Machine learning based deconvolution of microarray atrial samples from atrial fibrillation patients reveals increased fractions of follicular CD4+ T lymphocytes and gamma-delta T cells.

A potential relationship between T cell immunity and development of atrial fibrillation (AF) has been proposed. Historically in AF patients it has been reported that peripheral blood had elevated CD4+ T cells. However few studies have explored whether there is a direct increase of CD4+ T cells in atrial tissues with AF. In this study, public domain micro-array dataset of cardiac surgery patients with atrial tissue biopsies in AF and non-AF patients have been used to explore immune cell subsets. Machine learning based deconvolution of permanent atrial fibrillation microarray atrial samples was applied using Cibersort to enumerate the relative fractions of twenty-two different leukocyte sub-populations. Cibersort enumerated significantly increased fractions of follicular CD4+ T lymphocytes and gamma-delta T cells in the atria of permanent AF subjects. Gene expression analysis of permanent AF microarray tissue samples with elevated follicular CD4+ T cell fractions with gene pathways associated with myocardial substrate remodelling. That is both integrin and non-integrin mediated gene interactions between inflammatory cells and the extra cellular matrix, including infiltrating follicular CD4+ T cells that trafficked to the atria by virtue of the repertoire of cell surfaced expressed adhesion molecules. Additionally, IL-17 and other interleukin inflammatory gene heat maps were associated with enhanced CD4+ follicular T cell expression in our profiled atrial tissues with AF. These observations suggest that atrial structural remodelling was associated with the presence of pathogenic T cell mediated inflammation, present in AF atria but not in non-AF atria.

RUNX1T1 rs34269950 is associated with obesity and metabolic syndrome.

BACKGROUND: Runt-related transcription factor 1 (RUNX1T1) isoforms are involved in adipogenesis. RUNX1T1 is mediated by the fat mass and obesity-associated (FTO). However, the extent to which RUNX1T1 single-nucleotide polymorphisms (SNPs) are associated with obesity risk or metabolic abnormalities in a community population basis is unknown. METHODS: Samples were obtained from the Australian Crossroads study bio-bank. SNPs located in the coding region and 3'untranslated regions of RUNX1T1 with minor allele frequency ≥0.05 were analysed using Taqman genotyping assays. RESULTS: Eight candidate SNPs were genotyped successfully in 1440 participants. Of these SNPs only rs34269950 located in the 'RRACH' motif, the most common N6-methyladenosine (m6A) methylation modification site (recognized by FTO), was significantly associated with obesity risk and metabolic abnormalities. Specifically, compared to AA genotype, rs34269950 del/del genotype was associated with a 1.47 [95% confidence interval (CI): 1.01-2.14, P = 0.042] fold higher rate of obesity risk. Additionally, the del/del genotype was associated with a 60% increased risk of metabolic syndrome (MetS) [odds ratio (OR) = 1.60, 95% CI: 1.10-2.32, P = 0.015], in comparison to the AA genotype. Finally, rs34269950 del/del increased the risk of a larger waist circumference (OR = 1.65, 95% CI: 1.15-2.36, P = 0.007), but not other components of MetS. CONCLUSION: Our study demonstrates that RUNX1T1 rs34269950, located in a potential FTO recognition motif, is significantly associated with waist circumference. This provides novel evidence to suggest SNPs located in RRACH motif may be involved in RNA m6A modification and mechanistic pathways that influence abdominal obesity.

Sex-specific educational attainment is associated with telomere length in an Australian rural population.

BACKGROUND: There is limited understanding on whether and how socioeconomic status (SES), particularly educational attainment and household income, impacts on telomere length in an Australian rural context. Additionally, it is unknown whether access to health services via the Australian public or private health system influences telomere length. AIM: This study investigates whether there is a relationship between telomere length and SES indicators (income, education) as well as health insurance status in a rural Australian population. METHODS: Samples were drawn from the Australian Rural Victoria cross-sectional Crossroads Study. Leucocyte telomere length (LTL) was measured using a multiplex quantitative polymerase chain reaction method. RESULTS: Among 1424 participants, we did not find a significant main effect association with LTL across education, income level and health insurance. An exploratory finding was sex may influence the relationship between educational attainment and LTL (P = 0.021). In males, but not females, higher education was associated with longer LTL by 0.033 [95% confidence interval (CI) 0.002-0.063, P = 0.035]; in those with low education attainment, male participants had shorter LTL by 0.058 (95% CI -0.086 to -0.029) than female participants (P < 0.0001). CONCLUSION: Being male and having lower education attainment was associated with shorter telomere length in our rural population. Evidence from our study supports the importance of education on LTL in males in rural Australia. Our studies also support previous findings that LTL in later life may not be closely associated with indicators of SES.

rs9939609 FTO genotype associations with FTO methylation level influences body mass and telomere length in an Australian rural population.

