{"paper_id":"0e9c12fb-ee3d-4bfc-95f7-0c38b8927bf4","body_text":"Six intact female macaques aged between 14 and 31 years (mean age, 19.8\nyears) were identified during clinical evaluations at the Armed Forces Radiobiology\nResearch Institute (AFRRI) or at the Wake Forest Primate Center at the Wake Forest\nSchool of Medicine (WFSM). Macaque care and experimentation was approved by each\ninstitution’s Institutional Animal Care and Use Committee and maintained in\naccordance to federal and state laws including the National Academy of\nSciences’ Guide for the Care and Use of Laboratory Animals. 24  In addition, WFSM and AFRRI are\naccredited by the Association for Assessment and Accreditation of Laboratory Animal\nCare International. Two rhesus macaques were part of a chronic gastric\ncarcinogenesis study being conducted at AFRRI, were receiving nitrosating carcinogen\nethyl-nitro-nitrosoguanidine and were experimentally infected with\n Helicobacter pylori . A third rhesus was part of an opioid and\npain receptors study at WFSM. The remaining rhesus macaque and the cynomologus\nmacaques were either on holding protocols or part of a WFSM breeding colony. Of the\n6 cases, 3 had been treated with progesterone analogs (case Nos. 1, 3 and 6,\nmedroxyprogesterone acetate) and 1 with an aromatase inhibitor (case No. 4;\nletrozole) for endometriosis.\nAfter the initial clinical workup, cases were presented to pathologists at\nthe Division of Comparative Pathology at AFRRI or the Department of Pathology at the\nWake Forest University School of Medicine for pathologic evaluation. Diagnostic\nnecropsies were performed on each animal and tissues were collected. Affected\ntissues, to include grossly recognized decidua were stained with hematoxylin\n& eosin (HE) and various combinations of vimentin (Vector Laboratories,\nclone SP20, rabbit monoclonal, 1:40), common acute lymphoblastic leukemia antigen\n(CALLA, CD10, abcam®, clone 56C6, mouse monoclonal, 1:20), progesterone\nreceptor (Novocastra, Vision Biosystems, clone 1A6, mouse monoclonal, 1:100),\nestrogen receptor alpha (Leica Microsystems, clone 6F11, mouse monoclonal, 1:100),\ndesmin (Epitomics, clone Y66, rabbit monoclonal, 1:250), cytokeratin AE1/AE3\n(BioCare Medical, clone LU-5, mouse monoclonal, 1:50), kermix P8, human chorionic\ngonadotropin (HCG, BioCare Medical, rabbit monoclonal, 1:200), human placental\nlactogen (U22 HPL, Novocastra, Vision Biosystems, rabbit monoclonal, 1:200), and\ncalretinin. Secondary antibodies for the appropriate species were used at a 1:20\ndilution (Super Sensitive™ Kit, BioGenex). Vector® Red (Vector\nLaboratories, Burlingame, CA) or 3,3’-Diaminobenzidine (DAB) were used as\nchromogens and slides were counterstained with Mayer’s hematoxylin. Negative\ncontrols included slides treated with only secondary antibody. All\nimmunohistochemical stains were validated and performed by histology cores at each\nlocation. Board certified veterinary pathologists or a board certified medical\npathologist evaluated all cases and slides.\n\nA summary of clinical findings is shown in  Table 1 . The 6 cases included 4 rhesus macaques and 2\ncynomolgus macaques. The age of affected animals was 19.8 ± 6.0 years\n(mean ± standard deviation). Affected macaques commonly presented with\nanorexia with or without weight loss and abdominal pain. Only 3 of the 6\nmacaques had a history of progesterone treatment (medroxyprogesterone acetate,\nDepo-Provera). Serosanguineous or sanguineous abdominal effusion was present in\n3 of 6 macaques. One rhesus (case No. 6) was in heart failure at the time of\ndeath and had serous abdominal effusion. Only 1 macaque (case No. 5) had a\nhistory of heavy menses.\nAt necropsy, the uterus was commonly enlarged and disfigured or replaced\nby dense connective tissue. Fibrous adhesions were commonly observed especially\nbetween the omentum and the uterus, liver, spleen, and urinary bladder ( Fig. 1 ). In 3 cases, one or both ovaries were\nnot identified and were suspected to be incorporated or effaced by the fibrous\nadhesions. Diffuse tan, red or black solid or cystic nodules were identified on\nthe uterus, urinary bladder, intestines and ovaries of 4 of the 6 macaques. The\ngross lesions were similar in treated and untreated macaques ( Table 1 ).