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1.
Origin of bombesin-like peptides in human fetal lung 总被引:2,自引:0,他引:2
Four different forms of bombesin-like immunoreactive peaks were detected in extracts of human fetal lung by the use of reversed-phase high performance liquid chromatography (HPLC). Peaks I, II, III and IV, (increasing retention time), were eluted using a 14-38% of acetonitrile gradient containing 0.1% trifluoroacetic acid (TFA). Peak II was the major material found in the extract of human fetal lung obtained at 16-20 weeks gestation. None of the four compounds contained in the eluted peaks had the same retention time as amphibian bombesin or porcine gastrin releasing peptide (GRP). On reversed-phase HPLC using two different solvent systems TFA or heptafluorobutyric acid (HFBA) as a hydrophobic counter ion, and in gel filtration chromatography, the chromatographic behavior of the main peak (peak II) was the same as that of the carboxyl terminal fragments of GRP, GRP18-27 or GRP19-27. This suggested that the peptide(s) in peak II resembled in composition the carboxy terminal 9 or 10 amino acids of porcine GRP. Following tryptic digestion the material in peak IV was converted to the more polar compound present in peak II. Two other peptide peaks were eluted close to peak II and these were presumed to be a modification of this main peak. One of the possible biosynthetic steps in the formation of bombesin-like peptides in human fetal lung could be a tryptic conversion of a less polar peptide to a more polar form (peak IV to II). 相似文献
2.
Distribution of bombesin-like peptides in the alimentary canal of several vertebrate species 总被引:2,自引:0,他引:2
G H Greeley M Partin A Spannagel T Dinh F L Hill J Trowbridge M Salter H F Chuo J C Thompson 《Regulatory peptides》1986,16(2):169-181
The objective of this study was to quantitate and characterize the variants of bombesin-like immunoreactivity in the alimentary canal of the rat, rabbit, hawk, owl, dog, monkey and human. Bombesin-like immunoreactivity was found throughout the entire gastrointestinal tract of all species studied. In the rat, the highest concentration of bombesin-like immunoreactivity was found in the colon. Gel chromatography showed that bombesin-like immunoreactivity corresponded to gastrin-releasing peptide (GRP-27) and GRP-10. In the dog, the greatest concentration of bombesin-like immunoreactivity was observed in the mucosal layer of the fundus, whereas the concentration of bombesin-like immunoreactivity in the muscle layer of the dog did not vary significantly from region to region. Gel chromatography showed that bombesin-like immunoreactivity in the dog corresponded to GRP-27, bombesin, GRP-10, and a smaller fragment. In the human, the concentration of bombesin-like immunoreactivity did not vary significantly from region to region in the mucosal and muscular layers. Gel chromatography of human fundal mucosa showed that bombesin-like immunoreactivity peaks occur in the regions of GRP-27, bombesin and GRP-10. These findings substantiate the observation that bombesin-like peptides play a variety of roles in the regulation of gut function. 相似文献
3.
A sensitive radioimmunoassay for bombesin-like immunoreactivity (BLI) was developed and utilised in conjunction with G50 gel chromatography and reverse-phase HPLC, to study the content and molecular characteristics of bombesin-like peptides in acid extracts of human fetal lung. The antiserum, (B5), is directed towards the C-terminal region of the bombesin molecule and cross-reacts 70% with synthetic porcine GRP and the synthetic GRP fragment, GRP (14-27). Specimens of lung were collected from fetuses of gestational ages 15-22 weeks, following prostaglandin termination of pregnancy. The tissue was extracted into 0.1 N HCl at 90 degrees C. The mean BLI content was 50.2 pg/mg wet weight of tissue (range 15.5-136 pg/mg; n = 13). No correlation between gestational age and BLI content could be established. G50 gel chromatography of acid extracts, under dissociating conditions, revealed two peaks of BLI, one in the position of synthetic porcine GRP and the second, constituting greater than 90% of the immunoreactivity, eluting with synthetic amphibian bombesin. Reverse-phase ODS silica HPLC of this major G50 peak, utilising a methanol/trifluoroacetic acid gradient, indicated that this peptide was similar to the GRP C-terminal fragment, GRP (14-27). We have therefore (1) confirmed the presence and heterogeneity of BLI in human fetal lung, and (2) shown, for the first time, that the majority of this BLI more closely resembles a fragment of GRP than amphibian bombesin itself. 相似文献
4.
