Immunocytochemical demonstration of ornithine decarboxylase in the rat exocrine pancreas using the protein A–gold technique

1986 ◽  
Vol 64 (4) ◽  
pp. 444-448 ◽  
Author(s):  
Jean Morisset ◽  
Patrice Sarfati ◽  
Gilles Grondin
Keyword(s):  
Ornithine Decarboxylase ◽  
Protein A ◽  
Acinar Cells ◽  
Growth Stimulation ◽  
Pancreatic Acinar Cells ◽  
Saline Control ◽  
The Third ◽  
Group 4 ◽  
Injection Control ◽  
Rat Exocrine Pancreas

Previous studies from our laboratory have shown that caerulein, a cholecystokinin analog, can induce pancreatic growth. Because ornithine decarboxylase (ODC) could be involved in this process, it is of interest to localize and estimate ODC immunoreactivity in rat pancreatic acinar cells from control and caerulein-treated animals. This was carried out with the protein A–gold immunocytochemical technique. Rats received either saline (control) or caerulein at a dose of 1 μg∙kg−1 and were sacrificed 8 h after the first injection (control and caerulein group), 4 h after the second caerulein injection (second caerulein group), and 8 h after the third caerulein injection (third caerulein group). ODC immunoreactivity was revealed using a specific antibody. ODC was localized specifically in nuclei and rough endoplasmic reticulum (RER) of the pancreatic acinar cells and the number of gold particles was increased in both of these organelles by caerulein. Peak ODC immunoreactivity was observed in nuclei 4 h after the second caerulein injection, whereas it occurred 8 h after the third peptide injection in the RER. These studies are the first to demonstrate ODC localization in pancreatic acinar cells and show that the enzyme can be induced early upon growth stimulation of the organ by a cholecystokinin analog.

scholarly journals Double immunocytochemical labeling applying the protein A-gold technique.

1982 ◽  
Vol 30 (1) ◽  
pp. 81-85 ◽  
Author(s):  
M Bendayan
Keyword(s):  
Tissue Section ◽  
Protein A ◽  
Antigenic Site ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Secretory Proteins ◽  
Zymogen Granules ◽  
Double Labeling ◽  
Gold Particles ◽  
Antigenic Sites

In the present study we report the modifications and the different steps of the protein A-gold (pAg) technique that allow the simultaneous demonstration of two antigenic sites on the same tissue section. The labeling is carried out in the following manner: face A of the tissue section is incubated with an antiserum followed by a pAg complex prepared with large gold particles; face B of the same tissue section is then incubated with a second antiserum followed by a pAg complex prepared with small gold particles. Each of the pAg complexes reveals a different antigenic site on opposite faces of the tissue section. The transparency of the section in the electron beam allows the visualization of the gold particles present on both faces. The double labeling pAg technique was applied for the simultaneous demonstration of two secretory proteins in the same Golgi, condensing vacuoles, and zymogen granules of the rat pancreatic acinar cells.

scholarly journals Beneficial Effects of Pyridoxal Phosphate on Hydrogen Peroxide Induced Stress on Isolated Pancreatic Acinar Cells

2016 ◽  
Author(s):  
Rajanna Ajumeera ◽  
Vijayalakshmi Venkatesan
Keyword(s):  
Treatment Group ◽  
Pyridoxal Phosphate ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
H2o2 Treatment ◽  
Group 4 ◽  
Group 3 ◽  
Group 2 ◽  
Group 1

