E-cigarette use is disparately high among sexual minoritized populations. As e-cigarette advertising may influence product appeal, this study tested sexual orientation- and gender-based differences in response to e-cigarette advertisement exposure on advertisement perceptions and product appeal.We recruited 497 adults  living in the United States via the crowdsourcing platform Prolific. Participants viewed two randomly selected e-cigarette advertisements (from n=173 advertisements). Post-exposure, participants rated the perceived advertisement effectiveness, relevance, and product use intention. Associations between sexual orientation and outcomes were estimated using multivariable linear mixed-effects models. We tested interaction effects between sexual orientation, gender, and advertisement feature , and ran Tukey post hoc tests for pairwise comparisons.Post-exposure, heterosexual women, sexual minoritized men, and sexual minoritized women  rated perceived advertisement effectiveness and relevance lower after viewing advertisements featuring flavors . Sexual minoritized men and sexual minoritized women rated perceived advertisement relevance lower after viewing advertisements featuring humans  or fruit . Heterosexual women, sexual minoritized men, and sexual minoritized women reported lower product use intention after viewing advertisements featuring an e-liquid bottle .Sexual minoritized women and men reported lower e-cigarette advertisement appeal and product use intentions than heterosexual men. More evidence is needed to understand advertisement perceptions and product appeal in this group to inform e-cigarette advertising regulations and anti-tobacco messaging campaigns that aim to reduce tobacco-related health inequities. However, these differences vary across subpopulations of sexual minoritized adults. For example, research using the 2016–2018 Population Assessment of Tobacco and Health (PATH) Study data found that 21.7% of bisexual men aged 18–29 years and 13.2% of bisexual men aged 30–44 years reported past 30-day e-cigarette use 2. However, past 30-day e-cigarette use rates were lowest among gay men aged 18–29 years (12.9%) and 30–44 years (7.5%)2. Among women, past 30-day e-cigarette use rates were lowest among heterosexual women aged 18–29 years (7.9%) and 30–44 years (5.0%), compared to lesbian (17.6%) and bisexual (20.9%) women aged 18–29 years, and lesbian (16.4%) and bisexual (19.0%) women aged 30–44 years2. In general, tobacco product use tends to be higher among sexual minoritized women, particularly those who identify as bisexual2.E-cigarette use is disparately high among sexual minoritized populations , with 13.2% of US adults within this population reporting current use, over three times that of heterosexual adults (4.1%)4. Tobacco use is also described as a ‘counterculture’ strategy to resist social norms5, a means to build social connections within lesbian, gay, bisexual, and transgender (LGBT) communities6, and a way to signal one’s LGBT identity and belonging6. Big Tobacco’s historically targeted marketing to LGBT groups  is likely a contributing factor to marked e-cigarette use disparities among sexual minoritized people7. The tobacco industry has consistently tailored advertisements to LGBT populations while also placing targeted marketing in LGBT publications and event spaces8. Yet, until recently, tobacco prevention and control efforts have largely overlooked sexual minoritized populations, including the role of tobacco marketing in e-cigarette product appeal and use.The factors underlying e-cigarette use disparities among sexual minoritized populations are multilevel and complex. Generally, sexual minoritized adults describe tobacco use as a strategy to resist and cope with excess stress arising from structural violence and minority stress 9. However, there are differences by sexual orientation and gender. For example, in a study of US adults, sexual minoritized adults (both men and women) generally reported more exposure to tobacco industry marketing than their heterosexual peers, yet only lesbian and bisexual women were more receptive to this marketing 9. In contrast, in a nationally representative sample of young adults in the US, bisexual women reported greater exposure to e-cigarette marketing than heterosexual women, but these differences were not evident for young adult lesbian women or gay and bisexual men 10. These disparities in tobacco advertising exposure among sexual minoritized groups are further exacerbated by the intersectionality of race, ethnicity, and gender11, as the tobacco industry also has a history of targeting these groups11. For example, in a study of US young adults, bisexual Hispanic women had higher exposure to e-cigarette, cigarette, and cigar advertisements , whereas Black bisexual women had higher exposure to cigarette and cigar marketing  and Black bisexual men had higher exposure to cigar advertisements . However, there were no differences by race or ethnicity among lesbian women or gay men10. These findings underscore the importance of examining differences in tobacco marketing exposure among gender-based subgroups of sexual minoritized adults.Emerging literature has found that compared to their heterosexual peers, sexual minoritized adults have increased exposure to tobacco advertising and find tobacco industry marketing more appealing12. A study found that sexual minoritized adults were more likely to have viewed and interacted with (i.e. searched for and/or shared) tobacco advertisements, compared to their heterosexual peers13. This study also found that sexual minoritized participants reported significantly higher odds of past 30-day e-cigarette and tobacco use13.Excess exposure to tobacco industry marketing among the sexual minoritized populations is concerning, as prior evidence links e-cigarette advertisement exposure to product appeal and use15. An eye-tracking study found gender-based differences in time of viewing certain advertisements14, which may also suggest that there are differences in the impact of advertisement features based on gender and sexual identity. Thus, understanding how the perceived effectiveness and relevance of certain e-cigarette advertisement features appeal to other subgroups, such as sexual minoritized people, may help reduce e-cigarette and tobacco use disparities.However, little is known about how exposure to e-cigarette advertising is associated with product appeal among sexual minoritized  populations. This is a critical gap, as the US Food and Drug Administration (US-FDA) needs scientific evidence to regulate e-cigarette advertisements that may exacerbate disparities. Research on the appeal of specific e-cigarette advertisement features has mainly focused on the general population of youth and young adults. Such research has found that certain features, such as the presence of humans, flora imagery, and natural descriptors, are associated with increased appeal in this younger age categoryOur main research aim was to identify whether certain popular e-cigarette advertisement features were more appealing to sexual minoritized people. Within e-cigarette advertisements, key features might be more appealing than others in specific populations. Thus, the current study tested the effect of exposure to e-cigarette advertisements with key features on advertisement perceptions and product appeal among heterosexual and sexual minoritized women and men. We hypothesized that e-cigarette advertisements would be more appealing to sexual minoritized women and men, compared to heterosexual women and men.16. In this study, we used Prolific’s screening tools to determine the eligibility of participants, as Prolific only allows participants to opt-in to the study if they meet the specific requirements of the study. We aimed to recruit a sample of equal number heterosexual individuals (n=250) and sexual minoritized individuals (n=250) . Once participants met the eligibility criteria, they were able to opt-in to the study. Once quotas for each group were met, the study was not available to members of that specific group.We conducted an online experiment among 497 participants. Participants were enrolled through the online consumer research panel Prolific in October 2021. Participants were eligible to enroll in the study if they were aged ≥18 years and resided in the US. We oversampled sexual minoritized participants to have enough power to answer our research question17. Advertisements were selected from a previous content analysis of e-cigarette advertisement features17. Advertisements with features that appeared in at least 5% of the ads were chosen to further explore in this study15. This methodology was based on previous work done to examine cigarette advertising15. Multiple advertisements with the same feature were used in the study to account for one-message design error, which ensured that message responsiveness was attributed to the particular feature and not to any other aspect of a singular message 18. Advertisements came from media, including email, print magazines and mail, social media sites, and e-cigarette company websites17. Participants were allowed to view the advertisements for as long as they wanted. Participants then completed post-assessment measures, including perceptions of the advertisement and product use intentions. After completion, participants were compensated $3.96 according to Prolific’s policies.Consented participants began the online survey by reporting their e-cigarette, tobacco, and alcohol use. They then viewed two e-cigarette advertisements, randomly selected from a sample of 173 advertisements that were identified as including popular features , with an equivalent number of advertisements representative of each featureDemographics16.Participants reported their age, race/ethnicity , and individual income ($) (<50000 or ≥50000)Sexual orientation and gender identityFirst, participants were asked: ‘What is your gender?’ with possible answer options of ‘Female’, ‘Male’, ‘Trans female/Trans woman’, ‘Trans male/Trans man’, ‘Genderqueer/Gender non-conforming/Gender expansive’, and ‘Other’. For sexual orientation, participants were asked: ‘Which of the following best describes you?’ with possible answer options of ‘Straight or heterosexual’, ‘Lesbian or Gay’, ‘Bisexual’, and ‘Other non-heterosexual identity’.With respect to sexual orientation, ‘Lesbian and Gay’ (n=57), ‘Bisexual’ (n=142), and ‘Other non-heterosexual orientation’ (n=28) were combined and recoded as ‘Sexual minoritized’ to prevent small cell sizes at the intersection of sexual orientation and gender identity (SOGI) for our multivariable analyses. We then created a multi-categorical SOGI variable as follows: heterosexual man, heterosexual woman, sexual minoritized man, sexual minoritized woman. Our final analytic sample for multivariable analyses comprised 462 participants.Perceived advertisement effectiveness19. Scores on a 5-point Likert scale ranged from 1 (strongly disagree) to 5 (strongly agree). Scores were averaged across the five questions .Perceived advertisement effectiveness was assessed using a five-question instrument that asked whether participants thought the advertisement was memorable, attention grabbing, powerful, convincing, and meaningfulPerceived advertisement relevance20. Scores from a 5-point Likert scale ranged from 1 (strongly disagree) to 5 (strongly agree)16. Scores were averaged across the two questions 16.Perceived advertisement relevance was assessed using two questions: ‘The ad seemed to be written personally for me’, and ‘The ad was very relevant to my situation’Product use intention21.Product use intention was assessed using a single item: ‘This ad made me want to use the product’ on a 5-point Likert scale of 1 (strongly disagree) to 5 (strongly agree)E-cigarette advertisement features22. Complete information on the advertisements and procedures of the content analysis can be found elsewhere17. Features that appeared in at least 5% of the advertisements were tested in the current study15, including: the presence of non-tobacco flavor; the presence of a human(s); price reduction; product packaging; e-liquid bottle; actual product; product in use; ‘satisfying’ descriptor; claim that the product was an alternative to or exchangeable with a cigarette; fruit imagery ; flora imagery ; positive sensation appeal ; and camaraderie or friendship appeal (e.g. closeness with others)17.Advertisements were taken from four of the most popular e-cigarette brands  between 2019 and 202016. As the evidence indicates a marked heterogeneity in e-cigarette use among US adults by SOGI11, we assessed differences in outcomes at the intersection of these identities. We used linear mixed-effects models, fit with