Thymic granulomatous lesions in pigs.

Rare cases of thymic granulomatous lesions were found in pigs. The lesions consisted of epithelioid cells, multinucleated giant cells, and lymphocytes. Such lesions also were observed in the mesenteric lymph nodes, spleen, kidney, and stomach. The cytoplasm of the majority of giant cells and some epithlioid cells was periodic acid-Schiff (PAS) positive. All cells were positive for vimentin, lysozyme, and desmin. Ultrastructurally, the giant cells were rich in organella and attached to adjacent epithelioid cells by membrane interdigitation. The cells included numerous coated vesicles and granules. No etiologic pathogen, including porcine circovirus type 2, was detected in the lesions. This is the rare case of idiopathic thymic granulomatous lesion in pigs.

Two cases of feline malignant craniopharyngioma.

Tumors at the cranial base in 2 cats (a 9 1/2-year-old, castrated male Chinchilla and a 7-year-old, castrated male American Shorthair) were diagnosed as malignant craniopharyngioma. The tumor lesion was histopathologically divided into four parts: 1) a small acinus part, in which relatively large cells with a pale cytoplasm composed small acini; 2) a duct part, in which small cuboidal cells composed ducts; 3) a cyst part, in which there were large cysts lined with flat cells; and 4) a pavement part, in which large multi-angular-shaped cells proliferated in a pavement pattern. The epithelial cells of some parts were positive for keratin by immunohistochemistry. Histopathologic findings of the present feline cases were identical to those of malignant craniopharyngioma in other animal species.

GM2-gangliosidosis variant 0 (Sandhoff-like disease) in a family of Japanese domestic cats.

A five-month-old, female Japanese domestic shorthair cat with proportionate dwarfism developed neurological disorders, including ataxia, decreased postural responses and generalised body and head tremors, at between two and five months of age. Leucocytosis due to lymphocytosis with abnormal cytoplasmic vacuolations was observed. The concentration of G(M2)-ganglioside in its cerebrospinal fluid was markedly higher than in normal cats, and the activities of beta-hexosaminidases A and B in its leucocytes were markedly reduced. On the basis of these biochemical data, the cat was diagnosed antemortem with G(M2)-gangliosidosis variant 0 (Sandhoff-like disease). The neurological signs became more severe and the cat died at 10 months of age. Histopathologically, neurons throughout the central nervous system were distended, and an ultrastructural study revealed membranous cytoplasmic bodies in these distended neurons. The compound which accumulated in the brain was identified as G(M2)-ganglioside, confirming G(M2)-gangliosidosis. A family study revealed that there were probable heterozygous carriers in which the activities of leucocyte beta-hexosaminidases A and B were less than half the normal value. The Sandhoff-like disease observed in this family of Japanese domestic cats is the first occurrence reported in Japan.

Sunburn reaction in the dorsal skin of hypotrichotic WBN/ILA-Ht rats.

The dorsal skin responses to a single irradiation with a high-dose of UVB (10kJ/m2) were examined histologically and immunohistochemically in UVB-sensitive Wistar-derived hypotrichotic WBN/ILA-Ht rats (HtRs). Sunburn cells (SBCs) which were characterized by pyknotic nuclei and eosinophilic cytoplasm and had ultrastructual characteristics of apoptotic cells were first observed in the epidermis at 3 hours (h) after irradiation. The number peaked at 6 h, and then decreased rapidly. The expressions of p53 protein, which is known to be closely related to the formation of SBCs, and of p21 protein, which is one of the transcriptional target genes of p53, were immunohistochemically detected, and their labeling index (LI) in the epidermis peaked at 12 to 24 h (p53) or at 24h (p21) after irradiation. On the other hand, proliferating cell nuclear antigen (PCNA)-LI in keratinocytes was significantly lower than the control group at 6 h after irradiation and thereafter it increased and became significantly higher than the control group from 24 to 48 h. At 48 h, moderate hyperplasia with moderate numbers of mitotic keratinocytes was first observed in the epidermis. In the dermis, mild edema developed from 12 to 36 h and it accompanied mild lymphocyte infiltration at 36 h. Judging from the present results, it was suggested that some factors other than p53 might be involved in SBC formation, and that p53 might induce p21 protein and play an important role in cell growth arrest in keratinocytes after UVB irradiation.

Dorsal skin responses to subchronic ultraviolet B (UVB)-irradiation in Wistar-derived hypotrichotic WBN/ILA-Ht rats.

