Cross-sectional association of blood pressure variability and night-time dipping with cardiac structure in adolescents

Greater blood pressure variability (BP) and reduced night-time BP dipping are associated with cardiovascular disease independently of mean BP in adults. This study examines whether these associations are apparent in adolescents. A cross-sectional analysis was undertaken in 587 UK adolescents. We examined associations between measures of blood pressure dipping and variability (including standard deviation weighted for day/night (SDdn), average real variability (ARV) and variability independent of the mean (VIM)) with cardiac structure measures assessed by echocardiography: (1) left ventricular mass indexed to height2.7 (LVMi2.7), (2) relative wall thickness (RWT), (3) left atrial diameter indexed to height (LADi), and (4) left ventricular internal diameter in diastole (LVIDD)). Greater BP variability was associated with cardiac structures including higher RWT, which persisted after adjustment for mean BP. There was no evidence for an association between night-time dipping and cardiac structures. Measurement of BP variability might benefit cardiovascular risk assessment in adolescents.


Introduction
Higher blood pressure (BP) is associated with an increased risk of cardiovascular disease (CVD) 1 . However, BP is inherently variable, and under a typical circadian rhythm night-time BP is lower than daytime 2 . Loss of this nocturnal dipping pattern in the adult general population of adults has been shown to be associated with cardiovascular events and all-cause mortality, independent of 24-hour BP 2, 3 . In is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint addition, there is also evidence that non-circadian variability in BP may be associated with cardiovascular disease 2,4,5 .
Cardiovascular pathology starts in early life: childhood BP levels are known to track across life 6 and early adulthood BP associates with CVD mortality 7 . In adults, higher left ventricular (LV) mass and left atrial enlargement are both associated with higher risk of CVD 8,9 and are considered evidence of target organ damage 10 . Another measure of left heart function, relative wall thickness (RWT, a measure of remodelling 11 ), has been suggested to be predictive of stroke among adult populations 12,13 . We previously demonstrated that in 17 year-olds that higher body mass index (BMI) is causally related to higher LV mass indexed to height 2.7 (LVMi 2.7 ) 14 , suggesting that there is meaningful variation in cardiac structure measures in early adulthood. A study in adults from the general population indicated a positive association between BP variability and LVMi 15 . However, it is not known whether 24hour BP variability and night-time dipping in adolescents are related to cardiac structure.
In this study, we used data from a prospective cohort study of 587 UK adolescents to assess the cross-sectional associations of mean BP (from clinic measurements and ambulatory monitoring), BP variability, and night-time dipping, with measures of cardiac structure at age 17, determined by echocardiography. The measures of cardiac structure we consider are 1) LV mass (LVM), 2) RWT 11 , 3) left atrial diameter (LAD), and 4) left ventricular internal diameter during diastole (LVIDD, a measure of the initial stretching of cardiomyocytes before contraction (preload)) 16 . Together these . CC-BY 4.0 International license It is made available under a perpetuity.
is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint represent a comprehensive assessment of left heart structure, with functional significance 17 .

Participant characteristics
A total of 587 participants were included in our analysis. Figure 1 shows how this cohort size was reached from the participants enrolled in ALSPAC at baseline.
Compared with the full ALSPAC cohort, the participants included in our analysis tended to have mothers who were more educated and older when the participant was born and be from a family with a higher head of household occupational social class; females were also more likely to be included. Clinic blood pressure, minutes of moderate to vigorous physical activity at age 15 and DXA-determined fat mass were similar compared with the full ALSPAC cohort (Supplementary Table 1).
Males tended to have higher systolic blood pressure, pulse pressure and mean arterial pressure, while females had higher diastolic blood pressure. Night-time dipping was similar between sexes. Males tended to have higher systolic and diastolic BP . CC-BY 4.0 International license It is made available under a perpetuity.
is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint 4 variability than females. Ventricular measures were higher in males, while atrial index and wall thickness were similar between sexes (Table 1).
There was no evidence of associations with LADi or LVIDD.
Results were broadly similar in the age and sex only adjusted models (Supplementary Table 2).

