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J G Iversen

Publications and source records attributed to J G Iversen.

At least 37 records · Page 2Linked to original sources
PubMed1996

Growth-promoting effects of Ca(2+)-mobilizing agents in hepatocytes: lack of correlation between the acute activation of phosphoinositide-specific phospholipase C and the stimulation of DNA synthesis by angiotensin II, vasopressin, norepinephrine, and prostaglandin F2 alpha.

Although several hormones that promote hepatocyte proliferation also activate phosphoinositide-specific phospholipase C (PI-PLC) and mobilize Ca2+, the role of PI-PLC in the growth-stimulating effect of these agents is not clear. We have investigated this issue further, by exposing freshly isolated adult rat hepatocytes to vasopressin, angiotensin II, norepinephrine (in the presence of the beta-adrenoceptor blocker timolol) or PGF2 alpha, and examined both acute responses and the subsequent DNA synthesis when the cells were grown in monolayer culture. All the agonists elevated the level of inositol 1,4,5-trisphosphate (InsP3) and enhanced the DNA synthesis, amplifying the response to epidermal growth factor (EGF), and this comitogenic effect could be exerted by a single exposure of the cells 24 h prior to the addition of EGF. The acute activation of PI-PLC, measured as the early rise (peak 15-60 s) in InsP3, was 8-10-fold with vasopressin or angiotensin II, 3-4-fold with norepinephrine, and approximately 2-fold with PGF2 alpha. For all the agonists, a rise in cytosolic free Ca2+ in 100% of the cells and a maximal increase in glycogen phosphorylase activity were evoked at concentrations that approximately doubled the level of InsP3. However, the growth-stimulatory effects of these agonists showed a different order of efficacy as compared to the activation of PI-PLC; in terms of the maximal stimulation of DNA synthesis, the effects were: norepinephrine approximately PGF2 alpha > angiotensin II > vasopressin. Also, norepinephrine, PGF2 alpha, and angiotensin II, but not vasopressin, further enhanced the DNA synthesis when their concentrations were increased above those yielding maximal elevation of InsP3. In experiments where vasopressin and angiotensin II were combined, their effects on the DNA synthesis were additive while the InsP3 responses were not. The results show that the extent of the initial activation of PI-PLC is not the determinant for the magnitude of the growth effects of Ca(2+)-mobilizing hormones in hepatocytes. This suggests either (a) that the proliferative response to these agents is determined by the activity of PI-PLC at a later time, or its integral over an extended part of the prereplicative period, rather than by the acute activation, or (b) that additional, PI-PLC-independent, mechanisms are required.

Angiotensin II↗
PubMed1995

Binding of human factor VIIa to tissue factor induces cytosolic Ca2+ signals in J82 cells, transfected COS-1 cells, Madin-Darby canine kidney cells and in human endothelial cells induced to synthesize tissue factor.

Tissue factor (TF) is the most potent trigger of blood clotting known. It activates factor VII (FVII) thereby initiating a cascade of proteolytic reactions resulting in thrombin production. The cloning of TF revealed its structural characteristics to be those of a receptor related to the class 2 cytokine receptor superfamily, but until now no intracellular signal has been discovered related to binding of the ligand (FVIIa) to the putative receptor. We have studied possible intracellular signaling effects of the FVIIa-TF interaction by measuring cytosolic free Ca2+ in single fura-2-loaded cells and found that 200 nM FVIIa caused Ca2+ transients in about 30% of human umbilical vein endothelial cells treated with interleukin-1 beta to express TF, compared to below 5% in uninduced cells. A gradual increase of the basal Ca2+ level was also caused by binding of FVIIa. In the human bladder carcinoma cell line J82, which has a high constitutive TF activity, similar results were found. An antibody neutralizing TF activity decreased the response rate to control levels. COS-1 cells which do not make TF did not respond to FVIIa as opposed to COS-1 cells expressing TF after transfection with a human TF cDNA construct. The canine kidney cell line MDCK, a constitutive TF producer, responded especially well; up to 100% of the cells examined showed Ca2+ oscillations which were dose dependent with regard to frequency, latency, maximal amplitude, and recruitment of responding cells. The frequency was reduced by inhibition of Ca2+ influx with 100 microM LaCl3. In confluent MDCK cells the Ca2+ oscillations were synchronous, constituting the first evidence of a synchronous cytosolic Ca2+ oscillator generated by global application of agonist. Thus, TF mediates a cytosolic Ca2+ signal upon interaction with its ligand FVIIa, thereby suggesting a more complex biological role for TF.