BACKGROUND: The fat mass- and obesity-associated (FTO) gene influences energy homeostasis in humans. Although the obesity-related variant, rs9939609 has been replicated across a number of cohort studies, there remains significant variance and a low to modest association. Telomere length is another commonly reported obesity risk factor. We hypothesize understanding the associations between FTO rs9939609 with FTO methylation and telomere length will provide a more accurate assessment of obesity risk. METHODS: Overall, 942 participants free of diabetes or pre-diabetes were included in the retrospective study. Leukocyte genomic DNA was analyzed for rs9939609 genotyping, FTO gene methylation and leukocyte telomere length (LTL) measurement. RESULTS: In general linear models, rs9939609 AA genotypes were associated with increased fat percentage (3.15%, P=0.001), fat mass (4.16 kg, P=0.001), body mass index (BMI) (1.38, P=0.006) and waist circumference (3.35 cm, P=0.006), but not with FTO methylation or LTL in this overall population. However, when participants were stratified into higher and lower FTO methylation groups, the AA genotype possesses a 2.04-fold increased obesity risk in comparison to TT genotype (95%CI, 1.07-3.89, P=0.031) in participants with a higher FTO methylation level, but this association was absent in the lower FTO methylation sub-group. Moreover, AT and AA genotype carriers were associated with shorter LTL compared to TT carriers (P=0.020 and P=0.111, respectively) in the higher FTO methylation level group. However, this association was absent in the lower methylation group. Furthermore, FTO gene methylation level was significantly associated with LTL in the 942 samples (P=0.017). CONCLUSIONS: FTO rs9939609 is associated with obesity risk and LTL in this study, where this association is only observed at higher, but not lower, FTO methylation levels of participants. Our data suggest association of multiple factors, including FTO methylation level, may be involved in one of several mechanisms underlying the commonly reported obesity risk of this FTO polymorphism.

Cognitive impairment and reduced quality of life among old-age groups in Southern Urban India: home-based community residents, free and paid old-age home residents.

AIM: The purpose of the study were (i) to screen for cognitive impairment using Mini-Mental Status Examination among three old-age groups based on dwelling types in Chennai, India i.e. residential paid old-age homes, residential free (charitable) homes and home-based community-dwelling residents; (ii) secondly to investigate factors (demographic, psychological, medical and disability) associated with cognitive impairment in the these old-age; (iii) third, to investigate the independent association between cognitive impairment and health-related quality of life (QOL) among elderly across aged care dwelling types. METHODS: A total of 499 elderly from three old-age groups were interviewed in this cross-sectional study (173 elderly home-based community-dwellers, 176 paid-home and 150 free-home residents). All the participants were interviewed for their socio-economic condition, medical morbidity, self-reported worry and anxiety, disability and QOL. RESULTS: 42.7% free-home elderly residents were found to have cognitive impairment, whereas 32.4% of paid-home and 21.9% of community-dwelling elderly had cognitive impairment. The residents of free-home were less educated, had lower income and reported higher incidence of worry, anxiety, disability and poor QOL than community-dwelling or paid-home residents. Increasing age, low education, female gender, high blood pressure and disability were associated with cognitive impairment. Cognitive impairment had significant negative effect on their health-related QOL (b = -0.10, P = 0.01), independent of age, gender, education, chronic illness and dwelling type. CONCLUSION: The burden of cognitive impairment was high in all aged-care dwelling types in urban India; with free charitable home residents being worse affected. Cognitive impairment was associated with disability and poor health-related QOL in these age-care settings.

Chronic NG-nitro-l-arginine methyl ester (L-NAME) administration in C57BL/6J mice induces a sustained decrease in c-kit positive cells during development of cardiac hypertrophy.

