Comparative clinical study of cefcapene pivoxil and cefteram pivoxil in chronic respiratory tract infections by a double-blind method.

In a double-blind study, the efficacy and safety of the novel cephem antibiotic cefcapene pivoxil (CFPN-PI; 450 mg/day) was compared with cefteram pivoxil (CFTM-PI; 600 mg/day) in 171 patients with chronic respiratory tract infections. There was no significant difference between the clinical efficacy of the two drugs (80.2% for CFPN-PI versus 78.9% for CFTM-PI). There was no significant difference in the rate of elimination of the causative bacteria (60.5% for CFPN-PI versus 65.9% for CFTM-PI). Side-effects were observed in 6.0% of patients treated with CFPN-PI compared with 6.4% of patients treated with CFTM-PI. There were no significant differences in incidence of abnormal laboratory findings following treatment with the two drugs (13.9% for each), and none of the side-effects was severe. We conclude that CFPN-PI (450 mg/day) was as effective and as well tolerated as CFTM-PI (600 mg/day) in the treatment of chronic respiratory tract infections.

A novel gene, GliH1, with homology to the Gli zinc finger domain not required for mouse development.

The Sonic hedgehog (Shh)-Gli signaling pathway regulates development of many organs, including teeth. We cloned a novel gene encoding a transcription factor that contains a zinc finger domain with highest homology to the Gli family of proteins (61-64% amino acid sequence identity) from incisor pulp. Consistent with this sequence conservation, gel mobility shift assays demonstrated that this new Gli homologous protein, GliH1, could bind previously characterized Gli DNA binding sites. Furthermore, transfection assays in dental pulp cells showed that whereas Gli1 induces a nearly 50-fold increase in activity of a luciferase reporter containing Gli DNA binding sites, coexpression of Gli1 with Gli3 and/or GliH1 results in inhibition of the Gli1-stimulated luciferase activity. In situ hybridization analysis of mouse embryos demonstrated that GliH1 expression is initiated later than the three Gli genes and has a more restricted expression pattern. GliH1 is first detected diffusely in the limb buds at 10.0 days post coitus and later is expressed in the branchial arches, craniofacial interface, ventral part of the tail, whisker follicles and hair, intervertebral discs, teeth, eyes and kidney. LacZ was inserted into the GliH1 allele in embryonic stem cells to produce mice lacking GliH1 function. While this produced indicator mice for GliH1-expression, analysis of mutant mice revealed no discernible phenotype or required function for GliH1. A search of the Celera Genomics and associated databases identified possible gene sequences encoding a zinc finger domain with approximately 90% homology to that of GliH1, indicating there is a family of GliH genes and raising the possibility of overlapping functions during development.

Seasonal variations in growth rate of phytoplankton in Hakata Bay, Japan.

Recently, water pollution with high concentrations of organic matter has occurred frequently in Hakata Bay. It is said that a high level of primary production provides much organic matter and affects water quality of the bay, and it is proved by the appearance of phytoplankton throughout the year. In this study, we simulated variations of phytoplankton population with a box-type model using monthly data in a long-term measurement and we analyzed the final growth rate changes of phytoplanktons that explain the conditions for its appearance. Consequently, we found that the final growth rate varies with pattern of half-year periodicity and water temperature and/or ambient nutrient controls the final growth rate to be low every January and July.

Effects of recombinant human erythropoietin therapy on blood coagulation and fibrinolysis system.

This study was designed to examine the effects of recombinant human erythropoietin (rHuEPO) therapy on blood coagulation and fibrinolysis in patients scheduled for elective heart surgery and undergoing preoperative autologous blood donation. Twenty-seven patients were studied, of whom 16 patients received rHuEPO (group E) and 11 patients no rHuEPO therapy (group N). The patients in group E were given 6000 units of rHuEPO intravenously every other day, three times a week, beginning from two weeks prior to the operation. In both groups, 400 ml of blood was collected preoperatively for predeposit once a week for two weeks, and the self-donated blood was returned to the patient intra- and postoperatively. Blood samples were drawn at the beginning of the study, immediately before the operation and two weeks after the operation. They were analyzed to assess blood coagulation, fibrinolysis, platelet function and vascular endothelial cell function, in order to examine the effects of the administration of rHuEPO. No significant difference was observed between the two groups in the degree of changes in these parameters following the operation. As enhancement of blood coagulability and fibrinolytic activity was evident postoperatively in both groups, changes in these parameters during the preoperative autologous blood donation period were also assessed excluding the postoperative data. Again, there was no significant intergroup difference in any of the markers evaluated. It was concluded that the administration of rHuEPO during preoperative autologous blood donation is unlikely to affect coagulation and fibrinolysis.

