Secretogranin III stringently regulates pathological but not physiological angiogenesis in oxygen-induced retinopathy.

Conventional angiogenic factors, such as vascular endothelial growth factor (VEGF), regulate both pathological and physiological angiogenesis indiscriminately, and their inhibitors may elicit adverse side effects. Secretogranin III (Scg3) was recently reported to be a diabetes-restricted VEGF-independent angiogenic factor, but the disease selectivity of Scg3 in retinopathy of prematurity (ROP), a retinal disease in preterm infants with concurrent pathological and physiological angiogenesis, was not defined. Here, using oxygen-induced retinopathy (OIR) mice, a surrogate model of ROP, we quantified an exclusive binding of Scg3 to diseased versus healthy developing neovessels that contrasted sharply with the ubiquitous binding of VEGF. Functional immunohistochemistry visualized Scg3 binding exclusively to disease-related disorganized retinal neovessels and neovascular tufts, whereas VEGF bound to both disorganized and well-organized neovessels. Homozygous deletion of the Scg3 gene showed undetectable effects on physiological retinal neovascularization but markedly reduced the severity of OIR-induced pathological angiogenesis. Furthermore, anti-Scg3 humanized antibody Fab (hFab) inhibited pathological angiogenesis with similar efficacy to anti-VEGF aflibercept. Aflibercept dose-dependently blocked physiological angiogenesis in neonatal retinas, whereas anti-Scg3 hFab was without adverse effects at any dose and supported a therapeutic window at least 10X wider than that of aflibercept. Therefore, Scg3 stringently regulates pathological but not physiological angiogenesis, and anti-Scg3 hFab satisfies essential criteria for development as a safe and effective disease-targeted anti-angiogenic therapy for ROP.

Gastrointestinal neuroendocrine peptides/amines in inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a chronic recurrent condition whose etiology is unknown, and it includes ulcerative colitis, Crohn's disease, and microscopic colitis. These three diseases differ in clinical manifestations, courses, and prognoses. IBD reduces the patients' quality of life and is an economic burden to both the patients and society. Interactions between the gastrointestinal (GI) neuroendocrine peptides/amines (NEPA) and the immune system are believed to play an important role in the pathophysiology of IBD. Moreover, the interaction between GI NEPA and intestinal microbiota appears to play also a pivotal role in the pathophysiology of IBD. This review summarizes the available data on GI NEPA in IBD, and speculates on their possible role in the pathophysiology and the potential use of this information when developing treatments. GI NEPA serotonin, the neuropeptide Y family, and substance P are proinflammatory, while the chromogranin/secretogranin family, vasoactive intestinal peptide, somatostatin, and ghrelin are anti-inflammatory. Several innate and adaptive immune cells express these NEPA and/or have receptors to them. The GI NEPA are affected in patients with IBD and in animal models of human IBD. The GI NEPA are potentially useful for the diagnosis and follow-up of the activity of IBD, and are candidate targets for treatments of this disease.

CD56 antibody: old-fashioned or still trendy in endocrine lung tumors.

BACKGROUND: In 2010, the World Health Organization published a new classification of the endocrine tumors based on the mitotic rate and index. Concerning lung endocrine tumors, the classification of 2004 remains acceptable and widely approved. We noticed in many publications that the most used antibodies in these tumors are chromogranin and synaptophysin. This finding let us wonder about the diagnostic utility of the CD56 antibody which is widely used in our department. MATERIAL AND METHODS: Sixty-nine endocrine lung cancers were diagnosed over a 12-month period in our Department of Pathology. Immunohistochemical technique using the three antibodies: chromogranin, synaptophysin, and CD56 was performed. The sensitivity of the three antibodies was performed using the ratio: true negative cases/true negative cases + false positive cases. The specificity wasn't performed because the antibodies were used only in endocrine tumors. The comparison of the different percentages of expression of the three antibodies was made by the SPSS software 22.0. RESULTS: The sensitivity of the chromogranin, synpatophysin, and CD56 accounted for 69%, 77%, and 98%, respectively. The mean percentage of immunoreactive cells with CD56 was 70% towards 15% and 20% with chromogranin and synaptophysin antibodies, respectively. The comparison of the percentages of expression showed a significant statistical difference between the expression of CD56 versus synaptophysin and CD56 versus chromogranin with P<0.001. CONCLUSION: CD56 antibody seems to be of diagnostic value in endocrine lung tumors with the highest sensitivity. This fact highlights the necessity of using it as a first-line neuroendocrine marker in association to chromogranin which is considered as the most specific endocrine antibody.

