Computer-supported analysis (MegaBlot) of allopurinol-induced changes in the autoantibody repertoires of rats suffering from experimental lens-induced uveitis

Using a new quantitative immunoblot technique (MegaBlot and artificial neural network) sera of rats with lens-induced uveitis (LIU) were tested against Western blots (WB) of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separations of protein fractions from rat lenses. The blots were scanned using digital image analysis and evaluated by multivariate analysis of discriminance and an artificial neural network. Five groups of LIU rats (male Wistar rats, n = 28) were investigated: no treatment, single doses of methylprednisolone (MPR; 7.5 mg/kg body weight, intravenous), allopurinol (AL; 50 mg/kg body weight, intravenous), a combination of both drugs (AL and MPR), and repeated application of AL (ALFR; 50 mg/kg body weight intravenous every 2 weeks during the immunization period and a daily dose of approximately 25 mg/kg body weight, orally). ALFR, MPR, and MPRAL show statistically significant immunological effects in LIU treatment. Classification of WBs using both discriminance analysis or neural network resulted in 100% correct assignment; and 82% (discriminance analysis) and 86% (neural network) of WBs were correctly assigned using only unknown blots for classification (not studied before). The MegaBlot technique is able to detect therapeutically induced changes in autoantibody repertoires allowing therapeutic control and a prognostic view of treatment.     

Circadian changes of type II adenylyl cyclase mRNA in the rat suprachiasmatic nuclei

Circadian functions of the suprachiasmatic nuclei (SCN) are influenced by cyclic AMP (cAMP). Adenylyl cyclase type II (AC-II) is a cAMP-generating enzyme which, in the context of activation by Gsalpha, is further stimulated by protein kinase C or G protein betagamma subunits. Using in situ hybridization we have found a biphasic variation in AC-II mRNA within the rat SCN during the light-dark cycle (peaks at Zeitgeber time 6 and 18) and also in constant darkness (peaks at circadian time 2 and 14). The cingulate cortex showed no such variation. These findings suggest that circadian changes in AC-II expression may be pertinent to the rhythmic functions of the SCN.     

Sexually transmitted disease prevention: adolescents' perceptions of possible side effects

Forty-eight sexually active adolescents participated in an open-ended interview about the possible secondary consequences (side effects) of implementing measures to reduce the risk of contracting a sexually transmitted disease (STD). These adolescents noted 134 different consequences, which were grouped into 15 substantive categories. When four prevention measures (using condoms, being selective about sex partner(s), being monogamous, and abstaining from sexual activity) were analyzed, different patterns of consequences that were salient to these adolescents emerged. The results are discussed in terms of their implications for understanding and improving adolescent decision making about STD prevention.     

A prospective, double-blind, randomized study of the effects of perioperative steroids on palatoplasty patients

OBJECTIVE: To determine whether perioperative steroids affect the outcome of patients who undergo palatoplasty. DESIGN: A prospective, double-blind, randomized study. SETTING: A university medical center. PATIENTS: Twenty patients undergoing primary repair of a cleft palate. INTERVENTION: A prospective double-blind technique was used to randomly assign patients to receive a placebo or dexamethasone sodium phosphate perioperatively. MAIN OUTCOME MEASURE: Patients were monitored for postoperative airway distress, fever, oral fluid intake, days of hospitalization, and wound healing. RESULTS: The use of perioperative steroids was associated with shorter hospitalizations. No adverse sequelae from the administration of steroids were identified. CONCLUSIONS: In our current managed care environment, the use of perioperative steroids may play an important role in reducing health care costs.     

End-of-life decisions in the home care setting

Recent advances in the identification of genetic abnormalities associated with certain types of cancer have stimulated the development of screening and counseling programs for hereditary and familial forms of cancer. In 1995, such a program was established in a collaboration between three familial cancer clinics in Amsterdam. Given the potential impact of genetic screening and counseling on the psychosocial health of participants, it was considered essential that the program be evaluated from its inception to determine the participants' satisfaction with the services provided. A pilot study was initiated in which individuals who received genetic counseling for cancer were asked to provide feedback on the perceived quality of the services provided, and to identify areas in which additional services may be required. Preliminary results based on 36 counseled individuals indicated generally high levels of satisfaction with the care provided by the clinical geneticist and with the procedures at the familial cancer clinics. Several areas were identified that deserve additional attention: (1) the role of the family doctor in the genetic counseling; (2) communication of information regarding the possible impact of genetic counseling and testing on daily life; (3) communication between the clinical geneticist and other health care workers, and (4) psychosocial support during and after the process of genetic counseling.     

