Growth arrest and DNA damage-45 alpha (GADD45alpha).

Regulation of cell cycle and growth is integral for cell survival. The intricate mechanisms that control proliferation and cell cycle are numerous. The growth arrest and DNA damage (GADD)-inducible gene family is often up-regulated in response to various environmental stresses and drug therapies. GADD45alpha was the first stress-inducible gene determined to be up-regulated by p53 and is also a target for the p53 homologues, p63 and p73. When GADD45alpha is deleted or repressed, cells show uncontrolled proliferation. Furthermore, decreased GADD45alpha expression is also considered a survival mechanism, as cancer cells without this control can evade the apoptotic pathway leading to increased tumourigenesis. Drug therapies can act to directly or indirectly up-regulate GADD45alpha and promote apoptosis. As GADD45alpha is an essential component of many metabolic pathways that control proliferating cancer cells, it presents itself as an emerging drug target worthy of further investigation.

Role and mechanism of phosphatidylinositol-specific phospholipase C in survival and virulence of Cryptococcus neoformans.

Phospholipase B1 (Plb1) is secreted after release from its glycosylphosphatidylinositol anchor and is implicated in initiation and dissemination of infection of the pathogenic fungus, Cryptococcus neoformans. To investigate the role of phosphatidylinositol-specific phospholipase C (PI-PLC) in Plb1 secretion, we identified two putative PI-PLC-encoding genes in C. neoformans var. grubii (PLC1 and PLC2), and created Deltaplc1 and Deltaplc2 deletion mutants. In Deltaplc1, which expressed less PI-PLC activity than wild type (WT), three major cryptococcal virulence traits, Plb1 secretion, melanin production and growth at host temperature (37 degrees C) were abolished and absence of Plb1 secretion coincided with Plb1 accumulation in plasma membranes. In addition, Deltaplc1 cell walls were defective, as indicated by cell clumping and irregular morphology, slower growth and an inability to activate mitogen-activated protein kinase (MAPK) in the presence of cell wall-perturbing agents. In contrast to Deltaplc2, which was as virulent as WT, Deltaplc1 was avirulent in mice and exhibited attenuated killing of Caenorhabditis elegans at 25 degrees C, demonstrating that mechanism(s) independent of the 37 degrees C growth defect contribute to the virulence composite. We conclude that Plc1 is a central regulator of cryptococcal virulence, acting through the protein kinase C/MAPK pathway, that it regulates release of Plb1 from the plasma membrane and is a candidate antifungal drug target.

Cell wall-linked cryptococcal phospholipase B1 is a source of secreted enzyme and a determinant of cell wall integrity.

Phospholipase B (Plb1) is secreted by pathogenic fungi and is a proven virulence determinant in Cryptococcus neoformans. Cell-associated Plb1 is presumptively involved in fungal membrane biogenesis and remodelling. We have also identified it in cryptococcal cell walls. Motif scanning programs predict that Plb1 is attached to cryptococcal membranes via a glycosylphosphatidylinositol (GPI) anchor, which could regulate Plb1 export and secretion. A functional GPI anchor was identified in cell-associated Plb1 by (G)PI-specific phospholipase C (PLC)-induced release of Plb1 from strain H99 membrane rafts and inhibition of GPI anchor synthesis by YW3548, which prevented Plb1 secretion and transport to membranes and cell walls. Plb1 containing beta-1,6-linked glucan was released from H99 (wild-type strain) cell walls by beta-1,3 glucanase, consistent with covalent attachment of Plb1 via beta-1,6-linked glucans to beta-1,3-linked glucan in the central scaffold of the wall. Naturally secreted Plb1 also contained beta-1,6-linked glucan, confirming that it originated from the cell wall. Plb1 maintains cell wall integrity because a H99 deletion mutant, DeltaPLB1, exhibited a morphological defect and was more susceptible than H99 to cell wall disruption by SDS and Congo red. Growth of DeltaPLB1 was unaffected by caffeine, excluding an effect of Plb1 on cell wall biogenesis-related signaling pathways. Environmental (heat) stress caused Plb1 accumulation in cell walls, with loss from membranes and reduced secretion, further supporting the importance of Plb1 in cell wall integrity. This is the first demonstration that Plb1 contributes to fungal survival by maintaining cell wall integrity and that the cell wall is a source of secreted enzyme.

