Mutant herpes simplex virus-mediated suppression of retinoblastoma.

PURPOSE: To test the ability of a mutant herpes simplex virus (HSV) hrR3 to inhibit growth of Y79 human retinoblastoma in vitro and in vivo. METHODS: Cultured Y79 cells were infected with multiplicities of infection (MOI) ranging from 0.004 to 0.1 of hrR3. Surviving cells were counted using trypan blue dye exclusion. Using X-gal staining, expression of the lacZ gene was examined in vitro on day 3 postinfection to evaluate viral replication. Nude mice harboring Y79 tumors subcutaneously received an intraneoplasmic injection of 5 x 10(7) plaque-forming units of hrR3. The tumor sizes were measured weekly. Expression of the lacZ gene was also examined on one week postinfection. RESULTS: There are 31% and 13% cells surviving in cultured Y79 cells infected by hrR3 at an MOI of 0.1 on days 3 and 5 postinfection respectively compared to those of mock-infected cells. Also more than 70% of Y79 cells were stained with X-gal at an MOI of 0.1 which demonstrated active viral replication in vitro. Virus-treated subcutaneous tumors were smaller than control tumors (p<0.05, Student's t-test) on days 14, 21, and 28 postinfection. Positive X-gal staining was also observed in the tumor nodule which was challenged with this viral vector. CONCLUSIONS: We have demonstrated that hrR3 is capable of inhibiting Y79 tumor growth both in cell culture and in nude mice. These data suggest that gene therapy using this mutant HSV vector can be a new supplementary therapeutic modality for retinoblastoma.     

GABA(A) receptor-mediated inspiratory termination evoked by vagal stimulation in decerebrate cats.

To identify the GABAergic inhibitory mechanisms involved in inspiratory termination or off-switching (IOS), the effects of a specific enhancer of GABA(A) receptors, midazolam, and an antagonist, bicuculline, on vagally evoked inspiratory inhibitions and IOS were investigated in decerebrate cats. Stimulation of vagal afferents at late inspiration provoked either reversible inspiratory inhibition or IOS, depending on the stimulus intensity. Each response occurred at a constant latency (phase 1). The reversible response was triphasic, consisting of an early (phase 2) inhibition, a brief (phase 3) excitation and a late (phase 4) inhibition in the phrenic neurogram, and early (phase 2) IPSPs, brief (phase 3) EPSPs and late (phase 4) IPSPs in bulbar inspiratory (I) neurones. With an increasing stimulus intensity, phase 4 inhibitions were increased in amplitude and duration, leading to IOS. Midazolam (0.1 mg/kg i.v.) increased more selectively phase 4 IPSPs than phase 2 IPSPs in I neurones, and decreased the threshold for evoking IOS by producing an earlier and larger phase 4 IPSPs. Bicuculline (1.0 mg/kg i.v.) had an opposite effect. These results suggest that the late inhibitory response evoked by vagal stimulation in the I neuronal pool organizes an initial phase of IOS which is mediated by GABA(A) receptors.     

Effects of ethyl alcohol administration to rat dams during the gestation period on learning behavior and on levels of monoamines and metabolites in rat pup brain after birth

Pregnant rats were given 0%-, 5%-, 10%- and 20% ethyl alcohol in drinking water during the gestation period.We evaluated the brain function of pups born of alcohol-administered dams. Learning ability (Sidman avoidance behavior), the amounts of monoamines (noradrenalin, dopamine, serotonin) and metabolites (3,4-dihydroxyphenyl acetic acid [DOPAC], homovanillic acid [HVA] and 5-hydroxyindole acetic acid [5-HIAA])in whole brain were examined for neurobehavioral and neurochemical effects.There was no effect on Sidman avoidance behavior in 56-day-old offspring, but alterations of the amounts of monoamines and their metabolites were observed even in 66-day-old offspring as a result of the dams' exposure to ethanol during pregnancy.     

Changes in proton magnetic resonance spectroscopy and Wechsler adult intelligence scale revised after clipping of unruptured aneurysms

The purpose of this study is to evaluate the influence of surgery for unruptured aneurysms on neuropsychological status and cerebral metabolism. We studied 21 patients with unruptured aneurysms treated with direct surgery accompanied by craniotomy, rather than by the endovascular method. Patients were evaluated before and after surgery, using the Wechsler adult intelligence scale revised (WAIS-R) and proton magnetic resonance spectroscopy which measured the ratio of N-acetyl-aspartate to creatine (NAA/Cr). Although the results of WAIS-R (total IQ, verbal IQ and performance IQ) was not significantly different after surgery in any of the patients, the total IQ of the patients with anterior communicating artery aneurysms (AcoA) showed a significant decline compared with patients with other aneurysms after surgery. Five of eight AcoA patients showed a specific reduction in verbal IQ, suggesting deterioration of recent memory. The NAA/Cr remained within the normal range and was not significantly different before and after surgery. However, the NAA/Cr in the white matter of the frontal lobe of AcoA patients showed a significant reduction compared with that of non-AcoA patients. All three elderly patients older than 70 showed a reduction in NAA/Cr of more than one standard deviation from normal subjects in their frontal or parietal lobes. These results indicated that operation for unruptured aneurysms is reliable and well established but they also show that careful consideration should be given to possible deterioration in neuropsychological status and cerebral metabolism after operation in AcoA and elderly patients.     

