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C D Logsdon

Publications and source records attributed to C D Logsdon.

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

Stress-activated protein kinase activation is the earliest direct correlate to the induction of secretagogue-induced pancreatitis in rats.

We compared the cellular events induced by hyperstimulation of rats with caerulein which induces acute pancreatitis, to bombesin, which does not induce pancreatitis. Both secretogogues induced the intracellular activation of trypsinogen and the colocalization of lysosomal hydrolases and zymogen granules within 10-15 minutes. These data indicate that these parameters, previously thought to be crucial initiating events of pancreatitis, are not definitive cellular markers of the disease. We then compared the abilities of the two secretagogues to activate stress-activated protein kinase (SAPK). Significant effects of caerulein hyperstimulation on SAPK activity were observed within 5 minutes, the maximum (57-fold) activation was evident after 15 minutes, and levels remained above control for at least 3 hours. In comparison, hyperstimulation with bombesin induced a maximal 5-fold increase of SAPK activity which returned to basal within one hour. These data indicate that SAPK activity is the earliest and best correlated cellular marker associated with secretagogue-induced pancreatitis.

Animals↗
PubMed1996

Distinct cytoplasmic domains of the growth hormone receptor are required for glucocorticoid- and phorbol ester-induced decreases in growth hormone (GH) binding. These domains are different from that reported for GH-induced receptor internalization.

Glucocorticoids inhibit growth in children and antagonize the growth-promoting action of GH in peripheral tissues. Recently, they have been shown to decrease GH binding. In this study we examine the molecular mechanisms by which the glucocorticoid dexamethasone (DEX) and the phorbol ester phorbol myristate acetate (PMA) decrease cellular GH binding. In 3T3-F442A fibroblasts, DEX and PMA decrease the number of GH receptors (GHRs) capable of binding GH by 50% (t1/2 = 6 h) and 70% (t1/2 = 15 min), respectively. Neither appear to decrease the total number of cellular GHR. Rather, they appear to redistribute GHRs away from the plasma membrane or inactivate GHRs on the membrane such that they cannot bind GH. DEX and PMA also decrease GH-induced tyrosyl phosphorylation of GHR and JAK2 with a magnitude and time course correlating with that of inhibition of GH binding. DEX- and PMA-induced reductions of GH binding are also observed in a Chinese hamster ovary (CHO) cell line stably transfected with a rat liver GHR cDNA, further arguing that DEX and PMA act post-translationally on GHR. Using mutant GHRs stably expressed in CHO cells, amino acids 455-506 and tyrosines 333 and/or 338 of GHR were shown to be required for maximal DEX-induced inhibition of GH binding. DEX decreased GH binding to a GHR mutant F346A, which is reported to be deficient in ligand-induced internalization, suggesting that DEX decreases GH binding by a mechanism distinct from that of ligand-induced GHR internalization. PMA reduced GH binding to CHO cells expressing all GHR mutants tested. However, deletion of the C-terminal 132 amino acids decreased this effect, suggesting that at least one component of PMA action on GHR requires amino acids 507-638. These data suggest that distinct pathways mediate the effects of GH, DEX, and PMA on GHR number in the plasma membrane.

3T3 Cells↗
PubMed1996

Jun kinases are rapidly activated by cholecystokinin in rat pancreas both in vitro and in vivo.