cDNA cloning, characterization, and brain region-specific expression of a neuromedin-B-preferring bombesin receptor. 总被引:7,自引:0,他引:7
E Wada J Way H Shapira K Kusano A M Lebacq-Verheyden D Coy R Jensen J Battery 《Neuron》1991,6(3):421-430
Recent binding studies in the central nervous system and other tissues provide evidence that the mammalian bombesin-like peptides, gastrin-releasing peptide (GRP) and neuromedin-B (NMB), exert their numerous physiological effects through at least two different receptors. We describe the structure and expression of a cloned NMB-preferring bombesin receptor (NMB-R) with properties distinct from a GRP-preferring bombesin receptor (GRP-R) reported previously. In particular, the NMB-R shows higher affinity binding to NMB than to GRP in BALB 3T3 fibroblasts expressing the cloned NMB-R. The distinct regional distribution of NMB-R and GRP-R mRNA in the brain suggests that both bombesin receptor subtypes play independent roles in mediating many of the dramatic effects of bombesin-like peptides in the central nervous system. 相似文献
5.
Bombesin-like peptides in small cell lung cancer: biochemical characterization and secretion from a cell line 总被引:7,自引:0,他引:7
High intracellular levels of BN-like peptides are present in tumors and cell lines of small cell carcinoma of the lung (SCCL) as well as the putative precursor cells of this tumor, the pulmonary endocrine cell. In cell line NCI-H209 the density of bombesin-like peptides was 8.9 +/- 1.1 pmol/mg total protein. Gel filtration chromatography of an extract of these cells revealed one major peak of immunoreactivity which coeluted with synthetic bombesin (1620 daltons). Also, high pressure liquid chromatography revealed one major peak of immunoreactivity was present which eluted before synthetic peptide. Therefore, SCCL bombesin-like peptides may be of similar size but are more hydrophilic than synthetic peptide. Cells maintained in culture continuously release bombesin-like peptides into the growth medium. Also, high concentrations of K+ stimulated the secretion of immunoreactive bombesin from cell lines in a Ca++-dependent manner. These SCCL bombesin-like peptides may function as important regulatory agents in the malignant lung. 相似文献
6.
Bombesin-like immunoreactivity in human gastrointestinal tract 总被引:3,自引:0,他引:3
In the present study the distribution and molecular characteristics of bombesin-like immunoreactivity (BLI) were studied in acid extracts of human gastrointestinal tract. The highest levels were found in the fundus, antrum, pylorus and pancreas with lower levels in the duodenum, jejunum, terminal ileum and colon. BLI was also detected in both the muscle and mucosal layers of the antrum and colon. Sephadex G-50 gel chromatography under acid dissociating conditions revealed two peaks of immunoreactivity, one in the position of synthetic porcine gastrin releasing peptide (GRP) and the second eluting with synthetic amphibian bombesin. Variations in the proportions of the two molecular forms were seen in different regions of the gut. In the stomach and pancreas greater than 70% of the BLI eluted with the GRP marker while in pylorus, jejunum and terminal ileum only 20% was present in this form. Reverse-phase ODS silica HPLC of the major antral BLI peak, utilising a methanol/trifluoroacetic acid gradient indicated that this peptide was similar to porcine GRP. We have therefore (1) demonstrated the presence and heterogeneity of bombesin-like immunoreactivity throughout the human gastrointestinal tract and (2) shown for the first time that a proportion of this BLI closely resembles porcine GRP. 相似文献
7.
