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Lymphocyte subsets in cerebrospinal fluid (CSF) and peripheral blood were studied using monoclonal antibodies, in patients with subacute sclerosing panencephalitis, eight of whom were at stage 2 and seven at stage 4. Eighteen subjects affected with non immunological diseases constituted the controls. Regardless of the stage, patients with subacute sclerosing panencephalitis had lower percentages of OKT3+ (pan-T) cells in both CSF and peripheral blood, with an increase of OKIa+ cells (B cells, macrophages and active T cells) in peripheral blood. A difference was found in the proportion of OKT4+ (helper-inducer) and OKT8+ (suppressor/cytotoxic) cells in relation to the stage, the most striking finding being a significant decrease of OKT8+ with an increase of T4/T8 ratio in peripheral blood at an early stage. [ABSTRACT FROM AUTHOR]

We used arbitrarily primed polymerase chain reaction (AP-PCR) fingerprinting to identify chromosomal imbalances in six primary mediastinal B-cell lymphomas (PMBLs). Seventy-four chromosomal imbalances were detected, consisting of 49 sequence gains and 25 losses. Amplifications on chromosome X were seen in five cases, four of which involved the same chromosomal locus. Nonrandom gains at the same locus were also identified on chromosomes 2 and 7 in four cases and on chromosomes 5, 9, and 12 in three cases. Five PMBLs were also analyzed by comparative genomic hybridization (CGH), which found chromosome arm 9p amplification as the only nonrandom imbalance. Our data demonstrate that chromosomal amplifications outnumber losses in PMBL. These mainly involve chromosomes 9 and X and may reflect more complex phenomena, such as translocations or other chromosomal rearrangements, as AP-PCR found coexistent gains and losses on these chromosomes. Comparison between AP-PCR and CGH suggests that anomalies affecting the same chromosomal regions may occur at much higher frequencies than expected by CGH, suggesting that genomic amplifications are usually confined to DNA segments smaller than the megabase long segments required for detection in CGH. Modest increases in genetic material may be as effective as higher-level amplifications when affecting sites where a proto-oncogene resides., (Copyright 1999 Wiley-Liss, Inc.)

Fibroblast growth factors (FGFs) are believed to play a key role in tissue differentiation and maturation. Thus, the expression of the four members of the high-affinity tyrosine kinase FGF receptor family (FGFRs) and of the low-affinity heparan sulphate proteoglycan binding sites, syndecan-1 and perlecan, was studied in the human skeletal muscle during development. Northern blot analysis demonstrated a developmentally regulated expression of the mRNAs for FGFR-1, FGFR-3, FGFR-4, whereas only traces of FGFR-2 mRNA were found. Each receptor type had a different developmental pattern, suggesting an independent regulation. Signal for FGFR-3 was retained only in the adult muscle. Among the low-affinity FGF binding sites, perlecan was absent, whereas RNA transcript for syndecan-1 peaked at week 13 of gestation, after which a significant decrease was observed. Immunohistochemistry for FGFRs revealed that their localization changed with muscle maturation. At early embryonic stages, FGFR-3 and FGFR-4 had a scattered distribution in the tissue, and FGFR-1 was found on myotube and myofiber plasma membranes. At later stages, FGFR-1 positivity decreased and was found in a few areas of the muscle, FGFR-3 was concentrated in the nuclei of some, but not all, muscle fibers, and FGFR-4 maintained an association with plasma membrane. In adult tissue, weak positivity for FGFR-3 and FGFR-4 was observed in the connective tissue only. When immunocytochemistry was performed on human fetal myoblasts in culture, confocal microscope analysis revealed a nonhomogeneous cell membrane distribution of FGFRs. Taken together, the data strongly suggest that developmentally regulated expression and cell distribution of FGFRs play a role during muscle maturation.