Stimulation of pancreatic acini from male Sprague-Dawley rats by both cholecystokinin (CCK)-8 and anisomycin caused an increase in p46jnk and p55jnk activities. Both forms of c-Jun amino-terminal kinase (JNK) were slightly activated at 5 min, reached a maximum at 30 min, and remained significantly increased at 60 min of CCK stimulation. By contrast, p42mapkwas activated fully by 5 min. In pancreatic acini stimulated with different concentrations of CCK for 30 min, the minimal and maximal JNK responses were observed at 30 pm and 100 nM CCK, respectively; p42mapk activation was, as previously reported, much more sensitive, with maximal activation by 1 nm CCK. Carbachol and bombesin also stimulated JNK activity, while vasoactive intestinal peptide did not. Neither activating protein kinase C nor increasing intracellular Ca2+ significantly activated JNK. In in vivo experiments, rats were infused intravenously for 5 and 15 min with a secretory (0.1 microg/kg/h) or supramaximal (10 microg/kg/h) dose of the CCK analog caerulein (CER). Secretory doses of CER induced a 4-fold increase of both forms of JNK in pancreatic tissue at 5 and 15 min, while at the same time points, supramaximal stimulation with CER caused 4- and 27-fold increases, respectively, of these kinase activities. The secretory dose of CER slightly increased the activities of both forms of mitogen-activated protein kinase, while the supramaximal dose induced a 10-fold increase of p42mapk at 5 min. In conclusion, JNKs and mitogen-activated protein kinases are rapidly activated in rat pancreatic acini stimulated with CCK as well as in pancreatic tissue during in vivo stimulation with CER. The large response to supramaximal CER stimulation may be of importance in the early pathogenesis of acute pancreatitis.

Animals↗
PubMed1996

Nucleotides regulate the binding affinity of the recombinant type A cholecystokinin receptor in CHO K1 cells.

Cholecystokinin (CCK) receptors on rat pancreatic acinar cells display two binding affinity states in the presence of adeninine and guanine triphosphates with the effect of ATP mediated by the enzyme nucleoside diphosphate kinase. To determine whether this behavior was intrinsic to a single receptor protein we studied the binding affinity of CHO cells stably transfected with a cloned rat CCKA receptor. 125I-CCK binding to intact cells at 37 degrees C revealed two affinity states for CCK of Kd values 20 pM and 2.4 nM. Membranes prepared from these cells displayed a single affinity state for CCK but two affinity states could be restored in the presence of GTP[gamma S], ATP and ATP[gamma S] but not AMP-PCP. ATP and ATP[gamma S] but not AMP-PCP were substrates for nucleoside diphosphate kinase present in CHO cell membranes and transferred their terminal phosphate to GDP. These findings indicate that the interconvertible affinity states of the CCK receptor are inherent in a single receptor protein and that nucleoside diphosphate kinase mediates the effect of ATP to regulate these two affinity states.

Adenosine Triphosphate↗
PubMed1995

Muscarinic acetylcholine receptor down-regulation limits the extent of inhibition of cell cycle progression in Chinese hamster ovary cells.

Cellular desensitization is believed to be important for growth control but direct evidence is lacking. In the current study we compared effects of wild-type and down-regulation-resistant mutant m3 muscarinic receptors on Chinese hamster ovary (CHO-K1) cell desensitization, proliferation, and transformation. We found that down-regulation of m3 muscarinic acetylcholine receptors was the principal mechanism of desensitization of receptor-activated inositol phosphate phospholipid hydrolysis in these cells. Activation of wild-type and mutant receptors inhibited anchorage-independent growth as assayed by colony formation in agar. However, the potency for inhibition of anchorage-independent growth was greater for cells expressing the mutant receptor. Activation of either receptor also initially inhibited anchorage-dependent cell proliferation in randomly growing populations. Rates of DNA synthesis and cell division were profoundly reduced by carbachol in cells expressing either receptor at early time points. Analysis of cell cycle parameters indicated that cell cycle progression was inhibited at transitions from G1 to S and G2/M to G1 phases. However, mutant receptor effects on anchorage-dependent growth were sustained, whereas wild-type receptor effects were transient. Thus, receptor down-regulation restored cell cycle progression. In contrast, activation of either receptor blocked entry into the cell cycle from quiescence, and this response was not reduced by receptor down-regulation. Therefore, activation of m3 muscarinic acetylcholine receptors inhibited CHO cell anchorage-dependent and -independent growth. In anchored cells carbachol inhibited the cell cycle at three distinct points. Inhibitions at two of these points were eliminated by wild-type receptor down-regulation while the other was not. These results directly demonstrate that desensitization mechanisms can act as principal determinants of cellular growth responses.

