By contrast, ISD was not associated with UC . Our results suggest that consuming a more pro-inflammatory diet may contribute to the risk of CD, supporting that a healthy diet might be beneficial in its prevention. Further, larger studies are needed to verify these findings.Diet may influence the development of inflammatory bowel disease through the modulation of inflammation. We investigated whether the inflammatory potential of the diet is associated with the risk of Crohn’s disease (CD) and ulcerative colitis (UC) in the Spanish cohort of the European Prospective Investigation into Cancer and Nutrition (EPIC-Spain). The study included 32,633 participants aged 29–69 years. The inflammatory potential of the diet was measured by using an inflammatory score of the diet (ISD) based on a baseline dietary history questionnaire. Cox regression was used to estimate hazard ratios (HRs) and 95% confidence intervals (CIs). During 21 years  of follow-up, 32 and 57 participants developed CD and UC, respectively. In multivariable analysis, a one-standard deviation (SD) increment in the ISD (two-unit increase) was associated with a higher risk of CD (HR of 1.71; 95% CI: 1.05–2.80;  Inflammatory bowel disease (IBD) denotes a group of idiopathic immune-mediated disorders, characterized by chronic relapsing-remitting inflammation of the digestive tract. The two main types of IBD are ulcerative colitis (UC), which is a non-transmural disease limited to the colonic mucosa, and Crohn’s disease (CD), which is transmural and can cause inflammation throughout the entire gastrointestinal tract . In EuroIBD has been regarded as a disease of industrialized societies. Its incidence increased steadily in developed countries in the last decades of the twentieth century, and it is now rising rapidly in newly industrialized countries . The worThe possible effect of diet on the development of IBD is of great interest as it can be one of the risk factors most likely to be modifiable, and therefore a possible target for prevention. Available evidence suggests that diet influences gut inflammation through different mechanisms, which include alterations in the composition of gut microbiota and its interactions with the local immune system ,7. SeverA widely used approach to measure the inflammatory capacity of the diet has been the Dietary Inflammatory Index (DII) , which wThe objective of this study was to evaluate the association between the inflammatory potential of the diet, as measured by the ISD, and the incidence of UC and CD among participants of the Spanish EPIC cohort.EPIC is a multi-centre cohort study, conducted in 23 centres across 10 European countries, aiming to investigate the aetiological role of biological, lifestyle, and environmental factors in cancer and other chronic diseases. The full methodological details have been described elsewhere . The popn = 218 participants). In addition, we excluded 4 and 10 participants with prevalent CD and UC at recruitment, respectively. Thus, the final number of individuals included was 32,663.Participants with extreme values of the ratio energy intake to estimated energy requirement, defined as >3 standard deviations (SD) from the average log-transformed ratio, were considered to have implausible dietary data, hence they were excluded from the study  Ethics Committee approved the EPIC study protocol. All participants gave written informed consent.Participants were interviewed at recruitment to collect information on habitual diet during the previous year by using a validated dietary history questionnaire . The queThe ISD was calculated to measure the inflammatory capacity of the diet. This is a relative index that allows to rate an individual’s diet on a scale from maximally anti- to pro-inflammatory. The procedure followed to calculate the ISD in the EPIC cohort ,25, showThe DII is an index that includes 45 food parameters which were identified as having anti- or pro-inflammatory properties based on an extensive literature review. Each food parameter was assigned an inflammatory effect score according to its association with biomarkers . Of the The inflammatory effect scores (weights) for each food parameter were used as reported for the DII , with a Baseline data on the participants’ socio-demographic and lifestyle factors were collected by standardized questionnaires ,32. For The cohort was followed-up to identify those participants who developed IBD by linkage with hospital discharge and primary care databases and mortality registries, using the codes for CD and UC of the International Classification of Diseases—9th revision: 555 and 556, and 10th revision: K50 and K51—and the code D94 (chronic enteritis or ulcerative colitis) of the International Classification of Primary Care-2. For each potential case, the clinical records were thoroughly reviewed by local physicians to verify the diagnoses. Participants for whom the diagnosis of IBD was not confirmed were included in the analyses as non-cases.Standard descriptive statistics were used to summarize the baseline characteristics of the cohort, and differences across ISD tertiles were tested by Chi-squared or Kruskal–Wallis rank sum tests, as appropriate. Incidence rates of CD and UC were calculated and directly age- and sex-standardized using the 2013 European Standard Population. Cox proportional hazards regression models were fitted to assess the association of the ISD with CD and UC. Age was used as the underlying timescale, with entry time defined as the participant’s age at recruitment, and exit time as the age at IBD diagnosis, censoring, or death (whichever came first). The ISD was assessed as a continuous variable (1-SD increase) and as a categorical variable according to tertiles, using the highest tertile (most pro-inflammatory diet) as the reference. Linear trends across tertiles were tested by using the median value for each tertile as a continuous variable.Hazard ratios (HRs) and 95% confidence intervals (CIs) were obtained, adjusted for age, as time scale, and sex, and stratified by centre (model 1). A second model was additionally adjusted for energy intake as a continuous variable (model 2). Smoking status and physical activity did not change the HR by more than 10%, so they were not retained in the final model to avoid overfitting given the small number of cases. However, these covariates were retained in sensitivity analyses. The possibility of a nonlinear association of ISD with CD and UC risk was assessed by adding a quadratic term for ISD to the model. There was no evidence of a nonlinear relationship, and so a quadratic term was not retained in any of the fitted models. Effect modifications by age group (<50 and ≥50 years) and sex were evaluated by modeling interaction terms between these variables and ISD. To account for pre-diagnosis symptoms potentially influencing diet prior to diagnosis, we performed a lag analysis excluding the first two years of follow-up. For all models, the proportional hazards assumption was satisfied, evaluated by plotting the Schoenfeld residuals against the time variable in each model. p-values below 0.05 were considered significant. The analyses were performed using STATA 15.1 .Statistical tests were two sided, and A total of 32,663 participants, 20,168 (61.7%) women and 12,495 (38.3%) men, were included in the study. The median age at recruitment was 48.9 (interquartile range (IQR) 42.9–56.0) years. The inflammatory capacity of the diet in the cohort, as assessed by the ISD, had a mean (±SD) value of 0.24 (±2.02) and a median of 0.35, ranging from −6.08 to 4.98.The main baseline characteristics of the cohort by tertiles of the ISD are shown in p = 0.031). The HR for the lowest tertile was 0.32 (95% CI: 0.10–1.05) as compared with the highest tertile (P-trend = 0.067) (P for interaction = 0.234), nor between ISD and age group (P for interaction = 0.676).Among the 32 participants who developed CD, 12 had an ISD value below the cohort average. In multivariable Cox regression analysis, after adjustment by age, sex and energy intake and stratified by centre, one-SD increment in the ISD  was associated with a higher risk of CD with HR of 1.71  . Exclusi= 0.067) . We did p = 0.515)  . The sen= 0.515) . No inteIn this prospective cohort study in Spanish adults, we found that higher ISD values  were associated with an increased risk of CD, independently of age, sex, energy intake, and centre when ISD was examined as a continuous measure. However, no significant association was observed when ISD was analysed as a categorical variable, possibly due to the small number of cases in each category. On the other hand, no association was found between ISD and risk of developing UC. The finding of a possible association between inflammatory diet and UC is particularly relevant given the increasing incidence of this disease worldwide , and sinThere have also been some studies examining the effect of the inflammatory potential of the diet, as measured by a score, on gut inflammation and disease activity in patients already diagnosed with IBD, but both positive and null relationships have been reported ,39.Dietary patterns in relation to biomarkers of inflammation have been investigated in several epidemiological studies ,42,43,44Evidence also suggests that Western-type diets are positively whereas healthy patterns are inversely associated with the risk of IBD. For instance, an analysis performed in other EPIC sub-cohorts showed that a dietary pattern characterized by high consumption of sugar and soft drinks, and low intake of vegetables, was linked to an increased risk of UC . MoreoveA number of plausible explanations have been proposed to elucidate how the inflammatory potential of diet contributes to the inflammatory process underline the IBD aetiology, although the exact pathophysiological mechanisms are yet to be completely understood. It is well known that diet is a key factor in determining the structure and activity of the human gut microbiome ,47. A moOur study has a number of limitations. The small number of cases in the cohort, due to the relatively low incidence of IBD, limited the statistical power for some analyses. Children and very young adults were not represented in the study. Dietary data were collected at a single time point (at baseline), so we were not able to account for variations in dietary intake over time and this may have led to an underestimation of the associations. Although we used a previously validated dietary history questionnaire , and we Strengths of the current study include the prospective longitudinal design that avoids the possibility of reverse causation bias present in cross-sectional studies, the large study population involving participants from different Spanish geographic areas, and the long follow-up period that allowed accumulating a sufficient number of cases to detect a possible association for Crohn’s disease. Additional studies are needed to confirm this association, and in this respect, our results may contribute to future meta-analyses of the emerging literature on this topic. Other study strengths are the comprehensive evaluation of dietary intake, the availability of sociodemographic, anthropometric, and lifestyle data to adjust for, and the inclusion of only verified incident cases. Moreover, the ISD has been shown to be a valid tool in predicting different health outcomes ,24,25,26Our results suggest an association between a more pro-inflammatory diet, as measured by the ISD, and higher risk of developing Crohn’s disease. Our findings are consistent with the available literature showing a potential protective effect of consuming a diet rich in anti-inflammatory foods  and reducing the pro-inflammatory dietary intake  on the risk of Crohn’s disease. Studies using alternative methods for measuring the dietary inflammatory potential would provide further insight regarding this association and contribute to the verification of these findings.


