Inhibition of pilus-mediated adhesion of Pseudomonas aeruginosa to human buccal epithelial cells by monoclonal antibodies directed against pili.

The Pseudomonas aeruginosa PAK pilus is capable of mediating the binding of this strain to human respiratory epithelial cells. We have produced monoclonal antibodies (MAbs) to the PAK pilus in order to elucidate the location of the binding domain of the pilus for human buccal epithelial cells (BECs). Four MAbs are described. MAbs PK41C and PK34C were found to react with P. aeruginosa pilins produced by a large number of strains. The epitope recognized by PK41C was determined to lie within the N-terminal region of the pilin and is likely constituted by amino acid residues 22 through 33. The epitope for PK34C was located in the C-terminal region of the pilin and was partially dependent on an intact intrachain disulfide bridge between cysteine residues 129 and 142. PK99H and PK3B were found to react specifically with PAK pilin. The epitope for PK99H was also localized in the C-terminal region of the pilin protein and appears to reside between amino acid residues 130 and 138. The epitope for PK3B was not localized by using the methods of this study, but it is likely dependent on the three-dimensional structure of the pilin. Fab fragments of PK99H inhibited adhesion of strains PAK and 492c to BECs, but the adherence of five other strains was not affected. Fab fragments of PK34C inhibited adhesion of all piliated strains examined. Fab fragments from both of these antibodies inhibited PAK pilus binding to BECs. Fab fragments of PK41C and PK3B had no effect on P. aeruginosa binding to BECs. These results confirm that the C-terminal region of the pilin has adhesin qualities and that a conserved epitope lies within this region.     

Comparison of the Staph-Ident System with a Conventional Method for Species Identification of Urine and Blood Isolates of Coagulase-Negative Staphylococci

The Staph-Ident system (Analytab Products) for species identification of coagulase-negative staphylococci was compared with the conventional method of Kloos and Schleifer (21). A total of 101 clinical isolates from urine cultures and 95 clinical isolates from blood cultures were studied: overall agreement between the two methods was 86%. We concluded that the Staph-Ident system is a practical test for most clinical microbiology laboratories and that results obtained from this rapid test are comparable to those obtained from the more cumbersome conventional method. Additional investigations are needed to determine the clinical relevance of such species identification.     

EFFECT OF NITROGLYCERNE-INDUCED HYPOTENSION ON CANINE SPINAL CORD BLOOD FLOW

Twenty-four mongrel dogs were anaesthetized with pentobarbitoneand morphine sulphate. Neuromuscular blockade was achieved usingpancuronium. Spinal cord blood flow was measured using the radionuclidemicrosphere and hydrogen washout methods before, during, andfollowing nitroglycerine-induced hypotension. Heart rate, meanarterial pressure, cardiac output, pulmonary capillary wedgepressure, and acid-base balance were determined with each measurement.Mean arterial pressure was reduced by 50%. Spinal cord bloodflow, as measured by the microsphere method, increased duringthe period of hypotension, whereas values obtained using thehydrogen washout method were not significantly different fromthose at normotension. No significant change in spinal cordblood flow was detected by either method after the applicationof spinal distraction. Nitroglycerine acts predominantly onvenous capacitance vessels and it is postulated that perfusionpressure, and therefore flow, is maintained despite a reductionin arterial pressure. Presented in part at the Anual Meeting of the American Societyof Anesthesiologists, October 1985, San Francisco, California. ^*Shackleton Department of Anaesthetics, Southampton GeneralHospital, Shirley, Southampton, Hants SO9 4XY. Section of Orthopedic Surgery, Madison, Wisconsin.     

The Effects of Magnetic Resonance Imaging on Implantable Pulse Generators

The effects of magnetic resonance imaging were assessed on four dual chamber and two single chamber pulse generators. The tests were performed with a resistive, water-cooled magnet operating at 0.15 T. The 6.4-MHz radiofrequency (RF) field was operated at a maximum power of 1,000 watts with a period adjusted from 130 to 500 ms. Reed switch closure occurred in all six pulse generators tested when placed near the entrance of the magnetic resonance imaging scanner, and the generators reverted to asynchronous operation unless programmed to the "magnet off" mode. None of the pulse generators exhibited any alterations in programmed parameters or in the ability to be reprogrammed after RF pulsing. When the RF field was turned on, there was no change in the asynchronous paced cycle length in four pulse generators; however, during RF scanning there was rapid cardiac stimulation at the RF pulse period in one single chamber and one dual chamber pulse generator.     