Gu Q Lee LY 《American journal of physiology. Lung cellular and molecular physiology》2005,289(6):L1104-L1112
Small cell lung cancer (SCLC) patients suffer from pulmonary stresses such as dyspnea and chest pain, and the pathogenic mechanisms are not known. SCLC cells secrete a variety of bioactive neuropeptides, including bombesin-like peptides. We hypothesize that these peptides may enhance the sensitivity of the pulmonary chemosensitive nerve endings, contributing to the development of these pulmonary stresses in SCLC patients. This study was therefore carried out to determine the effects of bombesin and gastrin-releasing peptide (GRP), a major bombesin-like peptide, on the sensitivities of pulmonary chemoreflex and isolated pulmonary vagal chemosensitive neurons. In anesthetized, spontaneously breathing rats, intravenous infusion of bombesin or GRP significantly amplified the pulmonary chemoreflex responses to chemical stimulants such as capsaicin and ATP. The enhanced responses were completely abolished by perineural capsaicin treatment of both cervical vagi, suggesting the involvement of pulmonary C-fiber afferents. In isolated pulmonary vagal chemosensitive neurons, pretreatment with bombesin or GRP potentiated the capsaicin-induced Ca(2+) transient. This sensitizing effect was further demonstrated in patch-clamp recording studies; the sensitivities of these neurons to both chemical (capsaicin and ATP) and electrical stimuli were significantly enhanced by the presence of either bombesin or GRP. In summary, our results have demonstrated that bombesin and GRP upregulate the pulmonary chemoreflex sensitivity in vivo and the excitability of isolated pulmonary chemosensitive neurons in vitro. 相似文献
8.
T W Moody A Murphy S Mahmoud G Fiskum 《Biochemical and biophysical research communications》1987,147(1):189-195
The ability of bombesin-like peptides to elevate intracellular Ca2+ levels in small cell lung cancer cells was investigated using the fluorescent Ca2+ indicator Fura 2. Nanomolar concentrations of bombesin elevated cytosolic Ca2+ levels in the absence or presence of extracellular Ca2+. Potent bombesin receptor agonists, such as gastrin releasing peptide (GRP) or (GRP)14-27 elevated cytosolic Ca2+ levels whereas inactive compounds such as (D-Trp8)bombesin or (GRP)1-16 did not. Furthermore, the bombesin receptor antagonist (D-Arg1, D-Pro2, D-Trp7,9, Leu11) substance P (30 microM) had no effect on the Ca2+ levels by itself but antagonized the increase in Ca2+ caused by 10 nM or 100 nM bombesin. These data suggest that bombesin receptors may regulate the release of Ca2+ from intracellular organelles in small cell lung cancer cells. 相似文献
9.
Internalization and degradation of peptides of the bombesin family in Swiss 3T3 cells occurs without ligand-induced receptor down-regulation. 总被引:3,自引:1,他引:2
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The binding of [125I]gastrin releasing peptide ([125I]GRP) to Swiss 3T3 cells at 37 degrees C increases rapidly, reaching a maximum after 30 min and decreasing afterwards. The decrease in cell-associated radioactivity at this temperature is accompanied by extensive degradation of the labelled peptide. At 4 degrees C equilibrium binding is achieved after 6 h and [125I]GRP degradation is markedly inhibited. Extraction of surface-bound ligand at low pH demonstrates that the iodinated peptide is internalized within minutes after addition to 3T3 cells at 37 degrees C. The rate of internalization is strikingly temperature-dependent and is virtually abolished at 4 degrees C. In addition, lysomotropic agents including chloroquine increase the cell-associated radioactivity in cells incubated with [125I]GRP. The binding of [125I]GRP to Swiss 3T3 cells was not affected by pretreatment for up to 24 h with either GRP or bombesin at mitogenic concentrations. Furthermore, pretreatment with GRP did not reduce the affinity labelling of a Mr 75,000-85,000 surface protein recently identified as a putative receptor for bombesin-like peptides. These results demonstrate that while peptides of the bombesin family are rapidly internalized and degraded by Swiss 3T3 cells, the cell surface receptors for these molecules are not down-regulated. 相似文献
10.
