Enhancement of the primary antibody response by 2-mercaptoethanol is mediated by its action on glutathione in the serum

2-Mercaptoethanol (2-ME) is widely used in rodent lymphoid cell cultures as an enhancer of multiple cellular functions. We have confirmed that the action of 2-ME must be on a serum component(s), rather than a direct action on the cells. The serum component(s) is contained within the dialyzable fraction of fetal calf serum (FCS) since: (a) dialysis of FCS diminished the ability of FCS to support an antibody response even in the presence of 2-ME; and (b) FCS dialysate, pulsed with 2-ME, restored the ability of dialyzed FCS to support an antibody response. Diminution of the reduced glutathione content of FCS by heating reduced the capacity of FCS to support an antibody response, whereas addition of 2-ME-pulsed glutathione restored the supportive capacity of heated FCS. Conversely, oxidized glutathione inhibited the antibody response in the absence of 2-ME, but that inhibition was not seen in the presence of 2-ME. We have concluded that reduced glutathione is an essential component in FCS in order for 2-ME to produce its enhancing effect. The most plausible explanation for the enhancement of antibody responses, in vitro, by 2-ME is the concomitant reversal of the inhibitory effect of oxidized glutathione and the increased availability of reduced glutathione which can scavenge oxygen-derived radicals, thus protecting macrophages and lymphocytes from the deleterious effects of oxygen-derived free radicals.     

Working conditions and health behavior as causes of educational inequalities in self-rated health: an inverse odds weighting approach

Objective:Using a novel mediation method that presents unbiased results even in the presence of exposure–mediator interactions, this study estimated the extent to which working conditions and health behaviors contribute to educational inequalities in self-rated health in the workforce.Methods:Respondents of the longitudinal Survey of Health, Ageing, and Retirement in Europe (SHARE) in 16 countries were selected, aged 50–64 years, in paid employment at baseline and with information on education and self-rated health (N=15 028). Education, health behaviors [including body mass index (BMI)] and working conditions were measured at baseline and self-rated health at baseline and two-year follow-up. Causal mediation analysis with inverse odds weighting was used to estimate the total effect of education on self-rated health, decomposed into a natural direct effect (NDE) and natural indirect effect (NIE).Results:Lower educated workers were more likely to perceive their health as poor than higher educated workers [relative risk (RR) 1.48, 95% confidence interval (CI) 1.37–1.60]. They were also more likely to have unfavorable working conditions and unhealthy behaviors, except for alcohol consumption. When all working conditions were included, the remaining NDE was RR 1.30 (95% CI 1.15–1.44). When BMI and health behaviors were included, the remaining NDE was RR 1.40 (95% CI 1.27–1.54). Working conditions explained 38% and health behaviors and BMI explained 16% of educational inequalities in health. Including all mediators explained 64% of educational inequalities in self-rated health.Conclusions:Working conditions and health behaviors explain over half of the educational inequalities in self-rated health. To reduce health inequalities, improving working conditions seems to be more important than introducing health promotion programs in the workforce.     

Variability of the steady-state visually evoked potential: Interindividual variance and intraindividual reproducibility of spatial frequency tuning

At low contrast levels there is good agreement between the psychophysical contrast sensitivity function and the tuning curve of the visually evoked potential (i.e., VEP amplitude vs spatial frequency). At high contrast, however, some researchers have found bimodal VEP tuning curves whereas others have not. We studied the VEP in 22 subjects in a short-term cross-sectional study and in 13 subjects in a longitudinal study over 8 sessions covering 28 days. Grating stimuli with 60% contrast were square-wave modulated in time (7.8 reversals/s) and space (0.06–16 cycles/degree). We found large interindividual variance in the shape of the tuning curves; about half of the subjects showed a unimodal shape, while the other half showed a bimodal one (with a notch between 1 and 2 cycles/degree). These features turned out to be stable in the longitudinal study, where variability could mainly be ascribed to a multiplicative influence common to all spatial frequencies. The marked interindividual differences in the shape of the tuning curve, which seem to be intraindividually stable, may explain previous discrepancies. It is not yet clear why the notch exists in about half of our subjects.     