Associations between ambulatory averages of BP and cardiac structures
There was evidence of a positive association between 24-hour mean SBP and LVMi 2.7 (β = 0.17 SDs per SD higher 24-hour SBP, 95% CI 0.093 to 0.25, P=1.8x10 -5 ), which was slightly smaller in magnitude than the association for clinic SBP (Figure 2).
Daytime and night-time means for SBP also showed positive associations with LVMi 2.7 , with similar magnitudes to 24-hour mean SBP. The 24-hour mean SBP also showed a positive association with RWT (β = 0.18, 95% CI 0.089 to 0.26, P=8.1x10 -5 ), with similar magnitudes of association seen for daytime and night-time mean SBP.
There was no evidence of associations between 24-hour, day-time or night-time mean SBP and LADi or LVIDD. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint There was evidence for associations between all 24-hour DBP measures (mean, day and night) and RWT, with similar magnitudes of associations between the three exposures, but no evidence of associations for the other measures of cardiac structure.

2) Associations between 24-hour blood pressure variability and cardiac structures
ARV of SBP was associated with LVMi 2.7 after adjustment for confounders ( Table 2).
DBP variability measures were positively associated with RWT, although evidence of association was weaker for VIM than for SDdn and ARV. ARV and VIM of DBP were also positively associated with LVMi 2.7 and LADi.
After further adjustment for 24-hour BP ( Table 3), associations of SBP and DBP variability with LVMi 2.7 and RWT attenuated towards the null. Some associations with RWT remained: before adjustment for mean DBP the standardised association between ARV of DBP and RWT was 0.13 (95% CI 0.045 to 0.21, P=2.7x10 -3 ). After adjustment for mean DBP it was 0.11 (95% CI 0.022 to 0.19, P=0.014).

3) Associations between night-time BP dipping and cardiac structures
The results provided no evidence for associations between either of the dipping variables (percentage difference and categorical) and cardiac structures (Supplementary Table 2, Table 2, Table 3). This was true for both SBP and DBP. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint 6

Complete case analysis
For all analyses, there were similar magnitudes of estimates between the complete cases and imputed analyses (Supplementary Table 3 and Table 3). However, as there was less power in the complete case analysis, confidence intervals were wider.

Discussion
In this cross-sectional study of a general population of adolescents, we have shown Variability in BP over 24 hours in this sample of adolescents was similar in magnitude to that reported in studies of adults 38,39 . In contrast, the percentage of normal dipping was higher than in adult studies 26 . In adults, greater variability in BP and non-dipping is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint are associated with cardiovascular risk, independently of average BP 2,26,40,41 . Higher visit-to-visit BP variability in children has been shown to be associated with adult hypertension in the Bogalusa Heart Study 42 . Two previous studies, restricted to hypertensive children, did not find an association between 24-hour BP variability and LVMi 2.7 43, 44 . Similarly, several studies have found little association between night-time dipping and LVMi in hypertensive children 25,45,46 . To our knowledge, this is the first study to explore these associations in a general population cohort of adolescents.
Higher mean SBP is associated with higher LVMi 2.7 and RWT in our study. This finding, together with our previous finding that higher BMI is causally related to higher LV mass 14 , suggests that higher values of LVMi 2.7 and RWT are, on average, related to adverse cardiovascular health even in this young population, rather than due to high levels of fitness. This implies that the cardiac structures are meaningful markers of cardiac health in this young population. Both DBP and SBP were associated with RWT to a similar extent. However, unlike SBP, DBP did not show associations with LVMi 2.7 . This could reflect a greater importance of systolic pressure (and by implication pulse pressure on LV mass) or it may be at least partially driven by regression dilution bias because of the greater levels measurement error for DBP compared with SBP 47 .
Our results indicate that some associations between greater BP variability and cardiac structure remained once average BP was accounted for, such as diastolic measure of ARV with RWT. These findings support the notion that the influence of BP variability on cardiac structure may begin early in life 42 . We found no convincing evidence of an association between non-dipping and cardiac structure in young people. Findings in older adults are inconsistent 48 and most studies finding a positive association between . CC-BY 4.0 International license It is made available under a perpetuity.
is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint non-dipping and LV mass have been conducted in hypertensive individuals 49 . The majority of the participants in our sample had blood pressures in the normal range; other studies which included such participants have also not found evidence of an association 50 .
Previous research used smaller participant numbers than the current study and tended to focus on hypertensive children 43, 45, 46 . It is possible that our study may have lacked statistical power to detect some associations between BP variability and dipping and cardiac structure independently of mean BP. However, this is the only available dataset with a highly detailed range of clinic and ABPM measurements for each participant in this age group. One of the strengths of our analysis is the inclusion of adolescents from the general population which provides a more representative sample of UK adolescents. A possible limitation, however, is that the cohort are of European descent and in a localised area of the UK, which may limit its external validity.
Our study uses cross-sectional data from a birth cohort study. This limits our ability to determine the true direction of the association between blood pressure and cardiac structures, and whether this relationship may be causal. The participants included in our analysis are more affluent than the full ALSPAC cohort 18 . However, whilst this does affect the generalizability of the study, it does not necessarily lead to bias in the estimates of associations. There is some evidence that lack of generalisability on cohort studies does not bias exposure-outcome associations 51 .
Another possible limitation of this study is that ABPMs affect sleep quality due to the cuff inflating throughout the night, which may affect night-time dipping levels. A . CC-BY 4.0 International license It is made available under a perpetuity.
is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint previous study found that those with lower quality of sleep had higher nocturnal BP levels and a smaller BP dip 52 ; this could weaken associations. Despite this limitation, a recent large cohort study has confirmed that 24-hour ambulatory blood pressure monitoring is a stronger predictor of cardiovascular and all-cause mortality than clinic measurements, confirming the validity of its use 28 .
Our results show that, in adolescents, higher clinic and 24-hour SBP and DBP, as well as an increase in blood pressure variability, are associated with more adverse cardiac structure. Non-dipping was not found to be associated with cardiac structure. Our study implies that measurement of BP variability, but not night-time dipping, might add to the assessment of cardiovascular risk in adolescents. However, this finding would benefit from replication in larger studies. It would be valuable to explore whether BP variability and dipping in adolescents track across the life course, and whether these BP measurements in adolescents are predictive of longer-term cardiovascular outcomes.

