A spontaneous thymic carcinosarcoma in a young Sprague Dawley rat.

We encountered a case of spontaneous thymic carcinosarcoma in a young Crl:CD (Sprague Dawley) rat. Grossly, a white multinodular mass replaced the thymus in the thoracic cavity. Histologically, multiple nodules were separated by fibrous stroma, and each nodule included isolated regions that were composed of epithelial or non-epithelial tumor cells. The epithelial tumor cells were relatively large and round to polygonal cells with large nuclei and weakly eosinophilic cytoplasm. These cells were cytokeratin-positive and vimentin-negative. These cells infiltrated the lungs. The non-epithelial tumor cells were poorly differentiated, small, round to spindle-shaped cells with small nuclei and basophilic cytoplasm. These cells were vimentin-positive and mostly cytokeratin-negative. Many islands of cartilage were observed near non-epithelial cells. Based on these findings, the tumor was diagnosed as a primary thymic carcinosarcoma consisting of a malignant thymoma composed of epithelial tumor cells and a mesenchymal chondrosarcoma composed of non-epithelial tumor cells.

Selective Chemonucleolysis With Condoliase in Cynomolgus Monkeys.

Selective chemonucleolytic effects of condoliase, a glycosaminoglycan degrading enzyme, was investigated histopathologically in cynomolgus monkeys. Condoliase was administered once into the lumber intervertebral disc (IVD), and as a comparative control, chymopapain, a proteolytic enzyme, was administered in a similar manner. Histopathological changes of the IVD and the adjacent vertebral body (VB) were examined at 1 to 26 weeks after administration. Major changes induced by condoliase in the IVD were degenerative and necrotic changes in the nucleus pulposus, annulus fibrosus, cartilaginous endplate (CEP), and epiphyseal growth plate (EGP); focal disappearance of the EGP; and neovascularization and ossification of the CEP. Decreased/necrosis of bone marrow cells with new bone formation was observed in the VB. Cellular regeneration in the IVD was observed as a recovery changes on and after week 4. The changes in the IVD and VB subsided at week 26. Chymopapain induced qualitatively similar but more widely extended changes. The degrees of the changes in the IVD and VB were more severe than those of condoliase, and the changes were exacerbated even at week 26. These results indicated that histopathological changes caused by condoliase were less severe and more selective than those by chymopapain.

Effects of Repeated Intravenous Administration of Dextrans,Water-soluble Macromolecules, in Rats.

We investigated the influence of repeated intravenous administration of dextrans (DEXs) to rats. Seven-week-old Sprague Dawley rats (6 males/group) were given intravenously 10% saline solutions of dextrans (DEXs), 40 kDa or 200-300 kDa, at a dose level of 5 mL/kg/day for 28 days and they were examined histopathologically. Another group (3 males/group) was administered saline in a similar manner and served as the control. Histopathological changes indicating accumulation of DEXs in the mononuclear phagocyte system (MPS) and the liver were noted in the treated groups. The incidence and severity of the findings were molecular weight-dependent, except for the lungs. These results are considered useful in interpreting data from preclinical studies, in which DEXs or their derivatives are administered as test or control substances.

Telomerase-specific oncolytic adenoviral therapy for orthotopic hepatocellular carcinoma in HBx transgenic mice.

The telomerase-specific replication-competent oncolytic adenovirus, Telomelysin, was developed for virus-mediated preferential lysis of tumor cells. Its selectivity is derived from a human telomerase reverse transcriptase (hTERT) promoter-driven active viral replication, which occurs in cancer cells with high telomerase activity but not in normal cells lacking such activity. Because the TERT activity is elevated in most cases of hepatocellular carcinoma (HCC), the current study aims to investigate whether Telomelysin can be used for treatment of HCC. The oncolytic effect of Telomelysin has been investigated both in vitro using cell culture and in vivo using an immunocompetent in situ orthotopic HCC model. In this model, HCC developed spontaneously in the liver of HBx transgenic mice, which is pathologically and genetically similar to human HCC. In cell culture assay, Telomelysin lyses HCC cell lines at a low multiplicity of infection (MOI), ranging 0.77-6.35 (MOI [PFU/cell]). In the orthotopic HCC model, Telomelysin showed a potent oncolytic effect on HCC but spared normal liver tissue. Dose escalation analysis identified a safety dose of 1.25 × 10(8) PFU for this model. The effect of multiple injections of Telomelysin was also evaluated in this immunocompetent HCC model. We found that the virus replicates in HCC after a second intratumoral injection despite an immune response induced by the previous injection. This preclinical study shows that Telomelysin can be used for treatment of human HCC at an appropriate dosage and that its tumor-killing activity persists after multiple injections.

Selective CB1 cannabinoid receptor antagonist, SR141716A, attenuates liver injury induced by Concanavalin A.

