Effects of glutathione transferase activity on benzo[a]pyrene 7,8-dihydrodiol metabolism and mutagenesis studied in a mammalian cell co-cultivation assay

This study deals with the role of glutathione transferase (GST)-mediatedconjugation of (+)-7ß,8-dihydroxy-9,10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene(BPDE) in two mammalian cell lines, human mammary carcinomacells (MCF-7) and rat hepatoma cells (H4IIE), in relation tothen-capacity to metabolize (–)-trans-7,8-dihydroxy-7,8-dihydro-benzo[a]pyrene[(–)-BP-7,8-diol] to products that induce mutations inco-cultivated V79 cells. Both MCF-7 and H4IIE cells metabolized(–)-BP-7,8-diol to BPDE, but mutations in co-cultivatedV79 cells were only detected with MCF-7 cells. However, depletionof glutathione (GSH) in H4HE cells increased the mutagenidtyof (– )-BP-7,8-diol to a similar level to that found withMCF-7 cells. Measurements of GST activity using GSH and post-microsomalsupernatants from H4IIE, V79 and MCF-7 cells indicated a substantialdifference in conjugation capacity. Although preparations fromall three cell-lines showed GST activity with l-chloro-2,4-dlnitro-benzeneas the substrate, GST activity towards BPDE could only be detectedin supernatants from H4HE cells. This is consistent with thepresence of GST 7-7 an isoenzyme highly efficient hi catalysingBPDE-GSH conjugation. The difference in GSH-conjugation activitytowards BPDE was confirmed using intact H4IIE and MCF-7 cellsin culture. These results indicate that GSH-conjugation playsa pivotal role in mutagenesis induced by polycyclk aromatichydrocarbons (PAH). Accordingly, a deficiency in GSH-conjugationcapacity may be regarded as one important factor in defininga target cell population with an increased risk for tumour initiationfollowing exposure to PAH.     

Sleep and its homeostatic regulation in mice lacking the adenosine A1 receptor

Sleep deprivation (SD) increases extracellular adenosine levels in the basal forebrain, and pharmacological manipulations that increase extracellular adenosine in the same area promote sleep. As pharmacological evidence indicates that the effect is mediated through adenosine A1 receptors (A1R), we expected A1R knockout (KO) mice to have reduced rebound sleep after SD. Male homozygous A1R KO mice, wild-type (WT) mice, and heterozygotes (HET) from a mixed 129/C57BL background were implanted during anesthesia with electrodes for electroencephalography (EEG) and electromyography (EMG). After 1 week of recovery, they were allowed to adapt to recording leads for 2 weeks. EEG and EMG were recorded continuously. All genotypes had a pronounced diurnal sleep/wake rhythm after 2 weeks of adaptation. We then analyzed 24 h of baseline recording, 6 h of SD starting at light onset, and 42 h of recovery recording. Neither rapid eye movement sleep (REM sleep) nor non-REM sleep (NREMS) amounts differed significantly between the groups. SD for 6 h induced a strong NREMS rebound in all three groups. NREMS time and accumulated EEG delta power were equal in WT, HET and KO. Systemic administration of the selective A1R antagonist 8-cyclopentyltheophylline (8-CPT) inhibited sleep for 30 min in WT, whereas saline and 8-CPT both inhibited sleep in KO. We conclude that constitutional lack of adenosine A1R does not prevent the homeostatic regulation of sleep.     

Effects of age on volumes of cortex, white matter and subcortical structures

The effect of age was investigated in and compared across 16 automatically segmented brain measures: cortical gray matter, cerebral white matter, hippocampus, amygdala, thalamus, the accumbens area, caudate, putamen, pallidum, brainstem, cerebellar cortex, cerebellar white matter, the lateral ventricle, the inferior lateral ventricle, and the 3rd and 4th ventricle. Significant age effects were found for all volumes except pallidum and the 4th ventricle. Heterogeneous age responses were seen in that age relationships for cortex, amygdala, thalamus, the accumbens area, and caudate were linear, while cerebral white matter, hippocampus, brainstem, cerebellar white, and gray matter, as well as volume of the lateral, inferior lateral, and 3rd ventricles showed curvilinear relationships with age. In general, the findings point to global and large effects of age across brain volumes.     

Primary antitumor immune response mediated by CD4+ T cells

Gene-targeted mice have recently revealed a role for lymphocytes and interferon-gamma (IFNgamma) in conferring protection against cancer, but the mechanisms remain unclear. Here, we have characterized a successful primary antitumor immune response initiated by naive CD4+ T cells. Major histocompatibility complex class II (MHC-II)-negative myeloma cells injected subcutaneously into syngeneic mice were surrounded within 3 days by macrophages that captured tumor antigens. Within 6 days, naive myeloma-specific CD4+ T cells became activated in draining lymph nodes and subsequently migrated to the incipient tumor site. Upon recognition of tumor-derived antigenic peptides presented on MHC-II by macrophages, the myeloma-specific CD4+ T cells were reactivated and started to secrete cytokines. T cell-derived IFNgamma activated macrophages in close proximity to the tumor cells. Tumor cell growth was completely inhibited by such locally activated macrophages. These data indicate a mechanism for immunosurveillance of MHC-II-negative cancer cells by tumor-specific CD4+ T cells through collaboration with macrophages.     

Transcellular Passage of Neisseria gonorrhoeae Involves Pilus Phase Variation

Piliated and nonpiliated Neisseria gonorrhoeae organisms were added on top of confluent layers of HEC-1-B cells, each maintained on a microporous Transwell-COL membrane. The bacteria released into the lower chamber were characterized with respect to the following virulence determinants: pili, which mediate adherence to target host cells; PilE, the major pilus subunit protein; and PilC, which is involved in pilus biogenesis and adherence. Even if >99% of the added bacteria of N. gonorrhoeae MS11 were piliated, bacteria recovered on the other side of the cell layer were predominantly nonpiliated. The recovered clones still expressed unassembled PilE protein, but 50% had lost PilC production. Nonpiliated gonococci, in which the 5′ end of pilE had been deleted, were released in reduced numbers, and piliated recA bacteria added to the cell layer were not released at all, at time points when piliated recA⁺ clones were found at high numbers in the lower chamber. Our data indicate that bacteria producing unassembled PilE protein are selected for during passage through an epithelial cell layer. The finding that the pilE gene sequence was altered in the transmigrants suggests that pilin sequence variation is involved in the transcellular passage of N. gonorrhoeae.     

Marlex Mesh Gastric Banding: A 7-12 Year Follow-Up

This paper presents a 7-12 year (mean 9.8 years) follow-up of 92 extremely obese patients treated with Marlextrade mark mesh gastric banding (GB). The follow-up rate was 92% (85 patients). Weight loss was initially good (an average reduction in BMI of 13 during the first year), but late weight gain has been a common complaint and a reason for reoperation. Other complications that necessitated reoperation were severe vomiting and esophagus not amenable to medical treatment. Four patients have developed signs of Barrett's esophagus at late follow-up. Forty-six patients (50%) were reoperated 70 times for correction of the band or conversion to vertical banded gastroplasty (VBG). The most common reoperative procedure was conversion to VBG (38 patients). Only 25 (31%) of the 80 patients with long-term follow-up have an intact band. Our results show the need of long follow-up and that this GB cannot be recommended for the treatment of morbid obesity.     

Antibiotic resistance in hospitals: a ward‐specific random effect model in a low antibiotic consumption environment

Association between previous antibiotic use and emergence of antibiotic resistance has been reported for several microorganisms. The relationship has been extensively studied, and although the causes of antibiotic resistance are multi‐factorial, clear evidence of antibiotic use as a major risk factor exists. Most studies are carried out in countries with high consumption of antibiotics and corresponding high levels of antibiotic resistance, and currently, little is known whether and at what level the associations are detectable in a low antibiotic consumption environment. We conduct an ecological, retrospective study aimed at determining the impact of antibiotic consumption on antibiotic‐resistant Pseudomonas aeruginosa in three hospitals in Norway, a country with low levels of antibiotic use. We construct a sophisticated statistical model to capture such low signals. To reduce noise, we conduct our study at hospital ward level. We propose a random effect Poisson or binomial regression model, with a reparametrisation that allows us to reduce the number of parameters. Inference is likelihood based. Through scenario simulation, we study the potential effects of reduced or increased antibiotic use. Results clearly indicate that the effects of consumption on resistance are present under conditions with relatively low use of antibiotic agents. This strengthens the recommendation on prudent use of antibiotics, even when consumption is relatively low. Copyright © 2012 John Wiley & Sons, Ltd.     

Cholinergic basal forebrain structures are involved in the mediation of the arousal effect of noradrenaline

Cholinergic basal forebrain structures are implicated in cortical arousal and regulation of the sleep–wake cycle. Cholinergic neurones are innervated by noradrenergic terminals, noradrenaline excites them via alpha‐1 receptors and microinjection of noradrenaline into the basal forebrain enhances wakefulness. However, it is not known to what extent the cholinergic versus non‐cholinergic basal forebrain projection neurones contribute to the arousing effects of noradrenaline. To elucidate the roles of cholinergic basal forebrain structures we administered methoxamine, an alpha‐1‐adrenergic agonist into the basal forebrain, in intact animals and again after selective destruction of the basal forebrain cholinergic cells by 192 IgG‐saporin. In eight male Han–Wistar rats implanted with electroencephalogram/electromyogram electrodes, a microdialysis probe targeted into the basal forebrain was perfused with artificial cerebrospinal fluid for 6 h on a baseline day, and with cerebrospinal fluid in the first and with methoxamine in the second 3‐h period of the subsequent day. The sleep–wake activity was recorded for 24 h on both days. Saporin was then injected into the basal forebrain and 2 weeks later the same experimental schedule (with cerebrospinal fluid and methoxamine) was repeated. In the intact animals, methoxamine exhibited a robust arousing effect and non‐rapid eye movement (NREM) and REM sleep was suppressed. Lesioning of the basal forebrain cholinergic neurones abolished almost completely the NREM sleep‐suppressing effect of methoxamine, whereas the REM sleep‐suppressing effect remained intact. Thus, the basal forebrain cholinergic neurones mediate, at least in part, cortical arousal and non‐REM sleep‐suppression, but they are not involved in the REM sleep‐suppressing effects of noradrenaline.