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P Pradelles

Publications and source records attributed to P Pradelles.

At least 55 records · Page 3Linked to original sources
PubMed1991

[Variation of substance P-like immunoreactivity in plasma and cerebrospinal fluid in the course of arthritis induced by Freund adjuvant in rats, a model for the study of chronic pain].

Parallel time courses of clinical and behavioural parameters and levels of plasma substance P-like immunoreactivity (SPLI-PI) were studied in arthritic rats (adjuvant induced arthritis, AIA, a chronic pain model). Acute (14 and 21 post-inoculation days,PI) and post-acute (42 days PI) phases of the syndrome were investigated. These data were compared with those obtained in a control situation (inoculation day). In a second experimental series, levels of substance P-like immunoreactivity in cerebrospinal fluid (SPLI-CSF) were determined at the same stages of AIA. In arthritic rats SPLI-PI was strongly enhanced (X4) as early as 14 days PI and remained increased (X4) at all stages studied, whereas SPLI-LCR was significantly increased (X2) only 21 days PI and returned to control levels at 42 days PI. These data suggest that SP could be distributed in two different pools, a peripheral one of inflammatory origin, and a central one which could be more specific to the chronic pain situation.

Animals↗
PubMed1991

Human placental low molecular weight factors inhibit the entry into cell cycle of murine pluripotent stem cells.

In 1977, Frindel et al. reported the presence in fetal calf bone marrow of a low molecular factor, the tetrapeptide Ac-N-Ser-Asp-Lys-Pro (AcSDKP), capable of inhibiting in vivo the hematopoietic pluripotent stem cell (CFU-S) recruitment into DNA synthesis and to increase the survival of mice which had received lethal doses of cytosine arabinoside (AraC), a phase-specific drug. Considering the potential clinical importance of CFU-S proliferation inhibitor during anticancer chemotherapy and the importance of monitoring the inhibitor by immunological methods, we tried to determine if a similar inhibitor is present in humans. Preliminary results indicate the presence in human placenta of an inhibitor coeluted with AcSDKP and which is effective in inhibiting murine CFU-S.

Amino Acid Sequence↗
PubMed1990

Characterization of a B2-bradykinin receptor in rat renal mesangial cells.

The specific binding of bradykinin (BK) was investigated using membrane fractions from mesangial cells in primary culture, a cloned cell line, and in intact adherent cells with three different radiolabelled BK analogues: 125I-[Tyr0]BK, 125I-[Tyr5]BK and 125I-[Tyr8]BK. The best radioligand was 125I-[Tyr0]BK, and assay conditions were determined to ensure maximal stable binding. Binding appeared to be reversible and not to be inhibited by a wide variety of protease inhibitors including converting enzyme inhibitor and phosphoramidon. The maximum density of binding sites (Bmax) was about 88 +/- 18 fmol/mg protein, which is equivalent to about 6000 sites/cell, and the dissociation constant averaged 2 nM. No significant difference in Bmax was observed between membranes from cells in primary culture and those from cloned cells. Of the BK analogues tested, unmodified BK exhibited the highest inhibition constant (close to 10(-10) M). No displacement of 125I-[Tyr0]BK was observed in the presence of the B1 agonist des-Arg9-BK or several unrelated peptides, including atrial natriuretic factor and angiotensin I and II, whereas 50% inhibition of binding was achieved with the B2 antagonist [D-Arg,Hyp3,D-Phe7]BK (10(-9)M). Addition of BK for 3 min to the incubation medium of cloned mesangial cells induced a dose- and time-dependent increase in PGE2 unlike des-Arg9-BK, which showed no such effect. The secretion was strongly inhibited by prior incubation with the B2 antagonist [D-Arg,Hyp3,D-Phe7]BK. The pharmacological profile of the binding site determined with various BK agonists and antagonists, and the stimulating effect of binding site activation on prostaglandin release strongly suggest that B2-kinin-like receptors are present in rat mesangial cells.

Animals↗
PubMed1990

Anti-substance P anti-idiotypic antibodies modulate the secretory process in the rat parotid gland in vitro.

Anti-idiotypic antibodies (anti-Id) obtained in rabbits in response to immunization with polyclonal anti-substance P antibodies (anti-SP) were shown to bind specifically and with high affinity to membranes from rat parotid gland cells. Whereas substance P (SP) was unable to displace anti-Id from membrane binding sites, anti-Id partly inhibited the binding of radiolabelled substance P. Like substance P, anti-Id were able to trigger protein secretion by parotid cells i.e. to behave as physiological agonists of the neuropeptide. Under our experimental conditions, the biological effects of both ligands appear to be additive. Unlike substance P, however, anti-Id did not potentiate the secretory response induced by a beta-adrenoceptor agonist. Taken together, the present results might indicate that anti-Id interact with epitope(s) at or/and near the peptide-combining site on the substance P receptor. These data demonstrate further the possibility of raising pharmacologically active anti-receptor antibodies through the immunological anti-idiotypic approach.

