Sequential analysis of testicular lesions and serum hormone levels in rats treated with a Psoralea corylifolia extract.

In order to clarify pathogenetic targets for the testicular toxicity of a extract of Psoralea corylifolia (P. corylifolia), F344 rats were fed diet containing 3% P. corylifolia extract for up to 12 weeks and subjected to hormone assays and histopathological examination on the testis and epididymis at weeks 1, 2, 4, 8 and 12 (Exp 1). Similar analyses were performed on 1, 3, 7 and 14 days after a single gavage administration of the P. corylifolia extract at a dose of 10 g/kg b.w. (Exp 2). In Exp 1, increase in the numbers of degenerated and exfoliated germ cells and loss of elongated spermatids beyond steps 7 or 8 were initially observed in the seminiferous tubules at week 1, followed by more pronounced degeneration of germ cells with depletion of post-meiotic populations from week 2. The tubular degeneration was associated with Leydig cell atrophy and persistent reduction of serum testosterone and FSH levels throughout the treatment period and a slight reduction of serum LH in later stages. In Exp 2, reduction of serum testosterone and FSH levels preceded degeneration of germ cells in stage VII and VIII tubules at 3 and 7 days after the administration. The results suggest that rapid androgen deprivation reflecting direct interference with Leydig cell function and simultaneous disturbance of the pituitary-testicular axis play pivotal roles in P. corylifolia extract-induced germ cell injury in seminiferous tubules.

Anticlastogenic activity of cacao: inhibitory effect of cacao liquor polyphenols against mitomycin C-induced DNA damage.

Oxidative DNA damage has been implicated as a factor playing a role in mutagenesis and carcinogenesis. We investigated the anticlastogenic activity of cacao: the inhibitory effect of cacao liquor polyphenols on DNA strand cleavage induced by mitomycin C (MMC) in vitro and the anticlastogenic effect of cacao liquor extract against formation of micronuclei induced by MMC in bone marrow cells and peripheral blood cells of mice. In the DNA strand cleavage test, cacao liquor polyphenols inhibited cleavage of RFI DNA. In the micronuclei test, the frequency of occurrence of micronucleated cells among bone marrow cells and peripheral blood cells were reduced significantly when cacao liquor extract was administered orally to mice 6 h before intraperitoneal injection of MMC. These findings suggest that cacao liquor polyphenols are effective in preventing DNA damage, and one of the mechanisms of action might involve scavenging of active oxygen radicals generated in reactions initiated by MMC.

Cacao liquor polyphenols reduce oxidative stress without maintaining alpha-tocopherol levels in rats fed a vitamin E-deficient diet.

The effect of crude polyphenols (CLP) from cacao liquor on vitamin E-deficient rats was examined. The CLP fraction contained 49.8% antioxidative polyphenols such as catechins and their oligomers. Supplementation of the vitamin E-deficient diet with CLP for 7 wk did not prevent the decrease in alpha-tocopherol levels in the liver, kidney, heart, brain, and plasma. The lipid peroxide levels in these tissues increased in the group fed the vitamin E-deficient diet compared with the control group. However, these changes were inhibited in a dose-dependent manner as a result of supplementation of the vitamin E-deficient diet with 0.25, 0.5, or 1.0% CLP. The lipid peroxide levels in plasma increased in the group fed the vitamin E-deficient diet. This change tended to be suppressed as a result of supplementation of the diet with CLP, but the difference was not significant. There was no evidence of absorption and distribution of CLP to the tissues; however, CLP intake resulted in a decrease in oxidative stress without maintaining vitamin E levels in the plasma and the tissues.

[A 90-day repeated dose oral toxicity study of magnesium chloride in F344 rats].

