Andreas Gocht

A transgenic mouse model for the ductal carcinoma in situ (DCIS) of the mammary gland.

The ductal carcinoma in situ (DCIS) of the mammary gland represents an early, pre-invasive stage in the development of invasive breast carcinoma and is increasingly diagnosed since the introduction of high-quality mammography screening. Uncertainties in the prognosis for patients with DCIS have caused a controversial discussion about adequate treatment, and it is suspected that most patients undergoing mastectomy may be overtreated. In order to improve treatment and treatment decision, it therefore is highly desirable to identify prognostic markers and therapeutic targets for DCIS. We here introduce a set of transgenic mice (WAP-T and WAP-T-NP lines) presenting with various morphological forms of DCIS-like lesions. In these mice the SV40 large tumor antigen is specifically induced in epithelial cells of the terminal duct lobular units (TDLU). As a consequence of continuous expression of the oncogene, the animals develop multifocal DCIS and consequently invasive carcinoma within strain specific periods of latency. DCIS lesions in transgenic mice exhibit distinct architectural and cytological features which closely resemble those commonly present in humans. We therefore propose these transgenic lines as an experimental model to study the underlying molecular events leading to DCIS and its progression to invasive disease.

Changed expression of 9-O-acetyl GD3 (CDw60) in benign and atypical proliferative lesions and carcinomas of the human breast

Abstract Expression of gangliosides is affected in various ways by malignant cell transformation. In the present study, we investigated the expression of CDw60, a constituent of O-acetylated disialogangliosides, in benign and atypical proliferative breast diseases, and preinvasive and invasive carcinomas by immunohistochemistry and thin-layer chromatography (TLC). In normal ducts, antibodies to CDw60 (mAb M-T21) reacted to membranes of the Golgi apparatus in the juxtaluminal cell compartment. A similar polarized distribution of Golgi cisterns in epithelial cells was observed in several benign lesions, i.e., fibroadenomas, intraductal papillomas, and gynecomastia. In contrast, blunt duct adenosis and duct hyperplasia exhibited an abnormal cytosolic and cell surface staining, whereas atypical duct hyperplasia showed randomly dispersed immunoreactive Golgi cisterns, indicating loss of epithelial polarity. In mammary carcinomas and in two breast carcinoma cell lines (MCF-7 and EFM-19) the neoplastic cells contained CDw60-immunolabelled Golgi complexes, which were distributed in a disorderly fashion throughout the cytoplasm, thus reflecting a loss of epithelial polarity. Additionally, only well differentiated ductal carcinomas in situ or invasive ductal carcinomas disclosed a strong cell surface labelling, which was absent in lower differentiated carcinomas of the same types. In all carcinomas, the intensity of CDw60 immunostaining decreased with progressing loss of differentiation (grade of dedifferentiation), as demonstrated by staining intensity in paraffin sections and by evaluation of the relative amounts of extracted 9-O-acetyl GD3 by TLC. Our results indicate that abnormal CDw60 expression is already detectable in benign proliferative breast lesions with different risk rates to develop into malignant lesions. Downregulation of CDw60 expression in poorly differentiated invasive carcinomas may be the consequence of loss of cell functions usually associated with poor prognosis.

CDw60: An Antigen Expressed in Many Normal Tissues and in Some Tumours

CDw60 is a recently described T-cell antigen, which functionally delivers a costimulatory signal in T-cell activation. In addition, CDw60 has been regarded as a melanoma-associated antigen. To date, only limited information exists on the distribution of CDw60 in other normal and pathologically altered tissues in human. In the present study, the expression of CDw60 was analysed immunohistologically in a large panel of formalin-fixed and paraffin-embedded normal and pathological human tissues. The antigen was detected in several normal tissues, such as epithelia of the reproductive system, exocrine and endocrine glands, glial cells and neurons of the central and peripheral nervous systems, and lymphoid cells. These showed different subcellular distribution patterns, i.e. (1) cell surface labelling of peripheral lymphocytes and lymphocytes of the lymph node and thymus, (2) diffuse cytosolic staining in lymphocytes, subpial glial processes, and the outer plexiform layer of the retina, (3) granular cytoplasmic staining associated with the Golgi apparatus in epithelial cells of certain endocrine and exocrine glands, of the ductus epididymis and deferens, neurons of the peripheral and central nervous system, and lymphocytes and megakaryocytes of the bone marrow.In exocrine glands, e.g. of the prostate and uterine corpus, CDw60-positive Golgi fields were located in the juxtaluminal cell compartment, thus reflecting a polarized distribution. In some malignant tumours, the neoplastic cells contained CDw60-immunolabelled Golgi complexes, which were disorderly distributed throughout the cytoplasm, thus reflecting a loss of epithelial polarity. Only in mammary carcinomas was abnormal cell surface labelling detected. A putative de novo expression of CDw60 was observed in pleomorphic adenoma and mucoepidermoid carcinoma of the parotid gland, seminoma, embryonal and teratocarcinoma of the testis, small cell carcinoma of the lung, and malignant melanoma. These results define the CDw60 determinant as a broadly distributed antigen within a large panel of normal human tissues. The antigen is also detectable in some previously undescribed benign and malignant tumours, which may give importance to CDw60 as a possible diagnostic marker.