Animals↗
PubMed1990

Negative regulator of pluripotent hematopoietic stem cell proliferation in human white blood cells and plasma as analysed by enzyme immunoassay.

This paper describes the analysis, by a highly sensitive and specific enzyme immunoassay (EIA), of AcSDKP, a tetrapeptide recently isolated from fetal calf bone marrow and subsequently purified and identified which substantially inhibits entry into cycle of hematopoietic pluripotent stem cells (CFU-S). This molecule has a marked protective effect in mice during anticancer chemotherapy with phase-specific drugs and plays an essential role in maintaining CFU-S out of cycle in normal mice. Using acetylcholinesterase-AcSDKP conjugate as tracer, rabbit specific antiserum and 96-well microtiter plates coated with a mouse monoclonal anti-rabbit IgG antibody, this EIA allows detection of AcSDKP at 15 fmol levels with a coefficient of variation less than 10% in the 50-500 fmol range. When combined with high-performance liquid chromatography, this assay clearly reveals the presence of this peptide in normal human white blood cells whereas in supernatant from cultured lymphocytes and in plasma the immunoreactive material is distinct from standard AcSDKP.

Acetylcholinesterase↗
PubMed1990

Evidence for Pro-dermorphin processing products in rat tissues.

Dermorphin (Tyr-D.Ala-Phe-Gly-Tyr-Pro-Ser-NH2) and dermenkephalin (Tyr-D.Met-Phe-His-Leu-Met-Asp-NH2), two powerful opioid peptides issued from a common biosynthetic precursor, were recently isolated from the skin of the frog phyllomedusa sauvagii. Since many amphibian's skin secretory peptides or their homologues are found in the mammalian central nervous system and gastrointestinal tract, dermorphin and dermenkephalin may have counterparts in mammals. We have prepared antibodies directed against dermorphin, dermenkephalin and the spacer sequence lying between them in the progenitor form and developed sensitive enzyme immunoassays that detect under the picogram level to verify the occurrence of these peptides in rat tissues. Immunocytochemical studies of rat brain sections revealed a similar distribution of immunoreactivities both at the regional and the cellular levels when either one of these three anti-sera was used. Characterization of the immunoreactive peptides was achieved with molecular sieve filtration followed with Reverse Phase High Performance Liquid Chromatography of various rat tissues extracts. Identification was achieved by immunological analysis and chromatographic comparison with synthetic peptides. Immunoreactive materials corresponding to dermorphin, dermenkephalin and the spacer peptide were detected in either brain, stomach or intestine, indicating processing of the dermorphin precursor in these tissues. Immunoreactive species of higher Mr were also detected in all three tissues and may represent extended forms or homologous peptides.

Amino Acid Sequence↗
PubMed1990

Use of anti-idiotypic antibodies as probes for in vitro and in vivo identification of substance P receptor.

In order to develop immunological tools for studying the receptor of the neuropeptide substance P (SP), anti-idiotypic antibodies (anti-Id abs) were produced by immunization with anti-SP antibodies whose specificity was close to that of the SP receptor. Immunological studies revealed structural similarity between some anti-Id abs and SP itself. As a consequence, these anti-Id abs were able to bind to mammalian SP receptors. These antibodies were used in immunocytochemistry to label SP receptors both in the rat spinal cord and in rat and guinea pig peripheral tissues (parotid gland and trachea, respectively). Like SP, anti-Id abs were able to trigger protein secretion by isolated rat parotid gland cells. Finally, it was shown that anti-Id abs in vivo modulated reactivity to chemical stimuli. These antibodies therefore appear to be promising tools for further biochemical, cytochemical, and pharmacological characterization of SP receptors.

Animals↗
PubMed1990

Enzyme immunoassays for thyroliberin (TRH) and TRH-elongated peptides in mouse and rat hypothalamus.