Characterization of the muscarinic receptor subtype(s) mediating contraction of the guinea-pig lung strip and inhibition of acetylcholine release in the guinea-pig trachea with the selective muscarinic receptor antagonist tripitramine

1. The muscarinic receptor subtypes mediating contraction of the guinea-pig lung strip and inhibition of the release of acetylcholine from cholinergic vagus nerve endings in the guinea-pig trachea in vitro have previously been characterized as M₂-like, i.e. having antagonist affinity profiles that are qualitatively similar but quantitatively dissimilar compared to cardiac M₂ receptors. The present study sought to establish definitely the identity of these receptor subtypes by using the selective muscarinic receptor antagonist, tripitramine. Guinea-pig atria and guinea-pig trachea (postjunctional contractile response) were included for reference.
2. It was found that tripitramine antagonized methacholine-induced contractions of the guinea-pig lung strip with a pK_B value of 8.76±0.05. Both the parallel shifts of the concentration-response curves and the slope of the Schild plot being not significantly different from unity (when antagonist preincubation was for 2 h) indicated the involvement of a single population of receptors in the contractile response. From the pK_B values obtained with tripitramine and a range of other selective muscarinic receptor antagonists (cf. Roffel et al., 1993), this single population of receptors can only be classified as M₂-like.
3. Tripitramine antagonized methacholine-induced negative chronotropic and inotropic responses in guinea-pig right and left atria with apparent pK_B values of 9.4–9.6. However, such values were only obtained when antagonist preincubation was relatively long and/or antagonist concentration relatively high (e.g. with 1 h at 100 or 300 nM but 3 h at 30 nM). It thus appears that low concentrations of tripitramine do not readily equilibrate with M₂ receptors in guinea-pig atria nor with M₂-like receptors in the guinea-pig lung strip.
4. Tripitramine increased electrical field stimulation-induced cholinergic twitch contractions in guinea-pig trachea in concentrations of 0.3–100 nM, by blocking prejunctional muscarinic inhibitory autoreceptors; with higher concentrations, twitch contractions were progressively diminished, as a result of blocking postjunctional M₃ receptors (apparent pK_B value 6.07±0.15). The pEC₂₀ value (−log concentration that increases twitch by 20% of maximum) was 8.29±0.08, which would suggest that M₄ receptors are involved in this response.
5. Oxotremorine-induced inhibition of the release of prelabelled [³H]-acetylcholine from guinea-pig trachea, under conditions where there is no auto-feedback, was blocked by tripitramine (2 h preincubation) with a pK_B value of 8.56±0.06. The slope of the corresponding Schild plot was not significantly different from unity, which together with the parallel shifts of the concentration-response curves indicated the involvement of a single muscarinic receptor subtype.
6. Since the pK_B value for tripitramine at prejunctional receptors in guinea-pig trachea is in between the affinities towards M₂ and M₄ receptors, correlation plots were constructed to compare the pK_B values obtained with tripitramine and a range of other selective muscarinic receptor antagonists (cf. Kilbinger et al., 1995) to reported affinities at M₁–M₄ receptors. This showed rather similar distribution patterns of the data points around the line of equality in the case of M₂ and M₄ receptor subtypes. However, the correlation coefficient was markedly better for M₂ (0.9667) than for M₄ (0.5976). Since recent evidence suggests that M₄ receptors are not expressed in cholinergic nerves from guinea-pig trachea, it is concluded that prejunctional muscarinic autoinhibitory receptors in this tissue exhibit an atypical M₂ type character, with a pharmacological profile distinct from cardiac M₂ receptors.

     

Intraoperative HDR brachytherapy for rectal cancer using a flexible intraoperative template: standard plans versus individual planning.

