Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have identified and characterized receptors for the amino-terminal domains of PTH and PTH-like peptide (PLP) on an immortalized human keratinocyte cell line, RHEK-1. Binding of both PLP-(1-34) and PTH-(1-34) to the RHEK-1 cells was consistent with a two-site model; affinities and capacities for each site were similar for the two peptides. Both peptides also stimulated adenylate cyclase activity with an equal ED50 in this cell line. Pertussis toxin pretreatment enhanced this peptide-mediated enzyme activity, suggesting linkage of the receptor to an inhibitory guanyl nucleotide-binding protein (Gi). Adenylate cyclase activity was diminished by both homologous [PLP-(1-34)] and heterologous [epidermal growth factor (EGF)] effectors. Malignant conversion of the immortalized cells with an activated H-ras oncogene to produce the RHEK-ras cell line was associated with a reduction in binding at both PLP/PTH and EGF receptors as well as a postreceptor defect in PLP/PTH-stimulated adenylate cyclase activity. The defect in enzyme activity appeared to be due in part to a decrease in the activity of the stimulatory guanyl nucleotide-binding protein (Gs), but not to an increase in Gi activity. Activation of the keratinocyte amino-terminal PLP/PTH receptor resulted in a small increase in [3H]thymidine incorporation, which was associated with an increase in cell numbers. This mitogenic effect was enhanced in the presence of EGF and was markedly reduced when cells were cultured in a high extracellular calcium environment. These studies demonstrate that the amino-terminal region of PLP and PTH activates adenylate cyclase-linked receptors, which are associated with mitogenesis, in RHEK-1 cells and suggest that this cell line represents a suitable model in which to examine the actions of PLP in keratinocytes.
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PMID:Identification and functional characterization of adenylate cyclase-linked receptors for parathyroid hormone-like peptides on immortalized human keratinocytes. 130 43

Ca2+ and other polyvalent cations as well as polycations, such as neomycin, produce similar effects on intracellular second messengers and PTH release in dispersed bovine parathyroid cells, but it is unclear whether all of these agents share the same mechanism of action. The lectin Concanavalin-A (Con-A) and the activator of protein kinase-C tetradecanoylphorbol acetate (TPA) blunt the effects of elevated extracellular calcium (Ca2+) concentrations on several aspects of parathyroid function, including PTH release, the cytosolic calcium concentration, and the accumulation of cAMP and inositol phosphates. In the present studies we used these two agents as well as pertussis toxin as probes to investigate further whether neomycin acts on parathyroid cells through the same receptor-like mechanism used by extracellular Ca2+ to regulate parathyroid function. Con-A and TPA both enhanced PTH release by about 2-fold at 0.5-1 x 10(-4) M neomycin, concentrations that inhibited PTH release to an extent (40-50%) similar to that seen with high (1.5-2 mM) Ca2+. Con-A also reduced the inhibition of agonist-stimulated cAMP accumulation by the same concentrations of neomycin. Conversely, Con-A and TPA produced 70-80% decreases in the cytosolic calcium concentration transient and the accumulation of inositol phosphates stimulated by neomycin. The effects of these two agents on neomycin-regulated parathyroid function were similar in magnitude to their actions on the modulation of these same parameters by extracellular Ca2+. Pertussis toxin, however, which we have previously shown to block the inhibitory effects of high Ca2+ and neomycin on cAMP accumulation, had no effect on the inhibition of PTH release by these two agents. These results provide further indirect evidence that polyvalent cations and polycations act on the parathyroid cell through related pathways, which probably involve cell surface moieties containing carbohydrate(s).
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PMID:A comparison of the effects of concanavalin-A and tetradecanoylphorbol acetate on the modulation of parathyroid function by extracellular calcium and neomycin in dispersed bovine parathyroid cells. 131 77

