Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01275 (glucagon)
 26,492 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mechanisms that regulate collagen gene expression in hepatic cells are poorly understood. Accelerated Ca2+ fluxes are associated with inhibiting collagen synthesis selectively in human fibroblasts (Flaherty, M., and Chojkier, M. (1986) J. Biol. Chem. 261, 12060-12065). In suspension cultures of isolated hepatocytes, the Ca2+ agonist vasopressin increases cytosolic levels of free Ca2+ (Thomas, A.P., Marks, J.S., Coll, K.E., and Williamson, J. R. (1983) J. Biol. Chem. 258, 5716-5725). However, whether vasopressin's interactions with plasma membrane V1 receptors attenuate hepatic collagen production is unknown. We investigated this problem by studying vasopressin's effects on collagen synthesis and Ca2+ efflux in long-term primary cultures of differentiated and proliferation-competent adult rat hepatocytes. Twelve-day-old quiescent cultures were exposed to test substances and labeled with [5-3H]proline. Determinations of radioactivity in collagenase-sensitive and collagenase-resistant proteins were used to calculate the relative levels of collagen production. Synthetic [8-arg]vasopressin stimulated 45Ca2+ efflux within 1 min and inhibited hepatocyte collagen production within 3 h by 50%; overall rates of protein synthesis were not affected significantly. In cultures labeled with [35S]methionine, vasopressin also decreased the levels of newly synthesized and secreted albumin, but not fibrinogen, detected in specific immunoprecipitates analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Northern blot analyses using specific [32P]cDNA probes revealed 70% decreases in hybridizable levels of collagen alpha 1(I) mRNA in hepatocyte cultures treated with either vasopressin or Ca2+ ionophore A23187; hybridizable levels of albumin mRNA also fell approximately 50% following vasopressin treatment. Vasopressin did not affect collagen production in quiescent cultures of mouse Swiss 3T3, human myofibroblast or rat smooth muscle cells; and hepatocyte collagen production was unaffected by treatment with glucagon or dibutyryl cAMP. Thus, accelerated Ca2+ fluxes induced by vasopressin are associated with decreased production of hepatocyte collagen and albumin in primary cultures that simulate quiescent adult rat liver.
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PMID:Vasopressin inhibits type-I collagen and albumin gene expression in primary cultures of adult rat hepatocytes. 254 14

Isolation of islets of Langerhans from the pancreas by action of collagenase, a major breakthrough for physiological studies in vitro, has long appeared empirical, and the results were sometimes unpredictable. Isolation yields (number of islets obtained per pancreas) and their reproducibility, purification from exocrine remnants and vitality of the islets obtained, can improve owing to precise techniques, adapted to the architecture of collagen in the pancreas. We have tested four isolation-purification techniques in the rat pancreas. The best results were obtained by combining intra-ductal collagenase injection, with complete but moderate distension of the gland, avoiding leakage, multistep digestion (in situ and then in vitro), followed by purification on a discontinuous bovine serum albumin (BSA) gradient. Average yields were 670 +/- 40 islets per pancreas (range 570-800), versus 170 +/- 9 (range 40-350) with the technique used initially. The use of BSA discontinuous gradient improved the purification yield: 90-96% of islets obtained were concentrated in the 26/29% BSA interface. Furthermore, this technique shortened the duration of purification step: 1 hr (centrifugation gradient) vs 3 hrs (handpicked). It was verified that islets were morphologically free of exocrine tissue. Islet structure was well preserved either in conventional histology or insulin and glucagon immunoperoxidase staining. Islet vitality, as assessed by trypan blue exclusion test, was 100% of freshly isolated islets, and 89% after 24 hrs culture. Insulin secretory responses to a given stimulus were stronger than in the case of islets isolated by former techniques: 10-12 times the basal release (vs 5 times) with clear dose-response proportionality.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Reproducible high yields of rat islets of Langerhans. 254 70

The effect of various factors and substrates on the growth of a human hepatoblastoma cell line, HuH-6, which was inoculated at low density in a serum-free medium was examined. Several supplements were required to enhance cell growth of HuH-6. These included cholera toxin (CT), glucagon (Glu) and selenium (Se). Type IV collagen (C-IV) provided the most conductive environment tested for cell growth. These results suggest that CT, Glu, Se, and C-IV are important stimulators for the continuous growth of HuH-6 in a serum-free medium at low density.
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PMID:Effect of various factors and substrates on the growth of a human hepatoblastoma cell line, HuH-6 in a serum-free medium. 262 43