Dorsal skin responses to a subchronic UVB-irradiation (10kJ/m2/rat /day), were examined in Wistar-derived hypotrichotic WBN/ILA-Ht rats for up to 3 months. Hyperplasia of epidermal cells and hair follicle epithelial cells as well as parakeratosis developed at 1 month and progressed thereafter, resulting in a prominent epidermis thickening and formation of epidermal ingrowths projecting into the dermis. At the same time, the percentage of proliferating cell nuclear antigen (PCNA)-positive epidermal cells significantly increased after I month. In some portions of the hyperplastic epidermis, especially of the epidermal ingrowths, keratinocytes were somewhat pleomorphic and migrated into the dermis. In the upper dermis, edema with capillary congestion, mast cell infiltration and fibroblast proliferation developed at I month, and the intensity of edema and the number of dermal mast cells was most prominent at 3 months. Edema spread to the epidermis, resulting in intercellular edema and subsequent dissociation of epidermal cells. Degeneration of collagen fibers was also detected in the upper dermis, especially beneath the epidermis. In addition, although not significant because of a large individual difference, the serum IgE concentration, showed a tendency to increase after 2 months. The present study clarified the characteristics of the dorsal skin responses to a subchronic UVB-irradiation in rats.

Age-related changes in the dorsal skin histology in Mini and Wistar rats.

Mini rats (Jcl: WistarTGN(ARGHGEN)1Nts (MRs) are Wistar rat (WR)-derived transgenic rats in which the expression of growth hormone (GH) gene is suppressed by the presence of antisense transgene. The plasma GH level of MRs is reduced to 40 to 60% of that of WRs. In this study, to evaluate the influence of GH deficiency on the skin nature, age-related changes in the dorsal skin histology were compared between male MRs and WRs. Although there were no essential differences in the skin structures between the two strains, MRs had thinner skin with less collagens, more abundant subcutaneous adipose tissues and small-sized sebaceous glands compared with WRs. On the other hand, the hair cycle evaluated by the morphology and the depth of hair follicles was greatly different between them. Namely, two cycles of 4 weeks each were observed in both strains during the first 8 weeks after birth, but the cycle entered a long-lasting quiescence (telogen phase) in MRs while the 3rd cycle started in WRs afterwards. The lower level of serum insulin-like growth factor (IGF)-1 in MRs may be related to such a difference in hair cycle pattern, although the levels of IGF-1 and IGF-1 receptor mRNAs in the dorsal skin tissues were similar between MRs and WRs. MRs are considered to be a useful animal model for dermatopathy in patients suffering from GH deficiency and for grasping a clue to elucidate the exact effects of GH on the skin nature, especially on hair follicle development.

Prevention of cerebral vasospasm by a novel endothelin receptor antagonist, TA-0201.

This study was designed to examine the preventive effect of a novel endothelin (ET)-receptor antagonist TA-0201 on the cerebral vasospasm in a canine double-hemorrhage model. TA-0201 (10(-9)-10(-7) M) inhibited ET-1-induced vasoconstriction in the isolated canine basilar artery without endothelium in a concentration-dependent manner. Its pA2 value was 9.2 (ET(A) antagonism). In a canine double-hemorrhage model, intravenous treatment with TA-0201 (3 mg/kg, twice a day for 7 days) ameliorated the basilar artery narrowing significantly on day 7 compared with that in nontreated dogs. The reductions of the basilar artery diameter were 26.1+/-3.9% and 40.5+/-4.1% with and without TA-0201 treatment, respectively (p<0.05). Histologic study on day 7 indicated that treatment with TA-0201 inhibited vessel-wall damage such as disintegration of endothelium architecture and degeneration of medial smooth-muscle cells. We conclude that intravenous treatment with TA-0201 prevents the development of cerebral vasospasm and accompanying pathologic changes of the vessel wall, probably through blockade of ET(A) receptors.

Ecabet sodium, a novel locally-acting anti-ulcer agent, protects the integrity of the gastric mucosal gel layer from pepsin-induced disruption in the rat.

BACKGROUND: Ecabet sodium, a novel non-systemic anti-ulcer agent, possesses high affinity to gastric adherent mucus, which plays an important role in the protection of the gastric epithelium against acid and pepsin. AIM: To assess the effect of ecabet on pepsin-induced degradation of the structure of the mucus gel layer. METHODS: Everted sacs of rat stomach were incubated in HCl solution containing pepsin with or without ecabet. Pepsin-induced release of the cleaved peptides and hexosamine from the sacs was determined. Changes in the molecular size of glycoproteins in the adherent mucus (using gel filtration methods) and in the morphology of the epithelium (using both light and scanning electron microscopy) were also examined. RESULTS: Ecabet reduced the pepsin-induced release of peptides and hexosamine, depending on its content in the adherent mucus. Pepsin treatment partially lowered the molecular weight of native glycoproteins in the adherent mucus, caused exfoliation of the epithelial cells, and degraded the network-like ultrastructure of the mucus layer, giving it a lumpy, globular appearance. Ecabet prevented both the pepsin-induced molecular size shift in mucus glycoproteins, and morphological alteration of the epithelium, including ultrastructural derangement of the mucus gel layer. CONCLUSION: Ecabet protects the polymeric structure of mucus glycoproteins from proteolytic degradation by pepsin, and thus maintains integrity of the gastric mucus gel layer.