Thiamin is decomposed due to Anaphe spp. entomophagy in seasonal ataxia patients in Nigeria

A fairly high activity of a relatively heat-resistant thiaminase was detected and characterized from the pupae of an African silkworm Anaphe spp. which had been the putative cause of a seasonal ataxia and impaired consciousness in Nigerians. The thiaminase in the buffer extract of Anaphe pupae was type I (thiamin: base 2-methyl-4-aminopyrimidine methyl transferase EC 2.5.1.2), and the optimal temperature and pH were 70 degrees C and 8.0-8.5, respectively. Based on gel filtration chromatography, the molecules were estimated to be 200 kDa. Second substrates which could be utilized by the thiaminase were pyridoxine, amino acids, glutathione, taurine and 4-aminopyridine. Thiamin phosphate esters were inactive as substrates. This is the first report describing an insect thiaminase. Our results indicate the necessity of thorough heat treatment for the detoxification of the African silkworm, making the worm a safe source of high-quality protein.     

A study in cases of Brugada-type electrocardiogram and its management proposals in health examination]

In 1992, Brugada et al. reported a characteristic electrocardiogram (ECG) pattern and ST-segment elevation in leads V1 to V3 associated with sudden death in patients without demonstrable structural heart disease. That disease is now called Brugada Syndrome. The diagnostic criteria for the Brugada Syndrome have still not been decided on, and the prevalence of Brugada type ECG (B-ECG) varies widely in Japan. Therefore, we should consider B-ECG according to the consensus statement from the European Society of Cardiology and we proposed its management in health examinations. There were 35 B-ECG cases (0.9%), all male out of 3,875 Postal Service Trainees. There were 5 cases of Type I (Coved) (0.13%), 21 cases of Type II (0.54%), and 9 cases of Type III (0.23%), Only one case (0.026%) of Brugada Syndrome was found, and eventually, he received an Implantable Cardioverter Defibrillator (ICD). Type I (Coved) may be a more important electrocardiographic factor having a stronger causal relation to Ventricular Arrhythmia. Therefore, in management of health examinations, Type I patients with syncope or a family history of sudden cardiac death should visit a cardiologist for ICD-implantation, and even without any cardiac symptoms (syncope and a family history of sudden death), they are advised to visit a cardiologist for a program electrical stimulation (PES). Type II and III patients with any cardiac symptoms are advised to visit a cardiologist for PES or a drug challenge.     

In vitro metabolism of the calmodulin antagonist DY-9760e (3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate) by human liver microsomes: involvement of cytochromes p450 in atypical kinetics and potential drug interactions.

Human cytochrome P450 (P450) isozyme(s) responsible for metabolism of the calmodulin antagonist 3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate (DY-9760e) and kinetic profiles for formation of its six primary metabolites [M3, M5, M6, M7, M8, and DY-9836 (3-[2-[4-(3-chloro-2-methylphenyl)piperazinyl]ethyl]-5,6-dimethoxyindazole)] were identified using human liver microsomes and recombinant P450 enzymes. In vitro experiments, including an immunoinhibition study, correlation analysis, and reactions with recombinant P450 enzymes, revealed that CYP3A4 is the primary P450 isozyme responsible for the formation of the DY-9760e metabolites, except for M5, which is metabolized by CYP2C9. Additionally, at clinically relevant concentrations, CYP2C8 and 2C19 make some contribution to the formation of M3 and M5, respectively. The formation rates of DY-9760e metabolites except for M8 by human liver microsomes are not consistent with a Michaelis-Menten kinetics model, but are better described by a substrate inhibition model. In contrast, the enzyme kinetics for all metabolites formed by recombinant CYP3A4 can be described by an autoactivation model or a mixed model of autoactivation and biphasic kinetics. Inhibition of human P450 enzymes by DY-9760e in human liver microsomes was also investigated. DY-9760e is a very potent competitive inhibitor of CYP2C8, 2C9 and 2D6 (Ki 0.25-1.7 microM), a mixed competitive and noncompetitive inhibitor of CYP2C19 (Ki 2.4 microM) and a moderate inhibitor of CYP1A2 and 3A4 (Ki 11.4-20.1 microM), suggesting a high possibility for human drug-drug interaction.     

Pronounced synergistic promotion of N-bis(2-hydroxypropyl)nitrosamine-initiated thyroid tumorigenesis in rats treated with excess soybean and iodine-deficient diets.

We have reported that excess soybean treatment and iodine deficiency synergistically interact, resulting in remarkable induction of thyroid hyperplasias in rats. In the present study, modifying effects of excess soybean and iodine-deficient diets were investigated in the post-initiation phase of N-bis(2-hydroxypropyl)nitrosamine [DHPN]-initiated thyroid tumorigenesis in rats. AIN-93G in which casein was replaced with gluten was used as a basal diet to avoid possible iodine contamination. In Experiment 1, F-344 rats of both sexes were sc injected with DHPN at a dose of 2800 mg/kg body weight and then fed a diet containing 0%, 0.8%, 4%, or 20% defatted soybean for 12 weeks, with proportional replacement of gluten by soybean flour. Although no thyroid proliferative lesions were found in any group, the absolute thyroid weights were significantly (p < 0.01) elevated with the 20% soybean treatment. In Experiment 2, after similar sc injection of DHPN, rats were fed a basal diet or a diet containing 20% soybean under iodine normal or deficient conditions for 12 weeks. Soybean feeding to both sexes under iodine deficient but not normal conditions dramatically enhanced the development of thyroid follicular adenomas (p < 0.01) and adenocarcinomas (p < 0.05), in good agreement with decrease in thyroxine and increase in thyroid-stimulating hormone. Thus co-exposure to excess soybean and iodine deficiency results in synergistic promotion of DHPN-initiated thyroid tumorigenesis in rats, of which mechanisms appear to primarily involve effects on serum hormone levels.