A comparison of Ca2+ channel blocking mode between gabapentin and verapamil: implication for protection against hypoxic injury in rat cerebrocortical slices

1 The mode of Ca(2+) channel blocking by gabapentin [1-(aminomethyl)cyclohexane acetic acid] was compared to those of other Ca(2+) channel blockers, and the potential role of Ca(2+) channel antagonists in providing protection against hypoxic injury was subsequently investigated in rat cerebrocortical slices. 2 mRNA for the alpha(2)delta subunits of Ca(2+) channels was found in rat cerebral cortex. 3 Nitric oxide (NO) synthesis estimated from cGMP formation was enhanced by KCl stimulation, which was mediated primarily by the activation of N- and P/Q-type Ca(2+) channels. Gabapentin blocked both types of Ca(2+) channels, and preferentially reversed the response to 30 mM K(+) stimulation compared with 50 mM K(+) stimulation. In contrast, verapamil preferentially inhibited the response to depolarization by the higher concentration (50 mM) of K(+). 4 Gabapentin inhibited KCl-induced elevation of intracellular Ca(2+) in primary neuronal culture. 5 Hypoxic injury was induced in cerebrocortical slices by oxygen deprivation in the absence (severe injury) or presence of 3 mM glucose (mild injury). Gabapentin preferentially inhibited mild injury, while verapamil suppressed only severe injury. omega-Conotoxin GVIA (omega-CTX) and omega-agatoxin IVA (omega-Aga) were effective in both models. 6 NO synthesis was enhanced in a manner dependent on the severity of hypoxic insults. Gabapentin reversed the NO synthesis induced by mild insults, while verapamil inhibited that elicited by severe insults. omega-CTX and omega-Aga were effective in both the cases. 7 Therefore, the data suggest that gabapentin and verapamil cause activity-dependent Ca(2+) channel blocking by different mechanisms, which are associated with their cerebroprotective actions and are dependent on the severity of hypoxic insults.     

Protective effects of glutathione on 5-fluorouracil-induced myelosuppression in mice

The protective effects of glutathione (GSH) administration on myelosuppression induced by 5-fluorouracil (5-FU) were investigated in female BALB/c mice. Animals were allocated to four groups (16 mice/group). GSH was given orally at a dose of 800 mg/kg to groups 3 and 4 for 21 consecutive days (day 0 to day 20). 5-FU was repeatedly administered at a dose of 40 mg/kg to groups 2 and 3 for 1 week (day 7 to day 13) by gavage. Group 3 served as a combined treatment group and group 1 as a non-treated control group. The total observation period was 3 weeks. Body weight was measured once a week. A decrease in body weight due to 5-FU treatment was observed in groups 2 and 3 on day 14. Although the body weight in group 2 had not increased by 1-week after cessation of 5-FU treatment, the value in group 3 markedly recovered. Hematology, total nucleated myelocyte count and histopathology of bone marrow were carried out on day 14 and day 21. In groups 2 and 3, these examinations showed thrombocytopenia, leukopenia, reticulocytopenia and myelosuppression on day 14. However, platelets and bone marrow were less affected in group 3 than in group 2. On day 21, the thrombocytopenia in groups 2 and 3 was resolved. The myelosuppression, leukopenia and reticulocytopenia resolved in group 3, but not in group 2. Although simple microcytic anemia occurred delayed on day 21, it was less severe in group 3 than in group 2. Therefore, GSH may have preventive effects against 5-FU-induced hematopoietic toxicity, and accelerate recovery after cessation of 5-FU treatment.     

Inactivation and Aggregation of β-Galactosidase in Lyophilized Formulation Described by Kohlrausch-Williams-Watts Stretched Exponential Function

