of cytosolic alkalinization and prevented the secondary acidification. Suspending alkalinized and Cl(-)-depleted cells in Cl(-)-containing solutions resulted in cytosolic acidification. All these pHi changes were inhibited by 4',4',-diisothiocyano-2,2'-stilbene disulfonic acid (DIDS) and H2DIDS, and were not affected by manipulation of the membrane potential. The pattern of extracellular Cl- dependency of the exchange process suggests that Cl- ions interact with a single saturable external site and HCO3- (OH-) complete with Cl- for binding to this site. The dependencies of both net anion exchange and Cl- self-exchange fluxes on pHi did not follow simple saturation kinetics. These findings suggest that the anion exchanger is regulated by intracellular HCO3- (OH-). A rise in [Ca2+]i, whether induced by stimulation of protein kinase C-activated Ca2+ channels, Ca2+ ionophore, or depolarization of the plasma membrane, resulted in cytosolic acidification with subsequent recovery from acidification. The Ca2+-activated acidification required the presence of Cl- in the medium, could be blocked by DIDS, and H2DIDS and was independent of the membrane potential. The subsequent recovery from acidification was absolutely dependent on the initial acidification, required the presence of Na+ in the medium, and was blocked by amiloride. Activation of protein kinase C without a change in [Ca2+]i did not alter pHi. Likewise, in H2DIDS-treated cells and in the absence of Cl-, an increase in [Ca2+]i did not activate the Na+/H+ exchanger in UMR-106 cells. These findings indicate that an increase in [Ca2+]i was sufficient to activate the Cl-/HCO3- exchanger, which results in the acidification of the cytosol. The accumulated H+ in the cytosol activated the Na+/H+ exchanger. Kinetic analysis of the anion exchange showed that at saturating intracellular OH-, a [Ca2+]i increase did not modify the properties of the extracellular site. A rise in [Ca2+]i increased the apparent affinity for intracellular OH- (or HCO3-) of both net anion and Cl- self exchange. These results indicate that [Ca2+]i modifies the interaction of intracellular OH- (or HCO3-) with the proposed regulatory site of the anion exchanger in UMR-106 cells.

Animals↗
PubMed1990

Hydrogen peroxide inhibits iodide influx and enhances iodide efflux in cultured FRTL-5 rat thyroid cells.

This study describes the effects of hydrogen peroxide on the two iodide transport systems, I influx and I efflux, in the cultured FRTL-5 rat thyroid cells. I influx was measured by the amount of I taken up by the cells during incubation with Na125I and NaI for 7 min, and I efflux was measured by calculating the rate of 125I release from the 125I-loaded cells in the presence and absence of 5 mmol/l H2O2. Exposure to greater than 100 mumol/l H2O2 for 40 min caused a significant inhibition of I influx; the inhibition was reversible and non-competitive with iodide. Thyroid Na+K+ ATPase activity, a major mechanism to drive I influx, decreased by 40% after the cells were exposed to 5 mmol/l H2O2 for 10 min. H2O2 enhanced I efflux only when Ca2+ was present in the medium. The mechanism of an enhanced I efflux by H2O2 appears to be mediated through the elevation of free cytosolic Ca2+ concentration. Our data indicate that H2O2 can affect I transport by inhibiting I influx and enhancing I efflux.

Animals↗
PubMed1989

Classification of prostaglandin receptors based on coupling to signal transduction systems.

A wide spectrum of prostaglandins (PG) stimulate both the production of cyclic AMP and an increase in free cytosolic Ca2+ concentration [( Ca2+]i) in the osteogenic osteosarcoma cell line, UMR-106-01, which has characteristics compatible with osteoblasts. Using PG-stimulated determinations of the second messengers cyclic AMP and [Ca2+]i, a method for classification of PG receptors is presented. UMR-106-01 cells demonstrate three subclasses of PG receptors. One receptor interacts with PGF2 alpha, PGD2, and thromboxane B2 (TxB2) to increase [Ca2+]i. A second receptor binds PGE2, PGE1, PGI2, PGA2 and 6-oxo-PGF1 alpha to increase [Ca2+]i by stimulation of a second separate phospholipase C pool. A third receptor accepts PGE2, PGE1, PGA2, PGI2 and to a lesser extent PGF2 alpha, PGD2 and TxB2 to increase cyclic AMP. Such a classification system may be applicable to other cells responding to multiple PGs by inducing changes in cellular second messengers.

