A Sparsely Granulated, Nonsecreting Adenoma of the Pars Intermedia Associated with Galactorrhea in a Male Rhesus Monkey (Macaca mulatta)

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1 Vet. Pathol (1983) A Sparsely Granulated, Nonsecreting Adenoma of the Pars Intermedia Associated with Galactorrhea in a Male Rhesus Monkey (Macaca mulatta) L. V. CHALIFOUX, J. J. MACKEY, and N. W. KING Harvard Medical School, New England Regional Primate Research Center. Southborough, MA Abstract. A pituitary mass was found at necropsy ofa malemacma muluttu. Hematoxylin and eosin-stained sections were consistent with a chromophobe adenoma. Ultrastructural examination revealed the tumor to be comprised predominantly of sparsely granulated cells. The tumor cells were negative for prolactin, somatotropin, adrenocorticotropin, luteinizing hormone, and thyrotropin by the peroxidase anti-peroxidase method. Other major lesions were gynecomastia and galactorrhea, testicular atrophy, ankylosing spondylitis, and amyloid deposition in the liver, spleen, adrenal, and intestinal tract. There have been few reports of pituitary tumors in nonhuman primates. These reports include a tumor of the anterior pituitary in a Macaca mulatta [5], a tumor of the infundibular portion in a Cebus capucinus [ 11, and a chromophobe adenoma in a Macaca arctoides [8]. This report describes the microscopic, ultrastructural, and immunohistochemical features of a nonsecreting, sparsely granulated chromophobe adenoma associated with galactorrhea in a Macaca mulatta. Case History A male Mucuca mulutta (90-68) was received at the New England Regional Primate Research Center from another institution in January, In September, 1974, he was dehydrated and emaciated and had abdominal pain. A colon resection was done and approximately 15 cm of colon were removed. Chronic ulcerative colitis was diagnosed; recovery was complete. In December, 1979, a needle biopsy of liver was done as part of a study, and it was normal. In October, 1981, the monkey was sacrificed in poor condition. A complete blood count and serum chemistries were done about two weeks before death. The serum calcium was low (8.5 mg/dl), albumin was low (2.1 gm/dl), the globulin high (4.5 gm/dl), and the albumin/globulin ratio was reversed (0.5). The alkaline phosphatase and lactic dehydrogenase were elevated, 268 mu/ml and 342 mu/ml respectively All other values were normal. Hematology results were normal except for a mild anemia (hemoglobin: 10.0 gm/dl; hematocrit: 36.1 %). 54 1

2 542 Chalifoux, MacKey, and King Fig. 1: Low power magnification of sagittal section of entire tumor. Pars nervosa is thin band (arrow). Darker staining cells underlying this band are morphologically normal cells of the adenohypophysis (arrowheads). Ends of smallkr piece were attached to mass forming a circle (point A to point A and B to B). Space (S) was occupied by adenoma and colloid. C = colloid; N = neoplastic cells. HE. Bar = 2 mm. Fig. 2: Sinusoidal arrangement of neoplastic cells. HE. Bar = 20 pm. Fig. 3: Diffuse arrangement of neoplastic cells. HE. Bar = 20 pm.

3 Pituitary Adenoma in Mucacu mzdafcltu 543 Fig. 4: Ultrastructure of neoplasm. Most cells are sparsely granulated, but the few containing large granules probably are lactotrophs. Bar = 5 pm. Fig. 5: Sparsely granulated cells predominant within tumor. Granules are of various sizes. Bar = 2 pm. Fig. 6: Immunohistochemistry of neoplasm from monkey. Prolactin-secreting cells are confined to periphery (arrows). Peroxidase-antiperoxidase (PAP). Bar = 50 pm. Fig. 7: Gonadotrophs in neoplasm from monkey are very rare (arrows) and are found only in a few areas near pars nervosa as indicated by peroxidase-antiperoxidase for luteinizing hormone. PAP. Bar = 20 pm.

