Elevated serum levels of calcitonin gene-related peptide (CGRP) but no evidence for CGRP gene expression in non-small cell lung carcinomas.

Calcitonin gene-related peptide (CGRP) is a hormone formed by alternative post-transcriptional processing of the calcitonin gene. It is a neuropeptide localized to discrete regions of the central nervous system (CNS) and in nerve fibres associated with blood vessels. It is also expressed in medullary carcinomas of the thyroid and lung carcinoma cell lines. The latter finding suggests a possible value for CGRP as tumour marker in lung carcinomas. In this investigation of 22 patients undergoing operation for lung tumours, pre and post-operative levels of serum CGRP were measured. Preoperative as well as postoperative serum CGRP levels were significantly elevated when compared to age-matched normals. However, no evidence could be found for CGRP gene expression in tumour tissue from the same patients as judged by immunocytochemistry or in-situ hybridization using CGRP cRNA probes. CGRP has been localized to nerve fibres in relation to pulmonary blood vessels and has been shown to be a potent vasodilator. These findings, and the absence of evidence for synthesis in tumours, as opposed to cell lines derived from lung carcinomas, suggests that the lack of post-operative normalization of serum CGRP concentrations may be related to physiological changes in cardiovascular haemodynamics following surgery. Elevated pre-operative serum CGRP levels may also reflect a consequence of the lung carcinoma leading to increased release of CGRP from sites in the vasculature yet to be determined, but does not indicate synthesis de novo and secretion of CGRP by the tumours.

Arginine vasopressin and cyclic adenosine monophosphate during acute sodium loading in chronic glomerulonephritis.

The relationship of arginine vasopressin (AVP) in plasma to cyclic adenosine 3' 5'-monophosphate (cAMP), sodium excretion in urine, and arterial blood pressure were determined during intravenous infusion of hypertonic sodium chloride solution (500 ml of 50 g/l) in 10 normotensive control subjects and in 11 normotensive and 10 hypertensive patients with chronic glomerulonephritis and relatively well preserved kidney function. The concentration of AVP in plasma increased 2-4 fold, osmolality in serum increased 12-16 mosmol/kg, and urinary excretion of cAMP increased 20-40% during sodium loading to the same extent in all three groups. Sodium and water excretion were higher during the sodium loading in the hypertensive patients, but not in the normotensive patients when compared to the control subjects. Neither AVP nor changes in AVP correlated significantly with changes in cAMP excretion, sodium excretion or blood pressure. In the control subjects the level of parathyroid hormone in serum was unchanged during the sodium chloride infusion. Water loading without sodium loading in eight of the control subjects caused a decrease in the excretion of cAMP. In conclusion, the increase in cAMP excretion in urine during the sodium loading might be explained by an AVP-induced stimulation of renal cAMP production. The study does not suggest that AVP plays a role in the increased sodium excretion during sodium loading or in the development of hypertension or chronic glomerulonephritis.

In vivo synthesis of cholecystokinin in the rat cerebral cortex: identification of COOH-terminal peptides with labeled amino acids.

The synthesis of the COOH-terminal octa- and tetrapeptides of cholecystokinin (CCK) has been studied in rat cerebral cortex after intraventricular administration of radioactive amino acids characteristic of the porcine COOH-terminal octapeptide of CCK, CCK-8. After immunosorption with a COOH-terminal directed antibody, cortical CCK was fractionated on Sephadex G-50 columns. The experiments demonstrated newly synthesized CCK forms which coeluted with porcine CCK-8 and CCK-4. Except for threonine the amino acids employed, methionine, tryptophan, aspartic acid, glycine and phenylalanine were incorporated. The sequence-specific radioimmunoassay, the incorporation of the employed labeled amino acids, and the elution pattern by gel filtration, suggest an almost identical structure of porcine and rat cortical CCK-8, and a concomitant synthesis of CCK-8 and CCK-4 in rat cerebral cortex.

In vivo biosynthesis of cholecystokinin in hog cerebral cortex.

The biosynthesis of cholecystokinin (CCK) in the cerebral cortex of hogs was studied by intracisternal injections of [32S]methionine. At different times (15, 60 and 120 min) after the injection, cortex was isolated and extracted with boiling water and 0.5 M acetic acid. CCK in the extracts was immunosorbed, using an antiserum specific for the COOH-terminal sequence of CCK. Subsequently, the CCK-immunoreactivity was applied to Sephadex G-50 superfine columns. The fractionation showed incorporation in five molecular forms with elution constants of 0.08, 0.50, 0.90. 1.1 and 1.3. After a pulse period of 15 min, [32S]methionine was incorporated mainly into the largest form of CCK (Kav of 0.08). The incorporation in all forms increased during the first hour. After 2 hours, a decline occurred in the larger forms, whereas the incorporation in the octapeptide form and tetrapeptide-like form increased.