The prolactin (PRL)-releasing activity (PRA) in the bovine hypothalamic extract (BHE) was compared to that of known substances with PRA and further characterized by gel filtration and reversed-phase high performance liquid chromatography (RP-HPLC). Crude BHE produced marked dose-dependent stimulation of PRL secretion from the cultured rat adenohypophysial cells. Among the synthetic substances examined, vasoactive intestinal peptide (VIP), thyrotropin-releasing hormone (TRH) and beta-endorphin (END) showed significant PRA. However, the flatter dose-response slope for TRH compared with BHE or the small amounts of VIP and END in BHE suggested that these peptides could not account for the major active elements of BHE. Oxytocin and interleukin-1beta were also tested, but they exhibited no PRA in our assay system. Gel filtration of BHE on the Sephadex G-100 column yielded two peaks of PRA distinct from TRH, VIP and END. One eluted in the void and the other in more retarded fractions. The latter fractions were pooled and subjected to the two-step RP-HPLC. The PRA was separated into three peaks designated peaks I, II and III in the first RP-HPLC experiment. Furthermore, the second RP-HPLCs with finer resolution revealed that peak II as well as peak III consisted of three peaks, while peak I eluted as a single peak. Most of these seven PRA peaks exhibited different RP-HPLC profiles from those of the newly characterized PRL-releasing peptides. These findings again provide confirmatory evidence that BHE contained unique factors different from the above known substances. 相似文献
11.
Leu13-psi(CH2NH)Leu14]bombesin is a specific bombesin receptor antagonist in Swiss 3T3 cells 总被引:3,自引:0,他引:3
P J Woll D H Coy E Rozengurt 《Biochemical and biophysical research communications》1988,155(1):359-365
The pseudopeptide [Leu13-psi(CH2NH)Leu14]bombesin blocks bombesin-stimulated mitogenesis in Swiss 3T3 cells in a competitive and reversible manner, but not that of other mitogens. It inhibits the mobilization of intracellular Ca2+ and activation of protein kinase C by bombesin-like peptides. It acts at receptor level, as shown by inhibition of [125I]GRP binding and reduction in cross-linking of the Mr 75-85,000 receptor-associated protein. Thus [Leu13-psi(CH2NH)Leu14]bombesin is a specific bombesin receptor antagonist in Swiss 3T3 cells which blocks long-term growth promoting effects of bombesin-like peptides. 相似文献
12.
Ohki-Hamazaki H Iwabuchi M Maekawa F 《The International journal of developmental biology》2005,49(2-3):293-300
Amphibian bombesin and its related peptides consist a family of neuropeptides in many vertebrate species. Bombesin and two major bombesin-like peptide in mammals, gastrin-releasing peptide (GRP) and neuromedin B (NMB), have been shown to elicit various physiological effects. These include inhibition of feeding, smooth muscle contraction, exocrine and endocrine secretions, thermoregulation, blood pressure and sucrose regulations and cell growth. Receptors for GRP and NMB (GRP-R and NMB-R), as well as third subtype of bombesin-like peptide receptor (BRS-3) have been cloned. These receptors are G-protein-coupled receptors and are expressed in various brain regions and in the digestive tract. In this paper, we will summarize studies on these peptides and their receptors, with special reference to research using gene-knockout mice. These studies clearly demonstrated the role of three receptors in vivo and in vitro. We will also discuss the phylogeny of these receptors. 相似文献
13.
Immunohistochemical studies have established that fish gastrointestinal tissues contain peptides with gastrin-releasing peptide (GRP)/bombesin-like immunoreactivity, but the molecular nature of this material is unclear. In this study, the most abundant peptide that was immunoreactive towards an antiserum raised against pig GRP was isolated in pure form from an extract of the stomach of the rainbow trout (Oncorhynchus mykiss). The primary structure of the peptide was established as: Ser-Glu-Asn-Thr-Gly-Ala-Ile-Gly-Lys-Val10- Phe-Pro-Arg-Gly-Asn-His-Trp-Ala-Val-Gly20-His-Leu-Met-NH2. Although this amino acid sequence is shorter than those of mammalian GRPs by four residues, the COOH-terminal dodecapeptide is identical to the corresponding region in pig GRP. The data indicate, therefore, that the predominant molecular form of GRP in the stomach of a teleost fish is structurally more similar to mammalian GRP than to the amphibian skin peptide, bombesin. 相似文献
14.
