Xenomitochondrial mice: investigation into mitochondrial compensatory mechanisms.

Xenomitochondrial mice, harboring evolutionarily divergent Mus terricolor mitochondrial DNA (mtDNA) on a Mus musculus domesticus nuclear background (B6NTac(129S6)-mt(M. terricolor)/Capt; line D7), were subjected to molecular and phenotypic analyses. No overt in vivo phenotype was identified in contrast to in vitro xenomitochondrial cybrid studies. Microarray analyses revealed differentially expressed genes in xenomitochondrial mice, though none were directly involved in mitochondrial function. qRT-PCR revealed upregulation of mt-Co2 in xenomitochondrial mice. These results illustrate that cellular compensatory mechanisms for mild mitochondrial dysfunction alter mtDNA gene expression at a proteomic and/or translational level. Understanding these mechanisms will facilitate the development of therapeutics for mitochondrial disorders.

Effects of socialization in the household on youth susceptibility to smoking: a secondary analysis of the 2004/05 Canadian Youth Smoking Survey.

OBJECTIVE: To determine associations between younger youths' susceptibility to smoking and four household variables related to tobacco socialization: parental and sibling smoking, restrictions on smoking in the home and exposure to smoking in vehicles. METHODS: A secondary analysis of the 2004/05 Canadian Youth Smoking Survey used logistic regression to investigate the relationships between youth susceptibility to smoking, gender, and four household variables related to tobacco socialization. Susceptibility to smoking was operationalized by three levels of smoking experience and intention: non-susceptible non-smoker, susceptible non-smoker and experimenter/smoker. The national survey included 29 243 grade 5 to 9 students from randomly sampled public and private schools in ten provinces. RESULTS: For non-smokers, the odds of being susceptible to smoking increased with having a sibling who smokes, a lack of a total household smoking ban and riding in a vehicle with a smoker in the previous week, when adjusting for all other variables in the model. These variables also increased the odds of being an experimenter/smoker versus a susceptible non-smoker. Parent smoking status was not significant in these models. CONCLUSION: Denormalization messages, through enforced home and vehicle smoking bans, appear to support youth in maintaining a resolve to not smoke, regardless of parental smoking status.

Development and initial characterization of xenomitochondrial mice.

Xenomitochondrial mice harboring trans-species mitochondria on a Mus musculus domesticus (MD) nuclear background were produced. We created xenomitochondrial ES cell cybrids by fusing Mus spretus (MS), Mus caroli (MC), Mus dunni (Mdu), or Mus pahari (MP) mitochondrial donor cytoplasts and rhodamine 6-G treated CC9.3.1 or PC4 ES cells. The selected donor backgrounds reflected increasing evolutionary divergence from MD mice and the resultant mitochondrial-nuclear mismatch targeted a graded respiratory chain defect. Homoplasmic (MS, MC, Mdu, and MP) and heteroplasmic (MC) cell lines were injected into MD ova, and liveborn chimeric mice were obtained (MS/MD 18 of 87, MC/MD 6 of 46, Mdu/MD 31 of 140, and MP/MD l of 9 founder chimeras, respectively). Seven MS/MD, 1 MC/MD, and 11 Mdu/MD chimeric founder females were mated with wild-type MD males, and 18 of 19 (95%) were fertile. Of fertile females, only one chimeric MS/MD (1% coat color chimerism) and four chimeric Mdu/MD females (80-90% coat color chimerism) produced homoplasmic offspring with low efficiency (7 of 135; 5%). Four male and three female offspring were homoplasmic for the introduced mitochondrial backgrounds. Three male and one female offspring proved viable. Generation of mouse lines using additional female ES cell lineages is underway. We hypothesize that these mice, when crossbred with neurodegenerative-disease mouse models, will show accelerated age-related neuronal loss, because of their suboptimal capacity for oxidative phosphorylation and putatively increased oxidative stress.

Choosing to remain smoke-free: the experiences of adolescent girls.

PURPOSE: To develop an understanding of the experiences of nonsmoking girls and determine how these girls avoid smoking. METHODS: A grounded theory approach was used to direct the discovery of process(es) that nonsmoking females undertake to remain nonsmokers. Seventeen nonsmoking females between the ages of 13 and 17 years were interviewed using open-ended questions designed by the authors. The interviews were tape-recorded and then analyzed using the techniques described by Strauss and Corbin. RESULTS: The data revealed that the process of remaining a nonsmoker is composed of three phases: making sense of smoking, rejecting smoking, and declaring oneself to be a nonsmoker. The process of remaining a nonsmoker appears to be rooted in developing self-confidence. As girls rejected smoking, self-confidence was boosted because they learned that their choice would be accepted and their strategies were effective in rejecting smoking. CONCLUSIONS: The findings of this study offer new insights into the experiences of adolescent nonsmokers. Our assumption that there is a process that underlies remaining a nonsmoker was supported. The findings suggest that adolescents use several strategies in this process. These strategies are noteworthy because they offer those who work in the field of health promotion new insights into ways in which they might contribute to increasing the ranks of nonsmokers.

