Validation of a limited sampling model to determine etoposide area under the curve

STUDY OBJECTIVE: To validate the utility of a previously reported 3-point limited sampling model (LSM) for determining etoposide area under the curve to infinity (AUC(infinity)). DESIGN: Secondary analysis of data from two clinical trials of etoposide. SETTING: University medical center clinical research center. PATIENTS: Thirty-four patients with different malignancies. INTERVENTIONS: Etoposide was administered as a 2-hour infusion to 34 patients. Serial plasma samples were drawn over 24 hours after the infusion and analyzed for etoposide by high-performance liquid chromatography. MEASUREMENTS AND MAIN RESULTS: The 3-point LSM AUC was compared with a 14-point actual AUC calculated by the linear trapezoidal rule. Actual and predicted AUC(infinity) by the LSM were highly correlated (r=0.97, p<0.0001). The LSM predictions had a mean absolute error of 10.9% (95% CI -14.1, -5.3) and a mean error of -9.7% (95% CI 6.9, 14.9). Nine patients with poor AUC(infinity) estimations by the LSM (error > 12%) tended to have abnormally low or high peak concentrations. CONCLUSION: Our findings suggest the development of more robust LSM using other techniques, such as pharmacostatistical models, that can accommodate a greater degree of pharmacokinetic variability.     

Pharmacokinetics and distribution over the blood-brain barrier of zalcitabine (2',3'-dideoxycytidine) and BEA005 (2', 3'-dideoxy-3'-hydroxymethylcytidine) in rats, studied by microdialysis

Microdialysis was applied to sample the unbound drug concentration in the extracellular fluid in brain and muscle of rats given zalcitabine (2',3'-dideoxycytidine; n = 4) or BEA005 (2', 3'-dideoxy-3'-hydroxymethylcytidine; n = 4) (50 mg/kg of body weight given subcutaneously). Zalcitabine and BEA005 were analyzed by high-pressure liquid chromatography with UV detection. The maximum concentration of zalcitabine in the dialysate (Cmax) was 31.4 +/- 5. 1 microM (mean +/- standard error of the mean) for the brain and 238. 3 +/- 48.1 microM for muscle. The time to Cmax was found to be from 30 to 45 min for the brain and from 15 to 30 min for muscle. Zalcitabine was eliminated from the brain and muscle with half-lives 1.28 +/- 0.64 and 0.85 +/- 0.13 h, respectively. The ratio of the area under the concentration-time curve (AUC) (from 0 to 180 min) for the brain and the AUC for muscle (AUC ratio) was 0.191 +/- 0.037. The concentrations of BEA005 attained in the brain and muscle were lower than those of zalcitabine, with Cmaxs of 5.7 +/- 1.4 microM in the brain and 61.3 +/- 12.0 microM in the muscle. The peak concentration in the brain was attained 50 to 70 min after injection, and that in muscle was achieved 30 to 50 min after injection. The half-lives of BEA005 in the brain and muscle were 5.51 +/- 1.45 and 0.64 +/- 0.06 h, respectively. The AUC ratio (from 0 to 180 min) between brain and muscle was 0.162 +/- 0.026. The log octanol/water partition coefficients were found to be -1.19 +/- 0.04 and -1.47 +/- 0.01 for zalcitabine and BEA005, respectively. The degrees of plasma protein binding of zalcitabine (11% +/- 4%) and BEA005 (18% +/- 2%) were measured by microdialysis in vitro. The differences between zalcitabine and BEA005 with respect to the AUC ratio (P = 0.481), half-life in muscle (P = 0.279), and level of protein binding (P = 0.174) were not statistically significant. The differences were statistically significant in the case of the half-life in the brain (P = 0.032), clearance (P = 0.046), volume of distribution (P = 0.027) in muscle, and octanol/water partition coefficient (P = 0.019).     

Cellular interactions of 5-fluorouracil and the camptothecin analogue CPT-11 (irinotecan) in a human colorectal carcinoma cell line

CPT-11 (irinotecan) is a DNA topoisomerase I inhibitor active against metastatic colorectal carcinoma. We investigated, in a human colon carcinoma cell line, HT-29, the effects of CPT-11 and 5-fluorouracil (5FU) combinations. A strong synergism between CPT-11 and 5FU was observed after sequential exposure and only additivity or antagonism after simultaneous exposure. When cells were first exposed to 5FU, the product of cellular CPT-11 concentrations versus time (CxT) was 6895 +/- 1020 pmol x hr/10(6) cells, while it was 3875 +/- 121 pmol x hr/10(6) cells with CPT-11 alone (p < 0.01). The same phenomenon was observed with SN-38: 148.2 +/- 49.5 versus 83.4 +/- 23.6 pmol x hr/10(6) cells (p < 0.05). Consequently, the formation of protein-DNA complexes was 1.4 times greater with 5FU pretreatment than with CPT-11 alone (p = 0.03). Moreover, the incorporation of 5FU derivatives into DNA was multiplied by a factor of 1.5 24 hr after CPT-11 exposure. When cells were first incubated with CPT-11, the decrease in thymidylate synthase (TS) activity was identical to that obtained after 5FU exposure (1.09 to 0.023 pmol/min/mg protein), but this decrease persisted for 24 hr (0.014 pmol/min/mg protein) (p = 0.035). At the same time, a 1.8-fold increase in the incorporation of 5FU derivatives into DNA and a 2-fold increase in DNA-protein complex formation were evidenced. With the two sequential associations, we observed a persistent S-phase arrest, as compared with CPT-11 alone. These results suggest that CPT-11 and 5FU combinations are of clinical interest, and mechanisms of interaction between the two drugs seem to be multifactorial.     