A Hernanz 《Biochimie et biologie cellulaire》1990,68(9):1142-1145
Extracts of rat brain and gastrointestinal tract, analyzed by reverse-phase high-performance liquid chromatography and radioimmunoassay, contained two bombesin-like immunoreactivity peaks with similar retention times as porcine gastrin-releasing peptide (GRP) and its COOH-terminal decapeptide, neuromedin C or GRP(18-27). However, the GRP-like peptide peak did not elute with exactly the same retention time as porcine GRP. The highest concentration of bombesin-like immunoreactivity was found in extracts of antrum, whereas the lowest was found in whole brain. Neuromedin C was present at lower concentrations than the GRP in antrum, duodenum, and ileum, while similar amounts of each were found in brain. 相似文献
15.
Enzymatic cleavage prior to antibody incubation as a method for neuropeptide immunocytochemistry 总被引:4,自引:0,他引:4
Summary Five anti-gastrin releasing peptide (GRP) sera have been characterized against GRP, bombesin and related polypeptides spotted on cellulose acetate discs. Antibodies reacting with the C-terminal G-14 sequence of bombesin and the 19–27 sequence of GRP, were detected in all sera. Antibodies directed exclsively against the bombesin unrelated 1–17 sequence of GRP were found only in one serum (R-6902). With parallel immunohistochemical tests only the C-terminal immunoreactivity was detected in endocrine-paracrine cells of the chicken proventriculus, while both immunoreactivities were present in nerve fibres and a few nerve cell bodies of the mammalian gut. The distribution of GRP- and bombesin-like immunoreactive nerves in the gastric mucosa of both pyloric and oxyntic type the submucosal and myenteric plexus along the whole gastrointestinal wall and at sphincter regions is detailed. 相似文献
16.
M C Thorndyke J R Reeve S R Vigna 《Comp. Biochem. Physiol. C, Comp. Pharmacol. Toxicol.》1990,96(1):135-140
1. Ratfish (Hydrolagus colliei) intestines were boiled in water to inactivate proteases and then treated with cold 4% trifluoroacetic acid to extract bombesin-like peptides. 2. The extract was fractionated in several steps using reverse-phase and ion exchange HPLC, and bombesin-like immunoreactive peptides were detected by radioimmunoassay using an antiserum specific for the bioactive C-terminal region of bombesin. 3. A highly purified bombesin-like peptide-containing fraction stimulated amylase release in a dose-responsive fashion from rat pancreatic acini; the dose-response curve was parallel to a bombesin standard, and the ratfish peptide stimulated the same maximal rate of amylase secretion as the bombesin standard. 4. A potent, highly selective bombesin receptor antagonist completely abolished the stimulation of amylase release caused by the ratfish peptide, demonstrating the specificity of the response. 5. Estimates of the bombesin-like peptide concentration of this fraction by radioimmunoassay and by bioassay were nearly identical, indicating that ratfish bombesin is very similar biologically and antigenically to frog skin bombesin. 相似文献
17.
