scholarly journals Immunocytochemical localization of cathepsins B, H, and their endogenous inhibitor, cystatin beta, in islet endocrine cells of rat pancreas.

1988 ◽  
Vol 36 (7) ◽  
pp. 783-791 ◽  
Author(s):  
M Watanabe ◽  
T Watanabe ◽  
Y Ishii ◽  
H Matsuba ◽  
S Kimura ◽  
...  
Keyword(s):  
Endocrine Cells ◽  
Islet Cells ◽  
Secretory Granules ◽  
Double Immunostaining ◽  
Thin Sections ◽  
Cathepsin H ◽  
Immunocytochemical Techniques ◽  
The Difference ◽  
Immunocytochemical Reaction ◽  
Strong Immunoreactivity

To determine the characteristics of lysosomes in rat islet endocrine cells, we examined the precise localization of cathepsins B, H, and L and their specific inhibitors, cystatins alpha and beta, using immunocytochemical techniques. By use of serial semi-thin sections, we detected immunoreactivity for cathepsin B in insulin-, glucagon-, somatostatin-, and pancreatic polypeptide-positive (PP) cells. Strong immunoreactivity for cathepsin H was seen in A-cells and weak immunoreactivity in PP cells, but none in others. Immunodeposits for cystatin beta were demonstrated in B-cells. Brief dipping of thin sections in 1% sodium methoxide before the following immunocytochemical reaction enhanced specific deposits of immunogold particles on the target organelles. Use of a double-immunostaining technique showed co-localization of insulin with cystatin beta in many secretory granules. This suggests that cystatin beta may regulate converting enzymes participating in the maturation process of insulin. By use of an immunogold technique, heterogeneous localization of cathepsins B and H in lysosomes was also found among islet cells at the light microscopic level. This may be due to the difference in peptides degraded in lysosomes among the cells.

scholarly journals Embryogenesis of the murine endocrine pancreas; early expression of pancreatic polypeptide gene

Development ◽  
1991 ◽  
Vol 113 (4) ◽  
pp. 1257-1265 ◽  
Author(s):  
P.L. Herrera ◽  
J. Huarte ◽  
F. Sanvito ◽  
P. Meda ◽  
L. Orci ◽  
...  
Keyword(s):  
Pancreatic Polypeptide ◽  
Endocrine Cells ◽  
Islet Cells ◽  
Secretory Granules ◽  
Pcr Amplification ◽  
Immunogold Labelling ◽  
Pancreatic Ducts ◽  
Thin Sections ◽  
Embryonic Pancreas ◽  
Early Expression

By immunofluorescence on cytospin preparations and on semithin sections of mouse pancreatic buds, we have found glucagon and pancreatic polypeptide (PP)-containing cells at embryonal day 10.5 (E 10.5) in dorsal buds and at E 11.5 in ventral buds. Insulin-containing cells appear in dorsal buds at E 11.5, and one to two days later in ventral buds. Somatostatin-containing cells are detectable from E 13.5 in both dorsal and ventral buds. A quantitative analysis shows that up to E 15.5, PP-containing cells are relatively abundant in both buds. By PCR amplification of oligo(dT)-primed cDNAs prepared from total pancreatic RNA, we also detect PP mRNA from E 10.5 onwards, thus confirming the early expression of the PP gene in the developing mouse pancreas. Analysis of endocrine cells in situ suggests three major patterns of cell distribution in embryonic pancreas. First, individual hormone-containing cells are located within the epithelium of pancreatic ducts. In both dorsal and ventral buds, the majority of these endocrine cells contain PP, but many also contain glucagon, insulin or somatostatin. Secondly, clusters of endocrine cells are found in the pancreatic interstitium. Many of these cells contain both glucagon and PP which, by immunogold labelling of consecutive thin sections, can be shown to co-exist within individual secretory granules. Finally, starting on E 18.5, typical islets are formed with centrally located B cells and with the adult ‘one cell-one hormone’ phenotype. These results suggest an intriguing ontogenic relationship between A- and PP-cells, and also indicate that PP-containing cells may occupy a hitherto unexpected place in the lineage of endocrine islet cells.

scholarly journals Detection of nitric oxide synthase (NOS) in somatostatin-producing cells of human and murine stomach and pancreas.

