Detecting alterations in cell ultrastructure with optical imaging.

Understanding cell functioning at the nanoscale has been hampered in part by the diffraction limited resolution of optical microscopy. We developed partial wave spectroscopic (PWS) microscopy that is capable of quantifying statistical properties of cell structure at the nanoscale. Our animal and human studies demonstrated that alterations in the nanoscale cell architecture is one of the earliest events in carcinogenesis and precedes any other known morphological changes at larger length scales (i.e. microarchitecture).

Perineurioma intestinal / Intestinal perineurioma: report of four cases

Antecedentes: Los perineuriomas en el tracto gastrointestinalse describieron por primera vez en el año 2005como un subtipo especial de pólipos. Actualmente hay 15casos descritos en la literatura con esta denominación.Material y métodos: Presentamos cuatro casos (tres mujeresy un hombre de edades comprendidas entre los 47 y 80años) a los que les detectaron pólipos sésiles mediante colonospiay cuyo diagnóstico histológico fue de perineurioma.Resultados: El tamaño de los pólipos remitidos oscilabaentre 0,3 y 0,5 cm. Los pólipos mostraban una proliferaciónde células fusiformes, sin atipias, que se disponían en unpatrón fascicular y en el epitelio presentaban cambios hiperplásicos.Las células expresaban EMA, Glut-1, claudina 1 yvimentina y mostraban negatividad para S100, c-kit, cromogranina,desmina y actina alfa músculo lisa. En dos delos casos, se obtuvieron muestras del material en parafinapara estudio de ultraestructura observándose las característicastípicas del perineurioma. Discusión: Los perineuriomasgastrointestinales se presentan como pólipos, generalmentesésiles, que suelen ser detectados durante colonoscopiasde cribado en pacientes asintomáticos. Se localizanpreferentemente en colon y se observan con más frecuenciaen mujeres. Teniendo en cuenta los criterios actuales deperineurioma, cabe suponer que muchos pólipos intestinalesdiagnosticados previamente como pólipos fibroblásticos yneurofibromas eran realmente perineuriomas. Se asociancon cambios hiperplásicos en el epitelio adyacente. Se desconocesi esta asociación es coincidencia o si el perineuriomainduce la proliferación epitelial (AU)

Introduction: Gastrointestinal perineuriomas were firstdescribed in 2005 as a special subtype of polyps. Currently,there are 15 cases described in the medical literature withsuch denomination. Material and methods: We presentfour cases, three females and one male, from 47 to 80 years,who had sessile polyps detected during colonoscopy, diagnosedhistologically as intestinal perineuriomas. Results:The polyps ranged from 0.3 to 0.5 cm. Microscopically, theywere composed of uniform, bland spindle cells with a fasciculargrowth pattern and they exhibited hyperplastic changesin the epithelium. Cells were positive for epithelialmembrane antigen (EMA), Glut-1, claudin 1 and vimentinand negative for S-100 protein, C-kit, chromogranin andsmooth-muscle actin. Electron microscopy was performedin two cases, revealing typical features of perineurioma.Discussion: Gastrointestinal perineurioma is a distinctivetype of polyp frequently detected during screening colonoscopyas sessile lesions in asymptomatic patients. It appearsthey are most frequently seen in the colon and are morecommon in women. Taking into account the current criteriaof perineurioma, it seems likely that many intestinal polypspreviously diagnosed as fibroblastic polyps and neurofibromasare actually perineuriomas. They are frequently associatedwith hyperplastic changes in the adjacent epithelium.It is not known if this association is a coincidence or if theperineurioma induces the epithelial proliferation (AU)

Localised massive tumourous xanthomatosis of the small intestine.