S R Nagalla B W Gibson D Tang J R Reeve E R Spindel 《The Journal of biological chemistry》1992,267(10):6916-6922
On the basis of structural homology and similar biological activity, gastrin-releasing peptide (GRP) has been considered the mammalian equivalent of amphibian bombesin. In this paper we now show this to be incorrect. Chromatography of frog (Bombina orientalis) gut extracts demonstrated two peaks of bombesin-like immunoreactivity (BLI), one similar in size to GRP and one similar in size to amphibian bombesin. These peaks were purified by high pressure liquid chromatography then subjected to mass spectrometric analyses to determine molecular weights and amino acid sequence. Based on the amino acid sequence of the lower molecular weight BLI species, a mixed oligonucleotide probe was prepared and used to screen a B. orientalis stomach cDNA library. Sequence analysis showed that all hybridizing clones encoded a 155-amino acid protein homologous to the mammalian GRP precursor. The mass spectra of the high and low molecular weight peaks of frog gut BLI were consistent with their origin from the processing of the frog GRP (fGRP) precursor into GRP-29 and GRP-10, just like the processing of the rat GRP precursor. Sequence homology showed that the fGRP precursor is more homology showed that the fGRP precursor is more closely related to the mammalian GRP precursors than to either the frog bombesin or frog ranatensin precursors. Northern blot analysis showed that fGRP is encoded by a mRNA of 980 bases, clearly different from the 750-base mRNA which encodes frog bombesin. Northern blot analysis and in situ hybridization showed fGRP mRNA in frog brain and stomach and bombesin mRNA in frog skin, brain, and stomach. That frogs have independent genes for both GRP and bombesin raises the possibility that mammals have an as yet uncharacterized gene encoding a true mammalian bombesin. 相似文献
18.
Gastrin-releasing peptide (GRP) is a member of bombesin-like peptides and bombesin and neuromedin B are other members of this family. They act on receptors that belong to the GPCR superfamily and exert important physiological functions upon binding to their receptors. The biologically active C-terminal decapeptide of GRP (GRP10) was studied in explicit DMPC bilayers using molecular dynamics simulations. In the initial conformation, the peptide was placed perpendicular to the membrane plane and the peptide-membrane complex with approximately 20,000 atoms was simulated for a period of 8 ns. After a 5 ns simulation, GRP10 adopted a tilted orientation and the tilt angle with respect to the bilayer normal was approximately 60 masculine. Analysis of the interactions of individual residues indicated the role of histidine residues in maintaining a tilted orientation. 相似文献
19.
Dual effect of bombesin and gastrin releasing peptide on gastric emptying in conscious cats 总被引:1,自引:0,他引:1
The effect of bombesin (BBS) and gastrin releasing peptide (GRP) on gastric emptying was studied in conscious cats. This effect was measured simultaneously with antral motility. Acid and pepsin secretions as well as blood hormonal peptide release were additionally measured. A dual effect was observed. First, BBS and GRP slowed gastric emptying of liquids, while antral motility was decreased, then after 60 minutes of continuous intravenous infusion, antral motility returned to basal values and gastric emptying effect reversed. The mechanism of this peculiar action is independent of gastrin, pancreatic polypeptide, somatostatin and motilin release and most probably connected with a cholinergic stimulation induced by the peptides, the late predominance of which counterbalances the inhibitory effect of bombesin-like peptides on antral motility. 相似文献
20.
The occurrence of neuropeptides in the brain of larvae of the tobacco hawkmoth, Manduca sexta, and tomato moth, Lacanobia oleracea, was investigated using matrix-assisted laser desorption ionisation-time of flight (MALDI-TOF) mass spectrometry (MS) and post-source decay (PSD). Methanolic extracts of 100 brains separated by reversed-phase high performance liquid chromatography yielded numerous ion peaks, some of which were common to both species. In M. sexta six [M+H](+) ions were in agreement with peptides previously structurally characterised from M. sexta (FLRF-amides I, II and III, M. sexta allatostatin, CAP(2b) and myoinhibitory peptide VI), whereas a further five corresponded to other known lepidopteran peptides (cydiastatins 3 and 4, helicostatins 1 and 6 and helicokinin II). Of these the identities of FLRF-amide I, cydiastatins 3 and 4 and CAP(2b) were confirmed by PSD analysis. Fourteen [M+H](+) ions corresponding to known lepidopteran peptides (FLRF-amide I, cydiastatins 2, 3 and 4, helicostatins 1, 5, 6, 7 and 9, CCAP, CAP(2b), M. sexta allatostatin and myoinhibitory peptide VI) were measured in L. oleracea brain extracts. From this insect, cydiastatins 3 and 4, helicostatin 5 and FLRF-amide I were identified by PSD. These peptides had not previously been structurally characterised from L. oleracea. 相似文献