Antithrombotic action of TA-993, a new 1,5-benzothiazepine derivative, in a canine model of femoral arterial thrombosis.

TA-993 is a novel 1,5-benzothiazepine derivative of l-cis configuration, having a potent antiplatelet action and an increasing action on femoral blood flow. We evaluated the antithrombotic effect of TA-993 in a canine model of femoral arterial thrombosis. Thrombus was induced by both application of direct anodal current to the femoral artery and partial occlusion of the artery. The partial occlusion by placing an adjustable occluder on the artery and the current application were carried out 40 and 60 min after the intraduodenal administration of drugs, respectively. In control dogs, complete sustained occlusion of the femoral artery due to thrombus occurred 55.4 +/- 9.2 min after the onset of current application. TA-993 (3 and 10 mg/kg) dose-dependently prolonged the time for occlusion. Aspirin (30 mg/kg) also prolonged it. TA-993, 10 mg/kg, significantly inhibited whole-blood aggregation 60 min after the administration with a weaker potency than that of aspirin (30 mg/kg), whereas 3 mg/kg of TA-993 did not. The inhibitory effect of TA-993 (10 mg/kg) on platelet aggregation was maintained for >7 h. Moreover, TA-993 (10 mg/kg) increased femoral blood flow in spite of the partially occluded condition. These results indicate that TA-993 has an antithrombotic effect on femoral arterial thrombosis and suggest that an increasing action on femoral blood flow of TA-993 is more relevant than its antiplatelet action to the antithrombotic effect in this model.

Morphologic changes in hepatocyte nuclei of streptozotocin (SZ)-induced diabetic mice.

Morphological examinations were carried out on hepatocyte nuclei of streptozotocin (SZ)-induced diabetic mice. The area of hepatocyte nuclei in diabetic mice was about two times larger than that in control mice, and the incidence of hepatocytes with intranuclear inclusions was 3.4 +/- 0.2% in diabetic mice and 0% in control mice, respectively. Although the incidence of binuclear hepatocytes was not significantly different between diabetic (14.5 +/- 4.6%) and control mice (16.4 +/- 4.4%), the morphology of the nuclei of binuclear hepatocytes was apparently different between diabetic and control mice. Namely, the nuclei of binuclear hepatocytes of control mice were round and identical in ultrastructural appearance, and they did not differ from those of mononuclear diploid hepatocytes. On the other hand, the nuclei of binuclear hepatocytes of diabetic mice were not identical in distribution pattern of chromatin granules, and they frequently varied in size and showed irregular contours.

Bactericidal effect of ecabet sodium on clarithromycin- and metronidazole-resistant clinical isolates of Helicobacter pylori.

Ecabet sodium showed a bactericidal effect on Helicobacter pylori NCTC 11637 which is susceptible to antimicrobial agents (Antimicrob. Agents Chemother, 39: 1295-1299, 1995). In the present study, we investigated the bactericidal effect of ecabet sodium on clarithromycin- and metronidazole-resistant strains of clinical isolates of H. pylori under acidic conditions. In a buffer supplemented with urea at pH 3.0, ecabet sodium decreased the number of viable cells in both isolates. In the morphological study, ecabet sodium changed the isolates from the bacilliform to the horseshoe-shaped form and denatured the cytoplasm. Ecabet sodium also showed the bactericidal effect on both isolates in buffers at pH 4.0 and 5.0, and the bactericidal effect was dependent on pH, i.e., the lower the pH, the greater the effect. These results suggest that the susceptibility of H. pylori to antimicrobial agents does not affect the bactericidal effect of ecabet sodium on H. pylori.

Hydrogen peroxide-induced dermatitis in WBN/Kob-Ht rats.