1996 ◽  
Vol 44 (4) ◽  
pp. 339-346 ◽  
Author(s):  
M A Burrell ◽  
L M Montuenga ◽  
M García ◽  
A C Villaro
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Ganglion Cells ◽  
Secretory Granules ◽  
The Body ◽  
Gastric Epithelium ◽  
Thin Sections ◽  
Immunocytochemical Techniques ◽  
D Cells ◽  
Oxide Synthase

The aim of this study was to identify by immunocytochemistry the distribution of nitric oxide synthase (NOS) in human and murine gastric epithelium. Using two different antisera specific for neuronal NOS (nNOS), we detected nNOS immunoreactivity in endocrine cells of the epithelium of the body and pyloric regions as well as in ganglion cells of the intrinsic plexi of the stomach of the three species studied. Both immunocytochemistry of contiguous sections and double immunolabeling methods showed that the nNOS-immunoreactive cells were also immunoreactive for somatostatin. Co-localization of nNOS and somatostatin has also been found in the pancreatic islets, where strong nNOS immunoreactivity appeared in scattered cells, which were peripheral in rat and mouse islets and more randomly distributed in human. The possibility of crossreactivity between the antisera against nNOS and somatostatin was ruled out by means of absorption controls. Immunocytochemical techniques were also applied to thin sections, confirming the immunostaining of gastric D-cells, which was restricted principally to the secretory granules. The possible functional implications of these findings for gastric and pancreatic physiology are discussed.

scholarly journals Coexistence of renin and cathepsin B in secretory granules of granular duct cells in male mouse submandibular gland.

1993 ◽  
Vol 41 (3) ◽  
pp. 433-438 ◽  
Author(s):  
K Sano ◽  
S Waguri ◽  
N Sato ◽  
E Kominami ◽  
Y Uchiyama
Keyword(s):  
Submandibular Gland ◽  
Golgi Complex ◽  
Cathepsin B ◽  
Cysteine Proteinase ◽  
Secretory Granules ◽  
Interstitial Cells ◽  
Double Immunostaining ◽  
Duct Cells ◽  
Cathepsin H ◽  
Rat Pituitary

Cathepsin B, a representative lysosomal cysteine proteinase, has been demonstrated to coexist with renin in secretary granules of rat pituitary LH/FSH cells and renal juxtaglomerular cells. We investigated immunocytochemically the localization of cathepsins B, H, and L in the submandibular gland of male mice, in which active renin is also produced. By light microscopy, granular immunodeposits for cathepsin B were detected in epithelial cells of the gland, particularly in granular duct cells and interstitial cells. Immunoreactivity for cathepsins H and L was mainly found in interstitial cells, although that for cathepsin H was weakly seen in acinar cells. By electron microscopy, immunogold particles indicating cathepsin B intensely labeled small granules near the Golgi complex of granular duct cells and weakly labeled large secretory granules, whereas those showing renin labeled both granules. Double immunostaining co-localized immunogold particles showing renin and cathepsin B in small perinuclear granules near the Golgi complex. Some immunopositive granules seemed to be closely associated with the Golgi elements. These results indicate that the co-localization of renin and cathepsin B is also seen in secretory granules of granular duct cells in the mouse submandibular gland, as seen in rat juxtaglomerular and LH/FSH cells. This suggests that cathepsin B is one of the possible candidates for the renin-processing enzyme.

scholarly journals Cysteine proteinases in rat parathyroid cells with special reference to their correlation with parathyroid hormone (PTH) in storage granules.