Aims: To study the effects of pyridoxal phosphate (PLP) on oxidative stress in isolated pancreatic acinar cells. We have previously shown that PLP has cytoprotective and insulinotropic effects on mice islet cells in vivo and in vitro studies. Main methods: Acinar cells were isolated from three months old WNIN male rats and were cultured in vitro for a period of 24 h in CO2 incubator. Later the cells were divided into four groups as untreated (group 1), H2O2 treatment (group 2), PLP treatment (group 3) and PLP followed by H2O2 treatment (group 4). Cell viability was confirmed using MTT assays, oxidative stress levels were measured with ROS assay, change in different protein levels were recorded by flow cytometry. The acinar cells insulin secretion assay was performed with ELISA. The amylase protein expression was assessed using confocal microscopy. Key findings: The cell viability of acinar cell in group 1 was considered as 100%, while in group 2 it was reduced to 82% due to H2O2 effect, and in group 3 (99.8%) and group 4 (99.5%) were near to group 1 due to the cytoprotective effect of PLP. The ROS levels were increased by 1.47 folds in group 2, while PLP decreased to 1.02 fold in group 4, which was comparable with the changes in group 1. Beneficial effects of PLP were also observed from the increased expression levels of acinar cells are amylase -2.01, neurogenin-3-9.51, PDX-1- 23.6 and insulin-13.5 in group 3 compared to group 1. The specificity of PLP’s response was confirmed by amino oxy acetic acid (AOAA), a specific PLP inhibitor. The increased amylase protein localization with PLP was confirmed by confocal microscopy. Insulin secretion efficiency of acinar cells was observed to be 6.13 folds higher at basal and 24.63 fold higher at stimulated levels in group3 compared to group1. Significance: Our results advocate the antioxidant and cytoprotective effects of PLP on the pancreatic acinar cell along with increased pancreatic marker expressions of amylase,PDX-1, neurogenin-3 and insulin proteins.

scholarly journals Possible association of chaperonin 60 with secretory proteins in pancreatic acinar cells.

1996 ◽  
Vol 44 (7) ◽  
pp. 743-749 ◽  
Author(s):  
I M Le Gall ◽  
M Bendayan
Keyword(s):  
Secretory Pathway ◽  
Active Form ◽  
Acinar Cells ◽  
Morphological Data ◽  
Pancreatic Acinar Cells ◽  
Electron Microscopic Level ◽  
Secretory Proteins ◽  
Zymogen Granule ◽  
Electron Microscopic ◽  
Rat Exocrine Pancreas

Assembly and folding of newly synthesized polypeptides, acquisition of their biological active form, and their translocation in different cellular compartments are processes assisted by molecular chaperones. Because particular chaperones have been found to be present along the RER-Golgi-granule secretory pathway in pancreatic acinar cells, we presume that they should play important roles in secretion. In the present study, applying double immunogold labeling at the electron microscopic level on rat exocrine pancreas, we have revealed the existence of a topographical association between Hsp60 and particular pancreatic enzymes along the secretory pathway. The highest association was found for amylase, lipase, and chymotrypsinogen, whereas trypsinogen and carboxypeptidase B showed much lower association values. Immunoprecipitation of isolated zymogen granule content with an anti-Hsp60 antibody appears to confirm the morphological data, since amylase and lipase were found to co-precipitate with Hsp60. These findings support the hypothesis that Hsp60 is associated with certain pancreatic proteins along the secretory pathway. Hsp60 would assist the proper folding and assembly of pancreatic secretory proteins and could also prevent their autoactivation before secretion.

Coupled Na+-H+ exchange in isolated acinar cells from rat exocrine pancreas

1985 ◽  
Vol 249 (1) ◽  
pp. G125-G136 ◽  
Author(s):  
W. Hellmessen ◽  
A. L. Christian ◽  
H. Fasold ◽  
I. Schulz
Keyword(s):  
Intracellular Ph ◽  
Fluorescence Intensity ◽  
Exocrine Pancreas ◽  
Acinar Cells ◽  
Extracellular Ph ◽  
Pancreatic Acinar Cells ◽  
Intracellular Acidification ◽  
Extracellular Medium ◽  
Rat Exocrine Pancreas ◽  
Intracellular Fluorescence