Animals↗
PubMed1995

Carboxyl-terminal domains determine internalization and recycling characteristics of bombesin receptor chimeras.

To investigate the role of the carboxy terminus in the regulation of the bombesin (BN) receptor, we constructed two chimeric receptors with carboxyl termini transferred from either m3 muscarinic cholinergic (m3 ACh) (BMC) or cholecystokinin A (CCKA) (BCC) receptors and expressed them in Chinese hamster ovary cells. Previous studies showed that agonist treatment caused rapid internalization of CCKA but not m3 ACh receptors in these cells. In the current study we conducted separate analyses of ligand and receptor internalization and analyzed receptor recycling. Ligand internalization was assessed using acid washing. BN and CCKA receptors internalized ligand with 80 +/- 3 and 85 +/- 7% in an acid-resistant compartment at equilibrium. Ligand internalization of chimeric receptors generally assumed the properties of the donor receptors. Thus, BCC receptors internalized ligand to a similar extent as wild-type CCKA receptors (75 +/- 3%), whereas, BMC receptors showed reduced ligand internalization (38 +/- 1%). Receptor internalization was more directly assessed by determining agonist-induced loss of surface binding. BN and CCKA receptors were largely internalized (56 +/- 8 and 50 +/- 7%, respectively). BCC receptors were also extensively internalized (82 +/- 3%). In contrast, BMC receptors were minimally internalized (22 +/- 8%). Receptor recycling was assessed as recovery from agonist induced loss of binding. BN, CCKA, and BMC receptors showed rapid recycling. In contrast, BCC receptors did not recycle. These data indicate that carboxyl-terminal structures determine both internalization of ligand-receptor complexes and subsequent receptor recycling.

Amino Acid Sequence↗
PubMed1995

High- and low-affinity CCKA receptor states mediate specific growth inhibitory effects on CHO cells.

To relate specific effects on growth and transformation to activation of specific affinity states of the CCKA receptor stably expressed in CHO cells we compared responses to the CCK analogues JMV-180 and CCK8. CCK8 led to an inhibition of both cell proliferation and transformation. Effects on proliferation were indicated by a reduction of DNA synthesis and cell numbers. Effects on transformation were indicated by a reduction of colony formation in soft-agar. JMV-180 did not inhibit cell proliferation although a small inhibitory effect on DNA synthesis was observed. JMV-180 inhibited the maximal effects of CCK8 on cell proliferation and DNA synthesis. In contrast, JMV-180 substantially inhibited cell colony formation in soft-agar and did not inhibit the effects of CCK8 on this parameter. Collectively these data with receptor affinity state specific analogues indicated that inhibition of cell proliferation and growth in soft-agar can be attributed to activation of distinct affinity states. Thus, different second messengers are likely responsible for the inhibitory effects on anchorage-dependent and -independent growth.

Animals↗
PubMed1995

Influence of second and third cytoplasmic loops on binding, internalization, and coupling of chimeric bombesin/m3 muscarinic receptors.