Animals↗
PubMed1995

On the mechanisms of the growth-promoting effect of prostaglandins in hepatocytes: the relationship between stimulation of DNA synthesis and signaling mediated by adenylyl cyclase and phosphoinositide-specific phospholipase C.

While many observations indicate that prostaglandins may act as positive regulators of hepatocyte proliferation, the underlying mechanisms are not known. We have examined some of the signal pathways in the growth response induced by prostaglandins in hepatocytes, with particular focus on adenylyl cyclase and phosphoinositide-specific phospholipase C. Adult rat hepatocytes were cultured as primary monolayers in serum-free medium in the presence of EGF and insulin. PGE2 or PGF2 alpha (added 0-3 h after plating) enhanced the incorporation of [3H]-thymidine into DNA (measured at 50 h); at 100 microM the stimulation was about threefold PGI2 and PGD2 also showed significant but smaller stimulatory effects. No significant increase in the level of cyclic AMP (cAMP) was detected in response to any of the prostaglandins. Low concentrations of glucagon (0.1-10 nM), a potent activator of hepatic adenylyl cyclase, or 8-bromo-cAMP (0.1-10 microM) enhanced the DNA synthesis. When 8-bromo-cAMP was used in maximally effective concentrations, no further stimulation was obtained by combining it with glucagon, whereas the effects of PGE2 and 8-bromo-cAMP were completely additive. All the prostaglandins also showed additivity with the effect of glucagon on the DNA synthesis. PGE2, PGF2 alpha, PGI2, and PGD2 increased intracellular inositol-1,4,5-trisphosphate (InsP3), with a relative order of efficacy roughly corresponding to their activity as stimulators of DNA synthesis. Increases in cytosolic free Ca2+, as measured in single cells, were elicited in a majority of the hepatocytes by all these prostaglandins at 1 microM. Supramaximal concentrations of vasopressin, a strong activator of phospholipase C in hepatocytes, acted additively with PGE2 on the DNA synthesis. Pretreatment of the hepatocytes with a concentration of pertussis toxin that prevented the inhibitory effect of PGE2 on glucagon-induced cAMP accumulation did not abolish the ability of PGE2 to stimulate the DNA synthesis. The results do not support a role for adenylyl cyclase activation in the stimulatory effect of prostaglandins on hepatocyte growth. While the data are compatible with an involvement of phosphoinositide-specific phospholipase C in the growth-promoting effect of prostaglandins in cultured rat hepatocytes, they suggest this may not be the sole mechanism.

Adenylate Cyclase Toxin↗
PubMed1995

Interferon-gamma affects protein kinase C activity in human neutrophils.

