Potent effects of novel anti-platelet aggregatory cilostamide analogues on recombinant cyclic nucleotide phosphodiesterase isozyme activity

The inhibitory potential of novel anti-platelet aggregatory cilostamide analogues on phosphodiesterase (PDE) isozyme activities was investigated with recombinant PDE isozymes expressed in a baculovirus/ Sf9 expression system. The recombinant enzymes (PDE1-PDE5 and PDE7) showed Km values and sensitivities to selective inhibitors similar to those reported previously for native enzymes purified from tissues. The cyclooctylurea derivative OPC-33540 (6-[3-[3-cyclooctyl-3-[(1R*,2R*)-2-hydroxycyclohexyl]ureido]-propoxy]-2(1H)-quinolinone) inhibited recombinant PDE3A (IC50 = 0.32 nM) more potently and selectively than the classical PDE3 inhibitors cilostamide, cilostazol, milrinone, and amrinone. The cyclopropylurea derivative OPC-33509 [(-)-6-[3-[3-cyclopropyl-3-[(1R,2R)-2-hydroxycyclohexyl]ureido]-propoxy]-2(1H)-quinolinone] was less potent (IC50 = 0.10 microM) than OPC-33540, demonstrating that the cyclooctyl moiety was important for a potent inhibitory effect. In platelets, OPC-33540 potentiated cyclic AMP accumulation concentration-dependently in both the absence and the presence of 3 nM prostaglandin E1 (PGE1) (doubling concentrations: 32.5 and 6.2 nM, respectively). OPC-33540 inhibited thrombin-induced platelet aggregation potently (Ic50 = 27.8 nM). The anti-platelet aggregation effect also was stimulated in the presence of 3 nM PGE1 (IC50 = 6.0 nM). There was a good correlation between the IC50 values of PDE3 inhibitors in this study for recombinant PDE3A activity and their IC50 values for thrombin-induced platelet aggregation (r = 0.998). These data demonstrated that OPC-33540 is a highly selective and potent PDE3 inhibitor and a useful probe for identification of the intracellular functions of PDE3.     

Development of novel corneal storage medium: first report. Examinations of rabbit cornea

PURPOSE: To develop a new corneal storage medium with a simple formula and evaluate it by histological methods. METHODS: Two corneal storage media containing minimum essential medium and 2.5% chondroitin sulfate (pH 7.33), with osmolarity of 320 mOsm/kg, were compared to Optisol-GS. The differences in the two media were the molecular weight (MW) and the source of chondroitin sulfate. The MW of Medium I was 27,500 and the MW of Medium II was 33,700. Japanese albino rabbits were used in this study. A cornea with scleral rim obtained from a rabbit was stored in either Medium I or Medium II and the fellow cornea was stored in Optisol-GS for 7 or 14 days at 4 degrees C. Histological examination of corneal endothelial cells was performed both by scanning electron microscopy and by transmission electron microscopy. RESULTS: At day 7, there was no significant difference in histological findings among the rabbit corneas stored in Optisol-GS, Medium I, or Medium II. At day 14, corneas stored in Optisol-GS or Medium I showed similar histological findings. In Medium II, endothelial cells showed marked degeneration. CONCLUSIONS: The results of experiments with rabbit cornea indicated that Optisol-GS and Medium I could preserve endothelial cellular structure better than Medium II. The difference between Medium I and Medium II was only the MW of the chondroitin sulfate used. The MW may be an important factor for determining suitable chondroitin sulfate for use in a corneal storage medium.     

Lack of interaction between amiodarone and mexiletine in cardiac arrhythmia patients

Amiodarone has pharmacokinetic interactions with various therapeutic agents, including phenytoin, flecainide, and cyclosporine. Mexiletine is metabolized by CYP2D6 and CYP1A2. The objective of this study is to evaluate the effect of amiodarone on the pharmacokinetics of mexiletine through its inhibition of various cytochrome P450 (CYP) subtypes. In a series of 181 inpatients with supraventricular tachyarrhythmias, 26 inpatients received mexiletine and amiodarone therapy (MEX + AMD group), and the others received mexiletine therapy (MEX group). In 10 inpatients of the MEX + AMD group, the mexiletine clearance (CL(MEX)/F) before and after coadministration of amiodarone was compared. CL(MEX)/F was also compared in the MEX and MEX + AMD groups after the start of amiodarone therapy. Serum mexiletine, amiodarone, and desethylamiodarone concentrations were measured by an HPLC method. The CL(MEX)/F was estimated by the Bayesian method using population pharmacokinetic analysis. There was no significant difference in CL(MEX)/F before and after 1-month coadministration of amiodarone in 10 inpatients of the MEX + AMD group. Although serum amiodarone and desethylamiodarone concentrations gradually increased with time after the start of amiodarone therapy in these patients, CL(MEX)/F showed no change at 3 and 5 months after the start of amiodarone therapy. There was no significant difference in CL(MEX)/F of the MEX group and the MEX + AMD group. The results suggest that the pharmacokinetics of mexiletine is not affected by amiodarone in patients with cardiac arrhythmias.     