In order to investigate the molecular basis for differences in the characteristics of bombesin (Bn) and m3 muscarinic cholinergic (m3 ACh) receptors, chimeric Bn receptors possessing cytoplasmic domains from the m3 ACh receptor were produced. The receptors were expressed in CHO-K1 cells and binding, structural, and signal transduction characteristics were analyzed. Cell lines bearing chimeric Bn receptors possessing m3 ACh receptor domains in place of either the second cytoplasmic loop (BM2L), the third cytoplasmic loop (BM3L), or both loops (BM23L) each bound 125I-bombesin with a single affinity that was approximately the same as that of the Bn receptor (5-10 nM). However, Bn receptors possessing the m3 ACh third cytoplasmic loop were severely affected in other respects. Internalization of ligand in Bn and BM2L cells was rapid and extensive (> 80% of bound 125I-bombesin was acid-resistant). In contrast, internalization was dramatically reduced in BM3L and BM23L cells (approximately 20% of bound 125I-bombesin was acid-resistant). In Bn or BM2L cells 10 nM bombesin stimulated approximately 10-fold increases in phosphatidylinositol hydrolysis. Activation of Bn receptors also induced an increase in arachidonic acid release (478 +/- 32% of control, n = 3) and large increases in intracellular Ca2+. In contrast, in BM3L or BM23L cells, bombesin had no significant effect on phosphatidylinositol hydrolysis. Furthermore, BM3L receptor activation did not increase arachidonic acid release. However, BM3L and BM23L cells showed a small increase in intracellular Ca2+ at high concentrations of bombesin. These data indicate that the third cytoplasmic loop alone, or together with the second cytoplasmic loop, was not sufficient to transfer the characteristics of G protein interaction between m3 ACh and bombesin receptors. Furthermore, for the Bn receptor, ligand internalization does, whereas formation of the high affinity binding state does not, appear to require activation of G proteins.

Animals↗
PubMed1995

Ca2+ signaling through secretagogue and growth factor receptors on pancreatic AR42J cells.

Intracellular signaling by an increase in [Ca2+]i was observed in pancreatic AR42J cells in response to agonists whose receptors are G-protein coupled including cholecystokinin (CCK), bombesin, carbachol, substance P, pituitary adenylate cyclase activating peptide (PACAP), bradykinin, ATP, calcitonin gene related peptide (CGRP), and in response to growth factors EGF and FGF whose receptors are tyrosine kinases. The response to growth factors was smaller both in magnitude and in the percentage of cells responding but was independent of extracellular Ca2+. CCK and carbachol induced sizeable increases in inositol phosphates while growth factors did not. The responses to both carbachol and EGF, however, were blocked by the phospholipase C inhibitor U73122. The tyrosine kinase inhibitor, genestein, blocked the response to EGF but not that to CCK. These data are consistent with two types of signaling mechanisms in AR42J cells. Secretagogues act on receptors which couple through G proteins to induce a large amount of inositol phosphate production and subsequent intracellular Ca2+ mobilization. Growth factors act on receptors which signal through tyrosine kinase activity and in this cell type produced limited amounts of inositol phosphate and a smaller increase in intracellular Ca2+.

Adenosine Triphosphate↗
PubMed1995

Mutation of carboxyl-terminal threonine residues in human m3 muscarinic acetylcholine receptor modulates the extent of sequestration and desensitization.

We previously reported that a mutant human m3 muscarinic acetylcholine receptor in which threonine residues at positions 550, 553, and 554 in the carboxyl terminus had been substituted with alanines showed a significant blockage of receptor down-regulation when expressed in Chinese hamster ovary-K1 cells. Because Chinese hamster ovary cells showed little receptor sequestration, in the present study we investigated further the effects of these mutations on sequestration and desensitization in human embryonic kidney (HEK) 293 cells. Wild-type and mutant receptors were transiently transfected into HEK 293 cells. The level of m3 muscarinic acetylcholine receptor expression was approximately 300 fmol/mg protein, and the transfection efficiency was approximately 30% for all receptors. Also, wild-type and mutant receptors induced similar 4-fold increases in phosphoinositide (PPI) hydrolysis and showed similar Ca2+ responses after carbachol (CCh) treatment. However, the sequestration of wild-type receptors, determined as the difference between the extent of binding of lipophilic and hydrophilic ligands, occurred in a time- and dose-dependent manner to a maximum of approximately 40% of total receptors. In contrast, sequestration was almost totally blocked in cells expressing Ala550,553 or Ala550,553,554 mutant receptors. To determine the functional significance of sequestration and investigate its relationship to receptor desensitization, cells were preincubated with CCh and then washed free of agonist and restimulated with CCh. Desensitization was manifest as a time- and concentration-dependent decrease in the ability of the second stimulation to increase PPl hydrolysis. One-hour pretreatment with 1 mM CCh decreased PPl hydrolysis by 24% for wild-type receptors but had no effect on the ability of the mutant receptors to respond to a second CCh challenge. Furthermore, inhibition of wild-type receptor sequestration by treatment with conconavalin A also blocked desensitization to a 1-hr treatment with CCh. These results suggest that sequestration may be directly involved in m3 receptor desensitization at early times. More prolonged CCh treatment (3-9 hr) reduced the PPl hydrolysis response of the mutant and the wild-type receptors, indicating that the mechanism of m3 receptor desensitization at later times involves multiple components.

