Repair of site-specific double-strand breaks in a mammalian chromosome by homologous and illegitimate recombination.

In mammalian cells, chromosomal double-strand breaks are efficiently repaired, yet little is known about the relative contributions of homologous recombination and illegitimate recombination in the repair process. In this study, we used a loss-of-function assay to assess the repair of double-strand breaks by homologous and illegitimate recombination. We have used a hamster cell line engineered by gene targeting to contain a tandem duplication of the native adenine phosphoribosyltransferase (APRT) gene with an I-SceI recognition site in the otherwise wild-type APRT+ copy of the gene. Site-specific double-strand breaks were induced by intracellular expression of I-SceI, a rare-cutting endonuclease from the yeast Saccharomyces cerevisiae. I-SceI cleavage stimulated homologous recombination about 100-fold; however, illegitimate recombination was stimulated more than 1,000-fold. These results suggest that illegitimate recombination is an important competing pathway with homologous recombination for chromosomal double-strand break repair in mammalian cells.

Insertion of a telomere repeat sequence into a mammalian gene causes chromosome instability.

Telomere repeat sequences cap the ends of eucaryotic chromosomes and help stabilize them. At interstitial sites, however, they may destabilize chromosomes, as suggested by cytogenetic studies in mammalian cells that correlate interstitial telomere sequence with sites of spontaneous and radiation-induced chromosome rearrangements. In no instance is the length, purity, or orientation of the telomere repeats at these potentially destabilizing interstitial sites known. To determine the effects of a defined interstitial telomere sequence on chromosome instability, as well as other aspects of DNA metabolism, we deposited 800 bp of the functional vertebrate telomere repeat, TTAGGG, in two orientations in the second intron of the adenosine phosphoribosyltransferase (APRT) gene in Chinese hamster ovary cells. In one orientation, the deposited telomere sequence did not interfere with expression of the APRT gene, whereas in the other it reduced mRNA levels slightly. The telomere sequence did not induce chromosome truncation and the seeding of a new telomere at a frequency above the limits of detection. Similarly, the telomere sequence did not alter the rate or distribution of homologous recombination events. The interstitial telomere repeat sequence in both orientations, however, dramatically increased gene rearrangements some 30-fold. Analysis of individual rearrangements confirmed the involvement of the telomere sequence. These studies define the telomere repeat sequence as a destabilizing element in the interior of chromosomes in mammalian cells.

Role of the nucleotide excision repair gene ERCC1 in formation of recombination-dependent rearrangements in mammalian cells.

Spontaneous recombination between direct repeats at the adenine phosphoribosyltransferase (APRT) locus in ERCC1-deficient cells generates a high frequency of rearrangements that are dependent on the process of homologous recombination, suggesting that rearrangements are formed by misprocessing of recombination intermediates. Given the specificity of the structure-specific Ercc1/Xpf endonuclease, two potential recombination intermediates are substrates for misprocessing in ERCC1(-) cells: heteroduplex loops and heteroduplex intermediates with non-homologous 3' tails. To investigate the roles of each, we constructed repeats that would yield no heteroduplex loops during spontaneous recombination or that would yield two non-homologous 3' tails after treatment with the rare-cutting endonuclease I-SCE:I. Our results indicate that misprocessing of heteroduplex loops is not the major source of recombination-dependent rearrangements in ERCC1-deficient cells. Our results also suggest that the Ercc1/Xpf endonuclease is required for efficient removal of non-homologous 3' tails, like its Rad1/Rad10 counterpart in yeast. Thus, it is likely that misprocessing of non-homologous 3' tails is the primary source of recombination-dependent rearrangements in mammalian cells. We also find an unexpected effect of ERCC1 deficiency on I-SCE:I-stimulated rearrangements, which are not dependent on homologous recombination, suggesting that the ERCC1 gene product may play a role in generating the rearrangements that arise after I-SCE:I-induced double-strand breaks.

The relationship between diet, activity, and other factors, and postpartum weight change by race.

