Immunofluorescence and electron-microscopic observations of intermediate cells in the pancreas of mice, rats and humans.

The immunohistochemical method supplemented with a transmission electron-microscopic survey was used to investigate the pancreas of mice, rats and humans to demonstrate intermediate cells. Indirect immunofluorescence was applied to paraffin sections of pancreatic samples of normal rats and autopsied humans to localize amylase in the islets and excretory duct cells. The amylase immunofluorescent spots were occasionally detected in the apices and perinuclear cytoplasm of few islet and excretory duct cells, which completely disappeared in the specificity control sections. These cells, respectively, represented the islet amylase and duct amylase intermediate cells. The ultrastructural survey was conducted on pancreatic tissue samples from normal and streptozotocin-diabetic mice and rats to prove the existence of these intermediate cells. Three morphologically distinct types of intermediate cells were recognized and characterized into beta-acinar, alpha-acinar and duct acinar types. The beta-acinar and alpha-acinar cells contained numerous secretory granules with few large and dense, membrane-bound zymogen granules. No clear difference in the frequency of these cells between normal and diabetic tissues was qualitatively observed in either mice or rats. The duct acinar cells were occasionally seen in few small intercalated ducts of the normal mouse pancreas, which contained few dense, membrane-bound zymogen granules adjacent to a prominent Golgi complex. On the other hand, no delta-acinar, pancreatic polypeptide (PP)-acinar or acinoislet cells were observed in both normal and diabetic animals. In conclusion, the present study provides immunohistochemical and morphological evidence on the existence of different types of intermediate cells in the normal and streptozotocin-diabetic pancreas of mice, rats and humans. Although no prevalence of these cells in the streptozotocin-diabetic pancreas was observed qualitatively, their existence throws light on the possible transformation between structurally and functionally different pancreatic cells when subjected to pathological trauma. This concept might be of great significance in beta-cell regeneration in insulin-dependent diabetes mellitus.

A comparative immunohistochemical study on amylase localization in the rat and human exocrine pancreas.

OBJECTIVE: To localize amylase enzyme immunohistochemically in the pancreatic acinar cells of rats and humans using polyclonal sheep anti-human amylase antibody, and to compare between the intensities of their amylase-immunostaining. METHODS: Indirect immunofluorescence method was applied on formaldehyde-fixed, and paraffin-embedded pancreatic sections obtained from adult male Wistar rats and autopsied human samples. Primary incubation was performed using sheep anti-amylase antibody followed by secondary incubation with fluorescein isothiocyanate-labeled rabbit anti-sheep IgG serum. Control tests of amylase immunospecificity were also undertaken either by incubation with primary antibodies previously pre-adsorbed with an excess of human pancreatic amylase, or only with secondary antibodies. RESULTS: The amylase immunofluorescence was positively and homogenously detected in all acinar cells of both rat and human pancreatic stained sections. The immunostaining was clearly demonstrated in the cell apices and peri-nuclear areas, but it was consistently brighter and more intense in the human acinar cells compared with that of the rat pancreas. Control tests of amylase immunofluorescence revealed the specificity of the antibodies applied for amylase localization in rat and human pancreas. CONCLUSION: Although many previous immunohisto- and cytochemical reports have successfully localized amylase in the pancreas of different mammalian species, but all of them have used locally prepared anti-amylase antibodies. The present report successfully illustrates immuno-localization of amylase in the pancreatic acinar cells of rats and humans using commercial polyclonal sheep anti-human pancreatic amylase antibodies, and also suggests their useful application in the immunochemical studies on various mammalian species. Additionally, the results indicate a structural similarity between the human and rat pancreatic amylases, a concept required further exploration.

The ultrastructure of primary cilia in the endocrine and excretory duct cells of the pancreas of mice and rats.

A primary cilium was frequently observed in the endocrine alpha, beta and delta cells, as well as in the excretory duct cells of the pancreas of normal mice and rats. The characteristic components of the cilium including the basal body, axoneme (shaft), and terminal part were clearly recognizable. The basal body or distal centriole surrounded by Golgi vesicles was perpendicularly oriented to the proximal centriole, and a dense striated band was seen filling the gap between them. The microtubules of the basal body consisted of nine peripheral triplets exhibiting a 9 + 0 pattern, an appearance similar to that of the proximal centriole. Rootlets, basal feet and alar sheets associated with the basal body were occasionally seen. The axoneme usually consisted of a 9 + 0 pattern of microtubule doublets, but other irregular patterns of 7 + 2, 7 + 3, and 8 + 1 were also seen. The microtubules in the terminal part of the cilium became fewer in number and had no peculiar arrangement. The cilium of the endocrine cells always projected into the intercellular canaliculus and was covered by the ciliary sheath, and occasionally, double cilia were visualized in the vicinity of beta cells. In the excretory duct cells, the cilium showed similar features, but it was slightly longer and always projected into the dense secretory content of duct lumen. On the other hand, no primary cilium was ever observed in the acinar cells of mouse and rat pancreas. In conclusion, the present study describes the morphology of primary cilia and its associated components in the endocrine and excretory duct cells of the pancreas of mice and rats. The findings suggest that the primary cilium should be considered as a constant intracellular organelle though its function and significance remain speculative.