Methods
Because of the sensitive nature of the data collected for this study, requests to access the dataset from qualified researchers trained in human subject confidentiality protocols may be submitted via the Avon Longitudinal Study of Parents and Children is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020.  (Figure 1). Since birth, participants have been followed up, using questionnaires, links to routine data, and research clinics. The study website provides further details of the cohort and a data dictionary http://www.bris.ac.uk/alspac/researchers/data-access/data-dictionary/.
Approval was obtained from the local ethics committee and the ALSPAC Law and Ethics committee. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint

Inclusion/Exclusion Criteria
The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint and hourly at night. Participants were permitted to perform usual physical activities, although a diary of activities was recorded. Daytime and night-time were defined by the participant. The expected maximum number of total readings per participant therefore varied depending on the duration of the night-time period. For this study, we included participants with at least 14 readings during the self-defined daytime and at least 5 readings during the self-defined night-time 20, 21 .
We estimated the mean 24-hour SBP and DBP using the ABPM data, and also estimated the daytime and night-time means for SBP and DBP. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint

Confounders:
We considered variables as confounders if they had plausible relations with BP and cardiovascular risk 28 . Maternal confounders were self-reported in questionnaires completed during pregnancy: educational attainment (categorised as university degree or higher, Advanced-levels (exams usually taken around 18 years and necessary for university entry), Ordinary-levels (exams usually taken around 16 years, which was the minimum UK school leaving age at the time these participants were this age), or lower than Ordinary-levels, including vocational education); pre-pregnancy body mass index (BMI; in kg/m 2 ); age at delivery (categorised as <25 years, 25-35 years, and >35 years); parity, and highest head of household occupational social class. We selected these maternal variables as the mother's socioeconomic position (SEP) represents the participant's family SEP. SEP has been shown to influence BMI (a key determinant of both BP and LVM 14 ), blood pressure 29 , and left ventricular structure 30 . Maternal pre-pregnancy BMI has also been shown to affect offspring BP and cardiovascular outcomes 31 .
Child-based confounders were from a combination of self-reported questionnaire and clinic-based data: age (in months) at year 17 clinic visit; smoking at age 17 (<1 or ≥1 cigarette per week from self-report); minutes of moderate to vigorous physical activity  is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint 14 Statistical analysis: All analyses were performed using Stata version 15.1 (StataCorp, TX).
We used multivariable linear regression to estimate the associations between all blood pressure exposures and cardiac structure outcomes defined above. We standardised all exposures and outcomes before analysis to have a mean of zero and SD of one. additional adjustment for potential confounders: maternal education, age at delivery, parity, pre-pregnancy BMI; household socio-economic class; smoking at age 17; minutes of moderate to vigorous physical activity at age 15; DXA-determined fat mass and height and height 2 at age 17, iii) further adjustment for average 24-hour blood pressure (systolic or diastolic as appropriate for the exposure) to evaluate whether any associations between BP variability and dipping were independent of 24-hour average BP.
To test for interactions between sex and each exposure, we regressed each outcome on each exposure, with sex and an interaction term for the exposure and sex as covariables. There was no strong evidence of any interactions by sex from these analyses (p>0.1 for all interaction terms), and as such, all results are presented for . CC-BY 4.0 International license It is made available under a perpetuity.