Immunohistochemical and biochemical studies with region-specific antibodies to chromogranins A and B and secretogranins II and III in neuroendocrine tumors.

This short review deals with our investigations in neuroendocrine tumors (NETs) with antibodies against defined epitopes of chromogranins (Cgs) A and B and secretogranins (Sgs) II and III. The immunohistochemical expression of different epitopes of the granin family of proteins varies in NE cells in normal human endocrine and non-endocrine organs and in NETs, suggesting post-translational processing. In most NETs one or more epitopes of the granins were lacking, but variations in the expression pattern occurred both in benign and malignant NETs. A few epitopes displayed patterns that may be valuable in differentiating between benign and malignant NET types, e.g., well-differentiated NET types expressed more CgA epitopes than the poorly differentiated ones and C-terminal secretoneurin visualized a cell type related to malignancy in pheochromocytomas. Plasma concentrations of different epitopes of CgA and CgB varied. In patients suffering from carcinoid tumors or endocrine pancreatic tumors the highest concentrations were found with epitopes from the mid-portion of CgA. For CgB the highest plasma concentrations were recorded for the epitope 439-451. Measurements of SgII showed that patients with endocrine pancreatic tumors had higher concentrations than patients with carcinoid tumors or pheochromocytomas. SgIII was not detectable in patients with NETs.

Immunohistochemical staining of human islet cells with region-specific antibodies against secretogranins II and III.

Chromogranins and secretogranins belong to the granin family of proteins, which are expressed in neuroendocrine and nervous tissue. In earlier publications we have described the development of region-specific antibodies against CgA and CgB. In this study we describe antibodies to SgII and SgIII and their usefulness for immunohistochemical staining. Peptides homologous to defined parts of secretogranins II and III were selected and synthesized. Antibodies were raised and immunostainings were performed on normal human pancreas. The SgII 154-165 (N-terminal secretoneurin), SgII 172-186 (C-terminal secretoneurin) and SgIII antibodies immunostained all insulin-immunoreactive cells, most of the glucagon cells and some of the pancreatic polypeptide cells. The SgII 225-242 antibody immunostained only the insulin-containing cells. None of the antibodies immunostained the somatostatin cells. This study is the first observation of the expression of SgIII in human tissues, where we show expression of SgIII in three of the four major islet cell types in human pancreas.

Immunocytochemistry in the diagnosis of small blue cell tumours of childhood.

OBJECTIVE: This study attempts to define a limited, cost effective, reliable primary panel of antibodies for immunohistology, as an adjunct to morphological features, for the diagnosis of Small Blue Cell Tumors (SBCT) which would be convenient for use in low resource settings to improve their diagnostic accuracy. The choice of antibodies is based on the common childhood tumors in Ibadan and limited by financial constraints and availability of antibodies. MATERIALS & METHODS: Twenty-five representative cases of previously diagnosed small blue cell tumours of childhood were selected from the file of the Department of Pathology, University College Hospital, Ibadan. The retrieved blocks were cut and stained with antibodies to desmin, leucocyte common antigen, cytokeratin, chromogranin, neuron-specific-enolase, vimentin and neurofilament using the avidin-biotin technique as previously described. RESULTS: Of the 25 cases studied 24 (96%) gave interpretable immunostaining reaction and the immunophenotype of these were defined. The staining quality equaled that produced on the control well-fixed positive control sections. The final diagnosis of six of the 25 cases changed based on immunostaining. Four cases previously diagnosed as lymphoma were confirmed to be rhabdomyosarcoma (3 cases) and neuroblastoma, one case each of rhabdomyosarcoma and neuroblastoma were both reclassified as lymphoma. CONCLUSION: Based on our findings, the use of a small first-line panel of antibodies to leucocyte common antigen, desmin and neuron-specific-enolase are ideal for immunohistochemical discrimination of SBCT, as an adjunct to morphology, in low-resource settings.

Peptide microarrays for the characterization of antigenic regions of human chromogranin A.