Challenges and solutions to ultra-high-throughput screening assay miniaturization: submicroliter fluid handling.

The miniaturization of HTS assays is an important objective in the pharmaceutical industry. The ability to perform primary screening assays in high-density micro-well plates at volumes of 1-2 ml will accelerate the early stages of drug discovery and reduce costs. Ultra-HTS (uHTS) assays require an accurate and reliable means of fluid handling in the submicroliter volume range. This relates to the design of instrumentation for dispensing fluids, as well as assay plates. Fluid handling has been a major obstacle to the full implementation of miniaturized assays. This report focusses on current approaches to submicroliter fluid handling in high-density multi-well plates.

The relation of ERP components to complex memory processing.

The relation between various ERP components generated during encoding of a word and its subsequent recall were investigated using a "rote" serial-order and an "elaborative" category memory task. Words (flashed separately) were time-locked to EEG recordings from 21 cortical sites. ERP components from the five subjects having the highest recall scores were compared to the five lowest scoring subjects. Results based on the P200 peak amplitude data as well as the N400 and late positive component peak amplitude and latency data suggest that anterior and posterior distributional differences are elicited during encoding of words for rote and elaborative memory tasks. Furthermore, strong individual differences in these patterns were found as a function of task. A tentative argument was made that the obtained anterior and posterior differences may index different word feature selection and encoding processes, which are differentially utilized by high and low recallers.

T-cell receptor V beta expression in normal human skin.

The skin-associated immune system is the first line of defense against pathogenic attack from the environment and is simultaneously tolerant to localized autoantigens and to antigens of the normal microbial flora. The T-cell receptor (TCR) repertoire of skin lymphocytes may therefore be influenced by the skin microenvironment. We studied the expression of TCR beta-chain variable region (V beta) genes in normal skin by a polymerase chain reaction-based comparative method. When comparing the amplification of V beta genes in peripheral blood and normal skin, we found that TCR V beta 1, -7, -14, and -16 were often highly expressed in skin relative to peripheral blood, whereas V beta 5.1 was often highly expressed in peripheral blood mononuclear cells but not in skin. These results demonstrate that the TCR repertoire of skin lymphocytes is not determined by random sampling of peripheral blood mononuclear cells but may be molded by the interaction with self antigens and/or the normal microbial flora in the microenvironment of the skin.

Base-selective oxidation and cleavage of DNA by photochemical cosensitized electron transfer.

A photochemical mechanism for single-strand cleavage of DNA is proposed in which a photoexcited intercalator transfers an electron to an externally bound cosensitizer. Once formed, the oxidized intercalator oxidizes an adjacent base, creating a charge-separated complex from which reactions leading to cleavage of the sugar-phosphate backbone occur in competition with back electron transfer. Using ethidium bromide (EB) as the intercalator and methyl viologen (MV) as the externally bound cosensitizer, a 10-fold enhancement in the rate of single-strand break formation was found in pBR322 DNA over that for EB alone using 488-nm excitation. The rate of cleavage correlated with the amount of MV bound to DNA. In accord with the expected redox properties of the one-electron-oxidized EB and the DNA bases, cleavage occurs selectively at guanines. Although the reaction proceeds in nitrogen-purged solutions, the rate of cleavage in air-saturated solutions was enhanced 2-fold. Treatment of irradiated samples with alkali leads to a 2-fold increase in the yield of single-strand breaks. These results support a mechanism in which cleavage occurs by selective oxidation of guanines in DNA, initiated by photochemical cosensitized electron transfer from intercalated EB to externally bound MV, and may provide a basis for the development of light-activated base-selective DNA cleaving agents.

The role of ground state complexation in the electron transfer quenching of methylene blue fluorescence by purine nucleotides.