Up-regulation of splenic prohormone convertases PC1 and PC2 in diabetic rats.

Organisms respond to infection in a complex manner involving bidirectional interactions between the neuroendocrine and immune systems. Many of the bioactive endocrine/immune factors are synthesized in a precursor form and are expected to be activated by prohormone convertases (PCs). Since patients with both type 1 and type 2 diabetes have an increased incidence and severity of infections, we hypothesized that in a condition of hyperglycemia, these processing enzymes would be activated in an immune tissue, the spleen. To test this hypothesis, we treated rats with intraperitoneal streptozotocin (STZ) (50 mg/kg/day) daily for 5 days and measured splenic PC1 and PC2 mRNA by ribonuclease protection assay. We found that PC1 mRNA was increased 6.0+/-0.02-fold (P<0.05) and PC2 mRNA was increased 1.80+/-0.01-fold (P<0.005) in the spleen of rats that received STZ compared to rats that received vehicle. Western blot indicated that the 75-kDa form of PC1 was the only form of PC1 present in the spleen and that this form increased with STZ treatment. Immunohistochemistry revealed that PC1 was found in both the white pulp (T-lymphocytes) and red pulp (monocytes and macrophages) and that its increase in immunoreactivity occurred primarily in the white pulp. PC2 and pro-opiomelanocortin (POMC, a possible splenic substrate for PC1/PC2) immunoreactivity was found predominantly in the red pulp. STZ induced an increase in splenic PC1 and POMC, but not PC2 protein levels. We conclude that in the STZ model of diabetes, splenic PCs are induced, which could lead to an increased activation of many immune-derived hormones. We speculate that this up-regulation of prohormone converting enzymes may be related to the increased infections seen in patients with both type 1 and type 2 diabetes.

Antimutagens in gaiyou (Artemisia argyi levl. et vant.).

Antimutagens from gaiyou (Artemisia argyi Levl. et Vant., Compositae) were examined. The methanol extract prepared from aerial parts of this plant strongly reduced the mutagenicity of 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), when Salmonella typhimurium TA98 was used in the presence of the rat liver microsomal fraction. The antimutagens were purified chromatographically while monitoring the antimutagenic activity against Trp-P-2 with a modified Ames test employing a plate method. This purification resulted in the isolation of four strong antimutagens, 5,7-dihydroxy-6,3',4'-trimethoxyflavone (eupatilin), 5, 7,4'-trihydroxy-6,3'-dimethoxyflavone (jaceosidin), 5,7, 4'-trihydroxyflavone (apigenin) and 5,7, 4'-trihydroxy-3'-methoxyflavone (chrysoeriol) from the methanol extract. These antimutagenic flavones exhibited strong antimutagenic activity against not only Trp-P-2 but also against other heterocyclic amines, such as 3-amino-1,4-dimethyl-5H-pyrido[4, 3-b]indole (Trp-P-1), 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3, 8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeA(alpha)C) in S. typhimurium TA98. In contrast, they did not exhibit antimutagenic activity against benzo[a]pyrene (B[a]P), 4-nitroquinoline-1-oxide (4-NQO), 2-aminofluorene (2-AF), 2-nitrofluorene (2-NF) or furylfuramide (AF-2) in S. typhimurium TA98, or B[a]P, 4-NQO, 2-NF, AF-2, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or sodium azide (SA) in Salmonella typhimurium TA100, whereas they decreased the mutagenicity caused by aflatoxin B(1) (AFB(1)) and 2-aminoanthracene (2-AA) in both of these tester strains. Regarding the structure-activity relationship, the tested flavones had distinct differences in the intensities of their antimutagenic activities according to the differences of their substitution patterns. Namely, the intensity of antimutagenic activities against Trp-P-2 decreased in the order of: 5,7,3',4'-tetrasubstituted flavones (IC(50): <0.1 mmol/plate), 5,7,4'-trisubstituted flavones (IC(50): 0.120-0.260 mmol/plate), 5,6,7,3',4'-pentasubstituted flavones (IC(50): 0.440-0. 772 mmol/plate). The four isolated flavones were also studied regarding their antimutagenic mechanisms with preincubation methods of the modified Ames test and emission spectroscopic analysis. The results suggested that all isolated flavones were desmutagens which directly inactivated Trp-P-2 or inhibited its metabolic activation.

