Ultrastructural Pathology
ISSN: 0191-3123 (Print) 1521-0758 (Online) Journal homepage: http://www.tandfonline.com/loi/iusp20
Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland J. D. Harrison To cite this article: J. D. Harrison (1991) Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland, Ultrastructural Pathology, 15:2, 185-188, DOI: 10.3109/01913129109016237 To link to this article: http://dx.doi.org/10.3109/01913129109016237
Published online: 10 Jul 2009.
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Date: 22 April 2016, At: 22:17
Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland J. D. Harrison, PhD
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
Department of Oral Pathology, The RayW IT&&&&, King:? College S&()()l of Medicine and Dentistry, London, England
A pleomorphic adenoma was examined by electron microscopy. Calcification was found in a lumen and in epithelial cells and consisted of needle-shaped crystals that contained calcium and phosphorus and were probably apatite. Small collections of crystals in the lumen, which were often associated with membranous cellular debris, appeared to form larger calcified masses by fusion. Collections of crystals were seen in vacuoles in adluminal cells. The calcification in the lumen may have started on membranous cellular debris, and that in the adluminal cells may have arisen either by endocytosis of luminal material or by autophagocytosis. The calcification appears t o relate t o the old age of the tumor.
KEY WORDS: calctnosts, mixed tumor, parotid neoplasm, pleomorphtc adenoma, salivary glands. -
INTRODUCTION Pleomorphic adenomas only occasionally contain calcification,’ which may explain why it has not been described ultrastructurally. Therefore the fortuitous ultrastructural finding of calcification in a pleomorphic adenoma appears t o be of particular interest.
MATERIALS AND METHODS A tumor of the left parotid gland was found in a 47-year-old woman with a history of approximately 1 0 years of swelling. Examination of the surgical specimen, which was received immediately upon removal, showed an encapsulated tumor of I gratefully acknowledge the financial support of the Medical Research Council and the technical assistance of K J Davies, Katherine L Paterson. and P S A Rowley Request reprints from J D Harrison, Department of Oral Pathology, The Rayne Institute, 123 Coldharbour Lane, London, SE5 9NU. England
about 2 cm diameter and surrounding parotid gland. Pieces of less than 2 mm greatest dimension were immersed in a solution of 5% glutaraldehyde, 4 % formaldehyde prepared from paraformaldehyde, 0 . 0 5 % CaCI,, and 0.08 M cacodylate buffer (pH 7.2) at room temperature for 2 hours.’ They were rinsed in 7.5% sucrose in 0.05 M cacodylate buffer (pH 7.2) a t 0-4OC, dehydrated, cleared, and embedded in Araldite. The pieces were not osmicated because they were intended for mucosubstance histochemistry. Ultrathin sections were stained with uranyl acetate and lead citrate for morphological examination. Unstained ultrathin sections were examined in a JEOL IOOC scanning transmission electron microscope equipped with a Link 860 energy-dispersive x-ray analysis system for elemental analysis. It was operated a t an accelerating voltage of 60 KV, and analysis was undertaken in the spot mode. Histological examination of formalinfixed, wax-embedded tissue showed the
Ultrastructural Pathology, 15: 185-1 88, 1991 Copyright
0 1991 by Hemisphere Publishing Corporation
185
J. D. Harrison
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
186
FIG. 1 Calcification in the lumen and an adluminal cell (A). Small collections of crystals are associated with membranous luminal debris (MI, and larger masses of crystals consist of fused small collections (arrows). Two lamellar collections of crystals are present in a vacuole in an adluminal cell (A). Vacuoles containing debris (asterisks) are present nearby, x 5,300.
Calcification in a Pleomorphic Adenoma
tumor t o be a pleornorphic adenorna, and did not reveal calcification.
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
RESULTS Calcification was seen ultrastructurally in sections from one piece only. It was seen in a lumen, in some of the associated epithelial cells (Figs. 1 and 21, in degenerate cells, and associated with cellular debris in the adjacent strorna. The calcification consisted of crystals that usually appeared shaped like needles or occasionally as gray plates. Small collections of crystals were rounded, sometimes lamellar, and some-
187 times associated with membraneous debris in the lumen, which contained widespread cellular debris (Fig. 1). Larger calcified masses appeared t o consist of fused small collections of crystals. Collections of crystals were seen in vacuoles in adluminal cells, in which there were also vacuoles containing debris, and secretory granules (Figs. 1 and 2). Collections of crystals and individual crystals were seen lying free in degenerate cells. Microanalytical probes of the calcification revealed peaks at 3.68 and 4.00 keV corresponding t o the ka and KO lines of calcium and 2.01 keV corresponding to
FIG. 2 Adluminal cell containing a collection of crystals together with some debris in a vacuole (arrow). The crystals are mainly needle-shaped, and some are plateshaped. Secretory granules (Sland vacuoles containing debris (asterisks) are present, x 18,200.
