Ph. U. Heitz

Immunohistochemical characterization of an anti-epithelial monoclonal antibody (mAB lu-5)

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.

Senile dementia of Alzheimer type: Astroglial reaction to extracellular neurofibrillary tangles in the hippocampus

SummaryTwo types of Alzheimer neurofibrillary tangles may be found in the hippocampus in senile dementia of the Alzheimer type. Besides classical flameshaped 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.

Evaluation of methods for hepatitis C virus detection in archival liver biopsies. Comparison of histology, immunohistochemistry, in-situ hybridization, reverse transcriptase polymerase chain reaction (RT-PCR) and in-situ RT-PCR.

To evaluate reliable methods for detection of hepatitis C virus (HCV) infection in routinely processed liver biopsies we analyzed formaldehyde-fixed and paraffin-embedded liver specimens of 10 patients with serological confirmed HCV infection. We compared (1) conventional histology; (2) indirect immunofluorescence using the mAb TORDJI-22 (Clonatec, Paris, France); (3) RT-PCR using total RNA and Southern blotting with chemiluminescent detection; (4) non-radioactive in-situ hybridization (ISH) with digoxigenin-labeled oligo- and cRNA probes; (5) direct in-situ RT-PCR with incorporation of labeled nucleotides into PCR-products, and (6) indirect in-situ RT-PCR using subsequent ISH for the visualization of intracellular PCR-products. Our results indicate that: (1) using the histological criteria described by Lefkowitch et al. [Gastroenerology 1993;104:595] together with clinical data, most chronic HCV infections can be diagnosed by conventional histology, if liver biopsies specimens are adequate; (2) the commercially available mAb TORDJI-22 appears to crossreact with non-HCV epitopes, resulting in false positives; (3) molecular methods performed on routinely fixed and processed liver biopsies frequently yield false negative results due to sampling problems, low viral copy number and RNA degradation in infected cells; (4) analysis of HCV-RNA by RT-PCR of extracted total RNA is more sensitive than indirect in-situ RT-PCR or ISH; and (5) direct in-situ RT-PCR is not reliable despite the use of modifications such as DNase pretreatment and hot-start procedures. It is concluded, that several molecular methods for HCV detection must await further improvements of protocols to be suitable for routine diagnostics on paraffin-embedded liver biopsies.

The endocrine pancreas in chronic pancreatitis

The endocrine pancreatic tissue from patients with severe primary chronic pancreatitis (n=6), secondary chronic pancreatitis due to duct obstruction by carcinoma (n=6) and non-diabetic, non-pancreatitic controls (n=4) was studied qualitatively and quantitatively using specific immunocytochemistry and electron microscopy. Grouping of variously sized islets in the sclerotic tissue (sclerosis islets), islet neoformation by ductuloinsular proliferation, and intrainsular fibrosis were the main qualitative findings. Immunocytochemical quantitation of the distribution of insulin (B), glucagon (A), somatostatin (D) and pancreatic polypeptide (PP) producing cells revealed a significant relative increase in the number of A cells and a decrease in the number of B cells of the sclerosis islets in primary chronic pancreatitis (B-44.1±9.3%:A-38.3±2.4%:D-8.6±5.1%:PP-4.6±4.1%) as well as in secondary chronic pancreatitis (B-38.0±14.3%:A-38.4±19.0%:D-9.1±5.8%:PP-14.5±23.4%) compared with controls (B-71.1±8.1%:A-24.3±5.5%:D-8.0±2.8%:PP-0.5±0.4%). The number of PP cells was significantly increased in primary chronic pancreatitis only. It is suggested that scarring of the exocrine pancreas affects islet composition, probably by impairment of the local circulation and of glucose diffusion, thus leading to reduction of the number and glucose sensitivity of B cells. The hyperplasia of A and PP cells appears to be a secondary phenomenon due to the loss of B cells.

Solid-cystic (papillary-cystic) tumours within and outside the pancreas in men: Report of two patients

Solid-cystic (papillary-cystic) tumours (SCT) of the pancreas are distinctive neoplasms with a predilection for young female patients. This is the first detailed report describing the occurrence of SCT in two young male patients. Except for the extrapancreatic occurrence of one of the tumours (in the retroperitoneal region behind the head of the pancreas), all other clinicopathological features were identical to those characterizing the SCT in women. Immunostaining was (at least focally) positive for Lu 5 (broad spectrum keratin marker), vimentin and alpha-1-antitrypsin. The tumours were negative for neuroendocrine markers (except for neuron-specific enolase), pancreatic hormones and enzymes, pancreatic stone protein, carcinoembryonic antigen, CA 19-9 and nuclear oestrogen and progesterone receptors. This report does not support the suggested female sex hormone dependence of SCT.

Angiomatoid malignant fibrous histiocytoma : Evidence for the histiocytic origin of tumor cells

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.

Identification of the conserved, conformation-dependent cytokeratin epitope recognized by monoclonal antibody (lu-5)

The epitope recognized by the murine monoclonal antibody (mAB lu-5) recently described as a formaldehyde-resistant, “pan-epithelial marker” of great value in tumour diagnosis is located on the surface of cytokeratin filaments. It has been preserved during vertebrate evolution from amphibia to man. As this epitope is not reactive after SDS-polyacrylamide gel electrophoresis (SDS-PAGE), the epitope-bearing protein has been identified by a dot-blot antibody binding assay, using purified proteins in which the epitope is reconstituted. We show that the epitope is present in most cytokeratin polypeptides of both the acidic (type I) and basic (type II) subfamily but does not occur in other cytoskeletal proteins. The location of this widespread epitope is discussed with respect to homologies of amino acid sequences of cytokeratins and their conformations.

IGFII and MIB1 immunohistochemistry is helpful for the differentiation of benign from malignant adrenocortical tumours

Aims:  The differentiation of adrenocortical carcinomas from adenomas may be difficult based on morphology alone. Differential expression of insulin‐like growth factor (IGF) II and cyclin‐dependent kinase (CDK) 4 has recently been described in these tumours. The aim of this study was to investigate the diagnostic usefulness of these markers immunohistochemically.

Preferential HER‐2/neu overexpression and/or amplification in aggressive histological subtypes of invasive breast cancer

Aims:  To investigate whether alterations of the HER2 gene occur more frequently in histologically unfavourable subtypes of invasive breast cancer.

Immunocytochemical demonstration of intermediate filament cytoskeleton proteins in human endocrine tissues and (neuro-) endocrine tumours

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.