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The role of xanthine oxidase in paraquat toxicity was investigated using cultured bovine pulmonary artery endothelial cells. Exposure to paraquat 0.1 mM was done for 24 hr with or without tungsten pretreatment and in the presence or absence of xanthine oxidase inhibitors. Exposure to paraquat significantly increased O2- production and relative xanthine oxidase activity (xanthine oxidase activity divided by total xanthine dehydrogenase plus xanthine oxidase) while depressing cell growth. In contrast, tungsten and allopurinol inhibited the increase of xanthine oxidase activity and decreased O2- release. Cell injury was assessed by leakage of lactate dehydrogenase and by fluorescein diacetate staining; it was found that oxidase inhibitors (both allopurinol and tungsten) reduced paraquat cytotoxicity. Thus the toxicity of paraquat was at least partly due to intracellular O2- production mediated by xanthine oxidase and the subsequent formation of other free radicals.

Treatment of L929 cells with tumour necrosis factor-alpha (TNF-alpha) plus actinomycin D induced DNA damage (indicated by the appearance of a sub-G1 peak due to extracellular leakage of low molecular weight DNA following DNA fragmentation) before significant cell lysis occurred. The DNA damage occurred in parallel with a decrease of the intracellular total glutathione content and an increase of intracellular reactive oxygen intermediates (ROI), as indicated by increased dihydrorhodamine 123 oxidation. Because the inhibition of mitochondrial respiration suppressed the increase of dihydrorhodamine 123 oxidation and DNA damage as well as the decrease in the total glutathione content, it was suggested that increased mitochondrial formation of ROI was responsible for DNA damage after TNF treatment. Deferoxamine (a ferric iron chelator) and dithiothreitol (a sulfhydryl reagent) both prevented DNA damage and cell killing, indicate that hydroxyl radicals generated from O2- and H2O2 produced by the mitochondria in a process catalysed by iron contributed to DNA damage and that this pathway may be involved in TNF-alpha-induced cytotoxicity. An inhibitor of poly(ADP)-ribose polymerase (3-aminobenzamide), worsened DNA damage, but was protective against cell lysis, suggesting that DNA repair subsequent to injury was more important than DNA damage per se in development of TNF-alpha cytotoxicity.

Washed human platelets were incubated under hypoxic and normoxic conditions to investigate the influence of oxygen deprivation on energy metabolism and aggregation. The maximum aggregation rate, oxygen consumption, lactate production, and adenine nucleotide content were measured. Hypoxic incubation decreased the oxygen burst and maximum aggregation induced by thrombin, and accelerated anaerobic glycolysis. ATP and ADP levels were preserved, but the hypoxanthine level increased in the incubation medium and the platelet AMP level and adenylate energy charge decreased compared with normoxic incubation. Thus, anaerobic glycolysis failed to compensate for impaired oxidative phosphorylation during hypoxic incubation, suggesting that oxidative energy is essential for full platelet function.

The effects of diabetes on fatty acid oxidation in platelets was determined in streptozotocin-induced diabetic rats. In platelets isolated from diabetes, the oxygen consumption which reflects mainly the degree of fatty acid oxidation and ADP- and thrombin-induced aggregation were increased as compared to non-diabetic rat platelets. Carnitine palmitoyltransferase I (CPT I), the rate-limiting enzyme for fatty acid oxidation, in platelets obtained from diabetes showed a higher Vmax for palmitoyl-CoA and an increased I50 (concentration giving 50% inhibition of CPT I activity) for malonyl-CoA inhibition. These changes observed in fatty acid oxidation in platelets derived from diabetes returned to the control levels after insulin therapy. When platelets were stimulated with thrombin, platelet CPT I activity increased over time in both diabetic and non-diabetic rats. From these findings, fatty acid oxidation in platelets, as in the liver, is likely to be regulated by insulin and both increased CPT I activity and decreased sensitivity to malonyl-CoA inhibition are attributable to enhanced platelet fatty acid oxidation in diabetic rats.

We observed significant changes in serial aminograms during direct hemoperfusion (DHP) in clinical cases and decided to evaluate the effects of the procedure on amino acid profiles. DHP at 100 ml/min was carried out for 6 h in patients with drug intoxications. Pre- and post-column aminograms were obtained 0.5, 1, 2, 3, 4, 5 and 6 h from the beginning of DHP. Nonmolecular-weight-dependent adsorption of some amino acids to the column was observed. Methionine (MET), isoleucine (ILE), leucine (LEU), tyrosine (TYR), phenylalanine (PHE) and arginine (ARG) had high adsorption rates. Blood concentrations of TRY, PHE, TRY and histidine (HIS) were significantly reduced. Although both branched-chain amino acids (BCAA) and aromatic amino acids (AAA) were initially adsorbed by the column, AAA recovery was slow.

