Gabriel M. Groisman

CD10: a valuable tool for the light microscopic diagnosis of microvillous inclusion disease (familial microvillous atrophy).

Microvillous inclusion disease (MID) is a specific disorder of the intestinal brush border that leads to intractable secretory diarrhea in infants. At present, electron microscopic analysis is required for its definitive diagnosis. However, this technique is not always available or feasible, and the diagnostic microvillous inclusions may not be evident in all specimens. Accordingly, the availability of a panel of histochemical and immunohistochemical stains displaying a specific staining pattern for MID will allow pathologists to reach a definitive diagnosis of this disorder without recourse to electron microscopy. CD10 is a membrane-associated neutral peptidase, shown to have a linear brush-border staining pattern in normal small intestine. We studied the staining pattern of CD10 in small intestinal biopsies from six patients with MID and in 24 control cases (10 normal small intestine, 10 celiac disease, two autoimmune enteropathy, and two allergic enteropathy). All MID cases revealed prominent cytoplasmic CD10 immunoreactivity in surface enterocytes. In contrast, all control cases showed linear brush-border staining. Similar results were obtained with periodic acid–Schiff, polyclonal carcinoembryonic antigen, and alkaline phosphatase, three stains known to show cytoplasmic staining of surface enterocytes in MID. In conclusion, CD10 is a valuable tool for the diagnosis of MID. It may be used as part of a panel that includes other stains with a distinctive staining pattern in MID such as periodic acid–Schiff, polyclonal carcinoembryonic antigen, and alkaline phosphatase. We suggest that the definitive diagnosis of MID can be reached when small bowel biopsies from infants with intractable diarrhea display cytoplasmic staining of surface enterocytes with the above-mentioned stains.

Treatment of intractable gastrointestinal manifestations of chronic granulomatous disease with cyclosporine

Gastrointestinal manifestations of chronic granulomatous disease of childhood include granulomatous inflammatory bowel disease. Severe colitis and perirectal disease developed in a 12-year-old boy with chronic granulomatous disease while he was receiving interferon gamma therapy. The boy had a deficiency of the 22 kd light chain of the cytochrome b heterodimer. After conventional medical therapy proved to be ineffective, a rapid clinical response was obtained to cyclosporine.

Fibroblastic Polyp of the Colon and Colonic Perineurioma: 2 Names for a Single Entity?

Fibroblastic polyps of the colon and intestinal perineuriomas are unusual mucosal lesions with identical clinical and histologic features, and apparent different immunohistochemical and ultrastructural characteristics. However, immunohistochemical distinction was solely based on the results obtained with epithelial membrane antigen (EMA), an antibody whose reactivity on perineuriomas is difficult to demonstrate. Likewise, accurate ultrastructural diagnosis may be flawed by sampling error, preservation artifacts, or paucity of specific diagnostic features. In a recent short communication, it was suggested that both lesions may represent the same entity. To further evaluate this hypothesis, 28 colorectal polyps with clinical and histologic features of colonic fibroblastic polyps/perineuriomas (including 10 cases previously reported as fibroblastic polyps) were stained immunohistochemically for 4 markers of perineurial differentiation, that is, claudin-1, GLUT-1, collagen type IV, and EMA (the latter performed using an extended protocol for antigen retrieval and a kit for signal amplification). In addition, electron microscopy was performed in 4 cases. EMA and claudin-1 stained 26 of 28 (93%) polyps whereas GLUT-1 and collagen type IV were expressed in all of them. EMA reactivity was mostly focal and weak whereas the other markers displayed a diffuse and strong signal. Ultrastructural examination revealed elongated cells with features of perineurial differentiation including long, slender cytoplasmic processes with pinocytotic vesicles and an external lamina. Our findings support the hypothesis that fibroblastic polyps and perineuriomas of the colon represent the same entity. We suggest reclassifying fibroblastic polyps reactive to perineurial markers as perineuriomas. To reach an accurate diagnosis, we recommend employing at least 2 markers of perineurial differentiation, and performing EMA immunostaining with high antibody concentration, prolonged incubation time, and/or extended protocol for antigen retrieval.

Pleomorphic hyalinizing angiectatic tumor of soft parts : Immunohistochemical study including the expression of vascular endothelial growth factor

We report morphologic, flow cytometric, and immunohistochemical findings in two cases of pleomorphic hyalinizing angiectatic tumor of soft parts. Both patients were middle-aged women with subcutaneous lesions located in the lower extremity. The tumors consisted of sheets of spindled and pleomorphic cells with frequent intranuclear pseudoinclusions associated with clusters of ectatic vessels surrounded by prominent perivascular hyaline material. Numerous, nonhyalinized vessels were also present, mostly in the peripheral areas of the lesions. Some of these vessels had their walls permeated by numerous small capillaries. Immunostaining for vascular endothelial growth factor (VEGF), a secreted protein that has been implicated in tumor-associated angiogenesis, demonstrated positive staining in both tumoral and endothelial cells. Tumor cells were also reactive to vimentin and CD34. Focal positivity for CD99 and factor XIIIa was also present. Flow cytometry yielded a diploid DNA histogram with S-phase fraction of 7%. Our findings corroborate those from previously reported cases. They further suggest that angiogenesis and the angiogenic factor VEGF may play a role in the development of this peculiar tumor.