Enzyme immunoassays (EIAs) for Thyroliberin (TRH) and TRH-elongated peptides were developed. Three haptens less than E-H-P-NH2 (TRH). Less than E-H-P-OH (TRH-OH), and S-K-R-Q-H-P-G-K-R-F (P10) were conjugated by the use of different heterobifunctional cross-linking agents either to sun-flower globulin as carrier or to acetylcholinesterase as tracer. For a same hapten, the same chemical group in the peptide was used to prepare the immunogen and the enzyme conjugate. These EIAs were performed with a second antibody solid phase technique using acetylcholinesterase as label. They permitted the measurement of TRH and TRH-elongated peptides with a sensitivity threshold of 10 fmol/well for TRH and 2 fmol/well for P10. TRH EIA only detected authentic TRH whereas TRH-OH EIA detected TRH and TRH peptides elongated on C terminal part. Anti-P10 serum was specific of TRH peptides elongated both on C and N terminal parts and no cross reactivity was observed with TRH. Using these assays, TRH and TRH-elongated peptides were determined in crude or chromatographed mouse and rat hypothalamus tissular extracts. Several TRH extended forms were identified by P10 EIA, whereas TRH-OH EIA permitted detection of both TRH and TRH-elongated peptides in chromatographed extracts. Authentic TRH was measured by TRH EIA both in crude and chromatographed hypothalamic extracts. These assays can permit the study of the processing and maturation of TRH.

Amino Acid Sequence↗
PubMed1990

In vitro desensitization of isolated nephron segments to vasopressin.

Recent studies have demonstrated that in vivo administration of 1-deamino-8-D-arginine-vasopressin, an analog of arginine-8-vasopressin, induces homologous desensitization to vasopressin in the thick ascending limb of the loop of Henle. Desensitization has been documented by a decreased physiological response to vasopressin in vivo and by a reduced cAMP accumulation in the cortical thick ascending limb (CTAL). By measuring cAMP content in single isolated medullary thick ascending limbs (MTALs), we now report that desensitization can occur all along the thick ascending limb and, more importantly, that it can also be induced in vitro. In a first series of experiments, we observed that 1 hr after in vivo injection of 1-deamino-8-D-arginine-vasopressin, MTALs were desensitized by 80% to vasopressin, whereas the effects of the other hormones acting on the same cyclase pool (glucagon, calcitonin) were fully maintained. In a second set of experiments, desensitization was induced in vitro by vasopressin, the natural hormone. A 60-min preincubation of MTALs with vasopressin caused a marked (up to 86%) and highly reproducible desensitization. The process was dose and time dependent. The apparent Ka for desensitization was 0.2 nM, and the half-maximal effect was obtained within 20 min. The desensitization induced in vitro by vasopressin was again essentially homologous in nature, with 80% of the maximal stimulation of cAMP accumulation being obtained in the presence of glucagon. Desensitization to vasopressin was observed in the presence and absence of indomethacin, indicating that it is independent of prostaglandin synthesis. It is concluded that (i) vasopressin and its analog 1-deamino-8-D-arginine-vasopressin cause marked desensitization in the CTAL and MTAL and (ii) the low vasopressin concentrations required to induce desensitization and the rapid onset of the process suggest that it has a physiological significance.

1-Methyl-3-isobutylxanthine↗
PubMed1990

Distribution of 11 beta-hydroxysteroid dehydrogenase along the rabbit nephron.

It has been recently proposed that 11 beta-hydroxysteroid dehydrogenase (11 beta-OHSD) is responsible for aldosterone tissue specificity. A 11 beta-OHSD deficiency has been invoked as a cause of the syndrome of apparent mineralocorticoid excess, and 11 beta-OHSD inhibition by liquorice has been invoked to explain the hypertension induced by this drug. Since the renal tubule is composed of aldosterone-sensitive and insensitive segments, we determined the distribution of 11 beta-OHSD along the rabbit tubule. Pools of tubular segments isolated by microdissection were incubated for 2 h at 37 degrees C in the presence of [3H]corticosterone (3H-B, 8.10(-9) M). Afterwards, the amounts of 3H-B and of the metabolite 11-dehydrocorticosterone (3H-A) were determined using HPLC analysis. In the proximal tubule, in either its convoluted or straight portion, and in the medullary thick ascending limb, the amount of 3H-A was 19.6 +/- 3.8% (n = 12), 17.9 +/- 3.4 (n = 8), and 15.0 +/- 2.2 (n = 4), respectively, of the sum of 3H-A + 3H-B. In the cortical ascending limb and the collecting tubule in its cortical and medullary parts, it was 74.7 +/- 6.8% (n = 4), 74.1 +/- 4.9 (n = 9) and 64.6 +/- 14.1 (n = 3), respectively. In both proximal and cortical collecting tubule, addition of carbenoxolone 8.10(-4) M, an inhibitor of 11 beta-OHSD, almost completely inhibited the conversion of 3H-B to 3H-A. Thus, 11 beta-OHSD activity was high in the aldosterone-sensitive segments, and low in the aldosterone-insensitive segments. These results strongly favor the hypothesis that 11 beta-OHSD is a key enzyme in mineralocorticoid tissue specificity along the rabbit nephron. They reinforce the notion that a defect in 11 beta-OHSD plays a major role in the syndrome of apparent mineralocorticoid excess and liquorice-induced hypertension.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1990

Evidence for high molecular weight immunoreactive thyrotropin-releasing hormone (TRH) precursor forms in the developing mouse hypothalamus. Simultaneous immunolocalization with TRH in cultured neurons.

