scholarly journals A Cytochemical Study on the Pancreas of the Guinea Pig

1958 ◽  
Vol 4 (2) ◽  
pp. 203-218 ◽  
Author(s):  
Philip Siekevitz ◽  
George E. Palade
Keyword(s):  
Electron Microscopy ◽  
Guinea Pig ◽  
Pancreatic Tissue ◽  
Supernatant Fraction ◽  
Differential Centrifugation ◽  
Zymogen Granules ◽  
Cytochemical Study ◽  
Specific Activities ◽  
Main Components ◽  
Cell Fractions

Pancreatic tissue, (guinea pig) homogenized in 0.88 M sucrose, was fractionated by differential centrifugation into a nuclear, zymogen, mitochondrial, microsomal, and final supernatant fraction. The components of the particulate fractions were identified with well known intracellular structures by electron microscopy. The fractions were analyzed for protein-N and RNA, and were assayed for RNase and trypsin-activatable proteolytic (TAPase) activity. The zymogen fraction accounted for 30 to 40 per cent of the total TAPase and RNase activities, and its specific enzymatic activities were 4 to 10 times higher than those of any other cell fraction. The zymogen fraction was cytologically heterogeneous; zymogen granules and mitochondria represented its main components. More homogeneous zymogen fractions, obtained by successive washing or by separation in a discontinuous density-gradient, had specific activities 2 to 4 times greater than the crude zymogen fractions. Chymotrypsinogen was isolated by column chromatography from pancreas homogenates and derived cell fractions. The largest amount was recovered in the zymogen fraction. The final supernatant had properties similar to those of the trypsin inhibitor described by Kunitz and Northrop.

scholarly journals 3β-Hydroxy steroid dehydrogenase activity in the mitochondria of rat adrenal homogenates

1971 ◽  
Vol 125 (4) ◽  
pp. 983-989 ◽  
Author(s):  
R S Basch ◽  
M J Finegold
Keyword(s):  
Electron Microscopy ◽  
Microsomal Fraction ◽  
Total Activity ◽  
Mitochondrial Fraction ◽  
Differential Centrifugation ◽  
Radioisotopic Method ◽  
Cell Fractions ◽  
Kinetics Of ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

The activity of 3β-hydroxy steroid dehydrogenase (EC 1.1.1.51) in the mitochondrial fraction of rat adrenal homogenates was approx. 31% of the total activity recovered after differential centrifugation and washing of the particulate fractions. Some 45% of the total activity was found in the microsomal fraction. The activity was assayed by a radioisotopic method devised in this laboratory for the purpose of studying small quantities of tissue and cell fractions. Satisfactory separation of the two fractions was demonstrated by electron microscopy of the pellets and by comparative recoveries of RNA, steroid 21-hydroxylase and cytochrome c oxidase in the various compartments. Analyses of the kinetics of the enzyme activity in the two fractions revealed no significant differences in apparent Km for pregnenolone, dehydroepiandrosterone or NAD+, but demonstrated a distinct difference in the Km for NADP+. pH optima and susceptibility to cyanoketone inhibition were similar in both fractions.

scholarly journals A Cytochemical Study on the Pancreas of the Guinea Pig

1960 ◽  
Vol 7 (4) ◽  
pp. 619-630 ◽  
Author(s):  
Philip Siekevitz ◽  
George E. Palade
Keyword(s):  
Guinea Pig ◽  
Microsomal Fraction ◽  
Specific Radioactivity ◽  
Early Time ◽  
Acid Extraction ◽  
Time Intervals ◽  
Cytochemical Study ◽  
Cell Compartments ◽  
Exocrine Cell ◽  
Cell Fractions

Chymotrypsinogen synthesis in the exocrine cell of the guinea pig pancreas was studied under the following conditions: Animals fed after a fast of ∼48 hours received ∼1 hour after feeding an intravenous injection of DL-leucine-1-C14. At various time intervals (1 to 45 minutes) after the injection, the glands were removed and fractionated into a series of cell fractions of known cytological significance. Ten to twelve animals were used for each time point. From each cell fraction, the chymotrypsinogen was isolated by acid extraction and purified by (NH4)2SO4 fractionation, isoelectric precipitation, and chromatography. Because of the minuteness of the quantities involved, chymotrypsinogen amounts were calculated from enzymatic activity figures, and a carrier method was used to precipitate and count the enzyme. The chymotrypsinogen isolated from the attached ribonucleoprotein particles of the microsomal fraction had the highest specific radioactivity at the early time points (1 to 3 minutes). After long intervals (at 15 to 45 minutes), the specific radioactivity of the enzyme increased in the microsomal contents and finally in the zymogen granules. The results are compatible with the view that the chymotrypsinogen is synthesized in or on the attached RNP particles and subsequently transported to other cell compartments.