Chronic NG-nitro-l-arginine methyl ester (L-NAME) administration induces cardiac hypertrophy in rodent models. Our aims is to determine the role of c-kit expression in L-NAME induced cardiac hypertrophy. 12-20 week old C57BL/6J mice (5 per group) were administered L-NAME (0.325mg/ml) in the drinking water. Hearts were excised at 1-day, 2-days, 5-days, 2-weeks or 6-weeks; or controls which received no L-NAME. Ventricular cross-sectional wall thickness and individual cardiac myocytes cross-sectional area and cardiomyocyte/nuclear ratio to determine cardiac hypertrophy. Immuno-histochemical staining for c-kit, sca-1 and BCRP undertaken. Six weeks L-NAME administration induced significant cardiac hypertrophy compared to control hearts, evidenced by an increase in the thickness of the cross-sectional free ventricular wall (p<0.05) and an increase in mean individual cross-sectional area of cardiac myocytes in the LV wall (p<0.007). We observed c-kit(+) cells (predominately non-mast cell sub-types) in both healthy mice and in the L-NAME treated mice. C-kit staining in the left ventricular cross sections following L-NAME remained stable at 1 and 2 days compared to controls (p=NS). After 5 days of L-NAME we observed c-kit expression to decrease below control levels (p<0.05) and these lower levels were sustained at 2 and 6 weeks. C-kit expression does not decrease during two days of L-NAME administration, suggesting, firstly, that the later decrease in c-kit is not due to NOS inhibition directly and, secondly, there is the possibility for c-kit(+) cell differentiation into other cell types, possibly inducing myocardial cellular hyperplasia, without significant replacement of the original pool of c-kit(+) cells.

Chronic high dose captopril decreases total heart rate variability and increases heart rate in C57BL/6J mice.

The effects of high dose captopril, within the therapeutic range, on autonomic activity are unknown in those with normal cardiovascular function. Thus the study aims were to assess the effects of high dose captopril on autonomic function in mice. Autonomic activity was measured using heart rate variability (HRV). ECG recordings were obtained from 18 Male C57BL/6J mice (20-25 g) subdivided into control (N=8) or mice receiving oral captopril (0.688 mg/ml captopril in the drinking water for 6 weeks, N=10). HRV results for linear and non-linear parameters were attenuated following chronic captopril for 6 weeks compared to control. Captopril was associated with a trend for an increase in average heart rate and approximate entropy (ApEn), a non-linear measure of HRV decreased significantly compared to control (p<0.05). In conclusion high dose captopril reduces total HRV and increases heart rate in normotensive mice with normal cardiac function.

Co-ordinated intra-abdominal and intra-thoracic pressures regulate cough assisted cardiac circulatory support.

Periodically case studies emerge that demonstrate voluntary and forced coughing in ventricular fibrillation or asystole can maintain consciousness for up to 100 s. Forced cough is effective in producing co-ordinated abdominal muscle contraction and increases in intra-abdominal and thoracic pressures. These co-ordinated pressure changes are likely to be responsible for assisted circulation. In this short communication we discuss some of the possible abdominal physiological mechanisms for cough assisted circulatory support.

Differences in anxiety-related behavior between apolipoprotein E-deficient C57BL/6 and wild type C57BL/6 mice.

The influence of ApoE gene deletion on the anxiety state has not been previously investigated. The elevated plus maze was used in this study to determine differences in anxiety-related behavior between apoE-deficient and wild type C57BL/6 mice. The apoE-deficient mice demonstrated less anxiety on the elevated plus maze by spending more time in the open arms of the elevated plus maze compared to wild type mice (p<0.001). Additionally, female apoE-deficient mice visited the open arm of the maze more often than their apoE-deficient male counterpart (p<0.05). The anxiety state and/or sex are possible variables to be considered when designing physiological and/or behavioral studies involving mice that are apoE-deficient.

The CCK2 agonist BC264 reverses freezing behavior habituation in PVG hooded rats on repeated exposures to a cat.

