Shiro Fujihira

Comparison of the course of biomarker changes and kidney injury in a rat model of drug-induced acute kidney injury

Objective: To aid in evaluating the performance of biomarkers, we measured kidney injury biomarkers in rat models of drug-induced acute kidney injury. Methods and results: Rats were treated with site-specific nephrotoxins, puromycin, gentamicin, cisplatin, or 2-bromoethylamine. Fifteen biomarkers (β-2-microglobulin, calbindin, clusterin, cystatin-C, KIM-1, GST-α, GST-μ, NGAL, osteopontin, EGF, TIMP-1, VEGF, albumin, RPA-1, and urinary total protein) were examined in comparison with BUN, serum creatinine, and NAG. Some biomarkers, which were different depending in each nephrotoxin, showed ability to detect the prodromal stage of drug-induced kidney injury. Characteristic changing patterns of biomarkers were also found depending on the specific lesion site in the kidney. Conclusion: These data suggested that establishment of a suitable biomarker panel would facilitate detection of site-specific kidney injury with high sensitivity.

ULTRASTRUCTURAL STUDY ON CYTOTOXIC EFFECTS OF CYCLOSPORINE A IN SPERMIOGENESIS IN RATS

Cyclosporine A (CsA) is known to have testicular toxicity, leading to male infertility. The occurrence of numerous anomalous spermatids and residual bodies in the epididymal ducts of rats treated with CsA was observed in our previous studies. To examine the fine structural changes of impaired spermiogenesis induced by CsA, rats were treated s.c. with 40 mg/kg/day CsA for 2 weeks. Desquamation of round spermatids still surrounded by a slender Sertoli cell cytoplasm was occasionally observed. A prominent change in the seminiferous tubules was an occurrence of numerous residual bodies containing one or more flagella. They were not phagocytosed by the Sertoli cell, and accumulated in the epididymal ducts. Cellular components in the epididymal lumen included degenerating round spermatids and abnormal spermatozoa and residual bodies. Although separation of the head from the tail of the spermatozoa was rarely seen, various kinds of abnormalities of flagella were frequently encountered; compact aggregation of numerous flagella in a single spermatozoon or residual body, disarrangement of mitochondrial or fibrous sheath with outer dense fibers, and fusion of flagella with a single intervening mitochondrial layer. These findings indicate that CsA gives rise to toxic effects on the spermiogenesis by impairing directly the spermiogenic cell development and by impeding Sertoli cell function including a reduction of its phagocytic activity.

Effect of Prograf (FK506) on spermatogenesis in rats

Prograf (FK506) was given to male mature rats in a daily subcutaneous dose of 1 or 3 mg/kg/day for 2 weeks to investigate its effect on spermatogenesis. Prograf dose-dependently sperm counts and motility, but did not affect testosterone level in the serum of rats. Histopathologically, there were no abnormal changes in the testis, seminal vesicle or prostate in any rats dosed with Prograf, but intra-ductal eosinophilic globules, probably degeneration of the sperm cells, were observed in the epididymis of the 3 mg/kg/day group. Sperm counts and motility returned to the control levels after stopping of the drug. The results indicate that Prograf decreased sperm counts and motility through direct action on the sperm in the epididymis, but not the production of sperm in the testis. Cyclosporine A (CsA) was used as the reference drug in the present study. Thirty mg/kg/day of CsA also decreased sperm counts and motility, and stopping of the drug led to the recovery of these changes. The males dosed with Prograf for 2 weeks were mated with non-dosed females to investigate its effect on the fertility potential of the males. Prograf did not affect copulation or fertility index, but a decrease in the number of live fetuses associated with implantation loss was observed in the 3 mg/kg/day group. The changes were considered to be due to the decrease of sperm counts and motility, although 1 mg/kg/day of Prograf, 5-10 times the clinical dose, did not affect any fertility parameters including implantation index.