restricted maximum likelihood estimation, to assess interactions between the sexual orientation  and gender identity (man/woman) variables with each outcome variable. All models included random intercepts for each participant and fixed effects for age, race/ethnicity, and income. We conducted post-estimation pairwise comparisons, adjusting the alpha using the Tukey test. We used heterosexual men as our reference group, as research shows that the association between sexual identity and tobacco use is stronger for women11.Using descriptive statistics, we report distributions of all variables stratified by SOGI, including the outcome measures: perceived advertisement relevance, perceived advertisement effectiveness, and product use intentionTo assess the influence of advertisement features, we used linear mixed-effects models, fit with restricted maximum likelihood estimation, to estimate associations of the SOGI variable and advertisement feature with each outcome variable. All models included random intercepts for each participant and fixed effects for age, race/ethnicity, and income. Finally, we ran adjusted linear mixed-effects models that included product interaction terms between the SOGI variable and each feature (in separate models) and assessed their statistical significance using partial F-tests. Statistical significance of fixed effects, including the interaction terms, was assessed using a partial F-test with an alpha of 0.05 (two-tailed). In the models where the interaction between the SOGI variable and advertisement feature was statistically significant, we report stratified results and conduct post-estimation pairwise comparisons, adjusting the alpha using the Tukey test. We used R software [version 1.1.456] for all analyses.23. For multivariable analyses, TNB participants were excluded, and gender was recoded as ‘woman’ and ‘man’.With respect to gender, few respondents identified as non-binary (n=25) or transgender (n=10). While data from transgender and non-binary (TNB) participants was too limited for multivariable statistical analyses at the intersection of gender and sexual orientation, we included descriptive statistics , as thes16. Of the participants who identified as men in our study (n=238), 21% identified as sexual minoritized men, and 63.4% of those who identified as women in our study (n=224) identified as sexual minorized women.Our sample  includedPerceived advertisement effectiveness score  was loweAdvertisement relevance score  was loweWith respect to product appeal score  , heteroNo outcome differences were found, between heterosexual and sexual minoritized women, or sexual minoritized women and men.Perceived advertisement effectivenessSOGI modified the effect of flavors (p=0.004), humans present (p=0.045), and product packaging shown (p=0.045) on perceived advertisement effectiveness. Among heterosexual women, sexual minoritized men, and sexual minoritized women, perceived advertisement effectiveness was rated lower for advertisements with flavors compared to advertisements without flavors . Among sexual minoritized men and sexual minoritized women, perceived advertisement effectiveness was rated lower for advertisements with humans present compared to advertisements without humans . Among heterosexual women, sexual minoritized men, and sexual minoritized women, perceived advertisement effectiveness was rated lower for advertisements with product packaging shown compared to advertisements without product packaging .Perceived advertisement relevanceSOGI modified the effect of flavors (p=0.031), humans present (p=0.014), and fruit imagery (p=0.039) on perceived advertisement relevance. Among heterosexual women, sexual minoritized men, and sexual minoritized women, advertisements with flavors were rated lower in perceived advertisement relevance than those without that feature . Among heterosexual women, sexual minoritized men, and sexual minoritized women, perceived advertisement relevance was rated lower for advertisements with humans compared to those without humans . Among heterosexual men, heterosexual women, sexual minoritized men, and sexual minoritized women, advertisements with fruit imagery were rated as less relevant than advertisements without that feature .Product use intentionSOGI modified the effect of e-liquid bottle shown (p=0.046) and ‘Alternative to Cigarettes’ claim (p=0.026) on product use intention. Among heterosexual women, sexual minoritized men, and sexual minoritized women, product use intention was rated lower for advertisements with an e-liquid bottle shown compared to advertisements without that feature . Among sexual minoritized women, product use intention was rated lower for advertisements with an ‘Alternative to Cigarettes’ claim compared to advertisements without that feature .Contrary to our hypotheses, sexual minoritized women and men did not report greater perceived advertisement effectiveness or perceived advertisement relevance after viewing real-world e-cigarette advertisements, nor did they report higher intentions to use e-cigarettes. With respect to advertisement features, SOGI modified the effect of flavors and humans on both advertisement effectiveness and advertisement relevance. SOGI also modified the effect of featuring e-liquid bottles and ‘Alternative to Cigarettes’ claims in advertisements on product use intention. Overall, we found that sexual minoritized participants tended to find such features less appealing and had lower product use intentions after viewing these advertisements, compared to heterosexual men.25. Some evidence suggests that exposure to tobacco marketing increases the risk of cigarette smoking among sexual minoritized adults26. It is possible that, given the history of targeted marketing directed toward sexual minoritized communities and increased public skepticism toward ‘rainbow washing’27  that sexual minoritized adults in our study have developed a level of media literacy and accompanying skepticism toward tobacco industry marketing, including e-cigarette advertisements. Qualitative studies have described how sexual and gender minoritized participants have felt targeted by the tobacco industry and, thus, were skeptical towards pro- and anti-tobacco messages8. Survey data additionally suggest that advertising skepticism may be a protective factor among sexual minoritized adults for tobacco use28. Given these findings and our results, additional population-level studies are needed to understand why sexual minoritized adults use e-cigarettes more than other groups29, and if advertising skepticism is protective of e-cigarette use among sexual minoritized adults despite their increased exposure to targeted tobacco advertising.In the general population, exposure to e-cigarette advertisements is associated with subsequent e-cigarette use across all age groups (youth and adults)30, in which exposure to advertisements with flavors and people was positively associated with e-cigarette product appeal among mostly heterosexual young adults. Similar findings have also been shown for cigarette advertisements and product appeal with heterosexual adolescents and young adults15. The contrasting findings may have been due to the fact that participants in our study were only briefly exposed to two advertisements in an online study rather than having a longer exposure period in the real world. Given that these features are commonly used in e-cigarette advertising17, our findings suggest that regulating these features may reduce use in the general population while failing to attenuate e-cigarette use disparities.We found substantial heterogeneity in responses to e-cigarette advertisement features such that, unlike heterosexual men, sexual minoritized women and men found advertisements featuring flavors, fruit imagery, e-cigarette products, or humans, less relevant than advertisements that did not include those features. These findings contrast with findings by Chen-Sankey et al.31, and more men receive e-cigarette promotions by direct mail marketing32. Prior studies have identified gendered-appeals in online e-cigarette marketing targeting men, including the portrayal of e-cigarette use as more attractive to romantic/sexual partners and as a way to increase social status33. Consequently, our findings that message effectiveness and product appeal was highest among heterosexual men may reflect the use of marketing tactics that generally target men as the primary consumers of e-cigarettes.It is important to note that this study included advertisements from the most popular e-cigarette brands, which may not have been marketing to specific subgroups but aimed at the general population. In the US general population, uptake and use of e-cigarettes has been highest among menGiven the tobacco industry’s history of predatory marketing directed to sexual minoritized populations, it is possible that culturally specific advertisement features  may differently impact advertisement perceptions, product appeal, and use among this population. Future studies comparing the effect of culturally targeted e-cigarette advertisements versus general e-cigarette advertisements may illuminate advertisement features that unduly influence product appeal among sexual minoritized populations and, thus, may be targeted in regulatory efforts to reduce health inequities.16, but we could not assess these multiple intersections due to sample size. Descriptive statistics for these groups are included in Supplementary file Table 2. Additionally, the findings of this study may not represent those who do not use the recruitment platform Prolific, and may have an overrepresentation of White participants (71.2%), and a lack of information from Asian and Pacific Islander groups. Thus, our sample population may not be generalizable to a national level. Yet, studies have found Prolific data to be of high quality34 and have shown to mirror the demographics of the US national population35.Although our study explored a novel topic regarding the appeal of e-cigarette advertisements to sexual minoritized groups, it has the following limitations. We had to exclude non-binary and transgender participants from multivariable analyses due to small cell size; however, descriptives for this group are included in Supplementary file Table 2. Additionally, all gender minoritized participants in our sample also reported a minoritized sexual orientation, but sample size precluded descriptive statistics at the intersection of gender identity and sexual orientation. Given increased nicotine and tobacco use among gender minoritized populations, future research must recruit larger samples of non-binary and transgender people. There may be additional heterogeneity in responses to advertisement features according to age group or e-cigarette/tobacco use status within gender and sexual minoritized groups 36. We also used real world advertisements, which can present issues due to the inability to manipulate specific elements of the advertisement. We did not include any validity checks to ensure that participants noticed the features being tested and did not include an equivalent number of advertisements representative for each feature. Yet, utilizing real world advertisements provides ecological validity, and we showed each participant multiple advertisements to reduce message design error. In other words, the design of the study is such that message responsiveness, on average, is likely attributed to the feature of the advertisement and not to any other aspect of the message 18. We did not examine whether type of advertisement was associated with feature and study outcomes in our models.Our measure of product use intention was through self-report, not behavioral outcomes. However, research has shown that intentions are a strong predictor of actual future health behaviorsOur study identified substantial heterogeneity across sexual minoritized groups in appeal of various e-cigarette advertisement features. Although we found that sexual minoritized groups tended to rate certain features – including the presence of flavors, humans, and e-liquid bottles – as less appealing than do heterosexual men, this points to the need for additional research to identify which advertisement features sexual minoritized groups do find appealing, and which should be regulated to reduce disparities. Moreover, future studies should investigate longitudinal associations between exposure to advertising and subsequent e-cigarette use among sexual minoritized populations. Researchers, clinicians, and policymakers must continue to prioritize sexual minoritized groups when considering tobacco cessation, intervention work, and policy implementation, to help reduce tobacco use disparities.Click here for additional data file.