Digestive end products mobilize secretory proteins from subcellular stores in the pancreas

1981 ◽  
Vol 241 (1) ◽  
pp. G67-G73
Author(s):  
J. H. Grendell ◽  
S. S. Rothman
Keyword(s):  
Pancreatic Tissue ◽  
Pancreatic Acinar Cell ◽  
Supernatant Fraction ◽  
Tissue Homogenate ◽  
Secretory Proteins ◽  
Zymogen Granule ◽  
Zymogen Granules ◽  
Amylase Release ◽  
End Products ◽  
Effective Level

Two digestive end products, D-glucose and L-lysine, produced substantial concentration-dependent release of amylase and trypsinogen, respectively, from subcellular storage pools into a postmicrosomal supernatant fraction of rat pancreatic tissue homogenate. This process was selective in that D-glucose did not lead to trypsinogen release, while L-lysine did not effect amylase. An analogue of D-glucose, 2-deoxy-D-glucose, was much less potent than D-glucose on an equimolar basis. Half-maximal release for both end-product enzyme pairs occurred at concentrations within the range of normal plasma values for these end products in the rat. Although amylase release reached an apparent plateau when the concentration of glucose was increased beyond the maximally effective level, lysine concentrations higher than that maximally effective resulted in a fall in trypsinogen release that ultimately returned (at 3.0 mM L-lysine) to the level seen in its absence. When isolated zymogen granules were exposed to the same concentrations of D-glucose or L-lysine, a similar pattern of release was seen, indicating that the zymogen granules are a source of the enzymes released from the particulate phase of the homogenates. These findings can be explained most simply by the selective movement of digestive enzymes across zymogen granule membranes in response to the presence of appropriate end products. They are also consistent with the concept that digestive end products can act rapidly and directly on the pancreatic acinar cell to regulate the mixture of enzymes secreted in response to the specific hydrolytic needs of a meal.

scholarly journals A Cytochemical Study on the Pancreas of the Guinea Pig

1958 ◽  
Vol 4 (3) ◽  
pp. 309-318 ◽  
Author(s):  
Philip Siekevitz ◽  
George E. Palade
Keyword(s):  
Endoplasmic Reticulum ◽  
Guinea Pigs ◽  
Intracellular Transport ◽  
Enzymatic Activities ◽  
Differential Centrifugation ◽  
Zymogen Granules ◽  
Cytochemical Study ◽  
First Case ◽  
Microsomal Membranes ◽  
Exocrine Cell

Microsomes were isolated from the pancreas of starved and fed guinea pigs. In the first case, the gland was removed from animals starved for 48 hours; in the second, the pancreas was excised 1 hour after the beginning of a meal that ended a fast of 48 hours. These are referred to below as fed animals. In both cases the tissue was homogenized in 0.88 M sucrose and the microsomes obtained by centrifuging the mitochondrial supernatant at 105,000 g for 60 minutes. In starved animals the content of the endoplasmic reticulum of the exocrine cells and the content of the microsomes were found to be of low or moderate density. In fed guinea pigs the cavities of the reticulum frequently contained dense intracisternal granules and the microsomes were distinguished by a content of high density sometimes in the form of recognizable intracisternal granules. In starved animals, the microsomes were found to account for 5 to 20 per cent of the trypsin-activatable proteolytic activity and ribonuclease activity of the whole cell, whereas in fed animals they contained uniformly almost 30 per cent of these activities. In fed animals the dense, cohesive content of the microsomes (intracisternal granules) could be isolated by breaking up the microsomes with dilute (0.1 per cent) deoxycholate solutions and separating microsomal subfractions by differential centrifugation. The specific enzymatic activities of a heavy microsomal subfraction rich in intracisternal granules were almost equal to those of isolated purified zymogen granules. The ribonucleoprotein particles attached to the microsomal membranes could be isolated by the same technique and found also to exhibit some of the same enzymatic activities. Corresponding subfractions isolated from the microsomes of starved animals were considerably less active. The relevance of these findings for the synthesis and intracellular transport of protein in the exocrine cell of the pancreas is discussed.