Acquired renal cysts derive from terminally differentiated tubular epithelium in adults as a consequence of increased epithelial cell proliferation, fluid accumulation and extracellular matrix remodelling. To understand better how human epithelial cysts may be initiated and progressively expand, cells from primary cultures of normal human adult renal cortex were dispersed in polymerized type I collagen. The transparent matrix permitted repeated observation by light microscopy of cyst formation from individual renal cells. The cyst cells reacted strongly with distal nephron histochemical markers (cytokeratin antibodies AE1/AE3, epithelial membrane antigen, and Arachis hypogaea lectin) but inconsistently or not at all to markers of proximal tubules (Tetragonolobus purpureas lectin and Phaseolus vulgaris erthroagglutinin lectin). The number of spherical, fluid-filled epithelial cysts that developed in a standardized microscope field quantified cyst initiation. Cyst progression was determined from the increase in the diameter (surface area) of cysts and represents a hyperplastic event. EGF or TGF alpha, were required in serum-free defined medium to cause cysts to develop from individual epithelial cells dispersed in the matrix; insulin was required as a co-factor. The EC50 for EGF was approximately 0.1 ng/ml, and for insulin 1 microgram/ml. Early cultures of normal cortex formed cysts more efficiently when dispersed in collagen matrix than cells passaged several times before suspension in the gel. Agonists of adenylate cyclase (PGE1, AVP, VIP, PTH, forskolin, cholera toxin), methylisobutylxanthine, and 8-Br-cAMP, though incapable of causing cyst formation alone in defined medium, enhanced cyst initiation and progression in the presence of EGF and insulin. Angiotensin II, TNF alpha, beta-estradiol, and pertussis toxin had no effect in the absence or presence of EGF and insulin. Pertussis toxin inhibited cyst initiation and expansion caused by EGF and forskolin but potentiated cyst initiation and expansion caused by EGF and PGE1. Cyst formation and expansion were inhibited by TGF beta 1 and 2-chloroadenosine. Polarized monolayers of human renal cortical cells grown on permeable membranes were used to independently quantify the effects of agonists on the net secretion of solute and water from the basolateral to the apical surface of the cells. PGE1, forskolin, and 8-Br-cAMP stimulated net fluid secretion that was sustained for several days; EGF enhanced forskolin-stimulated fluid secretion. We conclude that the formation and expansion of in vitro cysts derived from solitary human cortex cells depends on the coordinated interplay between cellular proliferation and fluid secretion.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:In vitro formation and expansion of cysts derived from human renal cortex epithelial cells. 131 21

Transforming growth factor-beta (TGF beta) produced by osteoblasts is present in high levels in bone and influences bone formation, replication of bone cells, and expression of osteoblast protein products. Interactions between bone active hormones and locally released and activated TGF beta were studied by examining the influence of TGF beta preincubation on PTH, calcitonin (CT), and vitamin D receptors in an osteoblastic cell line (UMR 106-06). Preincubation of UMR 106-06 cells with 1 ng/ml TGF beta for 3 days increased specific binding of [125I]PTH-related protein (PTHrP)(1-84) to 140% of that in control cells, but [125I]salmon CT binding decreased to 50% of controls. Binding isotherms indicated that the changes in binding were due to altered receptor numbers since affinities for 125I-labeled PTH and CT remained unchanged. The effect on receptor levels was time dependent, requiring 24 h preincubation with TGF beta for measurable changes, and dose dependent, with maximal effects seen with 1 ng/ml TGF beta. Binding of [3H]1,25(OH)2 vitamin D3 was increased to 130% of control in cytosolic extracts of UMR 106-06 cells pretreated for 3 days with 1 ng/ml TGF beta. Scatchard plots suggested an increase in receptor number without change in affinity. The adenylate cyclase response to PTH increased to 150% of control cells after 3 days of treatment with 1 ng/ml TGF beta; however, the adenylate cyclase response to CT was little changed. Forskolin- and cholera toxin-stimulated adenylate cyclase responses were increased by TGF beta treatment to 130-160% of control, indicating an increase in the stimulatory subunit of the G protein. Increased abundance of both Gs and Gi proteins were indicated by increased cholera toxin- or pertussis toxin-dependent [32P] NAD ribosylation of 47-kilodalton (kDa) and 42-kDa or 40-kDa proteins, respectively, in TGF beta-treated cells. Our data support a complex regulatory effect of TGF beta on UMR 106-06 cells with increases in PTH receptors, vitamin D receptors, and G proteins, whereas there is an apparent down-regulation of CT receptors. TGF beta might induce a more differentiated osteoblast phenotype of these cells, which already express differentiated features such as high alkaline phosphatase activity, PTH and vitamin D receptors, and collagenase production. Since low doses of PTH stimulate bone formation in vivo, TGF beta released or activated at sites of new bone formation might locally modulate PTH activity be allowing increased PTH receptor and postreceptor effectiveness.
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PMID:Transforming growth factor-beta modulates receptor binding of calciotropic hormones and G protein-mediated adenylate cyclase responses in osteoblast-like cells. 132 61