Our previous studies (NeuroReport 12 (2001) 2717) showed that PVG hooded and not Sprague-Dawley (SD) rats exhibit remarkable freezing behavior on exposure to a cat in the cat freezing test apparatus. In the present study, we further examined the differences between these two strains of rats in response to repeated daily exposure to a cat in the cat freezing test apparatus. Freezing behavior habituation was observed in both PVG hooded (days 5-7) and SD rats (days 3-7). A selective CCK(2) agonist (BC264, 0.3 microg/kg, i.p.) on day 8 reversed habituated freezing behavior and locomotor activity in PVG hooded rats, but not in SD rats. These results suggest that CCK2 receptors mediate habituation to an anxiety-inducing stimulus in PVG hooded rats and further suggest that differential expression of these CCK2 receptors underlies this strain difference.

A method to determine the fractal dimension of the cross-sectional jaggedness of the infarct scar edge.

This study describes the use of a shareware software package available from the National Institutes of Health for computing the fractal dimension. Specifically, when fractal analysis is used in its correct context it provides for a quantitative description of the space filling properties of two-dimensional objects. A rabbit model of post myocardial infarction is described where the cross-sectional infarct edge is reconstructed and its jaggedness determined by calculating its fractal dimension via the pixel dilation method. The fractal dimensions of the anterior and posterior lateral infarct edges were calculated to have a mean of 1.16 and 1.29, respectively. In conclusion, the fractal technique can be used to describe the complex jaggedness of the infarct edge. This case study also illustrates the fact that the complexity of an infarcted area is not uniform across the scar. For example, we found that the space filling properties of the anterior and posterior borders of a myocardial infarct can differ by more than 2-fold.

Cross-sectional infarct edge jaggedness does not influence ventricular electrical stability in a rabbit model of late myocardial infarct healing.

Previous studies have suggested that the jaggedness of the healed or healing infarct edge influences cardiac electrical stability. However, these findings have been based on histological observations rather than quantitative measurements. The aim of this study was to assess infarct jaggedness by calculating its fractal dimension and to examine how this influences cardiac electrical stability during late infarct healing in the rabbit. Using programmed electrical stimulation, it was found that the fractal dimension did not differ significantly in 19 rabbits that had inducible ventricular tachycardia and 16 that did not. We conclude from these studies in the mature rabbit that infarct edge jaggedness does not influence the ease with which ventricular tachycardia is induced during late myocardial infarct healing.

A semi-quantitative PCR method for the detection of low levels of apoptotic DNA fragmentation in a heart failure model.

Cells that are apoptotic and comprise less than 2% of the total cellular population are difficult to detect by conventional methods (i.e., DNA ladder). We discuss a new methodological technique, PCR-amplified DNA ladder, to detect very low levels of DNA fragmentation (indicative of apoptosis) in a myocardial infarct heart failure model. Results and methodology are contrasted with the traditional DNA ladder technique.

Delay in opening the infarct related coronary artery increases plasma atrial natriuretic peptide levels.

Plasma atrial natriuretic peptide (ANP) levels were measured in rabbits during the late healing phase of myocardial infarcts. Significant differences in plasma ANP levels (P < 0.02) were found between rabbits that had undergone very late (6 h) or early reperfusion (20 and 45 min of ischemia) of the infarct related coronary artery. Differences in ANP levels were independent of infarct size, ventricular remodeling and infarct expansion. We conclude late reperfusion of infarct related artery, independent of myocardial salvage, is associated with increased circulating ANP plasma levels.

Immediate and sustained effects of smoking on autonomic arousal in human subjects.

The effects of smoking on sudomotor/autonomic activity were examined by measuring water transfer across the skin (sweat output) as an index of activity. Sweat output was measured in 14 subjects (11 male and 3 female) during the act of smoking and for about 60 min following this. Sweat output was measured in 5 (4 male, 1 female) controls over the same time period. Smoking had two effects on sweat output: In 12 subjects it caused an immediate increase in output; in 4 of these 12 the increase persisted for the duration of the recording period. In the other 2 subjects no increase was noted, but in no subject did smoking cause a decrease in sweat output. Mood state questionnaires were administered at the beginning and end of the experimental period. No clear association was found between scores on the mood questionnaires and the autonomic effects of smoking. In 2 subjects, transdermal blood flow was also measured during and after smoking. Smoking caused a decrease in blood flow in these subjects. These results are discussed in terms of the "arousal modulation" hypothesis of smoking.