Background: Smartphones may facilitate continuous and fine-grained monitoring of behavioral activities via automatically generated data and could prove to be especially valuable in monitoring illness activity in young patients with bipolar disorder (BD), who often present with rapid changes in mood and related symptoms. The present pilot study in young patients with newly diagnosed BD and healthy controls (HC) aimed to (1) validate automatically generated smartphone data reflecting physical and social activity and phone usage against validated clinical rating scales and questionnaires; (2) investigate differences in automatically generated smartphone data between young patients with newly diagnosed BD and HC; and (3) investigate associations between automatically generated smartphone data and smartphone-based self-monitored mood and activity in young patients with newly diagnosed BD.Methods: A total of 40 young patients with newly diagnosed BD and 21 HC aged 15–25 years provided daily automatically generated smartphone data for 3–779 days [median (IQR) = 140 (11.5–268.5)], in addition to daily smartphone-based self-monitoring of activity and mood. All participants were assessed with clinical rating scales.Results: (1) The number of outgoing phone calls was positively associated with scores on the Young Mania Rating Scale and subitems concerning activity and speech. The number of missed calls (p = 0.015) and the number of outgoing text messages (p = 0.017) were positively associated with the level of psychomotor agitation according to the Hamilton Depression Rating scale subitem 9. (2) Young patients with newly diagnosed BD had a higher number of incoming calls compared with HC  and lower self-monitored mood and activity (p's < 0.001). (3) Smartphone-based self-monitored mood and activity were positively associated with step counts and the number of outgoing calls, respectively (p's < 0.001).Conclusion: Automatically generated data on physical and social activity and phone usage seem to reflect symptoms. These data differ between young patients with newly diagnosed BD and HC and reflect changes in illness activity in young patients with BD. Automatically generated smartphone-based data could be a useful clinical tool in diagnosing and monitoring illness activity in young patients with BD. Bipolar disorder (BD) is a serious, recurrent, and disabling disorder often with an onset of symptoms during a young age . In addiDiagnostic work in children and adolescents with psychiatric disorders is especially challenging as it is often characterized by unspecific prodromal symptoms . CorrectSmartphones, equipped with sensors, such as accelerometers, are widely used all over the world, with 45% of people in the world owning a smartphone . This alFor many young people, smartphone interaction is a significant part of their everyday life and important for social interaction . Thus, sHowever, for a diagnostic tool to be useful, it needs foremost to be able to differentiate between patients and healthy controls (HC). Recent studies have shown that smartphone-based self-monitored data represent symptom burden according to clinical ratings and are also able to differentiate between adult patients with newly diagnosed BD and HC , 12, 15.Prior research has found automatically generated smartphone data to be useful, feasible, and valid for adult patients with BD , 17–20. The present pilot study aimed to (1) validate automatically generated smartphone data reflecting physical and social activity and phone usage against validated clinical ratings and questionnaires in young patients with newly diagnosed BD and HC; (2) investigate differences in automatically generated smartphone data reflecting physical and social activity and phone usage between young patients with newly diagnosed BD and HC; and (3) investigate associations between automatically generated smartphone data reflecting physical and social activity and phone usage and smartphone-based self-monitored activity and mood in young patients with newly diagnosed BD.Based on prior research on adults with BD, we hypothesized that (1) automatically generated smartphone data reflecting physical and social activity and phone usage would be associated with validated clinical ratings and questionnaires, among young patients with newly diagnosed BD and HC; (2) automatically generated smartphone data reflecting physical and social activity and phone usage differ between young patients with newly diagnosed BD and HC with lower scores in physical and social activity in BD than in the HC group; (3) automatically generated smartphone data reflecting physical and social activity and phone usage would be associated with smartphone-based self-monitored activity and mood, with positive associations between smartphone-based self-monitored activity and mood, and automatically generated smartphone data on physical and social activity, respectively, in young patients with newly diagnosed BD.The participants included in the present study were recruited as part of the Bipolar Illness Onset study (the BIO study) , a longiAmong the participants in the BIO study, we recruited individuals newly diagnosed with BD and control persons without a personal or psychiatric family history, aged 25 years or younger at the time of inclusion (HC). Participants newly diagnosed with BD were recruited from the Copenhagen Affective Disorder Clinic at Rigshospitalet in Copenhagen, Denmark. The Copenhagen Affective Disorders Clinic is a specialized clinic that offers a 2-year course of treatment to everyone newly diagnosed with BD in the larger region of Copenhagen over the age of 18. We also included young patients with newly diagnosed BD under the age of 18, from the Child and Adolescent Mental Health Center in Copenhagen. We recruited HC among blood donors from the Blood Bank at Rigshospitalet. The exclusion criterion for the latter was a history of a psychiatric disorder requiring treatment, personally or in a first-degree relative.All participants underwent a diagnostic interview using Schedules for Clinical Assessment in Neuropsychiatry (SCAN) , during The baseline interview consisted of a collection of general information about educational and work status in addition to diagnostic and clinical assessments. Information about time for onset of symptoms, diagnosis, start of treatment, and number and duration of affective episodes was collected among the young patients with newly diagnosed BD. All participants also completed a questionnaire addressing physical activity. After baseline, participants attended an annual follow-up interview as well as interviews every time they had a change from one affective episode to another, from an affective episode to euthymic stage, or from a euthymic stage to an affective episode.At baseline and follow-up interviews, the following rating scales were used: the severity of depressive symptoms was assessed using the 17-item Hamilton Depression Rating Scale (HAMD) , 29; theThe participants completed the International Physical Activity Questionnaire Short Form (IPAQ) at each visit with the researchers. In the IPAQ, participants report how many minutes of physical activity in different intensities  they had the prior week; these are then converted to metabolic equivalent task (MET) minutes per week, which then add up to a total MET score . MET is The Monsenso system is a smartphone-based monitoring system with a collection of automatically generated smartphone data as mobility, activity, and phone usage using the telephones sensors, in addition to daily self-monitoring of, e.g., mood, sleep, and activity , 35. TheThe automatically generated smartphone data collected daily by the Monsenso system were step counts; the total number of steps during a 24-h period detected by the accelerometer in the phone, reflecting the physical activity; the number of incoming and outgoing text messages during a 24-h period, reflecting social activity; duration and number of incoming and outgoing calls and the number of missed calls during a 24-h period, reflecting social activity; and the number of seconds the screen was on (screen time) and the number of times the screen was turned on, reflecting the smartphone usage.For self-monitoring, all participants scored their daily level of activity and their mood, reflecting how good or how bad their day had been on a 7-point scale . Additionally, young patients with newly diagnosed BD scored their affective state on a 9-point scale with scores from depressive to manic, which is a more fine-grained scale  .a priori. For continuous variables, we used linear mixed effect models to investigate between-group differences in the mean regarding background characteristics. We used chi-square tests for between-group differences of categorical data. For analyses concerning aims 1–3, for each measure of interest, we employed a linear mixed effect model, which accommodates both the variation of the variables of interest within young patients with newly diagnosed BD  and between individuals . For participants with data available from both baseline and up to nine follow-up visits, a linear mixed effect model analysis to account for repeated measurements within each participant was