Newly synthesized cholecystokinin in subcellular fractions of the rat brain.

The subcellular localization of in vivo synthesized cholecystokinin (CCK) in different parts of the rat brain was studied after intracisternal pulse injections of [35S]methionine. The rats were decapitated 1 h after the injection, and the brain was divided into cortex, hippocampus and remainder. Subcellular fractions were obtained according to Whittaker's method. De novo synthesized CCK in the crude mitochondrial-synaptosomal fraction, P2, and in the purified synaptosomal fraction was demonstrated by affinity chromatography, using antibodies specific for the COOH-terminal sequence of CCK. By subsequent gel chromatography two molecular forms of labelled CCK occurred, with elution constants, Kav, of 1.1 (corresponding to the COOH-terminal octapeptide) and of 1.40 (a component which may correspond to the COOH-terminal tetrapeptide amide, CCK-4). The findings support the idea that the small molecular forms are the transmitter forms of CCK.

Concentration and in vivo synthesis of cholecystokinin in subcortical regions of the rat brain.

The in vivo biosynthesis of cholecystokinin (CCK) in subcortical regions of the rat brain was studied by intracisternal pulse injections of [35S]methionine. The rats were killed 1.0 or 1.5 h after the injection. Subcortical CCK extracted in boiling water and acetic acid was immunoabsorbed by an antiserum specific for the COOH-terminal sequence of CCK. Gel chromatography of the absorbed CCK separated four molecular forms with elution constants (Kav) of 0.08, 0.50 (corresponding to the tritriacontapeptide amide, CCK-33), 1.10 (corresponding to the COOH-terminal octapeptide, CCK-8), and 1.40 (a component which may correspond to the COOH-terminal tetrapeptide amide). Significant incorporation of [35S]methionine occurred in the largest (Kav approximately 0.08) and octapeptide-like form only. Chasing with methionine indicated a precursor relationship between the largest form and CCK-8. The results demonstrate a substantial synthesis of CCK also in subcortical regions of the brain.

In vivo biosynthesis of cholecystokinin in rat cerebral cortex.

The biosynthesis of cholecystokinin (CCK) in the cerebral cortex of rats was studied by intracisternal pulse injections of [35S]methionine. The rats were killed with intervals varying from 15 min to 10 h after the injections. Cortical CCK extracted in boiling water and acetic acid was immunoabsorbed using an antiserum specific for the COOH-terminal sequence of CCK. After displacement from the immunoabsorbent by heptadecapeptide gastrin, which contains the same COOH-terminal sequence as CCK, chromatography on Sephadex G-50 columns showed four molecular forms of CCK with elution constants (Kav) of 0.08, 0.50 (corresponding to the tritriacontapeptide amide, CCK-33), 1.10 (corresponding to the COOH-terminal octapeptide amide, CCK-8) and 1.40 (a component which may correspond to the COOH-terminal tetrapeptide of rat CCK). Chasing with methionine demonstrated a biosynthetic pathway from the largest molecular form to the octapeptide-like form. The results indicate that a rapid and extensive synthesis of CCK takes place in the cerebral cortex, and that a precursor relationship exists between CCK-8 and the larger molecular forms of CCK.

Neural regulation of pancreatic hormone secretion by the C-terminal tetrapeptide of CCK.

In pancreatic islets a peptide corresponding to the C-terminal tetrapeptide amide of cholecystokinin and gastrin, Trp-Met-Asp-Phe-NH2, is present in nerve terminals. This tetrapeptide amide is uniquely potent as a releaser of insulin and the other islet hormones, whereas larger cholecystokinins and gastrins as well as tetrapeptide analogues are considerably less potent. We suggest that neural release of the tetrapeptide amide is implicated in regulation of pancreatic hormone secretion.

Somatostatin cell processes as pathways for paracrine secretion.

Somatostatin is produced by gastrointestinal endocrine cells that have long, nonluminal, cytoplasmic processes. Such processes terminate on other cell types, including gastrin-producing and hydrochloric acid-producing cells, whose functions are profoundly affected by somatostatin. The findings suggest that somatostatin cells control the functions of other cells through local release of the peptide by way of cytoplasmic processes. Also, certain other types of gastrointestinal endocrine cells have similar cytoplasmic prolongations, which may have analogous local (paracrine) regulatory functions.

