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A small subset of familial pancreatic endocrine tumors (PET) arises in patients with von Hippel–Lindau syndrome and these tumors may have an adverse outcome compared to other familial PET. Sporadic PET rarely harbors somatic VHL mutations, but the chromosomal location of the VHL gene is frequently deleted in sporadic PET. A subset of sporadic PET shows active hypoxia signals on mRNA and protein level. To identify the frequency of functionally relevant VHL inactivation in sporadic PET and to examine a possible prognostic significance we correlated epigenetic and genetic VHL alterations with hypoxia signals. VHL mutations were absent in all 37 PETs examined. In 2 out of 35 informative PET (6%) methylation of the VHL promoter region was detected and VHL deletion by fluorescence in situ hybridization was found in 14 out of 79 PET (18%). Hypoxia inducible factor 1α (HIF1-α), carbonic anhydrase 9 (CA-9), and glucose transporter 1 (GLUT-1) protein was expressed in 19, 27, and 30% of the 152 PETs examined. Protein expression of the HIF1-α downstream target CA-9 correlated significantly with the expression of CA-9 RNA (PPVHL deletion (PPPVHL alterations and signs of hypoxia signalling were characterized by a significantly shortened disease-free survival. We conclude that VHL gene impairment by promoter methylation and VHL deletion in nearly 25% of PET leads to the activation of the HIF-pathway. Our data suggest that VHL inactivation and consecutive hypoxia signals may be a mechanism for the development of sporadic PET with an adverse outcome.

Background: Patients with a multiple endocrine neoplasia type 1 (MEN1)-associated Zollinger–Ellison syndrome (ZES) show multifocal duodenal gastrinomas and precursor lesions. Aims: To test these lesions for loss of heterozygosity (LOH) of the MEN1 gene locus on chromosome 11q13, and to investigate whether the MEN1-related endocrine cell changes also involved somatostatin cells. Material and methods: Tissue specimens from six patients with MEN1 and ZES were analysed by immunohistochemistry and immunofluorescence. LOH analysis was performed by fluorescence in situ hybridisation (FISH), using probes containing the MEN1 gene locus and the centromere 11 (C11) region. For simultaneous analysis of hormones and allelic deletions, a combined FISH/immunofluorescence protocol was established. Results: 28 of a total of 33 duodenal neuroendocrine tumours (NETs) were gastrin-producing tumours; 13/28 (46.4%) revealed LOH on 11q13 and/or C11. Five of the NETs were somatostatin-expressing tumours, two revealing LOH. Allelic loss was detected in tumours as small as 300 μm (gastrin) and 400 μm (somatostatin) in diameter. The gastrin-producing tumours showed different deletion/retention patterns. Hyperplastic somatostatin cell lesions, similar to those of the gastrin cells, were present in all patients. The hyperplastic lesions of both cell lines consistently retained both 11q13 alleles. Conclusions: Allelic deletion of the MEN1 gene may reflect a pivotal event in the development of multifocal gastrin and somatostatin cell neoplasms in the duodenum of patients with MEN1. The observation of distinct deletion patterns in small synchronous tumours supports the concept that each gastrin-producing tumour in an individual MEN1 patient arises from an independent cell clone.

To clarify the role of the MEN1 gene in the tumorigenesis of sporadic adrenocortical tumors, we performed a molecular study on 35 adrenocortical lesions including 6 hyperplasias, 19 adenomas and 10 carcinomas. Loss of heterozygosity (LOH) of the MEN1 gene was assessed by PCR using an intragenic (D11S4946) and 2 flanking microsatellite markers (D11S4936, PYGM) and/or fluorescence in situ hybridization (FISH) with a 40-kb cosmid probe containing the MEN1 gene. The complete coding sequence of the MEN1 gene was screened for mutations using non-radioactive, PCR-based single-strand conformation polymorphism (SSCP) analysis and MDE heteroduplex gel electrophoresis. PCR-LOH and FISH analyses performed in 29 tumors (PCR-LOH in 4, FISH in 17 and both in 8 tumors) revealed allelic deletion of the MEN1 locus in 8 (27.5%) and at 11q13 in 9 (31%) tumors. Furthermore, the frequency of LOH at 11q13 was significantly higher in adrenocortical carcinomas (60%) than in benign lesions (11%). Mutation analysis of tumor samples revealed 9 polymorphisms in 7 tumors (S145S, R171Q, R171Q together with L432L) but no mutations, with the exception of one adrenocortical adenoma. The latter tumor contained a somatic E109X stop codon mutation in exon 2 and a 5178–9GA splice mutation in intron 4, which was also detectable in various non-tumorous tissues and blood indicative of a germ-line mutation. The patient, who had no clinical signs or family history of MEN1, later also developed a neuroendocrine carcinoma (atypical carcinoid) of the lung. Our findings indicate that inactivating mutations of the MEN1 tumor-suppressor gene appear not to play a prominent role in the development of sporadic hyperplastic or neoplastic lesions of the adrenal cortex and that the newly reported 5178–9GA splice mutation in intron 4 might cause a variant of the MEN 1 phenotype. Int. J. Cancer 80:373–379, 1999. © 1999 Wiley-Liss, Inc.