INTRODUCTION: There are various disorders of the intestine described with accumulations of vacuolated macrophages including single or multiple xanthelasmata, Wolman's disease, cholesteryl ester storage disease (CESD), xanthomatogranulomatotic disease and xantheloma disseminatum. CLINICAL CASE: In this paper, we report on an exceptional case of a 68-year-old male patient with a localised, massive accumulation of vacuolated, most likely lipid-loaded macrophages with an infiltrative pattern in the muscularis mucosa and propria of the small intestine leading to a tumourous mass requiring surgical removal due to impaired gut function. No enlargement of the liver or the spleen and no evidence of general abnormal lipid storage were detected elsewhere. No evidence of Wolman's or CESD was present. Also, on the ultra-structural level, the macrophages contained no cholesterol clefts typical for either Wolman's and CESD. Instead, largely empty, partly large vacuoles were seen, which most likely contained fatty acids removed during processing. DISCUSSION: The pathogenetic mechanism of the massive local accumulation of histiocytic cells in this part of the intestine in our case remains un-clear. In summary, this case demonstrates that on rare occasions histiocytic proliferations can mimic tumourous masses with severe functional impairment of the intestine and thus should be in the differential diagnosis of gastrointestinal motility disorders.

Retroviral inclusions in the enteric smooth muscle of a tumor-bearing young chicken.

A 15-cm segment of small intestine from a 7-wk-old broiler chicken presented for slaughter was encased by a firm, white mass. Other tissues were grossly unremarkable. Microscopically, the enteric serosa and peripheral muscularis of this segment of small intestine were replaced by a fibrosarcoma. Numerous linear, intracytoplasmic, eosinophilic inclusion bodies were present in smooth muscle cells of the muscularis of the small intestine, and a few similar inclusions were present in the muscularis of the proventriculus. In the heart, there were rare intracytoplasmic inclusions typical of viral matrix inclusions. Ultrastructurally, inclusion bodies in enteric smooth muscle were viral matrix inclusions, and virions resembling avian retroviruses were present in adjacent intercellular spaces. Polymerase chain reaction (PCR) of the deoxyribonucleic acid (DNA) extracted from tumor tissues indicated the presence of proviral DNA of subgroup J avian leukosis virus. This is the first description of the light microscopic appearance of these viral matrix inclusions in enteric smooth muscle.

Intracellular, intercellular, and stromal invasion of gastric mucosa, preneoplastic lesions, and cancer by Helicobacter pylori.

BACKGROUND AND AIMS: It is not clear how Helicobacter pylori, an apparently extracellular pathogen colonizing the luminal side of the gastric epithelium, invariably causes an immune-inflammatory response on the stromal side of the mucosa. Penetration of H pylori into epithelial cell lines and its interaction with immune-inflammatory cells have been documented in vitro. Several investigations also showed in vivo bacterial penetration into the epithelium up to the lamina propria; however, the identification as H pylori of the bacteria-like bodies observed in unchanged, metaplastic, or neoplastic mucosa remained sometimes questionable. METHODS: To search for bacteria-like organisms, we used transmission electron microscopy on endoscopic biopsy specimens from 20 dyspeptic subjects and surgical specimens of neoplastic and nonneoplastic mucosa from 20 cancerous stomachs. To ascertain the H pylori nature of the organisms found, we used 6 different antibodies directed against bacterial lysates, purified vacuolating cytotoxin A, or purified cytotoxin-associated antigen A in immunogold tests. The results were compared with those of H pylori strains cultivated in vitro. RESULTS: In nonmetaplastic gastric epithelium, cytochemically proven H pylori were detected, in the majority of cases, inside cytoplasm of epithelial cells, in intraepithelial intercellular spaces, and in underlying lamina propria, often in direct contact with immune-inflammatory cells and sometimes inside small blood vessels. Cytochemically proven H pylori were also observed inside 6 of 8 intestinal metaplasias and 9 of 20 cancers. CONCLUSIONS: H pylori penetrates normal, metaplastic, and neoplastic gastric epithelium in vivo, intracellularly, or interstitially to cause a strong immune-inflammatory response and promote gastric carcinogenesis.

Intestinal type adenocarcinoma: a previously unrecognized histologic variant of ductal carcinoma of the pancreas.