Carbachol↗
PubMed1993

Direct trophic effects of fibroblast growth factors on rat pancreatic acinar cells in vitro.

We examined the effects of fibroblast growth factors (FGFs) on rat pancreatic acinar cells in primary culture. Both basic and acidic FGF stimulated [3H]thymidine incorporation in a dose-dependent fashion. Maximum effects of 6-7 fold over control were seen with 1 nM bFGF or 100 nM aFGF. These data indicate that FGFs are potent stimulants of rat pancreatic acinar cell DNA synthesis. Therefore, FGFs may play an important role in long term regulation of the exocrine pancreas in vivo. We also examined the interaction of bFGF with the pancreatic secretogogues CCK and carbachol. Effects of CCK, which is itself an acinar cell trophic factor, were additive with those of bFGF. In contrast, carbachol, which has no growth stimulatory effect, did not affect bFGF mediated stimulation of DNA synthesis. These data suggest that the mechanisms involved in acinar cell growth regulation are independent from those involved in secretion.

Animals↗
PubMed1993

Identification and cloning of GP-3 from rat pancreatic acinar zymogen granules as a glycosylated membrane-associated lipase.

The protein components of highly purified secretory granule membranes and the granule contents from rat exocrine pancreas were characterized by two-dimensional polyacrylamide gel electrophoresis, protein staining, lectin absorption, and Western blotting with anti-secretory protein antibodies. NH2-terminal amino acid sequence was obtained for a approximately 53-kDa glycoprotein denoted GP-3, present only in granule membrane preparations where it was resistant to washing with Na2CO3 and KBr. The sequence of this protein showed homology to pancreatic lipase but was distinct from the NH2-terminal sequence of a 50-kDa content protein presumed to be secretory lipase. Polymerase chain reaction amplification with degenerate oligonucleotide primers to GP-3 and secretory lipase gave partial length subclones that were used to isolate clones from a rat pancreas cDNA library. Dideoxy sequencing of full-length subclones of GP-3 revealed the predicted amino acid sequence for a mature protein of 452 amino acids with a potential N-linked glycosylation site and a deglycosylated molecular weight of 50,860. The GP-3 sequence possesses the serine esterase consensus sequence G-X-S-X-G centered around Ser154 and the catalytic state triad Asp178-His265-Ser154 characteristic of pancreatic lipases. Northern blot analysis of various rat tissues showed GP-3 expression solely in pancreas. Comparison of GP-3 nucleotide and amino acid sequence, along with pancreatic lipases of various species including rat, shows extensive homologies to both proteins and reveals an underlying diversity in the pancreatic lipase family. Close homology is observed between GP-3 and a lipase molecule previously isolated from mouse cytotoxic T cells.

Amino Acid Sequence↗
PubMed1993

Both low- and high-affinity CCK receptor states mediate trophic effects on rat pancreatic acinar cells.