OBJECTIVE: To identify the impact of dietary intake and activity level on postpartum weight change. METHODS: White (n = 121) and black (n = 224) women, 7-12 months postpartum, participating in the Special Supplemental Feeding Program for Women, Infants, and Children were assessed for dietary intake, activity level, body weight, and other maternal characteristics. RESULTS: For both black and white women, the most important variables in predicting postpartum weight loss were pre-pregnancy weight, gestational weight gain, parity, and prenatal exercise. After these factors were controlled, race predicted that black women retained 6.4 lb more than white women. These results may be due to the finding that black women reported significantly higher mean energy intake (2039 versus 1552 kcal, P < .001), higher percent fat in diet (41 versus 38%, P < .001), and significantly lower amounts of prenatal and postpartum activity. CONCLUSION: Higher energy intake and lower activity levels in black postpartum mothers compared with white mothers may contribute to the significantly higher rates of obesity found in black mothers. This study suggests the need for intervention strategies in the prenatal and postpartum periods to help those at risk of retaining weight gained during pregnancy.

The relationship between knowledge of cardiovascular dietary risk and food shopping behaviors.

A 44-item knowledge and attitude questionnaire was administered to 176 randomly selected shoppers to determine their cardiovascular and nutrition knowledge and attitude toward the diet-heart relationship. An analysis of descriptive sales slips issued by the supermarket determined food purchasing behavior. Food items purchased were categorized according to their polyunsaturated-saturated (P/S) ratio less than 1 (PS0), P/S ratio greater than 1 (PS1), and high cholesterol (greater than 100 mg per serving) (HCHO). We also categorized purchases by cholesterol content per serving: high, moderate, low, and very low levels. We observed a significant (r = -.25, P = .0008) negative correlation between purchasing behavior of PS0 and knowledge. A partial correlation revealed a similar association when controlling for income (r = -.26, P = .0002), age (r = -.26, P = .0005) and education (r = -.22, P = .0038). Correlation for whites was significant and negative (r = -.29, P = .0003) and for blacks nonsignificant and positive (r = .02, P = .9069). Regression analysis found a significant negative slope (b = -.57, P = .0001) of PS0 on knowledge, a marginally significant positive slope (b = -.21, P = .0527) of PS1 on knowledge, and a significant negative slope (b = -.86, P = .0171) of HCHO on education. The interaction of race and knowledge was significant for PS0 (P = .0005).

Production of chromatid breaks by single dsb: evidence supporting the signal model.

PURPOSE: The signal model proposes that all chromatid breaks arise from a single DNA double strand break (dsb) via a recombinational exchange mechanism. Here the prediction that chromatid breaks arise from a single dsb is tested. METHOD: The genetically engineered Chinese hamster cell line GS19-43 containing a unique yeast I-SceI recognition site was treated with I-SceI endonuclease (Meganuclease) in the presence of the porating agent streptolysin O. Chromatid breaks were scored at 4h, chromosome breaks at 18 and 22h following treatment (cells used for a 4h fixation were prelabelled with BrdU over two cell-cycles). Positive controls were treated with the restriction endonuclease Pst 1. RESULTS: I-SceI endonuclease produced chromatid breaks and at higher enzyme concentrations isochromatid breaks but no chromatid interchanges. About 16% of the chromatid breaks had a 'colour-switch' between the sister-chromatids at the site of breakage, as revealed by FPG staining. At the longer fixation times (18 and 22 h) chromosome breaks were observed, but again no interchanges were seen. Chromatid and chromosome breaks always appeared on the same chromosome. CONCLUSIONS: The production of chromatid breaks from a single dsb fulfils the prediction of the signal model. Moreover, the production of colour-switch breaks at a similar frequency to that for ionizing radiation indicates that chromatid breaks are produced via recombinational exchanges, a significant proportion of which occurs between sister chromatids. The majority is intrachromatid, not involving strand-switches. The absence of interchromosomal exchanges at all fixation times indicates a requirement of two dsb in two different chromosomes for their formation.

Effect of iron supplementation on endurance capacity in iron-depleted female runners.