employed. All participants were identified with a unique ID number. For aim 1, we used a linear mixed effect model to analyze the association between automatically generated smartphone data, as the dependent variable, and scores from clinical rating scales and questionnaires. For these analyses, averages of automatically generated smartphone data for the current day and 3 days before ratings with the HAMD and the YMRS, 7 days prior for the IPAQ, and 14 days prior for the FAST were calculated. For aim 2, we used a linear mixed effect model to investigate the between-group difference in daily automatically generated smartphone data between young patients with newly diagnosed BD and HC, from every observation available. For aim 3, we used a linear mixed effect model to analyze the associations between smartphone-based self-monitored mood and activity, as the dependent variable, and automatically generated smartphone data on physical and social activity and phone usage, respectively. Individual ID number was added as a random factor for all analyses. For aim 1, we included pooled data available from all participants. For aim 3, we included data available from young patients with newly diagnosed BD only. Analyses were conducted first in an unadjusted model and secondly in models adjusted for age and sex, by adding sex and age (as a numeric covariate) as fixed factors.Hypotheses and statistical analyses for the present study were defined p-value ≤ 0.05 was considered statistically significant.As no prior studies have investigated differences in automatically generated smartphone data between young patients with newly diagnosed BD and HC, we were not able to perform statistical power analyses prior to the study. Since data were collected as part of a larger longitudinal observational study, the sample size was defined according to this. As this is the first study on automatically generated smartphone data in young patients with BD and, in this way, the study is explorative by nature, we did not correct for multiple analyses. We checked model assumptions by visually utilizing residuals and QQ plots for each of the statistical analyses. SPSS version 25  was used for all analyses. A p = 0.012), the groups did not differ on other background characteristics , the HAMD subitem 8 , and the HAMD subitem 9 . Thus, for every increase of 1 point in the HAMD total score, there was a 0.04 increase in the number of incoming calls.As can be seen in B = 0.18; 95% CI: 0.092, 0.26; p < 0.001), the YMRS subitem 2 , the YMRS subitem 6 , and the HAMD subitem 9 . Thus, for every increase of 1 point in the YMRS total score, there was a 0.18 increase in the number of incoming calls.As can be seen in B = 0.41; 95% CI: 0.083, 0.73; p = 0.015). Thus, for every increase of 1 point in the HAMD subitem 9 score, there was a 0.41-point increase in the number of missing calls. The number of incoming text messages was statistically significantly positively associated with the HAMD subitem 9 , and the same was the case between the number of outgoing text messages and the HAMD subitem 9 . Thus, for every increase of 1 point in the HAMD subitem 9 score, there was a 3.26-point increase in the number of incoming text messages and a 2.88-point increase in the number of outgoing text messages. For the associations between the number of incoming text messages and scores on the clinical rating scales, age was also a significant covariate. We found no associations between screen time, the number of step counts, the number of times the screen was turned on, total screen time, and scores on clinical ratings or questionnaires.The number of missed calls was statistically significantly positively associated with the HAMD subitem 9 (These results can be seen in p = 0.043). There were no other statistically significant differences in automatically generated smartphone data between young patients with newly diagnosed BD and HC. Furthermore, there was a statistically significant difference in smartphone-based self-monitored mood as well as activity between young patients with newly diagnosed BD and HC .As can be seen, there was a statistically significant difference in the number of incoming calls between young patients with newly diagnosed BD and HC , the number of time the screen was turned on , number of outgoing calls , and the number of incoming text messages . Furthermore, there was a statistically significantly negative association between smartphone-based self-monitored activity and automatically generated data on screen time  and call duration . Thus, for every increase of 347 increase in step count, there was 1-point increase in smartphone self-monitored activity.As can be seen in B = 367.48; 95% CI: 119.61, 615.35; p = 0.004) and numbers of time the screen was turned on . There was a statistically significant negative association between smartphone-based self-monitored mood and automatically generated data on screen time . Thus, for every increase of 367 in step count, there is a 1-point increase in smartphone-based self-monitored mood.There was a statistically significant positive association between smartphone-based self-monitored mood and automatically generated data on step count , and clinically evaluated symptoms differed between young patients with newly diagnosed BD and HC. However, in contrast with our hypotheses, there were no other differences in automatically generated smartphone data between young patients with newly diagnosed BD and HC. In addition, as hypothesized, automatically generated smartphone data on physical activity and phone usage were associated with smartphone-based self-monitored mood and activity in young patients with newly diagnosed BD.The findings that automatically generated smartphone data on social activity were associated with clinically evaluated depressive and manic symptoms are in line with prior studies on adult patients with BD –13.However, in the present study, we only found this association between social activity, reflected by the number of calls and text messages, and total scores and subitem scores on the HAMD and the YMRS. A possible reason for this could be that the participants included in the present study did not present with severe symptoms during the clinical evaluations, and thus, the associations may differ during more severe states. Also, a possible explanation could be that young patients with newly diagnosed BD may present with more complex clinical presentation with unspecific prodromal symptoms  or a morThe findings that smartphone-based self-monitored mood, activity, and automatically generated smartphone data on social activity  differed between young patients with newly diagnosed BD and HC are in line with prior studies on adult patients with BD  and may The findings that automatically generated smartphone data on physical activity and phone usage was associated with smartphone-based self-monitored mood and activity in young patients with newly diagnosed BD are also in line with prior studies on adult patients with BD –13, 20. In the present study, we chose a naturalistic approach where the participants used their own smartphones as they would naturally; hence, there are missing data from when participants, i.e., turned off their phone, left it at home, or deactivated the app. Due to the nature of automatically generated smartphone-based data, we cannot say anything about the time periods where data were not available (missing data). However, since most people carry their smartphone with them during most of the day and use it in most of their online communications, we find that the results from the present study are valid. Overall, results from the present study confirm that automatically generated smartphone data may be used to monitor illness activity in young patients with BD unobtrusively during naturalistic settings between outpatient visits in a fine-grained and valid manner.Firstly, due to a large portion of iPhone users among the participants included in the BIO study, the number of participants using Android phones and thus being eligible for the present study was relatively limited. Therefore, the results from the present study should be interpreted with caution, as negative findings may be due to type II errors. It is possible that inclusion of a larger sample size and inclusion of a larger control group would have revealed other associations and differences. However, the present study is the first of its kind and therefore hypotheses generating.Secondly, few young patients with newly diagnosed BD were clinically evaluated during severe depressive and manic episodes; this will lower the probability of finding associations between automatically generated smartphone data and clinically evaluated symptoms reflected by clinical rating scales. Future studies, with a higher number of established affective episodes, could do classification modeling investigating differences in smartphone data during euthymic and affective episodes. Thirdly, as the participants used their own smartphone, there was no standardized platform for data collection, possibly leading to heterogeneity in the data collected. Fourthly, several of the young patients with newly diagnosed BD were using psychotropic medicine; this was