Adenocarcinomas with intestinal differentiation have been described in a wide variety of anatomical sites. To our knowledge, however, ductal adenocarcinomas with intestinal phenotype have not been described in the pancreas. We report here 11 ductal carcinomas of the pancreas that were morphologically similar to colonic adenocarcinomas. These pancreatic carcinomas of intestinal type represented 10% of 110 consecutively removed ductal carcinomas of the pancreas. All intestinal type carcinomas expressed cytokeratin 7, carcinoembryonic antigen, CDX2, and MUC2. The pattern of reactivity of cytokeratin 7 and carcinoembryonic antigen was diffuse, whereas that of mucin 2 staining and CDX2 nuclear labeling was focal and confined predominantly to goblet cells and less frequently to columnar cells. Six carcinomas contained collections of endocrine cells admixed with the columnar and goblet cells. Five carcinomas were associated with high-grade pancreatic intraepithelial neoplasia of intestinal type. Six patients were female and 5 were male. Their ages ranged from 52 to 76 years (mean age, 61 years). The clinical presentation did not differ from that of the conventional ductal carcinomas. All carcinomas originated in the head of the pancreas, and 5 had metastasized to the regional lymph nodes at the time of surgical resection. Only 1 patient survived 5 years. Three patients are disease free from 2.8 to 8.9 months after surgery. Six patients died as a direct result of the carcinomas, and 1 was lost to follow-up. More studies are needed to determine the biologic behavior of this distinctive histologic variant of ductal adenocarcinoma of the pancreas.

Intestinal perineuriomas: clinicopathologic definition of a new anatomic subset in a series of 10 cases.

Benign peripheral nerve sheath tumors are uncommon in the gastrointestinal tract, and perineuriomas have not previously been reported to occur at this anatomic location. In this study, we analyzed the clinicopathologic and immunohistochemical features of 10 perineuriomas arising in the intestine. Eight patients were female and 2 male (median age, 51 years; range, 35-72 years). Eight of the lesions were intramucosal perineuriomas presenting as small sessile polyps detected during colonoscopy; 6 of these 8 patients were asymptomatic and undergoing colorectal cancer screening. The remaining 2 cases were submucosal masses, one each located in the colon and jejunum. Of the mucosal polyps, six were located in the rectosigmoid or sigmoid colon and one each was detected in the descending colon and transverse colon. The polyps ranged from 0.2 to 0.6 cm (median, 0.4 cm) in greatest dimension. The colonic and jejunal masses measured 3 cm and 4.5 cm, respectively. Histologically, the intramucosal perineuriomas were composed of uniform bland spindle cells having ovoid to elongated nuclei and pale indistinct cytoplasm, with no cytologic atypia, pleomorphism, or mitotic activity. The lesions had a fine collagenous stroma, demonstrated irregular borders with the adjacent lamina propria, and entrapped colonic crypts. Five cases exhibited hyperplastic changes in the adjacent or entrapped epithelium. The colonic submucosal tumor was microscopically well circumscribed, whereas the jejunal perineurioma showed focal infiltration through the muscularis propria into the subserosa. The stroma was collagenous in the colonic tumor and predominantly myxoid in the jejunal tumor. The spindle cells in the submucosal perineuriomas demonstrated tapered nuclei and elongated bipolar cytoplasmic processes. All tumors except one were positive for epithelial membrane antigen (EMA); 4 of 10 expressed claudin-1 and 2 of 10 expressed CD34. All tumors were negative for S-100 protein, glial fibrillary acidic protein, neurofilament protein, smooth muscle actin, desmin, caldesmon, KIT, and pan-keratin. Electron microscopy was performed on the tumor lacking EMA expression, revealing typical features of perineurioma, namely, spindle cells with long bipolar cytoplasmic processes and prominent pinocytotic vesicles, surrounded by discontinuous basal lamina. Clinical follow-up was available for 4 patients (median, 34 months; range, 8-53 months). No tumor recurred. In summary, perineuriomas may arise in the intestine, most often as intramucosal lesions detected as colorectal polyps with distinctive histologic features including entrapment of colonic crypts. Distinguishing perineuriomas from other spindle cell neoplasms of the gastrointestinal tract can be facilitated by immunostaining for EMA and claudin-1.

[Deposition of keinoid fibers in gastrointestinal stromal tumors of the small intestine. Report of two cases and review of the literature]. / Ablagerung von Skeinoidfasern in gastrointestinalen Stromatumoren des Dünndarms. Zwei Fallberichte und Literaturübersicht.

Skeinoid fibers are interstitial collections of a pathological collagen, most often seen in gastrointestinal stromal tumors. They were first described in 1991. We report two cases of intestinal stromal tumors, one in an exceptionally young patient with excessive skeinoid fiber deposition. The microscopic as well as the ultrastructural findings of skeinoid fibers are demonstrated and their role is discussed considering the newest literature.