The purpose of this investigation was to examine the effects of oral iron supplementation on endurance performance in initially iron-depleted, nonanemic female distance runners. Eighteen iron-depleted (serum ferritin less than 20 ng.ml-1, hemoglobin greater than or equal to 12 g.dl-1) women (22-39 yr) performed a VO2max test and an endurance run to exhaustion. Subjects were pair-matched on the basis of endurance time and then randomly assigned to an iron supplement or a placebo group. Following supplementation, the iron group had a significantly higher (P = 0.03) mean serum ferritin concentration (23.4 vs 15.7 ng.ml-1) and lower (P = 0.04) mean total iron-binding capacity than the placebo group. Both groups increased their time to exhaustion (25.5% and 22.2% for the iron and placebo groups, respectively) but were not significantly different (P = 0.72) from each other. There were also no differences (P greater than 0.05) between the groups with respect to lactate concentrations and physiological measures taken during the two exercise tests. The results of this study suggest that 8 wk of oral iron supplementation improves iron status in iron-depleted female distance runners, but does not enhance endurance capacity.

Black-white differences in body size perceptions and weight management practices among adolescent females.

OBJECTIVE: This study compares body size perceptions and weight management practices of black and white adolescent females. DESIGN: Subjects were selected through a statewide, three-stage sampling procedure designed to provide a sample statistically representative of high school students in South Carolina. SUBJECTS: Participants included black (n = 1824) and white (n = 2256) females, 14-18 years of age, enrolled in South Carolina public high schools. METHODS: Respondents were asked to assess their perceived body size as overweight, underweight, or about right. Self-reported weight management practices included dieting (reducing caloric intake), exercise, and other methods (including diet pills and vomiting). Chisquare analysis was used to assess the differences in body size perception and weight management behaviors. Polychotomous logistic regression was performed to examine association while controlling for socioeconomic status. RESULTS: Forty-one percent of the white adolescents and 29% of the black adolescents perceive themselves as overweight (p < 0.005). In the week prior to the survey, 28% of the white adolescents and 13% of the black adolescents reported dieting 34% of the while versus 23% of the black adolescents reported exercising to lose weight; and 45% of the white and 16% of the black students reported both dieting and exercising. Polychotomous logistic regression analysis showed that white adolescent girls were almost twice as likely to perceive themselves as overweight as black adolescent girls. The white students had 6.04 [95% confidence interval (CI), 1.77, 20.67] times the odds of using pills and vomiting and 3.76 (95% CI, 2.99, 4.72) times the odds of engaging in dieting and exercising as methods of weight management compared to the black students. CONCLUSIONS: These findings suggest that white adolescents are more likely to perceive themselves as overweight than black adolescents and are more likely to engage in unhealthy weight management practices than black adolescents.

Weight control and calorie expenditure: thermogenic effects of pre-prandial and post-prandial exercise.

The thermogenic effects of pre- and postprandial exercise was examined in seven lean active females. Energy expenditure was measured for 3 h via open circuit indirect calorimetry after four separate treatments: Exercise Only (25 min treadmill run at 60% VO2 max), Meal Only (910 kcal mixed meal), Exercise-Meal and Meal-Exercise. The thermogenic response to the Exercise-Meal treatment was similar to the Meal Only treatment. However, the Meal-Exercise treatment resulted in a greater energy expenditure than the Meal Only and Exercise-Meal treatments. The Exercise Only treatment showed the lowest thermogenic response. These data suggest that exercise following a meal would be more beneficial than exercise before a meal in increasing and maintaining an elevated energy expenditure.

A study of factors associated with weight change in women who attempt smoking cessation.

This study investigated behavioral and attitudinal relationships over a 12-week time period in 48 women enrolled in a commercial smoking cessation program who were categorized as non-recidivists (NR), early recidivists (ER) and late recidivists (LR) to cigarette smoking. NR had significantly higher weight gains (F = 6.70), significantly higher levels of physical activity (F = 6.42), and significantly less concern of postsmoking cessation weight gain (F = 5.08) than either two groups of recidivists, (p less than or equal to .05). Other findings, although not significant, were that NR exhibited lower caloric intake and more frequent snacking behaviors than either ER or LR. These results indicate an overall stronger commitment to more positive health behaviors in the NR than those who returned to cigarette smoking.

Thermogenic effects of pre-prandial and post-prandial exercise in obese females.