Interferon-gamma (IFN-gamma) is a priming agent of polymorphonuclear neutrophilic granulocyte (PMN) oxygen metabolism, and protein kinase C (PKC) is traditionally believed to play a central role in activation of this oxygen metabolism. In the present study, we have shown that the PKC activity in PMN is affected by IFN-gamma. After only 2 minutes exposure to IFN-gamma (100 U/ml), PKC activity was significantly increased in the noncytosolic fraction of the cells. This increase was transient, but toward the end of the priming period of 2 h, the membrane-associated PKC activity increased again to about 152% of control. In the cytosolic fraction, a small and hardly detectable decrease in PKC activity was observed. Treatment of PMN with granulocyte-macrophage colony-stimulating factor (GM-CSF), another PMN priming agent, showed no significant effects on the PKC activity. When the cells were stimulated with the bacterial peptide fMLP after a priming period with IFN-gamma or GM-CSF for 2 h, no significant difference between treated and control cells could be observed. PMN oxygen metabolism, measured by flow cytometry as an accumulation of the fluorescent compound dichlorofluorescein, was in these experiments significantly primed by IFN-gamma, both at baseline and when stimulated with fMLP. The protein kinase C inhibitors H7 and Ro31-8220 blocked the fMLP responses to some extent, but not completely. However, no significant difference between fMLP responses in control and IFN-gamma-treated cells could be detected after administration of inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Interferon-gamma modulates cytosolic free calcium in human neutrophilic granulocytes.

To investigate the role of cytosolic free calcium ([Ca2+]i in interferon-gamma (IFN-gamma) pre-activation (priming) of human neutrophilic granulocytes (PMN) we used three different fluorescence methods, i.e. digital imaging of single, adherent, Fura-2 loaded cells, flow cytometric measurements of single, non-adherent, Fluo-3 loaded cells, and spectrofluorometry of Indo-1 loaded PMN in suspension. IFN-gamma increased the [Ca2+]i level in single, adherent PMN during the second phase of the fMLP response. The bacterial peptide fMLP (N-formyl-L-methionyl-L-leucyl-L-phenylalanine) is a known stimulant of the calcium/inositol phosphate system. The [Ca2+]i increase was abolished in Ca(2+)-free test buffer. Furthermore, the baseline [Ca2+]i level was found to be slightly increased in IFN-gamma primed PMN as analysed with flow cytometry. On the other hand, these [Ca2+]i responses were not detectable with the other methods used. We suggest that IFN-gamma increases the plasma membrane permeability for calcium in PMN, and substantiate this by demonstrating compliance with a capacitative model for intracellular calcium regulation. Mathematical modeling also suggested that IFN-gamma primed human PMN may sequester 13% more Ca2+ than unprimed cells in fMLP-insensitive intracellular stores. Thus, the Ca2+ responses to IFN-gamma are modest and not easily detectable with some of the methods currently in use. They nevertheless explain why fMLP elicits brisker responses from PMN after IFN-gamma priming.

Calcium↗
PubMed1992

Thapsigargin reveals evidence for fMLP-insensitive calcium pools in human leukocytes.

To investigate the relationship between different intracellular Ca2+ pools, cytosolic free calcium ([Ca2+]i) was surveyed by means of a Fura-2 fluorescence ratio method on single isolated human leukocytes. Both monocytes and neutrophilic granulocytes (PMN) displayed long lasting spontaneous [Ca2+]i transient changes (1-2 min). In PMN stimulated with the bacterial peptide fMLP we observed transients with shorter duration (10-30 s) and smaller amplitude often superimposed on the long lasting transients. The time course of changes in [Ca2+]i was recorded in a large number (149) of single leukocytes prestimulated for 5 min with fMLP and then challenged with thapsigargin (a blocker of Ca2+ uptake in intracellular pools). Statistical analysis of [Ca2+]i responses revealed that fMLP-sensitive pools contributed to the long lasting [Ca2+]i transients seen in both leukocyte types. However, the existence of fMLP-insensitive calcium pools may explain the superimposed transients seen in PMN. Thapsigargin was also added together with EGTA (to impede contribution from extracellular Ca2+) to 198 fMLP prestimulated and 153 unstimulated PMN. Based on Ca2+ registrations in these cells and a mathematical model (supposing two separate first order responses) the amount of Ca2+ stored in the various pools and their release kinetics were estimated. The results indicate that fMLP-insensitive calcium pools exist in PMN but not in monocytes. Since the digital imaging technique also depicts cellular motility, an additional finding was that the leukocyte's ability to sequestrate the Ca2+ from the cytosol seemed important to locomotion.