Oestrus, ovulation and peri-ovulatory hormone profiles in tethered and loose-housed sows

Multiparous sows that had been tethered during lactation were put in two different housing conditions after weaning (Day 0); the sows were either tethered by neck chain, or individually housed in a pen of approximately 6 m2. After two months, ten tethered and eleven loose housed sows were used to assess stress and reproductive parameters. Stereotypic behaviour after the afternoon feeding was assessed from Day 18 onwards; at Day 53 stereotypic behaviour tended to occur during a higher percentage of time in the tethered sows (P = 0.11) and at Day 66, the differences were significant (tethered, 78 +/- 5 vs. loose-housed, 40 +/- 10% of time (mean +/- sem); P = 0.03). At Day 35 and 55, cortisol profiles after afternoon feeding were similar for the two groups of sows (P > 0.10). Around oestrus (approximately Day 64), the profiles of oestradiol-17 beta, luteinizing hormone and progesterone were measured and proved to be similar for both treatment groups (P > 0.10). The duration of oestrus was shorter in the tethered sows (42 +/- 4 vs. 63 +/- 2 h; P < 0.001) and, consequently, the timing of ovulation during oestrus (h after onset of oestrus) was advanced in the tethered sows (28 +/- 2 vs. 41 +/- 2; P < 0.001). The duration of ovulation did not differ (tethered, 2.9 +/- 0.5 vs. loose-housed, 2.1 +/- 0.2 h; P = 0.16). The sows were sacrificed at Day 5 after ovulation; ovulation rate, fertilization rate, embryo development and embryo diversity were similar for the two groups, as were adrenal weight and size of adrenal cortex. Duration of oestrus and the levels of stereotypies at Day 60 tended to be negatively related in the tethered sows (P = 0.07), but not in the loose-housed sows (P = 0.65). In conclusion, sows that had been tethered during pregnancy and lactation, and were housed loose or were tethered again at weaning within two months differed both in stereotypic behaviour and in duration of oestrus, without apparent effects on reproductive hormones.     

Pharmacokinetic characteristics of the novel anticancer agent CPT-11 and its active metabolite in plasma and lung lymph fluid following intravenous administration to sheep

7-Ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxycamptothecin (CPT-11, 100286-90-6) is one of the most promising novel anticancer agents, especially for lung cancer. 7-Ethyl-10-hydroxycamptothecin (SN-38), an active metabolic of CPT-11, has much stronger cytotoxicity than CPT-11. The present study was designed to evaluate the distribution and behavior of CPT-11 and SN-38 in lung lymph circulation following intravenous infusion. Awake sheep with chronically instrumented lung lymph fistulas were prepared. The concentrations of CPT-11 and SN-38 in plasma and lung lymph fluid were measured after intravenous infusion of 100 mg/body of CPT-11 for 90 min. SN-38 constantly showed higher lymph to plasma concentration ratios than those of CPT-11, and the % area under the curve (AUC) ratio of SN-38/CPT-11 in lymph fluid was significantly higher than that in plasma. These data indicated that SN-38 distributed in lung tissue moved more easily into lung lymph fluid than CPT-11, and might be more rapidly metabolized in lung tissue than plasma. CPT-11 may have favorable therapeutic effects on intrathoracic malignancies such as lung cancer and lymph metastasis.     

Effects of elevated concentrations of prostaglandin F2 alpha on pregnancy rates in progestogen supplemented cattle

An experiment was performed to determine the effect of elevated prostaglandin F2 alpha (PGF2 alpha) on pregnancy rates of progestogen-treated bred cows in the presence or absence of luteal tissue. Ninety-one beef cows were bred (Day 0) and assigned randomly to receive either 3 mL saline (CON), 15 mg PGF2 alpha, or 15 mg PGF2 alpha + lutectomy (P + L) administered intramuscularly (i.m.) at 8 h intervals on either Days 5-8, 10-13, or 15-18 postbreeding. Lutectomies were performed by transrectal digital pressure before initiation of treatment on Day 5, 10, or 15 for the respective treatment groups. All cows were fed 4 mg/day of melengesterol acetate from two days prior to initiation of treatment until Day 30 postbreeding. Mean concentrations of 13,14-dihydro-15-keto-PGF2 alpha (PGFM) were increased in cows administered PGF2 alpha and P + L treatments (398 +/- 23 and 413 +/- 22 pg/ml, respectively; p < 0.01) compared to the CON group (80 +/- 29 pg/ml) regardless of treatment group. Mean concentrations of oxytocin (OT) were increased in cows given PGF2 alpha on Day 10 and 15 (p < or = 0.0001) and tended to be increased on d 5 when compared to CON and P + L treatment groups on Day 5. Pregnancy rates were reduced (p < or = 0.03) in the PGF2 alpha treatment group (23%) and by Day 5-8 compared to CON (72%). Lutectomy tended to improve pregnancy rate in P + L (5-8; 55%) compared to PGF2 alpha (5-8; p = 0.1). Pregnancy rates tended (p < or = 0.07) to increase in the PGF2 alpha treatment groups on Days 5-8 treatment (23%, 50%, and 60% for Days 5-8, 10-13, and 15-18, respectively). The later the treatments were initiated pregnancy rates did not differ between treatments given on Days 10-13 and 15-18. In conclusion, the most susceptible period of embryonic growth to the negative effects of PGF2 alpha was during morula to blastocyst development. Removal of luteal tissue diminishes the negative effects of PGF2 alpha through interruption of the luteal oxytocin-uterine PGF2 alpha feedback loop.     

