Geminin regulates neuronal differentiation by antagonizing Brg1 activity

Precise control of cell proliferation and differentiation is critical for organogenesis. Geminin (Gem) has been proposed to link cell cycle exit and differentiation as a prodifferentiation factor and plays a role in neural cell fate acquisition. Here, we identified the SWI/SNF chromatin-remodeling protein Brg1 as an interacting partner of Gem. Brg1 has been implicated in cell cycle withdrawal and cellular differentiation. Surprisingly, we discovered that Gem antagonizes Brg1 activity during neurogenesis to maintain the undifferentiated cell state. Down-regulation of Gem expression normally precedes neuronal differentiation, and gain- and loss-of-function experiments in Xenopus embryos and mouse P19 cells demonstrated that Gem was essential to prevent premature neurogenesis. Misexpression of Gem also suppressed ectopic neurogenesis driven by Ngn and NeuroD. Gem's activity to block differentiation depended upon its ability to bind Brg1 and could be mediated by Gem's inhibition of proneural basic helix-loop-helix (bHLH)-Brg1 interactions required for bHLH target gene activation. Our data demonstrate a novel mechanism of Gem activity, through regulation of SWI/SNF chromatin-remodeling proteins, and indicate that Gem is an essential regulator of neurogenesis that can control the timing of neural progenitor differentiation and maintain the undifferentiated cell state.     

Comparative evaluation of nonradiometric BACTEC and improved oxoid signal blood culture systems in a clinical laboratory.

The BACTEC NR660 blood culture system, which uses infrared spectroscopy to detect carbon dioxide generated by bacterial growth, was compared with the new medium formulation of the Oxoid Signal system. Two trials were conducted: a comparative study of 88 organisms in simulated blood cultures and a clinical trial of 3,321 paired patient blood culture samples. Both trials showed that overall the BACTEC system performed better in the recovery of organisms. The Oxoid system was unable to detect by signal the growth of the majority of yeasts, nonfermentative gram-negative bacilli, Neisseria meningitidis, Nocardia spp., and Corynebacterium jeikeium. There were no significant differences in the yield of Staphylococcus spp., members of the family Enterobacteriaceae, Streptococcus spp., or anaerobic organisms. BACTEC detected growth more quickly than did the Oxoid system; 61% of the isolates were detected by BACTEC at 24 h, while 49% of the isolates were detected by Oxoid. The Oxoid system had a high proportion (58.5%) of false-positives, compared with 7.7% for the BACTEC system. Despite the new medium formulation of the Oxoid system, its performance is still not equivalent to that of the BACTEC system.     

Transfer of immunity to cryptococcosis by T-enriched splenic lymphocytes from Cryptococcus neoformans-sensitized mice.

Splenic enriched T-cells and sera were obtained from inbred CBA/J mice injected 7 or 35 days earlier with either 10(3) viable Cryptococcus neoformans or sterile physiological saline. The transfer of enriched T-cells collected 7 days after immunization or of normal enriched T-cells did not transfer immunity to C. neoformans or delayed-type hypersensitivity responsiveness to cryptococcal culture filtrate (CneF) antigen to the recipients. However, enriched T-cells harvested 35 days after immunization, when transferred to recipient mice, were able to confer immunity as indicated by the reduction in numbers of C. neoformans cells in the tissues, and they also transferred delayed-type hypersensitivity responsiveness to CneF antigens. Sera from either sensitized or normal mice were unable to transfer immunity to recipient animals. These results suggested that there was a time requirement for development of the immune response in the donor mice and that T-cells were crucial in the host defense against a cryptococcal infection. Culturing of day-35 C. neoformans-sensitized T-cells in the presence of homologous antigen (CneF) but not in the presence of heterologous antigen (purified protein derivative or 2, 4-dinitro-1-fluorobenzene) induced the production of migration inhibition factor, thus indicating that lymphocytes from C. neoformans-injected mice were specifically sensitized to CneF antigen.     

Functional tyrosine kinase inhibitor profiling: a generally applicable method points to a novel role of platelet-derived growth factor receptor-beta in tuberous sclerosis

A ubiquitous herpesvirus that establishes life-long infection, the Epstein-Barr virus (EBV) has yielded little insight into how a single agent in general accord with its host can produce diverse pathologies ranging from oral hairy leukoplakia to nasopharyngeal carcinoma, from infectious mononucleosis to Hodgkin's disease (HD) and Burkitt's lymphoma. Its pathogenesis is further confounded by the less than total association of virus with histologically similar tumors. In other viral systems, defective (interfering) viral genomes are known to modulate outcome of infection, with either ameliorating or intensifying effects on disease processes initiated by prototype strains. To ascertain whether defective EBV genomes are present in HD, we examined paraffin-embedded tissue from 56 HD cases whose EBV status was first determined by cytohybridization for nonpolyadenylated EBV RNAs (EBERs). Using both standard polymerase chain reaction (PCR) and PCR in situ hybridization, we successfully amplified sequences that span abnormally juxtaposed BamHI W and Z fragments characteristic of defective heterogeneous (het) EBV DNA from 10 of 32 (31%) EBER-positive tumors. Of 24 EBER-negative HD, 8 yielded PCR products indicating presence of het EBV DNA. Two of these contained defective EBV in the apparent absence of the prototype virus. Of the 42 tumors analyzed for defective EBV by both PCR techniques, there was concordance of results in 38 (90%). Detection of defective EBV genomes with the potential to disrupt viral gene regulation suggests one mechanism for pathogenic diversity that may also account for loss of prototypic EBV from individual tumor cells.     