Adenylyl Cyclases↗
PubMed1989

1-Oleoyl-2-acetylglycerol promotes immunoglobulin production independent of cell proliferation in human peripheral blood mononuclear cells.

1-Oleoyl-2-acetylglycerol (OAG) stimulated IgG and IgM production in a dose-dependent manner in human peripheral blood mononuclear cells (PBM) but not PBM proliferation. 12-O-Tetradecanoyl phorbol-13-acetate (TPA) did not stimulate Ig production. OAG did not stimulate an increase in IL-2 generation or IL-2 receptor expression. H-7, a protein kinase C blocker completely inhibited OAG-stimulated Ig production. The results suggest that OAG stimulation of Ig production is independent of cell proliferation; a generalized increase in T-cell activation does not appear to be necessary in the OAG stimulation of Ig production. Finally, PBMs respond differently to OAG and TPA although both are protein kinase C activators.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Characterization of volume-sensitive, calcium-permeating pathways in the osteosarcoma cell line UMR-106-01.

Measurements of cell volume changes, free cytosolic Ca2+ concentration [( Ca2+]i) with Fura 2 and cell membrane potential with 3,3'-dipropylthiodicarbocyanine iodide were used to study the effect of cell volume change on Ca2+ influx and the membrane potential of the osteoblastic osteosarcoma cell line, UMR-106-01. Swelling the cells by hypo-osmotic stress was followed by reduction in cell volume which was markedly impaired by removal of medium Ca2+. Accordingly, cell swelling resulted in [Ca2+]i increase only in the presence of medium Ca2+. The cell swelling-activated Ca2+ entry pathway was active at resting membrane potentials, and Ca2+ influx through this pathway markedly increased upon cell hyperpolarization. A linear relationship between Ca2+ entry and the potential across the plasma membrane was observed. Thus, the volume-activated Ca2+ permeating pathway in UMR-106-01 cells has conductive properties. These pathways do not spontaneously inactivate with time when the cells are not allowed to volume regulate. The pathway can be blocked by micromolar concentrations of nicardipine and La3+ but display very low sensitivity to diltiazem and verapamil. Activation of the volume-sensitive, Ca2+ permeating pathway was not dependent on an increase in [Ca2+]i. Likewise, activation of the pathway was independent of a change in membrane potential between -85 and -3 mV. The increase in [Ca2+]i resulted in hyperpolarization of the cells, probably due to activation of Ca2+-activated K+ channels. The volume-sensitive pathways were partially active under isotonic conditions. Their activity was inhibited by cell shrinkage and increased by cell swelling. The pathways were sensitive to small changes in cell volume, particularly around a medium osmolarity of 310 mosM.

Calcium↗
PubMed1989

Properties of the depolarization-activated calcium and barium entry in osteoblast-like cells.

Measurements of free cystolic Ca2+ ([Ca2+]i) and Ba2+ ([Ba2+]i) concentrations with Fura 2 were used to identify and characterize the properties of a depolarization-activated Ca2+ and Ba2+ entry in the plasma membrane of osteoblast-like cells. The presence of this pathway was demonstrated in two osteoblastic cell lines, UMR-106 and MC3T3-E1 and osteoblasts isolated from rat long bone and rat neonatal calvariae. Subsequent characterization of the pathway was performed in the osteosarcoma cell line UMR-106. Depolarization of the cells with high medium K+ was followed by an increase in [Ca2+]i which was dependent on medium Ca2+. Ba2+ ions depolarized the cells and were transported by this pathway. Mg2+ ions interfered with Ca2+ and Ba2+ entry. At 140 mM KCl and 1 mM MgCl2, the pathway could be saturated with Ca2+ or Ba2+. The apparent affinity for Ca2+ was 0.78 mM and for Ba2+ 1.82 mM. Ca2+ or Ba2+ entry into the cells was blocked by low concentrations of nicardipine, diltiazem, verapamil, and La3+. In the absence of an increase in [Ca2+]i or [Ba2+]i, the pathway inactivated within about 5 min after depolarization. When [Ca2+]i or [Ba2+]i was allowed to increase, the pathway inactivated within about 20 s. These properties suggest that Ca2+ and Ba2+ entry are mediated by an L-type, depolarization-activated Ca2+ channel in osteoblasts. The activity of these channels changes little with an increase or decrease in cell volume. Thus, it is concluded that these pathways do not provide the Ca2+ entry pathway required for initiation of volume decrease by osteoblasts.

