Urinary excretion of uric acid, allantoin, and 8-OH-Deoxyguanosine in uricase-knockout mice.

Although uricase-knockout (Uox KO) mice are reported to develop uric acid (UA) nephropathy, those that mature without severe nephropathy could be useful for research into purine metabolism in humans. In this study, we measured the urinary excretion of creatinine, UA, allantoin, and 8-hydroxy-2'-deoxyguanosine (8-OHdG) collected from Uox KO mice housed in metabolic cages. UA and allantoin were determined using liquid chromatography-mass spectrometry and creatinine and 8-OHdG were measured with a commercial kit. Uox KO mice excreted significantly higher levels of UA than wild-type mice (C57BL/6), while the excretion of allantoin was significantly lower. Urinary allantoin was detected in Uox KO mice despite a lack of uricase, which is the same as in humans. In contrast to the elevated levels of UA, the daily excretion of 8-OHdG, an oxidative stress marker, was lower in Uox KO mice. UA is thought to act as an anti-oxidizing agent in humans; thus, these results show that Uox KO mice are potential animal models for research into human purine metabolism.

Inhibitory effects of tranilast on the proliferation and functions of human pterygium-derived fibroblasts.

PURPOSE: We studied the possibility that tranilast, an antiallergic and antiproliferative drug, may be beneficial for the treatment of pterygium. METHODS: Pterygium-derived cells were identified by immunohistochemical methods. Growth rate of pterygium-derived cells was determined by using a hemocytometer. Chemotaxis was determined in a microchemotaxis chamber. Pterygium-derived cells were cultured on floating collagen gel, and the contracted diameter was measured. Collagen synthesis by pterygium-derived cells was determined by the collagenase digestive method. Tranilast was added to the culture medium at final concentrations of 0, 12.5, 25, 50, and 100 microg/ml. RESULTS: Pterygium-derived cells were stained with anti-prolylhydroxylase and anti-alpha-smooth muscle actin, and identified as fibroblasts. Tranilast inhibited the proliferation and chemotaxis of pterygium-derived fibroblasts, and the collagen-gel contraction induced by these cells, but it exerted no inhibitory action on collagen synthesis by pterygium-derived fibroblasts. CONCLUSION: Tranilast may be useful for suppressing the recurrence and, possibly, the development of pterygium.

Male reproductive toxicity of ethinylestradiol associated with 4 weeks daily dosing prior to mating in rats.

Parameters of male reproductive toxicity of ethinylestradiol were assessed by conducting a mating test, sperm assay, organ weight determination and histopathological examination. Male Sprague Dawley rats were orally administered 0.1, 0.3, 3 or 10 mg/kg/day ethinylestradiol for 4 weeks prior to mating. Body weight gain and food consumption were suppressed in all treated groups. Reproductive ability of the 3 and 10 mg/kg/day males disappeared. Slightly low copulation indices were observed in the 0.1 and 0.3 mg/kg/day groups, although fertility indices were not affected. Sperm could hardly be found in the epididymis of 3 and 10 mg/kg/day males. Sperm counts were also decreased in the other treated groups, but sperm motility was not affected. Decreased absolute and/or relative weights of testes, epididymides, prostate and seminal vesicles were observed in all treated groups along with testis, epididymis, seminal vesicle and prostate atrophy, and degenerative changes of spermatocytes, spermatids, Sertoli cells and Leydig cells. These results suggest that sperm quantification and histopathological assessment are more appropriate for assessing male reproductive toxicity of ethinylestradiol than performance of copulation and fertility tests.

In vitro study of teratogenic effects of caffeine on cultured rat embryos and embryonic cells.

The teratogenic potential of caffeine was examined in vitro by a whole embryo culture system (WECS) and an embryonic cell culture system (micromas teratogen assay: MTA) in the rat. In the WECS, hyperemia of the tail, and a reduction of the placental size was induced by caffeine at concentrations higher than 50 micrograms/ml; hypoplasia of the forelimb bud was induced at concentrations higher than 100 micrograms/ml; hematoma in the yolk sac and dysmorphogenesis of the fore- and hind-limb buds, prosencephalon and tail were induced by 200 micrograms/ml caffeine. In the MTA, even with 200 micrograms/ml caffeine, the toxicological parameters obtained by proliferation and differentiation assays of the midbrain and limb bud cells were almost the same as in the control. In conclusion, caffeine induced various morphological anomalies, but did not affect proliferation or differentiation of cells in these experimental systems.

Renal cell carcinoma with renal artery aneurysm.