We investigated the role of energy supplied by long-chain fatty acid oxidation in rat platelet function. Inhibition of the mitochondrial uptake of long-chain fatty acids was achieved by treating rats with 2-tetradecylglycidic acid (TDGA), a potent inhibitor of the overt form of carnitine palmitoyltransferase (CPT-I). The maximum aggregation rate (MAR), CPT-I activity, lactate production, oxygen consumption and adenine nucleotide content of isolated rat platelets were then studied in vitro. 4 h after the in vivo administration of TDGA, the CPT-I activity in saponin-permeabilized platelets was nearly completely inhibited along with a significant reduction in the MAR induced by ADP, thrombin and ionophore A23187. The ATP level, adenylate energy charge (ATP + 1/2 ADP)/(ATP + ADP + AMP) and ATP/ADP ratio in the platelet cytoplasmic pool were also reduced. Platelets from TDGA-treated rats showed lower oxygen consumption rates in both the basal respiratory and oxygen burst states. These results indicate that mitochondrial long-chain fatty acid oxidation coupled to oxidative phosphorylation is an important energy source in rat platelets and is probably involved in the maintenance of platelet function. Enhanced in vitro lactate production in platelets from TDGA-treated rats may have resulted from a compensatory increase in glycolysis which only partly compensated for impaired long-chain fatty acid oxidation.

Hepatic mitochondrial functions related to fatty acid metabolism, including the respiratory control ratio, fatty acid oxidative capacity and carnitine palmitoyltransferase I activity, were studied in vitro with mitochondria isolated from rats treated with thioacetamide for up to 12 wk. The levels of ketone bodies, carnitine, carnitine esters and malonyl-coenzyme A were also determined in liver extracts. Polarography of mitochondrial respiration from succinate or glutamate plus malate showed a lower respiratory control ratio in thioacetamide-treated rats, whereas uncoupled oxygen consumption was not altered. This suggests that the mitochondrial respiratory chain capacity remained intact in the thioacetamide-treated rats. The oxygen consumption associated with palmitoyl-coenzyme A and palmitoyl-L-carnitine oxidation by isolated liver mitochondria was increased by thioacetamide treatment on both a per-mitochondrial protein and a per-total liver basis. The carnitine palmitoyl-transferase I activity; the tissue levels of ketone bodies, carnitine and carnitine esters; and the beta-hydroxybutyrate/acetoacetate ratio were all higher in the livers of thioacetamide-treated animals than in control livers, whereas the hepatic malonyl-coenzyme A level was decreased by thioacetamide. These results indicate the increased diversion of cytosolic long-chain acyl-coenzyme As into the mitochondria for beta-oxidation rather than their esterification and use in lipogenesis. These intrahepatic metabolic changes induced by chronic thioacetamide administration may reflect the whole-body catabolic state and can be seen as adaptive for maintaining energy homeostasis under conditions of impaired glucose tolerance.

Using microdialysis, we studied brain extracellular fluid (ECF) amino acid metabolism in rats with experimentally induced hyperammonemia and regional elevation of brain ECF ammonia levels. The total brain ECF amino acid level was increased by an elevation of the blood ammonia level. Hyperammonemia elevated brain ECF aromatic amino acids and reduced arterial blood branched chain amino acids. When rats with hyperammonemia were intravenously administered norleucine, the brain ECF norleucine level rose markedly, suggesting increased permeability of the blood-brain barrier. When rats with hyperammonemia were infused with a branched chain amino acid-rich preparation, the elevated brain ECF aromatic amino acids level was not suppressed. Following local intracerebral ammonia infusion, only glutamate levels showed a marked elevation. These results suggest that impairment of the blood-brain barrier related to hyperammonemia increases the inflow of low molecular weight substances including amino acids. Furthermore, the ammonia-induced increase of glutamate in the cerebral ECF suggests that high ammonia levels may increase the excitability of the brain. Thus, ammonia may serve as a key factor in the onset of hepatic encephalopathy.