188
the ka! line of phosphorus. These peaks were not seen when adjacent structures were analyzed.
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
DISCUSSION Thackray and Lucas’ mentioned that calcification may be a feature of old pleomorphic adenomas and occurs in lumina and stroma associated with the epithelium, which was so in the present tumor. The membranous component of the cellular debris in the lumen would form an ideal nidus for the start of calcification, particularly if alkaline phosphatase, which has been demonstrated lining l ~ m i n a , ~ was . ~ associated with it5; and the calcification of degenerate cells is well e~tablished.~ Possibly in older tumors there is a greater chance of trauma or ischemia leading to cellular degeneration. The calcification in the adluminal cells is similar in appearance to microliths found in the parenchymal cells of feline and human salivary glands, which appear to form in autophagosomes that contain degenerate membranes and accumulated calcium and phosphate, and these microliths appear to pass into lumina.6-8 However, adluminal cells of the pleomorphic adenoma appear to be endocytose luminal material,’ so their microliths could be in endocytotic vacuoles and of luminal origin. The morphological appearance of the crystals and the microanalytical detection of calcium and phosphorus in them are similar to the properties of the crystals of the microliths of salivary glands”’ and breast carcinomas,1o which appear to be apatite. Ahrned” suggested that the calcification in breast carcinomas, which is common, may be the consequence of an active secretory process by the tumor cells, and involves an increased concentration of calcium ions in the presence of phospholipids and phosphoproteins such as casein t o act as a nidus for calcification. Possibly the level of calcium ions in the pleomorphic ad-
J. D. Harrison
enoma is usually too low for calcification, which would explain its infrequency. Calcification in the pleomorphic adenoma possibly is caused by the simultaneous occurrence of the following factors: an increased concentration of calcium ions, perhaps from secretory granules; exposed membranes, perhaps from necrosis or autophagocytosis, to act as a nidus; and alkaline phosphatase, which is only occasionally found.3t4
REFERENCES 1 . Thackray AC, Lucas RE. Pleomorphic adenoma (mixed tumor). In: Tumors of the Major Salivary Glands. Washington, DC: Armed Forces Institute of Pathology; 1974:16-39. 2. Karnovsky MJ. A formaldehydeglutaraldehyde fixative of high osmolality for use in electron microscopy. J Cel/ Biol.
1965;27:137A-l38A. 3. Harrison JD, Auger DW. Ultrastructural cytochemistry of phosphatases in the ductal component of pleomorphic adenoma of human parotid and submandibular salivary glands. Histochem J. 1982;14:703-711. 4. Auger DW. Ultrastructural and Histochemical Studies on Pleomorphic Adenoma of Parotid and Submandibular Salivary Glands of Man. University of London; 1985.PhD thesis. 5. Anderson HC. Calcific diseases. A concept. Arch Path01 Lab Med. 1 983;107:341-348. 6. Epivatianos A, Harrison JD, Garrett JR, Davies KJ, Senkus R. Ultrastructural and histochemical observations on intracellular and luminal microcalculi in the feline sublingual salivary gland. J Oral Pathol. 1986;15:513-517. 7. Epivatianos A, Harrison JD, Dimitriou T. Ultrastructural and histochemical observations on microcalculi in chronic submandibular sialadenitis. J Oral Pathol. 1987;16:514-517. 8. Epivatianos A, Harrison JD. The presence of microcalculi in normal human submandibular and parotid salivary glands. Arch Oral Biol.
1989;34:261-265. 9. Harrison JD, Auger DW. Ultrastructural observations on luminal structures of pleomorphic adenoma of parotid and submandibular salivary glands of man. Virchows Arch (A).
1989;415:559-563. 10. Ahmed A. Calcification in human breast carcinomas: ultrastructural observations. J Pathol.
1975;1 17:247-251. Accepted in revised form October 15, 1990.