We describe a patient who was diagnosed as renal cell carcinoma with renal artery aneurysm in the contralateral kidney. Right aneurysmectomy followed by simple arteriorrhaphy with termino-lateral anastomosis, and then left radical nephrectomy were performed in one session. The patient did well postoperatively. Generally, surgical indication of renal artery aneurysm itself has been yet a matter of debate. In such case like this, however, it seems better to resect any kind of aneurysm of the opposite side, taken into consideration the haemodynamic changes after nephrectomy. Surgical indications are commented on.

[Localized cardiac tamponade after aortic valve replacement: a case report].

We report a case of localized cardiac tamponade after aortic valve replacement. A 56-year-old man had an aortic valve replacement for his aortic valve steno-insufficiency. At 3-postoperative day, severe hypotension occurred, causing acute renal failure. There were no cardiomegaly, high central venous pressure, nor echo-free space. A mass shadow, appearing on chest X-ray at 37-postoperative day, was diagnosed as a localized tamponade by means of a computed tomography and a radioangiography at 38 postoperative day. After the spontaneous drainage of old bloody effusion from the partially opened wound in mid-line, his cardiac and renal failure improved rapidly. When the hematoma is localized, computed tomography is most diagnostic while conventional echo-cardiography often fails to show echo-free spaces.

[The change of polymorphonuclear elastase releasing from ischemic skeleton muscles after reperfusion].

This report evaluates the activity of Polymorphonuclear Elastase (PMNE), after reperfusion of the skeleton muscles during the infra-renal abdominal aortic aneurysmal surgery. We studied the following data to prevent post-operative complications of acute arterial occlusion: the change of PMNE and alpha 1-AT and the effect of Ulinastatin (U)-the inhibitor of PMNE. Twenty surgical cases of the abdominal aortic aneurysms were used in the study. Ten cases of non-U-treated served as group I, 10 cases of U-treated served as group II. PMNE level, in group I, showed a value of 233.7 +/- 34.8 micrograms/l pre-clump and 848.0 +/- 110.3 micrograms/l after releasing the clamp 2 hours later (p < 0.001). In group II, pre-clamping value was 204.8 +/- 21.0 and a relative low value of PMNE (416.7 +/- 36.6 micrograms/l) 2 hours later after releasing the clamp compared with group I. In group I, consistently positive correlations were found between the duration of clamping time and the value of PMNE. This study shows that PMNE increases statistically after re-perfusion of the skeleton muscle. U shows a significant inhibition of PMNE increasing. U may prevent further damages of the visceral organs after operation of acute arterial occlusion.

Determination and profiling of purines in foods by using HPLC and LC-MS.

Purines in food are known to raise serum uric acid levels. We determined the purine content of sweet potato and beef by high-performance liquid chromatography and liquid chromatography-mass spectrometry. The purine content of the samples was 118-1,034 µmol/100 g. The total purine content was also divided into purine bases, nucleosides, nucleotides, and nucleic acids. Our results suggest that differences in total purine content and in the ratio of purine types between vegetables and beef cause a difference in elevation of serum uric acid levels.

Simultaneous determination of purine and pyrimidine metabolites in HPRT-deficient cell lines.

Genetic mutations in the purine salvage enzyme, hypoxanthine-guanine phosphoribosyltransferase (HPRT), are known to cause Lesch-Nyhan syndrome and Kelley-Seegmiller syndrome. In patients, purine metabolism is different from that of normal persons. We have previously developed a method for simultaneously determining the concentration of purine and pyrimidine nucleosides and nucleotides. This system was applied to determine the concentrations of nucleosides and nucleotides in HPRT-deficient cell lines. The amount of inosine 5'-monophosphate (IMP) was different in Lesch-Nyhan syndrome, Kelley-Seegmiller syndrome, and control cell lines. The difference in the amount of IMP confirmed the mutation of the enzyme.

[Secondary tricuspid insufficiency and right atrial myosin ATPase activity].

Myosin of heart muscle shows ATPase activity. In the atrial myocardium, normal isozymic pattern was alpha dominant which converted to being beta dominant in an overloaded hypertrophy. In order to clarify the distribution of myosin isozymes in human heart, ATPase activity of the atrial myosin recovered from the patient underwent open heart surgery was determined. In the present study, ATPase activity of right atrial myosin from the heart with tricuspid regurgitation (TR) (group A (n = 6); 398.1 +/- 67.0 nmol pi/mg/min) was significantly less than that from the heart without TR (group B (N = 7); 533.9 +/- 62.4, p less than 0.05). The myosin ATPase activity showed correlation with systemic RA pressure (y = 0.019x + 19.6, r = -0.68429), systemic RV pressure (y = 0.039x + 58.67, r = 0.73484), SVI (y = 0.05x + 18.1, r = 0.87587) and RV maxDp/Dt (y = 0.42x + 589.9, r = -0.67493) (p less than 0.05). These data suggests that preoperative cardiac function involves in cardiomuscular structure with redistribution of contractile protein.