Light and electron microscopic examinations were carried out on the dorsal skin to which hydrogen peroxide (HPO) (3, 6, and 10%) was topically applied for 7 consecutive days in Wistar rat-derived inbred WBN/Kob-Ht rats which have an autosomal dominant gene responsible for their characteristics of hypotrichosis. In addition to focal epidermal thickening, keratinocyte necrosis, dermal mononuclear cell infiltration and focal detachment of the epidermis from the dermis by fluid-filled spaces were detected. This is thought to be brought about by edema due to prominent capillary endothelial damage in the superficial dermis. The damage to keratinocytes and capillary endothelial cells was thought to be induced by HPO itself and free radicals generated by HPO. In addition, these changes were apparently more severe in WBN/Kob-Ht rats than in Wistar rats used as controls.

Antithrombotic effect of TA-993, a novel 1,5-benzothiazepine derivative, in conscious rats.

Since reported experimental models of thrombosis are not suitable for comparison of several drugs by oral administration, we developed a convenient model for this purpose by applying direct current through an intravascular electrode. In conscious rats, which were implanted with anodal electrodes in the abdominal aorta on the day before the experiment, application of 200 microA of direct current induced the formation of a platelet-rich thrombus around the intravascular electrode. Using this model, we studied the antithrombotic effect of the novel antiplatelet agent TA-993, (-)-cis-3-acetoxy-5-(2-(dimethylamino)ethyl)-2,3-dihydro-8-methyl-2-(4- methylphenyl)-1,5-benzothiazepin-4(5H)-one maleate, and compared its effect with other antiplatelet agents. TA-993 at doses of 30 mg/kg, p.o. or more by single administration or at doses of 10 mg/kg or more by repeated administration dose-dependently suppressed the thrombus formation. Aspirin (10 mg/kg, p.o. or more), cilostazol (100 mg/kg, p.o.) and ticlopidine (30 mg/kg, p.o. or more) also suppressed the thrombus formation by single administration. These results suggest that TA-993 has a comparable antithrombotic effect with other antiplatelet agents, and thus it is a possible remedy for thrombotic and embolic diseases.

Bacterial activity of a new antiulcer agent, ecabet sodium, against Helicobacter pylori under acidic conditions.

Helicobacter pylori NCTC 11637, which is nonviable at pH 3.0, became viable after addition of 10 mM urea owing to ammonia production by urease. In a buffer supplemented with urea, ecabet sodium decreased both the production of ammonia and the number of viable cells of H. pylori NCTC 11637 and changed the bacteria from the bacilliform to the horseshoe or doughnut shape in a concentration-dependent manner. In particular, ecabet sodium (2 and 4 mg/ml) decreased the number of viable cells below the control level. Benzohydroxamic acid, a urease inhibitor, also caused a decrease in ammonia production accompanied by a decrease in the number of viable cells and changed the morphological form at pH 3.0, but the number of viable cells was not lowered below the control level. In buffers at various pHs without urea, ecabet sodium showed a concentration-dependent bactericidal effect on H. pylori at pHs 4.0 and 5.0 but not at pHs 6.0 and 7.0 while benzohydroxamic acid caused only a slight decrease in the number of viable cells at pH 4.0. These results suggest that ecabet sodium has strong bactericidal activity in addition to its urease-inhibiting activity under acidic conditions.

Comparison of atherogenicity of soybean oil and peanut oil, and effect of clentiazem on diet-induced atherosclerosis in rabbits.

Rabbits were fed with two kinds of atherogenic diet, one containing 0.5% cholesterol and 3% soybean oil and the other 0.5% cholesterol and 6% peanut oil, for three months to compare the atherogenic property of the diets. The soybean oil diet seemed to be superior to the peanut oil diet for evaluation of the anti-atherogenic effect of drugs, because the former caused milder vascular lesions than the latter. Using this rabbit model for atherosclerosis, the anti-atherogenic effect of clentiazem, a new calcium antagonist, was examined. Clentiazem at an oral dose of 30 mg/kg/day significantly reduced the size of atheromatous lesion in the aortic arch and thoracic aorta, and lowered the collagen content of the aortic intima and media, although it did not decrease serum lipid levels. On the other hand, clentiazem showed no clear effect on reducing the coronary atherosclerotic lesions. These results suggest that clentiazem may inhibit the progression of diet-induced aortic atherosclerosis without normalizing the serum lipid levels.

Ultrastructure of spontaneous glomerular lesions in syrian hamsters of APA strain.

Electron microscopic observations were carried out on the spontaneous glomerular lesions in male APA hamsters from 3 to 12 months of age. Until 6 months of age, focal expansion of mesangial region due to an increase of matrix material and mesangial cells was characteristic, and segmental thickening of capillary basement membrane and partial effacement of foot processes of podocytes were also sometimes observed. At 12 months of age, although all of these changes became more severe, the most prominent alteration was found in podocytes, which showed various degenerative changes. No deposition of amyloid fibrils was detected in any portion of the glomerulus.