Effect of ADH on Rat Renal Papilla, an Ultrastructural and Cytochemical Study

1973 ◽  
Vol 31 ◽  
pp. 410-411
Author(s):  
C. N. Sun ◽  
H. J. White ◽  
E. J. Towbin
Keyword(s):  
Electron Microscopy ◽  
Diabetes Insipidus ◽  
Renal Papilla ◽  
Milk Diet ◽  
Intercellular Spaces ◽  
Cytochemical Study ◽  
Collecting Tubule ◽  
Concentrate Urine ◽  
Staining Techniques ◽  
Collecting Tubules

Diabetes insipidus and compulsive water drinking are representative of two categories of antidiuretic hormone (ADH) lack. We studied a strain of rats with congenital diabetes insipidus homozygote (DI) and normal rats on an isocaloric fortified dilute milk diet. In both cases, the collecting tubules could not concentrate urine. Special staining techniques, Alcian Blue-PAS for light microscopy and lanthanum nitrate for electron microscopy were used to demonstrate the changes in interstitial mucopolysaccharides (MPS). The lanthanum staining was done according to the method of Khan and Overton.Electron microscopy shows cytoplasmic lesions, vacules, swelling and degenerating mitochondria and intercellular spaces (IS) in the collecting tubule cells in DI and rats on milk diet.

Electron Microscopy of Human Gallstones

1971 ◽  
Vol 29 ◽  
pp. 296-297
Author(s):  
C. Wolpers ◽  
R. Blaschke
Keyword(s):  
Electron Microscopy ◽  
Room Temperature ◽  
Bile Pigment ◽  
X Ray Diffraction ◽  
Triclinic Crystal System ◽  
X Ray ◽  
Follow Up Study ◽  
Infra Red ◽  
Main Components

Scanning microscopy was used to study the surface of human gallstones and the surface of fractures. The specimens were obtained by operation, washed with water, dried at room temperature and shadowcasted with carbon and aluminum. Most of the specimens belong to patients from a series of X-ray follow-up study, examined during the last twenty years. So it was possible to evaluate approximately the age of these gallstones and to get information on the intensity of growing and solving.Cholesterol, a group of bile pigment substances and different salts of calcium, are the main components of human gallstones. By X-ray diffraction technique, infra-red spectroscopy and by chemical analysis it was demonstrated that all three components can be found in any gallstone. In the presence of water cholesterol crystallizes in pane-like plates of the triclinic crystal system.

INCORPORATION OF IODIDE INTO ORGANIC COMPOUNDS IN THE GUINEA PIG THYROID

1963 ◽  
Vol 43 (1) ◽  
pp. 110-118 ◽  
Author(s):  
R. Ekholm ◽  
T. Zelander ◽  
P.-S. Agrell
Keyword(s):  
Guinea Pig ◽  
Protein Binding ◽  
Organic Compounds ◽  
Guinea Pigs ◽  
Microsomal Fraction ◽  
Thyroid Tissue ◽  
Particulate Fraction ◽  
Supernatant Fraction ◽  
Hydrolysis Of

ABSTRACT Guinea pigs, kept on a iodine-sufficient diet, were injected with Na131I and the thyroids excised from 45 seconds to 5 days later. The thyroid tissue was homogenized and separated into a combined nuclear-mitochondrial-microsomal fraction and a supernatant fraction by centrifugation at 140 000 g for one hour. Protein bound 131iodine (PB131I) and free 131iodide were determined in the fractions and the PB131I was analysed for monoiodotyrosine (MIT), diiodotyrosine (DIT) and thyroxine after hydrolysis of PB131I. As early as only 20 minutes after the Na131I-injection almost 100% of the particulate fraction 131I was protein bound. In the supernatant fraction the protein binding was somewhat less rapid and PB131I values above 90% of total supernatant 131I were not found until 3 hours after the injection. In all experiments the total amount of PB131I was higher in the supernatant than in the corresponding particulate fraction. The ratio between supernatant PB131I and pellet PB131I was lower in experiments up to 3 minutes and from 2 to 5 days than in experiments of 6 minutes to 20 hours. Hydrolysis of PB131I yielded, even in the shortest experiments, both MIT and DIT. The DIT/MIT ratio was lower in the experiments up to 2 hours than in those of 3 hours and over.