Low birth weight (LBW) is a significant public health concern given its association with early-life mortality and other adverse health consequences that can impact the entire life cycle. In many countries, accurate estimates of LBW prevalence are lacking due to inaccuracies in collection and gaps in available data. Our study aimed to determine LBW prevalence among facility-born infants in selected areas of Kenya and Tanzania and to assess whether the introduction of an intervention to improve the accuracy of birth weight measurement would result in a meaningfully different estimate of LBW prevalence than current practice.We carried out a historically controlled intervention study in 22 health facilities in Kenya and three health facilities in Tanzania. The intervention included: provision of high-quality digital scales, training of nursing staff on accurate birth weight measurement, recording and scale calibration practices, and quality maintenance support that consisted of enhanced supervision and feedback (prospective arm). The historically controlled data were birth weights from the same facilities recorded in maternity registers for the same calendar months from the previous year measured using routine practices and manual scales. We calculated mean birth weight , mean difference in LBW prevalence, and respective risk ratio (95% CI) between study arms.Between October 2019 and February 2020, we prospectively collected birth weights from 8441 newborns in Kenya and 4294 in Tanzania. Historical data were available from 9318 newborns in Kenya and 12,007 in Tanzania. In the prospective sample, the prevalence of LBW was 12.6%  in Kenya and 18.2% (12.2%–24.2%) in Tanzania. In the historical sample, the corresponding prevalence estimates were 7.8% (6.5%–9.2%) and 10.0% (8.6%–11.4%). Compared to the retrospective sample, the LBW prevalence in the prospective sample was 4.8% points (3.2%–6.4%) higher in Kenya and 8.2% points (2.3%–14.0%) higher in Tanzania, corresponding to a risk ratio of 1.61 (1.38–1.88) in Kenya and 1.81 (1.30–2.52) in Tanzania.Routine birth weight records underestimate the risk of LBW among facility-born infants in Kenya and Tanzania. The quality of birth weight data can be improved by a simple intervention consisting of provision of digital scales and supportive training. Low birth weight (LBW), defined by the World Health Organization (WHO) as a birth weight less than 2500 g , is a maGlobally, approximately 15% of live births are LBW, with the highest burden in southern Asia and Sub-Saharan Africa . HoweverWhat is the prevalence of LBW among infants born in health facilities in selected health areas in Kenya and Tanzania?Will the introduction of a support package  result in different LBW prevalence estimate compared to current practices in the same health facilities?Accurate birth weight measurement serves as a guide to appropriate care for newborns. Also measuring birth weight provides a low-cost, feasible method of monitoring newborn health which is globally applicable. It is also vitally important metric in monitoring neonatal outcomes at the population level and tracking national, regional, and global progress towards the Every Newborn Action Plan , the GloBy addressing these questions, we sought to contribute to the improved understanding of existing newborn weighing methods in low-resource settings and improve the accuracy of measurements with simple intervention.We conducted a prospective study with historical controls at 22 health facilities in Kenya and three health facilities in Tanzania to improve birth weight measurement and recording practices Fig. . We focuKenya and Tanzania are representative of LMICs where access to accurate and timely LBW data is a challenge , 21, 31.The study sites were selected to represent the whole range of delivery facilities in urban and rural settings capturing variation in numbers of births per year, infrastructure, average county-level birth rate, and staffing and supervision. We selected study facilities if their maternity registers contained complete birth information, including birth date, birth weight, and sex of the newborns for the year preceding the study period. In Kenya, the study was conducted in 22 rural health facilities in four counties . In Tanzania, the study was conducted in two referral hospitals (Amana and Temeke) and one health centre  in suburban Dar es Salaam Table . In KenyProvision of digital weighing scales with 10 g reading increment.Training of health workers in scale calibration and use and in precise birth weight recording.Monitoring and evaluation: Quality maintenance support via setting up a weekly supervision and feedback routine for senior nurses and facility managers on the intervention uptake and data quality.Scheduled mid-point retraining to reinforce the effectiveness of the intervention.The support package provided to the prospective group consisted of:The intervention was fitted into the existing newborn weighing and care routines of the selected health facilities as much as possible.Provision of scales Before the intervention, the facilities used traditional analogue weighing scales or hand-held scales. We provided each study facility a battery-operated digital scale  that measures in graduations of 10 g. Larger facilities with separate operating theatres were given two scales. We also provided standard weights  for calibration. In some facilities, official calibration stones of similar weight were already available. If these existing calibration stones were used, we ensured that they produced correct readings and were in good condition with no visible chippings, affecting the weight.Training Prior to data collection, we carried out a three-day training among staff members from labour, post-natal, and natal wards and obstetric theatres of the selected health facilities. For the larger facilities in Kenya, we trained health facility supervisors who then trained nurses, whereas in smaller facilities in Kenya and all facilities in Tanzania, we trained nurses. Participants were provided with a detailed outline of the study, highlighting the importance of accurate birth weight measurement. Nurses were trained to weigh naked newborns within one hour of birth, using the digital scale provided on a level hard surface and recording the result with 10 g precision. A standard operating procedure was provided to ensure uniformity of performance. We also trained the nurses to calibrate the digital scales and provided a daily calibration sheet in which the nurses were asked to record the status of calibration at the start of each working day. Scale readings within the limit of ± 20 g were considered acceptable, and any readings beyond the accepted range were reported. In addition, nurses were asked to record all birth data into the maternity register following the existing routine practices. We tested whether the amount of training was adequate at each health facility for a period of two days following the training. We randomly selected nurses from each facility who practised daily calibration of the scales, invited mothers for routine newborn weighing, and practised the improved birth weight measuring techniques using the digital scales.Monitoring and Evaluation As a method of monitoring and evaluation, we designed our own system to measure the efficacy of the intervention implementation and quality control. Health facility supervisors and district nurses were asked to reinforce the training of health workers and midwives through enhanced supervision and feedback on-site. They were also instructed to report to the research team on the quality of work using a daily calibration log and weekly supervision logs. Furthermore, they were asked to ensure with regularity the accuracy and completeness of the data by weekly reporting of training and supervision frequency received at study sites, numbers of correct scales calibrations completed, numbers of days when birth records were correctly completed, proportion of rounding of birth weight, and organizing mid-point interviews to obtain feedback on the process from health workers. In Kenya, depending on the site, health facility supervisors or district nurses visited the sites once a week supported and supervised by the central research team. In Tanzania, monitoring of quality maintenance was conducted by a member of the research team who visited the sites weekly.Retraining At the mid-point of the data collection, the central research team in each country visited the study locations to reinforce the effect of the support package by running a second training session at all sites. The training consisted of the same topics as the first session but was reduced to half a day. At smaller sites, the research team trained the nurses and midwives, whereas, at the larger sites, the research team trained the district nurses, i.e. the trainers of the nurses and midwives.We collected data from maternity records for the prospective and historically controlled groups. We obtained data on maternity ID, birth weight in grams, sex of the newborn , and date of birth . In Kenya, for prospective data, field supervisors collected data weekly and entered data into a tablet with CommCare data collection software . For historical data, the field supervisors photographed the historically controlled data and entered them into the CommCare application. In Tanzania, both historical and prospective data were photographed and entered into an Excel spreadsheet . In both countries, collected variables were the same, and the data were entered in the dataset by two people independently. The two datasets were linked using maternity ID and cross-checked by data managers in the research team. If any discrepancies were identified, the data were checked from the original health records.t-test . The outcome variable (birth weight) was grouped into LBW (< 2500 g) and normal birth weight (≥ 2500 g) in the categorical analysis. Although rounding and digit preference occur throughout the birth weight range, we focused our analyses at the 2500 g cut-off. This is an internationally recognized cut-off for LBW [We calculated mean birth weight and 95% confidence interval (CI). We also compared the mean birth weight between the prospective intervention group and the historical control groups using the Student’s independent  for LBW , 20, 23. for LBW .Between October 2019 and February 2020, we prospectively collected birth weights from 8441 newborns in Kenya and 4294 in Tanzania. Historical data were available from 9318 newborns in Kenya