Isolated acinar cells from the rat exocrine pancreas were loaded with 6-carboxyfluorescein diacetate (CFDA), and the intracellular pH (pHi) was estimated from the pH-dependent fluorescence intensity of trapped 6-carboxyfluorescein liberated from CFDA by intracellular esterases. The intracellular fluorescence intensity was calibrated by equilibrating the internal and external pH with nigericin in K+ buffers. In the absence of Na+ (130 mmol/l K+) a pHi of 6.86 +/- 0.04 was found; in its presence (130 mmol/l Na+) a pHi of 7.17. Acute addition of Na+ increased intracellular pH with increasing Na+ concentrations, reaching a maximum at 150 mmol/l with an apparent Km of approximately 40 mmol/l. Of the different cations tested on pHi, such as Li+, K+, Rb+, and Cs+, only Li+ showed an effect on pHi similar to that of Na+. Amiloride dose dependently inhibited both Na+- and Li+-induced alkalinization (apparent Km approximately 10(-5) mol/l). In the presence of ouabain pHi was decreased by 0.2 pH units. Intracellular acidification induced by permeable buffers such as acetic acid-acetate or CO2-HCO3- was dissipated more rapidly in the presence of Na+ compared with K+ or with Na+ and amiloride in the medium. In Li+-preincubated cells intracellular acidification was higher in the absence of Li+ in the extracellular medium than in its presence. This Li+ gradient-induced acidification was dependent on the extracellular pH, was highest at an extracellular pH of 7.05, and decreased with increasing pH to 7.5. The results allow the conclusion that a coupled Na+-H+ exchange is present in pancreatic acinar cells and that the intracellular pH rather than the extracellular Na+ concentration regulates this transport mechanism.

scholarly journals Immunocytochemical localization of kallikrein in the rat exocrine pancreas.

1982 ◽  
Vol 30 (1) ◽  
pp. 58-66 ◽  
Author(s):  
M Bendayan ◽  
T B Orstavik
Keyword(s):  
Secretory Pathway ◽  
Protein A ◽  
Pancreatic Tissue ◽  
Pancreatic Acinar Cells ◽  
Zymogen Granules ◽  
Rat Exocrine Pancreas ◽  
Pancreatic Exocrine ◽  
Embedding Medium ◽  
Lowicryl K4m ◽  
Acinar Lumen

The subcellular localization of kallikrein was studied in the rat pancreas using the immunocytochemical protein A-gold technique. Kallikrein was found at the level of the rough endoplasmic reticulum (RER), Golgi cisternae, condensing vacuoles, and zymogen granules of the pancreatic acinar cells as well as in the acinar lumen. The effect of various tissue processings on the immunocytochemical labeling of kallikrein was evaluated using pancreatic tissue fixed in glutaraldehyde and embedded in Epon, Lowicryl K4M, or glycol methacrylate (GMA). Compared to the results obtained with Epon, Lowicryl allowed improved resolution and specificity in the immunocytochemical labeling, while GMA retained greater amounts of kallikrein antigenicity leading to a higher intensity in the labeling; since it also gave a good ultrastructural preservation, GMA appeared to be the superior embedding medium for the localization of kallikrein. The quantitative evaluation of the labeling obtained under the three embedding conditions showed the presence of an increasing concentration gradient along the RER-Golgi-granule secretory pathway, suggesting that, like other pancreatic exocrine enzymes, kallikrein is synthesized in the RER, processed through the Golgi apparatus, and packed in the zymogen granules before being released into the acinar lumen.

Visualization of CCK-evoked exocytic events in rat pancreatic acinar cells

2001 ◽  
Vol 120 (5) ◽  
pp. A24-A24
Author(s):  
H GAISANO ◽  
L TANG ◽  
L SHEU ◽  
W TRIMBLE
Keyword(s):  
Acinar Cells ◽  
Pancreatic Acinar Cells

Effects of oncogenic K-ras on pancreatic acinar cells In vitro

2001 ◽  
Vol 120 (5) ◽  
pp. A722-A722
Author(s):  
Y BI ◽  
C LOGSDON
Keyword(s):  
Acinar Cells ◽  
Pancreatic Acinar Cells

Carbon tetrachloride induced ultrastructural alterations in pancreatic acinar cells and in the hepatocytes. Similarities and differences

Toxicology ◽  
1978 ◽  
Vol 11 ◽  
pp. 289-296 ◽  
Author(s):  
Carmen R. De Castro ◽  
Adriana S. Bernacchi ◽  
Elida C. De Ferreyra ◽  
Olga M. De Fends ◽  
José A. Castro
Keyword(s):  
Carbon Tetrachloride ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Ultrastructural Alterations ◽  
Similarities And Differences
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