Characterization of Transplantable Insulinoma Cells

2018 ◽  
pp. 213-222
Author(s):  
Paul C. Guest
Keyword(s):  
Insulinoma Cells

scholarly journals Comparative biosynthesis, covalent post-translational modifications and efficiency of prosegment cleavage of the prohormone convertases PC1 and PC2: glycosylation, sulphation and identification of the intracellular site of prosegment cleavage of PC1 and PC2

1993 ◽  
Vol 294 (3) ◽  
pp. 735-743 ◽  
Author(s):  
S Benjannet ◽  
N Rondeau ◽  
L Paquet ◽  
A Boudreault ◽  
C Lazure ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Brefeldin A ◽  
Pulse Labelling ◽  
Prohormone Convertases ◽  
Post Translational Modifications ◽  
Intracellular Degradation ◽  
Golgi Compartment ◽  
Carbonyl Cyanide ◽  
Insulinoma Cells

We present herein the pulse-chase analysis of the biosynthesis of the prohormone convertases PC1 and PC2 in the endocrine GH4C1 cells infected with vaccinia virus recombinants expressing these convertases. Characterization of the pulse-labelled enzymes demonstrated that pro-PC1 (88 kDa) is cleaved into PC1 (83 kDa) and pro-PC2 (75 kDa) into PC2 (68 kDa). Secretion of glycosylated and sulphated PC1 (84 kDa) occurs about 30 min after the onset of biosynthesis, whereas glycosylated and sulphated PC2 (68 kDa) is detected in the medium after between 1 and 2 h. Furthermore, in the case of pro-PC2 only, we observed that a fraction of this precursor escapes glycosylation. A small proportion (about 5%) of the intracellular glycosylated pro-PC2 (75 kDa) is sulphated, and it is this glycosylated and sulphated precursor that is cleaved into the secretable 68 kDa form of PC2. Major differences in the carbohydrate structures of PC1 and PC2 are demonstrated by the resistance of the secreted PC1 to endoglycosidase H digestion and sensitivity of the secreted PC2 to this enzyme. Inhibition of N-glycosylation with tunicamycin caused a dramatic intracellular degradation of these convertases within the endoplasmic reticulum, with the net effect of a reduction in the available activity of PC1 and PC2. These results emphasize the importance of N-glycosylation in the folding and stability of PC1 and PC2. Pulse-labelling experiments in uninfected mouse beta TC3 and rat Rin m5F insulinoma cells, which endogenously synthesize PC2, showed that, as in infected GH4C1 cells, pro-PC2 predominates intracellularly. In order to define the site of prosegment cleavage, pulse-chase analysis was performed at low temperature (15 degrees C) or after treatment of GH4C1 cells with either brefeldin A or carbonyl cyanide m-chlorophenylhydrazone. These results demonstrated that the onset of the conversions of pro-PC1 into PC1 and non-glycosylated pro-PC2 into PC2 (65 kDa) occur in a pre-Golgi compartment, presumably within the endoplasmic reticulum. In contrast, pulse labelling in the presence of Na(2)35SO4 demonstrated that the processing of glycosylated and sulphated pro-PC2 occurs within the Golgi apparatus. In order to test the possibility that zymogen processing is performed by furin, we co-expressed this convertase with either pro-PC1 or pro-PC2. The data demonstrated the inability of furin to cleave either proenzyme.

Characterization of serine/threonine protein phosphatases in RINm5F insulinoma cells

1993 ◽  
Vol 13 (6) ◽  
pp. 349-358 ◽  
Author(s):  
Åke Sjöhom ◽  
Richard E. Honkanen ◽  
Per-Olof Berggren
Keyword(s):  
Okadaic Acid ◽  
Potent Inhibitor ◽  
Protein Phosphatases ◽  
Marine Sponges ◽  
Rinm5f Cell ◽  
Ic50 Values ◽  
Marine Dinoflagellates ◽  
Insulinoma Cells

This study investigates the occurrence and regulation of serine/threonine protein phosphatases (PPases) in insulin-secreting RINm5F insulinoma cells. PPases types 1 and 2A were identified in crude RINm5F cell homogenates by both enzymatic assay and Western blot analysis. We then characterized and compared the inhibitory actions of several compounds isolated from cyanobacteria, marine dinoflagellates and marine sponges, (viz. okadaic acid, microcystin-LR, calyculin-A and nodularin) cation-independent PPase activities in RINm5F cell homogenates. It was found that okadaic acid was the least potent inhibitor (IC50 ≈ 10−9M, IC100 ≈ 10−6M), while the other compounds exhibited IC50 values of ≈ 5·10−10 M and IC100 ≈ 5·10−9 M. The findings indicate that the inhibitory substances employed in this study may be used pharmacologically to investigate the role of serine/threonine PPases in RINm5F cell insulin secretion, a process that is likely to be regulated to a major extent by protein phosphorylation.