Lipid rafts in Cryptococcus neoformans concentrate the virulence determinants phospholipase B1 and Cu/Zn superoxide dismutase.

Lipid rafts have been identified in the membranes of mammalian cells, the yeast Saccharomyces cerevisiae, and the pathogenic fungus Candida albicans. Formed by a lateral association of sphingolipids and sterols, rafts concentrate proteins carrying a glycosylphosphatidylinositol (GPI) anchor. We report the isolation of membranes with the characteristics of rafts from the fungal pathogen Cryptococcus neoformans. These characteristics include insolubility in Triton X-100 (TX100) at 4 degrees C, more-buoyant density within a sucrose gradient than the remaining membranes, and threefold enrichment with sterols. The virulence determinant phospholipase B1 (PLB1), a GPI-anchored protein, was highly concentrated in raft membranes and could be displaced from them by treatment with the sterol-sequestering agent methyl-beta-cyclodextrin (MbetaCD). Phospholipase B enzyme activity was inhibited in the raft environment and increased 15-fold following disruption of rafts with TX100 at 37 degrees C. Treatment of viable cryptococcal cells in suspension with MbetaCD also released PLB1 protein and enzyme activity, consistent with localization of PLB1 in plasma membrane rafts prior to secretion. The antioxidant virulence factor Cu/Zn superoxide dismutase (SOD1) was concentrated six- to ninefold in raft membrane fractions compared with nonraft membranes, whereas the cell wall-associated virulence factor laccase was not detected in membranes. We hypothesize that raft membranes function to cluster certain virulence factors at the cell surface to allow efficient access to enzyme substrate and/or to provide rapid release to the external environment.

Acute psychosis due to treatment with phenytoin in a nonepileptic patient.

The development of psychosis related to antiepileptic drug treatment is usually attributed to the interaction between the epileptic brain substratum and the antiepileptic drugs. The case of a nonepileptic patient who developed psychosis following phenytoin treatment for trigeminal neuralgia is described. This case suggests that the psychotic symptoms that occur following phenytoin treatment in some epileptic patients may be the direct result of medication, unrelated to seizures.

Hemispheric predominance of abnormal findings in electroencephalogram (EEG).

The EEGs of 13,560 patients have been reviewed in order to determine whether abnormal findings, epileptiform or not, have a hemispheric dominance. We have included outpatients and hospitalized patients as well. Eight hundred and thirty-five EEGs had generalized abnormal findings, and 414 EEGs had lateralized abnormal findings. The EEGs of 322 patients (77.7%) had a left predominance, and those of 92 patients (22.3%) had a right predominance, of abnormal findings. A strong left predominance has been noted for the epileptiform discharges, i.e. 128 (79%) vs. 34 (21%). These results raise the possibility that the left hemisphere may be more vulnerable to nosological processes.

Convulsive status epilepticus following abrupt high-dose benzodiazepine discontinuation.

The misuse of benzodiazepines (BNZ)s may result in serious side effects. Three cases of convulsive status epilepticus (CSE) following abrupt discontinuation of long-term use of 25 mg of lorazepam in one patient and more than 20 mg of flunitrazepam in two patients are presented; they were non-epileptics and free of other high-risk factors for seizures. A favorable outcome for all three cases was noted. They remain free of seizures without antiepileptic treatment. Nevertheless, because of the extensive use of benzodiazepines, such rare high-risk side effects must be emphasized.

The usefulness of sleep and sleep deprivation as activating methods in electroencephalographic recording: contribution to a long-standing discussion.