not accounted for in the analyses. Fifthly, the HC in the present study were recruited among blood donors, possibly representing a “super healthy” population , 39. NevAll young patients with newly diagnosed BD in the present study were diagnosed at a specialized mood disorder clinic, and the diagnosis or lack of diagnoses was verified for all participants with a SCAN interview conducted by trained PhD students in medicine or psychology. Additionally, all participants were assessed using clinically validated observer-based rating scales with the HAMD, the YMRS, and the FAST. The smartphone-based system used in the present study is well-validated, is useful, and fulfills the safety of data storage and privacy requirements. Although the collection of automatically generated data works on Android led to a smaller population in this study, it is an advantage regarding global generalizability, as Android is the most used OS worldwide.Prior research has shown smartphone-based monitoring in young patients with psychiatric disorders to be feasible and acceptable , and as The present innovative pilot study investigated the use of automatically generated smartphone data collected daily during a long-term period in young patients with newly diagnosed BD and HC. Automatically generated smartphone data on social activity reflected clinically evaluated depressive and manic symptoms, differed between young patients with newly diagnosed BD and HC, and reflected self-reported changes in mood and activity in young patients with newly diagnosed BD. Although a rather low number of participants with a rather low level of affective symptoms during the study were included, our results suggest that automatically generated data on physical and social activity and phone usage could represent a potentially useful tool in diagnosing and monitoring young patients with newly diagnosed BD.Future studies including young patients with newly diagnosed BD during more severe affective states, unaffected first-degree relatives, and different psychiatric disorders could provide interesting knowledge on early changes in communicative activities.https://www.psykiatri-regionh.dk/forskning/forskningsomraader/Neuropsykiatri/cadic/Sider/default.aspx.The datasets presented in this article are not readily available because the study is ongoing; and therefore, the research data are not shared. Requests to access the datasets should be directed to The studies involving human participants were reviewed and approved by The Bipolar Illness Onset (BIO) study has been approved by the Ethics Committee in the Capital Region, Copenhagen, Denmark (Ref. Nr. H-7-2014-007) and the Danish Data Protection Agency, Capital Region of Copenhagen (Protocol No.: RHP-2015-023). Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin. Written informed consent was obtained from the individual(s), and minor(s)' legal guardian/next of kin, for the publication of any potentially identifiable images or data included in this article.LK, MF-J, and MV conceived the study and were in charge of overall direction and planning. JB and MF were in charge of the development of the technology, which was further adjusted for clinical use in by LK, MF-J, MV, MF, and JB. SM and SS were in charge of recruiting and clinical data collections. SM, SS, and MF-J conducted the statistical analysis. SM, MF-J, and LK were in charge of the manuscript. All authors have contributed in the final configuration of the manuscript.LK has, within the preceding 3 years, been a consultant for Lundbeck. MV has in the last 3 years been a consultant for Sunovion, Janssen, and Lundbeck. JB and MF are co-founders and shareholders in Monsenso ApS. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Cachexia is a paraneoplastic syndrome that accompanies and compromises cancer treatment, especially in advanced stages, affecting the metabolism and function of several organs. The adipose tissue is the first to respond to the presence of the tumor, contributing to the secretion of factors which drive the systemic inflammation, a hallmark of the syndrome. While inflammation is a defensive innate response, the control mechanisms have been reported to be disrupted in cachexia. On the other hand, little is known about the role of NLRP3 inflammasome in this scenario, a multiprotein complex involved in caspase-1 activation and the processing of the cytokines IL-1β and IL-18.based on the evidence from our previous study with a rodent model of cachexia, we examined the activation of the NLRP3 inflammasome pathway in two adipose tissue depots obtained from patients with colorectal cancer and compared with that another inflammatory pathway, NF-κB.For CC we found opposite modulation in ScAT and PtAT for the gene expression of TLR4, Caspase-1 (cachectic group) and for NF-κB p50, NF-κB p65, IL-1β. CD36, expression was decreased in both depots while that of NLRP3 and IL-18 was higher in both tissues, as compared with controls and weight stable patients (WSC). Caspase-1 basal protein levels in the ScAT culture supernatant were higher in WSC and (weight stable patients) CC, when compared to controls. Basal ScAT explant culture medium IL-1β and IL-18 protein content in ScAT supernatant was decreased in the WSC and CC as compared to CTL explants.The results demonstrate heterogeneous responses in the activation of genes of the NLRP3 inflammasome pathway in the adipose tissue of patients with cancer cachexia, rendering this pathway a potential target for therapy aiming at decreasing chronic inflammation in cancer. Cancer is among the four major causes of premature death in the world. The risk of getting sick and dying from cancer is related to hereditary aspects, increased life expectancy, the process of aging, lifestyle habits, the influence of risk factors, and the environment in which people live in. In low- and middle-income countries, such as Brazil, there has been a reduction in the number of cases of infectious agents-related cancers, such as those in the oral cavity and cervix, and an increase in the types that are more common in developed countries, including colorectal cancer (CRC). The latter is the third most incident type of cancer in the world .Cancer cachexia is an overly debilitating and harmful paraneoplastic syndrome that compromises the therapeutic response and quality of life of patients. Its etiology is complex and not yet fully understood, but sustained systemic inflammation is a peculiar feature, stimulating exacerbated energy expenditure through cytokine secretion by the tumor and host tissues in response to the tumor, causing muscle and adipose tissue loss –7.Considered a multiorgan, white adipose tissue performs an important role in health and disease. It is an active endocrine organ, responsible for the secretion of steroid precursors of hormones, cytokines, autacoids, fatty acids, and derivatives, and the so called adipokines, with functions involved in the regulation of fat storage, energy metabolism, food intake, insulin sensitivity and immunity, amongst many other –14. UndeAdipose tissue acts at the crossroads between metabolism and immunity: as it is an endocrine organ, it is capable of regulating the immune system, and the signaling between these two compartments displays a delicate and intricate balance –4, 7. InWe have previously shown that there is a distinct pattern of secretion of inflammatory factors by the tumor and the adipose tissue  and differences in the tumor microenvironment in cachectic patients versus weight-stable patients , 19. In Due to the contribution of adipose tissue to several biological processes and to cancer cachexia, we sought to investigate the activation of components of the NLRP3 inflammasome pathway in the two white adipose tissue depots in patients with colorectal cancer. A PubMed search retrieves only 5 articles under “inflammasome and cachexia”. As demonstrated for obesity, different adipose compartments present heterogeneous function/contribution in health and disease. The present study was designed as to confirm that the changes reported previously for animal model of cachexia , with inThis research project was approved, following the Declaration of Helsinki, by the Research Ethics Committees of the “Faculdade de Medicina” (CAAE no. 54503716.1.0000.0065), of the University Hospital (CAAE no. 54503716.1.3001.0076) and the Institute of Biomedical Sciences (CAAE no. 54503716.1.3002.5467) of the University of São Paulo (USP). Twenty-eight volunteers were recruited into the study at diagnosis. Informed consent before to surgery was obtained from all participants. Inclusion criteria were diagnosis of colorectal cancer with an indication for surgery, or with a surgical indication for correction of abdominal wall hernias , cholecystectomies, and abdominal proctoplexy, and willingness to participate in the study. Exclusion criteria: having received antibiotic therapy, anti-inflammatory therapy, or anti-cancer treatment. extreme low weight or obesity , chronic heart failure, liver or kidney failure, chronic obstructive pulmonary disease, acquired immunodeficiency syndrome (AIDS), inflammatory bowel disease (IBD), hepatitis and autoimmune diseases.For the diagnosis of cachexia, the criteria established