and 12,007 in Tanzania (Table The mean (SD) birth weight in Kenya was 3080 g (590) in the prospective sample and 3190 g (570) in the historical sample. In Tanzania, the respective values were 2990 g (590) and 3030 g (530). Compared to the historical sample, the mean birth weight in the prospective sample was 110 g (95% CI: 100–130) lower in Kenya and 40 g (95% CI: 20–60) lower in Tanzania.N = 8,441) was 12.6% (95% CI: 10.9%–14.4%) compared to 7.8% (6.5%–9.2%) in the historical data (N = 9318) (absolute difference 4.8% points (95% CI: 3.2%–6.4%), risk ratio 1.61 (1.38–1.88)). The corresponding figures in Tanzania were 18.2% (95% CI: 12.2%–24.2%) prospectively (N = 4294) and 10.0% (95% CI: 8.6%–11.4%) retrospectively  (absolute difference 8.2% points (95% CI: 2.3%–14.0%), risk ratio 1.81 (1.30–2.52)). A sensitivity analysis using adjusted birth weights by reallocating 25% of infants with an exact birth weight of 2,500 g to the LBW category gave essentially similar results in both countries  in Kenya and 8.2 percentage points (from 10.0 to 18.2%) in Tanzania compared to historical sample produced using routine practices in a sample of approximately 34,000 facility births.The validity of the findings could theoretically have been affected by variation in birth weight measurement in the multiple study facilities, but we minimized this problem by enhanced supervision, communication and clear accountability structures. Moreover, the study facilities might be considered “high performers” given that they contained complete birth information from the previous year. The differences between prospective and historical data might have been greater in facilities with incomplete birth information. In Tanzania, the number of births was lower in the prospective sample than the historical one because a new birthing facility was opened in the study area during the period of prospective data collection. This might theoretically have led to the referral of low-risk mothers to this new facility and an increase in the proportion of high-risk deliveries in the study facilities and thus potentially have contributed to an increase in the proportion of LBW infants in our prospective sample. There was, however, no public recommendation to concentrate or refer certain types of deliveries to specific facilities. Because of this and the consistency of the findings, we believe that our findings are valid and indicate that routine birth weight recording has produced considerable underestimates in the prevalence of LBW.Observer errors have been reported in birth weight research . WeightsThe 12.4% LBW prevalence in Kenya and 18.2% in Tanzania are higher than the UNICEF reported national prevalence rates from 2015  was significantly higher compared to the analogue device (23.0%) . Data quThe LBW prevalences reported in the current study are not generalizable beyond the study areas and cannot be taken as indicators of national LBW prevalence in Kenya and Tanzania. This is because data collection took place only at health facilities and included no data from home deliveries that remain common in both countries. Furthermore, the study samples were not designed to be nationally representative, as they represented only limited numbers of geographic areas and only some types of maternity wards. However, while these issues obviously limit the generalizability of the reported LBW prevalences, they do not affect the main conclusions of the study.There has been a call for research to establish the efficacy and feasibility of interventions to improve the quality of birth weight data . Our stu


This study investigates whether preterm birth, small for gestational age, or large for gestational age are associated with increased risks of atrial fibrillation later in life. Are preterm birth, small for gestational age, and large for gestational age associated with increased risks of atrial fibrillation (AF) up to age 49 years?In this multinational cohort study with 8 million participants, preterm birth and large for gestational age were associated with increased risks of AF in childhood and up to age 49 years in adulthood, while an association between small for gestational age and an increased risk of AF was observed only in childhood. Similar findings were observed in sibling analyses.Preterm birth, excessive fetal growth, and reduced fetal growth may increase the risk of AF up to age 49 years. Adverse birth outcomes, including preterm birth, small for gestational age (SGA), and large for gestational age (LGA) are associated with increased risks of hypertension, ischemic heart disease, stroke, and heart failure, but knowledge regarding their associations with atrial fibrillation (AF) is limited and inconsistent.To investigate whether preterm birth, SGA, or LGA are associated with increased risks of AF later in life.This multinational cohort study included Danish, Swedish, and Finnish national health registries. Live singleton births in Denmark from 1978 through 2016, in Sweden from 1973 through 2014, and in Finland from 1987 through 2014, who were followed up until December 31, 2016, in Denmark, December 31, 2021, in Sweden, and December 31, 2014, in Finland were included. Data analyses were performed between January 2021 and August 2022.Preterm birth , SGA , and LGA  identified from medical birth registers.Diagnosis of AF obtained from nationwide inpatient and outpatient registers. The study team ran multivariable Cox proportional hazard models and flexible parametric survival models to estimate hazard ratios (HRs) and 95% CIs for AF according to preterm birth, SGA, and LGA. Sibling analyses were conducted to control for unmeasured familial factors.The cohort included 8 012 433 study participants . In 174.4 million person-years of follow-up, 11 464 participants had a diagnosis of AF . Preterm birth and LGA were associated with increased AF risk in both the full population cohort and in the sibling analyses; the multivariate HRs from the cohort analyses were 1.30  and 1.55 , respectively. Preterm birth was more strongly associated with AF in childhood than in adulthood. Children born SGA had an increased risk of AF in the first 18 years of life but not afterwards.Preterm births and LGA births were associated with increased risks of AF up to middle age independently of familial confounding factors. Individuals born SGA had an increased AF risk only during childhood. The incidence of AF in children and young adults has increased slightly during the past decades and its estimated prevalence is 0.12% to 0.16%.6 Having AF in young age may entail a higher socioeconomic burden than in old age because of the lifelong work productivity loss and the increased health care costs. The well-established risk factors for AF, such as age, male sex, genetic factors, smoking, obesity, hypertension, diabetes, and specific heart diseases,7 do not explain a substantial proportion of AF cases in children and young adults.9 Knowledge on the etiology of AF in children and young adults is lacking, prompting the need for further studies in this area.Atrial fibrillation (AF), the most common cardiac arrhythmia of clinical significance, is a growing global epidemic.10 ischemic heart disease (IHD),11 stroke,12 heart failure,13 and other cardiometabolic disorders in childhood and adulthood.15 Short gestational age, as reflected in preterm birth, was also found to be associated with increased risks of IHD, stroke, and heart failure, irrespective of fetal growth.19 Knowledge regarding associations between fetal growth and risks of AF is limited and inconsistent25; some studies reported positive, others inverse, U-shaped, or no associations between birth weight and AF.25 The 2 studies regarding preterm birth and AF found no association.23 All of the earlier studies were conducted in populations that were middle aged or older at baseline. Thus, several of these studies were prone to recall bias, survival bias, or did not consider important confounders, including familial risk factors. None of them examined both the individual and the joint effects of fetal growth and gestational age or the role of these exposures in pediatric and adult AF.Restricted and excessive fetal growth, as reflected in small and large for gestational age , have been linked to increased risks of cardiovascular diseases (CVD), including hypertension,26 we also examined their joint effects. To consider confounding by shared familial factors, we performed analyses with a sibling design.In this population-based study using prospectively recorded nationwide data on more than 8 million individuals from 3 Nordic countries who were followed up to age 49 years, we investigated whether preterm birth, SGA, or LGA are associated with AF risk in childhood and adulthood. Given the strong association between preterm birth and poor fetal growth,STROBE) reporting guidelines.We conducted a cohort study by linking several nationwide registers in Denmark, Sweden, and Finland through the unique personal identification number assigned to each resident. A detailed description of the registers included and the main variables retrieved is provided in eAppendix 1 in Information on gestational age and birth weight was retrieved from the medical birth registers. Estimation of gestational age was based primarily on ultrasound examinations performed in the early second trimester or otherwise on the date of the last menstrual period. Preterm birth was defined as birth before 37 completed weeks of gestation and was categorized as extremely and very preterm (22 to 31 weeks) and moderately preterm (32 to 36 weeks).27 and categorized it as SGA (less than 10th percentile of each country’s distribution), LGA (more than 90th percentile), and appropriate for gestational age ([AGA] 10th to 90th percentile). The SGA and LGA were further divided into severe SGA (less than 3rd percentile), moderate SGA (3rd to less than 10th percentile), moderate LGA (more than 90th to 97th percentile), and severe LGA (more than 97th percentile).We calculated birth weight for gestational age according to a Scandinavian sex-specific reference curve for normal fetal growth28 We identified individuals with a primary or secondary diagnosis of AF from the Danish National Patient Register, the Swedish Patient Register, and the Finnish Hospital Discharge Register using the International Classification of Diseases codes shown in eTable 1 in AF was defined as AF or atrial flutter, given their close interrelationships.We obtained information on children’s characteristics , and maternal