1993 ◽  
Vol 41 (2) ◽  
pp. 273-282 ◽  
Author(s):  
Y Hashizume ◽  
S Waguri ◽  
T Watanabe ◽  
E Kominami ◽  
Y Uchiyama
Keyword(s):  
Electron Microscopy ◽  
Parathyroid Hormone ◽  
Special Reference ◽  
Secretory Granules ◽  
Cysteine Proteinases ◽  
Double Immunostaining ◽  
Thin Sections ◽  
Storage Granules ◽  
Golgi Network ◽  
And Storage

To further understand the roles of storage granules in parathyroid cells, we examined by immunocytochemistry the localization of cathepsins B and H and of PTH in rat parathyroid gland. In semi-thin sections, small and large granular immunodeposits for cathepsins B and H appeared in the cells, whereas those for PTH were detected throughout the cells, especially in perinuclear regions. By electron microscopy, immunogold particles indicating cathepsins B and H labeled lysosomes and storage granules, whereas those showing PTH were localized in storage granules, small secretory granules, and the trans-Golgi network. Small vesicles labeled by immunogold particles showing these proteinases often appeared close to the storage granules. By double immunostaining, immunogold particles indicating these proteinases were co-localized with those for PTH in storage granules. By EDTA treatment, immunoreactivity for cathepsins B and H and for PTH was notably reduced in the cells, but immunoreactivity for the proteinases was still seen in lysosomes. These results suggest that storage granules in the rat parathyroid cells fuse with small vesicles containing cathepsins B and H, which may participate in regulating the intracellular PTH levels by degrading PTH in the granules.

scholarly journals Immunocytochemical localization of cathepsin B in rat anterior pituitary endocrine cells, with special reference to its co-localization with renin and prorenin in gonadotrophs.

1991 ◽  
Vol 39 (9) ◽  
pp. 1199-1205 ◽  
Author(s):  
Y Uchiyama ◽  
M Nakajima ◽  
T Watanabe ◽  
S Waguri ◽  
N Sato ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Luteinizing Hormone ◽  
Light Microscopy ◽  
Cathepsin B ◽  
Anterior Pituitary ◽  
Endocrine Cells ◽  
Secretory Granules ◽  
Juxtaglomerular Cells ◽  
Immunocytochemical Localization ◽  
Double Immunostaining

We examined by immunocytochemistry the localization of cathepsin B in endocrine cells of rat anterior pituitary lobe, using a monospecific antibody to cathepsin B. By light microscopy, granular immunodeposits for cathepsin B were detected in most endocrine cells of anterior pituitary lobe. Cells immunoreactive for luteinizing hormone (LH) were diffusely immunostained by anti-cathepsin B. By electron microscopy, immunogold particles for cathepsin B were localized in lysosomes of thyrotrophs, somatotrophs, and mammotrophs. In mammotrophs, immunogold particles for cathepsin B were also detected in crinophagic bodies. Double immunostaining co-localized immunogold particles for LH and cathepsin B in secretory granules of gonadotrophs. Immunocytochemistry was also applied to demonstrate localization of renin and prorenin in LH-producing gonadotrophs; immunogold particles for renin were co-localized with those for LH, cathepsin B, or prorenin in their secretory granules. Immunogold particles for prorenin were also co-localized with those for LH or cathepsin B in secretory granules, but prorenin-positive granules appeared less frequently than renin-positive granules. These results suggest that cathepsin B not only plays a role in the protein degradation in lysosomes of anterior pituitary endocrine cells but also participates in the activation of renin in gonadotrophs, as has been demonstrated in secretory granules of juxtaglomerular cells.

scholarly journals Anti-lymphocyte antibody Leu-7 (HNK-1) recognizes a constituent of neuroendocrine granule matrix.

1986 ◽  
Vol 34 (9) ◽  
pp. 1213-1216 ◽  
Author(s):  
A S Tischler ◽  
H Mobtaker ◽  
K Mann ◽  
G Nunnemacher ◽  
W J Jason ◽  
...  
Keyword(s):  
Anterior Pituitary ◽  
Endocrine Cells ◽  
Islet Cells ◽  
Secretory Granules ◽  
Pancreatic Islet Cells ◽  
Cell Surfaces ◽  
Intracellular Protein ◽  
Myelin Associated Glycoprotein ◽  
Granule Matrix ◽  
Leu 7

