Assignment of Opsonic Values to Pneumococcal Reference Serum 007sp for Use in Opsonophagocytic Assays for 13 Serotypes.

Opsonophagocytic assays (OPAs) are routinely used for assessing the immunogenicity of pneumococcal vaccines, with OPA data often being utilized for licensure of new vaccine formulations. However, no reference serum for pneumococcal OPAs is available, making evaluation of data among different laboratories difficult. This international collaboration was initiated to (i) assign consensus opsonic indexes (OIs) to FDA pneumococcal reference serum lot 007sp (here referred to as 007sp) and a panel of serum samples used for calibration of the OPA and (ii) determine if the normalization of the OPA results obtained with test samples to those obtained with 007sp decreases the variability in OPA results among laboratories. To meet these goals, six participating laboratories tested a panel of serum samples in five runs for 13 serotypes. For each serum sample, consensus OIs were obtained using a mixed-effects analysis of variance model. For the calibration serum samples, normalized consensus values were also determined on the basis of the results obtained with 007sp. For each serotype, the overall reduction in interlaboratory variability was calculated by comparing the coefficients of variation of the unadjusted and the normalized values. Normalization of the results substantially reduced the interlaboratory variability, ranging from a 15% reduction in variability for serotype 9V to a 64% reduction for serotype 7F. Normalization also increased the proportion of data within 2-fold of the consensus value from approximately 70% (average for all serotypes) to >90%. On the basis of the data obtained in this study, pneumococcal reference standard lot 007sp will likely be a useful reagent for the normalization of pneumococcal OPA results from different laboratories. The data also support the use of the 16 FDA serum samples used for calibration of the OPA as part of the initial evaluation of new assays or periodic assessment of established assays.

Snapshot coherence-gated direct wavefront sensing for multi-photon microscopy.

Deep imaging in turbid media such as biological tissue is challenging due to scattering and optical aberrations. Adaptive optics has the potential to compensate the tissue aberrations. We present a wavefront sensing scheme for multi-photon scanning microscopes using the pulsed, near-infrared light reflected back from the sample utilising coherence gating and a confocal pinhole to isolate the light from a layer of interest. By interfering the back-reflected light with a tilted reference beam, we create a fringe pattern with a known spatial carrier frequency in an image of the back-aperture plane of the microscope objective. The wavefront aberrations distort this fringe pattern and thereby imprint themselves at the carrier frequency, which allows us to separate the aberrations in the Fourier domain from low spatial frequency noise. A Fourier analysis of the modulated fringes combined with a virtual Shack-Hartmann sensor for smoothing yields a modal representation of the wavefront suitable for correction. We show results with this method correcting both DM-induced and sample-induced aberrations in rat tail collagen fibres as well as a Hoechst-stained MCF-7 spheroid of cancer cells.

Establishment of a new human pneumococcal standard reference serum, 007sp.

Lot 89SF has been the reference standard serum pool used in pneumococcal enzyme-linked immunosorbent assays (ELISAs) since 1990. In 2005, it was estimated that there remained between 2 and 5 years' supply of lot 89SF. Since lot 89SF was the reference standard used in the evaluation of the seven-valent pneumococcal conjugate vaccine Prevnar (PCV7), the link to clinical efficacy would be severed if stocks became completely depleted. Furthermore, demonstration of immune responses comparable to those elicited by PCV7 is a licensure approach used for new pneumococcal conjugate vaccines, so a replacement reference standard was required. A total of 278 volunteers were immunized with the 23-valent unconjugated polysaccharide vaccine Pneumovax II, and a unit of blood was obtained twice within 120 days following immunization. Plasma was prepared, pooled, and confirmed to be free from hepatitis B virus (HBV), hepatitis C virus (HCV), and HIV. The pooled serum was poured at 6 ml per vial into 15,333 vials and lyophilized. Immunological bridging of 007sp to 89SF was used to establish equivalent reference values for 13 pneumococcal capsular serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F and 23F) by five independent laboratories. Antibody concentrations in 007sp were established relative to the lot 89SF reference preparation using the WHO reference ELISA. Subsequently, 12 existing WHO calibration sera had concentrations reassigned for 13 pneumococcal serotypes using new serum 007sp as the reference, and these were compared to concentrations relative to the original reference serum. Agreement was excellent for the 12 WHO calibration sera. The 007sp preparation has replaced 89SF as the pneumococcal reference standard. Sufficient quantity of this new preparation is available such that, with judicious use, it should be available for at least 25 years.

Renal transplantation in children younger than 6 years old.

Herein we report our experience in renal transplantation in 38 children (40 transplants), ages 1 to 5 years, between 1989 and 2005. Demographics as well as patient and graft survivals are reported. Mean age at transplantation was 3.3 +/- 1.3 years, and mean weight was 14 kg (range, 5.7-25 kg); 92.5% were Caucasian, 7.5% African-Brazilian. The main etiology for end-stage renal disease (ESRD) was uropathic/vesicoureteral reflux (45%) followed by glomerulopathy (25%), congenital/hereditary diseases (10%), and hemolytic uremic syndrome (12.5%). Prior to transplantation, 5% were on hemodialysis, 85% on peritoneal dialysis, and 10% preemptive. All children were followed for at least 6 months posttransplantation, except 2 who died in the first month. In 75% of cases, kidneys were obtained from living-related donors, and in 25% from deceased donors. Thirty-nine kidneys were extraperitoneally placed. Primary immunosuppressant therapy consisted of cyclosporine (61%), tacrolimus (39%), mycophenolate (49%), and azathioprine (51%). A steroid-free protocol was used in 17% of patients. In the last 21 cases, basiliximab or daclizumab was added. There were 13 (32.5%) graft losses (4 artery/vein thromboses, 3 chronic rejections, 3 deaths, 3 other causes). The 5-year patient and graft survival rates were 89.6% and 72.2%. We have concluded that renal transplantation can be performed with good long-term results in children younger than 6 years old.

300 pediatric renal transplantations: A single-center experience.

UNLABELLED: Our objective was to relate the results of 300 consecutive kidney transplants performed in children at a single center. PATIENTS AND METHODS: An analysis of kidney transplants was performed on patients less than 18 years old engrafted from May 1977 to August 2005. RESULTS: Among 300 kidney transplants, 48% of the patients were female, 87% were Caucasian, and 13% were African-Brazilian. The mean age at transplant was 11.5 +/- 4.5 years with 39 (13%) less than 6 years of age. The most frequent etiology of renal failure was vesicoureteral reflux/obstructive uropathy (36%) followed by glomerulopathy (27%). The donor was deceased in 32.3% and living related in 77.7% (parents 82%). The mean posttransplant follow-up was 4.8 +/- 4.3 years. The initial immunosuppression was CyA + AZA + PRED in 45%; CyA + MMF + PRED in 9.6%; TAC + AZA + PRED in 7.3%; TAC + MF + PRED in 9.7%; or TAC + MF without PRED in 10%. Sirolimus was employed initially in three cases. Induction with OKT3/ATG occurred in three patients and 112 received an anti-IL2 receptor antibody. The 103 graft losses during 28 years of follow-up were secondary to chronic allograft nephropathy in 51 (49.5%), vascular thrombosis in 5 (4.8%), acute rejection in 12 (11.6%), and recurrence of original disease in 13 (12.6%). Sixteen (15.5%) died with functioning grafts. Graft survival in the first, fifth, and tenth year were 90%, 72%, and 59%, respectively. Patient survival in the first, fifth, and tenth years were 95%, 93%, and 85%, respectively, with infection as the main cause of death.

Conversion to sirolimus in pediatric renal transplantation recipients.

We retrospectively evaluated the efficacy and safety of sirolimus (SRL) in 16 pediatric renal transplant recipients, who were 9.4 +/- 4.1 years of age when they first received SRL. The indications for SRL therapy were rescue from steroid-resistant acute rejection (31.3%), neoplasia (31.3%), diabetes (12.5%), polyomavirus-associated nephropathy (6.3%), chronic allograft dysfunction (6.3%), calcineurin inhibitor nephrotoxicity (6.3%), and hemolytic uremic syndrome (6.3%). Mean follow-up after the switch to SRL was 17.7 +/- 15 months. The final immunosuppression was CNI + SRL + prednisone (PRED) in five patients, SRL + PRED in six, SRL + mycophenolate mofetil (MMF) + PRED in four, and SRL + MMF in one. The use of SRL in these selected pediatric renal recipients was successful, except when creatinine was high at the moment of conversion. Further studies are necessary to assess the beneficial outcomes versus adverse events among the pediatric transplant population receiving SRL for immunosuppression.

Plasmapheresis for recurrent posttransplant focal segmental glomerulosclerosis.

The high recurrence rate of focal segmental glomerulosclerosis (FSGS) in kidney transplant recipients suggests that such patients have a circulating factor that alters glomerular capillary permeability. Serum from patients with FSGS increases glomerular permeability to albumin. This permeability factor has been partially identified as a protein. The removal of this protein by plasmapheresis (PP) decreases proteinuria. In this study we report data on the therapeutic effects of PP in FSGS children with recurrence in the transplanted kidney. Three hundred pediatric (age <19 years) renal transplants were performed, including 21 patients (24 transplants) with FSGS as a cause of renal failure. Fourteen (58.3%) subjects experienced disease recurrence (proteinuria >1 g/m(2) per day) within 1 month after transplantation. Mean age patient was 12 +/- 4.3 years, including 83.3% Caucasians and 70.2% recipients of living donor grafts. Nine were treated with 10 cycles of PP (3 cycles/weekly), initiated immediately after recurrence (<48 hours). Immunosuppression included high doses of cyclosporine (C(2) levels of 1700-1800 ng/mL), mycophenolate sodium or mofetil, and prednisone. Thirteen patients were induced with anti-IL2 receptor monoclonal antibody (daclizumab/basiliximab). Among the patients who underwent PP, five (55.5%) achieved a complete remission and one (12%), a partial remission (1 g/24 hours). There were no cases of remission among the five patients who were not treated with PP. Those who achieved remission after PP experienced no recurrences during the 2.6 +/- 1.4 years follow-up. PP appears to be effective to treat recurrent FSGS following kidney transplantation. It should be started as soon as possible.

Optimal designs for the individual and joint exposure general logistic regression models.

Interest in administering compounds in combination lies both in enhancing efficacious effects and in limiting adverse effects. Although much statistical work has focused on developing mathematical functions to model the joint dose-response curves, relatively little work exists in regard to designing experiments for assessing joint action. A variety of parametric dose-response models based on either the normal or logistic probability distribution have been proposed in the literature. These models are typically nonlinear in the parameters, and as such, a nonlinear weighted least squares approach can be employed for the purpose of designing experiments. The approach is applicable across a wide variety of settings commonly associated with joint action data, including continuous and discrete responses, alternative error structures, and nonzero background response. Further, designs can be expressed in terms of proportionate responses associated with the individual compounds rather than dose levels, thereby providing for results that are applicable across compounds. As a precursor to this effort, optimal and minimal experimental designs for the case in which a single compound is administered have also been developed. Although the proposed methodology for deriving experimental designs can be applied to any nonlinear regression model, primary focus is given to the additive and nonadditive independent joint action (IJA) models for individual and combined exposures proposed by Barton, Braunberg, and Friedman (1).

Treatment of nonepileptic psychogenic events in adolescent patients using the systems model for intervention.

Nonepileptic psychogenic events in children, adolescents and adults can be difficult to treat. Using a systems framework intervention method resulted in significant reduction in nonepileptic psychogenic events. This method provides for a continuous flow of interaction, feedback and modification among the health care team, family and community. Common goals and plans are defined and implemented with a mechanism for feedback and modification. Using a systems model approach in educating the caregivers resulted in immediate and long-term reduction in nonepileptic psychogenic events.

Application of the tukey trend test procedure to assess developmental and reproductive toxicity. I. Measurement data.

Developmental and reproductive (DAR) toxicity studies typically include a series of increasing doses of a compound and a zero dose control. Given this framework, Tukey et al. (Biometrics, 41, 295-301, 1985) proposed a procedure (referred to as either the Tukey trend or TCH test procedure) for detecting a nonzero trend in response to increasing doses of the test compound. The procedure considers three candidate dosage scalings to ensure high power against relatively common dose-response patterns and appreciable power against most reasonable patterns. For toxicologic effects with near monotonic dose-response patterns, simulation studies have shown the TCH test to be overall more powerful than pairwise comparison procedures. The TCH test can be applied sequentially, eliminating the highest dose each time a statistically significant trend is observed, until a no-statistical-significance-of-trend dose is reached. This is the highest dose through which there is no statistically trustworthy evidence of the compound's impact on the response. Since DAR toxicity usually exhibits a progressive (monotonic) dose-response, we advocate routine use of Tukey's trend test for the evaluation of treatment effects in these studies. In this article, we discuss the procedure in detail and apply it to fetal body weight, a continuous measurement variable, from a developmental toxicity study.

Reversible effects of triamcinolone and lack of effects with aspirin or L-656,224 on external genitalia of male Sprague-Dawley rats exposed in utero.

Inhibitors of the arachidonic acid cascade were given to pregnant rats during the critical period for morphogenesis of the external genitalia. Groups treated subcutaneously (s.c.) with 0.1 or 0.25 mg/kg/day of triamcinolone acetonide (TA) on gestational days (GD) 14-19 had male fetuses on GD 20 with moderate decreases in absolute anogenital distance (AGD), but gross and histological examinations revealed no alterations to the genital tubercle (i.e., no hypospadias). The s.c. coadministration of arachidonic acid at 100 mg/kg/day had minimal to no effect on AGD in the TA-exposed groups. No effect on AGD was observed in male fetuses from groups administered aspirin orally at 150 mg/kg/day, and only a 6% decrease was observed in the 300-mg/kg/day group. Neither TA nor aspirin adversely affected AGD of female fetuses. In another study, TA was administered on GD 11-19 at dose levels of 0.05 and 0.1 mg/kg/day, and dams were allowed to deliver. High-dose male offspring examined on postcoitum day (PCD) 23, had moderate decreases in AGD. In both studies with TA, there were also significant decreases in offspring weights. The contribution of the decreased weight to the decrease in absolute AGD was examined by a variety of methods (ratio of AGD to cube root of weight or biparietal distance, comparison to weight-matched controls, and covariance analysis). We conclude that TA caused a specific decrease in AGD on GD 20 that was largely reversed by PCD 23. When examined as adults (8 weeks old), the external genitalia of TA-exposed offspring were normal. Thus, the TA-induced decreases in AGD on GD 20 did not predict irreversible malformation. TA also caused other effects, which included a somewhat flattened genital tubercle and apparently thinned and glossy skin between the tubercle and the anus in both sexes on GD 20 and PCD 23, but not as adults. In addition, there were high pup mortality and high incidences of micrognathia and omphalocele (in the 0.25-mg/kg/day group only). Aspirin at 75 or 150 mg/kg/day and a specific lipoxygenase inhibitor (L-656,224) at 1,000 or 2,000 mg/kg/day were also administered from GD 14 to 19, and no offspring effects were observed. Thus, of the three agents that potentially inhibit the arachidonic acid cascade, only triamcinolone produced moderate effects on rat external genitalia that were largely reversible.(ABSTRACT TRUNCATED AT 400 WORDS)

Selection of food allotment for New Zealand white rabbits in developmental toxicity studies.

In three initial studies, female rabbits were fed 125, 150, or 230 g of Purina Certified Rabbit Chow No. 5322 ("regular" chow) per day or 150 g/day of Purina Certified High Fiber Rabbit Chow ("high fiber" chow) for at least 5 weeks prior to artificial insemination and until Day 28 of gestation when fetuses were removed and examined. Animals allotted 230 g/day of regular chow ate approximately 180 g/day and gained more weight than the 150 g/day group until Day 14 of gestation after which food consumption declined and body weight decreased. Animals fed 150 g/day regular chow ate all food provided until after Day 22 of gestation when food consumption decreased dramatically in some animals. Animals in the 125 g/day regular chow and 150 g/day high fiber chow groups ate essentially all food provided throughout gestation. Ad lib feeding in the 230 g/day groups was associated with adverse reproductive consequences consisting of decreased numbers of implants and live fetuses and decreased fetal weight. In one study involving 3 groups fed 125 and 150 g/day regular chow and 150 g/day high fiber chow, reproductive parameters were similar in all 3 groups. However, fetal weight in the 150 g/day regular chow group was 50% more variable than the other groups in association with more variable maternal body weight change late in gestation in that group. In subsequent studies using 125 g/day, there has consistently been fewer animals going off feed late in gestation and a decrease in fetal weight variance of approximately 60% compared to previously when the standard daily allotment was 150 g/day.(ABSTRACT TRUNCATED AT 250 WORDS)

Reversible decreases of fertility in male Sprague-Dawley rats treated orally with finasteride, a 5 alpha-reductase inhibitor.

Finasteride, a 5 alpha-reductase inhibitor, was investigated for its effects on fertility in male rats as part of its preclinical safety assessment. Studies were initiated when the male Sprague-Dawley rats were either young (4 to 6 weeks old) or mature (15 weeks old). Treatment duration ranged from 6 to 32 weeks. Each male was cohabited with two untreated females at various periods during and after treatment. Litter parameters were evaluated on either day 14 or 20 of gestation. Males were necropsied at the end of treatment or 7 to 11 weeks following the end of treatment. The major findings of these studies were that 1) young rats given 20 to 80 mg/kg/day of finasteride first showed mild to moderate decreases in fertility after 12 weeks of treatment, whereas mature males (given only 80 mg/kg/day) did not show a similar decrease until 24 weeks of treatment, 2) fewer copulatory plugs and atrophy of prostates and seminal vesicles were associated with finasteride treatment, 3) the decreased fertility was only partial (ie, fertility index did not decrease below 48% of control in any study) and was not due to decreases in mating, 4) formation of copulatory plugs, organ weights, and fertility returned to normal levels after at least 6 weeks of drug withdrawal, and 5) the testes showed no histologic or weight changes that would explain the effect on fertility. These results show that the decreased fertility in male rats was associated with finasteride-induced inhibition of accessory gland secretions, an expected pharmacologic effect.

Decreased fertility in male rats administered the 5 alpha-reductase inhibitor, finasteride, is due to deficits in copulatory plug formation.

Oral administration of 80 mg/kg/day of finasteride, a potent specific inhibitor of 5 alpha-reductase, to sexually mature male Sprague-Dawley rats for 24 to 38 weeks caused an approximate 30% to 40% decrease in fertility. There were no effects on mating indices or implants per pregnant female. From the mating trials, a selected group of treated males with poor reproductive performance was compared to a selected group of control males with good reproductive performance. Observed matings showed no qualitative effects on mating behavior or ejaculation. However, finasteride-treated males did not form or formed small and improperly positioned copulatory plugs, which are required in rats to transport sperm into the uterus. Intrauterine insemination of epididymal sperm from males that were nonfertile by natural mating resulted in similar numbers of embryos and unfertilized oocytes recovered from controls and finasteride-treated males, confirming that there was no effect of finasteride on the ability of sperm to fertilize. Decreased fertility of finasteride-treated males was due to failure to form copulatory plugs and is related to decreased weight of seminal vesicles and prostate, an expected pharmacologic effect. Testes weight was unaffected. Decreased fertility in male rats after finasteride administration is considered a species specific effect. The mechanism of the decrease in rats is not likely to be relevant to species that do not form copulatory plugs.

External genitalia abnormalities in male rats exposed in utero to finasteride, a 5 alpha-reductase inhibitor.

A series of studies was conducted to determine the developmental toxicity of the 5 alpha-reductase inhibitor finasteride (MK-0906) in rats. This compound was administered orally once daily to pregnant rats during various extended treatment periods during gestation. F1 offspring were evaluated on Day 20 of gestation as well as postnatally through mating to produce an F2 generation. MK-0906 treatment induced dosage-related incidences of hypospadias (penischisis) in male offspring with a threshold dosage level near 0.1 mg/kg/day and a 100% effect level of 100 mg/kg/day (with dosing through Day 20 of gestation). MK-0906 also caused decreased anogenital distance in male offspring. The dosage response for this effect (ranging from a 4.2% decrease at 0.003 mg/kg/day to a 38% decrease at 100 mg/kg/day) was more shallow than that for hypospadias. The decreases in anogenital distance were at least partially reversible postnatally with essentially complete recovery at dosages up to 0.1 mg/kg/day. There was also a dosage-related, temporary induction of nipples in F1 males. All of these effects were apparent following treatment on Days 6 through 17 of gestation but were more pronounced when dosing extended to Day 20 of gestation. Slight maternal toxicity consisting of minor decreases in body weight gain occurred only at dosages of 3 mg/kg/day and higher, indicating the selective nature of the developmental toxicity. The 5 alpha-reductase enzyme located in the rat fetal genital tubercle was studied in vitro and compared to that in the adult ventral prostate. The values for Km, Vmax, and IC50 for inhibition by MK-0906 were similar in the two tissues, suggesting that the enzymatic proteins in the genital tubercle and ventral prostate may be similar.

Dose--response effect of famotidine on patterns of gastro-oesophageal reflux.

The present study attempts to assess the alteration in patterns of gastro-oesophageal reflux as assessed by 24-h oesophageal pH monitoring by varying degrees of H2-receptor blockade with famotidine. Subjects were 12 patients with complaints of daily heartburn who demonstrated at least 6% of acid mucosal contact time by 24-h ambulatory oesophageal pH monitoring. All subjects had a positive Bernstein test, and nine of the 12 subjects had erosive oesophagitis. The study was conducted as a double-blind crossover design utilizing 40 mg nocte, 20 mg b.d., and 40 mg b.d. and placebo treatments. Results indicated that all treatments significantly reduced the 24-h percentage acid contact time (P less than 0.05) compared to placebo. The two b.d. treatment regimens also significantly (P less than 0.05) reduced the number of episodes lasting longer than 5 min. Only the b.d. regimens successfully lowered the percentage of upright acid exposure. All treatments significantly (P less than 0.01) reduced the percentage of supine acid contact time, as well as the number of episodes lasting more than 5 min. It is concluded that gastro-oesophageal reflux disease may well require a b.d. dosing regimen with famotidine in order to achieve optimal mucosal healing and day time symptom control.

Effects of age and chronic renal failure on the urinary excretion kinetics of famotidine in man.

The plasma and urine concentrations of famotidine, a new, potent H2-receptor antagonist, have been measured in 16 healthy young adults, 8 healthy elderly people and 18 patients with varying degrees of renal dysfunction after intravenous administration. Both the plasma elimination and renal excretion of famotidine were decreased in the elderly volunteers and renal patients. The renal clearance of famotidine averaged 4.43 ml/min/kg (310 ml/min) in normal young volunteers, which exceeded the mean creatinine clearance 1.55 ml/min/kg (109 ml/min), suggesting net secretion is a significant mechanism for elimination of famotidine. The ratio of famotidine renal clearance to creatinine clearance decreased as creatinine clearance decreased; these results suggest that the deterioration in the secretion process was much faster than that in glomerular filtration and are incompatible with the "intact nephron hypothesis". Nevertheless, both total body clearance and renal clearance were significantly correlated with creatinine clearance. The apparent half-life was also significantly correlated with creatinine clearance. Since famotidine is essentially free of dose-related adverse effects, dose adjustment in patients with mild renal insufficiency and in elderly people is not required; however, either a prolonged dosing interval or a decrease in daily dose during long-term therapy may be adapted for the patients with severe renal insufficiency to avoid accumulation and the potential undesirable effects.

Single-dose pharmacokinetics and bioavailability of famotidine in man. Results of multicenter collaborative studies.

Pharmacokinetics and bioavailability of famotidine, a new H2-receptor antagonist, were investigated in healthy subjects in five clinical studies. Linear pharmacokinetics were observed following either intravenous or oral administration. Plasma clearance averaged 463 ml min-1. Renal clearance averaged 310 ml min-1, which exceeded the glomerular filtration rate. Renal excretion was the major route of elimination. Urinary recovery of unchanged drug following intravenous administration was about 67 per cent. Famotidine plasma half-life was approximately 2.6 h. Oral absorption was incomplete. The bioavailability averaged 43 per cent of the dose.