Calcium↗
PubMed1990

Effect of Diamphidia toxin, a Bushman arrow poison, on ionic permeability in nucleated cells.

The effect of Diamphidia toxin, isolated from pupae of Diamphidia nigro-ornata, was tested on two different cell lines (GH4C1 cells and HL-60 cells) and on human lymphocytes. The toxin raised intracellular Ca2+ concentration, as assessed with quin 2, in a dose-related manner in all three cell types. The effect was abolished when extracellular Ca2+ was chelated by EGTA. Low concentrations of the toxin evoked a delayed as well as a smaller response. The response time was also temperature-dependent, with a Q10 of about 2. Low, but effective concentrations of the toxin did not affect cell membrane integrity, as tested with Trypan blue, and induced a seemingly physiological release of prolactin from the GH4C1 cells. Diamphidia toxin's effect on the membrane permeability of GH4C1 cells was further investigated with patch-clamp techniques. The toxin appeared to increase the conductance for all small ions without affecting the normal ionic channels present in these cells. We conclude that Diamphidia toxin has a general effect on the plasma membrane of different cell types and thereby increases, probably non-specifically, the permeability for small ions.

Ammonium Sulfate↗
PubMed1990

A transient acidification linked to intracellular Ca2+ in anti-mu-stimulated human B-lymphocytes.

Mitogen-induced cellular proliferation is in many cell types preceded by rapid changes in intracellular pH and free Ca2+ concentration. We studied the patterns of pH and Ca2+ changes in normal resting human B-lymphocytes after exposure to anti-mu antibodies and the monoclonal antibody IF5, reactive with the CD20 antigen, both able to activate resting B-lymphocytes to enter the G1 phase of the cell cycle. Monitoring intracellular pH with the pH-sensitive, fluorescent dye, 2',7'-bis(carboxyethyl)-5,6-carboxy-fluorescein, we demonstrated that poly- and monoclonal anti-mu antibodies induced a rapid (maximum change within 2 min) intracellular acidification of 0.06 pH units followed by a slower (10-15 min) alkalinization towards, or slightly above, the resting pH of 6.88. The acidification response was amiloride-resistant, whereas the return to baseline was sensitive. Intracellular free Ca2+ was measured by using the fluorescent Ca2+ dye, indo-I. Exposure of cells to anti-mu resulted in a rapid increase (maximum change within 2 min) in cytoplasmic Ca2+ of 340 nM and a slower decline in fluorescence back to baseline of about 180 nM. In contrast to anti-mu, IF5 caused no change in cytoplasmic Ca2+ and pH. However, the Ca2+ ionophore ionomycin at low concentrations mimicked the Ca2+ response as well as the pH response to anti-mu. In Ca2(+)-free solutions the intracellular Ca2+ stores are usually rapidly depleted and, indeed, the Ca2+ and pH responses to anti-mu were reduced after 5 min and almost abolished after 35 min under such conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

Amiloride↗
PubMed1988

Somatostatin inhibits prolactin secretion by multiple mechanisms involving a site of action distal to increased cyclic adenosine 3',5'-monophosphate and elevated cytosolic Ca2+ in rat lactotrophs.