The ALL-1 gene is an important regulator of embryonal and hematopoietic development, and structural variants of the human gene generated by chromosomal translocations and other genomic alterations presumably act as oncogenes in the pathogenesis of acute leukemias and other hematological malignancies. Antisera against two different epitopes of the human ALL-1 protein (anti-ALL1-N and anti-ALL1-C) were produced. Both sera revealed indistinguishable patterns of antigen localization in human peripheral blood mononuclear cells (PBMCs). In resting PBMCs, the antigen was distributed in a speckled pattern across the nuclei, with an increased density at the nuclear envelope and the nuclear indentation. In mitotically stimulated PBMCs, the antigen surrounded the condensing chromosomes but did not colocalize with chromatin or the nuclear scaffold. The antigen is considered a marker for a novel nuclear subcompartment, a perichromosomal area termed the "chromosomal envelope." In Western blot experiments, the anti-ALL1-N serum reacted with a polypeptide corresponding to the expected full-length 430-kDa ALL-1 protein. Recombinant proteins representing the AT-hook and zinc binding subdomains of the ALL-1 protein interacted in vitro with a degenerate mixture of double-stranded oligodeoxynucleotides. Thus, the ALL-1 protein probably is a DNA-binding protein with both a sequence-unspecific (AT-hook) and a sequence-specific (zinc binding subdomains) double-stranded DNA binding mode.

Alpha 2 macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2 Mr/LRP) is a multi-functional cell surface receptor that has been implicated in important processes, such as atherogenesis, cellular migration, immune response and degenerative diseases. Its expression increases in human brain during Alzheimer's disease, tissue injury and neoplastic transformation. In the present paper we studied the regulation of alpha 2 Mr expression by interferon-gamma (IFN gamma) in human astrocytoma cell lines and in fetal astrocytes. Western blots demonstrated an increase of the alpha 2 Mr expression after 24 h of IFN gamma treatment. This effect paralleled the up-regulation of alpha 2 Mr mRNA, as detected by PCR. By prolonging incubation with IFN gamma, we observed a decrement of alpha 2 Mr in IFN gamma treated cells, both by western blot and cytometric analysis. Since in the same cells IFN gamma also up-regulates alpha 2 macroglobulin, this effect may be due to an augmented degradation of the receptor during its recycling.

The effects of basic fibroblast growth factor (bFGF) on differentiation of human fetal microglial cells were investigated. Human ramified microglial cells treated with human recombinant bFGF underwent a morphological change which resulted in a round-shape phenotype. bFGF was also able to induce a dose- and time-dependent increase in cell proliferation and enhanced phagocytic and non-specific esterase activity. These results indicate that ramified microglia, when properly stimulated, are regulated in their physiological and proliferative activities and are transformed into amoeboid forms. Growth factors, such as bFGF, are likely to play a key role in microglial transformation in both normal developing brain and in central nervous system injury.

Experiments were carried out to correlate the cytological localization of DNA polymerase alpha with the presence of its specific mRNA in human lymphocytes studied at different times after phytohaemagglutinin stimulation. Our data indicated that in resting cells it is not possible to detect DNA polymerase alpha protein or mRNA by Northern hybridization. By contrast, in stimulated cells the detection of mRNA specific for DNA polymerase alpha synthesis is possible after 16 h phytohaemagglutin stimulation, whereas immunolocalization is possible after only 4 h stimulation. Observation of cytological preparations from cells stimulated for times long enough to obtain mitoses surprisingly showed an intense immunoreaction in mitotic chromosomes treated with monoclonal antibodies to DNA polymerase alpha.

In this paper, we report that pure cultures of human microglia were obtained from long-term astrocytic cultures of human fetal brain. After five to six months and repeated cell passages, macrophage-like cells started to spontaneously form in vitro, so that in two to three weeks the whole culture was populated by them. These cells were grown up to over 50 passages in culture and analyzed for morphology, specific marker positivity, growth rate and major histocompatibility complex (MHC) antigen expression with or without gamma-interferon (IFN) stimulation. We found that, regardless of embryonic age of original cultures (10-15 weeks of gestation), cultures showed a remarkable homogeneity and purity and over 90 stained for typical microglial markers. Under basal conditions, two cell subpopulations similar to those described in vivo, we observed: the reactive 'ameboid' type and the resting 'ramified' one, the latter increasing with time in vitro and cell passages. Both cell subpopulations were capable of active phagocytosis and of high-rate proliferation. They spontaneously expressed low levels of MHC class II antigens, but were negative for MHC class I. Stimulation with gamma-interferon lymphokine upregulated the MHC class II expression as well as the MHC class I heavy chain form in ameboid, 'reactive' cells but not in the ramified ones. We also found that beta 2 microglobulin, already expressed in basal conditions, was dissociated from HLA A-B-C molecules in lymphokine-stimulated cells at early passages. The physiological significance of these data, as well as the possible correlation with in vivo ontogenetic modifications, are also discussed.