ISSN: 0191-3123 (Print) 1521-0758 (Online) Journal homepage: http://www.tandfonline.com/loi/iusp20
Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland J. D. Harrison To cite this article: J. D. Harrison (1991) Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland, Ultrastructural Pathology, 15:2, 185-188, DOI: 10.3109/01913129109016237 To link to this article: http://dx.doi.org/10.3109/01913129109016237
Published online: 10 Jul 2009.
Submit your article to this journal
Article views: 13
View related articles
Citing articles: 1 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=iusp20 Download by: [University of California, San Diego]
Date: 22 April 2016, At: 22:17
Ultrastructural Observation of Calcification in a Pleomorphic Adenoma of the Parotid Gland J. D. Harrison, PhD
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
Department of Oral Pathology, The RayW IT&&&&, King:? College S&()()l of Medicine and Dentistry, London, England
A pleomorphic adenoma was examined by electron microscopy. Calcification was found in a lumen and in epithelial cells and consisted of needle-shaped crystals that contained calcium and phosphorus and were probably apatite. Small collections of crystals in the lumen, which were often associated with membranous cellular debris, appeared to form larger calcified masses by fusion. Collections of crystals were seen in vacuoles in adluminal cells. The calcification in the lumen may have started on membranous cellular debris, and that in the adluminal cells may have arisen either by endocytosis of luminal material or by autophagocytosis. The calcification appears t o relate t o the old age of the tumor.
KEY WORDS: calctnosts, mixed tumor, parotid neoplasm, pleomorphtc adenoma, salivary glands. -
INTRODUCTION Pleomorphic adenomas only occasionally contain calcification,’ which may explain why it has not been described ultrastructurally. Therefore the fortuitous ultrastructural finding of calcification in a pleomorphic adenoma appears t o be of particular interest.
MATERIALS AND METHODS A tumor of the left parotid gland was found in a 47-year-old woman with a history of approximately 1 0 years of swelling. Examination of the surgical specimen, which was received immediately upon removal, showed an encapsulated tumor of I gratefully acknowledge the financial support of the Medical Research Council and the technical assistance of K J Davies, Katherine L Paterson. and P S A Rowley Request reprints from J D Harrison, Department of Oral Pathology, The Rayne Institute, 123 Coldharbour Lane, London, SE5 9NU. England
about 2 cm diameter and surrounding parotid gland. Pieces of less than 2 mm greatest dimension were immersed in a solution of 5% glutaraldehyde, 4 % formaldehyde prepared from paraformaldehyde, 0 . 0 5 % CaCI,, and 0.08 M cacodylate buffer (pH 7.2) at room temperature for 2 hours.’ They were rinsed in 7.5% sucrose in 0.05 M cacodylate buffer (pH 7.2) a t 0-4OC, dehydrated, cleared, and embedded in Araldite. The pieces were not osmicated because they were intended for mucosubstance histochemistry. Ultrathin sections were stained with uranyl acetate and lead citrate for morphological examination. Unstained ultrathin sections were examined in a JEOL IOOC scanning transmission electron microscope equipped with a Link 860 energy-dispersive x-ray analysis system for elemental analysis. It was operated a t an accelerating voltage of 60 KV, and analysis was undertaken in the spot mode. Histological examination of formalinfixed, wax-embedded tissue showed the
Ultrastructural Pathology, 15: 185-1 88, 1991 Copyright
0 1991 by Hemisphere Publishing Corporation
185
J. D. Harrison
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
186
FIG. 1 Calcification in the lumen and an adluminal cell (A). Small collections of crystals are associated with membranous luminal debris (MI, and larger masses of crystals consist of fused small collections (arrows). Two lamellar collections of crystals are present in a vacuole in an adluminal cell (A). Vacuoles containing debris (asterisks) are present nearby, x 5,300.
Calcification in a Pleomorphic Adenoma
tumor t o be a pleornorphic adenorna, and did not reveal calcification.
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
RESULTS Calcification was seen ultrastructurally in sections from one piece only. It was seen in a lumen, in some of the associated epithelial cells (Figs. 1 and 21, in degenerate cells, and associated with cellular debris in the adjacent strorna. The calcification consisted of crystals that usually appeared shaped like needles or occasionally as gray plates. Small collections of crystals were rounded, sometimes lamellar, and some-
187 times associated with membraneous debris in the lumen, which contained widespread cellular debris (Fig. 1). Larger calcified masses appeared t o consist of fused small collections of crystals. Collections of crystals were seen in vacuoles in adluminal cells, in which there were also vacuoles containing debris, and secretory granules (Figs. 1 and 2). Collections of crystals and individual crystals were seen lying free in degenerate cells. Microanalytical probes of the calcification revealed peaks at 3.68 and 4.00 keV corresponding t o the ka and KO lines of calcium and 2.01 keV corresponding to
FIG. 2 Adluminal cell containing a collection of crystals together with some debris in a vacuole (arrow). The crystals are mainly needle-shaped, and some are plateshaped. Secretory granules (Sland vacuoles containing debris (asterisks) are present, x 18,200.