AIM: The aim of this study was to investigate the hepatoprotective activity of a selective cannabinoid receptor 1 (CB1) antagonist, SR141716A, in a Concanavalin A (Con A)-induced mouse liver injury model and to determine whether SR141716A has an effect on the production of inflammatory cytokines and chemokines induced by Con A. RESULTS: Injection of Con A (20 mg/kg) to mice developed hepatitis determined by plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) elevation and necrosis in the liver. Pretreatment with SR141716A (30 mg/kg) significantly reduced plasma AST and ALT level, protected against necrosis in the liver, and significantly reduced plasma cytokine and chemokine levels, including TNFalpha, IFN-gamma, CXCL9, MIP1-alpha, and IL-10 and no change decreased in IL-4. CONCLUSIONS: The selective CB1 antagonist, SR141716A, exerts a hepatoprotective effect on Con A-induced liver injury in mice by attenuating the increase in cytokine and chemokine levels and inhibiting hepatocyte injury. These findings raise the possibility of using CB1 antagonists as anti-inflammatory drugs for treating hepatitis as well as other inflammatory diseases.

Effects of the selective EP4 antagonist, CJ-023,423 on chronic inflammation and bone destruction in rat adjuvant-induced arthritis.

Prostaglandin E2 (PGE2) produced by cyclooxygenase (COX) is a potent pro-inflammatory mediator. We have recently discovered CJ-023,423, a highly selective antagonist of EP4 receptors, one of the PGE2 receptors. This agent is suitable for exploring the effects of blocking EP4 receptors following oral administration in rats. In this study, CJ-023,423 was used in rats with adjuvant-induced arthritis (AIA) to investigate the role of the EP4 receptor in chronic inflammation and bone destruction. These effects were compared with those of rofecoxib, a selective COX-2 inhibitor. CJ-023,423 had significant inhibitory effects on paw swelling, inflammatory biomarkers, synovial inflammation and bone destruction in AIA rats. In particular, the inhibitory effect on paw swelling in AIA rats was comparable to that of rofecoxib. These results suggest that PGE2 acting via the EP4 receptor is involved in the development of chronic inflammation and bone destruction, particularly with respect to oedema in AIA rats. This is the first study to confirm the in-vivo effects of EP4 receptor blockade on inflammation and bone destruction in AIA rats with a small-molecule compound.

Nitrofurazone-induced gene expressions in rat hepatocytes and their modification by N-acetylcysteine.

The antibiotic nitrofurazone (NF) at a subtoxic dose has been shown to increase hepatocyte DNA synthesis with no preceding cell damage or necrosis. This was suppressed by concomitant administration of the antioxidant N-acetylcysteine (NAC), which suggests that free radical production is involved in the process. In this study, male F344 rats were given a single oral subtoxic dose of NF to investigate the changes in genes implicated in hepatocyte proliferation between 1 and 20h postdose by real-time PCR. Some rats were also given NAC to examine the involvement of free radicals. There were transient and sequential increases in mRNA levels of c-myc and c-jun shortly after the administration, followed by tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-alpha (TGF-alpha), c-Ha-ras, and cyclin E. The increases were blocked by concomitant administration of NAC. In contrast, there were no NF-specific increases in c-fos, hepatocyte growth factor, epidermal growth factor or cyclin D1 mRNAs. These results indicate that the induction of hepatocyte proliferation by NF is triggered by free radicals, with a pathway involving increases in c-jun, c-myc, TNF-alpha, TGF-alpha, c-Ha-ras, and cyclin E. The results also indicate that NF-induced proliferation resembles that of other mitogens.

Nitrofurazone induces non-regenerative hepatocyte proliferation in rats.

The antibiotic nitrofurazone (NF) has been known for its testicular toxicity; in contrast, much less is known about its effect on the liver. NF was given to male rats for up to 7 consecutive days to evaluate NF-induced effects on the liver. NF increased hepatocyte DNA synthesis and liver weight in a dose-dependent manner, with no apparent histological or biochemical evidence of cell damage or loss. The hepatocyte proliferation ceased after a few days despite the continuation of treatment. The absence of cell damage indicates that NF-induced hepatocyte proliferation is different from regenerative proliferation that is seen after partial hepatectomy or cell necrosis.

Immunohistochemical study on inducible type of nitric oxide (iNOS), basic fibroblast growth factor (bFGF) and tumor growth factor-beta1 (TGF-beta1) in arteritis induced in rats by fenoldopam and theophylline, vasodilators.