The release of prolactin (PRL) from a clonal cell-line of anterior pituitary cells (GH4C1) was inhibited by somatostatin (SRIH) in a dose-dependent manner (ED50 nM). The inhibition (20% of control levels) was detectable within 50 s and maximal within 90 s. Thyroliberin (TRH) enhancement of PRL secretion was biphasic. SRIH inhibited both phases equally. Ionomycin in combination with the phorbol ester, TPA, mimics the TRH-elicited PRL release, and SRIH partly inhibited this effect. SRIH had no effect on TRH-stimulated formation of inositol trisphosphate, and only small effects on TRH-activated adenylate cyclase. Vasoactive intestinal peptide (VIP) and forskolin stimulated cAMP formation and PRL release potently. SRIH inhibited both effects of VIP and forskolin, and there was a close correlation between the inhibition of PRL secretion and cAMP accumulation. 8-Bromo-cAMP enhanced PRL release, an effect that was also partly reduced by SRIH. The Ca2+ channel activator, BAY-K-8644 and high extracellular K+ increased PRL release, and SRIH caused a partial reduction in the release response to both secretagogues. SRIH lowered [Ca2+]i, and markedly reduced the rise in [Ca2+]i elicited by TRH, VIP and K+. SRIH did not influence the Ca2+ spikes recorded in Na+-free solution, and had no effect on the TRH-induced membrane potential changes. Our results demonstrate that SRIH may inhibit PRL release from GH4C1 cells by (1) inhibiting hormone-sensitive adenylate cyclase, (2) blocking the effect of cAMP and (3) lowering [Ca2+]i. None of these effects is, however, sufficient to explain all the effects of SRIH, suggesting that SRIH also exerts a major action at a step subsequent to cAMP accumulation and [Ca2+]i elevation. Since the GH4C1 cells possess one single class of binding sites, this implies that the same SRIH receptor is coupled to several cellular signalling systems.

Animals↗
PubMed1987

Bombesin stimulates prolactin secretion from cultured rat pituitary tumour cells (GH4C1) via activation of phospholipase C.

Bombesin (BBS) stimulated prolactin (PRL) secretion from monolayer cultures of rat pituitary tumour cells (GH4C1) in a dose-dependent manner with half maximal and maximal effect at 2 nM and 100 nM, respectively. No additional stimulatory effect on PRL secretion was seen when BBS was combined with thyroliberin (TRH) used in concentrations known to give maximal effects, while the effects of BBS and vasoactive intestinal peptide (VIP) were additive. Using a parafusion system, BBS (1 microM) was found to increase PRL secretion within 4 s and the secretion profiles elicited by BBS and TRH (1 microM) were similar. Both BBS and TRH increased inositoltrisphosphate (IP3) as well as inositolbisphosphate (IP2) formation within 2 s. BBS also induced the same biphasic changes in the electrical membrane properties of GH4C1 cells as TRH, and both peptides caused a rapid and sustained increase in intracellular [Ca2+]. These results suggest that BBS stimulates PRL secretion from the GH4C1 cells via a mechanism involving the immediate formation of IP3 thus resembling the action of TRH.

Animals↗
PubMed1987

Vasoactive intestinal peptide and peptide with N-terminal histidine and C-terminal isoleucine increase prolactin secretion in cultured rat pituitary cells (GH4C1) via a cAMP-dependent mechanism which involves transient elevation of intracellular Ca2+.

Vasoactive intestinal peptide (VIP) and peptide (P) with N-terminal histidine and C-terminal isoleucine (PHI) stimulated prolactin (PRL) secretion from GH4C1 cells equipotent with ED50 values of 30-50 nM. In a parafusion system optimized to give high time resolution both VIP and PHI increased PRL secretion with a delay of about 60 s and subsequent to the activation of the adenylate cyclase. Thyroliberin (TRH) increased PRL secretion within 4 s. The dose-response curves for VIP- and PHI-stimulated cAMP accumulation were superimposable on those for PRL secretion. At submaximal concentrations the effects of VIP and PHI on both cAMP accumulation and PRL secretion were additive, whereas the effects were not additive at concentrations giving maximal effects. VIP and PHI increased [Ca2+]i measured by quin-2 in a different way than TRH, without inducing changes in the electrophysiological membrane properties of the GH4C1 cells. We conclude that both VIP and PHI stimulate PRL secretion via a cAMP-dependent process involving an increase in [Ca2+]i.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1986

On the functional relationship between 45Ca2+ release and prolactin secretion in cultured rat pituitary tumour (GH3) cells.