Purpose. To examine whether the empirical Kohlrausch-Williams-Watts (KWW) equation is applicable not only to protein aggregation but also to protein denaturation in lyophilized formulations. Lyophilized -galactosidase (-GA) formulations containing polyvinylalcohol and methylcellulose were used as model formulations. The possibility of predicting storage stability based on the temperature dependence of the estimated parameters of inactivation/aggregation—time constant () and its distribution () is discussed. Methods. Protein aggregation in lyophilized -GA formulations at 10-70°C and 6-43% relative humidity was determined as a function of time by size exclusion chromatography. Enzyme activity was also determined using 2-nitrophenyl--D-galactopyranoside as a substrate. Results. Inactivation and aggregation of -GA were describable with the empirical KWW equation, regardless of whether the temperature was above or below the NMR relaxation-based critical mobility temperature (T_mc) or whether protein molecules with different degrees of deformation resulting from stresses during lyophilization exist in the formulation. The estimated parameter for protein aggregation decreased rapidly as temperature increased beyond T_mc because the mobility of polymer molecules increased in the initial stages of glass transition. The time required for 10% enzyme to aggregate (t₉₀) calculated from the and parameters exhibited a change in temperature dependence gradient near T_mc. In contrast, t₉₀ for protein inactivation exhibited temperature dependence patterns varying with the excipients. Conclusions. The t₉₀ calculated from the estimated and parameters was found to be a useful parameter for evaluating the stability of lyophilized -GA formulations. The prediction of t₉₀ by extrapolation was possible in the temperature range in which did not rapidly vary with temperature.     

Effect of gelatins on human cancer cells in vitro

Three types of gelatins were tested for their antiproliferative activities in vitro against three human tumor cell lines (K-562; erythroleukemia, HCT-15; colon carcinoma, AGS; gastric carcinoma) with viable cell count and tritium-thymidine ((3)H-TdR) uptake by those cells. Porcine skin (PS) gelatin exerted the strongest antiproliferative activity of all three gelatins. Bovine bone (BB) gelatin did not exert such an activity. PS gelatin exerted antiproliferative activity against K-562 cells also in a serum-free medium. The serum-free medium contains two growth factors, insulin and transferrin, as well as nutrients. The activity of PS gelatin was not interfered by addition of insulin and transferrin to the medium. Effect of diluting a K-562 cell-concentration on the activity of PS gelatin was tested. Diluting the cell concentration did not affect the activity of PS gelatin. Moreover, the conditioned medium in which K-562 cells had been cultured did not stimulate the proliferation of K-562 cells. In conclusion, PS gelatin suppress the proliferation of human tumor cell lines in vitro. The antiproliferative activity of PS gelatin might not be attributed to trapping growth factors or autocrine mediators.     

Human papillomaviruses in the normal oral cavity of children in Japan

The purpose of this study was to determine the frequency of human papillomavirus (HPV) infections in the normal oral cavity of children in Japan. Oral squamous cell specimens were collected from 77 children (44 boys and 33 girls), aged 3 and 5 years. Extracted DNA was evaluated for HPV infections by polymerase chain reaction (PCR) methods, using consensus primers for the L1 region, specific primers, and direct DNA sequencing analysis. Thirty-seven of 77 specimens (48.1%) were positive for HPV DNA. Positive rates of boys and girls in all specimens were 28.3 (22/77) and 19.5 (15/77)%, respectively. The positive rate in 3-year-old children was 45.2 (14/31)%, and positive rates in boys and girls were 52.6 (10/19) and 33.3 (4/12)%, respectively. The positive rate in 5-year-old children was 50.0 (23/46)%, and positive rates in boys and girls were 48.0 (12/25) and 52.4 (11/21)%, respectively. HPV types were determined by specific PCR and direct DNA sequencing analysis. Frequent HPV types in the specimens of all children were HPV-16 (11/37; 29.7%),-1 (6/37; 16.2%),-2 (6/37; 16.2%),−75 (6/37; 16.2%). The results of the present investigation indicate that many HPVs, including HPV-16 (a high-risk type for cancer), are present in the oral cavity of 3- and 5-year-old children. It is suggested, therefore, that the oral cavity is already a reservoir of HPVs in childhood where later HPV-associated diseases, such as oral cancer and other oral lesions, may develop.     

Successful Laparoscopic Right Gastroepiploic Aneurysmectomy: Report of a Case

A 75-year-old woman presented with a pulsatile, movable mass, about 5cm in diameter, in her lower abdomen. Abdominal ultrasonography revealed a circular mass with a variable hypo- and isoechoic border and a hypoechoic center. Color Doppler echography showed blood flow in the hypoechoic center, which was strongly enhanced on contrast-enhanced computed tomography. Based on these findings, we diagnosed a splanchnic artery aneurysm; however, celiac arteriography, performed twice, could not definitively identify it. An operation was performed under the tentative diagnosis of an aneurysm of the superior mesenteric artery or the gastroepiploic artery. On laparoscopic exploration, a globe-shaped mass, about 5cm in diameter, was found in the right side of the greater omentum, which was diagnosed as an aneurysm of the right gastroepiploic artery. We resected the aneurysm laparoscopically and the patient had an uneventful postoperative course. Thus, laparoscopic surgery was effective for this patient who required no vascular reconstruction.     