In order to examine the toxicity of magnesium chloride hexahydrate, four groups of 10 male and 10 female F344 rats received the compound by dietary supplementation at 2.5, 0.5, 0.1 or 0% for 90 days. No treatment-related death was observed during the study. Transient soft stool and sustained increase in water consumption were observed both in males and females of the 2.5% group and slight reduction in body weight gain was noted in the high-dose males. There were no toxic changes in food consumption, organ weights, hematology and biochemistry, and histopathological examinations in any treated-groups. Based on these results, the no-observed-adverse-effect-level was estimated to be 0.5%, and 2.5% is considered to be appropriate as highest dose for a 2-year carcinogenicity study.

In vivo absorption of calcium carbonate and magnesium oxide from the large intestine in rats.

This study was conducted to evaluate the ability of the large intestine to absorb calcium (Ca) and magnesium (Mg) from their sparingly water-soluble salts, and also to determine whether fructooligosaccharides (FOS) stimulate the absorption of these minerals in rat large intestine in vivo. Rats were fed Ca- and Mg-free diets with and without 5% FOS. An aqueous suspension of CaCO3 and MgO was infused into the stomach via a gastric tube or into the cecum via an implanted catheter. Coprophagy was prevented by using wire-mesh anal cups throughout the experiment so as to exclude the re-ingestion of feces as an oral mineral source. In rats fed an FOS-free diet, the absorption degrees of Ca and Mg infused into the cecum were the same as those infused into the stomach. The absorption degree of phosphorus (P) was slightly but significantly higher in rats with the infusion of Ca and Mg into the cecum than in rats with the infusion of Ca and Mg into the stomach. FOS-feeding increased the absorption of Mg to a similar extent in either case of infusion via cecal and oral routes, while FOS-feeding did not increase the absorption of Ca in rats with infusion of Ca and Mg into the cecum. We concluded that both CaCO3 and MgO are absorbed in the large intestine, and we ascertained that the increasing effect of FOS on the absorption of Mg took place mainly in the large intestine.

Quantitative RT-PCR for inhibin/activin subunits: measurements of rat hypothalamic and ovarian inhibin/activin subunit mRNAs during the estrous cycle.

Inhibins (alpha-beta(A) and alpha-beta(B)) and activins (beta(A)-beta(A), beta(A)-beta(B) and beta(B)-beta(B)) were originally isolated from ovarian follicular fluids as FSH secretion modifiers. Inhibin/activin subunits, alpha, beta(A) and beta(B), are widely distributed in several tissues, including gonads and brain, and inhibins and activins have been reported to be involved in ovarian or hypothalamic functions. In this study, we established and employed a competitive RT-PCR assay system for rat inhibin/activin subunits by capillary electrophoresis to determine rat hypothalamic and ovarian inhibin/activin subunit mRNA levels during the estrous cycle. Linearity of standards for alpha, beta(A), and beta(B) subunit assays were between 0.01-0.3 amol, 0.003-0.09 amol and 0.002-0.02 amol of each fragment DNA as a standard, respectively. Hypothalamic beta(A) subunit mRNA during the estrous morning (1000 h) tended to be increased compared with that of the proestrous evening (1700 h), although they were not significantly different. Ovarian alpha subunit mRNA levels tended to be increased during the proestrous morning (1000 h) and were significantly increased in the proestrous evening (1700 h), compared with diestrus and estrus (P < 0.05). Ovarian beta(A) subunit mRNA was also significantly higher in the proestrous evening, compared with diestrus and estrus (P < 0.05), but in the case of beta(B) subunit mRNA there was no difference among diestrus, proestrus and estrus. We thus established a sensitive competitive RT-PCR system for the measurement of inhibin/activin alpha, beta(A) and beta(B) subunits, and this assay system would be helpful for the study of inhibin/activin action in brain and other tissues where these factors are expressed at low levels.

Effects of fructooligosaccharides on the absorption of magnesium in the magnesium-deficient rat model.