An ultrastructural study on the effect of streptozotocin on the islets of Langerhans in mice.

The ultrastructural changes in the morphology of the islets of Langerhans in response to streptozotocin were studied in the mice pancreas. Male white albino CSI mice were given a single intravenous injection of 75 mg kg(-1) body weight streptozotocin, and were sacrificed at different time intervals up to 48 h following the treatment. Their pancreases were excised and randomly processed for electron microscopic examination. Hyperglycaemia and glucosuria were detected 8 h after treatment, became remarkably high at 24 h and persisted then after. Light and electron microscopic examination of the islets of Langerhans from treated mice revealed an early chromatin aggregation and cytoplasmic vesiculation in the central B cells during the first 2 h of treatment. Nuclear shrinkage and pyknosis with swelling of mitochondria and endoplasmic reticulum were evident 8 h later, and lysis of B cells occurred 12 h after treatment. The morphology of A and D cells at the margin of the islets and in between B cell debris looked perfectly unaltered. Macrophage infiltration among lytic B cells was seen 24 h after drug administration, which contained clear and large phagocytic vacuoles. The necrobiotic and phagocytic figures disappeared from the pancreatic sections of 48 h treated mice, and the islets were smaller in size and consisted entirely of intact A and D cells with occasional degranulated B cells. No features of apoptosis were ever recorded, and the exocrine pancreatic tissue was protected from the effect of streptozotocin. In conclusion, the present study illustrates the sequence of morphological changes that occurs in the islets of Langerhans of mice after streptozotocin administration. It also confirms that streptozotocin at a high single dose in mice produces a specific necrosis of B cells with no evidence of apoptotic figures as another mechanism of cell death.

Ultrastructural study of the gastric mucosa and Helicobacter pylori in duodenal ulcer patients.

OBJECTIVE: To investigate the relationship between Helicobacter pylori and gastric mucosa in control and duodenal ulcer patients at the electron microscopic level. METHODS: Three antral biopsies were taken from each of 20 normal control volunteers and 30 duodenal ulcer patients presented to the gastroenterology unit at Jordan University Hospital for upper endoscopic examination. Each specimen was fixed and processed for electron microscopic study. RESULTS: Two types of Helicobacter pylori were observed and identified by their morphology at electron microscopy. The first one was characterized by double external smooth membranes and homogeneous cytoplasmic contents, and the second type with a characteristic ring-shaped intracytoplasmic vacuole. Electron microscopic examination of normal controls showed normal gastric mucosa and a small number of Helicobacter pylori in 12 out of 20 controls. However, in duodenal ulcer patients, 5 different patterns of interaction between the Helicobacter pylori and gastric mucosa were observed in relation to the severity of the disease. In duodenal ulcer patients, various types of epithelial damage was seen accompanied with a decrease or absence of mucous secretion and with more colonization of bacteria. CONCLUSION: The morphology and pathogenesis of Helicobacter pylori was described in duodenal ulcer patients, and 5 different patterns of contact between Helicobacter pylori and surface epithelium were recognized causing variable degrees of microvillous atrophy and reduced mucous secretion. The vacuolated type of Helicobacter pylori was more adherent to the damaged epithelium and there was a direct relationship between the epithelial damage and bacterial load. In the normal controls, no epithelial damage and scanty bacteria were observed. The various types of epithelial changes of gastric mucosa has initiated more research at electron microscopic level on the immune mechanism of the gastric mucosa to determine the underlying cause of the varying severity of the disease.

Fibroadenoma of the supernumerary breast of the axilla.

Supernumerary breast or polymastia is a well documented anomaly of the breast, and commonly presents along the embryonic milk line extending between the axilla and groin. However, cases of polymastia have been recorded in the face, vulva and perineum. The clinical significances of these anomalies include their susceptibility to inflammatory and malignant changes, and their association with other congenital anomalies of the urinary and cardiovascular systems. The present article reports a case of fibroadenoma developing in the supernumerary breast of the right axilla in a 28 year old woman. Clinical and mammography examination of both breasts revealed no abnormalities and no lymph nodes were detected in the axillae or the neck. No associated urologic or cardiovascular abnormalities were found, and the histopathological examination of the excisional biopsy samples showed a well-defined, capsulated intracanalicular type of fibroadenoma similar to that of eutopic mammary tissue. The article also outlines the common congenital anomalies of the breast, and emphasizes on their proper clinical assessment for any other associated anomaly together with adequate surgical excision and regular follow up of the treated patients.

Correlative morphometric and biochemical study on pancreatic amylase in normal and streptozotocin-diabetic rats.