We have evaluated the role of cellular Ca2+ transport associated with stimulus-secretion coupling in prolactin (PRL) producing rat pituitary adenoma cells (GH3 cells). The action of different substances, known to modify PRL secretion, on release of 45Ca2+ from preloaded cells were examined. Surface-bound 45Ca2+ was removed by pretreatment with trypsin in EDTA buffer. During the first 6 min, basal efflux of 45Ca2+ occurred at a constant rate (0.24 min-1) at 37 degrees C. Addition of TRH (5 X 10(-7) M) resulted in an immediate enhancement of 45Ca2+ release representing about 20% of the remaining cellular 45Ca2+. In the same experiments PRL secretion increased by 45%. The EDTA in the external medium reduced the basal rate of 45Ca2+ release by 60%, but did not apparently affect the TRH-stimulated release. Somatostatin (10(-6) M) and verapamil (5 X 10(-5) M) inhibited both basal and TRH-stimulated PRL secretion, whereas high extracellular concentration of K+ (5 X 10(-2) M) had a stimulatory effect. However, neither of these treatments changed cellular 45Ca2+ release. Interference with energy-dependent Ca2+ transport by using metabolic inhibitors (iodoacetate, 6 X 10(-3) M; and antimycin, 2 X 10(-6) M) or by replacing Na+ in the medium by choline or by lowering the incubation temperature from 37 to 25 degrees C, had no effect on TRH-stimulated 45Ca2+ release although basal and TRH-stimulated PRL secretion were reduced. Thus, TRH apparently releases 45Ca2+ from calcium binding sites in the cell membrane.

Adenoma↗
PubMed1986

Effects of bromocriptine on prolactin release, electrical membrane properties and transmembrane Ca2+ fluxes in cultured rat pituitary adenoma cells.

The effects of the dopamine (DA) agonist bromocriptine on prolactin (Prl) release, electrical membrane properties and transmembrane Ca2+ fluxes have been studied in a clonal strain of rat pituitary adenoma cells (GH3). These cells generate Ca2+ dependent action potentials, and produce and secrete spontaneously both Prl and growth hormone. Prl release stimulated by thyroliberin (TRH) and elevated extracellular K+ concentration was completely blocked by bromocriptine, whereas the basal release was only moderately affected. The TRH and K+ evoked Prl release were half maximally inhibited by bromocriptine at 5-10 and 10-50 microM, respectively. The normal biphasic membrane response to TRH and the depolarizing effect of elevated K+ concentration were not altered by bromocriptine, whereas the Ca2+- spikes in Na+-free solution were suppressed by the drug. We therefore suggest that bromocriptine blocks the voltage sensitive Ca2+-channels of GH3 cell. In agreement with this notion, bromocriptine also suppressed the basal and TRH induced 45Ca2+ efflux from preloaded cells. We conclude that the inhibitory effect of bromocriptine on the voltage dependent Ca2+- channels is an important mechanism responsible for suppression of Prl release.

Action Potentials↗
PubMed1985

No correlation between membrane potential and increased cytosolic free Ca2+ concentration, 86Rb+ influx or subsequent [3H]-thymidine incorporation in neoplastic human B cells stimulated with antibodies to surface immunoglobulin.