characteristics , as described in eAppendix 2 in We first investigated the associations between gestational age and birth weight for gestational age as continuous variables and AF risk. We applied restricted cubic spline functions with 5 knots, located at the 5th, 25th, 50th, 75th, and 95th percentiles of the distribution of each variable. Given the nonlinearity of the associations , we furt29 to visualize the time-varying effects. In our main models, we adjusted for the study participants’ year of birth and country of birth and for maternal characteristics, ie, parity, age at delivery, education, marital status, hypertensive disorders, and diabetes before delivery. Analyses of preterm birth were also adjusted for sex and birth weight. To control for unmeasured familial confounders, we conducted sibling analyses. We ran stratified Cox models with a separate stratum for each family; only sibling pairs discordant for exposure and outcome contributed to the estimates. We also analyzed the joint effects of gestational age and birth weight for gestational age on AF risk.We estimated hazard ratios (HRs) and 95% CIs for the associations between preterm birth, SGA, and LGA and the risk of AF by Cox proportional hazard regression using attained age as the underlying time scale. The log-minus log-survival curves and the Schoenfeld residuals suggested that the proportional hazard assumption does not hold for our exposures. To account for the nonproportional effects, we performed analyses with the follow-up time split at 18 years. We also used flexible parametric survival modelsTo allow comparisons with earlier studies, we examined AF risk according to birth weight categorized as low (less than 2500 g), medium (2500 to 3999 g), or high (4000 g or more). To test whether the association between preterm birth or abnormal fetal growth and AF differed by the participants’ country at birth or sex, we conducted stratified analyses and formal tests of interaction with these variables. In sensitivity analyses we: (1) changed the start of follow-up to the age of 1 year to exclude the possibility that AF may be an immediate complication of treatments during neonatal care; (2) performed analyses among participants without congenital anomalies; (3) adjusted for maternal smoking or BMI in early pregnancy, maternal country of origin, or maternal family history of CVD in addition to covariates in our main model, restricting to individuals with data on these variables; (4) adjusted one by one for participants’ BMI, hypertension, diabetes, IHD, stroke, and heart failure, in addition to covariates in our main model to examine whether these factors could contribute to the observed associations; and (5) repeated analyses in the Danish subcohort after excluding individuals who underwent cardiac surgery during the follow-up period to consider that postoperative AF has a different origin than other AFs. Statistical analyses were performed using SAS version 9.4 (SAS Institute) and Stata version 15.1 (StataCorp).A total of 378 917 of our study participants were born preterm (4.7%), 800 959 SGA (10.0%) and 802 759 LGA (10.0%). The maximum age at the end of follow-up was 49 years . During the 174.4 million person-years follow-up, 11 464 participants had AF (0.14%); the median age at diagnosis was 29.3 years . Characteristics of the cohort according to exposures are presented in eTable 2 in The study team observed an inverse, largely linear association between gestational age and AF risk . PretermThe risk of AF increased substantially by birth weight for gestational age from the 50th percentile . CompareBeing SGA was associated with an increased risk of AF during the first 18 years of follow-up  but with a reduced risk later ; the association between LGA and AF risk was similar between the 2 periods. The flexible parametric models showed that the strength of the associations with SGA or LGA declined with increasing age at follow-up . Both loCompared with term AGA births, the risk of AF was increased by 71% in preterm LGA births, 55% in term LGA births, 31% in preterm AGA births, and 25% in preterm SGA births. The risk of AF was 9% lower for term SGA than term AGA births . In thesThe associations of preterm birth, SGA, and LGA with AF risk did not differ by the child’s sex or country of birth (eTable 4 and eFigure 3 in In this large population-based study, we found that preterm birth and LGA were associated with increased risks of AF up to the age of 49 years. The highest AF risk was observed in study participants who were both preterm and LGA. These associations persisted from childhood into early middle age and were also generally observed in the sibling analyses. Individuals born SGA had an increased risk of AF in childhood, both when compared with the general population and with their siblings.22 and the Helsinki Birth Cohort Study23 found no association between preterm birth and AF risk. A possible explanation for the differences in results may be related to survival bias, as the participants included in the 2 previous studies were born between 1914 and 1952, ie, when no advanced neonatal care was available. Thus, individuals who survived after a preterm birth in the early 20th century were likely to be healthier than the survivors of preterm birth from our study. The 2 earlier studies reported U-shaped associations between birth weight and the risk of AF after adjusting for gestational age, but the associations were generally weak. Similar to this study, the Women’s Health Study21 also found a link between high birth weight and an increased AF risk, but the inclusion of only healthy and well-educated women limited the generalizability of their findings. In contrast, the Atherosclerosis Risk in Communities Study20 found that low birth weight was associated with an increased risk of AF. These studies were limited by low statistical power, self-reported birth weight, and lack of data on several important confounders.Evidence regarding the link between gestational age or fetal growth and the risk of AF has been limited and inconsistent. In contrast to this study, a Swedish cohort study30 conditions that may be related to both preterm birth and restricted fetal growth.32 Nevertheless, when we excluded study participants with congenital anomaly, we observed similar associations to those in the main analyses. Second, an increased risk of AF in childhood could be a direct complication of or may be triggered by neonatal treatments, more commonly occurring in preterm or SGA births. To address this concern, we performed sensitivity analyses in which follow-up started at 1 year of age, but the results of these analyses were similar to those of our primary analyses.To our knowledge, this study is the first to investigate the association between birth outcomes and both pediatric and adult AF. Preterm birth, SGA, and LGA were associated with increased risks of pediatric AF and the associations for preterm birth and SGA had larger effect sizes than the corresponding associations with AF in young adulthood. We can only speculate about explanations for these findings. First, the etiology of pediatric AF and adult AF has been suggested to differ; children with AF are more likely to have congenital heart diseases,34 lifestyle,38 hypertensive disorders,40 and diabetes,42 or other confounders. Nevertheless, adjustment for several maternal characteristics did not change the associations considerably. The results of our sibling analyses suggest that the contributions of shared genetic and environmental factors to the associations of preterm birth and LGA with the risk of AF are likely to be modest but might be important in the case of the association between SGA and AF. Second, preterm birth, SGA, and LGA may cause cardiac remodeling and dysfunction during fetal life and/or persistent impairment of atrial structure or function during early childhood and adolescence,44 which in turn may lead to AF. Third, preterm birth, SGA, and LGA birth may lead to AF through adverse changes in cardiorespiratory fitness,45 blood pressure,48 insulin sensitivity,50 adiposity and lipid deposition,51 and other cardiometabolic disorders later in life.15 Fourth, AF occurring during cardiac surgery, potentially more common in participants with adverse birth outcomes, may also contribute to the observed associations. Nevertheless, our point estimates did not substantially change after we controlled for high BMI, diabetes, hypertension, IHD, stroke, heart failure, or cardiac surgery, suggesting that other unmeasured factors may explain the investigated associations. A potential explanation for the finding that the risk of AF was highest for the combination of preterm birth and LGA may be that maternal medical conditions, such as diabetes and/or obesity during pregnancy lead to preterm birth,52 and together induce lasting physiological alterations, such as insulin resistance, vascular endothelial dysfunction, and cardiac damage, that lead to AF.53There are several potential explanations for the link of preterm birth and abnormal fetal growth with the risk of AF. First, the associations may in part be due to shared genetic background or to maternal socioeconomic status,The large sample size of this study allowed us to evaluate more narrowly defined gestational age and birth weight for gestational age groups, to study their associations with both pediatric and adult AF, and to perform sibling analyses to consider genetic or environmental confounders shared by siblings. The prospectively collected data from registers effectively eliminated the possibility for recall or selection bias.55 we may have missed some asymptomatic, paroxysmal, or mild cases of AF. Second, we cannot rule out the possibility of residual confounding. Our sibling analyses could partly control for unmeasured familial risk factors, but this design itself has several limitations like the bias caused by the carry over effects56 or nonshared confounders.57 Third, because of the limited follow-up time, our findings apply only to children and young adults and cannot be generalized to the predominant group of individuals developing AF at an older age. Additionally, our results may only be generalized to countries with predominantly White populations and universal free health care systems.Several limitations of our study should be noted. First, although the diagnoses of AF in the Nordic patient registers has a high positive predictive value,58 the possible long-term health effects of being born LGA may become increasingly important.In this study, preterm birth and LGA birth were associated with increased AF risks up to the age of 49 years. Individuals born SGA have an increased AF risk only in childhood. Further studies with longer follow-up and that may elucidate the underlying mechanisms for the observed associations are warranted. As the prevalence of LGA births is reported to increase over time,


