Spontaneous histiocytic sarcoma originating from the epididymis in a CD-1 mouse.

We report a histiocytic sarcoma originating from the epididymis observed in a 110-week-old male CD-1 mouse in a carcinogenicity study. At necropsy, no lesions were observed in the epididymis. Histologically, a neoplastic lesion was observed in the cauda of the epididymis that was well demarcated from the surrounding tissues. The lesion mainly consisted of spindle-shaped tumor cells with oval to elongated nuclei and abundant eosinophilic or foamy cytoplasm. The tumor cells were arranged in a fascicular pattern, interlacing bundles, or a whorl pattern. The nuclei showed mild atypia with irregular shapes and varied sizes, whereas few mitotic figures and no typical multinucleated cells were observed. The epididymal ducts remained within the neoplastic lesion, and the tumor cells invaded between the epithelium and the smooth muscle layer of the epididymal duct. Immunohistochemically, the tumor cells were positive for vimentin and macrophage markers (Iba1, CD204, F4/80, and Mac-2) but negative for cytokeratin and other mesenchymal cell (α-smooth muscle actin, desmin, CD31, and platelet-derived growth factor receptor-ß), neural cell (S-100 and nestin), or Leydig cell markers (calretinin). Proliferating cell nuclear antigen-positive tumor cells were sporadically observed in the lesion. Based on these results, the tumor was diagnosed as a histiocytic sarcoma originating from the epididymis. This report provides additional histopathological evidence of spontaneous histiocytic sarcomas originating from the epididymis of aged mice.

Evaluation of the Carcinogenic Potential of Enarodustat (JTZ-951), a Hypoxia-Inducible Factor-Prolyl Hydroxylase Inhibitor, in 26-Week Tg.rasH2 Mouse Study and 2-Year Sprague-Dawley Rat Study.

Enarodustat (JTZ-951) is an oral hypoxia-inducible factor-prolyl hydroxylase (HIF-PH) inhibitor for the treatment of anemia with chronic kidney disease. Carcinogenicity of enarodustat was evaluated in a 26-week repeated oral dose study in Transgenic rasH2 (Tg.rasH2) mice and a 2-year repeated oral dose study in Sprague-Dawley (SD) rats. The highest dose levels were set at 6 mg/kg in the Tg.rasH2 mouse study and at 1 mg/kg in the SD rat study based on the maximum tolerated doses in the 3-month and 6-month dose-range finding studies, respectively. Enarodustat did not increase the incidence of any tumors or affect survival in these carcinogenicity studies. Pharmacology-related findings including increases in blood RBC parameters were observed at the highest dose levels for each study. The AUC-based exposure margins as protein-unbound drug base are 16.3-/26.0-fold multiple (males/females) for Tg.rasH2 mice and 1.6-/1.1-fold multiple for SD rats when compared with the estimated exposure in human with chronic kidney disease at 8 mg/day (maximum recommended human dose). In conclusion, enarodustat was considered to have no carcinogenic potential at the clinical dose.

Novel Biomarker Establishment for Evaluation of Excessive Fructose Consumption Using a Rat Model.

BACKGROUND/AIM: The habitual consumption of excessive fructose is associated with the onset and progression of lifestyle-related diseases, such as nonalcoholic fatty liver disease (NAFLD). In this study, we investigated the physiological changes observed when consuming a diet containing excessive fructose on the viewpoints of hepatotoxicity biological markers using a rat model and explored the biomarker candidates that could detect the effects of excessive fructose intake at an early stage. MATERIALS AND METHODS: Male rats were fed 63% high fructose diet (HFrD) ad libitum and their blood samples were collected before and at 1, 2, 3, and 4 weeks after allocation. The plasma biochemical parameters, hepatotoxic enzyme activities including alkaline phosphatase (ALP) isozymes were analyzed. RESULTS: HFrD consumption for 4-weeks created NAFLD-like symptoms, including elevated plasma lipid parameters and hepatotoxicity markers, as well as fat accumulation in the liver compared with rats consuming a control diet. Alanine aminotransferase (ALT) and glutamate dehydrogenase (GLDH) were increased from the 3rd and 2nd weeks, respectively, but no changes were observed on ALP activity. However, the daily consumption of the HFrD increased the plasma activities of liver-type ALP isozyme, and decreased plasma small intestinal-type ALP isozyme soon after the start of feeding. CONCLUSION: ALP isozyme analysis in combination with GLDH and ALT activities in the plasma samples could be a useful tool to detect the physiological changes induced by excessive fructose intake at an early stage of the development of NAFLD.