in international consensus by Evans and others were employed : unintenDuring the surgical procedure, fragments of approximately 1g of subcutaneous adipose tissue (ScAT) and visceral adipose tissue (mesenteric) immediately close to the tumor  were collected.Adipose tissue explants were distributed in 12-well culture plates, with approximately 500 mg per well and incubated in DMEM containing 10% fetal bovine serum (FBS), supplemented or not with lipopolysaccharide  (1 µg/ml), in a humidified atmosphere of 10% CO2 at 37°C for 24h , 23. The© reagent  observing the manufacturer’s instructions. The concentration of total RNA was evaluated with the spectrophotometer. The cDNA was obtained by reverse transcription (RT) using 1µg of total RNA from each sample per reaction, in a final volume of 20 µl. The target gene was normalized to the reference gene GAPDH. The mRNA levels were determined using the 2-ΔΔCt method, in which, for each sample, the ΔCt value was obtained by subtracting the control gene values from those of the target gene; then, the average ΔCt value of the control group was subtracted from the sample to get a -ΔΔCt value , ELISA Max™ Deluxe Set Human IL-1β , and RayBio® Human IL-18 ELISA Kit .The protein concentration of the culture medium supernatant was determined by the Bradford method . Afterward, the quantification of proteins  was performed using ELISA. The kits used for these tests were Human Caspase-1 SimpleStep ELISAX2) was used. For multiple comparisons, the ANOVA 2-way test was used. The Uncorrected Fisher’s LSD post-test was applied. Outliers were identified using the Rout method and removed from the analysis.Comparisons among groups were carried out using the ANOVA 1-way test, for data with normal distribution, or the Kruskal-Wallis test, for data with asymmetric distribution. To assess the dispersion of nominal variables, the Chi-square test . Data were expressed as mean ± standard error unless otherwise indicated.For all analyses, a significance level of p<0.05 was adopted, using the PrismWe grouped the colorectal tumors according to location, in the right colon  and left colon . As for the anatomical location of the tumor, most patients presented cancer in the left colon, both for WSC group (80%) and CC (60%). Regarding tumor staging, the percentages were higher in stages II and III, in both groups with cancer.2 are at a higher risk of developing CRC  . ClinicaCachexia not only impacts cancer progression but also compromises the therapeutic response, increases treatment costs and length of hospitalization , and is The aim of the study was to confirm in human cachexia what we have previously shown in animal models of the disease: white adipose tissue plays a relevant role for sustained systemic inflammation. In these earlier studies we have reported that adipose tissue response in cachexia was not uniform in terms of tissue architectural changes, nor did different anatomical depots contribute to the same extent to inflammation , 40–48. When assessing whether there was heterogeneity in the inflammatory response of adipose tissue depending on the anatomical location  in an animal model of cachexia, Neves and others  reportedScAT and VAT  are very distinct, so differences in gene and protein expression profiles may contribute to the observed disturbances. Different fat depots differ in cell size, physiological function, and contribution to disease states . In VAT,in vitro PtAT morphofunctional changes, for example, shrinkage in size and number of adipocytes with intense lipolysis and a higher rate of macrophage infiltration  and non-canonical (which regulates various immune functions), responding to different types of receptors and their ligands. Control the expression of several genes involved in inflammation, immunity, cell survival, proliferation, and differentiation –65.The gene expression of the two NF-κB domains was not significantly different Figure 2The expression of NLRP3 was increased in the cachectic group in both tissues of patients in the CC group. Experimental animal studies have linked NLRP3 inflammasome activation to inflammatory bowel diseases , 16. In in vitro and in animal models, as it can promote the epithelial-mesenchymal transition in intestinal cells .The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding authors.The studies involving human participants were reviewed and approved by Faculdade de Medicina, Hospital Universitário and Instituto de Ciências Biomédicas - University of São Paulo. The patients/participants provided their written informed consent to participate in this study.JJ, JO, and MS conceived the project. JJ performed the experiments, with the help of AM, KR, RM, RF, and AP. JJ wrote the manuscript. JO and MS ensured funding, supervised the project, and helped to correct the manuscript. JR-N and EM-N provided technical support. PA, FT, LM, FB, and FF provided the adipose tissue samples. All authors have contributed equally to this work. All authors contributed to the article and approved the submitted version.We thank CAPES  for the scholarship. Financial support was provided by FAPESP 12/50079-0 and 20/7765-6.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Despite the high prevalence of diabetic neuropathy, its early start, and its impact on quality of life and mortality, unresolved clinical issues persist in the field regarding its screening implementation, the understanding of its mechanisms, and the search for valid biomarkers, as well as disease-modifying treatment. Genetics may address these needs by providing genetic biomarkers of susceptibility, giving insights into pathogenesis, and shedding light on how to select possible responders to treatment. After a brief summary of recent studies on the genetics of diabetic neuropathy, the current review focused mainly on microRNAs (miRNAs), including the authors' results in this field. It summarized the findings of animal and human studies that associate miRNAs with diabetic neuropathy and explored the possible pathogenetic meanings of these associations, in particular regarding miR-128a, miR-155a, and miR-499a, as well as their application for diabetic neuropathy screening. Moreover, from a genetic perspective, it examined new findings of polymorphisms of miRNA genes in diabetic neuropathy. It considered in more depth the pathogenetic implications for diabetic neuropathy of the polymorphism of MIR499A and the related changes in the downstream action of miR-499a, showing how epigenetic and genetic studies may provide insight into pathogenetic mechanisms like mitochondrial dysfunction. Finally, the concept and the data of genotype-phenotype association for polymorphism of miRNA genes were described. In conclusion, although at a very preliminary stage, the findings linking the genetics and epigenetics of miRNAs might contribute to the identification of exploratory risk biomarkers, a comprehensive definition of susceptibility to specific pathogenetic mechanisms, and the development of mechanism-based treatment of diabetic neuropathy, thus addressing the goals of genetic studies. Both distal symmetric polyneuropathy (DPN) and cardiovascular autonomic neuropathy (CAN) are common complications of diabetes. DPN and CAN affect 30% and 20%, respectively, of the unselected population with diabetes, and their prevalence increases according to age and diabetes duration , 2. MoreMoreover, not all patients needing symptomatic relief receive appropriate treatment, the latter is not universally effective , 12, andEven when considering the most effective preventive strategy , it is a matter of fact that for many patients (2% yearly), current strategies for optimizing glucose control are insufficient to fully prevent or delay the development of neuropathic complications . Thus, tThe search for biomarkers, i.e., a measurable indicator of a pathophysiological condition, has become a main research topic in the field of DPN with the need to identify outcome biomarkers for clinical efficacy in clinical trials , sensitive biomarkers for the very early stages of disease , prognostic biomarkers for the tough consequences of DPN, and biomarkers pertaining to pathogenetic mechanisms in order to identify responsive patients to therapeutic agents targeted at these mechanisms , 15.Genetics may address diabetic neuropathy needs by providing exploratory genetic biomarkers of susceptibility for disease development, by giving insights into the pathogenesis of neuropathy and neuropathic pain, and by shedding light on how to select the responders to treatment.Genetics has in turn its own needs and requirements. First of all, two main different approaches are possible. The candidate gene approach starts with a hypothesis based on current knowledge of what kind of gene you intend to look for. It involves a limited number of variants, selected in candidate genes, and requires lower statistical power. Regarding diabetic neuropathy, candidate genes should encode proteins involved in the known mechanisms of nerve protection or damage in diabetes.p < 5 × 10−8), wide coordination between different researchers and centers (Consortia), and considerable resources and replication studies  and looking for consistent difference between frequencies in