The value of polyclonal carcinoembryonic antigen immunostaining in the diagnosis of microvillous inclusion disease

Microvillous inclusion disease is a specific disorder recognized as a cause of intractable diarrhea of infancy. We studied three cases by light microscopy, electron microscopy, and immunostaining for polyclonal carcinoembryonic antigen (CEA). Histologically, all cases had villous atrophy and abnormal accumulation of periodic acid-Schiff-positive material in surface enterocytes. Ultrastructurally, poorly developed brush-border and intracytoplasmic inclusions lined by intact microvilli were present in surface enterocytes. Crypt cells showed well-preserved surface microvilli. Carcinoembryonic antigen immunostaining showed prominent intracytoplasmic reactivity in surface enterocytes and linear brush-border reactivity in crypt cells. Normal and diseased small bowel biopsy specimens used as controls revealed linear brush-border reactivity without intracytoplasmic staining. Intracytoplasmic positivity for carcinoembryonic antigen in microvillous inclusion disease is explained by its presence in the glycocalyx within the microvillous inclusions. The demonstration of a distinct staining pattern for polyclonal carcinoembryonic antigen in routinely processed small bowel biopsy specimens provides a new useful criterion that complements other established techniques for accurate diagnosis of microvillous inclusion disease.

Juvenile Granulosa Cell Tumor of the Testis: A Comparative Immunohistochemical Study with Normal Infantile Gonads

This study concerns the nature of two different cell populations in a juvenile granulosa cell tumor (GCT) of the infantile testis. Immunohistochemical features of the tumor were compared with those of normal infantile testes (six cases) and ovaries (six cases). The testicular neoplasm showed follicles, cysts and solid nodules composed of an internal layer of polyhedral cells that expressed cytokeratin and vimentin. Most of the follicles and nodules were surrounded by an external layer of spindle cells that reacted to muscle-specific actin, vimentin, and focally to desmin. A neoplastic rather than reactive origin of the spindle cell population is favored by their concentric arrangement in a peritubular-like or theca-like fashion and by their immunohistochemical correlation with normal peritubular-myoid and theca external cells. Sertoli and granulosa cells of normal infantile gonads were positive for cytokeratin and vimentin; peritubular myoid and theca externa cells expressed muscle-specific actin, vimentin, and focally desmin. The occurrence of two well-differentiated components in the tumor favors its origin from the primitive specialized gonadal stromal cell that during neoplastic transformation develops bidirectional differentiation toward epithelial-like and smooth muscle-like lineages. The possibility that this tumor is composed of immature Sertoli and peritubular myoid cells is discussed.

Enterocyte apoptosis and proliferation are increased in microvillous inclusion disease (familial microvillous atrophy).

Microvillous inclusion disease (MID) is characterized by diffuse villous atrophy without inflammatory changes. While increased apoptosis has been related to mucosal flattening in celiac disease, the role of apoptosis in the pathogenesis of MID is unknown. The aim of this study was to assess the rates of apoptosis and cell proliferation in MID and to compare them with those of normal controls and celiac disease. Small intestinal biopsies from 5 infants with MID, 10 children with normal villous architecture, and 10 children with untreated celiac disease were stained with the terminal uridine deoxynucleotidyl nick end labeling (TUNEL) method to assess apoptotic activity, and with Ki-67 immunohistochemistry to assess cellular proliferation. TUNEL and Ki-67 positive enterocytes were counted in a minimum of 20 well oriented half crypts per section. The percentage of apoptotic cells per crypt (apoptotic index) in normal, MID, and celiac biopsies was 0.03 +/- 0.01%, 0.08 +/- 0.08%, and 0.16 +/- 0.3%, respectively. Significant differences were found between normal and MID, and between normal and celiac cases. The percentage of Ki-67 positive cells per crypt (proliferation index) in normal, MID, and celiac cases was 14 +/- 2.5%, 28 +/- 9.2%, and 56 +/- 14%. Significant differences were found between the 3 groups. In conclusion, (1) enterocyte apoptosis and proliferation are increased in MID; (2) apoptosis appears to be an important factor of cell loss and may be, at least in part, responsible for villous atrophy in MID; and (3) crypts in MID are hyperplastic and not hypoplastic. HUM PATHOL 31:1404-1410.