Failure of preventive effects of 2-deoxy-D-glucose on ischemia-induced gerbil hippocampal neuronal damage by induced hyperthermia.

Post-ischemic administration of 2-deoxy-D-glucose (2-DG), a glucose antimetabolite, markedly reduces the occurrence of ischemia-induced delayed neuronal death (DND) in the gerbil hippocampus. This means that the reduction of energy dependent metabolism after ischemia prevents ischemia-induced damages of hippocampal neurons. In the present study, we demonstrated hyperthermia during ischemia fails to preserve neurons in hippocampal CA1 of 2-DG treated gerbil following transient forebrain ischemia.

A placebo-controlled, single-blind study to determine the appropriate alendronate dosage in postmenopausal Japanese patients with osteoporosis. The Alendronate Research Group.

Alendronate (4-amino-1-hydroxybutylidene-1,1-bisphosphonate) is a potent inhibitor of bone resorption. The efficacy and safety of 36 weeks of treatment with alendronate were evaluated in Japanese women with osteoporosis, osteoporotic osteopenia or artificial menopause. The bone mineral density (BMD) of the lumbar vertebrae, markers of bone and calcium metabolism and clinical symptoms were monitored. A total of 113 randomly selected patients with osteoporosis or osteopenia were enrolled in the study, of whom 12 were excluded from the analyses because of lack of data. As a result, 101 patients were evaluated for the safety of the drug. Since eight patients were excluded from the efficacy analysis, 93 were evaluated. The incidence of adverse effects in the placebo (P), alendronate 2.5 mg/day (L) and alendronate 10 mg/day (H) groups increased with increasing dose of alendronate, being 6.1, 14.3 and 18.2%, respectively. The most common adverse effects were gastrointestinal symptoms, none of which was serious. Lumbar BMD increased after 36 weeks of drug administration to 5.21%, 5.64% and -0.90% in the L, H and P groups, respectively (P < 0.001, L vs. P and H vs. P). Serum alkaline phosphatase activity, serum osteocalcin and urinary deoxypyridinoline excretion were significantly decreased in a dose-related manner. Serum calcium and phosphorus were also significantly decreased after alendronate administration. Serum intact PTH was transiently increased. The present results indicate that alendronate effectively decreases bone turnover in a dose-related manner and increases lumbar BMD at a dosage of 2.5 mg/day, the lowest dose used in this study, in Japanese patients with osteoporosis.

Photochemically induced endothelial injury in the mouse as a screening model for inhibitors of vascular intimal thickening.

We have established a mouse model of intimal thickening and assessed its suitability for experimental studies of intimal thickening. Neointimal formation was observed after endothelial injury by photochemical reaction between transluminal green light and systemically administered rose Bengal, which represents a nonmechanical approach to vessel wall denudation. Intimal thickening began 7 days after endothelial injury, reached a maximum after 21 days, and then remained unchanged for as long as 42 days. Furthermore, as a consequence of neointimal proliferation, the luminal area gradually decreased. The cells in the neointimal layer were identified as smooth muscle cells by immunohistochemical staining with an alpha-actin-specific antibody. Extracellular matrix deposition in the neointima was markedly increased beyond 14 days after injury. Smooth muscle cell proliferation, as measured by pulse labeling of 5-bromo-2'-deoxyuridine, was identified initially in the media 2 days after vessel wall denudation, with the proliferative activity's shifting almost exclusively to the neointima within 7 days. Endothelial regeneration, as indicated by Evans blue staining, was complete within 21 days after injury. To assess the suitability of this model for experimental studies on intimal thickening, the effect of tranilast, an antiallergy drug with a broad spectrum of pharmacological actions on intimal thickening, was investigated. Tranilast (100 mg x kg(-1) x d(-1) p.o.) significantly (P<0.05) reduced smooth muscle cell proliferation in the neointima and media 7 days after injury and neointimal formation 21 days after injury in treated mice compared with vehicle-treated mice. This simple experimental mouse model is suitable for studying factors promoting or inhibiting intimal thickening after endothelial injury and for developing therapeutic strategies against intimal thickening.