Enhancement of acetaminophen-induced chronic hepatotoxicity in spontaneously diabetic torii (SDT) rats.

Some patients encounter hepatotoxicity after repeated acetaminophen (APAP) dosing even at therapeutic doses. In the present study, we focused on the diabetic state as one of the suggested risk factors of drug-induced liver injury in humans and investigated the contribution of accelerated gluconeogenesis to the susceptibility to APAP-induced hepatotoxicity using an animal model of type 2 diabetes patients. Sprague-Dawley (SD) rats and spontaneously diabetic torii (SDT) rats were each given APAP at 0 mg/kg, 300 and 500 mg/kg for 35 days by oral gavage. Plasma and urinary glutathione-related metabolites, liver function parameters, and hepatic glutathione levels were compared between the non-APAP-treated SDT and SD rats and between the APAP-treated SDT and SD rats. Hepatic function parameters were not increased at either dose level in the APAP-treated SD rats, but were increased at both dose levels in the APAP-treated SDT rats. Increases in hepatic glutathione levels attributable to the treatment of APAP were noted only in the APAP-treated SD rats. There were differences in the profiles of plasma and urinary glutathione-related metabolites between the non-APAP-treated SD and SDT rats and the plasma/urinary endogenous metabolite profile after treatment with APAP in the SDT rats indicated that hepatic glutathione synthesis was decreased due to accelerated gluconeogenesis. In conclusion, SDT rats were more sensitive to APAP-induced chronic hepatotoxicity than SD rats and the high susceptibility of SDT rats was considered to be attributable to lowered hepatic glutathione levels induced by accelerated gluconeogenesis.

Estimation of potential risk of allyl alcohol induced liver injury in diabetic patients using type 2 diabetes spontaneously diabetic Torii-Leprfa (SDT fatty) rats.

In order to estimate the potential risk of chemicals including drug in patients with type 2 diabetes mellitus (T2DM), we investigated allyl alcohol induced liver injury using SD rats and Spontaneously Diabetic Torii-Leprfa (SDT fatty) rats as a model for human T2DM. The diabetic state is one of the risk factors for chemically induced liver injury because of lower levels of glutathione for detoxification by conjugation with chemicals and environmental pollutants and their reactive metabolites. Allyl alcohol is metabolized to a highly reactive unsaturated aldehyde, acrolein, which is detoxified by conjugation with glutathione. Therefore, we used allyl alcohol as a model compound. Our investigations showed that SDT fatty rats appropriately mimic the diabetic state in humans. The profiles of glucose metabolism, hepatic function tests and glutathione synthesis in the SDT fatty rats were similar to those in patients with T2DM. Five-week oral dosing with allyl alcohol to the SDT fatty rats revealed that the allyl alcohol induced liver injury was markedly enhanced in the SDT fatty rats when compared with the SD rats and the difference was considered to be due to lower hepatic detoxification of acrolein, the reactive metabolite of allyl alcohol, by depleted hepatic glutathione synthesis. Taking all the results of the present study into consideration, the potential for allyl alcohol to induce liver injury is considered to be higher in diabetic patients than in healthy humans.

Pathophysiological analyses of skeletal muscle in obese type 2 diabetes SDT fatty rats.