We have followed the changes in the membrane potential of neoplastic B cells after addition of antibodies to cell specific surface immunoglobulin heavy chains (anti-Ig). Two methods were used, both based on the distribution of lipophilic fluorescent indicators: the anionic bis(1,3-diethylthiobarbiturate)-trimethine oxonol (fluorescence measurement made on whole cell suspensions) and the cationic 3,3-dihexyloxacarbocyanine iodide (diOC6-(3] (using flow cytofluorimetric analysis). By the latter method, the resting membrane potential was calculated to range from -40 to -63 mV. The stimulatory effect of anti-Ig was investigated on two B cell populations. In one, which may be stimulated in this way to [3H]-thymidine incorporation, the membrane potential was apparently unchanged, at least during the first hour of stimulation. In the other population, anti-Ig induced a rapid depolarization (oxonol method), but these cells did not, on the other hand, respond with [3H]-thymidine incorporation. The nature of this depolarization was further investigated. Both Na+ and K+ permeabilities appeared increased, while Ca2+ influx did not contribute to the change in membrane potential. Nor did depolarization itself change free cytosolic Ca+ concentration, [Ca2+]i, as estimated by the quin-2 method. Furthermore, anti-Ig stimulation increased [Ca2+]i in these cells, even when depolarization was abolished by lowering the extracellular Na+ concentration. Thus, in cells where depolarization could be demonstrated, it was linked neither to early changes in [Ca2+]i nor to late [3H]-thymidine incorporation. In contrast, such effects of anti-Ig stimulation were observed in other cells without any significant early change in the membrane potential.

Antibodies↗
PubMed1985

Intracellular Ca2+ buffering potentiates an 86Rb+ permeability response in human lymphocytes.

The effects of antibodies against immunoglobulin delta-heavy chains (anti-delta) on intracellular free Ca2+ concentrations, [Ca2+]i, and 86Rb+ influx in human neoplastic B-cells were tested in vitro. When preloading the cells with high concentrations of the fluorescent Ca2+ chelator quin 2 and subsequently stimulating in EGTA medium, the anti-delta induced rise in [Ca2+]i was strongly reduced or blocked. Nevertheless, 86Rb+ influx, also induced by anti-delta, was potentiated. In fact, in a population of cells in which anti-delta increased [Ca2+]i, but not 86Rb+ influx under standard conditions, the combination of quin-2 preloading and subsequent extracellular Ca2+ chelation by EGTA revealed an anti-delta induced 86Rb+ influx. Most of this influx was ouabain resistant, suggesting only a minor contribution from the Na+/K+ pump. Based on the Ca2+ buffer effect of quin 2 we suggest that the Ca2+ effect on 86Rb+ (K+ analogue) permeability is not mediated by increased [Ca2+]i but rather by the Ca2+ release per se from the plasma membrane.

Antibodies↗
PubMed1983

Early, anti-immunoglobulin induced events prior to Na+-K+ pump activation: an analysis in a monoclonal human B-lymphoma cell population.

Events following F(ab)2 anti-delta immunoglobulin stimulation of monoclonal (leukemic) human B cells prior to Na+-K+ pump activation were investigated in vitro. This pump activation, measured by ouabain-sensitive 86Rb+ uptake, appeared susceptible to the phospholipid-interacting drugs tetracaine and quinacrine, to the antioxydant nordihydroguaiaretic acid (NDGA), and to the calmodulin antagonist trifluoperazine, while much less susceptible to the methylation inhibitor-3-deazaadenosine. The Ca++ ionophore A 23187 appeared to induce pump activation in a way similar to anti-delta, as it was susceptible to the same drugs and as anti-delta had no additional stimulating effect on A 23187-stimulated cells. However, whereas the anti-delta-induced activations appeared independent of the extracellular Ca++ activity, [Ca++]e, the activation by A 23187 was potentiated by addition of the Ca++ chelator ethyleneglycol-bis (beta-aminoethyl ether) N, N'-tetracetic acid (EGTA). Estimations by fluorescent chelator method (quin 2) showed anti-delta to increase the intracellular Ca++ activity, [Ca++]i both in the absence and presence of EGTA. A 23187 increased [Ca++]i strongly in Ca++ medium, but was weaker, more similar to the anti-delta response, in EGTA medium. It is suggested that Na+-K+ pump activation after anti-Ig stimulation in B cells may follow Ca++ mobilization from internal stores. The trifluoperazine susceptibility suggests that calmodulin regulation is involved.

Antibody Formation↗