Viable Hepatocytes were isolated from adult canine liver by in situ collagenase perfusion, and cultured on collagen coated borosilicate glass plates (100 X 200mm) at confluent cell density. The medium of hepatocytes in the primary culture was L-15 supplemented with aprotinin 5000U/L, proline 30mg/L, insulin 10(-8)M, dexamethasone 10(-8)M, glucagon 10(-8)M, and h-EGF 10ng/ml. Long-stroke type bioartificial liver module consisted of 200 glass plates with hepatocytes. It contained 6 billion primary cultured cells in total, that is almost equivalent to 30% of the normal canine liver. All hepatocytes in the module were quite viable during 2 weeks in the perfusion culture, and maintained various liver functions at a high level. Gluconeogenesis was 368.0 +/- 15.4mg/module/hr, albumin synthesis was 19.1 +/- 2.5mg/module/day, ureogenesis was 3.7 +/- 0.1mg/module/hr, and ammonia metabolism was 8.4mg/module/hr. Moreover, those functions were maintained at least 2 weeks in the canine plasma as well as in the culture medium with hormones. This hybrid bioartificial liver may exert various liver functions like a liver in situ.
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PMID:[Hybrid bioartificial liver using canine hepatocytes in primary culture]. 276 24

Prolyl endopeptidase [EC 3.4.21.26] was purified 4,675-fold with a yield of 26.3% from porcine muscle. The purified enzyme was shown to be very similar to the liver enzyme with respect to its molecular weight (72,000-74,000), antigenicity, substrate specificity, and susceptibility to protease inhibitors. Among several bioactive peptides, angiotensins I, II, and III had the lowest Km of 0.6 to 3 microM with the lowest kcat of 0.19 to 0.85 s-1, while thyrotropin-releasing hormone had the highest Km of 98 microM with the highest kcat of 14.4 s-1. Interestingly, mastoparan was hydrolyzed at alanyl bonds, but insulin was only slightly hydrolyzed and glucagon was not hydrolyzed although the latter two peptides contain prolyl and/or alanyl bonds. Muscle prolyl endopeptidase failed to hydrolyze proteins with high molecular weight such as albumin, immunoglobulin G, elastin, collagen, and muscle soluble and insoluble proteins. However, 8 of 14 peptides with molecular weights lower than 3,000, which were isolated from muscle extract, were digested by this enzyme, and they were proved to contain prolyl and/or alanyl residues in their molecules. The data suggest that they are probable endogenous substrates for prolyl endopeptidase.
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PMID:Porcine muscle prolyl endopeptidase and its endogenous substrates. 285 85

Some effects of culturing adult rat hepatocytes on each of four different substrates--laminin (LN), collagen type I (C-I), collagen type IV (C-IV), and fibronectin (FN)--have been investigated under defined conditions. No differential effect on the attachment of the cells to the various substrates was noted; however, the spreading of hepatocytes shortly after initial plating was most strikingly enhanced by FN, whereas LN exhibited little or no such enhancement. The two collagen substrates enhanced the spreading of hepatocytes more than did LN, but less than FN. The different substrates had no differential effect on the induction of tyrosine aminotransferase by dexamethasone and glucagon for at least the first 10 d in culture. The longevity of the hepatocytes was not changed significantly by any of the substrates, at least through the 14th d of culture. During the culture periods the hepatocytes at high cell density were maintained as confluent monolayers, regardless of the substrate on which they had been cultured. After 14 d of culture, gamma-glutamyltranspeptidase activity was highest in cells cultured on C-IV, and lowest in those on FN. DNA synthesis in cultured hepatocytes at a low cell density was highest in cells cultured on FN, with decreasing levels of this parameter in cells cultured on C-IV, C-I, and LN, respectively. These results demonstrate that specific components of the extracellular matrix modulate both differentiated functions and the replication of hepatocytes cultured in serum-free medium.
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PMID:Effects of extracellular matrix components on the growth and differentiation of cultured rat hepatocytes. 288 70

This report describes a new method for detecting and quantitating those immunoglobulins G (IgG) in serum that are related to Graves' disease. The method is based on previous observations which indicate that the guinea pig fat cell membrane (FCM) is capable of binding Graves'-specific IgG, but does not bind the IgG common to Graves' disease and Hashimoto's disease, such as antimicrosomal antibodies. Crude FCM preparations were iodinated by a lactoperoxidase technique and were then treated with Triton X-100 to yield a solubilized radioiodinated FCM (SFCM) preparation. SFCM, which retained bovine (b) TSH binding and Graves'-IgG binding properties, provided a radioactively labeled receptor with which to test for the presence of fat cell-binding IgG (FBI) in immunoprecipitates prepared by reacting these IgG with antibody against the Fc fragment of human IgG. FBI values (percentage of added SFCM bound to immunoprecipitate; mean + SD) in IgG from 16 patients with thyrotoxicosis caused by Graves' disease (6.0 +/- 1.7) were completely separated from those in IgG from 16 normal subjects (0.4 +/- 0.3). IgG from 2 hypothyroid patients with Hashimoto's disease, which were strongly positive in the TSH binding inhibition (TBI) assay, yielded FBI values within the range in Graves' disease, but values in TBI-negative IgG from 15 other patients with Hashimoto's disease were normal (0.0 +/- 0.9). Moderately false positive FBI values were found in the IgG of 15 patients with rheumatoid arthritis or systemic lupus erythematosis, all rheumatoid factor positive, 3 of which were also TBI positive. In IgG from Graves' disease and those from patients with TBI-positive collagen-vascular disease, binding of SFCM was inhibited by bTSH in a dose-dependent manner. As with binding of TSH to thyroid plasma membranes, similar but less potent inhibition of binding of IgG to SFCM was produced by LH, FSH, and hCG, but not by insulin, glucagon, PRL, or ACTH. FBI values in TBI-negative IgG from patients with collagen-vascular disease were also decreased by TSH, but higher concentrations of bTSH were required. In 40 IgG from among the various clinical groups tested, a significant correlation was found between FBI values and TBI activity (r = 0.48; P less than 0.01). In addition, among 10 IgG from Graves' disease and 6 from collagen-vascular disease patients, a very close correlation (r = 0.89; P less than 0.001) was noted between their TBI activity and the extent to which their FBI values were decreased by a standard concentration of bTSH.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Detection and measurement of fat cell-binding immunoglobulins: a new method applicable to the diagnosis and study of Graves' disease. 299 73