Effects of astemizole on ventricular activation delay and RT intervals in a canine myocardial infarction model.

In order to clarify the arrhythmogenic effects of nonsedating antihistamines, we examined the effects of astemizole, a nonsedating antihistamine, on ventricular activation and RT intervals in a canine myocardial infarction model. Myocardial infarction was produced by two-stage ligation of the left anterior descending coronary artery in dogs. Seven days after ligation, bipolar electrodes were sutured on the ventricular surface of the infarcted and normal zones to apply an electrical stimulation or record the ventricular activation. An electrical stimulation with coupling intervals between 300 and 140 ms was applied on the ventricular surface of the normal zone during atrial pacing, and the ventricular activation delay was measured. The effect of astemizole on the RT interval was also determined during atrial pacing, sinus rhythm or after premature stimulation. The ventricular activation delay increased after astemizole at doses of 0.3 to 3 mg/kg in the infarcted and at 3 mg/kg in the normal zones, and the effect of astemizole was greater in the infarcted zone. Astemizole increased the RT interval in the normal zone to a greater extent at a long coupling interval. The increase in the RT interval was greater in the infarcted zone compared with that in the normal zone. In conclusion, astemizole increased the activation delay in the infarcted zone, probably through prolongation of the repolarization time, and its effect on the activation delay may be arrhythmogenic.

Inhibitory effect of a novel orally active GP IIb/IIIa inhibitor, SC-54684A on intimal thickening in the guinea pig femoral artery.

The inhibitory effect of a novel orally active platelet membrane glycoprotein receptor complex IIb/IIIa (GP IIb/IIIa) inhibitor, SC-54684A is studied on intimal thickening in the guinea pig femoral artery. A segment of the femoral artery was occluded by a platelet-rich thrombus induced by photochemical reaction between systemically administered Rose Bengal and transluminal green light which causes endothelial injury followed by platelet adhesion and aggregation at the site of photochemical reaction. Three weeks after successful thrombolysis by tissue-type plasminogen activator, intimal thickening occurred at the irradiated site. SC-54684A (30 mg/kg), administered 2 h before photochemical reaction, significantly (P < 0.05) prolonged the time to occlusion of the femoral artery; in this respect aspirin (100 mg/kg) was ineffective. A combination of SC-54684A and recombinant tissue-type plasminogen activator (rt-PA) increased the frequency of reopening and significantly (P < 0.05) prolonged the duration of reflow compared with rt-PA alone. Further, SC-54684A administered orally twice a day for 3 weeks significantly (P < 0.05) inhibited intimal thickening but aspirin, administered orally once a day for 3 weeks, was ineffective. Scanning electron microscopy revealed extensive platelet adhesion and aggregation on the denuded vessel walls in the untreated group 24 h after successful thrombolysis. In separate experiments, SC-54684A markedly inhibited platelet aggregation ex-vivo. Inhibition of platelet adhesion and aggregation at the site of endothelial injury by SC-54684A (via GPIIb/IIIa inhibition) may account for its inhibitory effect on intimal thickening.

Increase in shedding of intercellular adhesion molecule-1 in human malignant melanoma cell lines treated with hyperthermia in vitro.