The High Incidence of Atrial Thrombosis in Mice Given Doxorubicin

Doxorubicin (DX)-treated mice represent an animal model for studying new drugs for heart disease. Coincidentally, in the collection of damaged myocardial tissue, thrombosis was detected in the atrium. The incidence reached 75% in mice given 4 mg/kg DX iv 10 times. They were white thrombi consisting of the fibrin, platelets, and neutrophils. Cardiac muscle damage was more prominent in the atria than in the ventricles. Light microscopically, vacuolization and degeneration of atrial myocytes and interstitial inflammatory cell infiltration were observed. Electron microscopy revealed dilatation of the sarcoplasmic reticulum and an increase in number of normal and/or degenerate mitochondria. Inflammation extended from the cardiac muscle to the endocardium. The cause of atrial thrombosis in DX-treated mice is unknown but may relate to endocardial damage and changes of blood flow in the atrium secondary to cardiac muscle damage. DX-treated mice could serve as an experimental animal model for the evaluation of efficacy and toxicity of antithrombotic or antiplatelet drugs.

Abdominal cysticercosis in a cynomolgus monkey.

Cysticercus tenuicollis, the larval form of Taenia hydatigena, was observed in a 5-year-old male cynomolgus monkey used in a toxicity study for a safety assessment of a pharmaceutical. The animal was born and raised in a primate colony in China. A pale yellow cyst filled with more than 100ml of pale yellow fluid was found in the abdominal cavity in the autopsy. The cyst was found attached to the greater omentum, and it was double layered. Histopathologically, the outer layer was a part of the greater omentum, and the inner layer was the bladder wall of a cysticercus with a well developed scolex. A partial sequence of mitochondrial NADH dehydrogenase subunit 1 showed a high homology to the same region of Taenia hydatigena (1.5-3.3%).

Restoration of Leydig cells after repeated administration of ethane dimethanesulfonate in adult rats

Adult male rats were repeatedly treated with ethane dimethanesutfonate (EDS), an agent known to destroy Leydlg cells selectively. Following a second Injection, changes In serum testostemna levels and histological and morphametric changes of Leydlg cells showed the time course to be similar to those after the flrst treatment. The number and volume of Leydlg cells markedly decreased at day 2, began to increase from day 7, and recovered to the values of the contml rats at day 30, concomitant with the changes of serum testosterone levels. Cells in the interstitial tlssue labeled wlth bromodeoxyuridlne markedly increased In number at day 2, gradually decreased thereafter, and returned to the values of the controls at day 14. During this period, cells undergoing mltosls were seen, their type unable to be determined, but were presumed to be regenerating Leydlg cells. Even 30 days following four treatments with Intervals of 30 days each, serum testosterone levels were the same as those in the controls. Also the numerlcal and volume denstties of Leydig cells and the volume of an average Leydlg cell were the same as those of the controls. Mttosis was observed In mature Leydig cells at this perlod, if any. It appears that new Leydig cells began to proliferate by division earller than 14 days after EDS, allowing that there were several stages of proliferation, and that the source of reappearing Leydig cells may not be a limited number of precursor cells, impiying the presence of stem cells for Leydig cells.

Goblet Cell Hyperplasia and Muscular Layer Thickening in the Small Intestine of a Cynomolgus Monkey

We report here the interesting case of a 5-year-old male cynomolgus monkey with goblet cell hyperplasia and thickening of the muscular layer throughout the small intestine without exhibiting any clinical symptoms. Necropsy examination showed diffuse thickening of the intestinal wall from the jejunum to the ileum, with an appearance likened to a rubber tube. Histopathologically, marked thickening was observed in both the mucosal and muscular layers in the jejunum and ileum, and slight thickening was observed in the duodenum. Goblet cell hyperplasia with extension of the circular folds and villi was prominently observed. The mucosal surface was covered with a thick mucus layer containing desquamated mucosal epithelial cells, and both the inner and outer muscular layers were markedly thickened due to smooth muscle hypertrophy. Neither macroscopic nor histopathological examination identified any causative factors, such as infection, enteritis and intestinal stenosis, or obstruction that may have caused development of this lesion. Given these observations, this case may simply be considered of spontaneous goblet cell hyperplasia and muscular layer thickening in the small intestine of a cynomolgus monkey.