Correction: a method to estimate cell cycle time and growth fraction using bromodeoxyuridine-flow cytometry data from a single sample

BACKGROUND: Presently available flow cytometric methods of bromodeoxyuridine (BrdUrd) labelling do not provide information on the cell cycle time (T(C)) and the growth fraction (GF). In this paper, we describe a novel and simple method to estimate T(C )and GF from flow cytometric analysis of a single tumour sample after BrdUrd labelling. METHODS: The proposed method is based on two assumptions: (1) the number of labelled cells traversing the cell cycle per unit time is constant and (2) the total number of labelled cells is constant throughout the cycle, provided that cells produced after division are excluded. The total numbers of labelled divided G(1 )cells, labelled divided S cells, labelled undivided S cells, and labelled undivided G(2 )cells were obtained for DNA histograms of BrdUrd-positive cells in a collected sample. These cell numbers were used to write equations to determine the durations of cell cycle phases, T(C )and GF. To illustrate the application of the proposed formulae, cell cycle kinetic parameters were analysed in solid SL2 tumours growing in DBA/2 mice and in human T-leukaemia Jurkat cells in culture. RESULTS: The suitability of the proposed method for estimating durations of the cell cycle phases, T(C )and GF was demonstrated. T(C )in SL2 tumours was found to be relatively constant at 4 and 10 days after tumour implantation (20.3 ± 1.1 h and 21.6 ± 0.9 h, respectively). GF in tumours at day 10 was lower than GF at day 4 (54.2 ± 7.7% vs. 79.2 ± 5.9%, p = 0.0003). Approximate values of T(C )and GF of cultured Jurkat cells were 23.9 h and 79.3%, respectively. CONCLUSION: The proposed method is relatively simple and permits estimation of the cell cycle parameters, including T(C )and GF, from a single tumour sample after labelling with BrdUrd. We have shown that this method may be useful in preclinical studies, allowing estimation of changes in GF during growth of murine tumours. Experiments with human Jurkat cells suggest that the proposed method might also prove suitable for measurement of cell kinetics in human tumours. Development of suitable software enabling more objective interpretation of the DNA profile in this method would be desirable

How the risk of liver cancer changes after alcohol cessation: A review and meta-analysis of the current literature

<p>Abstract</p> <p>Background</p> <p>It is well established that drinking alcohol raises the risk of liver cancer (hepatocellular carcinoma). However, it has not been sufficiently established as to whether or not drinking cessation subsequently reduces the risk of liver cancer and if it does reduce the risk how long it takes for this heightened risk to fall to that of never drinkers. This question is important for effective policy design and evaluation, to establish causality and for motivational treatments.</p> <p>Methods</p> <p>A systematic review and meta-analysis using the current available evidence and a specific form of Generalised Least Squares is performed to assess how the risk of liver cancer changes with time for former drinkers.</p> <p>Results</p> <p>Four studies are found to have quantified the effect of drinking cessation on the risk of liver cancer. The meta-analysis suggests that the risk of liver cancer does indeed fall after cessation by 6-7% a year, but there remains a large uncertainty around this estimate both statistically and in its interpretation. As an illustration it is estimated that a time period of 23 years is required after drinking cessation, with a correspondingly large 95% confidence interval of 14 to 70 years, for the risk of liver cancer to be equal to that of never drinkers.</p> <p>Conclusion</p> <p>This is a relatively under researched area and this is reflected in the uncertainty of the findings. It is our view that it is not possible to extrapolate the results found here to the general population. Too few studies have addressed this question and of the studies that have, all have significant limitations. The key issue amongst the relevant studies is that it appears that current drinkers, abstainers and former drinkers are not composed of, or effectively adjusted to be, similar populations making inferences about risk changes impossible. This is a very difficult area to study effectively, but it is an important topic. More work is required to reduce both statistical uncertainty and tackle the various study limitations this paper highlights and until this is done, the current result should be considered preliminary.</p

Test-retest reproducibility of a food frequency questionnaire (FFQ) and estimated effects on disease risk in the Norwegian Women and Cancer Study (NOWAC)

BACKGROUND: The Norwegian Women and Cancer Study (NOWAC) is a national population-based cohort study with 102 443 women enrolled at age 30–70 y from 1991 to 1997. The present study was a methodological sub-study to assess the test-retest reproducibility of the NOWAC food frequency questionnaire (FFQ), and to study how measurement errors in the data can affect estimates of disease risk. METHODS: A random sample of 2000 women aged 46–75 y was drawn from the cohort in 2002. A self-instructive health and lifestyle questionnaire with a FFQ section was mailed to the same subjects twice (test-retest), about three months apart, with a response rate of 75%. The FFQ was designed to assess habitual diet over the past year. We assess the reproducibility of single questions, food groups, energy, and nutrients with several statistical measures. We also demonstrate the method of regression calibration to correct disease risk estimates for measurement error. Alcohol intake (g/day) and high blood pressure (yes/no) is used in the example. RESULTS: For single foods there were some indications of seasonal reporting bias. For food groups and nutrients the reliability coefficients ranged from 0.5–0.8, and Pearson's r, Spearman's r(s), and two intraclass correlation coefficients gave similar results. Although alcohol intake had relatively high reproducibility (r = 0.72), odds ratio estimates for the association with blood pressure were attenuated towards the null value compared to estimates corrected by regression calibration. CONCLUSION: The level of reproducibility observed for the FFQ used in the NOWAC study is within the range reported for similar instruments, but may attenuate estimates of disease risk

Re-Assembly of the Genome of Francisella tularensis Subsp. holarctica OSU18

Francisella tularensis is a highly infectious human intracellular pathogen that is the causative agent of tularemia. It occurs in several major subtypes, including the live vaccine strain holarctica (type B). F. tularensis is classified as category A biodefense agent in part because a relatively small number of organisms can cause severe illness. Three complete genomes of subspecies holarctica have been sequenced and deposited in public archives, of which OSU18 was the first and the only strain for which a scientific publication has appeared [1]. We re-assembled the OSU18 strain using both de novo and comparative assembly techniques, and found that the published sequence has two large inversion mis-assemblies. We generated a corrected assembly of the entire genome along with detailed information on the placement of individual reads within the assembly. This assembly will provide a more accurate basis for future comparative studies of this pathogen

The HY5-PIF regulatory module coordinates light and temperature control of photosynthetic gene transcription

The ability to interpret daily and seasonal alterations in light and temperature signals is essential for plant survival. This is particularly important during seedling establishment when the phytochrome photoreceptors activate photosynthetic pigment production for photoautotrophic growth. Phytochromes accomplish this partly through the suppression of phytochrome interacting factors (PIFs), negative regulators of chlorophyll and carotenoid biosynthesis. While the bZIP transcription factor long hypocotyl 5 (HY5), a potent PIF antagonist, promotes photosynthetic pigment accumulation in response to light. Here we demonstrate that by directly targeting a common promoter cis-element (G-box), HY5 and PIFs form a dynamic activation-suppression transcriptional module responsive to light and temperature cues. This antagonistic regulatory module provides a simple, direct mechanism through which environmental change can redirect transcriptional control of genes required for photosynthesis and photoprotection. In the regulation of photopigment biosynthesis genes, HY5 and PIFs do not operate alone, but with the circadian clock. However, sudden changes in light or temperature conditions can trigger changes in HY5 and PIFs abundance that adjust the expression of common target genes to optimise photosynthetic performance and growth

Blood DNA methylation and breast cancer risk: a meta-analysis of four prospective cohort studies

BACKGROUND: Environmental and genetic factors play an important role in the etiology of breast cancer. Several small blood-based DNA methylation studies have reported risk associations with methylation at individual CpGs and average methylation levels; however, these findings require validation in larger prospective cohort studies. To investigate the role of blood DNA methylation on breast cancer risk, we conducted a meta-analysis of four prospective cohort studies, including a total of 1663 incident cases and 1885 controls, the largest study of blood DNA methylation and breast cancer risk to date. METHODS: We assessed associations with methylation at 365,145 CpGs present in the HumanMethylation450 (HM450K) Beadchip, after excluding CpGs that did not pass quality controls in all studies. Each of the four cohorts estimated odds ratios (ORs) and 95% confidence intervals (CI) for the association between each individual CpG and breast cancer risk. In addition, each study assessed the association between average methylation measures and breast cancer risk, adjusted and unadjusted for cell-type composition. Study-specific ORs were combined using fixed-effect meta-analysis with inverse variance weights. Stratified analyses were conducted by age at diagnosis ( 10 years). The false discovery rate (q value) was used to account for multiple testing. RESULTS: The average age at blood draw ranged from 52.2 to 62.2 years across the four cohorts. Median follow-up time ranged from 6.6 to 8.4 years. The methylation measured at individual CpGs was not associated with breast cancer risk (q value > 0.59). In addition, higher average methylation level was not associated with risk of breast cancer (OR = 0.94, 95% CI = 0.85, 1.05; P = 0.26; P for study heterogeneity = 0.86). We found no evidence of modification of this association by age at diagnosis (P = 0.17), ER status (P = 0.88), time since blood collection (P = 0.98), or CpG location (P = 0.98). CONCLUSIONS: Our data indicate that DNA methylation measured in the blood prior to breast cancer diagnosis in predominantly postmenopausal women is unlikely to be associated with substantial breast cancer risk on the HM450K array. Larger studies or with greater methylation coverage are needed to determine if associations exist between blood DNA methylation and breast cancer risk