Finasteride and flutamide as potency-sparing androgen-ablative therapy for advanced adenocarcinoma of the prostate

OBJECTIVES: Androgen ablation with luteinizing hormone-releasing hormone (LHRH) agonists, orchiectomy, or oral estrogens has significant untoward sexual side effects. We evaluated a combination of finasteride and flutamide as potency-sparing androgen ablative therapy (AAT) for advanced adenocarcinoma of the prostate. In addition, we evaluated whether finasteride provided additional intraprostatic androgen blockade to flutamide. METHODS: Twenty men with advanced prostate cancer were given flutamide, 250 mg orally three times daily. Serum prostate-specific antigen (PSA) values were measured weekly. At a nadir PSA value, finasteride, 5 mg orally every day, was added. PSA values were then measured weekly until a second nadir PSA value was achieved. Sexual function was evaluated at baseline, at the second nadir PSA value, and every 3 months thereafter. Testosterone, dihydrotestosterone (DHT), and dehydroepiandrostenedione (DHEA) levels were measured at baseline and at the first and second nadir PSA values. RESULTS: The median follow-up period was 16.9 months. Therapy failed in 1 patient with Stage D2 disease at 12 months, but an additional response to subsequent LHRH agonist therapy was observed. One patient developed National Cancer Institute grade 3 diarrhea and was withdrawn from the study. Seven of 20 men developed mild gynecomastia, and 3 of 20 developed mild transient liver function test elevations. Mean PSA levels were 94.6 +/- 38.2 ng/mL at baseline and 7.8 +/- 2.7 and 4.7 +/- 2.2 ng/mL at the first and second PSA nadir values, respectively (P = 0.034). Mean percent decline in PSA value from baseline was 87.0 +/- 3.1% with flutamide alone and 94.0 +/- 1.9% with both flutamide and finasteride (P = 0.001). Eleven of 20 men were potent at baseline. At the second nadir PSA value, 9 (82%) of 11 were potent, whereas 2 (18%) of 11 were impotent. With longer follow-up (median 16.4 months), 6 (55%) of 11 men were potent, 2 (18%) of 11 were partially potent, and 3 (27%) of 11 were impotent. With flutamide alone, testosterone rose a mean of 77 +/- 14.7% of baseline (P = 0.0001), DHEA fell a mean of 32.4 +/- 4.6% (P = 0.0001), and DHT was unchanged. With the addition of finasteride, testosterone rose another 14 +/- 6% (P = 0.06, not significant), DHEA was unchanged, and DHT fell a mean of 34.8 +/- 4.7% (P = 0.0009). CONCLUSIONS: Finasteride and flutamide were safe and well tolerated as AAT for advanced prostate cancer. Finasteride provided additional intraprostatic androgen blockade to flutamide, as measured by additional PSA suppression. Sexual potency was preserved initially in most patients, although there was a reduction in potency and libido in some patients on longer follow-up. Further evaluation of this therapy is needed.     

The evaluation of eustachian tube function in patients with chronic otitis media

To assess the clinical usefulness of salivary monitoring of irinotecan (CPT-11) and its active metabolite (SN-38), we examined the clinical pharmacological profile of both drugs in 9 patients with thoracic malignancies who received 60 mg/m2 CPT-11 (21 courses). Plasma and unstimulated whole saliva were collected over a 24-h period, and concentrations of CPT-11 and SN-38 were measured by high-performance liquid chromatography. Both CPT-11 and SN-38 were detectable in saliva, and the concentration-time curves in plasma and saliva showed a very similar pattern. A good correlation was observed between the saliva concentration (C3) and the plasma concentration (Cp) for both CPT-11 and SN-38 (r = 0.732, P < 0.0001 and r = 0.611, P < 0.0001, respectively). The area under the concentration-time curve calculated for saliva (AUCs) correlated with that generated for plasma (AUCp) for both CPT-11 and SN-38 (r = 0.531, P = 0.012 and r = 0.611, P = 0.0025, respectively). These results suggest that it may be feasible to use saliva instead of plasma for pharmacokinetics/pharmacodynamics studies of CPT-11.     

