Inflammation-mediated retinal edema in the rabbit is inhibited by topical nepafenac.

The purpose of this study was to evaluate the ability of the nonsteroidal anti-inflammatory drug nepafenac to prevent development of mitogen-induced pan-retinal edema following topical ocular application in the rabbit. Anesthetized Dutch Belted rabbits were injected intravitreally (30 microg/20 microL) with the mitogen concanavalin A to induce posterior segment inflammation and thickening (edema) of the retina. The Heidelberg Retina Tomograph was used to generate edema maps using custom software. Blood-retinal barrier breakdown was assessed by determining the protein concentration in vitreous humor, whereas analysis of PGE2 in vitreous humor was performed by radioimmunoassay. Inhibition of concanavalin A-induced retinal edema was assessed 72 h after initiation of topical treatment with nepafenac (0.1-1.0%, w/v), dexamethasone (0.1%), VOLTAREN (0.1%), or ACULAR (0.5%). Concanavalin A elicited marked increases in vitreal protein and PGE2 synthesis at 72 h postinjection. Retinal thickness was also increased by 32%, concomitant with the inflammatory response. Topical application of 0.5% nepafenac produced 65% reduction in retinal edema which was correlated with 62% inhibition of blood-retinal barrier breakdown. In a subsequent study, 0.5% nepafenac significantly inhibited (46%) blood-retinal barrier breakdown concomitant with near total suppression of PGE2 synthesis (96%). Neither Voltaren nor Acular inhibited accumulation of these markers of inflammation in the vitreous when tested in parallel. This study demonstrates that nepafenac exhibits superior pharmacodynamic properties in the posterior segment following topical ocular dosing, suggesting a unique therapeutic potential for a variety of conditions associated with retinal edema.

Nosocomial infection rates in US children's hospitals' neonatal and pediatric intensive care units.

BACKGROUND: Few data are available on nosocomial infections (NIs) in US children's hospitals' neonatal or pediatric intensive care units. The Pediatric Prevention Network (PPN) was established to improve characterization of NIs in pediatric patients and to develop and test interventions to decrease NI. METHODS: Fifty participating children's hospitals were surveyed in 1998 to determine NI surveillance methods used and neonatal intensive care unit (NICU) and pediatric intensive care unit (PICU) 1997 NI rates. Data were collected on standardized forms and entered and analyzed by using SPSS for Windows. RESULTS: Forty-three (86%) children's hospitals returned a completed questionnaire. All reported conducting NICU and PICU NI surveillance (range, 2-12; median, 12 months). Nineteen children's hospitals provided NICU NI rate data in one or more formats suitable for comparison. Denominators used for NICU NI rate calculations varied: 17 reported overall NI by patient-days; 19 reported bloodstream infection (BSI) by central venous catheter (CVC)-days, and 8 reported BSI by patient-days. Sixteen (16) children's hospitals reported NICU BSI data stratified by CVC-days and birth-weight cohort, and ventilator-associated pneumonia (VAP) by birth weight cohort was reported by 12. Twenty-four children's hospitals reported PICU NI rate data in one or more formats suitable for comparison. Denominators used for PICU NI rate calculations also varied: 20 reported overall NI rates by patient-days; 23 reported BSI rates by CVC-days, and 10 reported BSI rates by patient-days; 24 reported VAP by ventilator-days; and 15 reported urinary tract infections (UTIs) by urinary catheter-days. Median overall NI rates per 1000 patient days were 8.9 in NICUs and 13.9 in PICUs. Median NICU NI device-associated rates by birth weight (>2500 g, 1501-2500 g, 1001-1500 g, and

Periorbital cellulitis secondary to Conidiobolus incongruus.