Neurotoxicity Evaluation of N,N-Diethyl-m-toluamide (DEET) in Rats

The neurotoxic potential of N,N-diethyl-m-toluamide (DEET) wasevaluated following acute oral administration or following multigenerationplus chronic dietary administration to the rat. For the acutestudy, rats were administered undiluted DEET at dose levelsof 50, 200, or 500 mg/kg by gavage. A dose level of 500 mg/kgwas considered to be the highest practical dose that could beevaluated in this study based upon observations of overt toxicityat 500 mg/kg and mortality at 1000 mg/ kg in a dose range-findingstudy. The two measures of neurotoxicity evaluated in the acutestudy were functional observational battery (FOB) and motoractivity measurements. An apparent treatment-related effectin thermal response time (increased) was noted for both sexes1 hr after dosing at the 500 mg/kg dose level. A questionableeffect on rearing activity (decreased) also was noted at thesame dose level. For the multigeneration plus chronic dietaryadministration study, rats were administered DEET at dietaryconcentrations of 0, 500, 2000, or 5000 ppm continuously overtwo generations and then chronically for 9 months. A dietaryconcentration of 5000 ppm meets the criteria for a maximum tolerateddose (MTD) based on traditional chronic toxicology assessments.Evaluations included FOB, motor activity, discriminative acquisitionand reversal in an Mmaze, acoustic startle habituation, passiveavoidance acquisition and retention, and microscopic examinationof central and peripheral nervous tissue. The only effect thatwas considered to be possibly treatment-related was a slightincrease in exploratory locomotor activity at the 5000 ppm doselevel. Based on the results of these studies, the nervous systemdoes not appear to be a selective target when DEET is administeredto rats either as a single oral dose at high dose levels orchronically at the MTD.     

Reproductive and Thyroid Effects of Low-Level Polychlorinated Biphenyl (Aroclor 1254) Exposure

As little information is available on the adverse effects ofpolychlorinated biphenyls (PCBs) on the reproductive systemof the male rat, the current study was conducted to evaluatethe effects of subchronic administration of the PCB mixtureAroclor 1254 on testicular gamete production and endocrine function.The thyroid hormone thyroxine (T4), which is critical for reproductionand development, was also measured because of the well-documentedeffects of PCBs on this hormone. Weanling (31-day-old) maleFischer rats were administered 0, 0.1, 1, 10, or 25 mg/kg/dayAroclor 1254 by gavage for 5, 10, or 15 weeks and necropsied.The hormones testosterone (T) and thyroxine were measured inthe serum, and body weight and weights of the liver, kidney,testes, seminal vesicle plus coagulating gland, cauda epididymides,and pituitary were taken. At 10 and 15 weeks, testicular interstitialfluid (IF) was collected and T concentration in the IF was measured.Sperm motility was measured from a caudal sperm sample and spermnumbers in the testis and cauda epididymis were determined.In addition, tissues were examined microscopically for histopathologicalalterations. In the high-dose group, body, seminal vesicle,cauda epididymal, and pituitary weights were depressed at 10and 15 weeks and cauda epididymal sperm numbers were reducedafter 15 weeks of dosing. In contrast, testes weights, testicularsperm numbers, sperm motility, and serum and testicular testosteronelevels were unaffected, even in the highest dose group (25 mg/kg/day).Aroclor 1254 administration produced histological alterationsin the liver and kidney at doses of 1.0 mg/kg/day and above.These results indicate that the testis of the rat is not a specifictarget organ for Aroclor 1254. In contrast, serum T4 levelswere reduced by Aroclor 1254 administration at a dose 250-foldbelow the dose that failed to alter testicular function. SerumT4 levels were depressed 25% in the 1 mg/kg dose group after5 weeks of exposure and 30% in the 0.1 mg/kg group following15 weeks of exposure. T4 levels were undetectable in the twohighest (10 and 25) dose groups at all intervals. The fact thatthe decreases in T4 were generally concurrent with increasesin liver weight suggested that Aroclor 1254 altered T4 levelsby increasing the turnover rate in the serum by enhancing themetabolism of T4 by the liver. The reduction in serum T4 reportedhere occurred at a dose 25-fold lower than the dose generallyrecognized as affecting thyroid hormone levels.     

Interactions between the effects of propranolol and nicotine on radial maze performance of rats with lesions of the forebrain cholinergic projection system