The expression and function of a group VIA calcium-independent phospholipase A2 (iPLA2β) in β-cells

2004 ◽  
Vol 82 (10) ◽  
pp. 824-832 ◽  
Author(s):  
John Turk ◽  
Sasanka Ramanadham
Keyword(s):  
Cell Proliferation ◽  
Β Cells ◽  
Proliferation And Apoptosis ◽  
Bromoenol Lactone ◽  
Suicide Substrate ◽  
Glucose Stimulated Insulin Secretion ◽  
Calcium Independent Phospholipase A2 ◽  
Insulinoma Cells ◽  
And Function

Many cells express a Group VIA phospholipase A2, designated iPLA2β, that does not require calcium for activation, is stimulated by ATP, and is sensitive to inhibition by a bromoenol lactone suicide substrate (BEL). Studies in various cell systems have led to the suggestion that iPLA2β has a role in phospholipid remodeling, signal transduction, cell proliferation, and apoptosis. We have found that pancreatic islets, β-cells, and glucose-responsive insulinoma cells express an iPLA2β that participates in glucose-stimulated insulin secretion but is not involved in membrane phos pho lipid remodeling. Additionally, recent studies reveal that iPLA2β is involved in pathways that contribute to β-cell proliferation and apoptosis, and that various phospholipid-derived mediators are involved in these processes. Detailed characterization of the enzyme suggests that the β-cells express multiple isoforms of iPLA2β, and we hypothesize that these participate in different cellular functions.Key words: signalling, apoptosis, isoforms, mass spectrometry.

Cloning and characterization of the rat GALR1 galanin receptor from Rin14B insulinoma cells

1995 ◽  
Vol 34 (2) ◽  
pp. 179-189 ◽  
Author(s):  
Eric M. Parker ◽  
Darcy G. Izzarelli ◽  
Henry P. Nowak ◽  
Cathy D. Mahle ◽  
Lawrence G. Iben ◽  
...  
Keyword(s):  
Galanin Receptor ◽  
Insulinoma Cells

Morphological Characterization of Mycobacteriophage R1

1974 ◽  
Vol 32 ◽  
pp. 254-255
Author(s):  
B. L. Soloff ◽  
T. A. Rado
Keyword(s):  
Carbon Source ◽  
Stationary Phase ◽  
Growth Phase ◽  
Minimal Medium ◽  
Morphological Characterization ◽  
Logarithmic Growth Phase ◽  
Complete Lysis ◽  
Lysogenic Culture ◽  
Logarithmic Growth

Mycobacteriophage R1 was originally isolated from a lysogenic culture of M. butyricum. The virus was propagated on a leucine-requiring derivative of M. smegmatis, 607 leu−, isolated by nitrosoguanidine mutagenesis of typestrain ATCC 607. Growth was accomplished in a minimal medium containing glycerol and glucose as carbon source and enriched by the addition of 80 μg/ ml L-leucine. Bacteria in early logarithmic growth phase were infected with virus at a multiplicity of 5, and incubated with aeration for 8 hours. The partially lysed suspension was diluted 1:10 in growth medium and incubated for a further 8 hours. This permitted stationary phase cells to re-enter logarithmic growth and resulted in complete lysis of the culture.

AEM analysis of crystallization in Ti-based amorphous alloys

1983 ◽  
Vol 41 ◽  
pp. 270-271
Author(s):  
A.R. Pelton ◽  
A.F. Marshall ◽  
Y.S. Lee
Keyword(s):  
Magnetic Properties ◽  
Amorphous Alloys ◽  
Amorphous Materials ◽  
Isothermal Annealing ◽  
Amorphous Matrix ◽  
Nucleation And Growth ◽  
Current Interest ◽  
Amorphous Films ◽  
Electrical And Magnetic Properties

Amorphous materials are of current interest due to their desirable mechanical, electrical and magnetic properties. Furthermore, crystallizing amorphous alloys provides an avenue for discerning sequential and competitive phases thus allowing access to otherwise inaccessible crystalline structures. Previous studies have shown the benefits of using AEM to determine crystal structures and compositions of partially crystallized alloys. The present paper will discuss the AEM characterization of crystallized Cu-Ti and Ni-Ti amorphous films.Cu60Ti40: The amorphous alloy Cu60Ti40, when continuously heated, forms a simple intermediate, macrocrystalline phase which then transforms to the ordered, equilibrium Cu3Ti2 phase. However, contrary to what one would expect from kinetic considerations, isothermal annealing below the isochronal crystallization temperature results in direct nucleation and growth of Cu3Ti2 from the amorphous matrix.