4 544 Chalifoux, MacKey, and King Results Necropsy revealed ankylosing spondylitis involving all vertebrae. The mammary glands and a left supernumerary mammary gland were about 3 cm in diameter and yellowish and conspicuous as seen from the deep surface. On sectioning, cysts were visible and milk exuded from the cut surface. A roughly spherical mass measuring 1.2 cm anteroposterior, 1.8 cm dorsoventrad, and 1.8 cm in width was found at the site of the pituitary attached to the hypothalamus. When cut, it was cystic and contained mucinous material. Microscopically, the ankylosing spondylitis was characterized by bridging of intervertebral spaces by bone that had proliferated from the periosteum of the ventral surface of adjacent vertebrae. The mammary glands were hyperplastic and cystic. Spermatogenesis in the seminiferous tubules was interrupted at the spermatocyte stage and there were no mature spermatozoa. There was marked amyloidosis of the liver with reduction of the width of hepatic cords. Amyloid deposits also were present in the splenic malpighian corpuscles, lamina propria of small intestine and colon, and adrenal. There was a small focus of cortical hyperplasia in one adrenal. There was a plasma cell infiltrate in the gastric mucosa and lymph nodes. The thyroids had been removed by an investigator after death and therefore were not available for examination. A very low magnification of a sagittal section of the tumor is shown in fig. 1. The pars nervosa was flattened and compressed, and a thin capsule surrounded the mass. The tumor was cystic and contained uniformly pink-staining colloid with most cells pushed out on the periphery and some embedded in the colloid. The empty space to the right had been occupied by a mass of cells similar to those comprising the remainder of the mass. It had its own capsule and became detached from this section during processing. Compressed aggregates of cells consistent with normal acidophils were present at the periphery of the tumor adjacent to the pars nervosa. The remaining cells exhibited three different patterns of growth. Some appeared to be forming sinusoids (fig. 2) and others formed follicles containing pink colloid material. Still others were less organized and were present in diffuse sheets (fig. 3). Many of these cells were pyknotic. There were foci of calcification and cholesterol clefts associated with old hemorrhage. Except for small aggregates of acidophils in the areas where the architecture was normal, all other cells regardless of their morphology, had pale pink-purple agranular cytoplasm and thus were regarded as chromophobes. A performic acid-alcian blue-periodic acid-schiff (PAS)-orange G stain was done in an attempt to distinguish cell types. With this stain, the granules of acidophils should stain with orange G and chromophobes have no visible granules. There was a diffuse orange staining throughout most areas, but no granules were seen. An occasional cell at the periphery stained with alcian blue. Presumably these were gonadotrophs. There was no cell with PAS-positive granules.

5 Pituitary Adenoma in Mucucu muluttu 545 Pituitary tumors have a tendency to expand, compressing and eroding adjacent structures. This tendency has led to great caution in the diagnosis of malignancy unless there are distant metastases. In this tumor there were, however, a few areas where cells appeared to be invading the pars nervosa. Ultrastructurally, the neoplastic cells were roughly polygonal with large round or slightly indented nuclei. Most had sparse granulation but a few were more heavily granulated. The granules within the cytoplasm of the more granular cells measured 600 to 700 nm which is within the size range of lactotrophic granules (fig. 4). Granules in the sparsely granulated cells were of various sizes measuring 200 to 500 nm (fig. 5). There was a suggestion of variable density between these granules. There was moderately abundant rough endoplasmic reticulum and numerous mitochondria. Some chromophobe adenomas which have few or no visible granules may still secrete hormone and these may be identified by immunohistochemistry. The peroxidase-antiperoxidase (PAP) method [ 31 was used to stain this neoplasm for prolactin, somatotropin, luteinizing hormone, thyrotropin, and adrenocorticotropin. In a normal human pituitary used as a control, there were many cells containing precipitate which indicated prolactin secretion. In the tumor from the monkey there were a few cells which were positive for prolactin (fig. 6), and these were at the periphery where acidophils were seen with hematoxylin and eosin (HE). The normal human pituitary had numerous cells which were positive for luteinking hormone. In the tumor from the monkey there were only rare positive cells (fig. 7). These were in the same location as those cells which stained with alcian blue. The tumor cells were also negative for adrenocorticotropin, somatotropin, and thyrotropin. Based upon its morphologic, ultrastructural, and immunohistochemical characteristics this neoplasm was identified as a sparsely granulated, nonsecreting adenoma of the pars intermedia. Discussion Because of the gynecomastia and galactorrhea, a prolactin-secreting adenoma was suspected in this male rhesus monkey. Light microscopy of HE- and performic acid-alcian blue-pas-orange G-stained slides demonstrated that the tumor was a chromophobe adenoma and ultrastructural examination showed that it was sparsely granulated. However, actively secreting tumors may become largely depleted of storage granules and therefore appear sparsely granulated [2, 4, 71. Such secretion might explain the galactorrhea in this monkey. Studies of human pituitary tumors correlating immunohistochemical staining with blood prolactin levels indicate that the peroxidase-antiperoxidase method is sufficiently sensitive to detect hormone in the cytoplasm and Golgi s apparatus of functional, sparsely granulated tumors [4] and therefore it is unlikely that this tumor was functioning. Furthermore, the development of the rough endoplasmic reticulum and Golgi s apparatus was not