Microarraying peptides is a powerful proteomics technique for studying molecular recognition events. Since peptides have small molecular mass, they are not easily accessible when adsorbed onto solid supports. Moreover, peptides can lack a well-defined three-dimensional structure, and therefore a correct orientation is essential to promote the interaction with their target. In this work, we investigated the suitability as a peptide array substrate of a glass slide coated with a copolymer of N,N-dimethylacrylamide, N,N-acryloyloxysuccinimide, and [3-(methacryloyl-oxy)propyl]trimethoxysilyl. This polymeric surface was used as substrate for peptides in the characterization of linear antigenic sites of human chromogranin A, a useful tissue and serum marker for neuroendocrine tumors and a precursor of many biologically active peptides. The microarray support provided sufficient accessibility of the ligand, with no need for a spacer, as the polymer chains prevent interaction of immobilized peptides with substrate. In addition, the polymeric surface constitutes an aqueous micro-environment in which linear epitopes are freely exposed despite peptide random orientation. The results reported in this article are in accordance with those obtained in conventional ELISA assays using biotinylated and non-biotinylated peptides.

High plasma levels of human chromogranin A and adrenomedullin in patients with pheochromocytoma.

AIMS AND BACKGROUND: The aim of our study was to investigate the plasma chromogranin A (CgA) and adrenomedullin (AM) levels in patients with pheochromocytomas. METHODS AND STUDY DESIGN: We collected blood samples for measurement of plasma CgA and AM in 21 patients with pheochromocytomas, 43 healthy subjects and 26 patients with solid non-functioning adrenocortical adenomas. In 11 patients with pheochromocytomas plasma CgA and AM were measured again four weeks after tumor removal. CgA and AM were measured by means of a novel solid-phase two-site immunoradiometric assay based on monoclonal antibodies (CgA-RIA CT, CIS bio international) and by a specific radioimmunoassay (RIA, Phoenix Pharm. Inc.), respectively. RESULTS: The mean plasma CgA level (+/- SD) in patients with pheochromocytomas (204 +/- 147.9 ng/mL) was significantly higher (P < 0.001) than that in healthy subjects (41.6 +/- 10.7 ng/mL) and in patients with non-functioning adrenocortical adenomas (47.3 +/- 17.6 ng/mL). The mean plasma AM concentration (+/- SD) in patients with pheochromocytomas (27.5 +/- 10.4 pg/mL) was significantly higher (P < 0.001) than that in HS (13.8 +/- 4.5 pg/mL) and in patients with non-functioning adrenocortical adenomas (16.6 +/- 7.3 pg/mL). Plasma CgA levels correlated with plasma AM levels (r = 0.501; P < 0.02) and with plasma metanephrine levels (r = 0.738; P < 0.0001) in patients with pheochromocytomas. In 11 patients with pheochromocytomas plasma CgA and AM concentrations significantly decreased after tumor removal (P < 0.001 for both). Circulating CgA and AM had a sensitivity of 76.2% and 81%, a specificity of 97.7% and 90.7%, and an accuracy of 91% and 88%, respectively. CONCLUSION: This study demonstrates that circulating CgA and AM levels are increased in pheochromocytoma patients compared with healthy subjects and patients with non-functioning adrenocortical adenomas. Moreover, at the time of diagnosis plasma CgA levels correlated with plasma AM levels and with plasma metanephrine levels in all patients with pheochromocytomas. In conclusion, plasma CgA and AM concentrations may represent additional biochemical parameters for clinical monitoring of patients with pheochromocytomas.

Changes of the gastric endocrine cells in the C57BL/6 mouse after implantation of murine lung carcinoma: an immunohistochemical quantitative study.

AIM: The regional distributions and relative frequencies of some gastric endocrine cells of C57BL/6 mice were studied by immunohistochemical method using seven types of specific antisera against chromogranin A (CGA), serotonin, somatostatin, gastrin, cholecystokinin (CCK)-8, glucagon and human pancreatic polypeptide (HPP) after subcutaneous implantation of murine lung carcinoma (3LL) cells. METHODS: The experimental animals were divided into two groups, one is non-implanted sham and the other is 3LL-implanted group. Samples were collected from the two regions of stomach (fundus and pylorus) at 28 d after implantation of 3LL cells (1 x 10(5) cell/mouse). RESULTS: In this study, all the seven types of immunoreactive (IR) cells were identified except for HPP. Most of these IR cells in the gastric portion were generally spherical or spindle in shape (open-type cell) while cells showing round in shape (closed-type cell) were found occasionally. The regional distributions of gastric endocrine cells in the 3LL-implanted group were similar to those of non-implanted sham. However, significant decreases of some types of IR cells were detected in 3LL-implanted group compared to those of non-implanted sham. In addition, the IR cells showing degranulation were numerously detected in 3LL-implanted group. CGA-, serotonin- and somatostatin-IR cells in the fundus and pylorus regions, and gastrin-IR cells in the pylorus regions of 3LL-implanted groups significantly decreased compared to those of non-implanted sham. However, no changes on frequencies of CCK-8- and glucagon-IR cells were demonstrated between 3LL-implanted and non-implanted groups. CONCLUSION: Endocrine cells are the anatomical units responsible for the production of gut hormones, and the change in their density would reflect a change in the capacity of producing these hormones. Implantation of tumor cell mass (3LL) induced severe quantitative changes of gastric endocrine cell density, and the abnormality in density of gastric endocrine cells may contribute to the development of gastrointestinal symptoms such as anorexia and indigestion, frequently encountered in patients with cancer.