variants associated with a disease. Without a priori-hypothesis, newly identified genetic markers can open to new knowledge. GWASs require large sample sizes , rigorous thresholds for statistical significance  gene, glutathione peroxidase-1 (GPx-1) gene, and catalase (CAT) gene] have received greater attention, and even meta-analyses are available, with their polymorphisms possibly involved in the increased renin–angiotensin system activity, hyperhomocysteinemia, and reduced defense against oxidative stress . Moreovereceptor , a purinreceptor , and thereceptor –28, possFinally, four GWASs are available in diabetic neuropathy, 3 from the datasets from UK Genetics of Diabetes Audit and Research in Tayside Scotland (GoDARTS) and pertinent to the phenotype of neuropathic pain and foot ulceration –31 and tThus, there is increasing attention given to the genetics of diabetic neuropathy, and although a suitable genetic biomarker has not yet been developed, promising findings can be seen on the horizon.This review is focused on the genetics and epigenetics of microRNAs in diabetic neuropathy, given the research done by our group on this topic in the last few years.MicroRNAs (miRs or miRNAs) are small RNA molecules (20–22 nucleotide length) that act as regulators in biological processes. At least 20–30% of all human genes are regulated by miRNAs through targeting sequences in their 3′ untranslated region. DNA regulatory regions might be involved in the control of development processes, hematopoietic cell differentiation, apoptosis, cell proliferation, and organ growth and in disease development. Increasing evidence supports the involvement of miRNAs in diabetes and its micro- and macrovascular complications .κB) signaling pathway [Regarding diabetic neuropathy, in the last decade, very few studies have addressed miRNA expression in diabetic neuropathy with it being almost exclusively preclinical , 35. Stu pathway .Very few studies have explored the association between the expression of miRNAs and diabetic neuropathy in humans. In a study hampered by an unclear definition of both type 2 diabetes and diabetic neuropathy, miR-199a-3p was found upregulated in the plasma of 60 patients with type 2 diabetes and in the lower limb skin of 30 patients with type 2 diabetes and DPN, compared to samples of 5 and 20 healthy volunteers, respectively . Additiop = 0.015) and a lower expression of both miR-155 (p = 0.04) and miR-499a (p = 0.05), whereas patients with CAN, compared to those without, displayed only a lower expression of miR-155 (p = 0.05) [In a collaborative study between diabetologists and geneticists, we evaluated in 49 patients with type 2 diabetes the expression of 6 candidate miRNAs and assessed the presence of (1) DPN using validated scoring systems for neuropathic symptoms and signs and quantitative sensory testing for vibratory and thermal perception thresholds and (2) CAN by four cardiovascular reflex tests (CARTs) . We defi = 0.05) .Using ROC analysis, we found fair diagnostic accuracy for DPN and diabetic neuropathy (DPN and/or CAN) with both a model including all three miRNAs and a model with miR-128a plus miR-155. For diabetic neuropathy, the area under the ROC curve (AUC) was 0.817 and 0.801 for the three-miRNAs and two-miRNA model, respectively, the sensitivity was 75.9% and 80.6%, and specificity was 76.5% and 70.6%, respectively. For DPN, AUC was 0.815 and 0.802, sensitivity was 74.1% and 80.6%, and specificity was 76.2% and 70.6% for the three-miRNAs and two-miRNA model . Thus, tTo understand the possible meaning of these associations, one might consider that in animal studies, miR-128a downregulates insulin signaling pathways , impedesOn the other hand, miR-155 is a multifunctional miRNA: in animal studies, it enhances insulin sensitivity, regulates inflammation and immunity , and exeFinally, miR-499a, preferentially expressed in heart, skeletal muscle, and areas of the central autonomic network (nucleus ambiguous), has been found in animal models to be a regulator of the apoptotic pathway and mitochondrial fission in cardiomyocytes in response to ischemia or mechanical stress , 60 and p = 0.008) and the G allele of rs531564 in MIR124A as a risk allele . This was the first report of genetic polymorphisms in miRNA regions as possible contributors to type 2 diabetes susceptibility [Although the majority of studies are focused on miRNA expression profile investigation, recent studies have shown that also polymorphisms in miRNA genes may alter a wide spectrum of biological mechanisms and could play a role in the susceptibility to several human diseases, including diabetic neuropathy. In this regard, in our earlier studies, we used a genomic approach to investigate the relationship between miRNAs and diabetic neuropathy. The first study explored genetic polymorphisms in miRNA regions in relation to the susceptibility to type 2 diabetes . This sttibility .p = 0.032), while the variant T allele of rs11888095 in MIR128A was associated with a high risk of developing DPN . The latter association was also confirmed after correction for BMI, age, disease duration, HbA1c, and gender . Moreover, the same SNP in MIR146A showed a protective effect for early CAN  and for confirmed CAN , while a polymorphism in MIR27A was associated with a higher risk of developing early CAN . An association of SNPs of MIR128A and MIR146A was also present in multiple linear regression analysis with the severity of DPN and CAN, namely, the scores for neuropathic symptoms/signs (p = 0.026 for MIR128A) and the score based on CARTs (p < 0.0001 for MIR146A) [Subsequently, we evaluated the possible contribution of genetic polymorphisms of miRNA genes in susceptibility to DPN and CAN . Nine poMIR146A) . This reκB [We described above that in animal models of diabetic neuropathy, the expression of miR-146a was found to be downregulated and inversely associated with levels of inflammatory cytokines  and, furκB . SimilarκB . On the κB .The polymorphisms in these miRNA genes might affect the expression or the downstream action of the corresponding miRNAs and lead in some way to changes in metabolic and inflammatory/immune mechanisms active in the scenario of diabetic neuropathy.p = 0.002), confirmed CAN , and DPN . In addition, MIR499A GG genotype independently contributed to early CAN together with duration and HbA1c and to DPN together with duration, HbA1c, and age. Finally, the GG genotype was associated with worse values of neuropathic deficit score, i.e., Michigan Diabetic Neuropathy Score (p = 0.017), vibration perception threshold (p = 0.01), thermal thresholds (p = 0.01), and CART score (p < 0.001): in a multiple linear regression, the GG genotype was the main variable contributing to the CART score (p = 0.001). Thus, the rs3746444 GG genotype might represent a marker of higher risk of DPN and CAN and of CAN severity. We described above the hypothesized actions of miR-499a and its ability to prevent mitochondrial fission as well as the imbalance in mitochondrial fusion and fission as a potential pathogenetic mechanism of diabetic neuropathy [In a more recent study, in 150 participants with type 2 diabetes, we analysed the rs3746444 SNP in the MIR499A gene to evaluate its association with susceptibility to DPN and CAN . We founuropathy .p = 2 × 10−10) and further differences between patients with and without DPN (p = 0.02) (not between those with and without CAN). In addition, the homozygous variant genotype for the rs3746444 polymorphism of MIR499A was associated with the number of mtDNA copies, particularly in patients with type 2 diabetes with values of 22.71 ± 8.65 in patients with AA+AG genotypes and 14.43 ± 5.29 in those with the GG genotype (p = 0.009).More insight into the meaning of the association between MIR499A polymorphism and diabetic neuropathy comes from a further study by our group  that meaMitochondrial biogenesis is a defense modality against hyperglycemic load and hyperglycemia-driven oxidative stress in diabetes through the increase in the mass of the mitochondrial network, with mtDNA being a marker of this process . ProlongThe study confirmed in people with type 2 diabetes the observations obtained in animal models of diabetic neuropathy and documented for the first time that the studied polymorphism in MIR499A might affect the number of mtDNA copies, thus impairing mitochondrial biogenesis.Thus, summarizing the findings regarding the miR-499a system in diabetic neuropathy, we have evidences of (1) an association of homozygous variant genotype for the rs3746444 polymorphism of MIR499A with CAN and DPN in type 2 diabetes , (2) a rMIR155 promoter region was associated with a higher expression of this miRNA compared with the wild-type allele (adjusted p = 0.013). The rs767649 variant allele is localized in the promoter region of the MIR155 gene and might alter the binding of transcriptional factors, which are mainly inflammatory mediators and bacterial or viral-derived toll-like receptor ligands [A question might arise: are the observed polymorphisms of miRNA genes able to change the corresponding miRNA expression? In the previously cited study , in 49 p ligands  in this  ligands  and typep = 0.022). The polymorphism of MIR128A had