A previously healthy, 18-month-old girl developed edema and erythema around her left eye 1 week after getting sand in that eye. The patient did not respond to oral or intravenous antibiotics. A mass developed around the eye, and biopsy revealed Conidiobolus incongruus. The patient failed to respond to amphotericin B 1 mg/kg, and susceptibility tests indicated multiantifungal resistance. A combination of antifungal therapy, hyperbaric oxygen, and surgery was required for successful treatment. Three months after treatment the child was disease free. There is no definitive therapy for Conidiobolus incongruus infections, although various drugs have been administered with some success. When susceptibility tests determine multidrug resistance, radical resection with antifungal chemotherapy and hyperbaric oxygen may be necessary as well as lifesaving.

Protection against Bordetella pertussis in mice in the absence of detectable circulating antibody: implications for long-term immunity in children.

Most vaccines used for humans work through humoral immunity, yet many appear to be protective even after specific circulating antibody levels have waned to undetectable levels. Furthermore, it has been difficult to define a serologic correlate of protection against a number of infectious diseases, including those caused by Bordetella pertussis. B. pertussis clearance in immunized mice has been shown to correlate with pertussis vaccine efficacy in children. This murine respiratory challenge model was used to demonstrate persistent vaccine-induced protection against B. pertussis in the absence of circulating antibody at the time of challenge. Whole-cell and acellular pertussis vaccines induced persistent memory T and B cells and anamnestic antibody responses after challenge. The findings suggest that immunologic memory is more significant in protection than is the induction of immediate antibody responses and imply that vaccinated children still may be protected against disease following the disappearance of specific serum IgG.

Nepafenac, a unique nonsteroidal prodrug with potential utility in the treatment of trauma-induced ocular inflammation: I. Assessment of anti-inflammatory efficacy.

Nepafenac, the amide analog of 2-amino-3-benzoylbenzeneacetic acid (amfenac), was examined in preclinical models for its potential utility as a topical ocular anti-inflammatory agent. Diclofenac was selected as the reference compound. In contrast to diclofenac (IC50 = 0.12 microM), nepafenac exhibited only weak COX-1 inhibitory activity (IC50 = 64.3 microM). However, amfenac was a potent inhibitor of both COX-1 (IC50 = 0.25 microM) and COX-2 activity (IC50 = 0.15 microM). Ex vivo, a single topical ocular dose of nepafenac (0.1%) inhibited prostaglandin synthesis in the iris/ciliary body (85-95%) and the retina/choroid (55%). These levels of inhibition were sustained for 6 h in the iris/ciliary body and 4 h in the retina/choroid. Diclofenac (0.1%) suppressed iris/ciliary body prostaglandin synthesis (100%) for only 20 min, with 75% recovery observed within 6 h following topical dosing. Diclofenac's inhibition of prostaglandin synthesis in the retina/choroid was minimal. Nepafenac's inhibitory efficacy and longer duration of action was confirmed in a trauma-induced rabbit model of acute ocular inflammation monitoring protein or PGE2 accumulation in aqueous humor. Results warrant further assessment of nepafenac's topical ocular efficacy in the treatment of postoperative ocular pain, inflammation, and posterior segment edema.

Fasciola hepatica infection downregulates Th1 responses in mice.