Integrin alphavbeta3 promotes anchorage-dependent apoptosis in human intestinal carcinoma cells.

A population of cells surviving during prolonged incubation in suspension (anoikis-negative cells) were selected from the original anoikis-positive human intestinal carcinoma cell line Caco-2. Anoikis-negative cells are characterized by a strong transcriptional downregulation of the alphav-integrin chain as detected by FACS analysis, RT-PCR and Northern blotting. This finding suggested that alphav-integrin generates a signal stimulating apoptosis of Caco-2 cells upon their detachment from the extracellular matrix. Two lines of evidence supporting this suggestion were provided. First, activation of the alphavbeta3 integrin on Caco-2 cells by their treatment with an alphavbeta3-specific monoclonal antibody resulted in marked stimulation of anoikis. Second, treatment of Caco-2 cells with alphav-specific antisense oligonucleotide resulted in downregulation of the expression of alphav chain and in elevated resistance of these cells to anoikis. Thus, for the first time, our data prove that alphavbeta3 integrin can be an active transducer of apoptosis-stimulating signals generated in response to disruption of the cell-matrix contacts.

IgG-mediated histamine release from canine mastocytoma-derived cells.

BACKGROUND: Recent data suggest that normal tissue mast cells can express functional receptors for IgG under certain conditions. However, little is known about IgG receptor expression and functional consequences in mast cell neoplasms. METHODS: In this study, neoplastic mast cells were obtained from a dog with cutaneous mastocytoma (CM-MC) and from a dog with visceral mastocytoma (VI-MC). Both cell populations were characterized morphologically and functionally. RESULTS: Most cells proliferated constantly in suspension without particular supplements. Doubling times of CM-MC and VI-MC were 52.2 and 27.5 h, respectively. Both cell types were sensitive to formalin fixation, did not contain heparin and were tryptase and chymase positive. Electron microscopy showed fine granules with electron-dense content in both cell populations. The total histamine content of CM-MC and VI-MC was 0.25 and 0.10 pg/cell, respectively. Calcium ionophore A23187 and substance P induced dose-dependent histamine release, whereas compound 48/80 had no effect. Most significantly, both cell types, when sensitized with monomeric dog IgG, released histamine upon stimulation by anti-dog IgG. CONCLUSIONS: Dog mastocytoma-derived cells may be useful to study the regulation of neoplastic mast cell growth and differentiation, as well as IgG receptor-mediated activation in neoplastic mast cells. Further research is required to clarify the pathophysiological significance of constitutive expression of IgG receptors in neoplastic (canine) mast cells.

Duodenal periampullary gangliocytic paraganglioma: report of two cases with immunohistochemical and ultrastructural study.

We report two cases of Gangliocytic Paraganglioma (GP) of the ampulla of Vater occurring in a 63-year-old and a 34-year-old individual. The patients were both admitted for a long history of intermittent gastrointestinal bleeding and abdominal discomfort, with no other symptoms. At endoscopy, the GP appeared as a polypoid, ulcerated mass in the ampullar region, measuring 2.5x1.8 and 2 cm, respectively. Microscopically, the tumors showed similar features and were composed of epithelial cells (more than 50%), spindle cells, and ganglion-like cells. The epithelial cells showed clear cytoplasm and formed nests (zellballen or paraganglioma-like groups), and less frequently, cords (carcinoid-like), extending to mucosa and submucosa. Ganglion cells were sparse, constantly associated with the spindle cells. Both epithelial and ganglion cells were synaptophysin, chromogranin A, and anti-neurofilament immunoreactive. The spindle cells were all S-100 positive. Ultrastructural studies revealed dark and light cells, rare elongated cellular processes, secretory granules, and fine fibrils resembling neurofilaments. The histogenesis of GP is still a matter of debate, however its neoplastic nature is supported by the occasionally reported malignant evolution.

Comparative ultrastructural study of cytotoxic granules in nasal natural killer cell lymphoma, intestinal T-cell lymphoma, and anaplastic large cell lymphoma.

