Spleen antibacterial peptides: high levels of PR-39 and presence of two forms of NK-lysin.

Antibacterial peptides were isolated from porcine spleen by acetic acid extraction, ion exchange chromatography and reverse-phase high-performance liquid chromatography. C-terminal ladder sequence analysis of a bioactive peptide with matrix-assisted laser desorption/ionization mass spectrometry after digestion with carboxypeptidases P and Y showed that it is identical to the antibacterial proline/arginine-rich intestinal peptide PR-39. It is present at high levels in granulocytes of the spleen, and peptides with C-terminal proline amide and internal adjacent Pro residues can be analyzed with this method.

Large-scale HPLC purification of calbindin D9k from porcine intestine.

Two efficient procedures for large-scale purification of calbindin D9k from porcine intestine by HPLC were developed. Both protocols start with heat treatment of the intestinal tissue followed by acetic acid extraction, a capture with alginic acid, NaCl precipitation of other proteins, and a concentration step on Amberlite XAD-2. In the first method, a single reverse-phase HPLC step completes the purification and results in milligram quantities of pure calbindin.

A cytotoxic, apoptotic, low-molecular weight factor from pineal gland.

Previous studies suggest that the pineal gland may play a role in tumour growth inhibition. In this respect, melatonin, as the major hormone of this gland, has been extensively studied. However, there is growing evidence for the existence of other yet unknown pineal factors that may have tumour growth inhibiting properties.

Isolation and characterization of sulphated and nonsulphated forms of cholecystokinin-58 and their action on gallbladder contraction.

Cholecystokinin (CCK) exists in multiple molecular forms with different polypeptide lengths and the absence or presence of sulphation. We have isolated sulphated and nonsulphated forms of CCK-58 from porcine intestine and have determined their bioactivities in a guinea-pig gallbladder contraction assay. Both forms co-eluted in cation-exchange chromatography and in several rounds of reverse-phase (RP)-HPLC, but separated upon RP-HPLC using a water/acetonitrile system with heptafluorobutyric acid as counter ion.

Modulating effects of sensory and autonomic neuropeptides on murine splenocyte proliferation and cytokine secretion induced by Leishmania major.

The intimate, bidirectional link between neuroendocrine and immune systems is now accepted. A modulating effect of the nervous system on immune and inflammatory responses has been corroborated by identification of neuropeptide receptors on immunocompetent cells and the finding that neuropeptides can regulate leukocyte functions. The present study was undertaken to investigate the possible immunomodulatory role of sensory (SOM, CGRP and SP) and autonomic (VIP and NPY) neuropeptides in a murine model of cutaneous leishmaniasis, using two genetically different inbred mouse strains, BALB/c and C57BL/6, respectively susceptible and resistant to Leishmania (L.

Inhibitory effect of vasoactive intestinal peptide on the challenge phase of allergic contact dermatitis in humans.

There is increasing evidence that the nervous system has influence on the immune response. The effect of vasoactive intestinal peptide (VIP) and of serotonin and its antagonists on the challenge phase of allergic contact dermatitis in humans were tested. The substances were injected intracutaneously shortly before and 6 h after application of patch tests with nickel sulphate in nickel-allergic patients and the test areas were measured after a further 18 h.

An endogenous peptide isolated from the gut, NK-lysin, stimulates insulin secretion without changes in cytosolic free Ca2+ concentration.

We have recently isolated and cloned a novel endogenous peptide from pig intestine, NK-lysin (NKL). In the present study we show that NKL (1-100 nM) potently and reversibly stimulates insulin secretion in rat pancreatic islets and in the beta-cell line HIT T15. This effect of NKL was not accompanied by changes in cytoplasmic free calcium concentration.

In vitro Leishmania major promastigote-induced macrophage migration is modulated by sensory and autonomic neuropeptides.

Recruitment, migration and adherence of macrophages and their interaction with inoculated promastigotes are key steps in the initiation of the inflammatory process in cutaneous leishmaniasis. Parasite- and nervous system-derived factors might be involved in this process. In the present study the chemotactic activities of live, killed and sonicated Leishmania major promastigotes and of the promastigote culture supernatant as well as the L.