Heparanase expression by Barrett's epithelium and during esophageal carcinoma progression

Enzymatic activity responsible for the cleavage of heparan sulfate, commonly known as heparanase, is abundant in tumor-derived cells. Heparanase cleaves heparan sulfate side chains, presumably at sites of low sulfation, thus facilitating structural alterations of the extracellular matrix and basement membrane underlying epithelial and endothelial cells. Traditionally, heparanase activity was correlated with the metastatic potential of tumor-derived cells, attributed to enhanced cell dissemination as a consequence of heparan sulfate cleavage and remodeling of the extracellular matrix barrier. More recently, heparanase upregulation was documented in an increasing number of human carcinomas and hematological malignancies, correlating with increased tumor metastasis, vascular density, and shorter post-operative survival of cancer patients. Although heparanase upregulation and its pro-malignant features are well documented, the instance of its induction in the course of tumor development was less investigated. Here, we used immunohistochemical analysis to investigate heparanase expression in normal esophagus, Barretts esophagus without dysplasia, Barretts esophagus with low-grade dysplasia, Barretts esophagus with high-grade dysplasia, and adenocarcinoma of the esophagus. We report that heparanase expression is already induced in Barretts epithelium without dysplasia, and is further increased during progression through distinct pathological stages, namely, low-grade dysplasia, high-grade dysplasia, and adenocarcinoma. Notably, heparanase induction correlated with increased cell proliferation index revealed by Ki-67 staining. These findings suggest that heparanase function is not limited to the process of tumor metastasis, but rather is engaged at the early stages of esophagus carcinoma initiation and progression.

Expression of the intestinal marker Cdx2 in secondary adenocarcinomas of the colorectum.

CONTEXT Secondary adenocarcinomas of the large bowel can closely mimic primary tumors. The differentiation of secondary from primary adenocarcinomas of the colorectum, however, is important because their clinical management and prognosis are different. Immunostaining with the nuclear transcription factor Cdx2, expressed in normal intestinal epithelia and colorectal adenocarcinomas, could be of potential diagnostic use. OBJECTIVE To investigate the diagnostic value of Cdx2 immunoexpression in distinguishing primary from common forms of secondary colorectal adenocarcinomas. DESIGN Cdx2 immunoexpression was analyzed in 20 primary colorectal adenocarcinomas and in 34 secondary colorectal adenocarcinomas and their corresponding primary tumors. All secondary tumors were diagnosed through endoscopic biopsies and included 8 cases of ovarian (4 serous, 2 mucinous, and 2 endometrioid), 6 of mammary (4 lobular and 2 ductal), 4 of gastric (2 intestinal and 2 diffuse), 4 of pulmonary, 4 of pancreatic (ductal), 3 of prostatic, 3 of colorectal, and 2 of endometrial origin. RESULTS Cdx2 was expressed in normal colorectal epithelium, in primary colorectal adenocarcinomas (20/20 cases), in secondary adenocarcinomas of colorectal (3/3) and gastric (3/4) origin, and in metastatic ovarian mucinous adenocarcinomas (2/2). In contrast, no Cdx2 immunoreactivity was observed in secondary colorectal tumors of ovarian (serous and endometrioid), mammary, pancreatic, pulmonary, prostatic, and endometrial origin. CONCLUSION Cdx2 immunostaining may be useful in discriminating primary colorectal carcinomas from frequent types of secondary colorectal adenocarcinomas of nongastrointestinal origin. We suggest including Cdx2 in any antibody panel put together to distinguish between primary and secondary epithelial colorectal malignancies.

The histiocytic marker PG-M1 is helpful in differentiating histiocytes and histiocytic tumors from melanomas.

Previous studies have shown that immunohistochemical stains for histiocytes are immunoreactive for melanomas. Accordingly, their value in differentiating histiocytes and histiocytic lesions from melanomas was questioned. PG-M1, the most specific histiocytic marker, was not evaluated in these studies. Our aims were to assess the reactivity of PG-M1 with a series of primary cutaneous and metastatic melanomas and to establish the potential usefulness of this antibody in the differentiation between histiocytes and histiocytic tumors and melanomas. PG-M1 staining was performed in 50 primary cutaneous and metastatic melanomas. For comparison, additional sections were stained with KP-1 and lysozyme (commonly used as histiocytic markers) and with S-100 and HMB-45 (commonly used as melanoma markers). The intensity (1+, 2+) and extent (1+ to 4+) were recorded semiquantitatively. PG-M1 stained weakly (1+) and focally (2+) only four cases of melanoma (8%). In contrast, histiocytes were strongly reactive for PG-M1 in all cases, being readily differentiated from melanoma cells including the positive cases. KP-1 stained melanoma cells in 44 cases (88%), lysozyme in 11 cases (22%), S-100 in 50 cases (100%), and HMB-45 in 48 cases (96%). No changes were found after restaining of selected KP-1 and lysozyme positive melanomas using an endogenous avidin/biotin blocking kit. PG-M1 is helpful in discriminating histiocytes and histiocytic lesions from melanoma cells. We recommend its inclusion in any antibody panel put together to distinguish between them.