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Involvement of the IEGs in brain injury and ischemia is under intensive investigation (Gubits et al., 1993). There are several families of the IEGs. They include the fos, jun, and zinc finger genes that encode transcription factors. Products of the fos family (c-fos, fra-1, fra-2, and fos B) bind to members of the jun family (c-jun, jun B, jun D) via leucine zippers, and this dimer then binds to the AP-1 site (consensus sequence -TGACTCA-) in the promoter of target genes, which in turn regulate the expression of late response genes that produce long-term changes in cells. For example, c-fos may regulate the long-term expression of preproenkephalin, nerve growth factor, dynorphin, vasoactive intestinal polypeptide, tyrosine hydroxylase and other genes with AP-1 sites in their promoters (Curran and Morgan, 1987; Sheng and Greenberg, 1990). It is likely that the c-fos gene up-regulation observed in ischemic astrocytes leads to the changes observed in the expressions of hsp and cytoskeleton protein genes in this experimental model. This is supported by the findings of Sarid (1991) and Pennypacker et al. (1994) who have shown that AP-1 DNA binding activity in hippocampus recognized an AP-1 sequence from the promoter region of the GFAP which is a potential target gene. van de Klundert et al. (1992) also suggested the involvement of AP-1 in transcriptional regulation of vimentin. IEGs can be induced within minutes by extracellular stimuli including transmitters, peptides, and growth factors. In this study, we have shown that c-fos induction by ischemia was rapid and transient.(ABSTRACT TRUNCATED AT 250 WORDS)

The beta-amyloid peptide (beta AP) has been characterized by protein sequencing techniques as a 39-43 amino acid protein with heterogeneous COOH-termini. Controversy exists regarding the predominant form of beta AP in neuritic plaques (NP) and cerebral vasculature of Alzheimer's disease (AD) brain. A monoclonal antibody was developed that selectively recognizes the free COOH-terminal of beta AP 1-42 but not beta AP species with shorter or longer COOH-termini. Brain sections from AD and related disorders were examined using this antibody. In AD samples, the antibody stained diffuse amyloid and NP cores, many intraneuronal and extraneuronal neurofibrillary tangles (NFT), but not cerebrovascular amyloid. Pick and Lewy bodies lacked immunoreactivity. These findings suggest that beta AP 1-42 is present in early and mature amyloid deposits and NFT, but that species of beta AP other than 1-42 comprise human vascular deposits.

The nucleus basalis of Meynert (nbM) was examined using immunocytochemistry for beta-amyloid precursor protein (beta APP) expression in Alzheimer's disease (AD). In mild AD cases, light labeling of the cell body and proximal processes was observed, and small intracellular structures were labeled rarely. In the more severe cases, intense cytoplasmic beta APP labeling was seen, often along with small beta APP-positive structures. Double-labeling experiments demonstrated that in the more severe cases these small structures were also decorated by a neurofibrillary tangle (NFT) antiserum. Other neurons in the severe cases showed incorporation of beta APP into large inclusions, which were also labeled with the NFT antiserum. However, some large inclusions in the severe cases were labeled by the NFT antiserum but contained no beta APP. Extraneuronal NFTs did not show beta APP labeling and did not react with an antibody to the beta-amyloid peptide. These results suggest that increased expression of beta APP coincides with intracellular NFT formation in the nbM, but that the formation of extraneuronal NFTs results in a loss of beta APP immunoreactivity.

The morphologic and biochemical characteristics of human surgical biopsy specimens taken from 17 patients with anaplastic human gliomas and of athymic mouse-grown tumors derived from them were examined. Fourteen were categorized as glioblastoma multiforme, one as an anaplastic astrocytoma, one as a recurrent glioblastoma multiforme, and one as a gliosarcoma. Fifteen of 17 tumors stained positively immunohistochemically for glial fibrillary acidic protein (GFAP), a glial-specific marker. When portions of the 17 surgical biopsy specimens were injected into the flank subcutaneous space of athymic mice, 16 produced tumors; different portions of a single biopsy specimen were used to establish three separate tumor lines; in toto, 18 tumor lines were established. Mouse-borne tumors contained various proportions of fibrillary and protoplasmic astrocytes, gemistocytes, small anaplastic cells, and multinucleated giant cells. Some were more homogeneous than the human tumors from which they were derived, while others contained a mixed population similar to that of the original biopsy specimen. Of these initial 18 tumors, 16 were stained for GFAP and 14 contained from fewer than 5% to almost 100% GFAP-expressing cells. Ten of the tumor lines were studied in serial passage, several demonstrating increased cellularity with increased passage. GFAP expression was followed through serial passage, and 7 of 10 tumor lines continued to express it, often in reduced amounts, and 2 of 10 ceased expression, one (the gliosarcoma) never having expressed it. These data demonstrate that while athymic mouse-borne human anaplastic gliomas retained some features of the human tumors from which they were derived, they varied from one another morphologically. These mouse-borne tumors also continued to evolve, often changing their levels of GFAP and demonstrating increased cellularity with passage.

Glial fibrillary acidic (GFA) protein was demonstrated by immunofluorescence in both primary and long-term cultures of adult human brain cells explanted from normal, neoplastic and non-neoplastic CNS diseases, and in mouse brain cells subcultivated in vitro. GFA protein was present in the cytoplasm of at least 4 distinct cell types and persisted throughout the finite lifetime of brain cell cultures.

The differentiation potential of the human retinoblastoma cell lines Y-79 and WERI-Rb1 was evaluated in vitro for up to 120 days in a matrix system and in rotary suspension for 30 days. Matrix cultures were grown with 10% fetal calf serum (FCS), with and without differentiation-promoting agents. The latter were applied for a total of 5-45 days (usually 30 days) and included 7S nerve growth factor, dibutyryl cyclic AMP, sodium butyrate, retinoic acid, hydrocortisone, and ascorbic acid. Fully defined, serum-free medium and medium containing 5 or 15% FCS were also used for matrix cultures, and medium with 5 or 10% FCS for suspension cultures. By immunoperoxidase (performed on matrix cultures, both untreated and treated for 30 days with differentiation-promoting agents), the cells of both lines were positive for neuron-specific enolase (NSE), microtubule-associated protein 2 (MAP2), class III beta-tubulin (human h beta 4) isotype, and synaptophysin. In addition, the WERI-Rb1 cells expressed 200 kd neurofilament protein (NFP-H) and retinal S-antigen. Both lines were invariably negative for glial fibrillary acidic (GFA) protein, myelin-associated glycoprotein, myelin basic protein, the epitope recognized by the Leu-7 monoclonal antibody, opsin, and hydroxy-indole-O-methyltransferase. In the Y-79 line the presence of NSE and the absence of NF proteins-H, -M and -L, of GFA protein, and of retinal S-antigen were confirmed biochemically. No differentiated features were found by electron microscopy in either line. Thus, in the matrix system employed, both lines exhibited solely a potential for neuroblastic differentiation, which was more advanced in the WERI-Rb1 line, as reflected by the antigenic expression of NFP-H and of retinal S-antigen.