Sarcopenia is the age-related decrease of muscle mass and function. Diabetes and obesity are known to be risk factors that exacerbate sarcopenia, but the underlying mechanism of diabetes-related sarcopenia is still unknown. Obese type 2 diabetes SDT fatty rats show early onset of severe diabetes and there have been no reports on the characteristics of their skeletal muscle. Therefore, pathophysiological analyses were performed for the skeletal muscle in these rats. Diabetic male SDT fatty rats were sacrificed at 8, 16, 24, 32 and 40 weeks of age. Age-matched Sprague Dawley (SD) rats were used as the normal control. In addition to biological blood parameters, the soleus and the extensor digitorum longus muscles were examined for muscle weight, histopathology, and protein synthesis and degradation. Muscle grip strength was also examined. These results revealed that the muscle weights of the SDT fatty rats were significantly decreased from 16 weeks of age. The mean cross-sectional area of muscle fibers in the SDT fatty rats decreased from 24 weeks of age. Increased intramyocellular lipid accumulation, identified by immunohistochemistry for adipophilin and TEM, was observed in the SDT fatty rats from 8 weeks of age. Plasma insulin-like growth factor (IGF)-1 levels and muscle strength in the SDT fatty rats decreased at 24 weeks of age and thereafter. These pathophysiological findings have been reported both in sarcopenia in aged humans and in patients with diabetes. In conclusion, the SDT fatty rat was considered to be a useful model for analysis of diabetes-related sarcopenia.

High fructose diet feeding accelerates diabetic nephropathy in Spontaneously Diabetic Torii (SDT) rats.

Diabetic nephropathy (DN) is one of the complications of diabetes and is now the most common cause of end-stage renal disease. Fructose is a simple carbohydrate that is present in fruits and honey and is used as a sweetener because of its sweet taste. Fructose has been reported to have the potential to progress diabetes and DN in humans even though fructose itself does not increase postprandial plasma glucose levels. In this study, we investigated the effects of high fructose intake on the kidney of the Spontaneously Diabetic Torii (SDT) rats which have renal lesions similar to those in DN patients and compared these with the effects in normal SD rats. This study revealed that a 4-week feeding of the high fructose diet increased urinary excretion of kidney injury makers for tubular injury and accelerated mainly renal tubular and interstitial lesions in the SDT rats but not in normal rats. The progression of the nephropathy in the SDT rats was considered to be related to increased internal uric acid and blood glucose levels due to the high fructose intake. In conclusion, high fructose intake exaggerated the renal lesions in the SDT rats probably due to effects on the tubules and interstitium through metabolic implications for uric acid and glucose.

A novel JAK inhibitor JTE-052 reduces skin inflammation and ameliorates chronic dermatitis in rodent models: Comparison with conventional therapeutic agents.

Janus kinases (JAKs) are required for several inflammatory cytokine signalling pathways and are implicated in the pathogenesis of chronic dermatitis, including atopic dermatitis and psoriasis. JAK inhibitors are therefore promising therapeutic candidates for chronic dermatitis. In this study, we evaluated the effects of the novel JAK inhibitor JTE-052 on inflammatory responses associated with chronic dermatitis, and compared its profile with those of conventional therapeutic agents in rodent models of chronic dermatitis. JTE-052 inhibited the Th1-, Th2- and Th17-type inflammatory responses of human T cells and mast cells in vitro. Oral administration of JTE-052 inhibited skin inflammation in hapten-induced chronic dermatitis in mice, associated with reduced levels of inflammatory cytokines in the skin and immunoglobulin (Ig) E in serum. In contrast, although ciclosporin partly inhibited skin inflammation, it did not reduce interleukin (IL)-4 production in skin, and enhanced IgE production in serum. Oral administration of JTE-052 also inhibited skin inflammation in mouse models of atopic dermatitis and psoriasis induced by a mite extract, thymic stromal lymphopoietin or IL-23. The maximal efficacy of JTE-052 in these dermatitis models was superior to the conventional therapeutic agents, ciclosporin and methotrexate. Topical application of JTE-052 ointment ameliorated hapten-induced chronic dermatitis in rats more effectively than tacrolimus ointment. Furthermore, JTE-052 ointment did not cause the thinning of normal skin associated with topical corticosteroids. These results indicate that JTE-052 is a promising candidate as an anti-inflammatory drug for various types of chronic dermatitis, with a distinctly different profile from conventional therapy following either oral or topical application.

Hepatocellular adenoma with severe fatty change in a male Spontaneously Diabetic Torii rat.