6 546 Chalifoux, MacKey, and King as prominent as has been reported in actively secreting human tumors [7]. Alternative explanations for the galactorrhea and other lesions, therefore, must be postulated. Cells within remnants of normal pituitary were demonstrated by the peroxidaseantiperoxidase method to contain prolactin. Galactorrhea has been reported in rare human nonsecreting adenomas and is thought to be caused by damage to the hypothalamus with resulting interference with production and transport of prolactin-inhibiting factor [2]. This probably was true in this monkey. The testicular atrophy probably was due to loss of gonadotrophs due to pressure and encroachment of the tumor on the normal adenohypophyseal tissue. There were certainly very few gonadotrophs present in comparison with the normal human control. Loss of gonadrotrophs occurs in the presence of an enlarging tumor before other cell types are affected [6]. Another possible explanation for the arrested spermatogenesis might be an excess of endogenous estrogen due to the failure of the impaired liver to metabolize it. The first explanation seems more likely as other monkeys with severe hepatic amyloidosis have had no testicular atrophy. However, the two etiologies cannot be differentiated by the morphology of the testis [2]. The ankylosing spondylitis was probably a separate disease process unrelated to the pituitary tumor. Ankylosing spondylitis has been associated with regional ileitis in man [6], and this monkey previously had a colitis resembling Crohn s disease. Amyloidosis also has been associated with Crohn s disease but less commonly. The abnormal albumin and globulin may have been related to the immune problems associated with this syndrome, however, elevated serum globulin levels and reversed albumin/globulin ratios also occur in liver disease. The elevated alkaline phosphatase and lactic dehydrogenase probably were caused by the hepatic destruction due to amyloidosis. The small nodule of adrenocortical hyperplasia was almost certainly an incidental finding. Acknowledgements The authors wish to thank Drs. Ronald DeLellis and Arthur Lee, Tufts New England Medical Center, for assistance with the immunohistochemical techniques; Dennis Walsh, June Armstrong, and Bettye-Jean Roy for technical assistance. This work supported by Division of Research Resources (NIH) grant #RROO168 to the New England Regional Primate Research Center. References 1 ANDERSON, M.P.; CAPEN, C.C.: The endocrine system. In: Pathology of Laboratory Animals, eds. Benirschke, K., Garner, F.M., Jones, T.C., pp Springer-Verlag, New York, ANDERSON, W.A.D.; KISSANE, J.D.: Pathology, pp. 1014, C. V. Mosbey Co., St. Louis, DELELLIS, R.A.: Basic techniques of immunohistochemistry. In: Diagnostic Immunohistochemistry, ed. DeLellis, R.A., pp Masson Publishing, USA, Inc., New York, 1981

7 Pituitary Adenoma in Mrrc,ucu mi//uttu DUELLO, T.M.: HALMI, N.S.: Immunocytochemistry of prolactin-producing human pituitary adenomas. Am J Anat 158: , KENT, S.P.; PICKERING, J.E.: Neoplasms in monkey (Macaca mulatta): Spontaneous and irradiation induced. Cancer 11: , ROBBINS, S.L.; COTRAN, R.S.: Pathologic Basis of Disease, pp. 1347, W.B. Saunders, Philadelphia, PA, ROY, S.: Ultrastructure of chromophobe adenoma of the human pituitary gland. J Pathol 122~ , SEIBOLD, H.R.; WOLD, R.H.: Neoplasms and proliferative lesions in 1065 nonhuman primate necropsies. Lab Anim Sci 23: , 1973 Request reprints from L. V. Chalifoux, Department of Pathology, New England Regional Primate Research Center, One Pine Hill Drive, Southborough, MA 0 I772 (USA).