This experiment investigated the hypothesis that nicotine-induced regional release of noradrenaline contributes to the improvements in radial maze performance following nicotine treatment in rats with lesions to the forebrain cholinergic projection system (FCPS), by examining whether pretreatment with the noradrenergic beta-receptor antagonist propranolol abolished the facilitative effects of nicotine. After S-AMPA (8.0mM) lesions to the nuclei of origin of the FCPS in the nucleus basalis and medial septal areas, rats displayed long-lasting impairment in long-term reference and short-term working memory in both spatial (place) and associative (cue) radial maze tasks. Performance of control and lesioned rats was assessed after administration of nicotine (0.1mg/kg), propranolol (either 0.5 or 5.0mg/kg) and both treatments. Nicotine reduced working memory error rates in lesioned animals, but did not affect the performance of controls. Propranolol dose-relatedly increased error rates in both control and lesioned animals. Adverse effects were more marked in controls, all four types of error being increased under the high dose of propranolol, whereas in lesioned rats significant increases in error rates above baseline were confined to working memory. The low dose of propranolol, in conjunction with nicotine, abolished the improvement in working memory seen with nicotine alone in lesioned rats. However, under joint treatment with the high dose, the substantial increases in working memory error rates seen in lesioned rats after propranolol alone were reduced to baseline level. In controls, reduction in errors to baseline was seen only in the cue task; place task errors remained significantly elevated. These results suggest that both cholinergic depletion and noradrenergic blockade exert disruptive effects on cognition, but that these effects are largely independent, since an additive or interactive mechanism would be predicted to produce greater disruption, following noradrenergic blockade, in lesioned rather than in control animals. Although facilitative effects of nicotine were abolished with the low dose of propranolol, the results further suggest that these effects are independent of release of noradrenaline, since nicotine continued to reduce errors in control and lesioned animals following blockade of beta receptors with the high dose of propranolol.     

Immunotoxicological Profile of Morphine Sulfate in B6C3F1 Female Mice

This study was undertaken to investigate a number of immuneparameters which may be compromised with exposure to morphinesulfate. Mice were implanted subcutaneously with 8-, 25-, or75-mg morphine sulfate pellets. Placebo pellets of identicalmakeup to the 75-mg morphine pellet (without morphine of course)were used as a control. Twenty-four hours after implantationof a 75-mg morphine pellet, blood levels reached a peak of 1610ng/ml. Corticosterone increased in parallel with morphine andreached a peak level of 966 ng/ml 24 hr after implantation.The dose response of morphine to increase corticosterone, however,was fiat. The weight of the lymphoid organs, spleen and thymus,and the liver were significantly reduced in the morphine-treatedgroups. Morphine treatment was associated with an increase inserum albumin, SGPT, BUN, and alkaline phosphatase indicativeof hepatic damage. In contrast to increased serum proteins,the C3 component of complement was reduced in a dose-dependentmanner. Leukocyte number in the peripheral blood was significantlyreduced, while erythro-cyte number and hematocrit were bothincreased. The number of B cells and T cells was decreased inmorphine-treated animals. However, the percentage of T cellsrelative to B cells was increased. The primary IgM antibodyresponse to the T-depen-dent antigen, sheep red blood cells,was decreased. Natural killer cell activity was reduced in responseto morphine, as was the phagocytic capacity of Kupffer cells.Host-resistance models of Listeria monocytogenes or Streptococcuspneumoniae showed an increased resistance following administrationof morphine. This increased host resistance, however, was notdue to an increase in antimicrobial action of sera obtainedfrom mice treated with morphine. The majority of morphine'seffects on the immune system exhibited a flat dose response,suggesting that these effects may be mediated secondarily throughcorticosterone.     

Developmental Neurotoxicity: Evaluation of Testing Procedures with Methylazoxymethanol and Methylmercury

Testing procedures for identification of potential developmentalneurotoxicants were evaluated using two prototypical developmentalneurotoxicants, methylazoxymethanol (MAM) and methylmercury(MeHg). Evaluation of offspring of LongEvans rats incorporatedassessments of developmental toxicity, neurochemistry, histology,and behavior, with most testing being completed near weaning.A number of endpoints in the testing strategy were sensitiveto the effects of prenatal exposure to MAM [30 mg/kg on GestationDay (GD) 15]: (1) MAM caused reduced neonatal body weights butdid not effect viability or postnatal survivorship; (2) measurementof total and regional brain weight and histological analysisshowed that a number of regions, the cortex and hippocampusin particular, were affected by MAM exposure; (3) an assay forglial fibrillary acidic protein (GFAP) showed that the concentrationof this protein was significantly increased in the cortex andhippocampus of treated offspring; (4) a T-maze delayed-alternationprocedure indicated that MAM-treated pups were slower in theacquisition phase of the task relative to control pups; (5)motor activity testing revealed hyperactivity in treated offspringthat persisted into adulthood; and (6) acoustic startle proceduresrevealed reduced startle amplitudes in preweanlings. Few endpointswere significantly affected by prenatal MeHg exposure (1, 2,or 4 mg/kg on GD 6–15). High fetal and neonatal mortalityand lower neonatal body weights were detected at the highestdose of MeHg. Although minimal effects of MeHg may reflect arelative insensitivity of the test species and/or the test methods,the combined results from both chemicals suggest that some proceduresnot currently required in the developmental neurotoxicity guidelinemay be useful in hazard identification, and further evaluationwith other chemicals, species, strains, and/or exposure paradigmsmay be warranted.