Effect of 48 h treatment with 17 beta-oestradiol or progesterone on follicular wave emergence and synchrony of ovulation in Bos indicus cows when administered at the end of a period of progesterone treatment

The objective of this study was to determine the effect of treatment with additional progesterone (P4) or 17 beta-oestradiol (E2) at the end of a period of P4 treatment on ovarian follicular development, ovulation time, and plasma gonadotrophin and steroid hormone concentrations of Bos indicus cows. Initially, two injections of PGF2 alpha were given 14 days apart, and at the time of the second injection (Day 0) all cows received a single P4-releasing controlled internal drug release (CIDR) device that was removed 10 days later. Control cows (Group 1, n = 8) received no other treatment. On Day 8, cows in Group 2 (n = 8) and Group 3 (n = 8) received either a s.c. implant containing E2, or a second CIDR device, respectively. All CIDR devices and E2 implants were removed at a similar time on Day 10. Treatment with E2 or P4 delayed mean (+/- SD) time of ovulation (113.1 +/- 25.6 h, 153.4 +/- 44.5 h and 150.8 +/- 25.1 h for Groups 1, 2 and 3, respectively; P < 0.05) and the mean time (+/- SD) of the luteinising hormone (LH) peak (87.4 +/- 24.5 h, 124.3 +/- 45.0 h and 122.3 +/- 25.04 h for Groups 1, 2 and 3, respectively; P < 0.05). Both treatments delayed the mean (+/- SD) day of emergence of the ovulatory follicle (7.7 +/- 3.6 days, 11.3 +/- 1.7 days and 11.1 +/- 1.5 days for Groups 1, 2 and 3, respectively; P < 0.05), and reduced the variability in the day of emergence of the ovulatory follicle (P < 0.05) compared with the control cows. Variability in age and duration of dominance of the ovulatory follicle was greater in control animals compared with treated animals (P < 0.05). Treatment with E2 on Days 9 and 10 did not alter mean concentrations of gonadotrophins in the cows in Group 2 compared with control cows (P > 0.05), whereas treatment of cows with an additional CIDR device resulted in greater mean concentrations of FSH and lesser concentrations of LH on Day 9 (P < 0.05) compared with cows in Groups 1 and 2. By Day 10 mean concentrations of gonadotrophins were similar among cows in all three groups. Concentrations of E2 were less in cows in Group 3 compared with cows in Groups 1 and 2 from Day 9 to Day 11 (P < 0.05). We conclude that treatment with either E2 or P4 can influence the pattern of ovarian follicular development and ovulation in cattle; however, the mechanism of action of the two treatments may differ. Atretogenic treatments for ovarian follicles applied at the end of a period of progesterone treatment did not improve synchrony of ovulation.     

Unexpected hepatotoxicities in patients with non-Hodgkin's lymphoma treated with irinotecan (CPT-11) and etoposide

BACKGROUND: Irinotecan (CPT-11) is a topoisomerase I inhibitor that has been confirmed to be active against a broad spectrum of neoplasms including non-Hodgkin's lymphoma (NHL). Because the combination of topoisomerase I and II inhibitors seemed to be an attractive therapeutic strategy owing to their complementary functions, we conducted a combination phase I study of CPT-11 and etoposide, a topoisomerase II inhibitor, in relapsed or refractory non-Hodgkin's lymphoma (NHL). METHODS: The starting doses of CPT-11 and etoposide were 30 mg/m2/day (days 1-3 and 8-10) and 40 mg/m2 (days 1-3), respectively. RESULTS: All three patients who received the starting dose developed dose-limiting toxicities including one case of grade 4 neutropenia lasting for > 7 days, one of grade 3 serum transaminase elevation and one of grade 3 hyperbilirubinemia. All three patients presented hepatotoxicity > or = grade 2. The starting dose level was judged to be the maximum tolerated dose (MTD) and further dose escalation of this combination was halted. The patient who developed grade 3 hyperbilirubinemia presented a second peak of plasma SN-38, an active metabolite of CPT-11, on the concentration-time curve for day 3, suggesting the possibility of the enterohepatic circulation of SN-38 and of a drug-to-drug interaction. No durable objective response was observed in the three patients treated at the starting dose. CONCLUSIONS: We conclude that etoposide is not recommended for combination with CPT-11 in NHL patients because of unexpected frequent hepatotoxicities.     