HDR intraoperative brachytherapy (IOBT) is applied to locally advanced rectal tumors using a 5 mm thick flexible intraoperative template (FIT). To reduce the procedure time, treatment planning is performed using standard plans that neglect the curvature of the FIT. We have calculated the individual treatment plan, based on the real geometry of the FIT, and the dose at clips placed during surgery. A mean treatment dose of 9.55+/-0.21 Gy was found for the individual plan, compared to the prescribed 10 Gy (P<0.0001) The mean central dose was 10.03+/-0.10 Gy in the standard plan and 9.20+/-0.32 Gy in the individual plan (P<0.0001) The mean dose at the corners of the FIT was 10.3 Gy in the standard plan and ranged between 10.3 and 10.5 Gy in the individual plan. In 63% of the clips, the dose was larger than 15.0 Gy, which is equivalent to a gap between the FIT and the target smaller than 5 mm. In 18% of the clips, the dose was smaller than 13.0 Gy indicating that locally the gap was larger than 5 mm. Clinical practice will have to prove if these small dose deviations influence the clinical outcome.     

Use of AcrySof acrylic foldable intraocular lens for secondary implantation in children.

PURPOSE: Secondary intraocular lens (IOL) implantation is an increasingly viable option in the management of pediatric aphakia. We report our experience of secondary IOL implantation in pediatric patients using the AcrySof (Alcon Surgical, Fort Worth, Texas) 3-piece foldable lenses through a small incision. METHODS: We reviewed the records of all our patients < 18 years undergoing secondary IOL implantation of the AcrySof lens from 1997 to 2001. All patients with a minimum of 6 months follow-up were included. Records were analyzed for age at surgery, postoperative acuity change, postoperative refractive error and anisometropia, surgical complications, and length of follow-up. RESULTS: Fifty-five eyes of 36 patients were included in the review. Mean age at surgery was 7.4 years (1.1 to 15.4), and mean follow-up was 28 months (6.3 months to 5 years). Vision decrease > 2 lines was noted in 3 eyes (5.8%) during the follow-up period. Complications included IOL decentration in 3 eyes (5%), wound leak in 3 eyes (5%), secondary membrane formation in 5 eyes (9%), pupillary block glaucoma in 1 eye (2%), and ptosis in 1 eye (2%). Four eyes (7%) required reoperation for complications. Mean postoperative refractive error was -0.1 +/- 3.2 diopters (D), and mean anisometropia was 2.01 +/- 1.44 D. Glaucoma subsequently developed in 6 eyes (11%), 2 of which required surgical correction. CONCLUSIONS: Secondary placement of the AcrySof IOL in the ciliary sulcus is a safe and effective method to correct aphakia in pediatric patients with adequate capsular support. The incidence of complications requiring reoperation is low.     

Gene expression changes induced by ochratoxin A in renal and hepatic tissues of male F344 rat after oral repeated administration

Ochratoxin A (OTA), a naturally occurring mycotoxin, is nephrotoxic in all animal species tested and is considered a potent renal carcinogen, particularly in male rats. Its mechanism of toxicity is still unknown, although oxidative stress appears to be a plausible mechanism. Therefore, the objective of this study was to identify the biological pathways that are modulated in vivo by OTA in male F344 rats in order to gain further insight into its mechanism of renal toxicity. Rats were gavaged daily with OTA (500 microg/kg bw) and gene expression profiles in target and non-target organs were analyzed after 7 and 21 days administration. As was expected, a time-dependent increase of OTA concentrations was found in plasma, kidney and liver, with the concentrations found in both tissues being quite similar. However, histopathological examinations only revealed changes in kidney; signs of nephrotoxicity involving single cell necrosis and karyomegalic nuclei were observed in the treated rats. The number of differentially expressed genes in kidney was much higher than in liver (541 versus 11 at both time points). Several similarities were observed with other in vivo gene expression data. However, great differences were found with previous in vitro gene expression data, with the exception of DNA damage response which was not observed at mRNA level in any of our study conditions. Down-regulation was the predominant effect. Oxidative stress response pathway and genes involved in metabolism and transport were inhibited at both time points. RGN (regucalcin) - a gene implicated in calcium homeostasis - was strongly inhibited at both time points and genes implicated in cell survival and proliferation were up-regulated at day 21. Moreover, translation factors and annexin genes were up-regulated at both time points. Apart from oxidative stress, alterations of the calcium homeostasis and cytoskeleton structure may be present at the first events of OTA toxicity.