[A case of treatment of the Paget-Schroetter syndrome with PTA and Gianturco' expandable metallic stent].

We report a effective management of the Paget-Schroetter syndrome. A 23-year-old man was seen with a complaint of arm swelling and venous engorgement of the left arm. A first-rib resection was performed through a infraclavicular approach with removal of compressive elements including scalenus anterior and subclavius muscles. He became asymptomatic. But 2 months later, recurrent symptoms developed and venograms demonstrated the presence of restenosis of the subclavian vein with extensive collaterals. To relieve symptoms a percutaneous transluminal angioplasty (PTA) and a insertion and placement of the Gianturco's expandable metallic stents was carried out for the remaining stenosis of the subclavicular vein and a residual compression. Venograms, 7 months after operation, showed widely the patent subclavian vein. Trans-infraclavicular approach allowed exposure of the stenotic or obstructed segments of the subclavian vein. The PTA and stenting, especially after removal of compressive elements or during chronic phase, was a very useful and effective procedure for the treatment of the Paget-Schroetter syndrome.

Pharmacokinetics and disposition of rosuvastatin, a new 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, in rat.

1. The pharmacokinetics and disposition of rosuvastatin, a new 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, were investigated following single administration of (14)C-rosuvastatin in the Sprague-Dawley rat. 2. Following oral administration of (14)C-rosuvastatin at 1, 5 and 25 mg kg(-1), the C(max) and AUC of the radioactivity in the plasma increased more than the increase in dose ratio. 3. Excretion of radioactivity was 98.0% of the dose in the faeces and 0.4% in the urine up to 168 h after oral administration in the intact rat, and was 55.1% in the bile and 0.5% in the urine up to 48 h post-dosing in the bile duct-cannulated rat. The unchanged compound mainly accounted for the radioactivity in the bile and faeces. 4. In the tissue distribution study, the concentration of the radioactivity in the liver was markedly higher than those in the other tissues, and the radioactivity concentration ratios of the liver to the plasma were between 8 and 25 up to 48 h after oral administration. The liver-specific distribution of rosuvastatin was similarly recognized in whole-body autoradiography. 5. Metabolic profiling studies indicated that rosuvastatin would not be metabolized by CYP enzymes. 6. These results clarified that rosuvastatin selectively distributed in the liver - the target organ - and was excreted in the bile mainly as the unchanged compound.

Disposition of moxalactam and N-methyltetrazolethiol in rats and monkeys.

The disposition of moxalactam (MOX) and N-methyltetrazolethiol (NMTT) in rats and monkeys after intravenous injection was investigated, focusing on the in vivo liberation of NMTT, by using [NMTT-14C]MOX and [14C]NMTT. After [NMTT-14C]MOX injection, MOX levels in plasma quickly became high in both rats and monkeys and then declined, with half-lives at the beta phase of 18.8 and 67.1 min, respectively. The levels of NMTT liberated from MOX were much lower than those of MOX, but the apparent elimination was significantly slow. The levels of MOX and NMTT in rat liver were almost comparable but lower than those in plasma. With [14C]NMTT administration, the level of NMTT in plasma declined, with half-lives at the beta phase of 21.5 min in rats and 54.0 min in monkeys. After [NMTT-14C]MOX injection, most of the radioactivity was excreted in urine as MOX, with 11% of the dose in rats and 8% of the dose in monkeys eliminated as NMTT until 24 h. Total biliary excretion was 26% of the injected radioactivity in rats, and most of it was due to MOX. In one monkey, the total biliary excretion was only 0.2% of the injected radioactivity. With [14C]NMTT administration, most radioactivity was excreted in the urine as unchanged NMTT in both animals. Oral administration in rats showed that part of the biliary-excreted MOX was degraded to NMTT in the intestine and then absorbed. Repeated administration of [NMTT-14C]MOX to rats did not change the levels of MOX and NMTT in plasma or liver nor did it change the excretion profiles. Thus, accumulation of MOX and NMTT did not occur.