Arteritis induced in rats by vasodilators, fenoldopam and theophylline, was examined immunohistochemically for expressions of inducible type of nitric oxide synthase (iNOS), basic fibroblast growth factor (bFGF) and tumor growth factor-beta1 (TGF-beta1). Rats were administered fenoldopam for 24 hours by intravenous infusion with or without following repeated daily oral administrations of theophylline. Irrespective of theophylline administration, iNOS antigens were remarkably abundant in ED-1-positive cells on day 5 and 8 post-fenoldopam-infusion (DPI); bFGF antigens were remarkably abundant in ED-1-positive cells on 1 and 3 DPI; TGF-beta1 antigens were observed in ED-1-positive cells on and after 5 DPI. These results suggest that the peak expression of iNOS antigen was followed by that of bFGF antigen, and bFGF may have a suppressive effect on iNOS expression in these rat arteritis models. On the other hand, TGF-beta1 was not considered to have a suppressive effect on iNOS expression in these models.

Anti-PDGF-B monoclonal antibody reduces liver fibrosis development.

AIM: To evaluate the usefulness of a platelet-derived growth factor (PDGF)-B specific monoclonal antibody (mAb) as a therapeutic agent to treat chronic liver fibrosis. METHODS: Liver fibrosis was induced in ICR mice by bile duct ligation (BDL) or BALB/c mice by weekly injection of concanavalin A (ConA) for 4 or 8 weeks. A mAb specific for mouse and human PDGF-B chain, AbyD3263, was generated, tested in vitro and administered twice a week throughout the experimental period. RESULTS: AbyD3263 showed neutralizing activity against mouse and human PDGF-B chain in cell-based assays, as measured in vitro by inhibition of phosphorylation of PDGF receptor ß and proliferation of hepatic stellate cells induced by PDGF-BB. The half life of AbyD3263 in mice exceeded 7 days and dosing of animals twice a week resulted in constant plasma levels of the mAb. Induction of liver fibrosis by BDL and ConA resulted in elevated levels of alanine aminotransferase (ALT) in plasma and hydroxyproline in the liver. Treatment with AbyD3263 did not modify ALT levels, but significantly reduced hydroxyproline content in the liver with a maximum reduction of 39% and 54% in the BDL and ConA models, respectively, compared to controls. Conclusios: Consistent with the notion that PDGF-BB plays an important role in the progression of liver fibrosis, AbyD3263 exhibits efficacy in pre-clinical disease models suggesting that pharmacological inhibition of PDGF-B chain may be a therapeutic approach to treat liver fibrosis.

Pathophysiological characteristics of dimethylnitrosamine-induced liver fibrosis in acute and chronic injury models: a possible contribution of KLF5 to fibrogenic responses.

Dimethylnitrosamine administration induces a rapid increase in collagen deposition with concomitant proliferation of hepatic stellate cells in rats. Here, we investigated the pathophysiological profiles of acute and chronic hepatic fibrosis states and attempted to determine the possible role of Kruppel-like factor-5 (KLF5) in this model. In acute study using a single drug injection, we observed a rapid transient increase of ALT and mRNA levels of KLF5 followed by increases in fibrosis-related genes. Repeated administration of dimethylnitrosamine once a week caused early damage with severe fibrosis and sustained hepatocyte injury, while intermittent injections at 2-week intervals induced only modest fibrosis from 3 weeks. Weekly administration also induced profound upregulation of collagen I, alpha-smooth muscle actin, and KLF5 mRNA. In contrast, such continued augmentation was not observed after intermittent injections; KLF5 increased only after 3 weeks. These results suggested that dimethylnitrosamine induced a rapid hepatic fibrogenic response with a possible participation of KLF5.

Temporal reduction in size of salivary acinus in rats induced by theophylline.

Repeated administration of theophylline, a phosphodiesterase inhibitor, induces the enlargement of the salivary glands in rats. Time-course changes after a single administration of theophylline were examined in the salivary glands, including phosphodiesterase enzyme activity, and the expression of aquaporin 5 (AQP5), a water channel. We also examined the contribution of beta-adrenergic receptors to theophylline-induced salivary changes. Male F344 rats were given 50 mg/kg of theophylline intraperitoneally either alone or concurrently with a 10 mg/kg subcutaneous injection of propranolol. After treatment with theophylline alone, the weight and histology of the submaxillary and parotid glands were examined. Phosphodiesterase activity and AQP5 were detected by enzyme- and immuno-histochemistry, respectively. At 4 hours, 8 hours, or both, organ weights were decreased with depletion of secretory vesicles in the acinar cells. In the submaxillary glands, reduced activity of phosphodiesterase and increased expression of AQP5 in the intercalated ducts were observed at 4 hours. When co-administered, propranolol partially abolished theophylline-induced glandular reduction. These results suggest that the theophylline-induced transient reduction in size of the salivary glands is attributable not only to phosphodiesterase inhibition but also to beta-adrenergic receptor activation and that the intercalated ducts in submaxillary glands play a role in the production of saliva.