The role of xanthine oxidase in the mechanism of paraquat toxicity was assessed by in vitro and in vivo experiments. Paraquat stimulated the reduction of cytochrome c by xanthine-xanthine oxidase system in vitro. Paraquat, when added in vitro, stimulated hypoxanthine-dependent superoxide production in the cytosol of rat lung. Tungsten-feeding inhibits xanthine oxidase activity in a variety of tissues in experimental animals. Its therapeutic effect on paraquat intoxication was studied in this paper. In rats fed a tungsten-enriched diet for 5 weeks prior to intraperitoneal injection of 50 mg/kg paraquat dichloride, the mortality decreased significantly compared with rats fed a standard diet. Pretreatment with oxypurinol (1000 mg/kg, s.c.) also ameliorated the paraquat toxicity in rats. We conclude that xanthine oxidase plays an important role in paraquat toxicity and that xanthine oxidase inhibitors may become antidotes for paraquat intoxication.

Saponin-permeabilization (30 micrograms/ml) of the platelet plasma membrane, which enables access of added compounds to mitochondrial overt carnitine palmitoyltransferase (CPT I), was applied to allow the rapid determination of CPT I activity in situ. The effects of diabetes and short-term incubation with insulin in vitro on the kinetic parameters and malonyl-CoA sensitivity of CPT I were also studied in rat platelets. CPT I exhibited ordinary Michaelis-Menten kinetics when platelets were incubated with palmitoyl-CoA. Malonyl-CoA showed an I50 (concentration giving 50% inhibition of CPT activity) of 0.92 +/- 0.11 microM in permeabilized platelets. Platelets obtained from diabetic rats (induced by streptozotocin injection) exhibited an increased Vmax and I50 for malonyl-CoA, and an unaltered Km for palmitoyl-CoA. In contrast, preincubation of platelets prepared from both fed control rats and diabetic rats with insulin (100 and 150 microU/ml) led to a decrease in enzyme activity when assayed with 75 microM palmitoyl-CoA and 0.5 mM L-carnitine as substrates. These in vivo and in vitro results suggested that insulin directly modulated rat platelet CPT I activity, as it does in the liver.

The purpose of this study was to clarify the involvement of adenine nucleotide metabolism and substrates of the xanthine-xanthine oxidase system as the source of oxygen free radicals in a rat model of restrained water immersion stress ulceration. The gastric mucosal concentrations of adenine-5'-triphosphate (ATP), adenine-5'-diphosphate (ADP), adenine-5'-monophosphate (AMP) and thiobarbituric-acid (TBA)-reactant substances were measured after 4, 8 and 12 h restrained water immersion stress. The gastric mucosal concentrations of the nucleoside adenosine, the purine bases xanthine and hypoxanthine, and the final metabolic product uric acid, were measured after 4 h of restrained water immersion stress. The concentrations of ATP diminished significantly after 4, 8 and 12 h of restrained water immersion stress. However, the observed stress-induced changes in ADP were not significant. AMP concentrations increased significantly after 4, 8 and 12 h of stress. The adenylate pool (ATP + ADP + AMP) dropped significantly from the prestress value after 4, 8 and 12 h of stress, and the concomitant energy charge (EC = ATP + 0.5 ADP/ATP + ADP + AMP) decreased significantly after 4 and 8 h of stress compared with the prestress value. Gastric mucosal concentrations of TBA-reactant substances displayed a significant increase after 4 h of stress, and remained unchanged after 8 and 12 h of stress from the level after 4 h. Four hours of restrained water immersion stress induced an increase in adenosine and uric acid concentrations and a decrease in the hypoxanthine concentration of the gastric mucosa.(ABSTRACT TRUNCATED AT 250 WORDS)

Cecal ligation and puncture (CLP) were performed in rats. After 4 hr (early sepsis) and 16 hr (late sepsis), platelet morphology and function were studied. At 16 hr, platelet counts for the CLP group were significantly lower than for the sham-operated control group. Low maximum aggregation rates (MAR) and decreased platelet counts were elicited in platelet-rich plasma with 4 M ADP and 2 micrograms/ml collagen. However, with platelet counts equalized, MAR for the CLP group increased significantly, especially after 16 hr. The platelet-large cell rate and platelet distribution width decreased temporarily at 4 hr, then rose significantly at 16 hr. No significant changes were observed in the mean platelet volume after 4 hr, but there were significant increases after 16 hr. Total adenine nucleotide (TAN) levels within the platelets rose significantly in the CLP group, suggesting the appearance during the late sepsis of large, heavy platelets or adenine nucleotide-rich platelets. The platelet adenylate pool was divided into granular and cytoplasmic fractions, respectively characterized by ADP and ATP increases. However, no septicemia-related differences were noted in the degree of binding between goat antirat fibrinogen and platelet surface glycoprotein IIb/IIIa complex. Internal environment changes in the platelets indicated that during septicemia hyperfunctional or hypersensitive platelets with a latent capacity for active aggregation and release appeared in the circulation. Hypercoagulability in septicemia involves activation of coagulation factors, stimulation of the coagulation cascade, volume changes accompanying increased platelet TAN content, and changes in AN distribution in the two pools. These findings significantly increase our understanding of the transition from the prethrombotic state to thrombosis in septicemia.