FR901724, a novel anti-human immunodeficiency virus (HIV) peptide produced by Streptomyces, shows synergistic antiviral activities with HIV protease inhibitor and 2',3'-dideoxynucleosides.

A novel tricyclic 21-amino-acid peptide, FR901724, was isolated from the cultured broth of Streptomyces sp. No. 73264. This peptide appears to possess potent anti-human immunodeficiency virus (HIV) activity in vitro and might represent a lead to a new class of anti-HIV agents; it qualifies as an HIV-cell fusion inhibitor because of its weak inhibition of virus-cell binding and strong inhibition of syncytium formation. From the time-of-addition experiments, the mode of action of FR901724 was found to definitely differ from that of KNI-272, a peptide mimetic allophenylnorstatine-derivative HIV protease inhibitor. FR901724 appears to interact with a stage of the virus replicative cycle that may well correspond to virus-cell fusion. We also found that FR901724 was synergistic or had a strong tendency toward synergism when combined with other antiviral drugs, such as KNI-272, AZT, ddI and dextran sulfate.

Disposition and metabolism of the new hypocholesterolemic compound S-8921 in rats and dogs.

S-8921 (methyl 1-(3,4-dimethoxyphenyl)-3-(3-ethylvaleryl)-4-hydroxy-6,7,8-trimeth oxy-2- naphthoate, CAS 151165-96-7) is a novel hypocholesterolemic agent which was found to inhibit ileal Na+/bile acid cotransporter. In this report, the pharmacokinetic profile of S-8921 was studied in rats and dogs. After dosing of 14C-S-8921 to rats at 1 to 25 mg/kg as 0.5% methylcellulose (MC) suspension, tmax was observed during 5-6 h, and AUCs increased with the dose, but not proportionally. The elimination half-lives were around 38-41 h for the doses examined. The apparent absorption ratio of 5 mg/kg of 14C-S-8921 as MC suspension was about 14%. Most of the radioactivity (98% of dose) was excreted into the feces and only 1-2% into the urine. Biliary excretion of radioactivity after dosing of 1, 5 or 25 mg/kg was 22, 20, 15%, respectively. Saturation of the absorption process was suggested. Even in case of intravenous dosing, about 88% was excreted into the bile. Enterohepatic circulation of biliary metabolites was also observed in rat. Its extent was small (6%), but, it may be contribute to the slow elimination of S-8921 from rat. The highest radioactivity was observed in the liver, with other tissues showing similar radioactivity profiles to that of plasma. The elimination half-lives of radioactivity from tissues were very long, e.g. 68 h for the liver and 58 h for the kidney. After 14 days multiple dosing, most tissues showed about two times higher radioactivity than that after a single dose. The simulation curves of liver and plasma showed a good fit with those of the observed values. These results suggested that there is no serious accumulation of radioactivity in tissues by multiple dosing of 14C-S-8921 in rats. The plasma radioactivity after oral dosing of 5 mg/kg of 14C-S-8921 to dogs as an MC suspension reached maximum concentration (c.a. 33 ng/ml) at 2 h, then decreased very slowly with a half-life of 169 h. The apparent absorption ratio was 4.6% for MC suspension. The excretion of radioactivity into bile, feces and urine after oral dosing of 14C-S-8921 at 5 mg/kg as an MC suspension were 3.0%, 94.6% and 0.3%, respectively. Even in the case of intravenous dosing, urinary excretion was very small (2.2%) and most of the radioactivity was excreted very slowly into the feces. The major metabolite of S-8921 in rat bile was its glucuronide. Other minor metabolites identified were the demethylated forms of 7-methoxy and 4'-methoxy moieties of S-8921. They were also excreted into bile as their glucuronides.

Biosynthesis of 20,20,20-trifluoroleukotriene B4 from 20,20,20-trifluoroarachidonic acid: a metabolically stable analog of leukotriene B4 and its application to a study of stimulation of leukotriene B4 synthesis by immunoglobulin G1,2.