The effect of three purine nucleotides on the fluorescence of methylene blue in aqueous buffer has been investigated. Guanosine-5'-monophosphate (GMP) and xanthosine-5'-monophosphate cause fluorescence quenching while adenosine-5'-monophosphate causes a red shift in the fluorescence maximum. All three nucleotides form ground state complexes with the nucleotides as indicated by absorption spectroscopy. The fluorescence changes at nucleotide concentrations less than 30 mM are best described by a static mechanism involving the formation of non-fluorescent binary and ternary complexes in competition with dimerization of the dye. Quenching of the fluorescence decay (tau = 368 ps) at high GMP concentrations (10-100 mM) occurs at the rate of diffusion. The mechanism of fluorescence quenching may involve electron transfer within the singlet excited dye-nucleotide complex although published values of the oxidation potentials of various purine derivatives would suggest that all three nucleotides should cause quenching. Evidence for electron transfer was obtained from flash photolysis experiments in which 100 mM GMP was found to cause the appearance of a long lived transient species absorbing in the region expected for semimethylene blue.

Influence of education and advertising on the uptake of smoking by children.

How effective are peer-led programmes in preventing the uptake of smoking by children? In 1981, we conducted a randomized controlled trial of a school-based educational programme for the prevention of smoking in children who were in their seventh year at school. In this article, the reported results of two years of follow-up confirm an earlier report that both teacher-led and peer-led programmes resulted in a reduction, to about the same degree, in the uptake of smoking by girls, while only the teacher-led programme appeared to be effective in boys. In girls, both the teacher-led and peer-led programmes maintained their effects over the two years of follow-up with adjusted differences in prevalence rates of the uptake of smoking relative to the control group of -6.6% (95% CL, -17.3%, 4.0%) and -8.1% (95% CL, -18.9%, 2.7%), respectively, after two years. In boys, the effect of the teacher-led programme was reduced substantially by the second year with a difference in the prevalence rate of -2.8% (95% CL, -11.2%, 5.6%); for the peer-led programme the difference in the prevalence rate was +6.4% (95% CL, -3.6%, 16.4%). Other variables which had a significant effect on the smoking behaviour were the perceived response to cigarette advertising, parental and sibling smoking status, the perceived parental sanctions on smoking behaviour, selected peer influences and the intention to smoke. The children's perceived responses to cigarette advertising showed the strongest and most consistent evidence of an effect on the uptake of smoking by children who initially were non-smokers. After adjustment for the effects of other variables there was an excess of 15.0% (95% CL, 2.1%, 27.9%) in the prevalence rate of smoking after two years for girls who thought that they were influenced by advertising compared with those who did not. The corresponding difference for boys was 15.3% (95% CL, 4.0%, 26.6%). As smoking-prevention programmes only may delay the onset of smoking in children, it is important that legislative measures be introduced to reduce the effects of cigarette advertising.

Aortoiliac reconstruction for occlusive disease: comparison of end-to-end and end-to-side proximal anastomoses.

Between 1968 and 1979, 192 aortofemoral bifurcation grafts were placed for aortoiliac occlusive disease. The proximal anastomosis was end-to-end in 101 and end-to-side in 91 patients. Anastomoses were end-to-end in 38 (37%) of 104 patients from 1968 to 1976 and 63 (72%) of 88 patients from 1977 to 1979. Embolization occurred intraoperatively in four end-to-end and five end-to-side anastomoses. The grafts became occluded in the early postoperative period in two patients with end-to-end proximal anastomoses. There were two aortoduodenal fistulas, both associated with end-to-side proximal anastomosis. The cumulative patency rate at 5 years was 87% for end-to-end and 85% for end-to-side anastomoses. An end-to-end proximal anastomosis is indicated for associated aneurysmal disease or in the presence of aortic occlusion, while an end-to-side anastomosis is indicated when there are low-lying accessory renal arteries or in the presence of occlusive disease in the external iliac arteries. The author's experience suggests that there is no difference in the incidence of intraoperative embolization or late occlusion between end-to-side and end-to-end proximal anastomosis. The incidence of aortoduodenal fistula appears to be lower with end-to-end proximal anastomosis possibly because of better tissue apposition at the anastomosis.

Exploring risks and benefits of the birth control pill.

PIP: An activity designed to assist students in comparing the known and suspected risks and benefits involved in taking oral contraceptives (OCs) is presented. It has been used in a college-level, health issues class and probably could be used at the secondary level. The activity could function as a catalyst to a discussion of the research findings on OCs or it could follow such a discussion and be used as a type of evaluation. Each student is given a handout containing profiles of 3 women who are trying to decide whether or not to take OCs. The student must evaluate each characteristic in the profile, provided here, and indicate whether it is a risk or a benefit for the woman in question. The reason(s) for their decision must be listed. After the students have had time to list their individual responses, ideas can be shared during a class discussion. At this time, the group will most likely discover that certain factors represent both a risk and a benefit. Following the discussion of risks and benefits the class may try to reach a consensus and recommend a decision for each women. The students should be encouraged to explain why certain factors carry more weight than others when making a final decision.^ieng