Comparative immunohistochemical and ultrastructural studies were performed on five nasal natural killer (NK) cell lymphoma cases, two intestinal T-cell lymphoma cases, and eight anaplastic large cell lymphoma (ALCL) cases to clarify morphological differences in cytotoxic granules among these cytotoxic lymphomas. Nasal NK-cell lymphomas and intestinal T-cell lymphomas had fine azurophilic granules and displayed dot-like immunostaining of granzyme B- and T-cell intracellular antigen 1 (TIA-1), predominantly in the central area of the cytoplasm. Ultrastructurally, these NK-cell lymphomas and intestinal T-cell lymphomas had two types of cytotoxic granules, type-I granules (dense core granules) and type-II granules (multivesicular bodies), which have been demonstrated in normal large granular lymphocytes in peripheral blood. However, ALCLs did not have azurophilic granules, and only type-II cytotoxic granules were found ultrastructurally, even though they showed similar dot-like immunostained patterns of granzyme B and TIA-1, as seen in NK-cell lymphomas and intestinal T-cell lymphomas. Immunoelectron microscopy revealed that TIA-1 was primarily located at the periphery of the cytoplasmic granules in the NK-cell lymphoma and ALCL cases. These findings suggest that malignant lymphomas with a cytotoxic phenotype can be divided into two types, (azurophilic granule)+, (type-I granule)+, (type-II granule)+ lymphomas and (azurophilic granule)-, (type-I granule)-, (type-II granule)+ lymphomas.

Nuclear localization of 111In after intravenous injection of [111In-DTPA-D-Phe1]-octreotide in patients with neuroendocrine tumors.

UNLABELLED: Treatment with tumor-targeting substances is currently being evaluated in clinical trials. For patients with neuroendocrine tumors expressing somatostatin receptors, the 111In-labeled somatostatin analog [diethylenetriaminepentaacetic acid (DTPA)-DPhe1]-octreotide has been used with promising results. To further investigate the clinical effect of the injected conjugate, we analyzed the cellular distribution of 111In by ultrastructural autoradiography. METHODS: Seven patients with somatostatin receptor-expressing midgut carcinoid tumors scheduled for abdominal surgery were investigated by somatostatin receptor scintigraphy. During operation, tumor tissue samples and samples of normal intestine were collected, fixed, and processed for electron microscopy. A thin layer of film emulsion was applied on sections and after the exposure film was developed. The cellular distribution of silver precipitations indicating the presence of isotope was evaluated. RESULTS: Cell surface receptor binding and internalization of [111In-DTPA-D-Phe1]-octreotide in the tumor cells was easily revealed by silver precipitations in the film. Multiple silver grains were seen at the plasma membrane, in the cytoplasmic area among secretory granules and vesicular compartments, and in the perinuclear area. Silver grains were also regularly located in the nucleus. For all patients, the silver precipitation patterns from 111In decay were identical in all examined cells from removed tumors, and in most cells 111In could be seen in the nucleus. The specificity of the silver reaction products is supported by the observation that enterocytes in intestinal tissue specimens from near the tumor did not show any silver grains and no background labeling was seen in the plastic. CONCLUSION: After internalization through the somatostatin receptor system, 111In is translocated to the perinuclear area and into the nucleus. Whether the nuclide is still conjugated to the intact somatostatin analog or to part of it cannot be evaluated in this study. Despite the short irradiation range of 111In, the nuclear localization can explain its clinical effectiveness. The results from this study suggest that [111In-DTPA-D-Phe1]-octreotide may act as a powerful tumor cell-targeting substance.

[Sample preparation for electron microscope with tissues fixed with formalin and embedded in paraffin].

A method of sample preparation for electron microscope is introduced. After paraffin-embedded tissues, fresh and fixed on time, were deparaffined, the tissue were fixed in 4% glutaraldehyde for 24 hours, in 1% osmic acid for 1-2 hours, and dehydrated with acetone and embedded in Epon812; then they were cut into ultrathin sections. By this method, better ultrastructure of paraffin-embedded tissues was obtained, and the method could satisfy the demand for observation under electron microscope. In addition, some important proceedings were also discussed in the paper.

Gastrointestinal autonomic nerve tumor (plexosarcoma): report of a case with fine needle aspiration biopsy and histologic, immunocytochemical and ultrastructural study.

