Clostridium perfringens type C causing necrotising enteritis.

A rapidly fatal case of enteritis necroticans in a 24 year old man with diabetes was caused by Clostridium perfringens type C. The role of beta toxin in the disease is discussed. This type has not been previously described as a causative agent in necrotising bowel disease of man outside endemic areas.     

Mitotic and post mitotic consequences of genomic instability induced by oncogenic Ha-Ras

Induced expression of a mutant human Ha-ras oncogene in NIH3T3 cells leads to the rapid production of multicentric chromosomes, acentric chromosome fragments, double minute chromosomes, increased heteroploidy, and increased capacity to undergo gene amplification. In this study we have used fluorescent-in-situ hybridization (FISH) to demonstrate that induction of the Ha-ras oncogene also leads to disruption of the mitotic machinery, resulting in aberrant mitoses and abnormal daughter cells. Cells induced to express an oncogenic Ha-ras transgene accumulate chromosomes that lag outside of the rest of the chromosomal architecture, chromosomes that form bridges between daughter nuclei at anaphase, and that form micronuclei. Many of these mitotic aberrations contain structurally abnormal chromosomes. Theseras-induced changes were suppressed by the introduction of a gene encoding the dominant negative effector ofras, raf 301. Expression ofraf301 in cells induced to express Ha-ras reduced the level of growth in soft agar, chromosome aberrations, mitotic aberrations, and frequency of gene amplification. These data provide evidence for an association between Ha-ras induced transformation, chromosome aberrations and gene amplification. Furthermore they offer insight into how the cell responds to the formation of aberrant chromosomes, and how disrupting chromosomal architecture could lead to further imbalances in the distribution of genetic material between daughter cells.     

Expression and regulation of small proline-rich protein 2 in allergic inflammation

Asthma is a complex inflammatory pulmonary disorder that is on the rise despite intense ongoing research. We aimed to elucidate novel pathways involved in the pathogenesis of asthma. Employing asthma models induced by different allergens (ovalbumin and Aspergillus fumigatus), we uncovered the involvement of two members of the small proline-rich protein (SPRR) family, SPRR2a and SPRR2b, known to be involved in epithelial differentiation but not allergic disease. In situ hybridization revealed induction of SPRR2 signal in a subset of bronchial epithelial cells and mononuclear cells associated with inflammation after allergen challenge. Allergen-induced SPRR2 mRNA accumulation in the lung occurred in a time-dependent manner, with peak expression 10-96 h after a second ovalbumin challenge. Transgenic overexpression of interleukin (IL)-13 in the lungs resulted in a marked increase of SPRR2 expression, and allergen-induced SPRR2 expression was significantly decreased in IL-13-deficient mice. Studies in gene-targeted mice revealed that allergen-induced SPRR2 was dependent upon STAT6. Finally, we aimed to determine if the induction of SPRR2 by allergen was tissue specific. Notably, SPRR2 was markedly increased in the small intestine after induction of allergic gastrointestinal inflammation. Thus, SPRR2 is an allergen- and IL-13-induced gene in experimental allergic responses that may be involved in disease pathophysiology.     

pH Sensitivity of non-synaptic field bursts in the dentate gyrus

Under conditions of low [Ca(2+)](o) and high [K(+)](o), the rat dentate granule cell layer in vitro develops recurrent spontaneous prolonged field bursts that resemble an in vivo phenomenon called maximal dentate activation. To understand how pH changes in vivo might affect this phenomenon, the slices were exposed to different extracellular pH environments in vitro. The field bursts were highly sensitive to extracellular pH over the range 7.0-7.6 and were suppressed at low pH and enhanced at high pH. Granule cell resting membrane potential, action potentials, and postsynaptic potentials were not significantly altered by pH changes within the range that suppressed the bursts. The pH sensitivity of the bursts was not altered by pharmacologic blockade of N-methyl-D-aspartate (NMDA), non-NMDA, and GABA(A) receptors at concentrations of these agents sufficient to eliminate both spontaneous and evoked synaptic potentials. Gap junction patency is known to be sensitive to pH, and agents that block gap junctions, including octanol, oleamide, and carbenoxolone, blocked the prolonged field bursts in a manner similar to low pH. Perfusion with gap junction blockers or acidic pH suppressed field bursts but did not block spontaneous firing of single and multiple units, including burst firing. These data suggest that the pH sensitivity of seizures and epileptiform phenomena in vivo may be mediated in large part through mechanisms other than suppression of NMDA-mediated or other excitatory synaptic transmission. Alterations in electrotonic coupling via gap junctions, affecting field synchronization, may be one such process.     

Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography

In this study, we have used time-lapse video cinematography to study fertilization in 50 human oocytes that had undergone intracytoplasmic sperm injection (ICSI). Time-lapse recording commenced shortly after ICSI and proceeded for 17-20 h. Oocytes were cultured in an environmental chamber which was maintained under standard culture conditions. Overall, 38 oocytes (76%) were fertilized normally, and the fertilization rate and embryo quality were not significantly different from 487 sibling oocytes cultured in a conventional incubator. Normal fertilization followed a defined course of events, although the timing of these events varied markedly between oocytes. In 35 of the 38 fertilized oocytes (92%), there were circular waves of granulation within the ooplasm which had a periodicity of 20-53 min. The sperm head decondensed during this granulation phase. The second polar body was then extruded, and this was followed by the central formation of the male pronucleus. The female pronucleus formed in the cytoplasm adjacent to the second polar body at the same time as, or slightly after, the male pronucleus, and was subsequently drawn towards the male pronucleus until the two abutted. Both pronuclei then increased in size, the nucleoli moved around within the pronuclei and some nucleoli coalesced. During pronuclear growth, the organelles contracted from the cortex towards the centre of the oocyte, leaving a clear cortical zone. The oocyte decreased in diameter from 112 to 106 microm (P < 0.0001) during the course of the observation period. The female pronucleus was significantly smaller in diameter than the male pronucleus (24.1 and 22.4 microm respectively, P = 0.008) and contained fewer nucleoli (4.2 and 7.0 respectively, P < 0.0001). After time-lapse recording, oocytes were cultured for 48 h prior to embryo transfer or cryopreservation. Embryo quality was related to fertilization events and periodicity of the cytoplasmic wave, and it was found that good quality embryos arose from oocytes that had more uniform timing from injection to pronuclear abuttal and tended to have a longer cytoplasmic wave. In conclusion, we have shown that time-lapse video cinematography is an excellent tool for studying fertilization and early embryo development, and have demonstrated that human fertilization comprises numerous complex dynamic events.      

Expression, Structure, and Location of Epitopes of the Major Surface Glycoprotein of Pneumocystis carinii f. sp. carinii

The major surface glycoprotein (MSG) of Pneumocystis carinii f. sp. carinii consists of a heterogeneous family of proteins that are encoded by approximately 100 unique genes. A genomic expression library was screened with a panel of MSG-specific monoclonal antibodies (MAbs) to identify conserved and rare epitopes. All of the antibodies reacted with epitopes that are encoded within the 5′ end of MSG. The results from the expression screening identified antibodies that recognize highly conserved, moderately conserved, and rare epitopes. Four MAbs (MAbs RA-F1, RA-E7, RA-G10, and RB-E3) reacted with a maltose binding protein–MSG-B fusion protein (^MBPMSG-B_41–1065) by immunoblotting and enzyme-linked immunosorbent assay. Three of the MAbs (MAbs RA-F1, RA-G10, and RA-E7) reacted with the same continuous epitope that was localized to amino acids 278 to 290 of MSG-B. Comparison of the sequence of the RA-F1-, RA-G10-, and RA-E7-reactive epitope to the deduced amino acid sequences of multiple MSGs demonstrated that it is highly conserved. The reactivity of RB-E3 with MSG-B was shown to be dependent on amino acids 184 to 192, which may comprise a portion of a discontinuous epitope.