Effects of estrogenic compounds on immunoglobulin production by mouse splenocytes

The effects of culture medium and serum components on immunoglobulin (Ig) production by mouse splenocytes were examined. In this study, we showed that culture medium containing charcoal-dextran-treated fetal bovine serum (CDFBS) supported Ig production more efficiently than culture medium containing FBS. In addition, RPMI 1640 medium supported Ig production more efficiently than Dulbecco modified Eagle medium. In addition, an increase in medium IgA production was observed with the increase in both CDFBS and FBS concentrations, whereas an increase in IgM level was observed in the presence of 5% and 10% FBS and 10% CDFBS. Dose-dependent effects of estrogenic compounds on Ig production were also examined. We found that 17beta-estradiol induced increases in IgM and IgE levels, but diethylstilbestrol enhanced only the IgE level. On the other hand, tamoxifen and bisphenol A enhanced medium IgM level at physiological concentrations. Among flavonoids, daidzein enhanced IgM and IgE levels at concentrations above 10 microM, and genistein induced a decrease in IgM level and an increase in IgE level at concentrations above 10 microm. In addition, quercetin and luteolin enhanced medium IgE level at all concentrations tested, whereas IgA, IgG, and IgM levels were not affected. These results suggest that environmental estrogens affect Ig production of mouse Splenocytes in a complex and class-specific manner.     

Relation between teeth clenching and grip force production characteristics

This study investigated the possible contributing effect of oral motor function on maximal and explosive grip force production characteristics. Fourteen healthy male subjects (age 22.5 +/- 2.1 years) were asked to exert maximal explosive grip strength with their dominant hands under the following four conditions: 1. Teeth clenching before and during grip strength exertion (C-C), 2. Teeth clenching before grip strength exertion and mandibular resting position during grip strength exertion (C-R), 3. Mandibular resting position before grip strength exertion and teeth clenching during grip strength exertion (R-C), and 4. Mandibular resting position before and during grip strength exertion (R-R). Maximal force (maxF), average force for every 0.1 s (aveF), maximal rate of force development (maxRFD) and time required to reach 90% of maxF (T 90% max) were analyzed for 1 s from the onset of grip force production. MaxF under C-C and R-C were significantly greater than that under R-R by 12.1% and 12.3%, respectively. AveF under C-C was significantly greater by 10.0-41.2% than that under R-R for all ten periods. AveF under C-R was significantly larger by 9.8-19.0% than that under R-R conditions from 0 to 0.4 s. Compared with under R-R conditions, maxRFD under C-C and C-R increased by 15.8% and 8.5%, respectively, and T 90% max under C-C, C-R and R-C decreased by 22.3%, 12.3% and 12.8%, respectively. These findings suggest that oral motor functions such as teeth clenching may influence not only maximal grip strength generation but also the rapidity of grip force production.     

Seminal lactate dehydrogenase C4 (LDH-C4) isozyme activity in infertile men

Lactate dehydrogenase C4 (LDH-C4) is the specific isozyme of LDH produced by germ cells. We measured LDH-C4 activity in seminal plasma from infertile men with oligozoospermia or azoospermia using gel electrophoresis. Total LDH activity in seminal plasma from infertile patients (n = 99) was 2,487 +/- 1,384 IU/l (mean +/- SD). LDH-C4 isozyme activity was detected in 63 out of 75 seminal plasma samples from infertile patients with a mean of 383 +/- 356 IU/l (13.8 +/- 8.6% of total LDH). Sperm count was positively correlated with LDH-C4 (r = 0.298; P < 0.05), but not with any other LDH isozymes or with total LDH. Seminal LDH-C4 was significantly lower in patients with varicoceles (253 +/- 223 IU/l) than without varicoceles (474 +/- 262 IU/l). Six azoospermia patients were treated with hCG and hMG. Three out of four patients whose seminal plasma revealed LDH-C4 activity responded to the treatment, whereas none of the other two patients without seminal LDH-C4 activity did. These results indicate the clinical usefulness of seminal LDH-C4 as a potential marker for seminal epithelium activity in the diagnosis and treatment of male infertility.     