Stimulation of gastric mucosal protein synthesis by different molecular forms of gastrin.

Protein synthesis in gastric mucosa was studied by measuring the incorporation of labeled amino acids into protein by isolated gastric mucosal ribosomes in a cell-free system. In 48-hour fasted rats, administration of the synthetic analogues pentagastrin, tetragastrin and gastrin-17 or naturally occurring molecular forms of human gastrin (G-14, G-34) markedly enhanced (23-123%) the capacity of the gastric mucosal ribosomes to synthesize endogenous mRNA-directed protein in a cell-free system. In the presence of exogenous mRNA (poly-U), the gastric mucosal ribosomes from the saline-treated controls showed a higher poly(U)-directed protein synthesis, compared to each fo the gastrin-treated groups. The protein/polyphenylalanine ratio which represents a ratio of polysomes to monosomes was found increased in ribosomes from the gastrin-treated groups.

Chronic administration of pentagastrin. Effects on pancreatic protein and nucleic acid contents and protein synthesis in rats.

Groups of rats were injected with either saline or pentagastrin (0.5, 1 or 2 mg/kg/day) for 14 days. Pancreatic weight, DNA, RNA and protein contents as well as the ability of the pancreatic polyribosomes to synthesize protein in a cell-free system were investigated. In all three pentagastrin-treated groups the weight of the pancreas was found to be significantly higher (33-73%) than in the control. DNA, RNA and protein contents were significantly increased by 47, 98 and 85%, respectively, in the group that received the highest dose of the hormone. With respect to ribosomal protein synthesis, while the pentagastrin dose of 1 and 2 mg/kg/day resulted in 25 and 72% augmentation, respectively, the dose of 0.5 mg/kg had no influence on protein synthesis. It is concluded that chronic administration of pentagastrin (2 mg/kg/day for 14 days) causes hyperplasia and hypertrophy of the pancreas.

Chronic administration of pentagastrin: effect on gastric mucosal protein synthesis in rats.

Groups of adult rats were injected with either saline or pentagastrin (500 microgramg/kg) for 14 days. The capacity of the gastric mucosal ribosomes to synthesize endogenous and exogenous mRNA-directed protein in a cell-free system was then investigated. Polyribosomal profiles were also analysed on sucrose gradients. The endogenous mRNA-directed incorporation of 14C-labelled amino acids into protein by the gastric mucosal ribosomes from the pentagrastrin-injected rats was found to be considerably greater than that of the control. In the presence of exogenous mRNA (poly-U) the polyribosomes as well as the endogenous mRNA-free ribosomes (run-off ribosomes) from the pentagastrin-treated group showed 86% and 136% increment in [14C]-phenylalanie incorporation as compared to the corresponding control preparation. The ribosomal ribonuclease activity between the groups was found to be the same. The results of the present investigation indicate that an enhancement in the translational capacity of the ribosomes is in part responsible for stimulation of gastric mucosal protein synthesis after chronic administration of pentagastrin.

Gastrin in the human fetus. Distribution and molecular forms of gastrin in the antro-pyloric gland area, duodenum and pancreas.

The ontogeny of human fetal gastrin has been investigated by immunocytochemistry and radioimmunochemistry. Gastrin and gastrin cells are shown to appear later in the antral than in the duodenal mucosa. At the ages studied (11--22 weeks of gestation) pancreatic gastrin could not be detected. Gastrin component III was found to predominate in the antrum, whereas component II was quantitatively very important in the duodenum. Circumstantial evidence suggests that fetal gastrin is delivered to the blood, and that it may exert trophic functions in the fetus.

IgM turnover in Crohn's disease.

Simultaneous turnover studies with radioiodine-labelled IgM and IgG were made in 12 patients with Crohn's disease. Intestinal protein loss was estimated by means of (59)Fe-labelled iron dextran. The serum levels of IgM, IgG, and IgA were normal in most cases. The catabolic rate of IgM was increased in all but one case. A positive correlation was present between the catabolic rate and serum concentration of IgM, an observation which, so far, has been made only in Crohn's disease. The synthetic rate of IgM was raised or high normal in four cases with an intraabdominal abscess. It was normal in the remaining cases. A strong positive correlation was found between the synthetic rates of IgM and IgG. The size of the protein loss was unrelated to the raised catabolic rates of IgM and IgG.Faecal radioiodine excretion from labelled IgM and IgG bore no relation to faecal (59)Fe excretion, nor did it indicate the site of the intestinal lesion. However, a close correlation was observed between faecal excretion of the labels from IgM and IgG.