Optimal treatment parameters and appropriate methods of examination for Nd:YAG laser coagulation of the human prostate in dependence of power-setting and time using the SideFocus side-firing delivery system have been determined. Transurethral free-beam laser coagulation was performed in 10 patients prior to planned radical prostatectomy or cystoprostatectomy. Thereof, 6 patients underwent transperitoneal laparoscopic lymphadenectomy and laser coagulation of the prostate between 4 and 9 days prior to open surgery. These same patients had undergone MRI examination with an endorectal coil during staging and prior to radical prostatectomy. Depth and volume of coagulated prostatic tissue of a 4-quadrant laser application were measured at power-settings of 40 W/90 s and 60 W/60 s. There was no difference in volume coagulation between the two treatment modalities. In contrast there was a significant difference in volume coagulation between those prostates removed at 4-9 days and those removed at 60-210 min after laser coagulation. In histological and MRI examination, however, there was a strong correlation with view to coagulation volume in histological and MRI examination. Using the SideFocus side-firing laser fiber, both treatment modalities showed comparable volume coagulation. Laser-induced changes are conclusively discernible by 4 days, MRI examination with an endorectal coil thus being perfectly suitable for assessment.

Parvovirus infection of pregnant women leading to a transplacentar infection of the fetus may result in hydrops fetalis, and ultimately in intrauterine death of the fetus. In situ hybridization with a biotinylated as well as with a 35S-labeled probe for human parvovirus B19 was performed on formalin-fixed paraffin-embedded (FFPE) tissue from a fetus suffering from non-immunologic hydrops fetalis. Histology was suggestive of viral infection probably with human parvovirus. Parvovirus DNA could be detected and precisely localized mainly in the nuclei of erythroid precursors cells within fetal blood vessels of all organs examined. There was no detection of B19 nucleic acid in parenchymal cells of the placenta or the fetal organs, nor within maternal blood cells. These findings are in agreement with the well-known properties of animal parvoviruses to replicate exclusively in proliferating cells. Taking into consideration the problems in diagnosing human parvovirus infection by light microscopy, we conclude that in situ hybridization with an appropriate non-radioactive probe is a valuable, rapid and safe complementary detection method for the diagnosis and study of human parvovirus infections. The 35S-labeled probe is more sensitive than the biotinylated probe, but has the disadvantages of lower resolution of the signal, longer duration of the assay, the hazard of radioactivity and the shorter shelf-life of the probe.

We investigated the utriculus prostaticus from the fetal period up to adulthood in 148 prostates. During the second half of gestation the utriculus had a simple tubular or a cystic form and was lined with metaplastic squamous epithelium which showed immunohisto-chemical positivity for different keratins, carcinoembryonic antigen, and peanut agglutinin binding sites. After birth, alveolar outgrowths of the utriculus developed. After puberty, the utriculus had become a complicated and variable structure. The epithelium no longer differed from that of the prostate glands either morphologically or immunohistochemically. Within the epithelium numerous endocrine cells were found containing neuron-specific enolase, chromogranin, and serotonin. The utriculus and ejaculatory ducts were embedded in a fibrous stroma with, after birth, numerous plexus-like blood vessels. This fibrous zone was peripherally bordered by a layer of smooth muscle. There was no evidence for a function of the utriculus differing from that of the prostate glands. Since the epithelium of both structures is identical immunohistochemically, the epithelium of the sinus urogenitalis most likely participates in the lining of the utriculus during embryogenesis.