After adjustment for genetic factors, is birth weight associated with neurodevelopmental conditions?In this case-control study of 393 twins in Sweden, the twin with a lower birth weight in monozygotic twin pairs, but not dizygotic pairs, had more autism and attention-deficit/hyperactivity disorder (ADHD) symptoms, lower IQ ratings, and higher odds of having a diagnosis of autism and ADHD compared with their co-twin.These findings suggest that birth weight contributes to neurodevelopmental conditions when adjusting for genetic factors. This case-control study estimates the associations between birth weight and neurodevelopmental conditions, such as autism, attention-deficit/hyperactivity disorder, and intellectual disability, among monozygotic and dizygotic twins. Low birth weight is associated with an increased likelihood of neurodivergence and neurodevelopmental conditions (NDCs) such as autism, attention-deficit/hyperactivity disorder (ADHD), and intellectual disability. However, it is unclear whether birth weight contributes independently to NDCs or whether the association is predominantly driven by genetic predisposition.To estimate the associations between birth weight and dimensional (trait) and categorical (diagnoses) NDC outcomes, while adjusting for genetic risks.A co-twin design was applied to this case-control study conducted in Sweden. Diagnostic assessments were conducted between August 2011 and March 2022, within the Roots of Autism and ADHD Twin Study in Sweden (RATSS) during a 2.5-day participant visit to the clinic. The RATSS sample comprised phenotyped monozygotic and dizygotic twins enriched for NDCs. Data analysis was conducted in November 2022.Birth weight.Categorical and dimensional operationalizations of autism, ADHD, and intellectual disability were assessed. Generalized estimating equation models were fitted across and within twin pairs.The study sample included 393 twins: 230 were monozygotic and 159 were dizygotic (zygosity was unknown for 4). Their median age was 15  years. There were 185 female participants (47.1%) and 208 male participants (52.9%). Across twin pairs, higher birth weight was associated with fewer autistic traits  and lower odds of autism diagnosis  and intellectual disability . Within pairs, the association between birth weight and dimensional autism  and categorical autism  remained among monozygotic pairs but not dizygotic pairs. In addition, higher birth weight was associated with lower odds of ADHD diagnosis , fewer ADHD traits , and higher IQ ratings  among monozygotic twins.The findings of this co-twin study suggest an association between low birth weight and NDCs, but they also acknowledge the importance of genetics because the associations observed were only statically significant among monozygotic twins. It is of pivotal importance to facilitate early identification of factors contributing to fetal growth restriction to minimize detrimental outcomes. Neurodevelopmental conditions are heterogenous in causation, neurobiology, and behavioral phenotypes; they tend to co-occur and increase susceptibility for mental and somatic health issues.3Neurodevelopmental conditions (NDCs) are present in approximately 18% of the general US population.4 Nonshared environmental factors are exposures that are unique to members within the same family and make them differ from each other. Low birth weight is a specific NSE factor that has been linked to autism,5 ADHD,6 and low IQ7 and may be associated with long-lasting outcomes in child development and health.8 However, the origins of the association between low birth weight and NDCs remain difficult to establish, owing to potential confounding factors. It is unclear whether low birth weight independently contributes to NDCs or if its association with NDCs can be better explained by factors such as genetic susceptibility to both low birth weight and NDCs. By applying a twin co-control design using dizygotic and monozygotic twins, it is possible to approach this research question because dizygotic twins share approximately 50% of their genome , whereas monozygotic twins share 100% of their genome. If the association between birth weight and NDCs remains among twin pairs, then NSE factors specific to each fetus should be directly involved in the underlying pathway of NDCs.Although NDCs have a strong genetic component, nonshared environmental (NSE) factors also play a substantial role in their etiology.4 the number of twin studies exploring the issue has been limited. Willfors et al9 reported a trend in which lower birth weight was associated with autism and autistic traits within twin pairs. Losh et al10 found that among twin pairs discordant for autism, the lower-weighted twin had higher odds of being categorized as having autism; however, a limitation was that autism diagnosis was based solely on a parent questionnaire and not on a full-scale diagnostic assessment. Interestingly, the odds of being categorized as autistic for the lower-weighted twin were similar for monozygotic and dizygotic twin pairs,10 which suggests a direct contribution of NSE factors to autism and not a genetic contribution to the association. For ADHD, Lehn et al11 conducted a twin study operationalizing the condition based on diagnostic interviews and ratings. They found that within monozygotic twin pairs, individuals with ADHD had lower birth weight compared with their co-twin without ADHD. This finding suggests a direct contribution of NSE factors. Studies using dimensional operationalizations of ADHD have produced mixed findings, with both reported associations15 and no associations16 in population-based cohorts. Reported within-pair differences remained in some studies for both monozygotic and dizygotic subsets, suggesting a direct contribution of NSE factors14; in another study, they only presented among monozygotic twins, although with similar estimates.13 Additionally, previous studies have reported no associations between lower birth weight and ID,17 while associations have been reported for lower IQ.19Although previous research provides some support for a direct contribution of low birth weight to NDCs,20 However, that study only included monozygotic twins and was limited to few discordant pairs and a categorical measure of birth weight discordance. There are also few studies exploring differences in the association between birth weight and dimensional and categorical NDCs stratified by zygosity. Birth weight discordance within monozygotic twins is more likely to reflect differences in placental function or blood and nutrient supply to the individual fetus than shared genetic or environmental factors. Unequal placental sharing is regarded as a major cause of fetal growth discordance in monozygotic twins, and, in extreme cases, an unbalanced intertwin blood transfusion may lead to twin-to-twin transfusion syndrome (TTTS), in which 1 twin gives away more blood than they receive, with increased risk of malnourishment and a wide range of detrimental outcomes.21A recent study found that in pairs with birth weight discordance of 18% or greater, the lower-weighted twin had a higher likelihood of presenting with traits of autism and ADHD than their co-twin.With the objective to expand knowledge within the field, we applied a co-twin control design within a sample of deeply phenotyped monozygotic and dizygotic twins enriched for NDCs. To enable estimation of the associations between birth weight discordance and neurodivergence for both clinical and typical phenotypes, dimensional (trait) and categorical (diagnoses) outcomes were assessed. We hypothesized that (1) birth weight difference is associated with clinical and trait-level NDCs and (2) the association between birth weight differences and NDCs would remain within twin pairs even after adjusting for familial confounders, indicating a direct association between low birth weight and neurodivergent phenotypes.22 A subsample of monozygotic twins was included in a previous publication.20 This study adds to the previous research (1) by including a larger sample of twins and (2) by comparing the link between low birth weight and NDCs in both monozygotic and dizygotic twins. Zygosity was determined with a panel of 48 single-nucleotide polymorphisms.23 In a few cases in which DNA results had not yet been analyzed (n = 15 pairs), a 4-item zygosity questionnaire based on physical appearance was used instead. We lacked data on zygosity for 4 participants. This study was approved by the National Swedish Ethical Review Board. All participants and/or their legal guardians provided written and oral informed consent. The study conformed to the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guideline.Twins were assessed between August 2011 and March 2022 within the Roots of Autism and ADHD Twin Study in Sweden (RATSS). All Swedish twin pairs are eligible to participate in the RATSS if 1 twin or both twins obtain a positive screening result for NDCs or typical development.Diagnostic and Statistical Manual of Mental Disorders (Fifth Edition) criteria during a 2.5-day participant visit to the clinic, using diagnostic interviews, medical history records, and standardized diagnostic tools. Diagnostic tools included the Autism Diagnostic Observation Schedule–2, the Autism Diagnostic Interview–Revised, the Diagnostic Interview for ADHD in Adults, the Structured Clinical Interview for DSM-IV, and the Adaptive Behavior Assessment System–2. A group of clinicians performed the diagnostic assessment according to Autistic traits were assessed with parent ratings on the Social Responsiveness Scale–2 (SRS-2). The SRS-2 targets interpersonal behavior, communication, and restricted, inflexible, and repetitive interests and behaviors. The scale includes 65 items and is rated on a 4-point Likert-type scale , with a total score ranging between 0 and 195. A higher SRS-2 score indicates more autistic traits.Attention-deficit/hyperactivity disorder traits were assessed with the parent-rated attention subscale from the Child Behavior Checklist (CBCL) and the Adult Behavior Checklist (ABCL), depending on participant age. Items in the CBCL and ABCL are rated on a 3-point Likert-type