Effects of urinary trypsin inhibitor (UTI) on the number, morphology and function of platelets under septic state were studied in rat models of cecal ligation and puncture (CLP). At formation of CLP, 5,000 U/kg/h of UTI was serially administered intraperitoneally and blood was sampled after 16 hours. Comparative study among sham-operation group, CLP group, and CLP + UTI group revealed: 1) inhibition of the platelets of platelet counts and appearance of large-sized, active platelets by UTI in the CLP + UTI group, 2) increase of platelet maximum aggregation rate (MAR) by ADP and increase of collagen in the CLP group, while inhibition in the CLP + UTI group and 3) by HPLC evaluation of adenine nucleotide in the platelet, increased levels of total ATP and ADP in the CLP group, particularly, increases of ATP in the metabolic pool and ADP in the granular pool. CLP + UTI group did not show these changes in the adenylate pool. UTI was thus considered to stabilize the platelet cycle in sepsis. Platelets under septic state might be hyperactive, and thrombosis is easy to occur. UTI administration might work for maintaining constancy of the platelet internal environment and improve septic state because adenine nucleotide level in the platelet did not change in the CLP + UTI group through changed in the CLP group.

A 69-year-old woman with typical thrombotic thrombocytopenic purpura is reported, who was treated successfully by plasma exchange. An initial chromosomal analysis of peripheral blood lymphocytes revealed abnormal patterns of 45, XO/46, XX/47, XXX; a second analysis showed 45, XO/46, XX/47, XXX/47, XX + 21, and a third, performed in remission, showed a normal pattern. Skin fibroblasts showed a normal pattern even during the acute disease. The association of transient chromosomal aberrations and thrombotic thrombocytopenic purpura might be related to a viral infection.

We reported a case of 69-year-old female presenting with clinically typical TTP which was treated with Ticlopidine and plasma exchanges four times in total and resulted in successful improvement of her clinical state. The first chromosomal analysis of lymphocytes in the peripheral blood of the patient revealed abnormal patterns of 45, XO/46, XX/47, XXX, the second, 45, XO/46, XX/47, XXX/47.XX, + 21. Measurement of FITC-labelled fibrinogen antibody against the fibrinogen combined with platelet glycoprotein GP IIb/IIIa complex using flow cytometry showed an apparently increased positive rate 76.2% for the platelet of the patient, compared with that of 31.9% for the control. From the comparative study of the platelet agglutination of the platelet rich plasma (PRP) derived from the citrated-added complete blood taken from the patient to whom Ticlopidine was given, added with adenosine-5'-diphosphate (ADP), collagen or acetyl glyceryl ether phosphoryl choline (AGEPC) as the reagents and that of normal control, it was shown that for AGEPC increased. Thus, increased reaction specific to AGEPC in addition to activated platelet was demonstrated in the present study.

Three cases of "spontaneous" pneumocephalus suspected to have resulted from aerobic bacteremia caused by Enterobacter cloacae, Escherichia coli, and Klebsiella aerogenes are reported. In two cases, the E. cloacae and K. aerogenes were isolated from the cerebrospinal fluid. These cases were characterized by a rapid accumulation of air, without niveau, in the subarachnoid space and ventricles.

Ninety-seven pulmonary metastatic cases in 141 osteosarcoma from 1960 to 1981 were analysed. The overall cumulative 5-year survival rate of the cases was 38.1%, whereas that of the recent 41 cases with systemic chemotherapy was 56.2%. Eighty percent of pulmonary metastases were found within one year since their first visit to our clinic. It was revealed that there were positive relationship between the doubling time of the pulmonary metastases and the post-metastatic survival (p less than 0.001), and between the period from their first visit to our clinic to metastases and the post-metastatic survival (p less than 0.001). The favorable effects of the systemic chemotherapy using adriamycin, vincristine, methotrexate and cis-platinum were recognized. The thoracotomy revealed effective and five cases are living without disease for average 80.8 months after surgery.

