Hiroto Miyata

Full-Field Electroretinography Obtained Using a Contact Lens Electrode with Built-In High-Intensity White-Light-Emitting Diodes Can Be Utilized in Toxicological Assessments in Rats

Full-field electroretinography (ERG) using contact lenses with built-in LED was performed on albino rats, and used to evaluate the visual toxicity of sodium iodate (NaIO3). Experiment 1 was carried out to determine the optimal conditions in rats relating to stimulus intensity, background illumination, and light adaptation period. As a result, we found that a full-field ERG was recorded under the following conditions: stimulus intensity: –3.5 log cd s/m2 in rod response; background intensity and light adaptation period: 10 cd/m2 and 10 min in cone and flicker responses. Experiment 2 was carried out to confirm the usefulness of full-field ERG using rats with retinal toxicities induced by NaIO3. Male rats were given NaIO3 intravenously at a dose of 50 mg/kg. ERG was recorded before administration and after 3, 8, 24 h, and 7 days of administration, and histopathological analysis was conducted after 8 h of administration. The rod response disappeared completely at 3 h, based on a reduced maximal response and oscillatory potentials. On the other hand, cone and flicker responses were still present at 8 h. All responses disappeared on the 7th day. These findings indicate that the retinal toxicity induced by NaIO3 was expressed first in rods, followed by cones. There were no microscopical changes after 8 h of administration, although the rod responses had completely disappeared by this time. These results suggest that full-field ERG in rats using an LED contact lens is useful for the separate evaluation of toxic effects on rods and cones.

Evaluation of Bone Toxicity in Various Bones of Aged Rats

The aim of the present study was to provide a method for evaluating bone toxicity induced by drugs in various bones in aged rats. Male Crl:CD (SD) rats at 46 weeks of age were administered 15 mg/m2 body surface area of doxorubicin, which effects the growth plate in weanling rats, weekly for 9 weeks by intravenous injection, and the femur, sternum, humerus and tibia were examined histopathologically. In the doxorubicin-treated group, thinning of the growth plate was remarkably observed in the proximal tibia and humerus; however, these changes were not observed in other regions. In addition, the osteoclast number per bone perimeter in the proximal tibia was significantly higher than others in control aged rat. Thus, recognizing the various histological reactions related to the time of epiphyseal closure is important for evaluating bone toxicity in aged rats.

Evaluation of Myelotoxicity in Dietary Restricted Rats

The purpose of this study was to clarify the effect of decreased food consumption on evaluation of myelotoxicity in routine general toxicity studies. Male rats were divided into the following 7 groups: 12, 15, and 18 mg/kg 5-fluorouracil (5-FU) treatment groups (FU12, FU15 and FU18); dietary restriction groups (R12, R15 and R18 receiving the same amount of food as the rats in the FU12, FU15 and FU18 groups, respectively); and a nontreated control group (NT). We compared the changes in body weight, hematology and the results of cytological analyses of bone marrow and histopathology among the groups after administration and recovery periods of 14 and 7 days, respectively. At the end of the administration period, the FU15 and FU18 groups showed decreases in many hematologic and bone marrow parameters that were all similar to those in the corresponding dietary restriction groups (R15 and R18). A granulocyte abnormality (polyploidy: frequency of 1% or less) was also observed in all 5-FU treated groups. At the end of the recovery period, increases in the reticulocyte and platelet counts and extramedullary hematopoiesis of the spleen were observed in the 5-FU treated groups. These results indicate that the results of general toxicity studies in rats should be evaluated in consideration of dietary restriction effects when food consumption is decreased at about 30-40% or more. Careful morphological observation of hemocytes would be helpful in distinguishing the effect of a drug from that of dietary restriction in relation to hematological and bone marrow parameters. Performance of a recovery test to determine the reactive response of hematopoiesis is also recommended.

Importance of Starting Age for Myelotoxicity Study in Dietary Restricted Rats

The aim of this study was to prove our hypothesis that adult rats with lowering of body weight gain, rats at 12 weeks of age as an example, are suitable for evaluation of myelotoxicity. Age-related differences between young rats (6-week-old study) and adult rats (12-week-old study) were analyzed in hematological examination values. The data of the young rats were reprinted from our previous report (Miyata et al., 2009) since our hypothesis was verified by comparison with that previous report. Several experimental groups were defined for the 12-week-old study as well as for the 6-week-old study; these included 5-fluorouracil (5-FU) treated groups receiving 12, 15 and 18 mg/kg/day (FU12, FU15 and FU18), pair-feeding groups (R12, R15 and R18 receiving the same amount of food as in the FU12, FU15 and FU18 groups, respectively) and a nontreated control group. Numerous hematologic and bone marrow parameters in the 5-FU treated groups were comparable to those in the corresponding pair-feeding groups in both age studies. Generally, the influences of undernutrition were more apparent in the young rats than in the adult rats. Histopathological examinations showed a decrease in hematopoiesis in the bone marrow in the 5-FU treated and pair-feeding groups. No apparent differences were observed in the decreased hematopoiesis between the 5-FU treated and pair-feeding groups in the 6-week-old study, but a difference between these groups was noted in the 12-week-old study; decreased hematopoiesis was more frequently noted in the 5-FU treated groups. These facts suggest that adult rats are more suitable than young rats for evaluation of 5-FU-induced myelotoxicity.

Evaluation of Short-term Myelotoxicity Study in Dietary Reduced Rats

This study attempted to prove our hypothesis that a short-term toxicity study, using a 4-day dosing regimen as an example, is suitable for evaluating myelotoxicity in rats. We compared the hematological, bone marrow cytological and histopathological results of 5-fluorouracil (5-FU) treated and pair-feeding groups after a 4-day administration period. Several experimental groups were defined for this 4-day study as well as for our previously reported 14-day study (Miyata et al., 2009); these included 5-FU treated groups receiving 12, 15 and 18 mg/kg/day (FU12, FU15 and FU18), pair-feeding groups (R12, R15 and R18 receiving the same amount of food as the FU12, FU15 and FU18 groups, respectively) and a nontreated control group. Although severe reductions in body weight gain and food consumption were reported in the 14-day study, only slight reductions were observed in the 4-day study. In the 4-day study, a decrease in blood reticulocytes and a decreasing trend of marrow erythroid cells were only observed in the FU18 group, and no effects were observed in the pair-feeding groups. The erythroblastic changes observed in this 4-day study were thought to reflect the direct influence of 5-FU administration. Since concerns regarding the influence of secondary changes related to undernutrition were minimized in the 4-day study, it was thought to clarify the direct influence of 5-FU administration on erythroblastic cells. Thus, a 4-day study protocol might be helpful for distinguishing secondary changes related to undernutrition.