Human malignant melanoma cell lines were found to increase shedding of soluble intercellular adhesion molecule-1 (sICAM-1) into the culture medium when the cells were treated with hyperthermia at 41-43 degrees C for 3-6 hr in vitro. The content of ICAM-1 in the cell lysate was also found to be increased after hyperthermia. The increased rate of ICAM-1 concentration in the cells was at maximum when they were incubated at 41 degrees C for 3 hr. Also, the melanoma cell lines heat-treated at 41 degrees C showed more intense immunofluorescence in the ICAM-1 expression on the cell surface. It remains to be investigated further whether the effects of hyperthermia on the ICAM-1 expression in melanoma cells is to augment membrane ICAM-1 expression, which in turn leads to shedding of soluble ICAM-1 or only to acceleration of shedding of sICAM-1 by unknown mechanisms.

A chemiluminescent detection of superoxide radical produced by adherent leucocytes to the subendothelium following thrombolysis: studies with a photochemically induced thrombosis model in the guinea pig femoral artery.

Reocclusion following thrombolysis is a major limitation of thrombolytic therapy with recombinant tissue-type plasminogen activator (rt-PA) because denuded vessel wall exposed to blood following thrombolysis is a favourable surface for platelet and leucocyte deposition. We have applied a chemiluminescence technique to detect superoxide radical (0(-2)) produced by leucocytes adherent to the femoral artery 24 h after photochemically induced thrombogenesis in the guinea pig in vivo and subsequent thrombolysis by rt-PA. Intravenous administration of MCLA, a specific chemiluminescence reagent for detecting O(-2), markedly increased photon emission. the photon emission was markedly potentiated by phorbol myristate acetate and was suppressed by superoxide dismutase. Reocclusion 24 h after rt-PA induced thrombolysis was observed in 10 of 16 animals. Histological observations revealed extensive polymorphonuclear leucocytes adherent to the vessel wall at the site of thrombogenesis and thrombolysis. A higher level of 0(-2) could be detected from the arteries in which thrombolysis was induced compared with those without thrombolysis. Further, the level 0(-2) detected was greater in reoccluded arteries compared with those in which reflow was established. These observations suggest that 0(-2) is produced by adherent leucocytes at the site of thrombolysis and that leucocytes are involved in reocclusion after thrombolysis.

Measurement of 5-fluorouracil in scalp hair: a possible index of patient compliance with oral adjuvant chemotherapy.

OBJECTIVE: Little is known about patient compliance with oral adjuvant chemotherapy. It is estimated to be poor especially in Japan, where it is still unusual for patients to be directly informed of their diagnosis of malignancy. 5-Fluorouracil (5-FU) was measured in hair samples to assess patient exposure to 5-FU, and its potential usefulness is discussed as an index of compliance with postoperative adjuvant chemotherapy. METHODS: Hair samples obtained from 55 patients, who had received oral 5-FU (total dose 27-41 g) as postoperative adjuvant chemotherapy over a 6-month period, were used for the analysis of 5-FU. The drug was extracted from the hair using ethyl acetate, and its fluorescence derivatization was employed for measurement with HPLC. The detection limit of 5-FU in hair was 0.01 ppm. RESULTS: In 22 out of 55 samples 5-FU content was under the detection limit, whereas in the remaining 33 samples the drug was detected in a range of 0.006-2.125 ng per hair strand; in addition, drug content showed a lognormal distribution. 5-FU was detected in the hair collected from those patients who were possibly compliant with the postoperative oral adjuvant chemotherapy. CONCLUSION: As many as 40% of the patients analysed were supposed to be much less compliant. Even in the possibly compliant patients, the degree of compliance with the therapy varied according to a log-normal distribution.

Effect of dietary docosahexaenoic acid in the rat middle cerebral artery thrombosis model.