Animals↗
PubMed1989

Modulation of osteoblast function by prostaglandins.

The naturally occurring prostaglandins (PGs) were studied with respect to their abilities to change free cytosolic Ca2+ concentrations ([Ca2+]i), adenosine 3',5'-cyclic monophosphate (cAMP) levels, and cell proliferation in the osteoblastic cell line, UMR-106-01, and primary cultures of osteoblasts prepared from neonatal rat calvariae. All PGs tested stimulated an increase in [Ca2+]i, which was mainly due to Ca2+ release from intracellular stores. Measurements of the 50% effective concentration for the different PGs show that the potency ranking for PG-evoked [Ca2+]i increase in these cells is F2 alpha greater than D2 much greater than E2 greater than TxB2 greater than E1 greater than I2 much greater than A2. The PGs also increase cAMP levels in osteoblasts. At the highest concentrations tested (10-25 microM), dose-response saturation of cAMP production was observed only by PGE2 and PGE1. The potency rank for PG-stimulated cAMP increase was E2 greater than E1 much greater than A2 greater than I2 greater than F2 alpha greater than D2 greater than TxB2. Measurements of the effect of the PGs on thymidine uptake showed that low concentrations of PGF2 alpha and PGD2 had either no effect or stimulated proliferation of osteoblast-like cells. Relatively low concentration of PGE2, PGE1, and PGA2 inhibited proliferation. The potency ranking for PG-mediated inhibition of cell proliferation was identical to that found for PG-stimulated cAMP production. We conclude that all the naturally occurring PGs tested can activate the two signal transduction systems in osteoblasts.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium↗
PubMed1988

Relationship of cAMP and calcium messenger systems in prostaglandin-stimulated UMR-106 cells.

The effect of prostaglandins (PG) on free cytosolic calcium concentrations [( Ca2+]i) and cAMP levels was studied in the osteosarcoma cell line UMR-106. PGF2 alpha and PGE2, but not 6-keto-PGF1 alpha, induced an increase in [Ca2+]i which was mainly due to Ca2+ release from intracellular stores. The EC50 for PGF2 alpha was approximately 7 nM, whereas that for PGE2 was approximately 1.8 microM. Maximal doses of PGF2 alpha increased [Ca2+]i to higher levels than PGE2. Both active PGs also stimulated phosphatidylinositol turnover in UMR-106 cells. The effects of the two PGs were independent of each other and appear to involve separate receptors for each PG. PGE2 was a very potent stimulator of cAMP production and increased cAMP by approximately 80-fold with an EC50 of 0.073 microM. PGF2 alpha was a very poor stimulator of cAMP production; 25 microM PGF2 alpha increased cAMP by 5-fold. The increase in cellular cAMP levels activated a plasma membrane Ca2+ channel which resulted in a secondary, slow increase in [Ca2+]i. High concentrations of both PGs (10-50 microM) inhibited this channel independent of their effect on cAMP levels. Pretreatment of the cells with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate inhibited the PG-mediated increase in phosphatidylinositol turnover and the increase in [Ca2+]i. However, pretreatment with 12-O-tetradecanoyl-13-acetate had no effect on the PGE2-mediated increase in cAMP. The latter finding, together with the dose responses for PGE2-mediated increases in [Ca2+]i and cAMP levels, suggests the presence of two subclasses of PGE2 receptors: one coupled to adenylate cyclase and the other to phospholipase C. With respect to osteoblast function, the cAMP signaling system is antiproliferative, whereas the Ca2+ messenger system, although having no proliferative effect by itself, tempers cAMP's antiproliferative effect.

6-Ketoprostaglandin F1 alpha↗
PubMed1988

Selective modification of the kinetic properties of Na+/H+ exchanger by cell shrinkage and swelling.