The Spontaneously Diabetic Torii (SDT) rat is a rat model of nonobese type 2 diabetes mellitus, and hepatocellular adenomas have not been reported in this model. We report a hepatocellular adenoma with severe fatty change in a male 42-week-old SDT rat fed a high-fat diet. At necropsy, the animal had a whitish nodular mass of approximately 2 cm in diameter in the right medial lobe. Histologically, the mass was well demarcated from the surrounding tissues, slightly compressing the adjacent hepatic parenchyma and widely compartmented by fibrous connective tissues. The mass consisted of vacuolated tumor cells resembling hepatocytes with a solid and occasionally trabecular growth pattern. Abundant neutral lipids, which were positive for fat with Oil Red O stain and which ultrastructurally had moderately dense material, were contained within the vacuoles of the tumor cells. Immunohistochemically, the tumor cells showed an increase in immunoreactivity or number for Cytokeratin 8/18 and proliferating cell nuclear antigen but were negative for mesenchymal markers. From these findings, the mass could be distinguished from hepatocellular hyperplasia and was diagnosed as hepatocellular adenoma. In rats, hepatocellular adenoma accompanied by severe fatty change is rare, and this is the first report of a hepatocellular tumor with severe fatty change in a SDT rat.

Contribution of hyperglycemia on diabetic complications in obese type 2 diabetic SDT fatty rats: effects of SGLT inhibitor phlorizin.

The spontaneously diabetic torii (SDT) fatty rat is a new model of type 2 diabetes showing overt obesity, hyperglycemia and hyperlipidemia. With early onset of diabetes mellitus, diabetic microvascular complications, including nephropathy, peripheral neuropathy and retinopathy, are observed at young ages. In the present study, blood glucose levels of female SDT fatty rats were controlled with phlorizin, a non-selective SGLT inhibitor, to examine whether and how these complications are caused by hyperglycemia. Phlorizin treatment adequately controlled plasma glucose levels during the experiment. At 29 weeks of age, urinary albumin excretion considerably increased in SDT fatty rats. Glomerulosclerosis and tubular pathological findings also indicate diabetic nephropathy. These renal parameters tended to decrease with phlorizin; however, effects were partial. Sciatic nerve conduction velocities were significantly delayed in SDT fatty rats compared with Sprague-Dawley (SD) rats. Intraepidermal nerve fiber density, an indicator of subclinical small nerve fiber neuropathy, significantly decreased in SDT fatty rats. Retinal dysfunction (prolongation of peak latency for oscillatory potential in electroretinograms) and histopathological eye abnormalities, including retinal folding and mature cataracts were also observed. Both nerve and eye disorders were prevented with phlorizin. These findings indicate that severe hyperglycemia mainly causes diabetic complications in SDT fatty rats. However, other factors, such as hyperlipidemia and hypertension, may affect diabetic nephropathy. These characteristics of diabetic complications will become helpful in evaluating new drugs for diabetic complications using SDT fatty rats.

Immunohistochemical Characterization of Spontaneous Sertoli Cell Clusters in the Seminiferous Tubules of C57BL/6J Mice.

Cell clusters were observed in the seminiferous tubules of C57BL/6J mice as a spontaneous lesion in a 2-week toxicity study, and they were demonstrated to be basically composed of Sertoli cells by immunohistochemistry for claudin-11 and GATA-4 (GATA-binding protein 4), which are both Sertoli cell markers. The clusters were composed of about 5 to 50 cells, which had eosinophilic and occasionally vacuolated cytoplasm with an unclear cell boundary. The cell clusters involved some sperm. No mitotic figures were observed and no immunoreactivity for proliferating cell nuclear antigen (PCNA) was detected in the clusters. In most cases, the cell clusters were observed in seminiferous tubules that also showed degenerative changes. In rare instances, cell aggregates immunohistochemically positive for claudin-11 were observed in the lumen of the epididymis, suggesting that some of the Sertoli cell clusters were sloughed off from the seminiferous epithelium into the epididymal ducts. To our knowledge, this is the first report of Sertoli cell clusters in any animal species except for transgenic or surgically altered animals.

Ocular inflammation in uveal tract in aged obese type 2 diabetic rats (Spontaneously Diabetic Torii fatty rats).