Identification, isolation, and characterization of daintain (allograft inflammatory factor 1), a macrophage polypeptide with effects on insulin secretion and abundantly present in the pancreas of prediabetic BB rats.

A bioactive macrophage factor, the polypeptide daintain/allograft inflammatory factor 1 (AIF1), has been isolated from porcine intestine. It was discovered when searching for intestinal peptides with effects on insulin release, and its purification was monitored by the influence of the peptide fractions on pancreatic glucose-induced insulin secretion. Daintain/AIF1 is a 146-aa residue polypeptide with a mass of 16,603 Da and an acetylated N terminus.

Characterization of dopuin, a polypeptide with special residue distributions.

A 62-residue polypeptide, dopuin, has been isolated from pig small intestine. It is distinguished by an N-terminal part with a high content of proline (7 in a 26-residue segment), a C-terminal part with a high proportion of histidine (3 in a 9-residue segment), and six half-cystine residues in three intrachain disulphide bridges (connecting positions 22-25, 23-54 and 35-44). The Cys and Pro distributions suggest a tight and special conformation.

Full-length and N-terminally truncated chicken intestinal diazepam-binding inhibitor. Purification, structural characterization and influence on insulin release.

Two forms of diazepam-binding inhibitor (DBI) have been purified from chicken intestine and identified as the intact avian polypeptide (residues 1-86) and a truncated variant (residues 35-86). At 10 nM concentration, both the intact and the truncated peptide suppress in vitro-monitored glucose-induced insulin release by 50 (p < 0.02) and 64% (p < 0.

Vasoactive intestinal polypeptide inhibits the established allergic contact dermatitis in humans.

There is increasing evidence indicating that the nervous system influences the immune response. In the present study the potential immunomodulatory role of vasoactive intestinal polypeptide (VIP) in established allergic contact dermatitis in humans was investigated. Positive patch-test reactions were elicited by application of nickel sulphate for 48 h.

Inhibitory effect of vasoactive intestinal polypeptide and ketanserin on established allergic contact dermatitis in man.

Neuromediators may influence the immune response. To investigate their potential immunomodulating role in established allergic contact dermatitis in man, the following neuromediators were tested: vasoactive intestinal polypeptide (VIP), serotonin, and the serotonin antagonists ketanserin, methiotepine and ICS-205-930. Positive patch test reactions were elicited by application of nickel sulphate for 48 h.

The secretory trypsin inhibitor like-peptide, PEC-60 increases dopamine D2 receptor agonist induced inhibition of GABA release in the dorsolateral neostriatum of the awake freely moving rat. An in vivo microdialysis study.

The effects of local perfusion with the secretory trypsin inhibitor like-peptide, PEC-60 on dopamine and gamma-aminobutyric acid (GABA) release in the dorsolateral neostriatum and GABA release in the globus pallidus were studied using in vivo microdialysis in the awake freely moving rat. Local perfusion with PEC-60 (500 nM and 1 microM) increased dopamine release in the dorsolateral neostriatum while the highest (1 microM) concentration of PEC-60 decreased striatal but not pallidal GABA release. An inactive form of the peptide, S-carboxyamidomethylated PEC-60 (1 microM) failed to influence either striatal dopamine and GABA or pallidal GABA release.

Endosulfine, endogenous ligand for the sulphonylurea receptor: isolation from porcine brain and partial structural determination of the alpha form.

Anti-diabetic sulphonylureas act via high affinity binding sites coupled to K-ATP channels. Endosulfine, an endogenous ligand for these binding sites, was shown to exist in two molecular forms, alpha and beta, in both the pancreas and the central nervous system. We describe here the isolation, and partial structural characterization of alpha endosulfine derived from porcine brains by means of a series of chromatography runs and gel electrophoresis.

Two alternative processing pathways for a preprohormone: a bioactive form of secretin.

An N-terminally 9-residue elongated form of secretin, secretin-(-9 to 27) amide, was isolated from porcine intestinal tissue and characterized. Current knowledge about peptide processing sites does not allow unambiguous prediction of the signal peptide cleavage site in preprosecretin but suggests cleavage in the region of residues -10 to -14 counted upstream from the N terminus of the hormone. However, the structure of the isolated peptide suggests that the cleavage between the signal peptide and the N-terminal propeptide occurs at the C-terminal side of residue -10.