The effect of volume perturbation on the interaction of Na+ and H+ with the intracellular and extracellular faces of the Na+/H+ exchanger was studied in UMR-106 cells, a rat osteosarcoma cell line. Osmotic shrinkage of the cells stimulated the activity of the Na+/H+ exchanger. Kinetic analysis of this stimulation demonstrated that in hyperosmotically stressed cells, the apparent affinities for intracellular H+ and intracellular Na+ are modified in opposite directions. While there is an increased apparent affinity for protons from 0.275 +/- 0.03 to 0.107 +/- 0.025 microM in isotonic and hypertonic conditions, respectively, the apparent affinity for intracellular Na+ decreases from 83 +/- 9 to 126 +/- 6 mM under the same conditions. Osmotic swelling induced a decreased exchanger activity which appeared to involve reduction in Vmax only without changes in the apparent affinities of either H+i or Na+i. We conclude that: 1) osmotic shrinkage and swelling modify the kinetic behavior of the Na+/H+ exchanger in different modes; 2) in hyperosmotically stressed cells, the interactions of intracellular H+ and Na+ are modified in a selective mode. The described phenomenon may serve as a general mechanism for activation of the exchanger by various stimuli.

Animals↗
PubMed1988

Cytosolic pH regulation in osteoblasts. Interaction of Na+ and H+ with the extracellular and intracellular faces of the Na+/H+ exchanger.

The interaction of Na and H ions with the extracellular and intracellular sites of the Na+/H+ exchanger of the osteosarcoma cell line UMR-106 was investigated. Na ions interact with a single, saturable extracellular transport site. H+ and amiloride appear to compete with Na+ for binding to this site. The apparent affinity for extracellular Na+ (Nao+) and amiloride was independent of intracellular H+ (Hi+), Nai+, or an outwardly directed H+ gradient. The interaction of H+ with the intracellular face of the exchanger had a sigmoidal characteristic with a Hill coefficient of approximately 2. The apparent affinity for Hi+ was independent of Nao+ between 25 and 140 mM. The apparent affinity for Hi+, but not the number of intracellular sites, increased with the increase in the outwardly directed H+ gradient across the membrane. Nai+/Ho+ exchange (reverse mode) is an electroneutral process with a Na+/H+ stoichiometry of 1. The dependence of Nai+/Ho+ exchange on Nai+ was sigmoidal, with a Hill coefficient of 2.16. Nai+ competes with Hi+ for binding to at least the transport site. The apparent affinity for Nai+ decreased with the increase in the outwardly directed H+ gradient. High Ho+ inhibited exchange activity in the reverse mode. We conclude that intracellular Na+ and H+ can activate the exchanger. The exchanger has two separate and asymmetric extracellular and intracellular transport sites. The relative apparent affinities of the internal transport site for Na+ and H+ are determined by the direction and magnitude of the H+ gradient across the membrane. Kinetic characterization of the exchanger suggests that Na+/H+ exchange is compatible with a simultaneous transport model, although a ping-pong transport model could not be excluded.

Amiloride↗
PubMed1987

Protein kinase C-activated calcium channel in the osteoblast-like clonal osteosarcoma cell line UMR-106.

The effects of protein kinase C stimulation on free cytosolic Ca2+ [( Ca2+]i) were studied in Fura 2-loaded UMR-106 cells. Stimulation of the protein kinase C with the tumor-promoting phorbol esters 12-O-tetradecanoylphorbol 13-acetate (TPA) and phorbol 12,13-diacetate or 1-oleoyl-2-acetylglycerol was followed by an increase in [Ca2+]i. The protein kinase C-induced increase in [Ca2+]i has a lag period, the duration of which was dependent on the stimulant and medium Ca2+ concentrations. With 2 microM TPA, the rise in [Ca2+]i peaked within 1.5 min, after which [Ca2+]i returned partially toward base line. The increase in [Ca2+]i was absolutely dependent on the presence of medium Ca2+ and was inhibited by the Ca2+ channel blockers nicardipine and verapamil. Cell stimulation also results in Ca2+ release from intracellular pool(s) which appears to be mediated by a Ca2+-dependent Ca2+ release mechanism. The reduction in [Ca2+]i was due to channel inactivation. Pretreatment of the cells with 1 nM TPA, 2 units/ml parathyroid hormone (PTH), or 15 microM forskolin blocked the effect of 2 microM TPA on [Ca2+]i. TPA and PTH were more potent inhibitors than was forskolin. The properties of this channel are compared to the cAMP-independent PTH-stimulated Ca2+ channel present in these cells.