Effects of ecabet sodium (TA-2711), a new antiulcer agent, on gastrointestinal mucosal prostanoid production and morphology in rats.

Effects of ecabet sodium (TA-2711), a locally acting antiulcer agent, on prostanoid production and the morphology of the rat gastrointestinal mucosa were studied in comparison with sucralfate. Ecabet, at therapeutic doses (25 and 100 mg/kg, p.o.), dose-dependently increased the gastric mucosal level of prostaglandin E2 (PGE2): sucralfate (100 mg/kg, p.o.) showed a tendency to increase the PGE2 level. In an ex vivo study, ecabet (25 and 100 mg/kg, p.o.) dose-dependently increased the capacity of the gastric mucosa to synthesize PGE2 and PGI2 without modifying tromboxane A2 (TXA2) synthesis, and the 100 mg/kg dose persisted for up to 3 h. Ecabet (400 mg/kg, p.o.) also significantly increased PGE2 synthesis and there was a tendency to increase PGI2 synthesis by the duodenal mucosa, without affecting TXA2 synthesis. PGE2 synthesis by the colonic mucosa was not affected, even at a high dose of ecabet (1000 mg/kg, p.o.). When the rat gastric mucosa was examined by light microscopy and scanning electron microscopy, ecabet (100 and 400 mg/kg, p.o.) had caused no morphological change to the gastric mucosa, while sucralfate (100 and 400 mg/kg, p.o.) produced apical rupture of the epithelial cells and subepithelial edema. The present study indicates that ecabet locally stimulates PGE2 and PGI2 production in the gastroduodenal mucosa and this effect is not attributable to a local irritant action accompanied by superficial epithelium damage.

Pathology of chemically induced chronic active hepatitis in mice.

Female Swiss mice were treated for 24 weeks, with 3-hydroxy-4-pyrone (Py) added to their powdered diet at 0.5% (wt/wt), and the effects of this agent on the liver were examined. Serum transaminases (especially GPT) rose continuously, while the GOT/GPT ratio remained at approximately 1.0 throughout the study period. The characteristic changes found from 8 weeks onward were piecemeal necrosis and bridging necrosis of the hepatocytes with dense lymphocytic infiltration. Proliferation of collagen fibers in the portal tracts and formation of narrow fibrous septa dividing the lobules into pseudolobules were also noted from 12 weeks onward. A large number of the infiltrating lymphocytes were identified as T cells by immunohistochemical and electron microscopic studies. These lymphocytes often surrounded or were closely attached to degenerating hepatocytes. Focal apoptosis and necrosis accompanied by a granulomatous reaction of the centrilobular hepatocytes were noted as early changes in the liver. Our findings indicate that the hepatic changes produced in mice by long-term Py administration have characteristics in common with those of human chronic active hepatitis. Immunological cytotoxic mechanisms, especially T cell-mediated ones, appear to play an essential role in the development of hepatic lesions in this murine model of chronic active hepatitis.

Rapid induction of atherosclerosis in rabbits.

Japanese white rabbits fed a restricted amount (100 g/head/day) of an atherogenic diet (AD) containing 0.2% cholesterol and 6% peanut oil showed mild and persistent hypercholesterolemia (338 +/- 79 mg/dl). They developed atherosclerotic lesions 4 weeks after deendothelialization of aorta carried out at the 4th week of AD-feeding. This rabbit model of atherosclerosis has such advantages as being able to be produced in a short period and having similar biochemical and pathological characteristics with those in human atherosclerosis.

REM sleep without muscle atonia (stage 1-REM) and its relation to delirious behavior during sleep in patients with degenerative diseases involving the brain stem.

Nocturnal sleep was examined in 12 patients with degenerative diseases involving the brain stem and in 2 patients with late cerebellar cortical atrophy (LCCA). A peculiar sleep state, characterized by the concomitant appearance of a low-voltage mixed frequency EEG, rapid eye movements (REMs) and tonic EMG in mental muscles, repeatedly appeared during nocturnal sleep in all of the 12 patients with degenerative diseases involving the brain stem and it was called stage 1-REM after Tachibana et al. In 8 of the 12 patients, delirious or oneiric behavior appeared during, or soon after, the episodes of stage 1-REM. Inner experiences reported by one of the subjects well corresponded to his behavior during the episode of stage 1-REM. Stage 1-REM was not observed during nocturnal sleep of the patients with LCCA. These results indicate that a degenerative lesion in the brain stem induced stage 1-REM and delirious behavior during nocturnal sleep through abolishing muscle atonia of REM sleep and causing dissociation of the functional components characterizing REM sleep.