188
the ka! line of phosphorus. These peaks were not seen when adjacent structures were analyzed.
Downloaded by [University of California, San Diego] at 22:17 22 April 2016
DISCUSSION Thackray and Lucas’ mentioned that calcification may be a feature of old pleomorphic adenomas and occurs in lumina and stroma associated with the epithelium, which was so in the present tumor. The membranous component of the cellular debris in the lumen would form an ideal nidus for the start of calcification, particularly if alkaline phosphatase, which has been demonstrated lining l ~ m i n a , ~ was . ~ associated with it5; and the calcification of degenerate cells is well e~tablished.~ Possibly in older tumors there is a greater chance of trauma or ischemia leading to cellular degeneration. The calcification in the adluminal cells is similar in appearance to microliths found in the parenchymal cells of feline and human salivary glands, which appear to form in autophagosomes that contain degenerate membranes and accumulated calcium and phosphate, and these microliths appear to pass into lumina.6-8 However, adluminal cells of the pleomorphic adenoma appear to be endocytose luminal material,’ so their microliths could be in endocytotic vacuoles and of luminal origin. The morphological appearance of the crystals and the microanalytical detection of calcium and phosphorus in them are similar to the properties of the crystals of the microliths of salivary glands”’ and breast carcinomas,1o which appear to be apatite. Ahrned” suggested that the calcification in breast carcinomas, which is common, may be the consequence of an active secretory process by the tumor cells, and involves an increased concentration of calcium ions in the presence of phospholipids and phosphoproteins such as casein t o act as a nidus for calcification. Possibly the level of calcium ions in the pleomorphic ad-
J. D. Harrison
enoma is usually too low for calcification, which would explain its infrequency. Calcification in the pleomorphic adenoma possibly is caused by the simultaneous occurrence of the following factors: an increased concentration of calcium ions, perhaps from secretory granules; exposed membranes, perhaps from necrosis or autophagocytosis, to act as a nidus; and alkaline phosphatase, which is only occasionally found.3t4
REFERENCES 1 . Thackray AC, Lucas RE. Pleomorphic adenoma (mixed tumor). In: Tumors of the Major Salivary Glands. Washington, DC: Armed Forces Institute of Pathology; 1974:16-39. 2. Karnovsky MJ. A formaldehydeglutaraldehyde fixative of high osmolality for use in electron microscopy. J Cel/ Biol.
1965;27:137A-l38A. 3. Harrison JD, Auger DW. Ultrastructural cytochemistry of phosphatases in the ductal component of pleomorphic adenoma of human parotid and submandibular salivary glands. Histochem J. 1982;14:703-711. 4. Auger DW. Ultrastructural and Histochemical Studies on Pleomorphic Adenoma of Parotid and Submandibular Salivary Glands of Man. University of London; 1985.PhD thesis. 5. Anderson HC. Calcific diseases. A concept. Arch Path01 Lab Med. 1 983;107:341-348. 6. Epivatianos A, Harrison JD, Garrett JR, Davies KJ, Senkus R. Ultrastructural and histochemical observations on intracellular and luminal microcalculi in the feline sublingual salivary gland. J Oral Pathol. 1986;15:513-517. 7. Epivatianos A, Harrison JD, Dimitriou T. Ultrastructural and histochemical observations on microcalculi in chronic submandibular sialadenitis. J Oral Pathol. 1987;16:514-517. 8. Epivatianos A, Harrison JD. The presence of microcalculi in normal human submandibular and parotid salivary glands. Arch Oral Biol.
1989;34:261-265. 9. Harrison JD, Auger DW. Ultrastructural observations on luminal structures of pleomorphic adenoma of parotid and submandibular salivary glands of man. Virchows Arch (A).
1989;415:559-563. 10. Ahmed A. Calcification in human breast carcinomas: ultrastructural observations. J Pathol.
1975;1 17:247-251. Accepted in revised form October 15, 1990.
