Differential expression of sense and antisense transcripts of the mitochondrial DNA region coding for ATPase 6 in fetal and adult porcine brain: identification of novel unusually assembled mitochondrial RNAs.

The mammalian mitochondrial genome is a double-stranded circular DNA molecule, which is transcribed from both strands as polycistronic RNAs, which are further processed to yield the mature polyadenylated mRNAs, rRNAs and tRNAs. We compared the gene expression patterns of foetal and adult porcine brains and identified a sequence tag from the ATPase 6 region of the mitochondrial genome which, in adult brain, was more abundant in the sense (H-strand) form, but, in foetal brain, more abundant in the antisense form (L-strand). By means of solution hybridisation/S1 nuclease protection assay, Northern blotting, and PCR based techniques, we demonstrated that the ATPase 6 region of the porcine mitochondrial genome is transcribed as co-existing, stable sense and antisense RNAs. Furthermore, we identified sense and antisense transcripts from this region consisting of inversely assembled fragments joined together at a direct repeat of 7 nucleotides. Our results suggest that transcription and post-transcriptional processing of mitochondrial RNAs are much more complex than presently thought.

Intravenous granulocyte colony-stimulating factor increases the release of tumour necrosis factor and interleukin-1beta into the cerebrospinal fluid, but does not inhibit the growth of Streptococcus pneumoniae in experimental meningitis.

Granulocyte colony-stimulating factor (G-CSF) possesses an antimicrobial effect in several animal models of infection. To evaluate a possible effect of G-CSF on the course of pneumococcal meningitis, rabbits infected intracisternally (i.c.) with Streptococcus pneumoniae type 3 (n = 7) received 50 microgram/kg of rhG-CSF intravenously (i.v.) 1 h prior to infection. Seven infected animals served as controls. Uninfected rabbits received 10 microgram of G-CSF (n = 3), 2 microgram G-CSF (n = 3) or saline (n = 3) i.c. G-CSF injected i.c. did not produce cerebrospinal fluid (CSF) leucocytosis. Compared with the control group, i.v. G-CSF given prior to i.c. infection increased the percentage of granulocytes in blood 6 h and 12 h after infection. Twelve hours after infection, CSF tumour necrosis factor (TNF) activity and CSF interleukin (IL)-1beta concentrations were significantly higher in G-CSF-treated animals. G-CSF did not decrease bacterial growth in the subarachnoid space and the CSF leucocyte densities were not influenced. At 24 h after infection, G-CSF did not reduce the CSF concentrations of neurone-specific enolase and the density of apoptotic neurones in the dentate gyrus of the hippocampus. In conclusion, i.v. G-CSF increased the concentration of pro-inflammatory cytokines in the CSF but did not decrease the growth of Streptococcus pneumoniae in the subarachnoid space.

Glycerol does not reduce neuronal damage in experimental Streptococcus pneumoniae meningitis in rabbits.

To study the effect of high-dose glycerol therapy on inflammation and neuronal destruction in a model of experimental pneumococcal meningitis, 14 New Zealand White rabbits were infected intracisternally with Streptococcus pneumoniae type 3. Sixteen hours after infection, 7 animals received intravenous glycerol therapy (1 g kg(-1) bolus and 0.5 g kg (1)h(-1) maintenance dose) and 7 animals served as untreated controls.After 8 h of therapy, the glycerol CSF:serum ratio exceeded the previously observed values in rabbits with an intact blood-CSF barrier (0.72+/-0.25 vs. 0.35), i.e. glycerol crossed the blood-CSF barrier more readily in animals altered by meningitis than in healthy animals. In contrast, the brain tissue:serum ratio of glycerol (grey matter 0.33+/-0.29, white matter 0.30+/-0.31) was substantially lower than the CSF:serum ratio (p=0.03 and p=0.047). There was no significant effect of glycerol on intracranial pressure, brain water content and neuron-specific enolase release into the CSF. Glycerol significantly increased the density of neuronal apoptosis in the dentate gyrus of the hippocampal formation. Therefore, glycerol does not appear to be beneficial in experimental pneumococcal meningitis.

Trovafloxacin delays the antibiotic-induced inflammatory response in experimental pneumococcal meningitis.

This study evaluates the ability of the new fluoroquinolone trovafloxacin to attenuate the inflammatory burst known to occur after initiation of antibiotic treatment in pneumococcal meningitis. After exposure to trovafloxacin or ceftriaxone for 3 h in vitro, Streptococcus pneumoniae was injected intracisternally (i.c.) into rabbits every 3 h over 9 h (n = 6 for each antibiotic). Ceftriaxone-treated S. pneumoniae induced consistently higher CSF leucocyte counts (median 2568/microL versus 543/microL at 6 h; P = 0.03; 4560/microL versus 2207/microL at 18 h; P = 0.03) than trovafloxacin-treated bacteria. Meningitis induced in rabbits by i.c. injection of live S. pneumoniae was treated with equal doses of trovafloxacin or ceftriaxone i.v. (ten per group). The bactericidal rates of both antibacterial agents in CSF were almost identical. In comparison with ceftriaxone, trovafloxacin resulted in lower tumour necrosis factor (TNF) and interleukin 1beta (IL-1beta) CSF levels 2 h after the initiation of treatment (TNF levels, median 26 U/mL versus 141 U/mL; P = 0.02; IL-1beta levels 455 pg/mL versus 1399 pg/mL; P = 0.02). Twelve hours after initiation of therapy, however, TNF and IL-1beta were higher in trovafloxacin-treated animals (TNF, 61 U/mL versus 7 U/mL; P = 0.001; IL-1beta, 4320 pg/mL versus 427 pg/mL; P = 0.006). The increase in CSF lactate was less during trovafloxacin therapy than with ceftriaxone (median: 2.0 mmol/L versus 4.0 mmol/L; P = 0.03). In conclusion, S. pneumoniae treated in vitro with trovafloxacin induced less CSF leucocytosis than ceftriaxone-treated S. pneumoniae. After i.c. inoculation of live S. pneumoniae, trovafloxacin therapy delayed, but did not inhibit, the release of the proinflammatory cytokines TNF and IL-1beta, probably by slowing the liberation of bacterial cell wall components into the subarachnoid space.

Increased glutamine synthetase immunoreactivity in experimental pneumococcal meningitis.

Glutamine synthetase (GS), glial fibrillary acidic protein (GFAP) immunohistochemistry and neuronal apoptotic cell death were evaluated in a rabbit model of pneumococcal meningitis. Meningitis caused an increase of GS immunoreactivity in the cerebral cortex, but not in the hippocampal formation. GFAP immunoreactivity remained unchanged. This may represent a protective mechanism for cortical neurons. The inability of hippocampal GS to counteract the detrimental effects of glutamate may be the cause of neural apoptosis observed in the dentate gyrus during meningitis.