Cholecystokinin (CCK) stimulates the growth of pancreatic acinar cells. However, the molecular mechanisms involved in this trophic action are unknown. CCK binds to both high- and low-affinity receptor states, and these two states appear to activate separate sets of intracellular messengers and have opposite effects on amylase release. JMV-180 is a CCK analogue that interacts in the rat with the high-affinity state as an agonist and the low-affinity state as an antagonist. In the current study, CCK octapeptide (CCK-8) and JMV-180 were tested for their ability to stimulate the growth of rat pancreatic acinar cells in primary culture. CCK-8 stimulated [3H]thymidine incorporation into DNA in a dose-dependent manner. Effects were observed with 0.3 nM, and maximal increases were seen at 3 nM CCK-8 (442 +/- 53% of control, n = 5, P < 0.01). JMV-180 also stimulated DNA synthesis. Effects were noted with 10 nM, and a maximal increase of 267 +/- 82% (n = 4, P < 0.01) of control was stimulated by 100 nM JMV-180. These data with JMV-180 indicate that the high-affinity receptor state for CCK is capable of stimulating DNA synthesis. However, within the same experiment the effects of CCK were always significantly greater than those of JMV-180. To test whether CCK has an additional effect through interactions with the low-affinity state, the effects of a combination of JMV-180 with a maximal dose of CCK-8 were examined. JMV-180 inhibited the maximal effect of CCK-8 in a dose-dependent manner with a maximal inhibition occurring with 1 microM JMV-180. The effects of the combination of 3 nM CCK-8 and 1 microM JMV-180 were no greater than those of JMV-180 alone. Taken together these data indicate that CCK-mediated increases in DNA synthesis in rat pancreatic acinar cells in vitro occur by interactions with both high- and low-affinity receptor states.

Animals↗
PubMed1993

Human m3 muscarinic acetylcholine receptor carboxyl-terminal threonine resides are required for agonist-induced receptor down-regulation.

The mechanisms involved in agonist-induced down-regulation of the human m3 muscarinic acetylcholine receptor were investigated by site-directed mutagenesis of the receptor cytoplasmic carboxyl terminus. Threonine residues (Thr550,553,554) were converted into alanines collectively and individually. The mutated and wild-type receptor cDNAs stably expressed in Chinese hamster ovary cells displayed similar antagonist- and agonist-binding properties. Furthermore, mutant receptors showed the same efficacy and potency for carbachol-induced activation of phosphoinositide hydrolysis as did the wild-type clone. In all cases the maximal increase in phosphoinositide hydrolysis was 8-9-fold. In contrast to normal intracellular signaling, however, the mutant receptor with all three threonines changed to alanines (Ala550,553,554) failed to undergo normal down-regulation in response to carbachol. After a 24-hr incubation in the presence of 1 mM carbachol, subsequent N-[3H]methylscopolamine binding was reduced by 66% for the wild-type clone but by only 12% for the mutant receptor. The Ala553,554 mutant also showed a profound reduction in receptor down-regulation. Subsequent studies showed that a small but significant blockage of receptor down-regulation also could be produced by converting a single threonine residue (Thr553) to alanine. The fact that these effects were not due to nonspecific conformational changes was suggested by the lack of effects on binding, signal transduction, and down-regulation of converting Thr550 to alanine or converting two cysteine residues (Cys561,563) to glycines in an adjacent region. A similar reduction in receptor number also was observed in binding studies using the membrane-permeant ligand [3H]scopolamine. These results show that threonine residues in the carboxyl-terminal domain of the human m3 muscarinic acetylcholine receptor are important in agonist-induced receptor down-regulation.

Binding Sites↗
PubMed1992

Bovine chromaffin granule membranes undergo Ca(2+)-regulated exocytosis in frog oocytes.