Rescue of thymocytes from cell death by vasoactive intestinal peptide.

Vasoactive intestinal peptide (VIP) has previously been shown to increase survival of cultured neurons and to prevent the neurotoxic effect of the envelope glycoprotein 120 of human immune deficiency virus (HIV). The present report shows that VIP also protects mouse and human thymocytes exposed to a cytolytic dose of prednisolone in vitro. The activity of VIP is dose-dependent, and specific, since the structurally related secretin has no effect.

NK-lysin, a novel effector peptide of cytotoxic T and NK cells. Structure and cDNA cloning of the porcine form, induction by interleukin 2, antibacterial and antitumour activity.

A 78 residue antimicrobial, basic peptide, NK-lysin, with three intrachain disulfide bonds was purified from pig small intestine and characterized. A corresponding clone was isolated from a porcine bone marrow cDNA library. The 780 bp DNA sequence had a reading frame of 129 amino acids which corresponded to NK-lysin.

PEC-60 increases dopamine but not GABA release in the dorsolateral neostriatum of the halothane anaesthetized rat. An in vivo microdialysis study.

The effect of striatal perfusion with the intestinal peptide PEC-60 on endogenous dopamine (DA) and gamma-aminobutyric acid (GABA) release in the dorsolateral striatum and GABA release in the globus pallidus was monitored using in vivo microdialysis in the halothane anaesthetized rat. The results show that PEC-60 (100 nM) increases DA release in the dorsolateral striatum without influencing GABA release in the dorsolateral striatum or in the globus pallidus. In addition, PEC-60 failed to influence the extracellular striatal 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) levels.

Solution structure and dynamics of PEC-60, a protein of the Kazal type inhibitor family, determined by nuclear magnetic resonance spectroscopy.

The three-dimensional solution structure of porcine PEC-60, a 60 amino acid residue protein of the Kazal type family of proteinase inhibitors, was determined by nuclear magnetic resonance (NMR) spectroscopy. The structure determination is based on nearly complete 1H, 13C and 15N resonance assignments including stereospecific 1H resonance assignments for 40 pairs of methylene protons and isopropyl methyl groups. The stereospecific resonance assignments of the beta-protons were supported by heteronuclear long-range correlation experiments recorded at natural 13C and 15N isotopic abundances.

Activation of Na,K-ATPase by an endogenous peptide, PEC-60.

Several peptidic and non-peptidic factors can modulate Na,K-ATPase activity, among them mainly inhibitors of this enzyme, ouabain being the most effective. In a very few cases only, activation of Na,K-ATPase by endogenous factors has been recorded. We have investigated the effect on Na,K-ATPase of a novel regulatory peptide, PEC-60, recently isolated from porcine intestine.

A porcine gut polypeptide identical to the pancreatic hormone PP (pancreatic polypeptide).

A peptide hormone has been isolated from porcine intestine. Its primary structure was found to consist of 36 amino acid residues in a sequence identical to that of the porcine pancreatic polypeptide, previously not isolated from intestines or a tissue other than pancreas. The gut polypeptide significantly suppresses glucose-induced insulin secretion in vitro.

Characterization of porcine intestinal cytochrome c oxidase subunit VIIc, purified by affinity chromatography.

Cytochrome c oxidase subunit VIIc was isolated from porcine intestine. Its amino acid sequence, starting at position 17 of the predicted precursor peptide, differs at eight positions from the human form, two positions from the bovine and at one from the mouse form. Although the peptide does not contain cysteine, it was bound specifically to thiopropyl-Sepharose 6B.

Isolation of three antibacterial peptides from pig intestine: gastric inhibitory polypeptide (7-42), diazepam-binding inhibitor (32-86) and a novel factor, peptide 3910.

Two antibacterial peptides, cecropin P1 and PR-39 (39-residue proline/arginine-rich peptide), from the upper part of pig small intestine have previously been isolated and characterized. We have now continued our search for antibacterial peptides in different side fractions generated during the isolation of intestinal hormones. Starting from one such fraction and monitoring activity against Bacillus megaterium, we isolated three homogeneous peptides by three consecutive chromatographic steps.