Female para-urethral adenocarcinoma: histological and immunohistochemical study.

BACKGROUND: Female urethral cancer with a diverticular form is assumed to originate from the para-urethral duct, which is embryologically homologous to the male prostate gland. The purpose of the present paper was to investigate the female para-urethral adenocarcinomas histologically and immunohistochemically. METHODS: Surgical specimens obtained from six female patients with para-urethral adenocarcinomas were examined histologically, and an immunohistochemical study using antibodies against carcinoembryonic antigen (CEA), prostate specific antigen (PSA), and chromogranin A was performed. RESULTS: On histologic examination, the female para-urethral cancers were divided into five cases of mucin-producing-type adenocarcinoma and one case of clear cell-type adenocarcinoma. All five mucin-producing-type adenocarcinomas were positive with anti-CEA, and two of them showed neuroendicrine differentiation. One of them showed a focally positive area with anti-PSA. The clear cell-type adenocarcinoma had no positive reactions to these antibodies. CONCLUSIONS: On the basis of histologic structure, positive CEA staining, and the presence of focal neuroendocrine differentiation, mucin-producing-type adenocarcinomas may arise from the proximal part of the para-urethral duct.

Constitutive activation of gp130 leads to neuroendocrine differentiation in vitro and in vivo.

BACKGROUND: Neuroendocrine (NE) differentiation in prostate tumors has been correlated with androgen independent disease and increased risk of death. In vitro, IL-6 initiates NE differentiation utilizing the signal transduction initiated by the interaction with IL-6R alpha and gp130. In this study we analysed the NE differentiation process in vitro and in vivo using the LNCaP androgen dependent cell line via ligand independent induction of NE differentiation. METHODS: LNCaP cells were transfected with a constitutively active gp130 subunit, gp130act. Cell proliferation rate was determined and clones were examined for neuroendocrine differentiation by morphological change, upregulation of CgA and serotonin and formation of dense core vesicles with LNCaP parental cells as the control. Xenograft formation was examined and compared in immunocompromised mice. RESULTS: Gp130act expression promoted significant neuroendocrine differentiation in vitro as determined by a NE like morphology change (increased neurite like extension formation), elevated CgA expression and the formation of dense core vesicles (DCV). These measures concurred with those examined in LNCaP cells following 100 ng/ml IL-6 treatment. Further investigation of the LNCaP gp130act cells in vivo, in immunocompromised androgen intact mice, confirmed that the NE like morphology, as determined by histological and high resolution transmission electron microscopy, was maintained. CONCLUSIONS: NE differentiation was initiated by the expression of gp130act in a ligand independent manner, highlighting the importance of gp130 in the neuroendocrine differentiation process. Further investigation of upregulated/downregulated gene expression in these cells may provide valuable insight into the NE differentiation process.

Modulation of prostate carcinoma cell growth and apoptosis by chromogranin A.