been previously associated in a larger group of patients with type 2 diabetes with a higher risk of developing DPN [Moreover, the rs11888095 SNP polymorphism in MIR128A was associated with higher expression level of miR-128a (adjusted ping DPN , thus thIn our study, we failed to observe an association of the polymorphism in the rs3746444 polymorphism of MIR499A and the miR-499a expression. It is possible that other mechanisms contribute to changing the expression of this miRNA or that the variant allele might be involved in changing the miRNA targets rather than in regulating its expression.Despite the small size of the studied population, this genotype-phenotype association study has shown for at least two miRNA genes that the genetic polymorphism corresponds to changes in miRNA expression with a consistency between what observed for the variant alleles and expression in their association with diabetic neuropathy. These results link the genetics and epigenetics of miRNAs and support their potential usefulness as exploratory predictive biomarkers and therapeutic targets.The genetics of diabetic neuropathy is starting to offer results towards the achievement of biomarkers of risk and pathogenetic mechanisms, to provide support to known hyperglycemia-related mechanisms and to suggest new pathogenetic pathways and to validate therapeutic targets.Genetic information might be integrated into a multibiomarker approach including clinical, metabolic, and imaging phenotyping to define individual susceptibility to diabetic neuropathy and its therapeutic options.Genetic research is challenging and requires a joint force strategy to improve sample size and phenotyping. It is worthwhile proceeding, and the results in some fields might be nearer to our goals than we suspect.In this scenario, the genetics of microRNAs is at a very preliminary stage. Notwithstanding, the study of genetics and epigenetics of miRNAs may well contribute to the identification of exploratory biomarkers of risk and pathogenetic mechanisms of diabetic neuropathy and to a comprehensive definition of susceptibility to specific pathogenetic mechanisms to favor a tailored mechanism-based treatment or prevention. The data presented here would appear to anticipate a promising development.


A Body Shape Index (ABSI) is a recently proposed index for standardizing waist circumference (WC) for body mass index (BMI) and height in adults, using 2/3 and 1/2 as scaling exponents, respectively. However, ABSI has limited applicability to children and adolescents, as the relationship between height and weight changes with age and varies according to sex. This study aimed to investigate whether ABSI can be applied to adolescents and to analyze the relationships among BMI, WC, height, weight, and body shape index (BSI) in Korean adolescents. The data of 1023 adolescents aged 10–19 years from the 2009–2012 Korea National Health and Nutrition Examination Survey were collected. Body measurements  were analyzed to estimate the BSI using log-linear regression. The scaling exponents for standardizing WC for weight and height were estimated according to age (per year) and sex. The scaling exponents for standardizing WC for weight and height were 0.698 and −1.090 for boys and 0.646 and −0.855 for girls, respectively. The exponents also differed according to age. BSI was negatively correlated with height, weight, and BMI in boys and girls, and these correlations differed in direction from those in adults. ABSI cannot be applied to adolescents. In adolescents, the BSI is dependent on age and sex and is associated with growth and puberty. Further studies are required to evaluate the association between BSI and other biomarkers, to improve its applicability as a parameter for predicting the risk of chronic diseases in adolescents. The prevalence of obesity among children and adolescents has increased worldwide . PediatrAn increasing prevalence of overweight and obesity in the pediatric population has been reported worldwide ,11. The Body mass index (BMI) has been considered an indicator of body fatness and is widely used as a screening method for obesity . HoweverHowever, in another study, ABSI unexpectedly showed a negative association with blood pressure in Portuguese adolescents aged 10–17 years . TherefoWe hypothesized that ABSI obtained from adults cannot be applied to adolescents and examined the age- and sex-specific body shape index (BSI) in adolescents. The aim of the present study was to estimate the scaling exponents for standardizing WC for weight and height in Korean adolescents and to analyze the relationships among BMI, WC, height, weight, and BSI.This study was performed using data from the 2009–2012 KNHANES. The KNHANES is a nationwide cross-sectional survey conducted annually by the Korea Centers for Disease Control and Prevention . Its tar2).Body measurements, including height, weight, and WC, were obtained by trained health technicians following standardized procedures. Height was measured to the nearest 0.1 cm with a wall-mounted stadiometer. Weight was measured, with the participants in light clothing, to the nearest 0.1 kg on a digital scale. WC was measured using a steel tape, at the midpoint between the lower border of the rib cage and the iliac crest, to the nearest 0.1 cm. BMI was defined as weight (kg) divided by height squared (mWe performed linear least-squares regression on log (WC) as a function of log (weight) and log (height). The coefficients from log-log regression were the scaling ex-ponents for weight and height that were used to calculate the BSI, as follows:While ABSI was developed by Krakauer and Krakauer based on the data of adults from the NHANES, BSI was determined as an adapted formula for adolescents by age based on the data of adolescents to estimate new scaling exponents for standardizing WC for weight and height. To verify our hypothesis, we applied ABSI to our study population, and compared it with BSI.t-test. Linear least-squares regression models were constructed with log (WC) as a function of log (height) and log (weight). Pearson’s correlation coefficients were used to estimate the associations among the different variables . In correlations analysis, the statistical tests were two-sided, with an alpha level of 0.05. All analyses were conducted using R software (version 3.5.3). Scatter plots for the study variables were created using MATLAB R2020a software.Descriptive analyses were performed to characterize the samples. The normality of the distribution of each variable was tested using the Shapiro–Wilk test, and logarithmic transformation was used when needed. Mean values were compared using an independent A total of 1023 adolescents was included in the analysis. 2 = 0.842). The estimated exponents for weight and height were 0.698 and -1.090 for boys (R2 = 0.876) and 0.646 and −0.855 for girls (R2 = 0.794), respectively (1 and b2 for BSI also varied according to age in adolescents (In the 1023 adolescents, the BSI was calculated using the above equations , with 0.ectively . The estlescents .The sex-specific mean BSI showed a different pattern . The meaThe relationships among height, weight, and WC according to age in boys and girls are presented in However, in a subgroup analysis according to age and sex, BSI had a positive correlation with WC in all ages but showed no correlation with height, weight, and BMI . On the We estimated scaling exponents for weight and height to calculate the BSI in adolescents using data from the KNHANES. The exponents differed according to age and sex. The BSI in adolescents is age and sex dependent and is associated with growth and puberty.An increase in pediatric obesity is a global health issue ,10,11,18BMI is a widely accepted measure of general obesity; however, it does not distinguish between muscle and fat accumulation and does not provide information on fat distribution ,16. TherA Portuguese study showed a negative correlation between ABSI and systolic and diastolic blood pressure, and this negative correlation may be due to the application of inappropriate scaling exponents in adolescents . TherefoIn previous studies, the mean BSI steadily increased from midlife to old age in adults ,14. ConvIn our present study, the scaling exponent for height in Korean adolescents was higher than that in American and Korean adults ,14. ThisThis study had some limitations. First, we estimated the scaling exponents for BSI only in the Korean adolescent population. Thus, the results should be confirmed in pediatric groups of other ethnicities. Second, we could not investigate the relationship between BSI and obesity-related mortality or morbidity in this study. In addition, we could not differentiate between fat mass and fat-free mass and could not evaluate the correlation between BSI and muscle mass.The ABSI used in the adult population cannot be applied to adolescents. BSI in adolescents is age and sex dependent and is associated with growth and puberty. Further studies are required to confirm our results in other ethnic groups of different ages, and to evaluate the association between BSI and other biomarkers to improve its applicability as a parameter for predicting the risk of chronic diseases in adolescents.