We have devised a new method that permits the investigation of exogenous secretory vesicle function using frog oocytes and bovine chromaffin granules, the secretory vesicles from adrenal chromaffin cells. Highly purified chromaffin granule membranes were injected into Xenopus laevis oocytes. Exocytosis was detected by the appearance of dopamine-beta-hydroxylase of the chromaffin granule membrane in the oocyte plasma membrane. The appearance of dopamine-beta-hydroxylase on the oocyte surface was strongly Ca(2+)-dependent and was stimulated by coinjection of the chromaffin granule membranes with InsP3 or Ca2+/EGTA buffer (18 microM free Ca2+) or by incubation of the injected oocytes in medium containing the Ca2+ ionophore ionomycin. Similar experiments were performed with a subcellular fraction from cultured chromaffin cells enriched with [3H]norepinephrine-containing chromaffin granules. Because the release of [3H]norepinephrine was strongly correlated with the appearance of dopamine-beta-hydroxylase on the oocyte surface, it is likely that intact chromaffin granules and chromaffin granule membranes undergo exocytosis in the oocyte. Thus, the secretory vesicle membrane without normal vesicle contents is competent to undergo the sequence of events leading to exocytosis. Furthermore, the interchangeability of mammalian and amphibian components suggests substantial biochemical conservation of the regulated exocytotic pathway during the evolutionary progression from amphibians to mammals.

Animals↗
PubMed1992

Chinese hamster ovary mRNA-dependent, Na(+)-independent L-leucine transport in Xenopus laevis oocytes.

In freshly prepared uninjected folliculated oocytes, Na(+)-independent leucine uptake is mediated predominantly by a system L-like transport system. Removal of follicular cells, however, results in an irreversible loss of this transport activity. When total poly(A)+ mRNA derived from Chinese hamster ovary (CHO) cells was injected into prophase-arrested stage V or VI Xenopus laevis oocytes, enhanced expression of Na(+)-independent leucine transport was observed. The injected mRNAs associated with increased levels of leucine uptake were between 2 and 3 kb in length. The newly expressed leucine transport activity exhibited important differences from the known characteristics of system L, which is the dominant Na(+)-independent leucine transporter in CHO cells as well as in freshly isolated folliculated oocytes. The CHO mRNA-dependent leucine uptake in oocytes was highly sensitive to the cationic amino acids lysine, arginine, and and ornithine (> 95% inhibition). As with the leucine uptake, an enhanced lysine uptake was also observed in size-fractionated CHO mRNA-injected oocytes. The uptakes of leucine and lysine were mutually inhibitable, suggesting that the newly expressed transporter was responsible for uptakes of both leucine and lysine. The inhibition of uptake of lysine by leucine was Na+ independent, thus clearly distinguishing it from the previously reported endogenous system y+ activity. Furthermore, the high sensitivity to tryptophan of the CHO mRNA-dependent leucine transport was in sharp contrast to the properties of the recently cloned leucine transport-associated gene from rat kidney tissue, although leucine transport from both sources was sensitive to cationic amino acids. Our results suggest that there may be a family of leucine transporters operative in different tissues and possibly under different conditions.

Animals↗
PubMed1992

Role of polyamines in glucocorticoid effects on pancreatic acinar AR42J cell growth and differentiation.

We previously found that glucocorticoids inhibit growth and increase differentiation in rat pancreatic acinar AR42J cells. In the current study, we examined the role of polyamines in these effects. Treatment of AR42J cells with the ornithine decarboxylase (ODC) inhibitor difluoromethylornithine (DFMO) inhibited DNA synthesis. Thus polyamines are required for AR42J cell growth. However, we have previously shown that dexamethasone (Dex) increased AR42J cell ODC activity and mRNA levels. In the current study, we found that Dex treatment increased cellular putrescine levels. These increases in ODC and putrescine occurred during Dex-induced inhibition of DNA synthesis. Therefore, in AR42J cells, ODC activity and polyamine levels are not strictly growth related. To examine the requirement for glucocorticoid induction of ODC activity in glucocorticoid stimulation of differentiation, we examined the effects of DFMO on amylase gene expression and cholecystokinin binding. DFMO reduced cell amylase content while having little effect on mRNA levels in both Dex-treated and untreated cells. In contrast, DFMO had little effect on control CCK binding but inhibited the Dex-induced increase. Thus polyamines are necessary for growth and glucocorticoid-induced differentiation of AR42J cells; however, effects of glucocorticoids on AR42J cell growth and differentiation are not mediated by effects on ODC.

Amylases↗