Magnesium (Mg) is an essential dietary element that plays important roles, acting as a cofactor of many enzymes. Rats fed a Mg-deficient diet have been reported to exhibit auricular and facial peripheral hyperemia and hemorrhage. Moreover, increased intake of calcium (Ca) or phosphorus (P) has been reported to impair apparent absorption of Mg. We tried to induce such typical inflammation in Mg-deficient rats by feeding low-Mg, high-Ca, and high-P diets. Increasing concentrations of Ca or P in the experimental diets significantly decreased the apparent absorption of Mg. And all rats fed the low-Mg (0.25 mg/g diet), high-Ca (10.4 mg/g diet), and high-P (12.0 mg/g diet) diet exhibited auricular and facial peripheral-hyperemia and hemorrhage. Then, we used the low-Mg, high-Ca, and high-P diet to investigate the effects of the fructooligosaccharides (FO) on absorption of Mg and skin inflammation. In the rats fed FO-containing (1 or 5%) diet, apparent absorption of Mg was significantly increased as compared with that of the control (FO 0%) group. In the rats fed a 5% FO-containing diet and sufficient Mg (0.50 mg/g), auricular and facial peripheral hyperemia and hemorrhage were significantly reduced. We concluded that FO increased the Mg absorption in rats fed a low-Mg, high-Ca, and high-P diet. Moreover, FO reduced inflammation in Mg-deficient rats, such as peripheral hyperemia and hemorrhage.

[A case of transient vertical gaze palsy following right thalamic and midbrain infarction].

The authors report a 71-year-old male who suffered from vertical gaze palsy due to infarction localized on MRI in the right midbrain. Vertical gaze palsy was observed 1 hour after the onset of his stroke. Upgaze palsy was noted the following day. Four days later, the disturbances of ocular movement disappeared. T2 weighted MR imaging showed a high signal intensity lesion, which was localized in the right side of the thalamo-mesencephalic portion. However, since the vertical gaze palsy appeared only transiently for a short time in the initial stage and since the level of consciousness decreased during that time, it was suggested that there was a bilateral functional disorder caused by mild edema, etc., in the hyperacute stage, which was not demonstrated on MRI. This disorder had been overlooked either because the duration of the vertical gaze palsy due to the unilateral lesion was very short, i.e. improvement was seen 4 days after onset, or the lesion was not clearly delineated in a CT scan of the head. It appeared possible that this sign might not be very rare if careful observations are performed in the hyperacute stage of midbrain infarction.

Administration of alpha 1-proteinase inhibitor ameliorates bleomycin-induced pulmonary fibrosis in hamsters.

The effect of alpha 1-proteinase inhibitor (alpha 1Pi) administration on the acute lung injury and subsequent fibrosis induced by bleomycin (BLM) was examined in hamsters. Pulmonary lesions were quantitatively reduced in alpha 1Pi-administered BLM-treated (BLM-alpha 1Pi) animals compared with animals treated by BLM alone (BLM-control) at both 7 days (acute stage) and 30 days (fibrotic stage) after BLM treatment. Analysis of intraalveolar cells from bronchoalveolar lavage (BAL) fluid revealed that neutrophils and lymphocytes were significantly decreased in the BLM-alpha 1Pi animals at 7 days after BLM treatment and that 30 days after BLM treatment macrophages as well as neutrophils and lymphocytes were remarkably decreased in the BLM-alpha 1Pi animals. The elastase activity in supernatants of BAL fluid during 7 days following BLM treatment was detected, but there was no difference between the two groups. In vitro studies on neutrophil responsiveness to stimulation of BAL fluid at 3 days after BLM treatment revealed noticeable chemotaxis and generation of superoxide anion of isolated neutrophils, but alpha 1Pi did not show any inhibitory effects on neutrophil responsiveness. We suggest that alpha 1Pi administration ameliorates pulmonary fibrosis preceded by acute lung injury induced by BLM treatment in hamsters and that the inhibitory effects of alpha 1Pi on lung injury may not be brought about by altered elastase activity, chemotaxis, or superoxide generation in neutrophils. Alternative mechanisms are discussed.

[Effect of saibokuto on mucociliary transport system in the airway--basic and clinical assessments].