PURPOSE: We elucidated the effect and possible pathways of chromogranin A in regulating prostatic cancer cell growth. MATERIALS AND METHODS: Chromogranin A expression in prostatic cancer cell lines were detected with immunofluorescence flow cytometry (FCM) and the inhibition of cell growth due to chromogranin A antibody was measured using microculture tetrazolium. Cell cycle and RNA changes were evaluated with acridine orange FCM. Intracellular chromogranin A variations were detected with FCM, as were apoptotic changes, p53, Fas and tumor necrosis factor-alpha. Apoptosis and caspase-3 expression of tumor cells was assessed with dual immunohistochemical staining. RESULTS: Increased chromogranin A expression was observed in PC-3, DU145 and LNCap cells independent of hormone dependence. Dose and time dependent growth inhibition occurred at 12 hours. Chromogranin A antibody arrested PC3 cells in the S-phase immediately after treatment. The number of G0/G1 and G2/M cells subsequently decreased. PC3 tumor cells had transiently increased RNA at 12 hours with a marked decrease at 48 hours. Decreasing chromogranin A expression started at 12 hours and was most prominent at 48 hours. Apoptotic cells markedly increased at 12 hours with an increase in p53, Fas and tumor necrosis factor-alpha (Fas more than the others). Increased apoptotic cell and caspase-3 expression was observed on immunohistochemical stains. CONCLUSIONS: Chromogranin A is an important neuropeptide regulating the growth of prostate cancer cells. Specific antibodies can suppress its function through apoptotic pathways (Fas and caspase-3), leading to programmed cell death. Chromogranin A antibody mediated apoptosis may be a specific alternative treatment for prostate cancer.

[Primary hepatic vipoma]. / Vipome hépatique primitif.

Vipoma is a rare neuroendocrine tumor most frequently localized in the pancreas. When it is extrapancreatic, it is most often neurogenic. We report a case of primary extrapancreatic vipoma that is non neurogenic localized in the right liver in a patient with severe diarrhea and hypokaliema. Computed tomography, magnetic resonance imaging, intraoperative tomography and surgical exploration did not show any other extrahepatic primary lesion. The diagnosis was performed by immunochemistry, tumorous cells were positives with anti-VIP antibody. Forty two months after right hepatectomy, the patient was asymptomatic.

Immunohistochemical localization of human pineal tissue antigens in normal retina and retinoblastomas.

The normal human retina and retinoblastomas were examined immunohistochemically to assess the localization of pineal antigens in the retina and the oncogenesis and differentiation of retinoblastoma. In the present study, 41 eyes excised from children (aged 4 months to 7 years, all unilateral occurrence) diagnosed to have retinoblastoma and 4 eyes with normal retinas, were used. Retinoblastomas were histopathologically classified into well-differentiated, moderately-differentiated, and poorly-differentiated types. The antibodies used were 9 monoclonal antibodies to human pineal antigens and 6 antibodies to neural tissues. In the normal retina, staining patterns characteristic of retinal cell layers were observed with PP1, PP3, PP5, PP6, PI1, and PI2 antibodies. In retinoblastomas, PP5 antibody, which reacts with horizontal cells and ganglion cells, and PP6 antibody, which reacts with part of the bipolar cells in the inner nuclear layer, showed intense staining in well-differentiated retinoblastomas, but the intensity of staining and the positivity decreased with the degree of dedifferentiation. Antigens recognized by PP3 and PP4 antibodies were positive in all retinoblanstomas. Reactions to GFAP antibody and antibodies that recognize Müller cells were negative. Retinoblanstomas may express markers of not only photoreceptor cells but also other retinal nuclear cells. These results suggest that the retinoblastoma might be developed from visual stem cells, which are common progenitor cells of photoreceptor cells, intermediate neurons, and ganglion cells.

Chromogranin a processing in human pituitary adenomas and carcinomas: analysis with region-specific antibodies.

The expression of various chromogranin A (CgA) peptide fragments was examined with region-specific antisera in benign and malignant pituitary tumors. Analysis of the proconvertases responsible for proteolytic processing of CgA, prohormone convertase 1/3 (PC1/3), and PC2 was also performed. Adenomas were studied using tissue microarrays, and a larger tissue section of a subset of the prolactin (PRL) adenomas was used to compare to the tissue microarray analysis. Carcinomas were analyzed using larger tissue sections. There were differences in CgA proteolytic products detected between the functional (PRL, adrenocorticotropic hormone [ACTH], and growth hormone tumors and the nonfunctional (gonadotroph and null cell) tumors, with the former group expressing lower levels of many peptides. These differences were most notable in the PRL adenomas and carcinomas in which the region-specific antisera against vasostatin I and vasostatin II detected these fragments in the lowest percentage of tumors and/or had the weakest immunoreactivity. The CgA peptide fragment detected by CgA 176-195 (chromacin) antiserum was expressed by the highest percentage of most functional and nonfunctional benign and malignant pituitary tumors. ACTH carcinomas (n = 3) were more strongly immunoreactive compared to the ACTH adenomas. These results show that there is differential expression of CgA peptide fragments and PC1/3 among different types of pituitary tumors and that ACTH pituitary carcinomas have higher levels of immunoreactive CgA peptide fragments compared to ACTH adenomas. This study also shows the utility of tissue microarrays in the analysis of a large group of tumors with regionspecific antisera.