A female patient suffering from a bronchial neuroendocrine tumor with unilateral left-sided carcinoid heart disease is reported. Repeated x-ray films of the chest showed a slowly growing lung tumor in the left lower lobe. The patient refused any diagnostic or therapeutic procedure to define the type of the tumor. During the follow-up of 24 years she developed severe mitral and moderate to severe aortic insufficiency, both invasively quantified by thermodilution techniques. During surgery for double valve replacement the patient died from left ventricular heart failure. Necropsy revealed the typical pattern of a bronchial neuroendocrine tumor without metastases. Examination of the heart disclosed the characteristic deposits of fibrous tissue on the cusps of both the mitral and the aortic valves whereas the right heart showed no abnormalities. Review of the literature suggests the unilateral left-sided carcinoid heart disease to be a very rare finding, its pathogenesis remains to be elucidated.

Eighteen granular cell tumors from various sites were examined with antisera directed against protein S-100, neuron specific enolase (NSE), alpha-1-antichymotrypsin, and alpha-1-antitrypsin, glial fibrillary acidic protein (GFAP), lysozyme, factor VIII-related antigen, myoglobin and vimentin, as well as with a monoclonal antibody (lu-5) directed against a panepithelial marker. The immunocytochemical reaction pattern of the tumors was heterogeneous. The brain and pituitary tumors and one thyroid tumor reacted for alpha-1-antichymotrypsin and alpha-1-antitrypsin, but not for S-100 protein and NSE. However, tumors from other sites showed immunoreactions for S-100 protein and NSE and some also for vimentin. Reactions for alpha-1-antichymotrypsin and alpha-1-antitrypsin were not observed. All other reactions were similarly negative. We conclude that the morphologically homogeneous group of granular cell tumors is biologically heterogeneous.

A mouse monoclonal antibody (mAB lu-5) was prepared using a lung cancer cell line as an antigen. The selected clone produces an IgG with a gamma-1 heavy chain and a kappa-light-chain. Immunohistochemical testing of mAB lu-5 on 117 normal tissue biopsies and 474 tumours revealed reactivity with an intracytoplasmic, formaldehyderesistant antigen present in most epithelial and mesothelial cells, but absent in mesenchymal cells. The antibody can therefore be used as a first order, pan-epithelial marker. It proved also useful for fast tumour diagnosis on frozen sections.

The results of an histological, immunocytochemical and electron microscopic study of an angiomatoid malignant fibrous histiocytoma are reported. Our results support an histiocytic, rather than an endothelial origin for the tumor cells.

One case of each of the following human leukodystrophies was examined immunocytochemically with antisera against myelin basic protein (MBP), myelin associated glycoprotein (MAG) and gliofibrillary acidic protein (GFA): Metachromatic leukodystrophy (MLD), connatal adrenoleukodystrophy (ALD), sudanophilic leukodystrophy of the adult (SLD) and connatal Pelizaeus-Merzbacher disease (PMD). A case of canine globoid cell leukodystrophy (GLD) was also included under the assumption that this disease was the same in the dog as in man. It was shown that the storage process in MLD and GLD did not involve MBP or MAG and that the breakdown of myelin with the formation of fat granule cells containing droplets of neutral fat in ALD and SLS proceeds in a similar way as in experimental Wallerian degeneration. In PMD, MBP is present in the vicinity of axons not surrounded by a myelin sheath demonstrable with conventional means. The globoid cells of GLD could be demonstrated to be of non-astrocytic origin.

Classical acute allergic encephalomyelitis (EAE) was provoked in Lewis rats with bovine spinal cord (BWM) in complete Freund's adjuvant (CFA). An efficient immunohistologic technique (peroxidase-antiperoxidase (PAP) was used to trace exsudates of fibrinogen and immunoglobulin as well as their coexistence with cellular infiltrates and clinical signs. Exsudation was restricted to the vessels exhibiting cellular infiltrates. The findings do not lend support to the assumption that exsudation of circulating factors is the initial local event in EAE. It also remains open, whether the exsudation of fibrinogen and gamma globulin are responsible for the clinical symptoms.