Venoarterial bypass in children with congenital heart disease--a retrospective study

To investigate prognostic factors for patients supported by venoarterial (VA) bypass, we analyzed retrospectively 31 patients with congenital heart diseases supported by VA bypass between 1997 and 2000. Median age was 3.6 months and median body weight was 4.0 kg. Causes of VA bypass were difficult weaning from cardiopulmonary bypass in 8 patients, postoperative cardiac dysfunction in 5 and cardiopulmonary resuscitation in 18. Six (19.4%) of 31 patients were weaned successfully from VA bypass, and 2 (6.5%) survived to hospital discharge. In patients who were weaned from VA bypass successfully, VA bypass was instituted more quickly (41.6 +/- 5.0 vs 62.5 +/- 35.9 min), and good tissue perfusion was established in association with lower lactate levels at 12 hr (7.0 +/- 5.9 vs. 16.4 +/- 15.4 mmol.l-1) and larger urine output during first 24 h (81 +/- 68 vs. 22 +/- 43 ml.kg-1.day-1), compared to those who failed to be weaned. Major complications were intracranial hemorrhage in the newborn (50%), and hypoxic brain damage in patients with palliative operation (35%). It is necessary to establish guidelines of entry, weaning, and withholding of this support.     

Dose-dependent effect of dietary conjugated linoleic acid on the growth of rat hepatoma dRLh-84 cells in vivo

In this study, the effect of varying doses of conjugated linoleic acid (CLA) on the growth of transplanted hepatoma dRLh-84 cells and the relationship between tumor growth and prostaglandin (PG) E2 production or cyclooxygenase (COX)-2 expression were examined. Donryu rats were fed an experimental diet containing 0, 0.1, 0.5, or 2 wt.% CLA for 3 wk, and then dRLh-84 cells were transplanted into the liver. Results show that dietary CLA (0.5 and 2 wt.%) significantly enhanced the growth of the transplanted hepatoma cells compared to the non-CLA diet group at 20 d after cell transplantation. Tumor weight at 10 d after transplantation was also significantly higher in the 2 wt.% CLA group than in non-CLA fed rats. Ten days after transplantation, the PGE2 level in the tumor tissue was shown to be depressed in a CLA dose-dependent manner. Cyclooxygenase-2 (COX-2) mRNA expression in the tumor also tended to be lower in the CLA group than in the non-CLA diet group 10 d after transplantation. Dietary CLA did not affect the tumor phospholipid arachidonic acid level, which is a substrate for PG synthesis. These results indicate that dietary CLA of at least 0.5 wt.% enhances the growth of transplanted dRLh-84 cells in vivo. It is believed that growth promotion of dRLh-84 cells in vivo by CLA cannot be clarified by the PG synthesis dependent mechanism.     

Phase I study of combination chemotherapy with 5-fluorouracil (5-FU) and nedaplatin (NDP): adverse effects and eecommended dose of NDP administered after 5-FU

When nedaplatin (NDP) was used as a single agent in the phase I study, the dose-limiting toxicity (DLT) was thrombocytopenia and the recommended dose (RD) was 100 mg/m2. However, the DLT, maximum tolerated dose (MTD) and RD of NDP used in combination with 5-fluorouracil remained unknown. Therefore, we performed this study to assess the DLT and RD of NDP administered after 5-fluorouracil (5-FU). In this study, 5-FU was administered to 38 patients at a fixed dose (700 mg/m2/d on days 1-5) and NDP administered on day 6 at an initial dose of 80 mg/m2, which was subsequently increased to 100, 120, 130, 140, 150, and 160 mg/m2. The DLT of NDP was leukopenia and its MTD and RD were 160 and 150 mg/m2, respectively. Concerning impairment of renal function, only two patients had a grade I increase in serum creatinine. There were 19 responders (50%, 19/38) achieving partial response or complete response in the evaluation of antitumor effect. The result of this study is notable in that administration of 5-FU before NDP allows the dose of NDP to be substantially increased.