Determinants of CPT-11 and SN-38 activities in human lung cancer cells

Irinotecan (CPT-11) is a semisynthetic camptothecin derivative with a broad spectrum of anti-tumour activity. Carboxylesterase (CE) catalyses the conversion of CPT-11 to SN-38 (7-ethyl-10-hydroxycamptothecin), the active form of CPT-11. The antiproliferative effects of CPT-11 and SN-38, CE-activity and topoisomerase I protein expression were investigated in five human small-cell lung cancer (SCLC) cell lines and four human non-small-cell lung cancer (NSCLC) cell lines. Antiproliferative activity, expressed as IC50 values, was determined using the MTT assay. CPT-11 was significantly more active in SCLC than in NSCLC cell lines (P = 0.0036), whereas no significant difference between histological types was observed with SN-38. A significant correlation (r2 = 0.52, P = 0.028) was observed between CE activity and chemosensitivity to CPT-11 but not to SN-38, and significantly higher CE activity was observed in SCLC compared with NSCLC cell lines (P = 0.025). Western blotting experiments showed topoisomerase I protein expressions within a factor of 2, and a granular nuclear staining was detectable in all cell lines by immunocytochemistry of cytospins. No correlation was observed between protein expression and sensitivity to CPT-11 or SN-38. Cellular and medium concentrations of CPT-11 and SN-38 were measured by high-performance liquid chromatography (HPLC) in one SCLC cell line with high CE activity and high sensitivity to CPT-11, and one NSCLC cell line with low sensitivity to CPT-11 and CE activity. Intracellular concentrations of CPT-11 and SN-38 were higher in the SCLC cell line, and this was associated with an increase in cellular uptake of CPT-11 compared with the medium, and an increased intracellular formation of SN-38. In conclusion, CE activity appears to be associated with higher sensitivity to CPT-11 in human lung cancer cell lines and may partly explain the difference in the in vitro sensitivity to CPT-11 between SCLC and NSCLC cells. The assessment of CE activity in clinical material of lung cancer patients undergoing treatment with CPT-11 may be warranted. However, other mechanisms may influence sensitivity to CPT-11, possibly including drug transport.     

Methylprednisolone pharmacokinetics, cortisol response, and adverse effects in black and white renal transplant recipients

It is generally assumed that chronic glucocorticoid therapy is similar pharmacologically when administered to either black or white renal transplant recipients, resulting in adrenal suppression, low circulating plasma cortisol concentrations, and a similar degree of drug exposure and toxicity. To examine this theory and to investigate the relationship of glucocorticoid metabolism to steroid-induced adverse effects among specific ethnic groups of renal transplant recipients, 9 black and 9 white male patients chronically receiving methylprednisolone were enrolled. All patients had stable renal function and were matched for age, weight, and time since transplant. Standard pharmacokinetic parameters for methylprednisolone were determined and cortisol responses were characterized by total cortisol area under the concentration curve (AUC), return cortisol AUC, and cortisol suppression half-life. All patients received their daily oral dose of methylprednisolone (mean daily dose = 11 mg for blacks and 11 mg for whites) as an intravenous infusion with serial plasma samples obtained over 24 h. The patients were assessed for the presence of specific cushingoid manifestations (buffalo hump, moon facies) and steroid-associated diabetes. Methylprednisolone and cortisol were analyzed via HPLC. In the black patients, the mean clearance of methylprednisolone (206 +/- 70 ml/hr/kg) was significantly slower with a smaller volume of distribution (0.95 +/- 0.32 L/kg) when compared with the white group (327 +/- 129 ml/hr/kg, P = 0.03; volume of distribution = 1.33 +/- 0.27 L/kg, P = 0.015). Despite chronic methylprednisolone therapy, a definite 24-hr cortisol response pattern was noted in 15 of the 18 patients with a mean total cortisol AUC of 732 +/- 443 ng.hr/ml in blacks and 539 +/- 361 ng.hr/ml in whites (P = 0.17, black vs. white). The mean cortisol suppression half-life was 4.31 +/- 1.54 hr in black recipients and 4.11 +/- 1.49 hr in whites (P = 0.48). The mean return cortisol AUC for the black patients was 327 +/- 279 ng.hr/ml and 370 +/- 207 ng.hr/ml for white patients (P = 0.28). The serum cortisol nadir for black patients was 12.3 +/- 7.2 ng/ml, which was significantly higher than the cortisol nadir in white patients (6.4 +/- 4.4 ng/ml; P = 0.03). A majority (94%) of patients (9 black, 8 white) had moon facies and 27% of patients (3 black, 1 white) had a buffalo hump. While 5 of 9 black patients had steroid-associated diabetes, no white patients manifested this adverse effect. The black patients with diabetes had higher cortisol AUCs with lower methylprednisolone clearances than the white group.(ABSTRACT TRUNCATED AT 400 WORDS)     

Levels and patterns of alcohol consumption using timeline follow-back, daily diaries and real-time "electronic interviews"