The carbohydrate epitope 3-fucosyl-N-acetyllactosamine is developmentally regulated in the human cerebellum

SummaryThe carbohydrate epitope 3-fucosyl-N-acetyllactosamine (CD15) is involved, as a constituent of glycoconjugates, in cell-cell interactions and cell sorting during rodent CNS morphogenesis. The present study was designed to test whether CD15 is also involved in the development of the human CNS. Human cerebellar hemispheres and vermes from the 24th week of gestation (wg) to the 26th postnatal month (pnm) and from adults were investigated for CD15 immunoreactivity, using the monoclonal antibody MMA. Our findings establish that the carbohydrate moiety is developmentally regulated in neuronal and glial cells during their differentiation. First, the parallel fibers of granule cells are CD15+ during the epoch of synaptogenesis with Purkinje cell dendrites. Second, a subpopulation of neurons from the dentate nucleus is transiently CD15 from the 32nd wg until the 15th pnm. Third, at the onset of myelination (around the 35th wg), CD15 immunoreactivity is discernible in the cytoplasm of young oligodendrocytes. Immunoreactivity on protoplasmic astrocytes of the inner granular layer and on fibrous astrocytes of the white matter progressively increases during fetal development. In addition, the CD15 epitope is persistently present on Bergmann glial processes and ependymal cells. Within the three subdivisions of the cerebellum, i.e., hemispheres, vermis, and flocculonodular lobe, the CD15 expression follows a different timing of morphogenesis. For example, diminution of immunoreactivity in the parallel fibers occurs first in the phylogenetically older flocculonodular lobe and vermis, and later in the phylogenetically younger hemispheres. This study shows that in the human cerebellum the distribution of CD15 undergoes marked developmental changes. This epitope may also act in cell-to-cell recognition, and perhaps could play a role in controlling CNS development.

The Carbohydrate Epitope 3-Fucosyl-N-Acetyllactosamine Is Region-Specifically Expressed in Astrocytes of the Rat Brain

The carbohydrate epitope 3-fucosyl-N-acetyllactosamine (CD15) is involved in cell-to-cell recognition processes in various tissues. In the CNS of the adult rat, immunoreactivity for CD15 reveals a region-specific distribution pattern by light microscopy. In the present study we investigated the ultrastructural localization of CD15 in the rat brain using preembedding immunocytochemical methods. In addition we studied CD15 expression in cultured astrocytes from optic nerves of 11-day-old rats. In optic nerve sections, immunostaining was found on the surface of astrocytes at various contact sites, i.e. astrocyte-astrocyte, astrocyte-oligodendrocyte, astrocyte-axon myelin, and astrocyte-blood vessel contacts. Oligodendrocyte-oligodendrocyte contacts, however, were always negative. In the telencephalic cortex, CD15 immunoreactivity was found in glial cell processes around synapses and in the cerebellar cortex in Bergmann glial cells. In astrocytes grown in serum-containing medium, CD15 was expressed on the surface of fibroblast-like glial fibrillary acidic protein-positive astrocytes, which were identified as type 1 astrocytes as well as on process-bearing A2B5-positive cells, representing type 2 astrocytes. The present data support the assumption that in the adult rodent brain, CD15 is exclusively expressed by astrocytes. The in vivo distribution of this carbohydrate molecule on distinct astroglial contact sites supports the notion that CD15 could act in cell-to-cell recognition processes.

Fate of developing astrocytes in the optic nerve of the myelin-deficient rat.

There is considerable debate on the development of a glial cell line in the rat optic nerve, which is characterized by the specific expression of the A2B5 and HNK‐1 epitopes. This cell line has been assumed to give rise to oligodendrocytes and so‐called type 2 astrocytes. However, it is doubtful that the latter cell type really exists in vivo. In the present study, we have addressed this question by investigating the development of astrocytes in the myelindeficient (md) rat, which is characterized by dysmyelination and loss of oligodendrocytes. Defective oligodendrocytes were observed by the third postnatal day, well before the generation of type 2 astrocytes. Consequently, the number of type 2 astrocytes was reduced in cultures prepared from optic nerves of md rats vs. controls. This finding was not paralleled in vivo; i.e., no dying astrocytes were observed in md sections by conventional electron microscopy. However, immunoreactivity against the HNK‐1 epitope was enhanced in md compared to control sections. Ultrastructurally, HNK‐1 immunoreactivity was detected predominantly on the axonal surface at astroaxonal contact sites, which were found only at the nodes of Ranvier within controls but extended to the whole axonal surface in md animals. Only a minor portion of the immunoreactivity derived from glial cells, presumably from oligodendrocytes at the paranodal region in controls. Thus, the HNK‐1 epitope is not a useful antigen for distinguishing astrocytes in the rat optic nerve. Accordingly, our results do not provide evidence for the existence of specialized type 2 astrocytes in vivo. In vitro, these cells are probably only oligodendrocytes that mimic some astroglial features if grown in serum‐containing media. J. Comp. Neurol. 378:105–116, 1997.