We report uveitis observed in an obese type 2 diabetes rat model, Spontaneously Diabetic Torii Lepr(fa) (SDT fatty) rats aged over 50 weeks. The eyes of SDT fatty rats (16 animals: 7 males and 9 females with 50 or 60 weeks of age) were examined histopathologically. Infiltration of inflammatory cells in the uveal tract was observed in 13 of 16 animals. One female showed severe inflammation affecting the entire uveal tract including the iris, ciliary body, and choroid with a variety of inflammatory cells (neutrophils, lymphocytes, and macrophages). Those changes clinically mimic the findings of diabetic iridocyclitis in diabetic patients. Uveitis associated with diabetes can occur in diabetic patients but the pathogenesis still remains unknown. Since increased extramedullary hematopoiesis in the spleen and abscess in the genital and lower urinary tracts were observed in some SDT fatty rats, increased susceptibility to infection, prolongation of inflammatory states, and disorders of the immune system were considered to be possible factors of the uveitis in aged SDT fatty rats. There have been few reports on how diabetes has influence on the development of uveitis associated with bacterial infection. The SDT fatty rat can be an animal model to investigate diabetes-associated uveitis.

Enhancement of acetaminophen-induced chronic hepatotoxicity in restricted fed rats: a nonclinical approach to acetaminophen-induced chronic hepatotoxicity in susceptible patients.

Acetaminophen (APAP) is a commonly used and effective analgesic and antipyretic agent. However, some patients encounter hepatotoxicity after repeated APAP dosing at therapeutic doses. In the present study, we focused on the nutritional state as one of the risk factors of APAP-induced chronic hepatotoxicity in humans and investigated the contribution of undernourishment to susceptibility to APAP-induced chronic hepatotoxicity using an animal model mimicking undernourished patients. Rats were divided into 2 groups: the ad libitum fed (ALF) and the restricted fed (RF) rats and were assigned to 3 groups (n = 8/group) for each feeding condition. The animals were given APAP at 0, 300 and 500mg/kg for 99 days under each feeding condition. Plasma and urinary glutathione-related metabolites and liver function parameters were measured during the dosing period and hepatic glutathione levels were measured at the end of the dosing period. In the APAP-treated ALF rats hepatic glutathione levels were increased and hepatic function parameters were not changed, but in the APAP-treated RF rats hepatic glutathione levels were decreased at 500mg/kg and hepatic function parameters were increased at 300 and 500mg/kg. Moreover the urinary endogenous metabolite profile after long-term treatment with APAP in the ALF and RF rats was similar to that in human non-responders and responders to APAP-induced chronic hepatotoxicity, respectively. In conclusion, the RF rats were more sensitive to APAP-induced chronic hepatotoxicity than the ALF rats and were considered to be a useful model to estimate the contribution of the nutritional state of patients to APAP-induced chronic hepatotoxicity.

Inflammatory myopathy with severe tongue atrophy in Pembroke Welsh Corgi dogs.

A disease characterized by tongue and facial muscle atrophy has been recognized sporadically among Pembroke Welsh Corgi (PWC) dogs in Japan. The present study describes the pathologic findings of this canine syndrome. Histopathologic examinations were performed in 2 dogs, including a case of muscular biopsy. Identification and characterization of autoantibodies were attempted by fluorescent antibody test (FAT) and Western blot (WB) by using sera from 7 PWC dogs with typical clinical features, 6 PWC dogs with other clinical signs, and 2 from other breeds with polymyositis. Clinically, the 7 affected PWC dogs exhibited dysphagia with severe tongue atrophy, facial muscular atrophy, and occasional walking difficulty. Histopathologic examinations of the 2 dogs with clinical symptoms revealed moderate to severe inflammatory lesions characterized by lymphohistiocytic infiltration and muscular atrophy in the tongue and/or femoral muscles. The tongue lesions were very severe and accompanied by diffuse fatty infiltration. There were no major lesions in the nervous tissues examined. By FAT, an autoantibody against the cross striation of skeletal muscle was detected in sera from 5 affected PWC dogs. By using WB analysis, the autoantibodies recognized a 42-kDa molecule in striated muscle but not in the nervous tissues. All of the findings indicated that the unique disease of PWC dogs might be generalized inflammatory myopathy, whereas the detailed etiology concerning the dominant involvement of tongue muscles and the role of the autoantibody in the canine disease remain to be clarified.