BACKGROUND: Basic knowledge of the substances involved in wound healing after photorefractive keratectomy (PRK) is essential for development of pharmacological intervention. We present preoperative and postoperative analysis of tear fluid extracellular matrix proteins and cytokines after PRK. METHODS: Tear fluid samples from 70 patients (72 eyes) who had PRK (38 women and 32 men, mean age 31.5 yr) were studied. Samples from 18 patients (18 eyes) were analyzed in two different studies. RESULTS: Mean preoperative tear fluid flow in the collection capillary (volume divided by tear collection time) varied from 4.5 to 22.5 microliters/min in five series of patients. It increased significantly during the first two postoperative days (range of means, 55.5 to 88.8 microliters/min, p < 0.01), and decreased to the preoperative level by day 7 (range of means, 9.7 to 18.2 microliters/min). The tenascin and cytokine release rates increased significantly during the first two days after PRK and returned to the preoperative level by day 7. Mean +/- standard error for tenascin: day 0 (5.2 +/- 1.9 ng/min); day 2 (22.7 +/- 6.1 ng/min; p = 0.02). Mean +/- standard error for HGF: day 0 (3.2 +/- 0.7 pg/min); day 1 (22.8 +/- 4.2 pg/min; p = 0.0003). Mean +/- standard error for TGF-beta 1: day 0 (63.3 +/- 19.6 pg/min); days 1-2 (826.2 +/- 253.7 pg/min; p = 0.001). Mean +/- standard error for VEGF: day 0 (166.0 +/- 29.6 pg/min); days 1-2 (824.4 +/- 165.1 pg/min; p = 0.0007). Mean +/- standard error for PDGF-BB: day 0 (0.42 +/- 0.19 pg/min); day 2 (27.6 +/- 5.8 pg/min; p = 0.0000). Mean +/- standard error for TNF-alpha: day 0 (9.5 +/- 2.6 pg/min); day 2 (28.6 +/- 5.9 pg/min; p = 0.003). Excluding PDGF-BB, all substances studied were present in normal human tear fluid. PDGF-BB was present in only 17% of the preoperative samples. CONCLUSION: Corneal wounding induces an increased release of several growth modulating cytokines which may be involved in healing processes.     

Histamine and eicosanoid levels in plasma and peripheral blood leucocyte cultures during experimental infection of cattle with bluetongue virus serotype 11

Three calves were sensitized with three doses of inactivated BTV-11 UC8 strain and then experimentally infected with the homologous virus. In addition, four BTV-seronegative heifers were also experimentally infected with BTV-11. Granulocyte rich fractions of peripheral blood leucocyte (PBL-GRF) cultures from BTV-sensitized/infected calves and from control unexposed cattle were exposed in vitro with BTV-11. Histamine, leukotriene (LT) C4 and prostaglandin (PG) D2 were assayed in supernatant fluids. Plasma histamine levels increased in BTV-infected heifers from 10.1 +/- 2 ng/ml at Day 0 to 23.1 +/- 6.6 ng/ml at Day 12 following virus exposure. In addition, in this experimental group the concentration of PGF2 alpha (mean 551.97 +/- 243.54 pg/ml) increased significantly (P < or = 0.05) compared with control cattle (mean 467.3 +/- 73.9 pg/ml). Bluetongue virus induced histamine and LTC4 release after in vitro infection of PBL-GRF. Release of LTC4 was significantly (P < or = 0.05) higher in PBL-GRF cultures from sensitized and control animals than in unexposed PBL-GRF cultures. In contrast to these results, PGD2 was not released after BTV infection of PBL-GRF in vitro. The histamine release caused by BTV was virus-specific and mainly mediated by an immunological reaction, since the release was significantly reduced by removal of cell surface immunoglobulins.     

Pediatric hepatic trauma: does clinical course support intensive care unit stay?

PURPOSE: The objective of this study is to determine if grade of liver injury predicts outcome after blunt hepatic trauma in children and to initiate analysis of current management practices to optimize resource utilization without compromising patient care. METHODS: A retrospective review of 36 children who had blunt hepatic trauma treated at a pediatric trauma center from 1989 to present was performed. Hepatic injuries graded (AAST Organ Injury Scaling) ranged from grade I to IV. Injury Severity Score (ISS), Glasgow Coma Score (GCS), transfusion requirements, liver transaminase levels, associated injuries, intensive care unit (ICU) length of stay, and survival were analyzed. RESULTS: Mean (+/-SEM) age was 6.6+/-0.8 years, mean grade of hepatic injury was 2.4+/-0.2, mean ISS was 17+/-2.6, mean GCS was 13+/-1, and mean transfusion was 15.4 mL/kg of packed red blood cells (PRBC). There were three deaths with a mean ISS of 59+/-9 and a mean GCS of 3+/-0. Death was not associated with a high-grade liver injury, survivors versus nonsurvivors, 2.3+/-0.2 versus 2.7+/-0.3, but was associated with ISS, 13+/-1.4 versus 59+/-9 (P = .005) and GCS, 14+/-1 versus 3+/-0 (P = .005). Only one patient (grade III, ISS = 43) underwent surgery. There were no differences in mean ISS or GCS between grades I to IV patients. The hepatic injury grades of patients requiring transfusion versus no transfusion were significantly different, 3.4+/-0.2 versus 2.2+/-0.2 (P = 0.04). Abused patients had high-grade hepatic injuries and significant laboratory and clinical findings. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were significantly higher in grade III and IV injuries than in grades I and II, 1,157+/-320 versus 333+/-61 (P= .02) and 1,176+/-299 versus 516+/-86 (P= .04), respectively. No children with grade I or II injury had a transfusion requirement or surgical intervention. There were no liver-related complications. CONCLUSIONS: Mortality and morbidity rates in pediatric liver injuries, grades I to IV, correlate with associated injuries not the degree of hepatic damage. ALT, AST, and transfusion requirements are significantly related to degree of liver injury. Low-grade and isolated high-grade liver injuries seldom require transfusion. Blunt liver trauma rarely requires surgical intervention. In retrospect, the need for expensive ICU observation for low-grade and isolated high-grade hepatic injuries is questionably warranted.     