scale . The attention subscale includes 10 items in the CBCL and 17 items in the ABCL. In this study, CBCL and ABCL attention total scores were calculated as mean values, with a higher mean indicating more ADHD symptoms.Intelligence quotient ratings were assessed with the General Ability Index (GAI) of the Wechsler Intelligence Scale for Children–IV or the Wechsler Adult Intelligence Scale for Adults–IV, depending on participant age. The GAI is a full score measure that provides an estimate of intellectual functioning less influenced by processing speed and working memory, which are often compromised by NDCs. The GAI provides a composite score based on 3 verbal comprehension subtests and 3 perceptual reasoning subtests.9 There was high agreement between birth weight information and medical record data .The RATSS parent-report questionnaire was designed to cover prenatal, perinatal, and postnatal factors and include information about a twin’s birth weight. The questionnaire has been validated against medical registry data and was completed during the participant visit to the clinical research unit.25 First, we examined the association between the independent variable, birth weight , and the dependent variables, categorical neurodivergence  and dimensional neurodivergence , across individuals, while also adjusting for age and sex. In this model, included twins were treated as singletons, not adjusting for familial confounders. Second, we examined within-pair associations between birth weight and categorical or dimensional assessments of NDCs. In this co-twin design, the difference in birth weight within a pair was correlated with the difference in NDCs within the same pair, and we adjusted for all factors shared between twins (genes and shared environment). Third, we repeated the within-pair analyses among the dizygotic and monozygotic twin pairs, respectively. For the dimensional outcome , we used the identity link within the drgee function in R and reported unstandardized regression estimates. For binary outcomes , we used odds ratios (ORs) with the logit link and present the log ORs in back-transformed format. Information on the generalized estimating equation framework and the twin statistics is presented in the eMethods and the eFigure in P < .05 were considered significant. Data analysis was conducted in November 2022.All associations between birth weight and NDCs were examined using conditional regression analyses within the generalized estimating equation framework implemented in the drgee package of R . These analyses did not make any distributional assumptions , were appropriate for continuous and binary outcomes, and included calculations of doubly robust SEs.This study included 393 participants, with a median age of 15  years. There were 230 monozygotic same-sex twins and 159 dizygotic twins (1 triplet); zygosity was unknown for 4 participants. There were 185 female participants (47.1%) and 208 male participants (52.9%). A total of 110 participants (28.0%) had been diagnosed with ADHD, 92 (23.4%) with autism, 20 (5.1%) with ID, 66 (16.8%) with other NDCs , and 102 (26.0%) with other psychiatric disorders (including 46 with depressive disorders and 22 with anxiety disorders). Numbers of pairs concordant and discordant for NDCs, stratified by zygosity, are presented in P = .009), ADHD traits , and IQ .P = .02), although the association with ADHD diagnosis was lost .In this study of phenotyped monozygotic and dizygotic twins enriched for NDCs, we observed that lower birth weight was significantly associated with NDCs. A lower birth weight was foremost associated with more autistic traits as well as an autism clinical diagnosis across individuals. Within the monozygotic pairs, but not the dizygotic pairs, the twin with a higher birth weight had fewer symptoms of autism and ADHD, higher IQ ratings, and lower odds of having a diagnosis of autism and ADHD. These results emphasize birth weight as an NSE factor for NDCs, since the association was most evident when implicitly adjusting for all unmeasured familial confounding .5 ADHD,6 and low IQ.7 Partly in line with these findings on singletons, our results across individuals suggested that lower birth weight was associated with autism and ID, but not with a diagnosis of ADHD or traits of ADHD. This finding regarding ADHD contradicts other studies in which low birth weight was associated with ADHD.26 Yet as indicated in the review by Sciberras et al,6 the causal effect of factors such as birth weight on ADHD cannot be confirmed using observational designs compared with twin designs.Previous research has reported low birth weight being associated with a wide range of neurodivergent outcomes such as NDCs, including symptoms of autism,27 In our within-pair analyses, the associations observed were driven by the monozygotic pairs, indicating an NSE factor and genetic contribution to the associations. More specifically, the twin with a lower birth weight had more trait-level and categorical autism and ADHD as well as lower IQ ratings. None of these associations was present among the dizygotic twins. If there were an equal intrapair correlation in monozygotic and dizygotic pairs, the results would suggest NSE factors and no genetic contribution to the association.9 Here, the association was only present among monozygotic twins—a reason why the genetic contribution could mask NSE factors in the dizygotic pairs—resulting in a greater association in monozygotic pairs when adjusting for genetic confounding. Previous studies exploring differences in the association between birth weight and dimensional and diagnostic NDCs stratified by zygosity have been few and were mostly conducted with population-based nonclinical phenotyped samples, with mixed findings.14 Our findings foremost stress the importance of birth weight as a contributing factor to dimensional and categorical autism and ADHD as well as IQ, but they also acknowledge the importance of genetics for the association between birth weight and NDCs.The discordant co-twin design offers opportunities to investigate possible causal pathways to NDCs by focusing on the differences within twin pairs with respect to unique environmental experiences, thereby automatically controlling for both genetic factors and shared environment.28 Monozygotic twins often share a single placenta and are connected with each other through vascular anastomoses. Birth weight discordance within monozygotic twins may reflect uneven placental sharing, as in selective fetal growth restriction, and a more unbalanced intertwin blood transfusion  or slow intertwin blood transfusion .21 Although TTTS may result in a more extreme form of birth weight discordance, our results remained unaltered when pairs with reported TTTS were excluded, indicating that TTTS did not explain the result and that low birth weight is an important factor for NDCs per se.Monozygotic twins, compared with dizygotic twins, have a higher risk for adverse birth outcomes, including low birth weight and shorter gestations.8 However, as stated by O’Donnell and Meaney,8 previous studies failed to account for underlying genetic factors and for the transition of DOHaD from observational studies to those of biological mechanisms, and an integration of genetic information into developmental models is required. In this study, we expanded previous research by providing support for the DOHaD hypothesis within a co-twin control design, adjusting for unmeasured familial confounders. The applied discordant twin design is a powerful method for investigating the contribution of NSE factors to NDCs. Other strengths of this study include its relatively large and rare sample of well-phenotyped twins, concordant and discordant for NDCs.According to the developmental origins of health and disease (DOHaD) hypothesis, intrauterine processes alter fetal growth program tissue differentiation in a manner that predisposes to different outcomes, including NDCs.9 Other complications during pregnancy, such as preeclampsia, anemia, diabetes, infection, and defective implantation of the placental vessels, may affect birth weight; these were not controlled for, as they are likely to affect both twins comparably, thus not introducing bias. Twin fetuses have a substantially higher risk of low birth weight than nontwins, and a twin pregnancy is also associated with more complications than singleton pregnancies; thus, concerns have been raised that findings in twin samples may not be generalizable to nontwin samples. However, previous research suggests that twins do not systematically differ from the general population of nontwins on measures of behavior and development.30 Comorbidity was common in our sample, which may have affected the reported associations but also increased the generalizability of our findings, as comorbidity is common in NDCs.3 Our sample was biased toward monozygotic twins discordant for NDCs. Although this made it inappropriate to use bivariate twin models such as ACE , it was appropriate for investigating the association between NSE factors and outcomes while controlling for familial confounders. Although a co-twin design implies a high degree of genetic and shared environment control, there may be post-twining de novo alterations that may contribute to discordance.This study has some limitations. All data about birth weight were based on retrospective parental questionnaires, which may be affected by recall bias. However, a subsample of the data has been validated against medical records data, with excellent agreement.The findings of this case-control study suggest that as an indicator for fetal growth alterations, lower birth weight is an NSE factor for neurodivergent outcomes, including both trait and categorical autism, ADHD, and ID. Since this association was only observed among the monozygotic twin pairs, a genetic confounding of the association, besides the nonshared factor, was also evident. Given this link between low birth weight and future NDCs, it is important to acknowledge birth weight as influential when assessing NDCs. Furthermore, it is of pivotal importance to facilitate early identification of factors contributing to fetal growth restriction, such as uneven placental sharing, unbalanced intertwin blood transfusion, or slow intertwin blood transfusion, and offer monitoring and treatments  to minimize detrimental outcomes. Future studies are needed to obtain deeper knowledge about the biological mechanisms that underly the associations between low birth weight and NDCs.