We have reported that pretreatment of human SaOS-2 osteoblast-like cells with forskolin (Fsk; 10(-5) M) for 4 h strikingly inhibited subsequent cAMP responsiveness to a second challenge with Fsk (Fsk-induced homologous desensitization) without altering the responses to PTH or vasoactive intestinal peptide (VIP). Pretreatment with PTH acutely augmented Fsk responsiveness, despite desensitizing the cells to rechallenge with PTH. The present studies were performed to investigate the mechanism of this differential desensitization. Fsk-induced desensitization was not mimicked by 1,9-dideoxyforskolin, a Fsk analog that does not activate adenylate cyclase (AC) but does reproduce certain cAMP-independent effects of Fsk. Fsk-induced homologous desensitization was also completely blocked in a cAMP-resistant mutant SaOS-2 cell line (Ca 4A), in which protein kinase-A (PKA) is not activated by endogenous cAMP. However, pretreatment with PTH (or VIP), which induced a large increase in cAMP, did not attenuate, but, rather, increased, the subsequent cAMP response to Fsk. Potentiation by PTH was also observed in Ca 4A cells. Pretreatment of SaOS-2 cells with pertussis toxin (100 ng/ml) for 12 h strikingly inhibited the initial cAMP response to Fsk, although Fsk-induced homologous desensitization was still clearly observed. Pretreatment with cholera toxin (1 microgram/ml) completely prevented Fsk-induced homologous desensitization. Combinations of maximal concentrations of Fsk plus hormones such as human PTH, human PTH-related peptide, or VIP elicited cAMP responses that were much more than additive, an effect not observed with combinations of hormones alone. We conclude that 1) Fsk-induced homologous desensitization of the AC response of SaOS-2 cells to a second challenge with Fsk is dependent upon activation of PKA; 2) one or more pertussis toxin-sensitive G-proteins contribute to full AC activation by Fsk, but are not involved in homologous desensitization; 3) augmentation by PTH (or VIP) pretreatment of Fsk-dependent AC activation involves an effector(s) other than PKA. These results provide further evidence that the regulation of AC responsiveness in SaOS-2 cells by PTH or VIP is complex and cannot be explained by activation of PKA alone.
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PMID:Forskolin-induced homologous desensitization via an adenosine 3',5'-monophosphate-dependent mechanism(s) in human osteoblast-like SaOS-2 cells. 132 19

In UMR-106 osteosarcoma cells we found that PTH activated both the cAMP/protein kinase A and the Ca(2+)-dependent phosphoinositide/protein kinase C (PKC) pathways, but prostaglandin E2 (PGE2) activated only the cAMP pathway. Activation of PKC by the phorbol ester PMA had no effect on cAMP production but enhanced PTH-stimulated cAMP production by 50% or more; the effect on PGE2-induced cAMP was negligible. Inhibition of the alpha-subunit of the inhibitory guanine nucleotide binding protein (Gi) by pertussis toxin pretreatment also enhanced PTH-mediated cAMP production but had no effect on PGE2-induced cAMP production. These results suggest that although PTH-mediated adenylate cyclase activity is regulated via both the stimulatory (Gs) and inhibitory (Gi) guanine nucleotide binding proteins, only Gs regulates PGE2-mediated adenylate cyclase activity in UMR-106 cells. Costimulation with pertussis toxin and PMA did not increase PTH-stimulated cAMP production above that obtained with PMA alone. This implies a similar target of action for pertussis toxin and PMA, that is, the alpha-subunit of Gi. The alpha-subunit of Gi was found to be a substrate for in vitro PKC phosphorylation of membrane fractions from UMR-106 cells, seen as a +/- 40 kD band on SDS-PAGE. Stimulation of in situ 32P-labeled cells with either PMA or PTH also enhanced incorporation of 32P into the 40 kD band. Using the peptide antisera AS/7 and EC/2, we showed that pertussis toxin-labeled subunits of both Gi1 alpha/Gi2 alpha and Gi3 alpha could be immunoprecipitated, respectively, but immunoprecipitation of membrane proteins after in situ phosphorylation and stimulation with PMA precipitated only Gi2 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Protein kinase C modulates parathyroid hormone- but not prostaglandin E2-mediated stimulation of cyclic AMP production via the inhibitory guanine nucleotide binding protein in UMR-106 osteosarcoma cells. 133