Outcome of cardiac valve ring abscesses after medical treatment: attempt to identify criteria of favorable prognosis

OBJECTIVES: Identify factors predicting favorable outcome after medical management of valve ring abscesses in order to propose a surveillance schedule for conservative treatment. METHODS: A multicentric study conducted from July 1989 to February 1996 included 28 patients (mean age 64 +/- 16 years, range 26-83) hospitalized for active endocarditis and valve ring abscesses diagnosed at transthoracic or transesophageal echography. Conservative medical therapy was given because of a decision of the medico-surgical team (n = 9), high surgical risk (n = 12), or patient refusal of surgery (n = 7). Outcome was favourable in 18 patients (Group I) and unfavorable in 10 (Group II) due to death (n = 9) or subsequent surgery (n = 1). Univariate and multivariate analysis were used to determine differences between the groups in terms of clinical and laboratory data. RESULTS: Mean follow-up in Group I was 33 +/- 18 months and 15 +/- 10 months in Group II. Univariate analysis showed significant differences between Group I and II respectively for age (59 +/- 18 yr vs 72 +/- 10, p = 0.04), delay to apyrexia after antibiotics (4.3 +/- 2.8 vs 8.3 +/- 2.4 days, p < 0.0008), heart failure (5% vs 70%, p = 0.003), grade III or IV valvular regurgitation (5% vs 60%, p < 0.04), and mean surface area of the abscess (1.5 +/- 1.2 vs 5.4 +/- 6.4 cm2, p < 0.03). Independent factors at multivariate analysis were by decreasing order: lack of heart failure at admission, delay to apyrexia, abscess surface area, and age. Outcome was favorable (mean follow-up 33 +/- 10 months) in all patients with an abscess surface area < 1.5 cm2, no signs of heart failure, no grade III or IV valvular regurgitation, apyrexia after less than 8 days on antibiotics and no staphylococcus positive blood culture. CONCLUSION: Medical management of valve ring abscesses may be indicated in selected patients in care units with rigorous surveillance facilities. Further studies are needed to precisely identify surveillance and treatment criteria.     

Plasma pharmacokinetics of 7-ethyl-10-hydroxycamptothecin (SN-38) after intravenous administration of SN-38 and irinotecan (CPT-11) to rats

We studied the plasma pharmacokinetics of the lactone form and the carboxylate form of 7-ethyl-10-hydroxycamptothecin (SN-38), the active metabolite of irinotecan (CPT-11), after intravenous bolus administrations of each form of SN-38 and of CPT-11 to rats. After the SN-38 lactone injection, SN-38 lactone was predominant at first, and then the lactone to the carboxylate concentration ratio (LC ratio) was maintained from 30 to 90 min after the injection. The carboxylate was predominant throughout the period after the carboxylate dosing. Model-dependent analyses showed that the SN-38 lactone had greater plasma clearance and a greater distribution volume than the carboxylate. The CPT-11 administration resulted in a predominant plasma SN-38 lactone concentration. The contribution of the SN-38 lactone AUC to the total SN-38 AUC (57%) was independent of the dose of CPT-11. These results suggest that it is possible to estimate the SN-38 lactone concentration and AUC from the total SN-38 concentration without separate determination of the lactone and carboxylate. Our results showed that both SN-38 lactone and CPT-11 administration gave the predominant SN-38 lactone in plasma; however, only CPT-11 could sustain the lactone concentration at a high level, which is necessary for antitumor activity.     

Identification and properties of a major plasma metabolite of irinotecan (CPT-11) isolated from the plasma of patients

Irinotecan [7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11)] is a promising water-soluble analogue of camptothecin [S. Sawada et al., Chem. & Pharm. Bull. (Tokyo), 39: 1446-1454, 1991]. We have reported previously the presence of an important polar metabolite, in addition to 7-ethyl-10-hydroxycamptothecin (SN-38) beta-glucuronide, in samples of plasma taken from patients undergoing treatment with CPT-11 (L.P. Rivory and J. Robert, Cancer Chemother. Pharmacol. 36: 176-179, 1995; L. P. Rivory and J. Robert, J. Cromatogr., 661: 133-141, 1994). Plasma samples (0.5 ml) containing comparatively large amounts of this metabolite were extracted by solid-phase columns and subjected to high-performance liquid chromatography and mass spectrometry in parallel to fluorometric detection. The metabolite yielded [M + 1] ions with a m/z of 619, representing the addition of 32 atomic mass units to CPT-11. Purified fractions were subjected to proton nuclear magnetic resonance, and the structure determined, 7-ethyl-10-[4-N-(5-aminopentanoic acid)-1-piperidino]carbonyloxycampothecin (APC), was further validated following synthesis. Like CPT-11, APC was found to be only a weak inhibitor of the cell growth of KB cells in culture (IC50, 2.1 versus 5.5 micrograms/ml for CPT-11 and 0.01 microgram/ml for SN-38, the active metabolite of CPT-11) and was a poor inducer of topoisomerase I DNA-cleavable complexes (100-fold less potent than SN-38). In contrast to CPT-11, APC was not hydrolyzed to SN-38 by human liver microsomes or purified human liver carboxylesterase. Furthermore, APC did not inhibit the hydrolysis of CPT-11 in these preparations. Interestingly, APC was only a weak inhibitor of acetylcholinesterase in comparison to CPT-11 and neostigmine. It appears likely, therefore, that APC does not contribute directly to the activity and toxicity profile of CPT-11 in vivo.     