is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint males and females combined. We did not correct the results for multiple testing, as multiple testing correction emphasises the inappropriate dichotomisation of p values into significant versus non-significant [32][33][34][35] . Furthermore, in this analysis, exposures are correlated measures of a single underlying construct BP, and outcomes are measures of a single underlying construct, cardiac structure. A Bonferroni multiple testing correction would therefore be over-conservative. We interpret the overall pattern of results rather than focusing on single P values, and use the magnitude of coefficients and confidence intervals to assess the strength of associations.

Missing Data
Of the 587 participants with complete data on all 18 exposures and 4 outcomes, 196 (33.3%) also had complete data including all confounders. In the full dataset, individual confounder variables were missing between 0% and 43.4% of observations, with eight of 11 variables having less than 13% missingness (Supplementary Table 4). We used multivariate multiple imputation by chained equations to impute missing confounder data 36,37 . The imputation model included all exposures (excluding dipping variables, which were derived from other variables in the imputation model), outcomes and confounding variables, as well as weight and BMI at age 17, and maternal height. Fully conditional specification was used, with linear regression for continuous variables, multinomial regression for categorical variables and logistic regression for binary variables (Supplementary Table 4). We created twenty imputed datasets and used Rubin's rules to combine analysis results. Variable distributions were consistent between the imputed and the observed data sets (Supplementary Table 4).
We also conducted a complete case sensitivity analysis in the 196 participants with complete data for all variables (Supplementary Table 3). is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint correlation between visit-to-visit variability and 24-h variability of blood pressure. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint weighted for day and night, ARV = average real variability, VIM = variability independent of the mean, DBP = diastolic blood pressure, 4 LVMi 2.7 = left ventricular mass indexed to height 2.7 , LADi = left atrial diameter indexed to height, LVIDD = left ventricular internal 5 diameter during diastole, RWT = relative wall thickness. 6 7 8 Table 2. Associations of blood pressure measurements with cardiac structure, N=587 9 Analysis of multiply imputed data. Adjusted for sex, age at outcome assessment; maternal age at delivery, education, parity, and 10 maternal pre-pregnancy BMI; household social class; smoking at age 17; minutes of moderate to vigorous physical activity at age 11 15; DXA-determined fat mass, height and height 2 at age 17. Regression coefficients for continuous exposures are standardised, i.e. 12 they represent the change in SDs of the outcome (cardiac structure measurement) per one SD higher blood pressure.  Table 3. Associations of BP variability and dipping with cardiac structure after adjustment for 24-hour mean BP, N=587 19 Analysis of multiply imputed data. Adjusted for sex, age at outcome assessment; maternal age at delivery, education, parity, and 20 maternal pre-pregnancy BMI; household social class; smoking at age 17; minutes of moderate to vigorous physical activity at age 21 15; DXA-determined fat mass, height and height 2 at age 17; mean 24-hour blood pressure (systolic or diastolic, as appropriate for 22 the exposure). Regression coefficients are standardised, i.e. they represent the change in SDs of the outcome (cardiac structure 23 measurement) per one SD higher blood pressure. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted November 23, 2020. ; https://doi.org/10.1101/2020.11.20.20235473 doi: medRxiv preprint