We investigated the effect of dietary docosahexaenoic acid (DHA) supplementation on the thrombolytic efficacy of recombinant tissue-type plasminogen activator (rt-PA), platelet aggregability, serum cholesterol and phospholipids. Male Wistar rats (6 weeks old) received dietary DHA supplementation (300 mg/kg per day) for 8 weeks. The rat middle cerebral artery (MCA) was occluded by a thrombus induced by photochemical reaction between rose bengal and green light which cause endothelial damage followed by platelet adhesion, aggregation and formation of a platelet and fibrin-rich thrombus at the site of photochemical reaction. The MCA blood flow was monitored using a laser Doppler flowmeter. rt-PA was administered 30 min after the middle cerebral artery had been occluded by a thrombus. This regimen produced a significant (P < 0.05) decrease in serum free-cholesterol and phospholipids levels, inhibited platelet aggregation ex-vivo induced by collagen in whole blood (P < 0.05), reduced thromboxane (TX) B2 formation (P < 0.01) in whole blood and prolonged the time for thrombotic MCA occlusion (P < 0.01) as compared with values obtained from animals on standard diet. Further, dietary DHA enhanced thrombolytic efficacy of rt-PA and reduced the size of ischaemic cerebral lesions. Our findings suggest that dietary DHA produces antithrombotic effects via metabolic conversion to non-atherogenic and non-platelet stimulant metabolites.

Enhancement of thrombolytic efficacy of tissue-type plasminogen activator by adjuvants in the guinea pig thrombosis model.

Reocclusion following thrombolysis is a major limitation of thrombolytic therapy with recombinant tissue-type plasminogen activator (rt-PA). We investigated the effects of vapiprost ((1R-(1 alpha(Z),2 beta,3 beta,5 alpha))-7-(5-((1,1'-biphenyl)-4-yl-methoxy)- 3-hydroxy-2-(1-piperidinyl)cyclopentyl)-4-heptenoic acid, a thromboxane A2 receptor antagonist); argatroban ((2R,4R)-4-methyl-1-[N2-(3-methyl-1,2,3,4-tetrahydro-8-quinolinyl)sulfon yl] - L-arginyl)]-2-piperidine-carboxylic acid, a specific thrombin inhibitor) and MK-886 (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2- dimethylpropanoic acid, a specific leukotriene biosynthesis inhibitor) on the thrombolytic efficacy of rt-PA. The guinea pig femoral artery was thrombotically occluded by photochemical reaction between rose bengal and green light. Thirty min after the occlusion, rt-PA was administered and the time (T1) for reopening of the vessel and the frequency of reocclusion (Fro) 24 h after thrombolysis were monitored. With rt-PA alone, T1 was 28 +/- 7 min (n = 10) and Fro was 70%. T1 was reduced to 9 and 20 min by a combination of rt-PA with vapiprost and argatroban respectively. Fro was reduced by all three adjuvants. Histological observations revealed extensive adherence of polymorphonuclear leucocytes to the damaged endothelium at the site of thrombolysis. It is concluded that thromboxane A2, thrombin and leucocytes are involved in reocclusion after thrombolysis.

Effect of dietary docosahexaenoic Acid supplementation on platelet function: studies in the rat femoral artery thrombosis model.

We have developed a model whereby the femoral artery in an experimental animal can be occluded by a photochemical reaction between rose bengal and green light which causes endothelial injury followed by platelet adhesion, aggregation and formation of a platelet rich thrombus at the site of the reaction. Using this model, we investigated the effect of dietary docosahexaenoic acid (DHA) supplementation on platelet aggregation and on serum cholesterol and lipids. Male Wistar rats (iweeks-old) received dietary DHA supplementation (300 mg/kg/d) for 8 weeks. This regimen produced a significant (p < 0.05) decrease in serum free-cholesterol and phospholipids levels, inhibited platelet aggregation ex vivo induced by collagen in whole blood (p < 0.05) but not in platelet rich plasma, reduced thromboxane B(2) formation (p<0.01) induced by collagen in washed platelets and prolonged the time for thrombotic arterial occlusion (p<0.01) as compared with values obtained in animals on standard diet. In conclusion, dietary DHA produces antithrombotic effects such as reduction in platelet aggregability and lowering of plasma cholesterol. Whole blood where red and white blood cells can exert their influences on platelet function is a more sensitive and physiological medium than platelet rich plasma for studying the effects of antithrombotic treatments on platelet aggregability ex vivo.

[New animal models for drug discovery research: focus on cardiovascular diseases].