Leukotriene B4 is rapidly metabolized through omega-oxidation, preventing its detection when it is produced under certain biological conditions. To investigate leukotriene B4 production in various physiological conditions, analogs of arachidonic acid which are converted to metabolically stable analogs of leukotriene B4 would be useful. We have synthesized 20,20,20-trifluoroarachidonic acid by the cis-selective Wittig reaction of the C12-C20 fragment with phosphonium salt. 20,20,20-trifluoroarachidonic acid was transformed into 20,20,20-trifluoroleukotriene B4 when incubated with human neutrophils in the presence of the calcium ionophore A23187. The product was identified by uv absorption spectrophotometry, gas chromatography-mass spectrometry, and coelution on high-performance liquid chromatography with 20,20,20-trifluoroleukotriene B4, which was enantioselectively synthesized by the reaction of the fluorine-containing C11-C20 fragment with the C1-C10 phosphonate. The fluorinated leukotriene B4 demonstrated as much chemotactic activity on human neutrophils as natural leukotriene B4 and was metabolically stable when incubated with human neutrophils, probably by blocking omega-oxidation. Also, enzymes catalyzing the transformation of arachidonic acid (AA) into leukotriene B4 did not discriminate the fluorinated precursors from the natural, nonfluorinated AA, thus 20-F3-AA is a valid analog of AA to be used in the study of AA metabolism. When 50 microM of the fluorinated acid was incubated with neutrophils stimulated with heat-aggregated human immunoglobulin G, a significant amount of fluorinated leukotriene B4 (4.3 ng/10(6) cells/40 min, at most) was formed in a dose-dependent manner while little leukotriene B4 was detected with incubation with 50 microM arachidonic acid, probably due to omega-oxidation of the product, leukotriene B4. 20,20,20-Trifluoroarachidonic acid appears to be a useful tool for studying the capacity of leukotriene B4 synthesis in various biological systems while long-lasting 20,20,20-trifluoroleukotriene B4 would serve as an excellent analog of leukotriene B4 in pharmacological studies to understand functions of leukotrienes B4.

Ocular absorption, distribution, and systemic absorption of a novel antiglaucoma medication, prostaglandin derivative, in male white rabbits.

A prostaglandin derivative, (5Z,9 alpha,11 alpha,13E)-9,11-dihydroxyprosta- 5,13-dienoic acid sodium salt (S-1033), that lowers intraocular pressure with little adverse effect, may have clinical value in the treatment of glaucoma. After [14C]S-1033 (0.2% solution) was instilled into the eye of a white rabbit, radioactivity and S-1033 appeared in systemic plasma so rapidly (tmax, 5 min) and S-1033 was eliminated very rapidly with half-lives of 2.8 and 11.0 min at alpha- and beta-phases, respectively. The metabolite, M-1, [1R-[1 alpha,2 beta-(1E),3 alpha,5 alpha]]-3,5-dihydroxy-2-(1- octenyl)-cyclopentanepropanoic acid (tetranor-S-1033), appeared in plasma very rapidly (tmax, 5 min), suggesting that a fast metabolism was a major factor in the rapid elimination of S-1033 from plasma. The values for the ratios of the area under the curve of ocular instillation to intravenous administration for radioactivity and S-1033 were 1.01 and 0.52, respectively, indicating that more than half of the S-1033 instilled was transported into the systemic circulation. To clarify the contributing pathway of the massive and rapid systemic absorption of S-1033 after topical dosing, plasma levels of S-1033 were investigated after instillation to rabbits in which the nasolacrimal ducts were occluded. Plasma concentrations of S-1033 were slightly higher than those in intact rabbits, suggesting that conjunctiva would be as important as nasal mucosae for the systemic absorption under the physiological condition. As for the intraocular distribution, the highest levels of radioactivity were found in the cornea, conjunctiva, and anterior sclera.(ABSTRACT TRUNCATED AT 250 WORDS)

A novel assay system for anti-human immunodeficiency virus type 1 (HIV-1) activity using a subclone of a monocytic cell line, U937.

A subclonal cl.1-14 cell was established from a monocytic cell line U937 by a limiting dilution method. The anti-HIV-1 activity of some antiviral compounds was evaluated in HIV-1-infected cl.1-14 cells. The results demonstrated that although AZT was a potent inhibitor of HIV-1 replication in cl.1-14 cells, its 50% effective concentration (EC50) values was 80 times higher than that in HIV-1 infected MT-4 cells; the EC50 of AZT was 0.16 microM and 0.002 microM in cl.1-14 and MT-4 cells, respectively. In contrast, the anti-HIV-1 activity of ddA, ddI and ddC in cl.1-14 cells was comparable to that in MT-4 cells. The antiviral activity of nevirapine, dextran sulfate, curdlan sulfate and T22 did not differ significantly between the cl.1-14 and MT-4 cells. The antiviral activity of several compounds in the HIV-1-infected cl.1-14 cells was similar to that in the HIV-1JR-FL-infected human peripheral macrophages. Our results suggest that cl.1-14 cell cultures are very useful for estimating antiviral activity and more advantageous than the use of peripheral blood macrophages.