\nHistopathological features of the exuberant decidualized endometriosis\ntissue were very similar to lesions previously described in humans.\nEndometriotic cysts and islands of decidualized stroma were separated by\nvariably dense bands of fibrous connective tissue ( Fig. 2 ). Hemosiderin-laden macrophages were often identified near\ncysts ( Fig. 3 ). Cysts were lined by\npseudostratified, ciliated columnar epithelium and filled with erythrocytes,\ncellular debris, and degenerate neutrophils and macrophages ( Fig. 4 ). Decidualized endometriotic stromal\ncells were polygonal with distinct cell borders, abundant pale eosinophilic\ncytoplasm and brightly eosinophilic, intracytoplasmic globules, which\noccasionally marginalized the nucleus. Nuclei were round to oval with finely\nstippled chromatin and 1 to 2 nucleoli ( Fig.\n5 ). No mitoses were noted in any of the cases but anisocytosis and\nanisokaryosis were present.\nEndometrial granular leukocytes, a specialized type of gamma/delta T\ncell, were scattered in the decidualized endometriosis stroma. These were small\ncells with a high nuclear to cytoplasmic ratio ( Fig. 5 , small arrows). They contained eosinophilic granules\neccentrically clustered within scanty cytoplasm. The nuclei were dense and\ndeeply basophilic without recognizable nucleoli.\nDecidualized stroma in these cases consistently stained positive for\nnuclear progesterone and estrogen receptors, C10 and cytoplasmic vimentin ( Table 2 )( Figs. 6–9 ). Desmin, calretinin, cytokeratin, chorionic\ngonadotropin, cytokeratin AE1/AE3, U22 HPL, and kermix P8 were inconsistent or\nnegative.\n\nDecidualization of the endometrium or ovarian tissue is a normal\nphysiological process and occurs in up to 100% of human pregnancies.\nHowever, exuberant decidualized tissue, especially when it occurs in combination\nwith endometriosis, may endanger the mother and/or the fetus by causing pregnancy\ncomplications or being mistaken for neoplasia. 1 , 18 , 20  Stroma of endometriosis lesions and endometrium\nwith hormonal stimulation from high progesterone may transform into decidualized\ncells. The typical microscopic appearance of the transformed endometrial stroma\nwithin the myometrium (internal), peritoneum, thoracic cavity, or skin (external) is\nseen as islands of large polygonal cells with abundant eosinophilic cytoplasm\nfrequently containing intracytoplasmic droplets. These droplets have been described\nas sulfated mucin and glycogen. 12 , 17  Decidualized stroma cells have\noval, centrally located nuclei with distinct magenta nucleoli and finely stippled\nchromatin. While no mitoses are typically observed, there is often pronounced\nanisocytosis and anisokaryosis. The population of transformed cells is often\nembedded in swaths of histopathologically typical endometriosis: endometrial stroma,\nendometrial glands lined by ciliated epithelium, pools of hemorrhage and\nhemosiderosis. Furthermore there is often abundant lymphoplasmacytic inflammation,\nhemorrhage and varying degrees of necrosis. 26\nThe cellular and nuclear atypia as well as the implantation of these cells\ninto the serosa of a myriad of abdominal organs may resemble a malignant neoplastic\nprocess such as deciduoid mesothelioma, squamous cell carcinoma, mucin-producing\nadenocarcinoma or carcinomatosis. 8 , 17 , 27 , 35  Gross\ndifferential diagnoses for a decidualized stroma within the abdomen include\ndecidualized mesothelioma, endometriosis with granulomatous inflammation,\nfibroplasia, granulation tissue and carcinomatosis, especially if tissue invades and\ndestroys the structural integrity of abdominal organs. 27  Atypical extra-placental trophoblastic\nimplantation of the peritoneal cavity is another differential in animals with a\nhistory of pregnancy. 