Calcium↗
PubMed1987

Parathyroid hormone-activated calcium channels in an osteoblast-like clonal osteosarcoma cell line. cAMP-dependent and cAMP-independent calcium channels.

Changes in free cytosolic calcium were measured in UMR-106 cells in response to parathyroid hormone (PTH) stimulation. Bovine PTH-(1-34) induced an increase in [Ca2+]i with the contour of the rise in [Ca2+]i occurring in three successive phases: a rapid increase in [Ca2+]i occurring within seconds, rapid decrement in [Ca2+]i to near-resting levels within 1 min, and slow increment in [Ca2+]i. Phase one and phase three increases in [Ca2+]i were dependent on medium calcium. The phase one rise in [Ca2+]i was inhibitable by the calcium channel blockers lanthanum and verapamil. Only the phase one rise in [Ca2+]i was blocked by preincubation of the cells with the phorbol ester, phorbol 12-myristate 13-acetate. This channel was also blocked when cellular cAMP levels were increased prior to PTH stimulation. The phase two decrement of [Ca2+]i was due to the rapid inactivation of the phase one calcium channel. The phase three rise in [Ca2+]i was mediated by cellular cAMP levels. This cAMP-dependent Ca2+ channel was insensitive to pretreatment of the cells with phorbol diesters and showed low sensitivity to Ca2+ channel blockers. It is concluded that UMR-106 cells respond to PTH stimulation by the activation of a cAMP-independent Ca2+ channel. This channel rapidly inactivates. The subsequent PTH-dependent increase in cellular cAMP is followed by activation of a cAMP-dependent Ca2+ channel resulting in a slow rise in [Ca2+]i.

Animals↗
PubMed1986

Amiloride inhibition of DNA synthesis and immunoglobulin production by activated human peripheral blood mononuclear cells is independent of sodium/hydrogen antiport.

Activation of sodium/proton (Na+/H+) antiport activity has been shown to occur as an early event in mitogenesis. Because amiloride inhibits Na+/H+ antiport activity, it is hypothesized that mitogenesis may be inhibited by amiloride. In this work, we examined the effect of amiloride on DNA synthesis as measured by [3H]thymidine uptake and immunoglobulin (Ig) production as measured by an ELISA system in human peripheral blood mononuclear cells (PBM). Amiloride at 100 microM concentration inhibited irradiated Raji cell (*R)-activated and phytohemagglutinin-P (PHA-P)-stimulated DNA synthesis by 50 +/- 11% and 72 +/- 12%, respectively. IgG production was inhibited by 71% at 100 microM amiloride concentration in *R-activated PBM. This concentration of amiloride inhibited Na+/H+ antiport activity by 92%. Because amiloride is known to inhibit other pre-replicative cellular functions such as protein synthesis, we used an amiloride analogue, dimethylamiloride, which inhibited Na+/H+ antiport activity by 90% at a concentration of 1 microM without inhibition of PBM Ig or DNA synthesis. Furthermore, neither PHA-P nor *R-stimulated PBM demonstrated an intracellular alkalinization even after 6 hr of stimulation. Similarly, T cell-enriched or B cell-enriched populations did not show intracellular alkalinization after PHA-P or *R activation. Thus, it appears that Na+/H+ antiport activation is not an early event in PBM mitogenesis. The inhibition of mitogenesis by amiloride may be due to abrogation of premitotic events such as protein synthesis.

Adult↗
PubMed1979

Ultrasonic evaluation of the fetal heart. A report of experience and anatomic correlation.

Identification of fetal intracardiac structures in utero is accomplished with B-scan, gray-scale ultrasonography. Sagittal and transverse ultrasonographs of the interventricular septum, aortic root, tricuspid, and mitral valves are presented. These structures are characterized with respect to their relations to each other, atrial and ventricular chambers, and acoustic appearances. Correlation of these cardiac structures is made with gross anatomic fetal specimens sectioned in longitudinal and transverse planes. Recognition of fetal intracardiac detail may be of aid in future noninvasive investigations of human fetal cardiovascular dynamics.

Female↗