Sedated sleep and sleep deprivation are commonly used methods to increase the diagnostic yield of the electroencephalogram (EEG), especially in the evaluation of people with epilepsy, but the rate of activation achieved by them is controversial, as is the issue of whether it is sleep itself, or sleep deprivation which is responsible for their alleged efficacy. We retrospectively studied the EEGs of epileptic patients, examined in our laboratory, who, after having undergone an inconclusive initial routine recording, had then been examined with a second recording. This was after either: (1) sleep deprivation with evidence of drowsiness in the recordings, (2) sleep deprivation without drowsiness (indicative of the effect which sleep deprivation per se has in eliciting abnormal patterns), or (3) drug-induced sedation. The activation rates found were (1) 22.5%, (2) 24% (22.6% for sleep deprivation collectively, regardless of the presence or not of subsequent drowsiness) and (3) 27% respectively. Only the sleep deprivation rate was statistically different from the 9.6% increased rate of abnormal patterns elicited by the simple repeating of a second routine recording, while the rate of drug-induced sleep was not. Although, sleep deprivation appeared to be more effective as an activating method of EEG compared with sedated sleep, no conclusions could be drawn about which stage of sleep, wakefulness or drowsiness, is primarily responsible for the method's efficacy.

A case of fever following antiepileptic treatment.

A 23-year-old female patient treated with 900 mg oxcarbazepine for complex partial seizures is presented. Good seizure control and slight fever were noted a few weeks after drug administration. Reduction of oxcarbazepine and replacement with valproate resulted in a transient normothermia. Because of fever reappearance, vigabatrin was added and valproate was gradually reduced. Seizures reappeared, but the body temperature fell below 37 degrees C. Substitution of valproate for lamotrigine resulted in seizure control but abnormal body temperature (37- 37.6 degrees C) was noted again. Repeated hospital admission for clinical and laboratory investigation before any change of treatment revealed no other abnormal findings. The patient's abnormal temperature possibly reflects a derangement of high-level temperature control.

Electrophysiological study (EEG, VEPs, BAEPs) in patients with Charcot Marie Tooth (type HMSN I) disease.

A laboratory investigation consisting of EEG recordings, BAEPs and VEPs evaluation as well as estimation of the facial nerve distal latency was performed in 9 patients who had Charcot Marie Tooth (type HMSN I) disease. 3 patients showed VEP's abnormalities (2 of them had prolonged P100 latency and one had an abnormal interocular latency difference). Another patient showed an upper normal limit value in the interocular latency difference. Abnormal BAEPs were found in 8 patients (one had I-III/IPLD prolonged, 2 of them had the latency of wave I prolonged and the remaining 6 had no readable or repeatable responses unilaterely or bilaterally). 7 out of 9 patients had the facial nerve distal latency prolonged, without any evident clinical facial weakness. Abnormal EEG recordings were found in 2 of all tested patients. Our results provide some evidence that in Charcot Marie Tooth disease the involvement of the II, VII and VIII nerves is more frequent than clinically expected and is probably related to a demyelinated process.

[Contribution of visual evoked potentials, electronystagmography and the blink reflex to the diagnosis of multiple sclerosis]. / Apport des potentiels évoqués visuels, de l'électro-nystagmographie et du réflexe de clignement au diagnostic de la sclérose en plaques.

One hundred and forty six cases of multiple sclerosis were investigated and classified according to McAlpine's criteria into 3 group: probable (40 cases), possible (51 cases) and definite (55 cases). In every patient visual evoked potentials, electronystagmography and blink reflex were performed in the same session. Percentages of abnormal findings were 66 p. cent for VEP, 63 p. cent for ENG and 53 p. cent for blink reflex. These 3 methods can detect subclinical lesions and in reveal new sites of demyelination. Since in only 14 cases (10%) were the results of all 3 methods normal, it is concluded that simultaneous examination by these methods can contribute to the diagnosis.

Side-effects of lithium.

An account of the side-effects of 30 patients treated with lithium at Athens University Department of Psychiatry during the last two years is presented. The most frequent side-effects were: tremor of bands, polydipsia, polyuria, increase in appetite, dryness of mouth, general muscular weakness and memory reduction. Most side-effects were infrequent, mild and in no case necessitated discontinuation of treatment.