A technique has been devised to attach adult rat hepatocytes to collagen-coated dextran microcarriers. Cells were cultured serum-free for 2 d and their viability, enzyme activities, glucose metabolism, and hormone responsiveness were compared to data obtained from conventional dish cell culture. The two different culture methods showed no difference in cell viability and morphology. Microcarrier-cultured cells exhibited hormone responsiveness comparable to dish cultures; glycolysis could be activated three-fold by the sole addition of insulin, and gluconeogenesis was increased by 40 to 50% by glucagon. During the 48-h culture glucokinase and phosphoenolpyruvate carboxykinase activities declined at a similar rate in both culture systems. Long-term culture with 0.1 microM insulin prevented the decrease of glucokinase activity. Insulin responsiveness (activation of glycolysis) was still pronounced after 48 h in culture. The microcarrier technique establishes a new in vitro liver system in which acute and long-term hormonal actions can be investigated using the technical advantages of a suspension culture.
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PMID:Adult rat hepatocyte microcarrier culture. Comparison to the conventional dish culture system. 305 97

(i) Hepatocytes isolated from adult rats were cultured for 2 to 3 weeks on collagen in a modified, serum-free Waymouth medium containing fatty acids and varying concentrations of glucocorticoid, insulin and glucagon. (ii) In the presence of all three hormones, it was possible to maintain the content of DNA, the activity of glucokinase, pyruvate kinase, hexokinase and lactate dehydrogenase at initial levels for 2 to 3 weeks. The activity of glucokinase and pyruvate kinase was affected by the concentration of insulin. (iii) The activity of alcohol dehydrogenase was stable for 3 days and declined to about 25% of the initial level after 2 weeks of culture, irrespective of the presence of hormones. (iv) Maintenance of albumin secretion was dependent on the presence of glucocorticoid, and glucocorticoid and insulin showed an additive or, at some time points, a synergistic effect on its secretion. (v) The content of cytochrome P-450 could be kept at 65% of the initial level, provided that a relatively high concentration of dexamethasone was present (10(-6) M). (vi) In the absence of hormones, urea synthesis was 70% of initial levels throughout the experimental period. With insulin and glucocorticoid present, a high concentration of glucagon (10(-8) M) was required to maintain the synthesis of urea at this level. (vii) It is concluded that hepatocyte cultures as described in the present study may be a useful, well-defined system for long-term metabolic, pharmacologic and toxicologic studies.
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PMID:Long-term culture of hepatocytes: effect of hormones on enzyme activities and metabolic capacity. 327 89

Cell suspensions prepared by collagenase digestion of pancreases obtained from rat fetuses (21.5 d old) and newborns (2.5 d old) were mixed with a collagen solution and inoculated on a collagen base layer. At the onset of the culture, most acinar cells became necrotic, whereas other epithelial cells proliferated. Most of the cell clusters arranged themselves into simple polarized structures composed of epithelial cells forming hollow spheres, and from these budded neoformed endocrine islets. Scarce fibroblasts were located close to these structures. Immunocytochemical localization of insulin and glucagon, as well as ultrastructural characteristics of the cell types revealed an intrainsular distribution similar to the in vivo localization. Tridimensional matrix of collagen offers, to perinatal pancreatic cells in culture, an environment close to the in vivo conditions: cells reorganize themselves in tissuelike structures and cell interactions concerned in the cytodifferentiation of pancreatic islets occur. This system allows for the study of undifferentiated epithelial cells--the presumed stem cells--differentiating and differentiated endocrine cells in the same preparation.
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PMID:In vitro cytodifferentiation of perinatal rat islet cells within a tridimensional matrix of collagen. 327 39


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