Characterisation of WE-14 in porcine ocular tissue.

WE-14 is derived from the cell-specific posttranslational processing of chromogranin A (CgA) in subpopulations of neuroendocrine cells and neurons. Region- and site-specific chromogranin A, pancreastatin and WE-14 antisera were employed to study the generation of WE-14 in porcine ocular tissues. No chromogranin A or pancreastatin immunostaining was detected in ocular tissue. Immunohistochemistry detected WE-14 immunostaining in a network of nerve fibre bundles and nerve fibres throughout the limbus, cornea, iris and ciliary body with sparse nerve fibres detected throughout the choroid and sclera. Retinal analysis detected intense WE-14 immunostaining in large ovoid cells in the ganglion cell layer with weak immunostaining in a population of small cells in the inner nuclear layer; weak immunostaining was detected within the fibre layers in the inner plexiform layer. Quantitatively, the highest WE-14 tissue concentration was recorded in aqueous retinal and corneal extracts with lower concentrations in the sclera, choroid and anterior uveal tissues. Chromatographic profiling resolved a minor chromogranin A-like immunoreactant and a predominant immunoreactant co-eluting with synthetic human WE-14. This is the first study to demonstrate that WE-14 is generated in neuronal fibres primarily innervating the anterior chamber and in select cell populations in the retina.

Signal amplification by combining two advanced immunohistochemical techniques.

The immunohistochemical techniques known as EnVision trade mark + System (EVS) and Mirror Image Complementary Antibody (MICA) were recently introduced into laboratory practice because of their high sensitivity. In this paper these techniques were compared and their sequences combined to obtain a new method possibly more sensitive than the original ones. The immunohistochemical staining employing the avidin-biotin complex (ABC), largely used as routine, was adopted as a term of comparison. Samples from the small and large intestine of pigs and sheep were fixed in Bouin and embedded in Paraplast. The primary antibodies utilized were directed against the neuronal nitric oxide synthase (nNOS), vasoactive intestinal polypeptide (VIP) and chromogranin A (Cr A). Targets of these antibodies were nerve structures of the intestinal wall, as well as endocrine cells scattered in the mucosa of the bowel, defined neuroendocrine cells or paraneurons. The EVS method appeared as slightly superior to the MICA method regarding sensitivity of detection. The EVS/MICA (combined) method resulted four/eight times more effective than the original techniques regarding sensitivity of detection and staining intensity, both at low and high dilutions of the primary antibodies. Of these latter, immunopositive structures were still clearly identifiable, at a dilution of 1:256,000. Such efficiency could be explained by the high number of revealing molecules of peroxidase contained in the new sequence. The application of the combined method is recommended when a small quantity of tissue antigens needs to be detected immunohistochemically.

Loss of cell cohesion in breast cytology as a characteristic of neuroendocrine carcinoma.

OBJECTIVE: To characterize a specific group of breast cancers displaying a scattered single cell pattern in cytology and correlate it with histologic and immunohistochemical findings. STUDY DESIGN: Of 135 consecutive malignant breast cytologic specimens, 12 cases were selected for their scattered single cell pattern on aspiration cytology. Immunohistochemical staining for neuroendocrine markers and prognostic parameters was performed on paraffin sections of corresponding primary breast carcinomas. RESULTS: In the smears of the 12 cases, highly cellular neoplastic cells with a single cell pattern were observed predominantly. The tumor cells had relatively wide, granular cytoplasm and a low to moderate nuclear/cytoplasmic ratio. Histologically, they were arranged mainly in relatively large, solid nests and occasionally contained a tubular pattern with small amounts of stromal tissue. Five of the 12 cases demonstrated neuroendocrine differentiation with a positive immunoreaction for chromogranin A and synaptophysin. Except for the small mean size of the tumors (P < .01), no significant differences were identified among the prognostic parameters, including a nodal status, estrogen receptor status, growth fraction by Ki-67 or immunoreactivity for c-erbB-2, as compared with the other 123 cases. CONCLUSION: Loss of cell cohesion in breast cytology is a good morphologic marker for identifying neuroendocrine breast carcinoma.