The endocrine pancreatic tissue from 13 patients with severe chronic pancreatitis, 5 patients with pancreatic duct carcinoma and 4 non-diseased pancreases was analysed by immunocytochemistry and morphometry. The controls revealed two distinct islet types with different regional distribution. The lower dorsal part of the pancreatic head contained islets with irregular outlines and a high number of PP cells (PP-cells 60.4±4.1%; B-cells 29.4±4.6%; A-cells 7.4±1.5%; D-cells 2.8±0.6%). The other parts of the pancreas contained compact islets with only a few PP cells (PP-cells 1.0±0.4%; B-cells 69.3±3.0%; A-cells 24.1±2.1%; D-cells 5.8±0.5%). In chronic pancreatitis the sclerotic tissue of the body and the tail region contained compact islets with altered cell inter-relationships when compared with controls. While the number of B-cells was diminished (48.5%), A and PP cells appeared to be increased in number (42.7 and 4.1%, respectively). Furthermore, ductulo-insular proliferations were conspicuous (nesidioblastosis) with budding-off of small endocrine cell clusters made up predominantly of A and PP cells. In 3 patients with pancreatic carcinoma increased numbers of PP cells and of A cells were found along the advancing edge of the carcinoma.

A new semiautomatic image analyzing system based on an APPLE II or IIe personal computer is described. The configurations of the hardware and software are extensively explained. The possibilities of applying the system are demonstrated by different examples taken from various projects in biological research and clinical pathology. The system is desingned to fulfil the following requirements: (1) stepwise realization as a unit-by-unit system according to individual needs and interests, (2) acquisition and calculation of morphometric parameters, (3) provision of solutions for the greatest possible number of given problems in clinical and experimental pathology, (4) direct connection of the system to host computers for on-line data transfer and evaluation, and (5) optimum cost-benefit ratio by manifold application possibilities for the individual hardware and software elements. The advantages of the unit-by-unit system with personal computers outweigh in many respects the disadvantages (for example, increased file handling and limited capacity) in comparison with the closed automatized image evaluation system, particularly with regard to costs, flexibility and compatibility. The system is being further developed in collaboration with Kontron Bildanalyse GmbH, Eching/Munich, FRG.

Two types of Alzheimer neurofibrillary tangles may be found in the hippocampus in senile dementia of the Alzheimer type. Besides classical flame-shaped intraneuronal tangles, there are less compact tangles representing extracellular remnants of destroyed neurons with neurofibrillary change. Strong immunoreactivity for glial fibrillary acidic protein (GFA) was found in the second type of tangles, which was due to penetration of fine processes of fibrous astrocytes into bundles of paired helical filaments (PHF). PHF appear to be a strong stimulus for astrocytic reaction when they are not segregated from the neuropil by the neuronal cell membrane.

This is the first description of a connatal case of adrenoleukodystrophy. The clinical picture consisted of severe psychomotor retardation, convulsions and hypsarrhythmia, but no obvious signs of adrenal insufficiency. Pathologically, the adrenals were small. The entire cortex was largely replaced by large round cells. Ultrastructurally, some cells in the adrenal cortex contained inclusions with electron-lucent clefts surrounded by a membrane. The anterior pituitary lobe could be demonstrated to have produced ACTH. The central nervous system showed extensive zones of demyelination in the brainstem, the cerebellum and the right-sided capsula interna. In the demyelinated areas there was sudanophilic breakdown and an intense gliosis. Ongoing demyelination could also be demonstrated by the chemical analysis. In the gray matter there waere micropolygyria of the insular cortex and swollen nerve cells in the nucleus arcuatus. Ultrastructure revealed the type of inclusions in the microglia of the same type as in the adrenals, and a different type of inclusions in unidentifiable cells, possibly neurons. These latter inclusions consisted of loosely stacked lamellar material. The findings are interpreted as further evidence of storage taking place in this disease.

A case of small cell neuroendocrine (oat cell) carcinoma of the breast in a 52-year old male is presented. Oat cell carcinomas have been reported in various extrapulmonary sites, but this is the second case of a primary oat cell carcinoma of the breast and the first one to have been documented in a male. The tumor was investigated histologically, immunocytochemically and ultrastructurally. The relationship to so-called "carcinoid" mammary tumors is discussed.