The number, volume, and size of zymogen granules in pancreatic acinar cells of normal and streptozotocin-diabetic rats were measured using stereological techniques. These morphometric data were then correlated with the amylase activity of the acinar cells. In the normal rats, the acinar cells had a mean volume of 1,253.5 microns 3 and contained 343 zymogen granules, which occupied a volume of 103 microns 3 of the cell (8.28%). In the diabetic rats, the mean acinar cell volume was estimated as 1,017 microns 3 and the cell contained 220 zymogen granules, which occupied a volume of 55.8 microns 3 (5.38%). The cell volume and zymogen granule number and volume were 19, 36, and 46%, respectively, more in normal rat pancreas, while no difference in the size of zymogen granules between normal and diabetic rats was observed. On the other hand, the volume density and numerical densities of acinar cell nuclei were slightly larger in the diabetic rats, but no differences in the nuclear size between normal and diabetic rats were recorded. Biochemically, the amylase activity of diabetic rat pancreas was 37% less than that of normal rats. The present results indicate the impairment of pancreatic amylase production in streptozotocin-diabetic rats, and the correspondence between the morphometrical and the biochemical data indicates that amylase is processed intracellularly in membrane-bound compartments.

Immunohistochemical localization of amylase in peri- and tele-insular acinar cells of the human exocrine pancreas.

An immunohistochemical localization of amylase was demonstrated in human pancreatic acinar cells using a commercial anti-human pancreatic amylase antibody. The immunofluorescence was mainly localized in the cell apices, and some differences in the intensity of the fluorescence was observed among the acinar cells in respect to their location from the islets of Langerhans. The peri-insular acinar cells showed a brighter fluorescence than the cells of tele-insular acini. This inhomogeneity of pancreatic amylase distribution in the human exocrine pancreas adds a further clue to the concept of insulo-acinar interaction.

Morphometric studies on the juxta-insular and tele-insular acinar cells of the pancreas in normal and streptozotocin-induced diabetic rats.

In the present study on the exocrine pancreas, stereological techniques were applied at the level of electron microscopy to confirm morphological differences between juxta-insular and tele-insular acinar cells of normal rats and to evaluate the effect of B cell secretion on these differences in streptozotocin-diabetic rats. As no similar data have been previously published, we conducted a systematic sampling analysis with special reference to the acinar cell volume, the granule content and the nuclear size. In the normal rats, the juxta-insular acinar cell had an average volume of 918 microns3 and contained 420 zymogen granules which occupied 10.2% of the cell. In tele-insular acini, the average cell volume was estimated at 755 microns3 and the cell contained 231 zymogen granules which amounted to 6.8% of the cell. The juxta-insular acinar cell was 18% larger in size and contained 45% more zymogen granules, but no difference was observed in the size of zymogen granules. In addition, the numerical density of cell nuclei was slightly larger in the tele-insular acini in spite of the similarity in the volume density and diameter of the nuclei in the cells of the two areas. In the streptozotocin-induced diabetic rats, no differences were observed in the cell volume and the zymogen granule content between the juxta- and tele-insular acini. These data suggest that the morphological inhomogeneity in the pancreatic acini is caused by the influence of B cell secretion on the activity of the juxta-insular acini.

Changes in the size and number of secretion granules in the rat exocrine pancreas induced by feeding or stimulation in vitro. A morphometric study.

The size, number and volume per cell of secretion granules in rat exocrine pancreas have been measured using stereological techniques. The changes which occur as a result of feeding starved animals (90 min) or stimulating lobular fragments in vitro with carbachol are documented. In fasted animals mean acinar cell volume was estimated as 1670 micron 3 and the cells contained an average of around 450 secretion granules with a corrected mean diameter of 0.70 micron. They occupied around 7% of cell volume. After feeding mean cell volume was about 1300 micron 3 and the cells contained an average of about 190 granules per cell with a mean diameter of 0.58 micron. They occupied 3% of cell volume. A shift in the size frequency distribution of granule diameters occurred as a result of feeding. In vitro experiments in which lobules were induced to secrete with carbachol (10 microM, 3 h, 37 degrees C) had a similar effect. Mean cell volume was reduced from around 1760 micron 3 to 1360 micron 3, mean granule number from around 420 per cell to 180 per cell and the volume density of granules was reduced from about 8% to 3% of cell volume. There was no significant change in mean granule diameter or shift in the size-frequency distribution of granule diameters. Incubation of tissues with cycloheximide (1 mM, 3 h, 37 degrees C) did not prevent secretion by carbachol but it prevented replacement of granules. As a consequence, depletion by carbachol was greater in the presence of cycloheximide, the granules being reduced to around 110 per cell and to only 2.5% of cell volume. We conclude that feeding causes a preferential loss of larger granules and that during secretion replacement of granules occurs. Some of these granules are smaller than those evident in the glands of starved animals.