An immortalized rat ventral mesencephalic cell line, RTC4, is protective in a rodent model of stroke.

One therapeutic approach to stroke is the transplantation of cells capable of trophic support, reinnervation, and/or regeneration. Previously, we have described the use of novel truncated isoforms of SV40 large T antigen to generate unique cell lines from several primary rodent tissue types. Here we describe the generation of two cell lines, RTC3 and RTC4, derived from primary mesencephalic tissue using a fragment of mutant T antigen, T155c (cDNA) expressed from the RSV promoter. Both lines expressed the glial markers vimentin and S100beta, but not the neuronal markers NeuN, MAP2, or beta-III-tubulin. A screen for secreted trophic factors revealed substantially elevated levels of platelet-derived growth factor (PDGF) in RTC4, but not RTC3 cells. When transplanted into rat cortex, RTC4 cells survived for at least 22 days and expressed PDGF. Because PDGF has been reported to reduce ischemic injury, we examined the protective functions of RTC4 cells in an animal model of stroke. RTC4 or RTC3 cells, or vehicle, were injected into rat cortex 15-20 min prior to a 60-min middle cerebral artery ligation. Forty-eight hours later, animals were sacrificed and the stroke volume was assessed by triphenyl-tetrazolium chloride (TTC) staining. Compared to vehicle or RTC3 cells, transplanted RTC4 cells significantly reduced stroke volume. Overall, we generated a cell line with glial properties that produces PDGF and reduces ischemic injury in a rat model of stroke.

Gene expression profile in early stage of retinoic acid-induced differentiation of human SH-SY5Y neuroblastoma cells.

PURPOSE: The human SH-SY5Y cell line is an established model for retinoic acid (RA)-induced neural differentiation. We employed a broad human 15K microarray (15,000 genes) and focused Neuroarray (1152 genes) to examine changes in gene expression early in the process of differentiation (6 hr), before morphology or growth changes are observed. METHODS: 33 P-labeled CDNA probes prepared from RNA extracts of RA-treated and control cultures were hybridized to array membranes, and levels of expression were quantified and compared. RESULTS: In the 15K array, 19 % of the genes were decreased (0.4 % were named genes and the remainder were expressed sequence tags (ESTs) or unknowns), and 9 % were increased (4.2 % named genes). In the Neuroarray, 3 % were decreased and 8 % were increased. CONCLUSIONS: Summary gene profiles are presented, which include transcription factors, genes associated with cell cycle, cell shape, neurotransmission, intermediary filaments, and others. The prevalence of down-regulated genes in the broad 15K array and up-regulated genes in the neuro-focused array suggests a pattern shift in gene expression associated with differentiation. The predominance of ESTs among the down-regulated genes indicates a great number of as-yet-unidentified genes are repressed in early stage neural differentiation.

Human cortical neuronal cell line: a model for HIV-1 infection in an immature neuronal system.

HCN-1A is a human cerebral cortical neuronal cell line having properties consistent with cells of immature neuronal origin. This article details evidence for productive low-level infection of HCN-1A cells with human immunodeficiency virus type 1 (HIV-1). In vitro exposure to HCN-1A monolayers to a high titer of either LAV/HTLV-IIIB or HTLV-IIIMN resulted in HIV-1 p24 antigen production and a moderate increase in reverse transcriptase activity in cell-free supernatants. The cells in both LAV/HTLV-IIIB- and HTLV-IIIMN-infected cultures were passaged and proliferated as long as 5 weeks while continuing to express low levels of viral antigen. Virus-positive cells were detected by indirect immunofluorescence, using serum from an individual with acquired immune deficiency syndrome (AIDS) as well as with a gp120 monoclonal antibody. Confirmation of HCN-1A infection was provided by polymerase chain reaction analyses of both nuclear and cytoplasmic DNA and by de novo synthesis of viral proteins as shown by metabolic labeling and immunoprecipitation. Virus in cell-free supernatants from infected HCN-1A cultures was passaged to a permissive human T cell line (A3.01). HCN-1A cells had no detectable surface CD4 protein or CD4 message. However, the cells expressed the membrane glycolipids, galactocerebroside and sulfatide, possible receptors for gp120 on cells of neuronal origin. Undifferentiated HCN-1A cells provide an in vitro model for investigating potential interactions of HIV-1 with a homogeneous population of immature cortical neurons.

The continued search for evidence of retroviral infection in schizophrenic patients.

Retroviral infection has been proposed as an etiologic factor in schizophrenia. In an effort to unmask a latent retrovirus, short term cultures of peripheral lymphocytes from 15 schizophrenic subjects and nine normal controls were exposed to ionizing radiation and co-cultured with indicator cells. Reverse transcriptase activity, a marker of retroviral infection, could not be detected in any of the cultures. Our findings are further evidence against a role for retroviral infection in the etiology of schizophrenia.

HLA antigens in schizophrenia.

Human leukocyte antigen (HLA) typing was performed on 55 white schizophrenic patients, who were subdivided into groups on the basis of clinical subtype, response to neuroleptic treatment, enlargement of the lateral ventricles, presence of increased prefrontal or parieto-occipital markings, and presence of reversed frontal or occipital lobe asymmetry. No observed differences in antigen frequencies between the group as a whole and controls or between any subgroup and the remaining group or controls remained significant after correction for the number of antigens tested.

Failure to find interference between anti-HLA antibodies and chlorpromazine.

Chlorpromazine has been reported to interfere with the action of alloantibodies directed against HLA-A1. We attempted to replicate this finding using peripheral blood lymphocytes from 3 healthy donors in a complement-mediated lymphocytotoxicity assay. We were unable to find evidence of interference between chlorpromazine and the anti-HLA sera tested. Possible reasons for the difference between our finding and the previous report, as well as the implications for schizophrenia, are discussed.

Binding of 3H-spiperone to human peripheral lymphocytes: absence of stereospecific high-affinity binding.

The binding of 3H-spiperone to human peripheral lymphocytes (PBL) was characterized. The (+)-butaclamol displaceable binding of 3H-spiperone was not saturable, and both (+) and (-)-butaclamol were equally potent in displacing 3H-spiperone binding. We did not find evidence for the existence of a high-affinity specific binding site of 3H-spiperone on human PBL.

Reverse transcriptase activity in 5-azacytidine-treated lymphocyte cultures of patients with schizophrenia.

We investigated the involvement of human lymphotropic retroviruses in the etiology of schizophrenia. Short-term cultures of peripheral lymphocytes of 11 chronic schizophrenic patients and six controls were established and treated with 2.5 and 5 microM 5-azacytidine. No reverse transcriptase activity could be detected in any of the 5-azacytidine-treated cultures indicating that 5-azacytidine did not activate a putative retrovirus in lymphocytes of patients with schizophrenia. Therefore, our findings do not support a role of lymphotropic retroviruses as an etiology in schizophrenia.