Histopathological, immunocytochemical, and electron microscopical investigations were carried out in a man with a protracted history of spastic paraparesis, adrenal insufficiency and hypogonadism. Pathological findings were identical with those of the few previously reported cases of adrenomyeloneuropathy (AMN) including cytoplasmic lamellar inclusions consisting of two parallel 2.5 nm leaflets separated by a clear space of variable extent, in the brain, spinal roots, adrenal gland, and interstitial cells of the testis. No inclusions could be found in oligodendrocytes. In brain macrophages they are thought to represent breakdown products of pathological myelin stored in lysosomes, whereas in other localizations they might be an expression of the primary metabolic defect of the cell. Special attention was paid to the pseudosystematic type of fiber tract degeneration in the spinal cord. The dying-back pattern of axonal destruction was interpreted as a possible result of the multisegmental demyelination observed in these tracts. All known hormones could be localized in the pituitary by immunocytochemistry. Corticotrophs and gonadotrophs were numerous. The structural damage of the adrenal cortex and the interstitial cells of the testis is, therefore, considered to result from the inborn error of metabolism on the one hand and from an enhanced stimulation exerted by ACTH and gonadotrophins on the other.

Histochemical methods were used to obtain information on the chemical constituents of brain amyloid in senile dementia of the Alzheimer type. The staining properties of brain amyloid (senile plaque and amyloid angiopathy) were compared with those of extraneural amyloidosis and endocrine amyloid. We found no histochemical differences between amyloid in senile plaques and in amyloid angiopathy. The content of aromatic amino acids was higher in amyloid of plaques and in amyloid angiopathy than in endocrine amyloid. Furthermore, we found persistent birefringence and affinity of brain amyloid for Congo red after exposure to potassium permanganate, suggesting that AA amyloid is not a major constituent of cerebral amyloid.

The presence and distribution of intermediate filament proteins, such as cytokeratins, vimentin, neurofilament proteins and glial fibrillary acidic protein were assessed immunohistochemically in pituitary adenomas, medullary thyroid carcinomas, endocrine pancreatic tumours, gastric, intestinal and bronchial carcinoids, parathyroid adenomas, pheochromocytomas, paragangliomas and related non-neoplastic tissues. In some cases, immunohistochemical results were correlated with cytoskeletal proteins as analysed by SDS-polyacrylamide gel electrophoresis. Cytokeratin antibodies with broad range of immunoreactivity (i.e. to murine liver cytokeratin component D) reacted with epithelial cells in all non-neoplastic endocrine tissues and related neuroendocrine tumours studied, except for adrenal medulla, pheochromocytoma and paraganglioma, independently of hormone production and biological behaviour. In contrast, antibodies to epidermis-derived cytokeratins failed to stain endocrine tissues and tumours. Paranuclear cytokeratin accumulations were seen in bronchial, gastric, and intestinal carcinoids and seem to be a common feature of neuroendocrine tumours. One- and two-dimensional SDS-polyacrylamide gel electrophoresis of non-neoplastic endocrine tissues and related tumours revealed two major keratin polypeptides corresponding to cytokeratins No. 8 and 18 of the cytokeratin catalog of human cells (Moll et al. 1982). According to this cytokeratin polypeptide composition, endocrine tissues and related tumours conform to the "simple type" of epithelia. Vimentin-related immunoreactivity was restricted to stromal cells and to folliculo-stellate cells in normal pituitary gland, Schwann cells in carcinoids and satellite cells in normal adrenal medulla and in pheochromocytomas. Neurofilament protein- (70 kD)-antibodies only stained nerve fibers in normal tissues and at the periphery of carcinoid tumour cell complexes, and to a variable degree, cells in nontumorous adrenal medulla, pheochromocytomas and paragangliomas. Furthermore, neurofilament reactivity was observed along with cytokeratin expression in two bronchial carcinoids.