An in vitro evaluation of the glycation potential of a natural disaccharide,trehalose

Background. Glucose, an osmotic agent generally used in continuous ambulatory peritoneal dialysis (CAPD) dialysate, has a critical characteristic of forming advanced glycation endproducts (AGEs). We undertook this study to investigate whether a possible osmotic agent, trehalose, formed fewer AGEs than glucose. Methods. Hemoglobin (Hb), a counter-protein of AGE, was incubated in four kinds of medium; glucose-phosphate buffered saline (PBS), autoclaved glucose-PBS, trehalose-PBS, and autoclaved trehalose-PBS, for 3, 7, 14, and 30 days, respectively. Polymerization of the Hb molecule was detected by sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and carboxymethylated Hb was detected by Western blotting, using specific mono-clonal antibody for carboxymethylated N-terminal valine-Hb (CMV-Hb). Results. PBS containing glucose showed bands of polymerized Hb molecule, a phenomenon which was markedly exaggerated by autoclaving. Likely, PBS containing glucose showed the formation of CMV-Hb in the long incubation of 30 days, and PBS containing autoclaved glucose showed accelerated formation of CMV-Hb in an incubation as short as 3 days. By contrast, PBS containing trehalose showed much less increase in a band of 30k Dalton and in CMV-Hb formation even in autoclaved medium. Conclusions. Our present in vitro study clearly showed the superior characteristic of trehalose to produce fewer AGEs. Based upon the results of this study, we propose that the application of trehalose should be considered for CAPD solution.     

Relationship between attention and arousal level in schizophrenia

The purpose of the present paper was to clarify the link between the attention and arousal level that supports the basis of the cognitive dysfunction in schizophrenia, by investigating the relationship between the simple reaction time and the closed-eye eye movements in 30 patients with schizophrenia and 20 healthy controls. In terms of closed-eye eye movements during the simple reaction time test, healthy controls showed an increase of s-type (small and slow) eye movements after the end of the preparatory interval (PI) in both regular and irregular series, while the patients with schizophrenia, particularly those in whom the cross-over phenomenon was observed, showed no changes and maintained a hyperarousal level during the regular PI test. These results indicate that the patients with schizophrenia could not maintain appropriate attention during the burden tasks and their hyperarousal level persisted. It is therefore suggested that there is a close relationship between attentional deficit and hyperarousal among patients with schizophrenia.     

Three components of obstructive sleep apnea/hypopnea syndrome

The aims of this study were to calculate the apnea-hypopnea index (AHI), which represented as the number of apnea-hypopnea occurrences per hour, the 4% oxygen desaturation index (ODI4) and the breathing-related arousal index (B-ArI) in polysomnographic studies of obstructive sleep apnea/hypopnea syndrome (OSAHS) patients and to investigate whether there was any relationship between each pair of scoring schemes. Thirty-four cases of OSAHS were studied. Total OSAHS patients were subdivided into those with a high AHI (> 25), and those with a low AHI (< 25). The correlation between each pair of scoring schemes for OSAHS with a high AHI showed high value. The correlation between AHI and ODI4 for OSAHS with a low AHI was 0.18 and that between AHI and B-ArI showed a weak correlation of 0.59, while that between ODI4 and B-ArI was only -0.078. Our results mean that oxygen desaturation and arousal occur separately in mild or moderate OSAHS patients, even though they are diagnosed with the same level of OSAHS by means of AHI. Breathing-related arousal without oxygen desaturation often occurs in mild or moderate OSAHS patients. We previously reported that AHI does not accurately reflect the severity of the increase in negativity of esophageal pressure manifested as respiratory efforts. We consider that the comprehension and assessment of OSAHS can be improved by the systematic differentiations among the three components: oxygen desaturation, arousals and respiratory efforts.