NO3--induced pH changes in mammalian cells. Evidence for an NO3--H+ cotransporter

The effect of NO3- on intracellular pH (pHi) was assessed microfluorimetrically in mammalian cells in culture. In cells of human, hamster, and murine origin addition of extracellular NO3- induced an intracellular acidification. This acidification was eliminated when the cytosolic pH was clamped using ionophores or by perfusing the cytosol with highly buffered solutions using patch-pipettes, ruling out spectroscopic artifacts. The NO3-- induced pH change was not due to modulation of Na+/H+ exchange, since it was also observed in Na+/H+ antiport-deficient mutants. Though NO3- is known to inhibit vacuolar-type (V) H+-ATPases, this effect was not responsible for the acidification since it persisted in the presence of the potent V-ATPase inhibitor bafilomycin A1. NO3-/HCO3- exchange as the underlying mechanism was ruled out because acidification occurred despite nominal removal of HCO3-, despite inhibition of the anion exchanger with disulfonic stilbenes and in HEK 293 cells, which seemingly lack anion exchangers (Lee, B. S., R.B. Gunn, and R.R. Kopito. 1991. J. Biol. Chem. 266:11448- 11454). Accumulation of intracellular NO3-, measured by the Greiss method after reduction to NO2-, indicated that the anion is translocated into the cells along with the movement of acid equivalents. The simplest model to explain these observations is the cotransport of NO3- with H+ (or the equivalent counter-transport of NO3- for OH-). The transporter appears to be bi-directional, operating in the forward as well as reverse directions. A rough estimate of the fluxes of NO3- and acid equivalents suggests a one-to-one stoichiometry. Accordingly, the rate of transport was unaffected by sizable changes in transmembrane potential. The cytosolic acidification was a saturable function of the extracellular concentration of NO3- and was accentuated by acidification of the extracellular space. The putative NO3--H+ cotransport was inhibited markedly by ethacrynic acid and by alpha-cyano-4-hydroxycinnamate, but only marginally by 4, 4'-diisothiocyanostilbene-2,2' disulfonate or by p-chloromercuribenzene sulfonate. The transporter responsible for NO3--induced pH changes in mammalian cells may be related, though not identical, to the NO3--H+ cotransporter described in Arabidopsis and Aspergillus. The mammalian cotransporter may be important in eliminating the products of NO metabolism, particularly in cells that generate vast amounts of this messenger. By cotransporting NO3- with H+ the cells would additionally eliminate acid equivalents from activated cells that are metabolizing actively, without added energetic investment and with minimal disruption of the transmembrane potential, inasmuch as the cotransporter is likely electroneutral.     

Phosphorylation of growth factor receptor binding protein-2 by pp60c-src tyrosine kinase

A positive influence of magnesium in the prevention and treatment of hyperactivity in children is more and more frequently raised in the literature. The aim of our work was to estimate magnesium contents in children with attention deficit hyperactivity disorder, (ADHD). The investigations comprised 116 children (94 boys and 20 girls), aged 9-12 years, with recognized ADHD. In 68 out of 116 patients examined ADHD occurred with other coexisting disorders specific to the developmental age and in the remaining 48 patients it occurred together with disruptive behaviour. Magnesium levels have been determined in blood serum, red blood cells and in hair with the aid of atomic absorption spectroscopy. Magnesium deficiency was found in 95 per cent of those examined, most frequently in hair (77.6 per cent), in red blood cells (58.6 per cent) and in blood serum (33.6 per cent) of children with ADHD. The conclusion from the investigations is that magnesium deficiency in children with ADHD occurs more frequently than in healthy children. Analysis of the material indicated the correlation between levels of magnesium and the quotient of development to freedom from distractibility.     