Visceral obesity is directly linked to increased cardiovascular risk, including heart failure.This study explored the ability of human epicardial adipose tissue (EAT)-derived microRNAs (miRNAs) to regulate the myocardial redox state and clinical outcomes.This study screened for miRNAs expressed and released from human EAT and tested for correlations with the redox state in the adjacent myocardium in paired EAT/atrial biopsy specimens from patients undergoing cardiac surgery. Three miRNAs were then tested for causality in an in vitro model of cardiomyocytes. At a clinical level, causality/directionality were tested using genome-wide association screening, and the underlying mechanisms were explored using human biopsy specimens, as well as overexpression of the candidate miRNAs and their targets in vitro and in vivo using a transgenic mouse model. The final prognostic value of the discovered targets was tested in patients undergoing cardiac surgery, followed up for a median of 8 years.2.–) by targeting myocardial expression of WNT5A, which regulated Rac1-dependent activation of NADPH oxidases. Finally, high miR-92a-3p levels in EAT were independently related with lower risk of adverse cardiovascular events.EAT miR-92a-3p was related to lower oxidative stress in human myocardium, a finding confirmed by using genetic regulators of miR-92a-3p in the human heart and EAT. miR-92a-3p reduced nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase–derived superoxide (OEAT-derived miRNAs exert paracrine effects on the human heart. Indeed miR-92a-3p suppresses the wingless-type MMTV integration site family, member 5a/Rac1/NADPH oxidase axis and improves the myocardial redox state. EAT-derived miR-92a-3p is related to improved clinical outcomes and is a rational therapeutic target for the prevention and treatment of obesity-related heart disease.  The mechanisms by which EAT affects myocardial redox signaling must still be fully clarified, as that will give new therapeutic options for the prevention and treatment of the cardiac complications of obesity.Dysregulated redox signaling plays a role in the pathogenesis of many CVD, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs) are the major enzymatic source of superoxide . The study population consisted of patients undergoing cardiac surgery, all of whom were recruited under the Oxford Heart Vessels and Fat (ox-HVF) program at Oxford University Hospitals NHS Foundation Trust. Exclusion criteria included any inflammatory, neoplastic, renal, or hepatic disease.Description of the study arms is presented in the Human EAT  and its The screening phase and the confirmation phase of the miRNA profiling are described in the Patient genotyping and genome-wide genetic screening are described in the The H9c2 cardiomyocytes+) and C57BL/6, FVB CAG-rtTA (rtTA+) mouse lines have been previously described.A doxycycline-inducible wingless-type MMTV integration site family, member 5a (Wnt5a) knock-in mouse model was used to determine the in vivo effects of Wnt5a on myocardial NADPH oxidase activity. C57BL/6, FVB/N Tg(tetO-Wnt5a)17Rva/J -enhanced chemiluminescence, as we have previously described.,2.– was also quantified in cell lysates. Detailed protocols for O2.– quantification are presented in the Myocardial ORNA isolation and quantitative reverse transcription polymerase chain reaction methods are described in the Rac1 activation was assessed by using an active Rac1 detection kit . Details are provided in the Membrane translocation of Rac1 was estimated by differential centrifugation of cell lysates or tissue homogenates to isolate membrane proteins as previously described.Western immunoblot analysis was performed on myocardium homogenates and cell lysates .,Continuous variables were tested for normal distribution by using the Kolmogorov-Smirnov test. Non-normally distributed variables are presented as median (25th-75th percentile) and whiskers (from 10th to 90th percentile) for n ≥ 30. In experiments with a small sample size, variables are presented as scatterplots with median values.P < 0.05 was considered statistically significant.All statistical tests were 2-tailed and were performed by using SPSS version 20.0 . A value of A microarray analysis was performed on EAT and EAT-derived supernatants from patients who underwent cardiac surgery at Oxford University Hospitals (study arm 1)  to ident2.– production, as detected by lucigenin-enhanced chemiluminescence  to identify the leading single-nucleotide polymorphisms (SNPs)  associated with the levels of miR-92a-3p in EAT (EAT-miR-92a-3p) or myocardium (MYO-miR-92a-3p)  and 2E. oduction , whereasoduction .Figure 22.– generation in this cell type  and Frizzled class receptor 5 (FZD5) in the human myocardium was increased in patients with high myocardial NADPH-stimulated O2.–  . WNT5A induced a significant increase in myocardial NADPH oxidase–dependent Oby SFRP5 . Furtherby SFRP5 . To furtA+ mice) .12 Treatcontrols .12 In acneration  as well neration  in myocativation . Identictivation . Further2.– generation, we preincubated cardiomyocytes with NSC23766, a Rac1 inhibitor, before exposing them to WNT5A. NSC23766 abolished the ability of WNT5A to increase basal   (P = 0.043) after adjusting for age, sex, hypertension, smoking, hypercholesterolemia, body mass index, ethnicity, and ejection fraction.To explore whether these newly described effects of EAT-derived miR-92a-3p on myocardial redox state would translate to clinical phenotypes associated with cardiovascular outcomes, we followed up the population of study arm 5 for up to 8 years, through nationwide data collection via the NHS Digital coding system, to ascertain adverse clinical events. Patients with higher miR-92a-3p levels in EAT at baseline had a significantly lower risk for the composite outcome of cardiac mortality, nonfatal myocardial infarction (MI), and nonfatal cardioembolic stroke after adjusting for age, sex, hypertension, body mass index, and diabetes  . In linehe rest) . PatientWe also compared the demographic characteristics and other clinical factors among patients grouped according to tertiles of EAT miR-92a-3p levels. Although there was no significant difference in most of the risk factors between groups, patients with high miR-92a-3p levels were more likely to be taking antiplatelet medication or to have hypercholesterolemia .,,2.– from NADPH oxidases in the human heart. Causality was documented by using a Mendelian randomization approach in humans, as well as by showing that miR-92a-3p decreases NADPH oxidase–derived O2.– generation in cardiomyocytes in vitro. We then identified Wnt5a as a possible mediator of miR-92a-3p effects on myocardial redox state. Indeed, miR-92a-3p reduces the expression of Wnt5a through direct binding to its 3′-untranslated region.2.– production through Rac1-mediated NADPH oxidase activity . We also proved that this novel function of miR-92a-3p is mediated by Akt and possibly by downregulation of its target Wnt5a. The Akt inhibitor perifosine could indeed counteract the miRNA-mediated decrease in Rac1 activation/membrane translocation and consequent O2.– production, and miR-92a-3p transfection in cardiomyocytes downregulated protein levels of Wnt5a. The concept that EAT-derived miR-92a-3p could regulate myocardial Wnt5a is supported by a negative correlation between miR-92a-3p levels in EAT and myocardial WNT5A. Furthermore, it has been shown that miR-92a-3p directly binds to the 3′-untranslated region of Wnt5a to exert its extracellular repressive function; for example, Mao et alWNT5A/SFRP5 in the myocardium are negatively and positively associated, respectively, with risk of MACE.miR-92a-3p belongs to the miR-17-92 cluster and, in addition to playing a role in tumor biology, it is a key regulator of angiogenesis after MI.Although this study identified EAT-derived miR-92a-3p and its myocardial targets (Wnt5a/Rac1/NADPH oxidase activity) as predictors of cardiac outcomes, it is unclear how we could intervene pharmacologically. It is also unclear whether remote adipose tissue depots  could also secrete miR-92a-3p that might exert endocrine effects on the human heart. Furthermore, miRNA-92a-3p from human EAT may affect the myocardium in a paracrine way, but a definitive causal relationship has not been shown, and this is a limitation of the study. Moreover, in this study, we used differentiated H9c2 cardiomyocytes instead of primary human cardiomyocytes as an in vitro model for the mechanistic experiments. Finally, the paired RAA and the EAT from the atrioventricular groove were in anatomical proximity but not directly adjacent to each other; however, samples from the right atrioventricular groove have similar miR-92a-3p expression compared with samples directly adjacent to the RAA .PerspectivesCOMPETENCY IN MEDICAL KNOWLEDGE: EAT-derived microRNAs exert paracrine effects on human myocardium, miR-92a-3p improves regulation of the human myocardial redox state, and increased miR-92a-3p levels in EAT reduce cardiovascular risk.TRANSLATIONAL OUTLOOK: Further research is needed to clarify the role of miR-92a-3p in various cardiac cell types.The current study shows that high EAT-derived miR-92a-3p is associated with improved clinical cardiovascular outcomes and that this microRNA decreases myocardial oxidative stress, possibly by targeting the Wnt5a/Rac1/NADPH oxidase axis .PerspectThis study was supported by a Marie Skłodowska-Curie Early Stage Researcher fellowship to Dr Carena; the CATCH ME  consortium (grant number 633196); the British Heart Foundation ; Oxford BHF Centre of Research Excellence RE/18/3/34214, the Oxford NIHR Biomedical Research Centre, the National Institute for Health Research Oxford Biomedical Research Centre, and the Novo Nordisk Foundation (NNF15CC0018486) to Dr Antoniades. Dr Antoniades has had consultancy agreements with Mitsubishi Tanabe and Silence Therapeutics; has received grants from Sanofi and Novo Nordisk; is the Chair of the British Atherosclerosis Society; and has received honoraria from Amarin and Covance. Drs Antoniades and Channon are founders, shareholders, and directors of Caristo Diagnostics. Dr Casadei is the past president of the European Society of Cardiology. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose.