Joint modeling of performance and subjective reporting to assess sensitivity to drug‐induced sleepiness

In an NASA ground study, two forms of cognitive tests were evaluated in terms of their sensitivity to sleepiness induced by the drug promethazine (PMZ). Performance for the two test modes (Y₁ and Y₂), PMZ concentration, and a self‐reported sleepiness using the Karolinska Sleepiness Scale (KSS) were monitored for 12 h post dose. A problem arises when using KSS to establish an association between true sleepiness and performance because KSS scores are discrete and also because they tend to concentrate on certain values. Therefore, we define a latent sleepiness measure X^* as an unobserved continuous random variable describing a subject's actual state of sleepiness. Under the assumption that drug concentration affects X^*, which then affects Y₁, Y₂, and KSS, we use Bayesian methods to estimate joint equations that permit unbiased comparison of the performance measures' sensitivity to X^*. The equations incorporate subject random effects and include a negativity constraint on subject‐specific slopes of performance with respect to sleepiness. Published in 2010 by John Wiley & Sons, Ltd.     

Estimation of rank correlation for clustered data

It is well known that the sample correlation coefficient (R_xy) is the maximum likelihood estimator of the Pearson correlation (ρ_xy) for independent and identically distributed (i.i.d.) bivariate normal data. However, this is not true for ophthalmologic data where X (e.g., visual acuity) and Y (e.g., visual field) are available for each eye and there is positive intraclass correlation for both X and Y in fellow eyes. In this paper, we provide a regression‐based approach for obtaining the maximum likelihood estimator of ρ_xy for clustered data, which can be implemented using standard mixed effects model software. This method is also extended to allow for estimation of partial correlation by controlling both X and Y for a vector of other covariates. In addition, these methods can be extended to allow for estimation of rank correlation for clustered data by (i) converting ranks of both X and Y to the probit scale, (ii) estimating the Pearson correlation between probit scores for X and Y, and (iii) using the relationship between Pearson and rank correlation for bivariate normally distributed data. The validity of the methods in finite‐sized samples is supported by simulation studies. Finally, two examples from ophthalmology and analgesic abuse are used to illustrate the methods. Copyright © 2017 John Wiley & Sons, Ltd.     

Model-based analyses of bioequivalence crossover trials using the stochastic approximation expectation maximisation algorithm

In this work, we develop a bioequivalence analysis using nonlinear mixed effects models (NLMEM) that mimics the standard noncompartmental analysis (NCA). We estimate NLMEM parameters, including between‐subject and within‐subject variability and treatment, period and sequence effects. We explain how to perform a Wald test on a secondary parameter, and we propose an extension of the likelihood ratio test for bioequivalence. We compare these NLMEM‐based bioequivalence tests with standard NCA‐based tests. We evaluate by simulation the NCA and NLMEM estimates and the type I error of the bioequivalence tests. For NLMEM, we use the stochastic approximation expectation maximisation (SAEM) algorithm implemented in monolix . We simulate crossover trials under H₀ using different numbers of subjects and of samples per subject. We simulate with different settings for between‐subject and within‐subject variability and for the residual error variance. The simulation study illustrates the accuracy of NLMEM‐based geometric means estimated with the SAEM algorithm, whereas the NCA estimates are biased for sparse design. NCA‐based bioequivalence tests show good type I error except for high variability. For a rich design, type I errors of NLMEM‐based bioequivalence tests (Wald test and likelihood ratio test) do not differ from the nominal level of 5%. Type I errors are inflated for sparse design. We apply the bioequivalence Wald test based on NCA and NLMEM estimates to a three‐way crossover trial, showing that Omnitrope®; (Sandoz GmbH, Kundl, Austria) powder and solution are bioequivalent to Genotropin®; (Pfizer Pharma GmbH, Karlsruhe, Germany). NLMEM‐based bioequivalence tests are an alternative to standard NCA‐based tests. However, caution is needed for small sample size and highly variable drug. Copyright © 2011 John Wiley & Sons, Ltd.     

Influence of vitamin intake and MTHFR polymorphism on the levels of DNA damage in tobacco farmers

Background: Genetic damage may occur spontaneously under normal metabolic circumstances, inadequate intake of nutrients, and excessive exposure to environmental mutagens. Objectives: To evaluate the influence of the intake of micronutrients vitamin B₁₂, vitamin B₆, and folate and of the polymorphism methylenetetrahydrofolate reductase (MTHFR) C677T on the induction of DNA damage in tobacco farmers. Methods: The study involved 66 men and 44 women engaged in tobacco cultivation in the region of Venâncio Aires (Rio Grande do Sul state, Brazil). Peripheral blood samples were collected to analyze DNA damage using the Comet assay, the micronucleus (MN) test and MTHFR C677T polymorphism. Dietary intake was evaluated based on the mean values obtained from three 24-h diet recall questionnaires, and nutrient intake data were computerized and estimated in the Food Processor SQL 10.9 program. The statistical tests used to generate the stated results were Kruskal–Wallis test, Exact Fisher’s test, and multivariate linear regression analysis. Results: DNA damage was significantly higher in individuals who had an inadequate intake of folate, vitamin B₁₂, and vitamin B₆ (P < 0.01) assessed by Comet assay. In relation to MN test results, buccal cells showed MN frequency higher in individuals with inadequate intake of vitamin B₆ (P < 0.01). No difference was observed in MN lymphocytes frequency. No significant association was detected between MTHFR C677T polymorphism and DNA damage in tobacco farmers. Conclusion: Our results suggest that folate, vitamin B₁₂, and vitamin B₆ deficiency may be associated with genotoxic effect in individuals exposed to pesticides.     

B vitamins deficiency and decreased anti-oxidative state in patients with liver cancer

Background This study examined the status of oxidative stress and B vitamins in hepatocellular carcinoma (HCC) patients in different tumor-node-metastasis stages. Patients were divided into two groups as I + II (n = 21) and III + IV (n = 19). Methods Plasma levels of lipid oxidation, α-tocopherol, β-carotene, vitamin C, glutathione and the activity of antioxidant enzymes (glutathione peroxidase, superoxide dismutase, catalase, and xanthine oxidase) were determined for evaluating oxidative status. Blood B vitamins (B₁, B₂, B₆, B₁₂, and folate) and serum ghrelin were analyzed, and the relationship between serum ghrelin and vitamins B₂ (or B₆) was evaluated. Results HCC patients at III + IV stage showed significantly lower ghrelin, higher cholesterol, triglyceride, and uric acid than patients at I + II stage and healthy subjects (P < 0.05). Plasma lipid oxidation level in HCC patients was significantly greater than healthy subjects (P < 0.05). The activity of glutathione peroxidase, superoxide dismutase or catalase was significantly decreased, but xanthine oxidase activity was significantly elevated in HCC patients (P < 0.05). Plasma level of glutathione and vitamin C, not α-tocopherol and β-carotene, in HCC patients was significantly lower (P < 0.05). Vitamins B₂ and B₆ levels in red blood cells from these HCC patients were significantly lower (P < 0.05). Conclusion This study provided novel clinical findings regarding the status of oxidative stress and B vitamins in HCC patients. Plasma glutathione level may be a proper biomarker for evaluating oxidative status for HCC patients. Our data indicate that HCC patients might need B vitamins supplementation. The increased serum level of triglyceride and cholesterol might be a consequence of an impaired hepatic fat metabolism, and might be improved by a lower fat administration to these patients.     

Likelihood ratio and score tests to test the non‐inferiority (or equivalence) of the odds ratio in a crossover study with binary outcomes

We consider the non‐inferiority (or equivalence) test of the odds ratio (OR) in a crossover study with binary outcomes to evaluate the treatment effects of two drugs. To solve this problem, Lui and Chang (2011) proposed both an asymptotic method and a conditional method based on a random effects logit model. Kenward and Jones (1987) proposed a likelihood ratio test (LRT_M) based on a log linear model. These existing methods are all subject to model misspecification. In this paper, we propose a likelihood ratio test (LRT) and a score test that are independent of model specification. Monte Carlo simulation studies show that, in scenarios considered in this paper, both the LRT and the score test have higher power than the asymptotic and conditional methods for the non‐inferiority test; the LRT, score, and asymptotic methods have similar power, and they all have higher power than the conditional method for the equivalence test. When data can be well described by a log linear model, the LRT_M has the highest power among all the five methods (LRT_M, LRT, score, asymptotic, and conditional) for both non‐inferiority and equivalence tests. However, in scenarios for which a log linear model does not describe the data well, the LRT_M has the lowest power for the non‐inferiority test and has inflated type I error rates for the equivalence test. We provide an example from a clinical trial that illustrates our methods. Copyright © 2016 John Wiley & Sons, Ltd.     

Primary hepatocellular carcinoma in Aboriginal Australians

Abstract: We identified incident cases of primary hepatocellular carcinoma (PHC) in the Northern Territory from 1980 to 1989: there were 18 Aboriginal and six non-Aboriginal cases, yielding incidence rates of 5.2 per 100 000 (Aboriginal) and 0.5 per 100 000 (non-Aboriginal) with a relative risk of 10.4 (95 per cent confidence interval (CI) 4.0 to 26.6). The carcinoma was more frequent in males (2.3 per 100 000) than in females (0.7 per 100 000), with a relative risk of 3.4 (CI 1.3 to 9.3). Incidence increased with age; the trend was statistically significant in Aborigines (X²₁, = 4.7, P < 0.05) but not in non-Aborigines (X²₁ = 3.4, P > 0.05). Hepatitis B virus (HBV) serology was available for 11 Aboriginal and four non-Aboriginal cases; seven of the Aboriginal cases and two of the non-Aboriginal cases were positive for hepatitis B surface antigen (HBsAg). The prevalence of HBsAg in Aboriginal patients with the carcinoma (63.6 per cent) was much higher than that (13.1 per cent) in Aborigines surveyed from communities in the Northern Territory (X²₁ = 21.7, P < 0.001). Our results show that the age-specific incidence of PHC in Aborigines in the Northern Territory (30.9 for ages 40 and over) is comparable to that in high-incidence countries such as China (36.9 for ages 40 and over), and that hepatitis B is of major aetiological importance in the Aboriginal population. This underlines the importance of universal immunisation for prevention of HBV infection and for long-term prevention of PHC.     

Nutritional Supplements for Older Adults: Review and Recommendations—Part II

The use of nutritional supplements (NS) with the intention of improving health and delaying age-related chronic disease is a common practice among older adults; however, randomized controlled trials have yielded mixed results regarding the likelihood that these NS provide true health benefits. We reviewed the findings of these studies regarding the effects of NS of folic acid, vitamin B₁₂, vitamin B₆, and omega-3 fatty acids on health outcomes in older adults. Our conclusions include the following: Supplements of the B vitamins folate, B₁₂ and B₆ have been studied with regards to primary and secondary prevention of a number of major age-related chronic diseases, including cardiovascular disease (CVD), stroke, cognitive decline, and cancer. While there are some encouraging findings with regards to stroke, depression, and macular degeneration (although in only one study in the latter case), there is little evidence of benefit of B vitamin NS for delaying CVD or age-related cognitive changes. In the few cancer-related studies, the evidence of benefit is coupled with concerns about enhancing the growth of existing undiagnosed cancers. In contrast, clear health benefits have been shown with modest increases in consumption of fatty fish or fish oil supplements, including a reduction in the risk of sudden cardiac death. In addition, there is evidence that high dose fish oil supplements may lower serum triglyceride levels.     

Distribution of the admixture test for the detection of linkage under heterogeneity

The admixture test for the detection of linkage under heterogeneity is considered. We show that the null distribution of this test statistic has half its weight concentrated on zero and the other half on a complicated distribution that can be approximated by max (X₁, X ₂ ) where X₁ and X₂ are independent χ variables. We also investigate the stability of the size of the test for small samples. The power of this test to detect linkage, when heterogeneity is present, can be substantially greater than the standard test that assumes homogeneity. Even when heterogeneity is not present, the test is only slightly less powerful than the homogeneous test. This would suggest the use of the admixture test in preference to the homogeneous test if the presence of heterogeneity is at all suspected. © 1993 Wiley-Liss. Inc.     

The silent point mutations at the cleavage site of 2A/2B have no effect on the self-cleavage activity of 2A of foot-and-mouth disease virus

The 2A region of the foot-and-mouth disease virus (FMDV) polyprotein is 18 amino acids in length, and 2A self-cleavage site (2A/2B) contains a conserved amino acid motif G_2A/P_2B. To investigate the synonymous codon usage for Glycine at the 2A/2B cleavage site of FMDV, 66 2A/2B₁ nucleotide sequences were aligned and found that the synonymous codon usage of G_2A is conserved and GGG was the most frequently used. To examine the role of synonymous codons for G_2A in self-cleavage efficiency of 2A/2B, recombinant constructs which contains the chloramphenicol acetyltransferase protein (CAT) and enhanced green fluorescent protein (EGFP) linked by the FMDV 2A sequence with four synonymous codons for G_2A were produced. The activities of all the F2As based plasmids were determined in CHO cells. The results showed that the synonymous codon usage patterns for G_2A at the cleavage site (2A/2B) have no effect on the cleavage efficiency. This suggests that the synonymous codon usage of 2A peptide has no effect on the cleavage efficiency of FMDV 2A element.     

Promoter Methylation of E-Cadherin,p16, and RAR-β2 Genes in Breast Tumors and Dietary Intake of Nutrients Important in One-Carbon Metabolism

Aberrant DNA methylation plays a critical role in carcinogenesis, and the availability of dietary factors involved in 1-carbon metabolism may contribute to aberrant DNA methylation. We investigated the association of intake of folate, vitamins B₂, B₆, B₁₂, and methionine with promoter methylation of E-cadherin, p16, and RAR-β₂ genes in archived tumor tissues from incident, primary breast cancer cases in a population-based case-control study. Real-time methylation-specific PCR was performed on 803 paraffin-embedded samples; usual dietary intake was queried from a food frequency questionnaire. Unconditional logistic regression was used to derive adjusted odds ratios and 95% confidence intervals for likelihood of promoter methylation for high compared to low intake of those 1-carbon nutrients. Overall, in case-case comparisons, dietary intakes of folate, vitamins B₂, B₆, B₁₂, and methionine were not associated with likelihood of promoter methylation of E- cadherin, p16, and RAR-β₂ for all cases combined or within strata defined by menopausal status and estrogen receptor status in this study. This finding, however, does not exclude the possibility that intake of such nutrients might have the ability to modulate promoter methylation in normal or premalignant (dysplastic) breast tissue.     

Evaluation of Protective Ensemble Thermal Characteristics Through Sweating Hot Plate,Sweating Thermal Manikin,and Human Tests

The purpose of this study was to evaluate the predictive capability of fabric Total Heat Loss (THL) values on thermal stress that Personal Protective Equipment (PPE) ensemble wearers may encounter while performing work. A series of three tests, consisting of the Sweating Hot Plate (SHP) test on two sample fabrics and the Sweating Thermal Manikin (STM) and human performance tests on two single-layer encapsulating ensembles (fabric/ensemble A = low THL and B = high THL), was conducted to compare THL values between SHP and STM methods along with human thermophysiological responses to wearing the ensembles. In human testing, ten male subjects performed a treadmill exercise at 4.8 km and 3% incline for 60 min in two environmental conditions (mild = 22°C, 50% relative humidity (RH) and hot/humid = 35°C, 65% RH). The thermal and evaporative resistances were significantly higher on a fabric level as measured in the SHP test than on the ensemble level as measured in the STM test. Consequently the THL values were also significantly different for both fabric types (SHP vs. STM: 191.3 vs. 81.5 W/m² in fabric/ensemble A, and 909.3 vs. 149.9 W/m² in fabric/ensemble B (p < 0.001). Body temperature and heart rate response between ensembles A and B were consistently different in both environmental conditions (p < 0.001), which is attributed to significantly higher sweat evaporation in ensemble B than in A (p < 0.05), despite a greater sweat production in ensemble A (p < 0.001) in both environmental conditions. Further, elevation of microclimate temperature (p < 0.001) and humidity (p < 0.01) was significantly greater in ensemble A than in B. It was concluded that: (1) SHP test determined THL values are significantly different from the actual THL potential of the PPE ensemble tested on STM, (2) physiological benefits from wearing a more breathable PPE ensemble may not be feasible with incremental THL values (SHP test) less than approximately 150–200 W·m², and (3) the effects of thermal environments on a level of heat stress in PPE ensemble wearers are greater than ensemble thermal characteristics.     

Four year immunogenicity of the RTS,S/AS02(A) malaria vaccine in Mozambican children during a phase IIb trial

Previous studies with the malaria vaccine RTS,S/AS02_A in young children in a malaria endemic area of Mozambique have shown it to have a promising safety profile and to reduce the risk of Plasmodium falciparum infection and disease.In this study, we assessed the antibody responses to the P. falciparum and hepatitis B components of the RTS,S/AS02_A vaccine over a 45 months surveillance period in a large phase IIb trial which included 2022 children aged 1-4 years at recruitment.The RTS,S/AS02_A vaccine induced high anti-circumsporozoite antibody levels with at least 96% of children remaining seropositive during the entire follow-up period. IgG titers decayed over the first 6 months of follow-up to about 25% of the initial level, but still remained 30-fold higher until month 45 compared to controls. Children with higher levels of naturally acquired immunity at baseline, assessed by blood stage indirect fluorescent antibody test, had slightly higher anti-circumsporozoite levels, after adjusting for the effect of age.The RTS,S/AS02_A vaccine also induced high levels of anti-hepatitis B surface antigen antibodies (seroprotection >97%).RTS,S/AS02_A vaccine is immunogenic and induces long-lasting anti-circumsporozoite antibodies, persisting at least 42 months after immunization. These antibodies may play a role in protection against malaria.     

Power for <Emphasis Type="Italic">T</Emphasis>-test comparisons of unbalanced cluster exposure studies

Studies of individuals sampled in unbalanced clusters have become common in health services and epidemiological research, but available tools for power/sample size estimation and optimal design are currently limited. This paper presents and illustrates power estimation formulas for t-test comparisons of effect of an exposure at the cluster level on continuous outcomes in unbalanced studies with unequal numbers of clusters and/or unequal numbers of subjects per cluster in each exposure arm. Iterative application of these power formulas obtains minimal sample size needed and/or minimal detectable difference. SAS subroutines to implement these algorithms are given in the Appendices. When feasible, power is optimized by having the same number of clusters in each arm k _A =k _B and (irrespective of numbers of clusters in each arm) the same total number of subjects in each arm n _A k _A =n _B k _B . Cost beneficial upper limits for numbers of subjects per cluster may be approximately (5/ρ) −5 or less where ρ is the intraclass correlation. The methods presented here for simple cluster designs may be extended to some settings involving complex hierarchical weighted cluster samples.     

Strong and persistent CD4 T-cell response in healthy adults immunized with a candidate HIV-1 vaccine containing gp120, Nef and Tat antigens formulated in three Adjuvant Systems

This randomized double-blind study aimed to determine the safety and immunogenicity of a gp120/NefTat candidate human immunodeficiency virus type 1 (HIV-1) vaccine formulated with one of three different Adjuvant Systems (AS02_A, AS02_V and AS01_B) in healthy HIV-seronegative adults. All vaccine formulations induced strong HIV-specific CD4⁺ T-cell responses characterized by high lymphoproliferative capacity and IL-2 production that were still detectable 18 months after last immunization, with strongest responses seen in the AS01_B group. Broad coverage was demonstrated against gp120, and to a lesser extent Nef, derived from the most common circulating clades (B, C and circulating recombinant form [CRF]-01). All vaccine formulations exhibited acceptable safety and reactogenicity profiles. The demonstration of superior CD4⁺ T-cell induction by AS01_B provides important guidance for future HIV vaccine development.     

Bayesian noninferiority test for 2 binomial probabilities as the extension of Fisher exact test

Noninferiority trials have recently gained importance for the clinical trials of drugs and medical devices. In these trials, most statistical methods have been used from a frequentist perspective, and historical data have been used only for the specification of the noninferiority margin Δ>0. In contrast, Bayesian methods, which have been studied recently are advantageous in that they can use historical data to specify prior distributions and are expected to enable more efficient decision making than frequentist methods by borrowing information from historical trials. In the case of noninferiority trials for response probabilities π ₁,π ₂, Bayesian methods evaluate the posterior probability of H ₁:π ₁>π ₂?Δ being true. To numerically calculate such posterior probability, complicated Appell hypergeometric function or approximation methods are used. Further, the theoretical relationship between Bayesian and frequentist methods is unclear. In this work, we give the exact expression of the posterior probability of the noninferiority under some mild conditions and propose the Bayesian noninferiority test framework which can flexibly incorporate historical data by using the conditional power prior. Further, we show the relationship between Bayesian posterior probability and the P value of the Fisher exact test. From this relationship, our method can be interpreted as the Bayesian noninferior extension of the Fisher exact test, and we can treat superiority and noninferiority in the same framework. Our method is illustrated through Monte Carlo simulations to evaluate the operating characteristics, the application to the real HIV clinical trial data, and the sample size calculation using historical data.     

An algorithm for the design of group sequential triangular tests for single‐arm clinical trials with a binary endpoint

Consider the problem of testing H₀:p?p₀ vs H₁:p>p₀, where p could, for example, represent the response rate to a new drug. The group sequential TT is an efficient alternative to a single‐stage test as it can provide a substantial reduction in the expected number of test subjects. Whitehead provides formulas for determining stopping boundaries for this test. Existing research shows that test designs based on these formulas (WTTs) may not meet Type I error and/or power specifications, or may be over‐powered at the expense of requiring more test subjects than are necessary. We present a search algorithm, with program available from the author, which provides an alternative approach to triangular test design. The primary advantage of the algorithm is that it generates test designs that consistently meet error specifications. In tests on nearly 1000 example combinations of n (group size), p₀, p₁, α, and β the algorithm‐determined triangular test (ATT) design met specified Type I error and power constraints in every case considered, whereas WTT designs met constraints in only 10 cases. Actual Type I error and power values for the ATTs tend to be close to specified values, leading to test designs with favorable average sample number performance. For cases where the WTT designs did meet Type I error and power constraints, the corresponding ATT designs also had the advantage of providing, on average, a modest reduction in average sample numbers calculated at p₀, p₁, and (p₀ + p₁)/2. Copyright © 2010 John Wiley & Sons, Ltd.     

<Emphasis Type="SmallCaps">l</Emphasis>-Arginine and B vitamins improve endothelial function in subjects with mild to moderate blood pressure elevation

Purpose

The aim of this trial was to investigate the influence of a dietetic product consisting of a unique combination of l-arginine with the vitamins B₆, folic acid and B₁₂ (Telcor^® Arginin plus) on endothelial dysfunction.

Methods

Subjects aged 40–65 years with mild to moderate blood pressure (BP) elevation not treated with anti-hypertensive drugs were randomly assigned to either the dietetic product (n = 40) or a matching placebo (n = 41) for 3 months with open follow-up for a further 3 months. Postprandial change in endothelial function was assessed using the validated reactive hyperaemia index (RHI) at 3 months compared to the study onset (RHI post–pre, visit 3–visit 1; ΔΔRHI). Secondary parameters included BP and plasma homocysteine concentration.

Results

The primary efficacy analysis revealed superiority of the nutritional intervention over placebo (p = 0.0349) in reducing the deterioration of endothelial function. While in the active group ΔΔRHI increased (0.371 ± 0.122), almost no change could be detected in the placebo group (0.031 ± 0.100), thus demonstrating a significant improvement in vascular function in the intervention group. Moreover, the intervention reduced BP and homocysteine levels. Non-serious adverse events were equally distributed in both groups, and none of the events were assessed as possibly intervention-related by the investigators.

Conclusions

This trial confirmed the effective and safe use of dietary management with l-arginine in combination with B vitamins. The primary efficacy analysis demonstrated a statistically significant superiority of the combination of l-arginine with B vitamins over placebo in improving and restoring impaired endothelial function and lowering BP in patients with mild to moderate blood pressure elevation.

     

Association of Methylation Signatures at Hepatocellular Carcinoma Pathway Genes with Adiposity and Insulin Resistance Phenotypes

Obesity and type 2 diabetes mellitus are independent risk factors for the onset and progression of hepatocellular carcinoma (HCC). This study aimed to analyze the association of DNA methylation signatures at HCC pathway genes with obesity and related metabolic disturbances. A population of 474 adults within the Methyl Epigenome Network Association (MENA) project was included. DNA methylation levels were measured in white blood cells by microarray. The identification and discrimination of HCC pathway genes were performed using KEGG and PathDIP databases. Anthropometry measurements, the blood metabolic profile, and clinical data were analyzed. The methylation patterns of 20 CpG sites at HCC pathway genes strongly correlated with BMI (FDR <0.0001). These genes encompassed GADD45A, MTOR, FRAT2, E2F3, WNT7B, FRAT1, LRP5, DPF3, GSTA2, APC, MYC, WNT10B, ARID1B, AKT1, GSTA1, WNT5A, CDK4, GAB1, TCF7, which statistically contributed to the regulation of the HCC pathway (P?=?2.10e-07). The main biological process where these genes were implicated included uncontrolled cell proliferation, DNA damage, increased survival, and altered oncogenic expression. Interestingly, 9 out of 20 BMI-associated CpGs also correlated with waist circumference and HOMA-IR index. In conclusion, pathway analysis revealed potential associations of DNA methylation signatures at HCC pathway genes with adiposity and insulin resistance phenotypes.     

Biological Variability and Impact of Oral Contraceptives on Vitamins B6, B12 and Folate Status in Women of Reproductive Age

Vitamins B₆, B₁₂ and folate play crucial metabolic roles especially during the reproductive years for women. There is limited reporting of within-subject variability of these vitamins. This study aimed to determine the within and between subject variability in serum vitamins B₆, B₁₂, folate and erythrocyte folate concentrations in young women; identify factors that contribute to variability; and determine dietary intakes and sources of these vitamins. Data were obtained from the control group of a trial aimed at investigating the effect of iron on the nutritional status of young women (age 25.2 ± 4.2 year; BMI 21.9 ± 2.2 kg/m²). The coefficients of variability within-subject (CV_I) and between-subject (CV_G) for serum vitamins B₆, B₁₂ and folate, and erythrocyte folate were calculated. Food frequency questionnaires provided dietary data. CV_I and CV_G were in the range 16.1%–25.7% and 31.7%–62.2%, respectively. Oral contraceptive pill (OCP) use was associated (P = 0.042) with lower serum vitamin B₁₂ concentrations. Initial values were 172 ± 16 pmol/L and 318 ± 51 pmol/L for OCP and non-OCP users, respectively; with differences maintained at four time points over 12 weeks. BMI, age, physical activity, alcohol intake and haematological variables did not affect serum or erythrocyte vitamin concentrations. Vitamin B₁₂ intakes were derived from traditional and unexpected sources including commercial energy drinks. Young women using OCP had significantly lower serum vitamin B₁₂ concentrations. This should be considered in clinical decision making and requires further investigation.