Es werden 2 Falle von Pneumocystose beim Erwachsenen mitgeteilt. Die Affektion tritt fast nur im Gefolge von schweren Krankheiten mit Resistenzverminderung auf. Sie wird wesentlich begunstigt durch langdauernde Therapie mit Corticosteroiden, eventuell in Kombination mit Cytostatika, Rontgenbestrahlung oder Immunosuppressiva nach Organtransplantation. Diagnostisch wegleitend ist die Diskrepanz zwischen der starken Dyspnoe und den rontgenologisch eindrucklichen Lungeninfiltraten einerseits und dem sehr diskreten Auskultationsbefund anderseits. Die Diagnose wird gesichert durch die Komplementbindungsreaktion und dem Nachweis von Pneumocystis carinii in Sputum, Trachealsekret oder bioptisch gewonnenem Lungengewebe. Die Mortalitat der Erkrankung ist seit der Einfuhrung von Hydroxystilbamidin und Pentamidin von 20–50% auf ungefahr 5% gesunken.

A female suffering from a complete Carney's triad (two gastric epithelioid leiomyomas, recurrent and metastasizing laryngeal paraganglioma, and pulmonary chondroma or hamartoma), and from a myxoma is described. The time elapsed between the detection of the gastric tumors and the metastases of the paraganglioma was 19 years.

Using an immunoreactive technique the two peptides, motilin and Substance P, have been localized at the ultrastructural level in enterochromaffin (EC) cells. Motilin occurs in cells containing a mixed population of biconcave and round secretory granules whereas Substance P is found in cells with exclusively round granules. These observations confirm the existence of at least two functionally and morphologically different types of EC cell in rabbit bile duct, both of which contain 5-hydroxytryptamine. Classification of the endocrine cells of the gut on a purely morphological basis is clearly impossible, however.

Epithelial endocrine cells containing 5-hydroxytryptamine, substance P, somatostatin, enteroglucagon and vasoactive intestinal polypeptide-immunoreactivity were localized by immunocytochemistry in the mucosa of normal appendices, ileum and proximal colon, and in neurogenic appendicopathy. In neurogenic appendicopathy a large number of proliferating nerves were visualized independently of neurotransmitters by immunostaining for neuron-specific enolase. A large number of nerve fibers were shown to contain substance P-immunoreactivity and to be of intrinsic origin. Stromal endocrine cells containing 5-hydroxytryptamine, somatostatin- and possibly substance P-immunoreactivity, were observed in substantial numbers in neurogenic appendicopathy. Substance P may be involved as a neurotransmitter and/or as a paracrine/endocrine peptide in the pathogenesis of spastic contractions and abnormal peristalsis of the appendix, which are characteristic of neurogenic appendicopathy. Stromal endocrine cells may be considered to be the origin of certain carcinoids in the appendix.

Immunoperoxidase positive HPP-producing cells have been examined ultrastructurally using the serial semithin-thin section technique. The features of the HPP cells were found to be identical with those of the previously described D1-cell. The D1-cells are scattered throughout the exocrine parenchyma and occur to a minor extent in the islets.

Clinical, biochemical, light- and electron microscopic, and immunocytochemical findings of a 13 1/2 year old girl with delayed menarche and signs of virilization due to massive oedema of the left ovary with activation of stromal cells (hyperthecosis) are presented. Testosterone and oestradiol production by large cells in the voluminous ovary was demonstrated by immunocytochemistry and radioimmunoassay. Massive ovarian oedema may result from partial or intermittent torsion of the mesovary interfering with venous and lymphatic drainage, but not with arterial blood flow.

Bei 100 Patienten mit primarem Hyperparathyreoidismus (pHPT) waren das Serum Calcium und das Parathormon (PTH) meist erhoht, bei 35 Patienten mit Hypercalciamie und Malignomen dagegen das PTH normal oder nicht mesbar. Bei den 100 Patienten mit pHPT fanden wir our in 11 % die klassischen radiologischen Befunde (subperiostale Usuren und Cysten), in 64 % dagegen eine Nephrolithiasis. Postoperativ konnten wir 42 Patienten nachuntersuchen (Beobachtungsdauer 2–12 Jahre). 82% der Patienten mit praoperativer Nephrolithiasis zeigten postoperativ keine Symptome mehr, bei 75 % war auch radiologisch keine Nephrolithiasis mehr nachweisbar; 63 % der Patienten gaben spontan eine sehr deutliche Besserung ihres subjektiven Befindens an.

(1988). Special Tinctorial Stains or Histochemistry or Electron Microscopy or Immunocytochemistry or Hybridization or…? Ultrastructural Pathology: Vol. 12, No. 2, pp. iii-iii.