Two patients with hepatic fascioliasis are presented. Computed tomography demonstrated multiple low density areas located in the center and periphery of the liver. In one patient a spontaneous decrease in the size of the low-density areas occurred but the abdominal symptoms were persistent before bithionol treatment. After the treatment both patients became asymptomatic. The computed tomography scan can be a useful tool for the diagnosis and follow-up of the patients with hepatic fascioliasis.

A high-performance liquid chromatographic (HPLC) method for the determination of L-carnitine in clamped and frozen rat livers is described. L-carnitine + acetyl-CoA in equilibrium with acetyl-L-carnitine + CoASH Using the above enzymatic reaction, release of CoASH is stoichiometric with the L-carnitine added. The present method has made possible the determination of carnitine in liver tissues, which is difficult by the conventional enzymatic spectrophotometric method using 5,5'-dithiobis(2-nitrobenzoic acid), owing to acetyl-CoA hydrolysis during prolonged incubations at pH 7.8.

The occurrence of rhabdomyolysis and acute renal failure associated with viral infections is uncommon. We report a case of a 17-year-old female suffering from rhabdomyolysis caused by echovirus 9 in which the patient recovered after being treated for acute renal failure with hemodialysis. Among 42 cases of virus-induced rhabdomyolysis review in Japanese and English literatures, influenza virus is the most common cause of rhabdomyolysis, coxsackie virus tends to be next most common cause. Renal failure was recognized in 31 of the cases.

A method for the determination of carnitine palmitoyltransferase I (CPT I; EC 2.3.1.19) in isolated rat liver mitochondria by reversed-phase high-performance liquid chromatography is described. Enzyme activity is assayed by direct determination of coenzyme A (CoA) released from palmitoyl-CoA within 60 min by a linear gradient system. CPT 1 in rat liver mitochondria can be assayed from only 30 micrograms of mitochondrial protein per millilitre of assay mixture. The changes in the kinetic parameters of CPT I, including Ki for malonyl-CoA, resulting from the fasting-feeding cycle are also discussed.

A case of a 73 year old man who was preoperatively diagnosed as possibly having myelolipoma is presented herein. Adrenal myelolipoma is an uncommon benign tumor of which only 74 foreign cases and 43 Japanese cases have been previously reported in the literature dealing with surgically removed myelolipomas. The most consistent complaint of the adrenal myelolipoma sufferer is abdominal pain, caused by hemorrhaging in the tumor, and many cases are associated with obesity, hypertension and/or diabetes mellitus. There is now an increasing number of such cases being diagnosed during abdominal scanning with ultrasonography or computerized tomography for unrelated problems, whereupon the myelolipoma is usually surgically resected. With the availability of modern scanning techniques and fine needle biopsy, however, it should be possible to adopt a more conservative approach to the management of asymptomatic adrenal myelolipoma. Nevertheless, symptomatic or large tumors, must be removed since there is a high risk of spontaneous hemorrhage.

Rats having a non-necrotic damaged liver or a necrotic damaged liver produced by D-galactosamine administration were restrained in water for 0.5, 1, 2, 4, 6, 8 and 12 hours. Serial changes in intrahepatic energy metabolism were compared with those in normal liver. Energy charge, which represents the degree of equilibrium between the energy producing and consuming systems, and cyclic AMP, an intracellular messenger mediating the action of hormones, showed biphasic increases before and after 2 hr in the damaged livers. The lactate/pyruvate ratio, which reflects the cytosolic redox state, markedly increased at 0.5 hr in the damaged livers but returned to the pre-stress level after 1 hr in the non-necrotic damaged liver and after 4 hr in the necrotic damaged liver, showing a transient reduced state. The beta-hydroxybutyrate/acetoacetate ratio, which represents the mitochondrial redox state, decreased at 0.5 hr and returned to the pre-stress level at 1 hr in the non-necrotic damaged liver, exhibiting a transient oxidized state. However, in the necrotic damaged liver, the value decreased at 0.5 hr remained low thereafter, demonstrating a persistent oxidized state. These findings show that, in severely damaged liver, stress has more marked effects on hepatic energy metabolism.

The incidence of duodenal varices is exceedingly rare. A case of bleeding duodenal varices located in the third portion of the duodenum, secondary to idiopathic portal hypertension, which was successfully treated by surgery, is presented herein. Diagnosis was suspected at superior mesenteric angiography and was subsequently confirmed by endoscopy. A review of the literature reveals only 105 such cases in the world. While the duodenal bulb is the most common site of duodenal varices, the second portion of the duodenum appears to be the next most common site but duodenal varices in the other portions are extremely rare. From all of the possible causes of duodenal varices, liver cirrhosis remains the predominant etiological factor in 31 cases.