Signal transduction by the PTH receptor is now known to involve generation of multiple second messengers. Desensitization of the adenylate cyclase response to PTH is a common feature of bone- and kidney-derived target cells; however, no single mechanism appears to explain desensitization in the different cell types studied. To examine the role of protein kinase-A (PKA) in homologous desensitization to PTH, we employed human SaOS-2 osteoblast-like cells and a mutant subclone (Ca 4A), which expresses an inducible cAMP-resistant form of PKA. Pretreatment of SaOS-2 cells with PTH for 4 h reduced by 60-80% the cAMP response to subsequent rechallenge with the hormone. This homologous desensitization was significantly, but not completely, inhibited in Ca 4A cells. Desensitization was not mimicked by pretreatment of the cells with forskolin. PTH binding to its receptor was reduced 50% in both SaOS-2 and Ca 4A cells after 4-h incubation with PTH (homologous down-regulation), whereas forskolin did not cause receptor down-regulation. Pretreatment with the ionophore ionomycin for 4-24 h did not mimic desensitization to PTH. Both desensitization to PTH and receptor down-regulation were induced, however, by pretreatment with a phorbol ester (12-O-tetradecanoyl phorbol-13-acetate), and these effects were blocked completely by staurosporine. PTH-induced desensitization was not blocked by staurosporine, and receptor down-regulation was enhanced by the drug. Pertussis toxin did not prevent desensitization induced by either PTH or 12-O-tetradecanoyl phorbol-13-acetate. We conclude that homologous desensitization to PTH in SaOS-2 cells involves both cAMP-dependent and -independent mechanisms. Homologous PTH receptor down-regulation apparently is mediated by mechanisms independent of PKA activation. Neither pathway of homologous desensitization to PTH involves the action of pertussis toxin-sensitive G-proteins.
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PMID:Mechanisms of desensitization to parathyroid hormone in human osteoblast-like SaOS-2 cells. 139 21

PTH activates multiple second messengers in its target cells, but the level at which the hormonal signal splits into different pathways is still unknown. To achieve insights on this issue, we have studied the structure-function relationship of PTH by analyzing the effects of bovine PTH-(1-34) [bPTH-(1-34)] and PTH fragments truncated at the N-terminus on the intracellular calcium concentration [( Ca2+]i) and cAMP production in the rat osteogenic sarcoma cell line UMR 106-01. [Ca2+]i was measured in single cells using fura-2. When exposed to 10(-7) M bPTH-(1-34), 20% of the cells responded with a transient increase in [Ca2+]i of variable amplitude. Equimolar doses of bPTH-(2-34), propionyl bPTH-(2-34) [(pbPTH-(2-34)], and bPTH-(3-34) also transiently increased [Ca2+]i, whereas both [tyrosine34]bPTH-(7-34) amide [bPTH-(7-34)] and bPTH-(30-34) were ineffective. The amplitude of the [Ca2+] i transients was dose-dependent, with threshold concentrations of 10(-10) M for bPTH-(1-34) and bPTH-(2-34), and 10(-9) M for bPTH-(3-34). The response rate to the active peptides ranged between 10-30%, without a clear dose-relatedness. A second addition of 10(-7) M bPTH-(1-34) to cells prestimulated with equimolar doses of bPTH-(2-34), pbPTH-(2-34), or bPTH-(3-34) produced another transient, whereas after exposure to 10(-7) M bPTH-(1-34), the cells were completely desensitized to a second homologous stimulation, suggesting that the binding affinity of the truncated peptides for the PTH receptor is lower than that of the intact bPTH-(1-34) fragment. In addition, both bPTH-(1-34) and bPTH-(2-34) dose-dependently stimulated cAMP production, but the former was more potent (ED50 = 10(-9) vs. 10(-7) M, respectively). On the contrary, pbPTH-(2-34), bPTH-(3-34), and bPTH-(7-34) had no effect on cAMP. Pretreating the cells with pertussis toxin to enhance cAMP responses via inhibition of Gi potentiated the effect of bPTH-(1-34) and bPTH-(2-34) and disclosed weak but detectable agonist action of pbPTH-(2-34). These results indicate that specific domains of the PTH molecule are linked to activation of different second messenger pathways; while the first two amino acids are indispensable for activating the cAMP system, generation of the [Ca2+]i signal appears to involve a longer domain, including the amino acid residue in position 3.
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PMID:Dissociation of second messenger activation by parathyroid hormone fragments in osteosarcoma cells. 164 59