BACKGROUND: Gastrointestinal stromal tumors (GISTs) encompass a large group of mesenchymal neoplasms that display common cytologic spindle-shaped morphology on light microscopy. Immunocytochemical and ultrastructural studies can demonstrate several patterns of differentiation. CASE: A 70-year-old male presented with two intraabdominal small bowel masses. The cytopathologic features of a fine needle aspiration biopsy (FNAB) included plump spindle cells in densely populated aggregates or in a fasciculated pattern, without significant pleomorphism. An epithelioid component in a lobular arrangement with abundant, eosinophilic cytoplasm was also noted. The nuclei were vesicular, with a very evident, eosinophilic nucleolus and finely distributed chromatin. Groups of loosely cohesive cells with slender, dendritic-like cytoplasm were evident. Immunocytochemical study of the embedded, fine needle aspirated fragments of the neoplasm demonstrated immunoreactivity for vimentin and neuron-specific enolase. Cytokeratin immunoreactivity or muscular, vascular, neuroendocrine or nerve sheath differentiation failed to be demonstrated. The cytologic and immunocytochemical findings correlated well with the histologic features of the neoplasm. The morphologic diagnosis was confirmed by ultrastructural study. CONCLUSION: FNAB and immunocytochemistry can be valuable in making the correct diagnosis between gastrointestinal stromal tumors.

Multiple small intestinal stromal tumors with skeinoid fibers in association with neurofibromatosis 1 (von Recklinghausen's disease).

A case of multiple small intestinal stromal tumors (SIST) with skeinoid fibers of the jejunum arising in a 50 year old male with neurofibromatosis 1 (NF-1) is reported. Seven small tumors of the jejunal wall were incidentally found and excised during an operation for abdominal and retroperitoneal neurofibromas. Histologically, the tumors were composed of uniform spindle-shaped cells with fascicular pattern, almost indistinguishable from the histology in leiomyoma. Periodic acid Schiff stain-positive hyaline globules were observed among the tumor cells. Ultrastructurally, these globules were stromal tangles of curvilinear, fluffy fibrils, consistent with skeinoid fibers. The electron-dense granules, possibly neuro-secretory granules, were found in the cytoplasm of the tumor cells. Immunohistochemically, the tumor cells were positive for vimentin, neuron specific enolase and CD34, but negative for muscle markers and S100 protein. The association of NF-1 and multiple SIST with skeinoid fibers may have clinical implications. The multiple occurrence of SIST with skeinoid fibers seems to be often cited as one of the gastrointestinal manifestations of NF-1. The possible site of origin of SIST with skeinoid fibers in NF-1 may be the enteric autonomic nerve plexus in the small intestinal wall.

Endometrial stromal sarcoma with focal smooth muscle differentiation: recurrence after 17 years: a follow-up report with discussion of the nomenclature.

In 1977, a case report was published describing a 28-year-old women with an endometrial stromal tumor that showed foci of myogenic differentiation. The term "stromomyoma" was introduced to encompass both this type of neoplasm as well as "uterine neoplasms resembling ovarian sex-cord tumors" (UTROSCTs). More than 17 years later, the tumor recurred, involving the right ovary, sigmoid colon, small bowel, abdominal wall and omentum. The histologic and electron microscopic similarities between the recurrent tumor and the primary neoplasm were confirmed. Applying the recent classification and diagnostic criteria of endometrial mesenchymal neoplasms, we have concluded that this tumor was a low-grade endometrial stromal sarcoma (LGSS). The formerly proposed term "stromomyoma" implies a benign tumor, in contrast to the obviously malignant nature of this particular tumor. Focal myogenic differentiation of LGSS is not an uncommon finding and does not warrant a separate diagnostic or prognostic entity. UTROSCTs and endometrial stromal sarcomas are two separate diagnostic entities, and combining them under an inclusive terminology is not appropriate.

Differential expression of VIP/PACAP receptor genes in breast, intestinal, and pancreatic cell lines.

Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) are structurally-related neuropeptides that function as trophic factors in addition to their more classical roles as neurotransmitters. Binding and molecular cloning studies have shown that their actions are mediated by receptors encoded by at least three different genes. VIP binding has been demonstrated on many tumor types, and radiolabeled VIP has recently been used as a novel method to localize intestinal tumors in humans and their sites of metastasis. To determine the receptor subtype and level of gene expression, we screened breast, intestinal, and pancreatic, cell lines by Northern blot analysis. Breast lines expressed VIP/PACAP1 receptor mRNA levels comparable to intestinal lines, in agreement with the studies showing particularly high VIP binding in these tumors and their derived cell lines. Pancreatic cell lines expressed mRNA for several receptor types. This extends the potential utility of VIP and PACAP in the localization of tumors, and because VIP and PACAP may regulate the growth rate of some tumors by autocrine or other mechanisms, the identification of receptor subtypes on these lines sets the stage for studies in which the activity of these individual receptors in growth and other processes can be investigated.

Well-differentiated endocrine tumours of the middle ear and of the hindgut have immunocytochemical and ultrastructural features in common.

The immunocytochemical analysis of two cases of well-differentiated endocrine tumours (carcinoids) of the middle ear revealed predominant cell populations producing pancreatic polypeptide (PP)-related peptides, glucagon-related peptides, and serotonin (the latter only in one case). In consecutive sections PP- and glucagon-related immunoreactivities mainly colocalized in the same tumour cells. Ultrastructurally tumour cells were characterized by medium-sized to large granules of moderate to high density, on which PP and glicentin were localized by the immunogold technique. No amphicrine cells were found. These features are consistent with those of similar tumours in the rectal mucosa that are mainly composed of L cells coexpressing both PP-related and glucagon-related peptides. Additional tumour antigens of hindgut type detected immunohistochemically were prostatic acid phosphatase and CAR-5 mucin. Expression of the CAR-5 antigen was also found in samples of normal middle ear mucosa, in which endocrine cells have not been identified. In case 1 peritumoral mucosal invaginations showed a proliferation of endocrine cells identical immunophenotypically to tumour cells, possibly representing a precursor lesion. It is concluded that well-differentiated endocrine tumours of the middle ear are a distinct pathological entity characterized by multiple hormone production, typically involving three classes of hormones (pancreatic polypeptide-related peptides, glucagon-related peptides, and serotonin) of the hindgut endocrine system.

Activation of proglucagon gene transcription by protein kinase-A in a novel mouse enteroendocrine cell line.

The gene encoding proglucagon is expressed predominantly in the pancreas and intestine. The physiological importance of glucagon secreted from the islets of Langerhans has engendered considerable interest in the molecular control of proglucagon gene transcription in the endocrine pancreas. In contrast, little is known about the molecular control of proglucagon gene expression in the intestine. The recent demonstration that glucagon-like peptide-1 (GLP-1) secreted from the intestine is a potent regulator of insulin secretion and glucose homeostasis has stimulated renewed interest in the factors that control GLP-1 synthesis in the intestinal L-cell. To develop a model for the analysis of intestinal proglucagon gene expression, we have targeted expression of a proglucagon gene-simian virus-40 large T-antigen fusion gene to enteroendocrine cells in transgenic mice. These mice develop intestinal tumors that were used to derive a novel cell line, designated GLUTag, that expresses the proglucagon gene and secretes immunoreactive GLP-1 in vitro. GLUTag cells demonstrate morphological characteristics of enteroendocrine cells by electron microscopy and are plurihormonal, as shown by immunocytochemistry and RNA analyses. GLUTag cells express the proglucagon and cholecystokinin genes, consistent with the pattern of lineage-specific enteroendocrine differentiation described for mouse intestine. Proglucagon gene expression was induced by activators of the protein kinase-A pathway, and a combination of messenger RNA half-life and nuclear run-on experiments demonstrated that the protein kinase-A-induction is mediated by an increase in proglucagon gene transcription. In contrast, activators of protein kinase-C stimulated secretion, but not biosynthesis of the PGDPs in GLUTag cell cultures. Analysis of proglucagon processing in GLUTag cells demonstrated the liberation of glucagon, oxyntomodulin, glicentin, and multiple forms of GLP-1. These observations provide evidence for the direct induction of proglucagon gene transcription by a cAMP-dependent pathway and suggest that the GLUTag cell line represents a useful model for the analysis of the molecular determinants of enteroendocrine gene expression.