Immune responses induced with helminth parasites have been extensively studied, but there is limited information on those to Fasciola hepatica, especially on the subtype of T cell induced with this parasite. We investigated the local and systemic T cell responses of different strains of mice following oral infection with doses of metacercariae from F. hepatica. Spleen cells from BALB/c and 129Sv/Ev mice given a low-dose (5 metacercariae) infection exhibited a Th2 response, producing high levels of the cytokines IL-4 and IL-5, and low levels of IFN-gamma and IL-2. In contrast, C57BL/6 mice showed a mixed Th1/Th2 response. A more marked polarization to a Th2 response was observed in BALB/c, 129Sv/Ev exposed to a high-dose (15 metacercariae) infection and the C57BL/6 mice also exhibited a clear Th2 response. IL-4 defective (IL-4-/-) C57BL/6 mice infected with 5 metacercariae produced less IFN-gamma and more IL-5 compared to their wild-type C57BL/6 counterparts, suggesting that IL-4 is important in establishing the Th2 type response in murine fasciolosis. However, the secretion of IFN-gamma and IL-2 was completely suppressed in the high-dose infection and this was also observed in IL-4-/- mice. Thus, liver flukes may secrete molecules that downregulate Th1 responses. T cell responses in the mesenteric (MLN) and hepatic lymph nodes (HLN) were also examined since newly excysted juveniles infect through the intestinal wall of their host before migrating to the hepatic tissue. Cells from both MLN and HLN secreted higher levels of IL-4 and IL-5 compared to spleen cells. We also observed a difference in cytokine profiles secreted by the MLN and HLN, which may reflect responses to antigens liberated by newly excysted juveniles and hepatic stage parasites, respectively.

Fasciola hepatica suppresses a protective Th1 response against Bordetella pertussis.

Fasciolosis, like other helminth infections, is associated with the induction of T-cell responses polarized to the Th2 subtype. Respiratory infection with Bordetella pertussis or immunization with a pertussis whole-cell vaccine (Pw) induces a potent Th1 response, which confers a high level of protection against bacterial challenge. We have used these two pathogens to examine bystander cross-regulation of Th1 and Th2 cells in vivo and provide evidence of immunomodulation of host T-cell responses to B. pertussis by a concomitant infection with Fasciola hepatica. Mice with a coinfection of F. hepatica and B. pertussis exhibited a Th2 cytokine profile in response to F. hepatica antigens, similar to those infected with F. hepatica alone. By contrast, the Th1 response to B. pertussis antigens was markedly suppressed and the bacterial infection was exacerbated following infection with F. hepatica. Furthermore, an established Th1 response induced in mice by infection with B. pertussis or by parenteral immunization with Pw was also suppressed following infection with F. hepatica. This immunomodulatory effect of B. pertussis-induced responses by F. hepatica infection is significantly reduced, but not completely abrogated, in IL-4 knockout mice. Our findings demonstrate that Th2-inducing parasites can exert bystander suppression of protective Th1 responses to infection or vaccination with a bacterial pathogen and that the modulation is mediated in part by IL-4 and, significantly, is effective at both the induction and effector stages of the Th1 response.

Improved myeloperoxidase assay for quantitation of neutrophil influx in a rat model of endotoxin-induced uveitis.

Previously described models of endotoxin-induced uveitis quantify neutrophil influx into the eye using biochemical or direct cell count methods that result in an underestimation of ocular leukocyte accumulation following the inflammatory stimulus. We have optimized the rat model of endotoxin-induced uveitis by first overcoming interference in the biochemical assay of myeloperoxidase due to endogenous ocular reductants and cellular constituents containing free thiol functional groups. This was accomplished by simultaneously 1) extensively diluting soluble, interfering substances and 2) blocking tissue sulfhydril functional groups during tissue homogenization. Uveitis was induced in rats by subplantar injection of endotoxin. Twenty-four hours later, eyes were enucleated, homogenized, fractionated, and myeloperoxidase activity of neutrophils sedimenting with the membranous pellet was extracted. Previously published extraction procedures yielded only 40% of total assayable myeloperoxidase activity. Optimal recovery of myeloperoxidase activity (>twofold increase) was achieved only with two sequential extractions using 50 mM phosphate buffer (pH 7.4) containing 10 mM N-ethylmaleimide, and subsequent solubilization of myeloperoxidase activity by extraction with 0.5% hexadecyltrimethylammonium bromide in 50 mM phosphate buffer (pH 6.0). This modified extraction procedure and optimized myeloperoxidase assay conditions (300 microM hydrogen peroxide and 1.5 mM o-dianisidine) were then used to enhance the uveitis model. Maximum ocular neutrophil accumulation was observed at endotoxin doses of 100-200 microg. Total ocular neutrophil infiltrations ranged from 250,000 to 800,000 cells/globe. This leukocyte influx was inhibited dose-dependently by topical ocular administration of dexamethasone, with half-maximal inhibition observed at a concentration of 0.01%, w/v. Further validated by the correlation of biochemical results with histological evaluation, the refined methodology described in this report has application in assessing the ophthalmic therapeutic potential of antiinflammatory agents.