Development of novel drugs relies on research to discover new drugs. Testing and evaluating new drugs on human subjects are usually impossible because of ethical concerns. Therefore, for drug discovery research, it is essential to establish animal models of human diseases. Numerous animal models have been developed and used in drug discovery and evaluation studies. Such animal models have had important roles in developing new drugs as well as understanding the etiologies of diseases. In the field of cardiovascular drugs, several interesting animal models are currently in use. These include, transgenic mice carrying both human renin and human angiotensinogen genes, Watanabe's heritable hyperlipidemic rabbits, rats with pulmonary hypertension induced by monocrotaline, myocardial infarction model, cardial hypertrophy model and photochemical thrombosis models. It is envisaged that for drug discovery and development, the search for more physiological animal models will continue in the future.

Effect of a Ca2+ entry blocker, nilvadipine, on hearing disturbances and equilibrium dysfunction caused by microcirculatory disorders of the rat inner ear.

We evaluated the effects of Ca2+ entry blockers, nilvadipine and flunarizine, on microcirculatory disorders of the inner ear and on blood flow in the inner ear of rats. Under sodium pentobarbital anesthesia, the middle ear was opened by a ventrolateral approach. A green light (wave length 540 nm) was applied to the cochlea or the vestibule to induce a hearing disturbance or equilibrium dysfunction as a result of inner ear microcirculatory disorders, while rose bengal solution was infused intravenously. In a hearing disturbance model, a compound cochlear nerve action potential was recorded by electrocochleography every minute after the beginning of illumination. The sound stimulus was an 8 kHz sine wave 100 dB normal hearing level. The action potential was calculated 128 times. The action potential disappeared about 12 min after the beginning of illumination. In another model of equilibrium dysfunction, the photoillumination was applied for 40 min under the infusion of rose bengal. The behavior of rats was observed in the swimming test and nystagmus was recorded 24 h after the completion of photoillumination. In a separate experiment, blood flow in the inner ear was measured with a laser Doppler flowmeter under sodium pentobarbital anesthesia. In this study, both nilvadipine and flunarizine prolonged the time required for complete suppression of the action potential, prevented equilibrium dysfunction in the swimming test and reduced the occurrence of nystagmus. Flunarizine significantly increased inner ear blood flow and nilvadipine failed to decrease blood flow in the inner ear, despite a reduced systemic blood pressure. In conclusion, Ca2+ entry blockers may prevent microcirculatory disorders of the inner ear in rats.

Evaluation of the combination of a tissue-type plasminogen activator, SUN9216, and a thromboxane A2 receptor antagonist, vapiprost, in a rat middle cerebral artery thrombosis model.

BACKGROUND AND PURPOSE: We aimed to evaluate a modified tissue-type plasminogen activator, SUN9216, and the combination of SUN9216 and a thromboxane A2 receptor antagonist, vapiprost, in a rat middle cerebral artery thrombosis model. METHODS: Under anesthesia, the left middle cerebral artery was observed under an operation microscope without cutting the dura mater via a subtemporal craniotomy. Photoillumination (wave length, 540 nm) was applied to the middle cerebral artery, and then rose bengal (20 mg/kg) was administered intravenously. The reopening of the middle cerebral artery by SUN9216, injected 30 minutes after middle cerebral artery occlusion, was observed under an operation microscope for a 60-minute observation period. Twenty-four hours after the operation, sections of the cerebrum were stained with triphenyltetrazolium chloride, and the area of cerebral infarction was analyzed by a computer. RESULTS: The combination of SUN9216 and vapiprost caused reopening of the middle cerebral artery in 58.8% of the rats, which was a greater percentage than that achieved with SUN9216 alone (31.6%). In contrast, saline did not cause reopening of the middle cerebral artery during the 60-minute observation period. The area of cerebral infarction in rats reperfused with SUN9216 was significantly reduced compared with that in the control group. The infarction area in rats treated with the combination of SUN9216 and vapiprost was reduced compared with that in rats treated with SUN9216 alone; this was the case whether or not the occlusion was reperfused. There was a significant correlation between the time of reopening of the middle cerebral artery and area of cerebral infarction. CONCLUSIONS: A single injection of SUN9216 was effective in recanalizing the vessel and reducing the area of cerebral infarction.