We studied the effects of Saibokuto, an antiasthmatic agent, on the airway mucociliary transport system. The addition of Saibokuto to a Rose chamber containing rabbit cultured tracheal epithelium dose-dependently increased ciliary beat frequency (CBF) as assessed by a photoelectric method: the maximal increase from the baseline value was 31.5 +/- 5.2% (p less than 0.001) and the concentration of Saibokuto required to produce a half-maximal effect (EC50) was 10(-4) mg/ml. This effect was not affected by the beta-adrenergic receptor antagonist propranolol or Ca(2+)-free medium, but was inhibited by the cyclooxygenase inhibitor indomethacin. Intracellular cyclic AMP levels were increased from 45.6 +/- 8.2 to 72.1 +/- 12.6 pmole/mg protein by 1.0 mg/ml Saibokuto (p less than 0.05). Thus, Saibokuto enhances ciliary motility probably through the synthesis of cyclooxygenase products and cyclic AMP. Additionally, administration of this drug to patients with chronic bronchitis, asthma and bronchiectasis improved their problems with sputum expectoration and reduced the amount of daily sputum (p less than 0.001). Therefore, Saibokuto may be of value in the treatment of patients with impaired mucociliary transport function in the airway.

Stimulation of Na absorption by the antiasthmatic kampo drug Saiboku-to in cultured airway epithelium.

To study the effect of the Kampo drug Saiboku-to (TJ-96) on ion transport function of airway epithelial cells, we studied bioelectric properties of cultured tracheal epithelium from dogs under short-circuit conditions in vitro. Addition of TJ-96 (1 mg/ml) to the mucosal solution of the Ussing chamber increased the epithelial short-circuit current (SCC) from 6.5 +/- 0.7 to 11.4 +/- 1.6 microA/cm2 (P less than 0.001). This effect was dose-dependent, with the maximal increase from the baseline value and the concentration required to produce a half-maximal effect (EC50) being 70.5 +/- 12.6% (P less than 0.001) and 3 micrograms/ml, respectively; and there were corresponding increases in transepithelial potential difference and cell conductance. Submucosal addition of TJ-96 likewise increased SCC, although the magnitude of the response was smaller as compared with the response to the mucosal addition. The TJ-96-induced increase in SCC was not affected by diphenylamine-2-carboxylate or furosemide but abolished by amiloride. Intracellular cyclic AMP levels were dose-dependently increased by TJ-96. These results indicate that TJ-96 may selectively stimulate Na absorption across the tracheal epithelium, probably through intracellular accumulation of cyclic AMP.

Immunopharmacological studies of the aqueous extract of Cinnamomum cassia (CCAq). II. Effect of CCAq on experimental glomerulonephritis.

Effect of the aqueous extract of Cinnamomum Cassia (CCAq) on experimental glomerulonephritis was studied and compared with that of cobra venom factor (CoVF). In rat nephrotoxic serum (NTS) nephritis, CCAq clearly inhibited the excretion of protein into the urine and the increase of peripheral leucocyte counts. The histological score in CCAq administered animals was significantly lower than that in control animals. However, CCAq did not inhibit or lower the serum complement level. Contrary to CCAq, hypocomplementation was observed by the administration of CoVF, and the excretion of protein into urine was inhibited in a high dose group. In immune complex (IC) and autologous IC nephritis in rats, CCAq clearly inhibited the excretion of protein into urine and the elevation of blood urea nitrogen (BUN). The administration of CoVF caused hypocomplementation, but little inhibition of the excretion of urinary protein was observed in both types of immune complex nephritis. The histological score was slightly inhibited by a low dose of CCAq and a high dose of CoVF. In the experiment employing NZB/NZW F1 mice, the proteinurea, the elevation of BUN level, and the production of antibodies were clearly inhibited by the administration of CCAq. Similar inhibition was observed by CoVF at a high dose. However, the histological changes of the kidney in NZB/NZW F1 mice were not prevented by the administration of CCAq or CoVF.