Anti-lymphocyte monoclonal antibody HNK-1 (Leu-7) reacts with the cell surfaces of natural killer (NK) lymphocytes and with myelin-associated glycoprotein (MAG). This antibody reacts intensely with normal and neoplastic adrenal medullary cells. A small proportion of normal pancreatic islet cells, anterior pituitary, and gastroenteropancreatic endocrine cells also show Leu-7 immunoreactivity. In adrenal medulla, ultrastructural immunocytochemical studies and immunoblot analyses reveal that Leu-7 reacts with an intracellular protein of MW 75 KD which is localized within the matrices of the chromaffin granules. The MW of this protein differs from those of MAG and chromogranin A. The findings suggest that Leu-7 immunoreactivity might be a new marker for specific subsets of secretory granules.

scholarly journals Redistribution of alpha-granules and their contents in thrombin-stimulated platelets.

1984 ◽  
Vol 98 (2) ◽  
pp. 748-760 ◽  
Author(s):  
P E Stenberg ◽  
M A Shuman ◽  
S P Levine ◽  
D F Bainton
Keyword(s):  
Plasma Membrane ◽  
Tannic Acid ◽  
Extracellular Space ◽  
Plasma Membranes ◽  
Platelet Factor 4 ◽  
Platelet Factor ◽  
Thin Sections ◽  
Immunocytochemical Techniques ◽  
Morphologic Changes ◽  
Stimulation Of

The redistribution of beta-thromboglobulin (beta TG), platelet Factor 4 (PF4), and fibrinogen from the alpha granules of the platelet after stimulation with thrombin was studied by morphologic and immunocytochemical techniques. The use of tannic acid stain and quick-freeze techniques revealed several thrombin-induced morphologic changes. First, the normally discoid platelet became rounder in form, with filopodia, and the granules clustered in its center. The granules then fused with one another and with elements of the surface-connected canalicular system (SCCS) to form large vacuoles in the center of the cell and near the periphery. Neither these vacuoles nor the alpha granules appeared to fuse with the plasma membrane, but the vacuoles were connected to the extracellular space by wide necks, presumably formed by enlargement of the narrow necks connecting the SCCS to the surface of the unstimulated cell. The presence of fibrinogen, beta TG, and PF4 in corresponding large intracellular vacuoles and along the platelet plasma membrane after thrombin stimulation was demonstrated by immunocytochemical techniques in saponin-permeabilized and nonpermeabilized platelets. Immunocytochemical labeling of the three proteins on frozen thin sections of thrombin-stimulated platelets confirmed these findings and showed that all three proteins reached the plasma membrane by the same pathway. We conclude that thrombin stimulation of platelets causes at least some of the fibrinogen, beta TG, and PF4 stored in their alpha granules to be redistributed to their plasma membranes by way of surface-connected vacuoles formed by fusion of the alpha granules with elements of the SCCS.

scholarly journals Distribution of glucagon-like peptide I in canine and feline pancreas and gastrointestinal tract.

1986 ◽  
Vol 34 (9) ◽  
pp. 1117-1121 ◽  
Author(s):  
C R Vaillant ◽  
P K Lund
Keyword(s):  
Gastrointestinal Tract ◽  
Endocrine Cells ◽  
Northern Blot Analysis ◽  
Islet Cells ◽  
Cleavage Product ◽  
Antigenic Determinants ◽  
Normal Pancreas ◽  
Fetal Rat ◽  
Islet Tissue ◽  
Glucagon Like Peptide