Two antisera (Anti-P7 and Anti-P10) were raised against (-Gln-His-Pro-Gly-) elongated peptides: P7 Gln-His-Pro-Gly-Lys-Arg-Phe) and P10 (Ser-Lys-Arg-Gln-His-Pro-Gly-Lys-Arg-Phe). They recognized TRH extended peptides but not TRH. A RIA against P7 and a highly sensitive enzyme immunoassay against P10 were used to identify two major high mol wt forms of 25-35 K and 6-8 K in chromatography fractions of adult and fetal mouse as well as adult rat hypothalami. The existence of the largest form was confirmed by immunoblotting with Anti-P7. During mouse hypothalamus development in vivo and in vitro, the ratio of TRH content vs. P10-associated immunoreactivity increased several times. This suggests that these Pro-TRH peptides are precursors of TRH biosynthesis and indicate an acceleration of TRH processing during development. Double immunostaining with A-TRH and A-P7 of hypothalamic cells taken on the 16th fetal day and cultured for 6, 12, and 18 days in vitro (DIV) revealed three populations of neurons: 1) a very minor population (approximately 2%) of small round cells positive with A-TRH only; 2) a major population of neurons positive with both A-TRH and A-P7. 3) multipolar neurons positive with A-P7 only (up to approximately 45% after 18 DIV). The respective distribution of TRH and P7 along neurites also varied with time in culture. Whatever perikarya staining, TRH was restricted to short neurites and growth cones before synapse formation and, during synapse development, to varicosities and terminal boutons. However even at the latest stage examined some varicosities and terminal boutons were positive with A-P7 only. These results suggest a preferential processing of pro-TRH at a post-Golgi step during axonal transport to growth cones and synaptic boutons.

Amino Acid Sequence↗
PubMed1989

Production of monoclonal antibodies against interleukin-1 alpha and -1 beta. Development of two enzyme immunometric assays (EIA) using acetylcholinesterase and their application to biological media.

We describe two series of monoclonal antibodies (mAbs) directed against human interleukin-1 alpha (36 mAbs) and -1 beta (11 mAbs). The binding compatibility of each of mAb was studied using biotin-labelled mAbs in immunometric tests. Among the different pairs of compatible mAbs, we selected one pair for each interleukin-1 (IL-1) with optimal properties for a two-site immunometric assay. In these assays, covalent conjugates of mAb coupled to the tetrameric form of acetylcholinesterase (mAb-AChE) were used as tracers. The tests were performed in 96-well microtiter plates coated with the complementary mAb. Both assays appeared sensitive and specific since minimum detectable concentrations as low as 1 pg/ml were determined for each IL-1 without any significant cross-reactivity (less than 0.01%). The intra-assay precision was also very good with a coefficient of variation of less than 10% over a wide range (between 3 and 500 pg/ml depending on the time devoted to the enzymatic reaction). The high sensitivity and precision of the assays can be ascribed to the high affinities of the mAbs as well as the optimal catalytic properties of AChE. The specificity of the determination performed in culture medium was demonstrated using different validation tests including a comparison with a bioassay and the fractionation of samples by molecular sieve chromatography. Evidence is presented that the assay could be used for the determination of IL-1 levels in biological media such as plasma or serum.

Acetylcholinesterase↗
PubMed1989

Isolation of dermenkephalin from amphibian skin, a high-affinity delta-selective opioid heptapeptide containing a D-amino acid residue.

The predicted amino acid sequence of the biosynthetic precursor of dermorphin, a highly potent and nearly specific mu-opioid peptide from amphibian skin, contains four repeats of the dermorphin progenitor sequence and one single copy of a different heptapeptide sequence. We have developed a specific enzyme immunoassay and used synthetic peptides to detect and purify the new predicted heptapeptide (2.4 micrograms/g dry skin) from the skin of the Phyllomedusa sauvagei frog from which dermorphin was originally isolated. The identity of the novel pro-dermorphin related peptide, Tyr-D-Met-Phe-His-Leu-Met-Asp-NH2, was established by co-chromatography with synthetic peptides on reverse-phase HPLC, immunological analysis, gas-phase sequencing, mass spectrometry and by pharmacological assays. Opioid-binding assays in vitro demonstrated that both the natural and synthetic heptapeptides displayed exceptionally high selectivity and affinity towards the delta-opioid receptors. Because of its origin and its delta-opioid (enkephalin) activity and specificity, this novel D-amino acid containing peptide is named dermenkephalin.

Acetylcholinesterase↗