Regional differences in action potential characteristics and membrane currents of guinea-pig left ventricular myocytes

Regional differences in action potential characteristics and membrane currents were investigated in subendocardial, midmyocardial and subepicardial myocytes isolated from the left ventricular free wall of guinea-pig hearts. Action potential duration (APD) was dependent on the region of origin of the myocytes (P < 0.01, ANOVA). Mean action potential duration at 90 % repolarization (APD90) was 237 +/- 8 ms in subendocardial (n = 30 myocytes), 251 +/- 7 ms in midmyocardial (n = 30) and 204 +/- 7 ms in subepicardial myocytes (n = 36). L-type calcium current (ICa) density and background potassium current (IK1) density were similar in the three regions studied. Delayed rectifier current (IK) was measured as deactivating tail current, elicited on repolarization back to -45 mV after 2 s step depolarizations to test potentials ranging from -10 to +80 mV. Mean IK density (after a step to +80 mV) was larger in subepicardial myocytes (1.59 +/- 0.16 pA pF-1, n = 16) than in either subendocardial (1.16 +/- 0.12 pA pF-1, n = 17) or midmyocardial (1. 13 +/- 0.11 pA pF-1, n = 21) myocytes (P < 0.05, ANOVA). The La3+-insensitive current (IKs) elicited on repolarization back to -45 mV after a 250 ms step depolarization to +60 mV was similar in the three regions studied. The La3+-sensitive tail current, (IKr) was greater in subepicardial (0.50 +/- 0.04 pA pF-1, n = 11) than in subendocardial (0.25 +/- 0.05 pA pF-1, n = 9) or in midmyocardial myocytes (0.38 +/- 0.05 pA pF-1, n = 11, P < 0.05, ANOVA). The contribution of a Na+ background current to regional differences in APD was assessed by application of 0.1 microM tetrodotoxin (TTX). TTX-induced shortening of APD90 was greater in subendocardial myocytes (35.7 +/- 7.1 %, n = 11) than in midmyocardial (15.7 +/- 3. 8 %, n = 10) and subepicardial (20.2 +/- 4.3 %, n = 11) myocytes (P < 0.05, ANOVA). Regional differences in action potential characteristics between subendocardial, midmyocardial, and subepicardial myocytes isolated from guinea-pig left ventricle are attributable, at least in part, to differences in IK and Na+-dependent currents.     

Alterations in growth and body composition during puberty: III. Influence of maturation, gender, body composition, fat distribution, aerobic fitness, and energy expenditure on nocturnal growth hormone release

We examined the relationships among gender, sexual maturation, four-compartment model estimates of body composition, body fat distribution (magnetic resonance imaging for abdominal visceral fat and anthropometrics), aerobic fitness, basal and total energy expenditure, and overnight GH release in an ultrasensitive chemiluminescence assay in healthy prepubertal and pubertal boys (n = 18 and 11, respectively) and girls (n = 12 and 18, respectively). Blood samples were withdrawn every 10 min from 1800-0600 h to determine the area under the serum GH-time curve (AUC), sum of the GH peak heights (sigma GH peak heights), and the mean nadir GH concentration. GH release was greater in the pubertal than prepubertal subjects due to an increase in sigma GH peak heights (43.8 +/- 3.6 vs. 24.1 +/- 3.5 ng.mL-1, P = 0.0002) and mean nadir (1.7 +/- 0.2 vs. 0.7 +/- 0.2 ng.mL-1, P = 0.0002), but not peak number (4.3 +/- 0.2 vs. 4.5 +/- 0.2). The girls had a greater sigma GH peak heights (39.0 +/- 3.5 vs. 28.8 +/- 3.6 ng.mL-1, P = 0.05) and mean nadir concentration (1.4 +/- 0.2 vs. 0.9 +/- 0.2 ng.mL-1, P = 0.05) than the boys. Significant inverse relationships existed between sigma GH peak heights (r = -0.35, P = 0.06) or mean nadir (r = -0.39, P = 0.04) and four-compartment percent body fat for all boys but not for all girls or when combining all subjects. For all girls, significant inverse relationships existed between sigma GH peak heights (r = -0.39, P = 0.03) or mean nadir (r = -0.37, P = 0.04) and waist/hip ratio. Similar inverse relationships in all boys or all subjects were not significant. Forward stepwise regression analysis determined that bone age (i.e. maturation, primary factor) and gender were the significant predictors of AUC, sigma GH peak heights, and mean nadir. The influence of maturation reflects rising sex steroid concentrations, and the gender differences appear to be because of differences in estradiol concentrations rather than to body composition or body fat distribution.