Secretagogue-induced changes in subcellular Ca2+ distribution in isolated pancreatic acini

1981 ◽  
Vol 240 (2) ◽  
pp. G130-G140
Author(s):  
R. L. Dormer ◽  
J. A. Williams
Keyword(s):  
Atomic Absorption Spectrometry ◽  
High Speed ◽  
Microsomal Fraction ◽  
Absorption Spectrometry ◽  
Subcellular Fractionation ◽  
Differential Centrifugation ◽  
Zymogen Granules ◽  
Pancreatic Acini ◽  
Induced Changes ◽  
Stimulation Of

In a prior study, we demonstrated that pancreatic secretagogues increased both the uptake into and washout of 45Ca2+ from isolated mouse pancreatic acini. The net result of these processes was an initial fall in total acinar cell Ca2+ content. In the present study, we have employed subcellular fractionation of acini under conditions that minimized posthomogenization redistribution of Ca2+ in order to localize those organelles involved in intracellular Ca2+ fluxes. Homogenization and differential centrifugation of acini, preloaded with 45Ca2+ and subjected to a period of washout, showed that carbachol induced an increased loss of 45Ca2+ from all fractions isolated. The high-speed microsomal fraction lost 45Ca2+ to a greater extent than did whole acini; measurement of total Ca2+ by atomic absorption spectrometry showed a net loss of Ca2+ from this fraction. Purification of the lower-speed fractions indicated that carbachol increased 45Ca2+ exchange with both zymogen granules and mitochondria, but net Ca2+ levels in these organelles were unchanged. It was concluded that stimulation of pancreatic acini by carbachol results in the release of calcium from a microsomal compartment leading to a rise in cytoplasmic Ca2+, increased exchange with granule and mitochondrial Ca2+, and increased efflux of Ca2+ from the cell.

Carbohydrase activity in the pancreatic tissue and small intestine mucosa of sheep fed dried-grass or ground maize-based diets

1985 ◽  
Vol 104 (2) ◽  
pp. 435-443 ◽  
Author(s):  
A. N. Janes ◽  
T. E. C. Weekes ◽  
D. G. Armstrong
Keyword(s):  
Small Intestine ◽  
Amylase Activity ◽  
Total Activity ◽  
Pancreatic Tissue ◽  
Proximal Region ◽  
Small Intestinal Mucosa ◽  
Intestine Mucosa ◽  
Ruminant Animals ◽  
Specific Activities ◽  
Small Intestinal

SummaryTwo groups of six sheep were fed either dried-grass or ground maize-based diets for at least 4 weeks before slaughter. Samples of the small intestinal mucosa and spancreatic tissue were assayed for a-amylase, glucoamylase, maltase and oligo-l,6-glucosidase.The pancreatic tissue contained high activities of α-amylase and much lower activities of glucoamylase, maltase and oligo-1,6-glucosidase. There was no effect of diet on the specific activities of any of these enzymes in the pancreatic tissue.The activity of α-amylase adsorbed on to the mucosa of the small intestine was greatest in the proximal region of the small intestine, the activity generally declining with increasing distance away from the pylorus. There was no diet effect on the absorbed α-amylase activity.Similar patterns of distribution along the small intestine were observed for maltase, glucoamylase and oligo-1,6-glucosidase with the highest activities in t he jejunum. There was no overall effect of diet on glucoamylase or maltase specific activities and glucoamylase total activity, although the total activities of maltase and oligo-1,6-glucosidase were significantly greater for the sheep fed the ground maize-based diet (P < 0·05).It is suggested that ruminant animals may be capable of digesting large amounts of starch in the small intestine through an adaptation in the activity of the host carbohydrases.

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