The thermogenic effects of pre- and postprandial exercise was examined in six obese premenopausal females. Using open circuit calorimetry, energy expenditure was measured for 3 hours following five separate treatments: Resting Control, Exercise Only (25 min cycle at 60% VO2 max), Meal Only (902 kcal mixed meal), Exercise-Meal, and Meal-Exercise. Meal Only, Exercise-Meal, and Meal-Exercise were significantly higher than Rest Only and Exercise Only treatments. However, Meal-Exercise resulted in the greatest energy expenditure. These results suggest that in obese female subjects exercise following a meal will produce the greatest thermogenic response and could possibly influence weight maintenance or weight loss if implemented in food and exercise patterns of behavior.

The impact of a required college health and physical education course on the health status of alumni.

The impact of a required college health and physical education course on selected health knowledge, attitudes, and behaviors of alumni was evaluated. The design was a cross-sectional study (mail out/mail back survey) using a stratified random sample of 2,000 college alumni. The overall response rate was 50%. Results were analyzed by college of graduation (college with a required health/physical education course versus no required course). Alumni who took the course were more likely to know their blood pressure, blood cholesterol, and recommended dietary fat intake than the comparison group were. They also reported that the course positively influenced their attitudes toward exercise, eating, and smoking They were more likely to engage in aerobic exercise and less likely to smoke; and they had lower intakes of dietary fat, cholesterol, and sodium. The results suggested that a required college course enhanced selected health-related knowledge, attitudes, and behaviors of alumni.

Body dissatisfaction among White and African American male and female college students.

Body size perception has been shown to be highly influenced by cultural factors including race. This study assessed body size perceptions of a convenience sample of college students (N=630). Included in a paper-and-pencil survey were current height and weight (body mass index (BMI)=wt [kg]/ht(2) [m]), perceived BMI, desired MI and perceptions of BMIs desired by others. Also assessed were perceptions of underweight, acceptable weight and overweight/obesity. African American females were found to have a higher current BMI than Whites (P< or =.001), higher desired BMI (P< or =.001) and higher BMI perceived to be desired by others (P< or =.001). African American and White males did not differ on any of these measures. Most (83%) underweight (BMI<19) African American females perceived themselves as underweight while only about one-half of underweight (56%) White females perceived themselves as underweight. Almost half (43%) of White females and about a fourth (27%) of African American females of acceptable BMI range (BMI=19-25) desired to be in the underweight range. Of those who were overweight (BMI=25-30), 20% of the Whites and only 3.0% of the African Americans chose an underweight silhouette as being desired. While African Americans and White males tended to have similar perceptions that there remains sociocultural influences in African American females regarding acceptance of a large body size.

Expanding the criteria for defining success when evaluating weight loss intervention programs.

In order to evaluate outcomes among former participants in a residential weight loss program, attendees were surveyed from 1 to 5 years post-intervention. A total of 187 respondents were studied for weight changes and behavioral practices. Diet practices were assessed by number of servings per day using the Food Guide Pyramid recommendations as a guideline for indicating fruit and vegetable intake. Exercise behaviors were measured in frequency, duration, and intensity, and a weighted score was computed to obtain units for describing physical activity. Maintaining a 10% weight loss from the program entry weight, consuming five or more servings per day of fruits and/or vegetables (5-a-day) and maintaining an "active" level of physical activity were included in criteria for describing intervention success. Those meeting two of the three criteria were categorized as "successful," which included 35.8% of the study population. Because each of these behaviors (5-a-day, active lifestyle, and modest weight loss maintenance) result in independent risk reduction, it is recommended that future weight loss intervention evaluations expand the criteria for describing successful impacts and outcomes to include not only weight maintenance, but also physical activity and diet compliance behaviors.

Measurements of body image in clinical weight loss participants with and without binge-eating traits.