Recently, a putative hormone, glucagon-like peptide I (GLP I), has been identified in the predicted sequences of the precursors to pancreatic glucagon in human, rat, hamster, and ox. The distribution of GLP I immunoreactivity in canine and feline pancreas and gastrointestinal tract was examined immunohistochemically and was compared with that of two other antigenic determinants of pancreatic pro-glucagon, i.e., glucagon and the NH2 terminus of glicentin. All three determinants occurred in the same population of islet cells in normal pancreas and in pancreas consisting predominantly of islet tissue from dogs with canine pancreatic acinar atrophy. Northern blot analysis of mRNA from the latter tissue, using a rat pre-pro-glucagon complementary DNA probe, revealed a single mRNA species similar in size to the pre-pro-glucagon mRNA detected in fetal rat pancreas. The three antigenic determinants of pancreatic pro-glucagon were co-localized also in intestinal L-cells and in canine gastric A-cells. Canine and feline pancreatic pro-glucagons therefore resemble those identified in other mammals and may also occur in gastrointestinal endocrine cells. Although there is evidence that the GLP I sequence is not liberated from pancreatic pro-glucagon, our results raise the possibility that this putative hormone may be a cleavage product of pro-glucagon in the gastrointestinal tract.

scholarly journals Immunocytochemical localization of histamine in secretory granules of rat peritoneal mast cells with conventional or rapid microwave fixation and an ultrastructural post-embedding immunogold technique.

1992 ◽  
Vol 40 (9) ◽  
pp. 1247-1256 ◽  
Author(s):  
G R Login ◽  
S J Galli ◽  
A M Dvorak
Keyword(s):  
Electron Microscopy ◽  
Mast Cells ◽  
Guinea Pig ◽  
Secretory Granules ◽  
Fixation Method ◽  
Structural Localization ◽  
Thin Sections ◽  
Aldehyde Fixation ◽  
Peritoneal Mast Cells ◽  
Rat Peritoneal Mast Cells

We used a post-embedding immunogold labeling approach to define the fine-structural localization of histamine in rat peritoneal mast cells that were fixed using either standard aldehyde fixation or a fast microwave-aldehyde fixation method. Specimens were processed routinely for electron microscopy. Thin sections were exposed first to guinea pig antihistamine antiserum and then to gold-conjugated goat IgG directed against guinea pig IgG. By transmission electron microscopy, gold particles were localized to the matrix of cytoplasmic granules. Control sections treated with non-immune sera did not show labeling of mast cells. Adsorption of antihistamine antiserum with purified histamine or histamine bound to agarose showed a significant reduction (p less than 0.005) in granule staining. We also confirmed that our isolation procedures yielded functionally competent mast cells which released histamine when stimulated with sheep anti-rat IgE antiserum or with compound 48/80. These studies define the conditions of fixation for electron microscopy that are appropriate for the localization of histamine in the granule matrix of rat peritoneal mast cells.

scholarly journals Three-dimensional electron microscopy of ribosomal chromatin in two higher plants: a cytochemical, immunocytochemical, and in situ hybridization approach.

1991 ◽  
Vol 39 (11) ◽  
pp. 1495-1506 ◽  
Author(s):  
P M Motte ◽  
R Loppes ◽  
M Menager ◽  
R Deltour
Keyword(s):  
Electron Microscopy ◽  
In Situ Hybridization ◽  
Spatial Organization ◽  
Higher Plants ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Thin Sections ◽  
Cytoplasmic Dna ◽  
Immunocytochemical Techniques

We report the 3-D arrangement of DNA within the nucleolar subcomponents from two evolutionary distant higher plants, Zea mays and Sinapis alba. These species are particularly convenient to study the spatial organization of plant intranucleolar DNA, since their nucleoli have been previously reconstructed in 3-D from serial ultra-thin sections. We used the osmium ammine-B complex (a specific DNA stain) on thick sections of Lowicryl-embedded root fragments. Immunocytochemical techniques using anti-DNA antibodies and rDNA/rDNA in situ hybridization were also applied on ultra-thin sections. We showed on tilted images that the OA-B stains DNA throughout the whole thickness of the section. In addition, very low quantities of cytoplasmic DNA were stained by this complex, which is now the best DNA stain used in electron microscopy. Within the nucleoli the DNA was localized in the fibrillar centers, where large clumps of dense chromatin were also visible. In the two plant species intranucleolar chromatin forms a complex network with strands partially linked to chromosomal nucleolar-organizing regions identified by in situ hybridization. This study describes for the first time the spatial arrangement of the intranucleolar chromatin in nucleoli of higher plants using high-resolution techniques.

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