Genetic diversity among Mycobacterium avium complex strains recovered from children with and without human immunodeficiency virus infection.

The genetic diversity and molecular epidemiology of Mycobacterium avium complex (MAC) infections in children with and without human immunodeficiency virus (HIV) infection were evaluated. Isolates recovered from 136 children were subtyped by sequence analysis of a 360-bp region of the gene (hsp65) encoding a 65-kDa heat-shock protein. Twenty-one distinct hsp65 alleles were identified. On the basis of hsp65 genotype, 6 isolates were not MAC organisms. Of the remaining 130 samples, 61% were M. avium, 37% were Mycobacterium intracellulare, and 2% were species nonspecific MAC. Eighty-eight percent of the isolates obtained from HIV-infected children were M. avium. In contrast, only 38% of the isolates obtained from children without HIV infection were M. avium (chi2 test, P < .001). M. avium isolates were further subtyped by Southern blot analysis with insertion element IS1245. Taken together, no evidence for a single clonal M. avium strain causing infection was detected.

Transient loss of prostaglandin synthetic capacity in rabbit iris-ciliary body following anterior chamber paracentesis.

The trauma-induced acute ocular inflammatory response has been characterized by investigating the kinetics of blood-aqueous barrier (BAB) breakdown, prostaglandin (PG) accumulation in the aqueous humor, and cyclooxygenase (PGH synthase) activity of the iris-ciliary body (ICB) following paracentesis in the NZA rabbit. BAB breakdown was assessed by quantifying plasma protein extravasation into the anterior chamber. PGE2 and 6-keto-PGF(1alpha) concentrations in the aqueous humor were quantified by radioimmunoassay. The capacity of ICB tissue homogenates to generate eicosanoids from exogenously supplied [I-14C]-arachidonic acid was assessed radiometrically by HPLC. Paracentesis resulted in a rapid and dramatic increase in aqueous humor PGE2 concentrations. Within 10 minutes, PGE2 concentrations increased 937-fold, from 6.2+/-4.9 pg/ml to maximal concentrations of 5810+/-3829 pg/ml. PG synthesis was followed temporally by an increase in aqueous humor protein, with peak levels (53.1 mg/ml) achieved within 30 minutes post paracentesis. Both PGE2 and protein levels gradually declined to near baseline levels 48 hours after trauma. ICB homogenates from naive animals produced significant amounts of eicosanoids (total PG=2.95 nmol/ 10 min/100 mg tissue). HHT (12 hydroxy-heptadecatrienoic acid) was produced in the greatest quantity, followed by PGE2alpha, PGI2, and TXB2/ PGF2 . Notably, following paracentesis, eicosanoid synthesis by the isolated ICB was observed to diminish abruptly. Formation of all eicosanoids was uniformly reduced by approximately 40% five minutes following paracentesis, with an 81% decrease in synthetic activity at 15 minutes. Eicosanoid synthetic capacity was only restored to baseline 48 hours post paracentesis. These findings suggest that, following ocular trauma, temporal changes occur in ICB PG synthetic activity that may impact on the selection of an optimal dosing paradigm for efficacy testing of topically administered NSAIDs.

Phosphorylated zidovudine concentrations in mononuclear cells in pediatric patients with human immunodeficiency virus infections.

PURPOSE: The efficacy of zidovudine (ZDV) in patients with HIV-1 infection may decrease over time due to its decreased activation. The objectives of this study were to determine ZDV concentrations in plasma, active phosphorylated zidovudine (pZDV) concentrations in mononuclear cells, and assess the markers of immune function and drug toxicity during extended therapy. METHODS: Pediatric patients (aged 3 months to 18 years) with HIV-1-infection were enrolled in the study. For each patient, one blood sample was collected at each of eight routine visits to measure plasma ZDV and ZDV concentrations by a radioimmunoassay. Data including demographic information, immunological markers (CD2+, CD3+, CD4+, CD5+/19+, CD8+, CD16+, CD19+, CD38+/8+ lymphocytes), hematologic function (absolute neutrophil count, white blood cell with differential, hemoglobin, and red blood cell count), concurrent medications, and dosage regimens were obtained. RESULTS: The data from 13 patients were as follows: age: 2-18 years; range of ZDV dose: 76-238 mg/m2, total ZDV daily dosage: 264-720 mg/m2; duration of ZDV therapy prior to study: 1 to 37 months; time in study: 180-394 days; plasma ZDV concentration range: 5-1021 ng/ml; and pZDV concentration range: 0-5.382 pmol/10(6) cells. Both plasma ZDV and intracellular pZDV concentrations had a marked inter- and intrapatient variability. The pZDV concentrations decreased significantly over time in one pediatric patient (p < 0.05), tended to decrease but not significantly in three patients, and no decrease was detected in nine patients due to high variability. In our population, neither immunological nor drug toxicity markers changed over time. CONCLUSIONS: Marked inter- and intrapatient variability in pZDV concentrations was observed. The ability to phosphorylate ZDV, however, did not appear to change significantly in 12 of 13 pediatric patients with HIV-1 infection during the study period of 6-13 months.

Pancreatitis in children infected with human immunodeficiency virus.

The incidence of pancreatitis in HIV-infected children is not well known. Medical records of 42 children with HIV infection followed at Children's Hospital during a 6-year period were reviewed. Pancreatitis (elevated serum lipase levels) developed in 10 children (23.8%). Three children acquired HIV infection from vertical transmission and seven from contaminated blood products (hemophiliacs). Nine were severely immunosuppressed (CD4+ of < 100 cells/mm3). Lipase values were more often elevated than amylase values. The clinical course was protracted and severe in two children, one had four recurrences, and seven had only a single episode of pancreatitis lasting a few weeks. Opportunistic infections were present in four children and seven were receiving medications previously implicated as cause of pancreatitis. Discontinuation of dideoxynosine (ddI) in one child led to rapid resolution of pancreatitis, but continuation of medications in the other children did not alter the course. The etiology of pancreatitis may be multifactorial. Severe and prolonged clinical course is associated with advanced HIV infection. Determination of serum lipase is more useful than serum amylase for identifying those with pancreatitis.

Randomized study of the tolerance and efficacy of high- versus low-dose zidovudine in human immunodeficiency virus-infected children with mild to moderate symptoms (AIDS Clinical Trials Group 128). Pediatric AIDS Clinical Trials Group.

The current dosage of zidovudine for children is 180 mg/m2 every 6 h. To investigate whether a lower dosage was equally effective, human immunodeficiency virus (HIV)-infected children (3 months to 12 years) with mild to moderate symptoms were randomly assigned to receive either high-dose (180 mg/m2/dose) or low-dose (90 mg/m2/dose) zidovudine (double-blind). Treatments were compared with respect to neuropsychologic function, survival, clinical and laboratory evidence of disease progression, and safety and tolerance. Four hundred twenty-six HIV-infected children were enrolled; median time for receipt of study drug was 35 months. Zidovudine in either dose was well tolerated, with no difference in efficacy or tolerance by treatment group using any clinical or laboratory parameter. In children with mild to moderate disease, a reduction of zidovudine to 90 mg/m2/dose will result in substantial cost savings and should be the recommended dose.

Success inhibitions in neurotic and character-disordered patients.

Freud's initial understanding of "wreckage by success" involved oedipal conflicts. Since that time, preoedipal issues have also been identified as underlying success inhibitions. Work inhibitions in character-disordered patients may be ignored because of our general understanding of how pathological ego and superego functioning interferes in work, as in all areas. However, after significant psychotherapeutic work has been done, the author has often found that such patients have remaining work problems that should be viewed as inhibitions rather than deficits. Separation fears often underlie success inhibitions in more disturbed patients. Careful understanding of work inhibitions in more disturbed patients is crucial, because in many such patients, more freeing of functioning is possible in the work sphere than in intimate relationships. Two cases are presented to illustrate oedipal-level and preoedipal conflicts related to success.

The in vitro and in vivo ocular pharmacology of olopatadine (AL-4943A), an effective anti-allergic/antihistaminic agent.

Olopatadine (AL-4943A; KW-4679) [(z)-11-[3-(dimethylamino)propylidene]-6, 11-dihydrodibenz[b,e]oxepine-2 acetic acid hydrochloride] is an anti-allergic agent which inhibits mast cell mediator release and possesses histamine H1 receptor antagonist activity. Studies were conducted to characterize the in vitro and in vivo pharmacological profile of this drug relevant to its topical ocular use. AL-4943A inhibits histamine release in a concentration-dependent fashion (IC50 = 559 microM) from human conjunctival mast cell preparations in vitro. Histamine release was not stimulated by AL-4943A at concentrations as high as 10 mM. In contrast, ketotifen stimulated histamine release at concentrations slightly higher than effective inhibitory concentrations. AL-4943A did not display any in vitro cyclooxygenase or 5-lipoxygenase inhibition. Topical ocular application of AL-4943A effectively inhibits antigen- and histamine-stimulated conjunctivitis in guinea pigs. Passive anaphylaxis in guinea pig conjunctiva was attenuated by AL-4943A applied 30 min prior to intravenous or topical ocular antigen challenge (ED50 values 0.0067% and 0.0170%, w/v, respectively). Antihistaminic activity in vivo was demonstrated using a model of histamine-induced vascular permeability in guinea pig conjunctiva. AL-4943A applied topically from 5 min to 24 hrs prior to histamine challenge effectively and concentration-dependently inhibited the vascular permeability response, indicating the compound has an acceptable onset and a long duration of action. Drug concentrations 5-fold greater than those effective against histamine-stimulated conjunctival responses failed to inhibit vascular permeability responses induced with either serotonin or Platelet-Activating-Factor. These data indicate that the anti-histaminic effect observed with AL-4943A is specific. These anti-allergic/antihistaminic activities of AL-4943A observed in preclinical model systems have been confirmed in clinical trials in allergic patients.

Pharmacokinetics of fluconazole after oral administration in children with human immunodeficiency virus infection.

The objective of this study was to determine the pharmacokinetics of fluconazole after oral administration in children with human immunodeficiency virus (HIV) infection. After an overnight fast, a single dose of either 2 mg.kg-1 or 8 mg.kg-1 was administered in a suspension; five children received 2 mg.kg-1 and four 8 mg.kg-1 (ages 5-13 years). Blood samples were collected at various times on day 1, and once daily on days 2-7 after the dose. Fluconazole serum concentrations were measured by gas chromatography. At the dose of 2 mg.kg-1, the Cmax, AUC (0-infinity), and t1/2 ranged from 2.3-4.4 micrograms.ml-1, 84.9-136 micrograms.h.ml-1, and 19.8-34.8 h, respectively. At the dose of 8 mg.kg-1 the Cmax, AUC (0-infinity), and t1/2 ranged from 5.4-12.1 micrograms.ml-1, 330-684 micrograms h.ml-1, and 25.6-42.3 h, respectively. When compared with published data in healthy adults, fluconazole achieved similar serum concentrations in the present group of children, indicating a nearly complete degree of absorption.