Characterization of Coherent, Ordered Precipitates in Nickel-Base Alloys

1973 ◽  
Vol 31 ◽  
pp. 144-145
Author(s):  
B. H. Kear ◽  
J. M. Oblak
Keyword(s):  
Stable Phase ◽  
Nickel Base Superalloy ◽  
Alloy 718 ◽  
Commercial Alloy ◽  
Precipitate Morphology ◽  
Continuous Precipitation ◽  
Nickel Base ◽  
Base Superalloy ◽  
Strengthening Precipitate

A nickel-base superalloy is essentially a Ni/Cr solid solution hardened by additions of Al (Ti, Nb, etc.) to precipitate a coherent, ordered phase. In most commercial alloy systems, e.g. B-1900, IN-100 and Mar-M200, the stable precipitate is Ni3 (Al,Ti) γ′, with an LI2structure. In A lloy 901 the normal precipitate is metastable Nis Ti3 γ′ ; the stable phase is a hexagonal Do2 4 structure. In Alloy 718 the strengthening precipitate is metastable γ″, which has a body-centered tetragonal D022 structure.Precipitate MorphologyIn most systems the ordered γ′ phase forms by a continuous precipitation re-action, which gives rise to a uniform intragranular dispersion of precipitate particles. For zero γ/γ′ misfit, the γ′ precipitates assume a spheroidal.

The Use of Advanced Analytical Techniques for Characterization of the Chemical, Physical, and Crystallographic Nature of a Surface

1973 ◽  
Vol 31 ◽  
pp. 132-133 ◽  
Author(s):  
R. E. Herfert
Keyword(s):  
Surface Characterization ◽  
Analytical Techniques ◽  
X Ray Diffraction ◽  
Depth Of Penetration ◽  
X Ray ◽  
The Past ◽  
Transmission Electron ◽  
Scanning Electron

Studies of the nature of a surface, either metallic or nonmetallic, in the past, have been limited to the instrumentation available for these measurements. In the past, optical microscopy, replica transmission electron microscopy, electron or X-ray diffraction and optical or X-ray spectroscopy have provided the means of surface characterization. Actually, some of these techniques are not purely surface; the depth of penetration may be a few thousands of an inch. Within the last five years, instrumentation has been made available which now makes it practical for use to study the outer few 100A of layers and characterize it completely from a chemical, physical, and crystallographic standpoint. The scanning electron microscope (SEM) provides a means of viewing the surface of a material in situ to magnifications as high as 250,000X.

Skeletal elements of crinoid echinoderms: High-resolution structural and microanalytical results

1983 ◽  
Vol 41 ◽  
pp. 194-195
Author(s):  
D. F. Blake ◽  
L. F. Allard ◽  
D. R. Peacor
Keyword(s):  
High Resolution ◽  
Marine Invertebrates ◽  
Sea Urchins ◽  
Structural Features ◽  
Sea Stars ◽  
High Resolution Tem ◽  
Relationship Of ◽  
The Relationship ◽  
Insight Into

Echinodermata is a phylum of marine invertebrates which has been extant since Cambrian time (c.a. 500 m.y. before the present). Modern examples of echinoderms include sea urchins, sea stars, and sea lilies (crinoids). The endoskeletons of echinoderms are composed of plates or ossicles (Fig. 1) which are with few exceptions, porous, single crystals of high-magnesian calcite. Despite their single crystal nature, fracture surfaces do not exhibit the near-perfect {10.4} cleavage characteristic of inorganic calcite. This paradoxical mix of biogenic and inorganic features has prompted much recent work on echinoderm skeletal crystallography. Furthermore, fossil echinoderm hard parts comprise a volumetrically significant portion of some marine limestones sequences. The ultrastructural and microchemical characterization of modern skeletal material should lend insight into: 1). The nature of the biogenic processes involved, for example, the relationship of Mg heterogeneity to morphological and structural features in modern echinoderm material, and 2). The nature of the diagenetic changes undergone by their ancient, fossilized counterparts. In this study, high resolution TEM (HRTEM), high voltage TEM (HVTEM), and STEM microanalysis are used to characterize tha ultrastructural and microchemical composition of skeletal elements of the modern crinoid Neocrinus blakei.

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