6 , 10 , 31  The rhesus macaques in cases Nos. 1 and 2 ( Table 1 ) were both exposed to a nitrosamine\ncarcinogen. Genital papillary mesotheliomas have been described in male rats given\nnitrosamines. 15  Finally,\ndifferential diagnoses for abdominal neoplasms following aromatase inhibitor\ntreatment, as in case 3, should include atypical mesenchymal proliferation induced\nby the aromatase inhibitor treatment or potentially a neoplastic mesenchymal\ntransformation as is noted in cases of retroviral induced retroperitoneal\nfibromatosis. 13  Retroperitoneal fibromatosis is histologically\ndistinct from deciduosis since it is composed of dense connective tissue without the\nlarge, polygonal decidualized stromal cells. 28\nIn nonhuman primates, decidualization of endometriosis tissue occurs most\noften under the influence of exogenous progestins or during pregnancy. 5 , 25 , 34  Here only 3 of\nthe 6 cases had received exogenous progestins (medroxyprogesterone acetate). None of\nthese animals were pregnant so we anticipate the endometriotic tissue decidualized\nunder the influence of elevated endogenous progesterone.\nRecognizing the histopathological appearance of the decidualized cells and\ncorrelating their presence with endometriosis are important criteria towards\nappropriate diagnosis. Furthermore, immunohistochemical markers aid in confirming\nthe transformed endometrial stroma. We found the decidualized stroma stained\npositive for nuclear progesterone and estrogen receptors, and cytoplasmic vimentin\nas well as CD10 ( Table 2 ). These\nhistopathological characteristics were consistent with reported results in studies\nof affected humans and nonhuman primates. Typical endometrial stroma stains\npositively for common acute lymphoblastic leukemia antigen, or CD10, which is\nexpressed by hematopoietic neoplasms as well as the endometrium. 2 , 4 , 5 , 32  Additionally, the nuclei of the stroma have\npositive staining for progesterone receptor and estrogen receptor. HCG and U22 HPL\nstaining are often assessed to rule out a trophoblastic implantation while\ncytokeratin markers are evaluated to reject carcinomas as possible\ndiagnoses. 6 , 31  Scattered immunopositivity to both HCG and U22 HPL\nobserved in 2 of the 5 cases stained may represent a variation in\nlaboratory-specific sensitivity. Both positive cases were processed within the same\nimmunohistochemistry laboratory. Desmin has been reported to have variable staining\nin decidualized endometriosis in humans, interpreted as metaplastic transformation\nof the stroma into myofibroblasts or leiomyocytes, reinforcing the argument of\nendometrial stroma plasticity. 7  In\nall of our cases, the stromal cells were negative to desmin. A recent report of\npleural endometriosis in a rhesus macaque found that the endometrial stroma was\npositive to calretinin, which is an immunohistochemical marker noted to demonstrate\nstrong nuclear and cytoplasmic staining of the normal endometrial stroma, while\nshowing decreased or negative reactivity in pathological uterine tissues to include\ndeciduosis, a finding which was supported in the immunohistochemical evaluation of\nall 4 of the cases evaluated with calretinin. 3 , 4\nIn summary, endometrial stromal decidualization was confirmed based on the\nhistopathological features and immunohistochemical analyses of each case.\nMacroscopically, the extensive, invasive spread of the endometriosis, multifocal\nislands of decidualized stromal cells, abundant hemorrhage and variable, regional\nnecrosis mimicked malignancy. Histopathological atypia noted within the decidualized\ncells represents a further confounding feature. These observations aid in\nidentifying and diagnosing such lesions in primates to serve as models for\ndecidualized endometriosis lesions in humans. Understanding endometrial\ndecidualization in nonhuman primates with endometriosis will aid to elucidate the\netiology of decidualized endometriotic stroma in women and further the investigation\nof deciduosis diagnosis and treatment.","source_license":"CC0","license_restricted":false}