This study measured body image disturbances of individuals in a residential weight loss program who were identified as having binge-eating disorder (BED) traits. The study population (N=97) was a convenience sample of 74 men (76%, mean age=51.0) and 23 women (24%, mean age=49.6) in the program who completed the Eating Questionnaire-Revised (EQ-R), the Attitudes About Weight and Dieting (AAWD), the Physical Appearance State and Trait Anxiety Scale (PASTAS): State Version, and the Contour Drawing Rating Scale (CDR). Fifty-five individuals reported having binge traits (57%) while 42 (43%) had no binge traits. Individuals with the binge traits had a significantly higher BMI than nonbinge trait individuals (P=.008). The binge trait group scored higher on the total AAWD (P=.004), the AAWD factor "Fear of Fat" (P=.002), the total PASTA (P=.001), and the PASTA factor "Weight" (P=.001) than the nonbinge trait group. Logistic regression analysis indicated that the odds of having a binge trait were 1.44 times more likely for a person at a given score on the PASTA subscale Weight vs. a person at a score of 5 units less. Feelings of being unable to control eating among individuals seeking weight control is associated with several characteristics related to body image. Individuals showing greater concern about weight and dieting and specifically greater fears of becoming fat were more likely to have a problem with binge eating than those without these concerns. The results of this study suggest that a negative body image is an important factor to consider when treating individuals indicating binge-eating traits.

Patterns of tobacco and alcohol use among sedentary, exercising, nonathletic, and athletic youth.

Through a statewide random design, 7,846 9th to 12th grade students in 81 South Carolina public schools were selected to complete the Youth Risk Behavior Survey. Mantel-Haenszel Chi-Square, general linear models, and logistic regression were used to examine the relation between the independent variables of race, gender, and physical activity status with the dependent variables of tobacco and alcohol use while controlling for race, gender, and participation in physical education classes. The six physical activity levels ranged from sedentary nonathletes to athletes participating on two or more sports teams. Highly active athletes drank more frequently than did low-activity nonathletes (p = .002) and sedentary nonathletes (p = .006), and they were more likely to binge drink (p = .004). Athletic groups were less likely to smoke than were the nonathletic groups (p < .0001) with "sedentary nonathletes" the heaviest smokers of all groups. Athletes were more likely to use smokeless tobacco (p < .0001) than nonathletes although this difference disappeared when controlling for gender and race. This research supports other findings about prevalence of smokeless tobacco use among athletic youth. In addition, it reveals that athletic youth are at an increased risk of alcohol use and binge drinking. Possible factors such as the risk-taking tendencies of team sport athletes should be investigated.

Racial and gender differences in weight status and dietary practices among college students.

A questionnaire was used to assess nutritional intake, weight status, and dietary practices among 630 U.S. college students. Body mass index (BMI) was calculated using self-reported heights and weights, and nutritional status was assessed via 24-hour recall. Sixty-four percent of the students had acceptable BMI levels (between 19 and 25); 16% of African-American females and 15% of African-American males had BMIs indicating obesity (30 or above). Approximately 18% of the students consumed 5 servings per day of fruits and vegetables, 7% consumed 6 or more grain products, and 53% consumed 2 or more dairy products. Twenty-seven percent reported never/rarely eating fast foods. Recommendations are presented for the development of culturally appropriate health education/promotion programs in order to help college students meet Healthy People 2010 objectives.

Black and white adolescent females' pre-pregnancy nutrition status.

The dietary intakes of energy, protein, calcium, and iron of 408 randomly selected nonpregnant, black and white female adolescents were analyzed to determine their pre-pregnancy nutritional status. Pre-pregnancy weight for height was calculated and used as an indicator of nutritional status. After controlling for race, socioeconomic status, and age, results indicated that black females had significantly higher mean intakes of energy (p = .0001), protein (p < .0001), calcium (p = .0205), and iron (p = .0001) than did white females. Distribution of white and black females in the percentages of RDA categories differed significantly for energy (p < .0001), protein (p < .0001), and iron (p < .0001). A higher percentage of white females were found in the three lower categories (< 100% of RDA) than of black females. No significant differences were found in the distribution of black and white females in the three Body Mass Index categories. A large proportion of both black and white females' intakes of energy, calcium, and iron were below the recommended allowances when categorized according to the percentages of recommended intakes. Thirty-seven percent of the black females and 42.6% of the white females were classified by their BMIs as being underweight. These findings suggest that the majority of black and white females surveyed had poor pre-pregnancy nutritional status.