Alterations in adhesion, transport, and membrane characteristics in an adhesion-deficient pseudomonad

A stable adhesion-deficient mutant of Burkholderia cepacia G4, a soil pseudomonad, was selected in a sand column assay. This mutant (ENV435) was compared to the wild-type strain by examining the adhesion of the organisms to silica sand and their transport through two aquifer sediments that differed in their sand, silt, and clay contents. We compared the longitudinal transport of the wild type and the adhesion mutant to the transport of a conservative chloride tracer in 25-cm-long glass columns. The transport of the wild-type strain was severely retarded compared to the transport of the conservative tracer in a variety of aquifer sediments, while the adhesion mutant and the conservative tracer traveled at similar rates. An intact sediment core study produced similar results; ENV435 was transported at a faster rate and in much greater numbers than G4. The results of hydrophobic interaction chromatography revealed that G4 was significantly more hydrophobic than ENV435, and polyacrylamide gel electrophoresis revealed significant differences in the lipopolysaccharide O-antigens of the adhesion mutant and the wild type. Differences in this cell surface polymer may explain the decreased adhesion of strain ENV435.     

Enhancement of outcome from intracytoplasmic sperm injection: does co-culture or assisted hatching improve implantation rates?

In two separate prospectively randomized trials, intracytoplasmic sperm injection (ICSI) cycles were studied in a controlled manner to monitor the effects of either bovine oviductal epithelial cell co-culture (n = 119) or assisted hatching by zona drilling (n = 100). In the first study, immediately following ICSI, all eggs were placed directly either onto partial monolayers of bovine oviductal cells or into regular culture medium. Although the embryo developmental rate was apparently compromised in part by the presence of the co-culture cells, ultimately there were no significant differences in either the viable pregnancy rate (31.6% co-culture versus 29.0% control) or the embryonic implantation rate (11.4% co-culture versus 13.6% control). Assisted hatching also had no significant impact on ICSI cycle outcome in terms of either the viable pregnancy rate (30.0% assisted hatching versus 32.0% control) or the embryonic implantation rate (8.5% assisted hatching versus 13.5% control). However, in female patients aged > or = 35 years, assisted hatching appeared to convey a marginally significant benefit in terms of both the viable pregnancy rate (35.5% assisted hatching versus 11.1% control) and the embryonic implantation rate (10.3% assisted hatching versus 3.1% control). It seems that the overall improvement of ICSI cycle outcome cannot be achieved by the general application of either co-culture or assisted hatching. Nevertheless, it is possible that there remain specific patient groups that might benefit from selected use of either of these modalities.     

Decreased sodium ion absorption across nasal epithelium of very premature infants with respiratory distress syndrome

OBJECTIVE AND STUDY DESIGN: Successful adaptation to air breathing at birth depends on rapid absorption of fetal lung liquid that is mediated by activation of amiloride-sensitive sodium ion channels. To test the relationship between respiratory epithelial Na+ transport and development of respiratory distress syndrome (RDS), we measured nasal transepithelial potential difference (PD) in 31 very premature (< or = 30 weeks of gestation) newborn infants. Infants were retrospectively assigned to RDS (22 infants) and non-RDS (9 infants) groups on the basis of clinical and chest x-ray criteria. RESULTS: Maximal nasal epithelial PD increased with birth weight (-1.2 mV/100 gm) and was lower in infants with RDS (-16.5 +/- 0.6 mV) than in those without RDS (-22.0 +/- 1.3 mV). Infants without RDS had PD values similar to normal fullterm infants. Amiloride inhibition of PD, an index of Na+ absorption, was significantly lower, within the first 24 hours of life, in infants in whom RDS developed (3.8 +/- 0.2 mV; 29.5% +/- 0.8% inhibition) than in those without RDS (6.1 +/- 0.6 mV; 38.6% +/- 0.5% inhibition). Maximal and amiloride-sensitive PD returned to normal during the recovery phase of RDS. CONCLUSIONS: We conclude that Na+ absorption across nasal epithelium increases with increasing birth weight and that impairment of Na+ absorption across the respiratory epithelia of very premature infants may contribute to the pathogenesis of RDS.