Role of L3T4+ and 38+ T-cell subsets in resistance against infection with Treponema pallidum subsp. pertenue in hamsters.

The protective immunity conferred by T-cell subsets against infection with Treponema pallidum subsp. pertenue was studied. We demonstrated that hamster T cells can be separated into two subsets by monoclonal antibody (MAb) GK 1.5 (anti-L3T4) and MAb 38. Eighty-five percent of hamster thymocytes were L3T4+ and 87% were 38+ cells; 84% were dual positive for MAbs anti-L3T4 and 38. In the peripheral lymph nodes, however, the L3T4+ and 38+ T cells were mutually exclusive according to two-color immunofluorescence analysis. The two T-cell subsets were found to be functionally distinct according to their secretion of interleukin 2 (IL-2) when stimulated with concanavalin A. The L3T4+ cells secreted IL-2 and had characteristics of T helper cells, while the 38+ cells did not secrete IL-2 and appeared to be T cytotoxic-suppressor cells. Transfer of 4 x 10(6) helper or cytotoxic-suppressor T lymphocytes from T. pallidum subsp. pertenue-immune hamsters protected irradiated naive hamsters against challenge with this subspecies. IL-2 production could still be detected in the irradiated recipients 12 days after irradiation of naive recipients, although at a low level. This suggests that the remaining lymph node cells could support the survival and expansion of the infused cytotoxic-suppressor T cells. No accumulation of macrophages was observed in regional lymph nodes of immune T-cell recipients within 10 days of infection. Instead, there was an influx of polymorphonuclear neutrophils in all animals injected with T. pallidum subsp. pertenue. This report demonstrates that hamster T cells can be separated into two phenotypically and functionally distinct subsets and that both T-cell subsets confer protection against challenge with T. pallidum subsp. pertenue.     

Elimination of multiple reactions of the Phadebact Streptococcus coagglutination test.

Strong multiple reactions often occur with the Phadebact Streptococcus test when the culture contains blood. These reactions interfere with the identification of the Lancefield groups of streptococci. Group B streptococci from the vagina of pregnant women are difficult to identify by slide coagglutination because of the frequent presence of blood on culture swabs. Elimination of these multiple reactions caused by blood would permit rapid identification of group B streptococci in pregnant women. Vaginal broth cultures were examined to determine the cause of multiple reactions with slide coagglutination and to eliminate them from the testing procedure. Of 245 maternal broth cultures, 135 (55%) yielded multiple reactions when tested by coagglutination. Such reactions were either eliminated or greatly diminished by heating the broth sample to 90 degrees C for 10 min. It was also found that globulins in the serum may be responsible for multiple reactions with blood. This heating protocol will permit vaginal broth cultures to be rapidly tested for group B streptococci by slide coagglutination.     

The distribution of fetal nuchal translucency thickness in normal Korean fetuses

The aim of present study was to establish normative data for the distribution of nuchal translucency (NT) thickness in normal Korean fetuses. The data were collected from pregnant women with singleton pregnancies in whom fetal ultrasound was performed and the fetal NT thickness was measured between 11 and 14 weeks of gestation. Among them, a total of 2,577 fetuses with a known normal outcome were included in this study. The distribution of multiple of median (MoM) values of the NT thickness with crown-rump length (CRL) in 10-mm intervals and the 95th percentile of MoM were calculated with the linear regression method. The present study showed that NT measurements increase with increasing CRL and a false positive rate increases with increasing gestational age. Therefore, a fixed cut-off point through the first trimester was not appropriate and each NT measurement should be examined according to the gestational age. The present study offers normative data of the fetal NT thickness in a Korean population, which can be used as reference for screening chromosomal aberrations or other congenital abnormalities in the first trimester.     

Distribution of glycoconjugates during cochlea development in mice: light microscopic lectin study

During development, different epithelial cells in the mouse cochlea express different cell surface glycoconjugates, which may reflect membrane specialization. Some of the lectins tested in this study (SBA, succ-WGA, and PSA) labeled the sensory cells of the cochlea around birth. Other lectins (WGA, Con A, RCA-II, and PHA-E) labeled surfaces of the sensory cells, particularly the stereocilia, from early stages of development (gestation day (GD) 16) through 21 days after birth. These may be adhesion molecules needed to attach the newly forming tectorial membrane (TM) to the stereocilia. Lectin staining of the developing TM revealed that the substructures of the TM are biochemically distinct. Lectin staining also showed the temporal sequence of the expression of cytoplasmic glycoconjugates of the cochlear epithelium during development. Biochemical changes during development are probably the result of different cells being involved in the production of glycoconjugates, and may have functional significance, specifically with regard to the expression of adhesion and/or signaling molecules.     

Functionally active complement proteins C6 and C7 detected in C6- and C7-deficient individuals

Two sensitive sandwich ELISAs based on monoclonal antibodies directed to native C6 and C7 allowed the detection and quantitation of these complement proteins in 20 out of 37 serum samples from individuals who had previously been classified as deficient in these proteins as assessed by immunochemical and/or functional assays. Furthermore, serum from four C6-deficient and one combined C6-/C7-deficient individual showed an increase in the terminal complement complex (TCC) and a decrease in native C6 and C7 after complement activation as assayed by specific ELISAs. Despite their (incomplete) deficiencies, these individuals therefore possess functionally active terminal complement proteins with respect to their ability to generate the TCC. As these individuals have no history of a susceptibility to neisserial infections, even low concentrations of functionally active C6 and C7 may provide sufficient protection against those micro-organisms whose destruction requires TCC formation.