We examined the effects of the polycationic antibiotic, neomycin, on the function of dispersed bovine parathyroid cells. Neomycin caused a reversible, dose-dependent inhibition of low calcium (Ca++)-stimulated PTH release, with half-maximal inhibition at 30 microM. Maximal inhibition (with 200 microM neomycin) was not additive with the suppressive effects of high (2 mM) Ca++. Neomycin also inhibited dopamine-stimulated cAMP accumulation by 90-98% at 100-200 microM, with a half-maximal effect at 40-50 microM. This action was reversible and was blocked by preincubating the cells overnight with 0.5 microgram/ml pertussis toxin. In addition to its suppressive effects on cAMP metabolism and PTH release, neomycin stimulated the accumulation of inositol phosphates and produced a transient increase in the cytosolic Ca++ concentration (Cai) in fura-2-loaded parathyroid cells. The neomycin-evoked spike in Cai persisted despite removal of extracellular Ca++, indicating that it arises from intracellular Ca++ stores. Exposure of cells to elevated magnesium (Mg++) concentrations elicited a similar spike in Cai but blocked the spike in Cai in response to subsequent addition of neomycin and vice versa. Thus, Mg++ and neomycin mobilize Ca++ from the same intracellular store(s). These results indicate that a polycation, neomycin, closely mimics the effects of polyvalent cations on parathyroid function, suggesting that both agents regulate parathyroid function via similar biochemical pathways.
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PMID:Neomycin mimics the effects of high extracellular calcium concentrations on parathyroid function in dispersed bovine parathyroid cells. 164 60

We investigated the effects of the basic peptides polyarginine, protamine, and polylysine on dispersed bovine parathyroid cells. All three peptides produced a dose-dependent inhibition of dopamine-stimulated cAMP accumulation, with half-maximal inhibition at 4 x 10(-8), 1.5 x 10(-7), 3 x 10(-7), and 2 x 10(-6) M, respectively, for polyarginine, protamine, and two preparations of polylysine of molecular weights 10,200 and 3800. The inhibition of cAMP accumulation was reversible and was blocked by preincubating the cells overnight with 0.5 micrograms/ml of pertussis toxin. The same peptides also inhibited PTH release at similar concentrations, markedly stimulated the accumulation of inositol phosphates at two- to threefold higher concentrations, and produced transient increases in the cytosolic Ca2+ concentration (Cai) in fura-2-loaded parathyroid cells. The polylysine-evoked spike in Cai persisted despite the removal of extracellular Ca2+, indicating that it arose from intracellular Ca2+ stores. Exposure of the cells to elevated extracellular magnesium (Mg2+) concentrations elicited a similar spike in Cai but blocked the Cai transient in response to subsequent addition of polylysine, or vice versa. Thus, Mg2+ and polylysine mobilize Ca2+ from the same intracellular store(s). These results indicate that highly basic peptides closely mimic the effects of polyvalent cations on parathyroid function, suggesting that both agents may regulate parathyroid function via similar biochemical pathways.
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PMID:Polyarginine, polylysine, and protamine mimic the effects of high extracellular calcium concentrations on dispersed bovine parathyroid cells. 166 8


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