Background: Data on the existing evidence for the association between blood retinol and transient ischemic attack (TIA)/stroke risk are limited, and the results are inconclusive. This study aimed to further assess the associations between the blood retinol levels and the risk of TIA/stroke after controlling the lifestyle factors and age-related confounders.Methods: The cross-sectional data from 1,113 individuals (aged 34–84 years old) were obtained from the Midlife in the United States (MIDUS) study. The multivariable analyses were performed to investigate the association of blood retinol levels with ever and currently TIA/stroke.Results: There was an inverse association between the blood retinol levels and the risk of ever TIA or stroke  and currently diagnosed TIA or stroke  after controlling the lifestyle factors and age-related confounders. The significance of these associations was maintained after a sensitivity analysis and involving “ever chronic respiratory diseases” as a covariate. Moreover, the stratified analyses suggested that the inverse associations could be affected by overweight , hypertension, and diabetes.Conclusions: A significant inverse association between blood retinol and the risk of TIA or stroke was found. This inverse association did not change even after adjustment for many potential confounders. Moreover, the potential protective effect of retinol on TIA/stroke could be blunted by overweight , hypertension, and diabetes. A transient ischemic attack (TIA) or stroke is a global public health problem that leads to the second leading cause of death . HoweverBlood retinol is a lipid-soluble micronutrient involved in the processes of vision, cell differentiation and proliferation, and immune system regulation . In addiTo our knowledge, few previous studies have comprehensively examined the relationship between blood retinol and the risk of TIA/stroke. The Midlife in the United States (MIDUS) study, a large population-based cohort study of adult individuals (aged 34–84 years) that includes detailed information on the levels of blood nutrients, incidence of cardiovascular diseases (CVDs), and potential confounders, such as the lifestyle factors and age-related confounders. Hence, this study aimed to analyze the impact of blood retinol on TIA/stroke and identify the modifying factors of this impact.http://www.midus.wisc.edu/. After excluding the missing values, we included 1,113 participants for analysis, as shown in This study contains 1,113 subjects from the MIDUS II study, which is a 9-year follow-up of the MIDUS I cohort from 2004 to 2006, such as the demographic, behavioral, and social factors of an adult American population . The welZ-score. Then, a multivariate logistic regression analysis was performed. The significance levels were corrected in each model to account for the relationships among the blood retinol levels and ever TIA/stroke and currently diagnosed TIA/stroke, respectively. No adjustment was made in a Crude Model. In Model 1, the blood retinol levels were regressed on both ever TIA/stroke and currently diagnosed TIA/stroke after adjusting for age, gender, and marital status. In Model 2, hemoglobin A1c %, HOMA-IR, fasting glucose levels, fasting insulin levels, total cholesterol, triglycerides, HDL cholesterol, LDL cholesterol, creatinine, and CRP were further adjusted. The categorical variables were defined as n (%). The subjects with missing data for specific variables were removed from the analysis.In this study, SPSS 26.0  was used to perform all the analyses. All continuous variables were non-normally distributed, we represented them by using the median (interquartile range [IQR]) and standardized them by using the 2],” “current smoking,” “number of years of drinking (≥6),” “exercise frequency ≥3/week,” “heart disease,” “hypertension,” and “diabetes”. These stratified variables were used to examine whether the lifestyle factors and CVDs affect the association between the blood retinol levels and TIA/stroke. A P-value ≤ 0.05 was considered statistically significant.In addition, we then used “chronic respiratory diseases (asthma and emphysema/COPD)” as a covariate for sensitivity analysis to further clarify the impact on blood retinol and its relationship with TIA/stroke. Finally, we performed the stratified analyses using the covariates, such as “overweight : 0.90; 95% CI: 0.84–0.95) and currently diagnosed TIA/stroke decreased 13% . Similarly, with the increment of blood retinol per 1-SD increase, the incidence of ever TIA/stroke decreased 17% , and currently diagnosed TIA/stroke decreased 20% . When adjusted for model 1 , as the blood retinol increased per 1 μmol/L, the risk of ever TIA/stroke was reduced by 9%  and currently diagnosed TIA/stroke was reduced by 10% . Likewise, with the blood retinol increased every 1-SD, the risk of ever TIA/stroke was reduced by 15%  and currently diagnosed TIA/stroke was reduced by 18% . In model 2, with the increment of blood retinol per 1 μmol/L, the incidence of ever TIA/stroke decreased by 7% , and currently diagnosed TIA/stroke decreased by 9% . The incidence of ever had TIA/stroke decreased by 11%  and currently diagnosed TIA or stroke decreased by 16%  when the blood retinol increased per 1-SD.The results of the multivariate logistic regression analysis for the association between the blood retinol levels, and ever and currently diagnosed TIA/stroke are shown in ORs were little changed by further adjustment for “ever chronic respiratory diseases” in the Crude Model, as well as in the model 1 and model 2 . These results further suggested that the significant and independent associations between the blood retinol levels and ever TIA/stroke and currently diagnosed TIA/stroke.The sensitivity analyses  showed tP < 0.05), but not decreased in the individuals with the three covariates . However, our results showed that the association between the blood retinol and TIA/stroke were not affected by the current smoking status, number of drinking years, frequency of exercises ≥3/week, and heart disease. We first found that overweight, hypertension, and diabetes were modifiers that affect the association between blood retinol and ever TIA/stroke and currently diagnosed stroke/TIA.We additionally performed the stratified analyses to assess the effect of blood retinol (per 1 μmol/L and 1-SD increment) on ever TIA/stroke and currently diagnosed TIA/stroke by adding several factors of lifestyle and CVDs as a covariate, respectively . Our resWe found that the increased blood retinol levels were associated with a reduced risk of TIA/stroke. This dependent association did change little after the adjustments for many lifestyle factors and age-related confounders. The potential protective effect of retinol on TIA/stroke can be attenuated by overweight, hypertension, and diabetes.As early as 1992, it was reported that the patients with ischemic stroke with the blood retinol concentrations above 2.27 μmol/L had increased rates of recovery and decreased mortality . In contOR: 0.92; 95% CI: 0.86, 0.97) and first ischemic stroke , and homocysteine was a modifier factor for the association. However, the participants in this study were all patients with hypertension, and the results may not apply to the generality of other populations  at the University of California-Los Angeles, Wisconsin, or Georgetown. The patients/participants provided their written informed consent to participate in this study.KH was incharge of the entire project and revised the draft of the manuscript. LG and YH analyzed the data. LG drafted the first version of the manuscript. All authors took part in the interpretation of the results and in the preparation of the final version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


This study investigates the reliability and precision of anthropometric measurements collected from 3D images and acquired under different conditions of head rotation. Various sources of error were examined, and the equivalence between craniofacial data generated from alternative head positions was assessed. 3D captures of a mannequin head were obtained with a stereophotogrammetric system (Face Shape 3D MaxiLine). Image acquisition was performed with no rotations and with various pitch, roll, and yaw angulations. On 3D images, 14 linear distances were measured. Various indices were used to quantify error magnitude, among them the acquisition error, the mean and the maximum intra- and inter-operator measurement error, repeatability and reproducibility error, the standard deviation, and the standard error of errors. Two one-sided tests (TOST) were performed to assess the equivalence between measurements recorded in different head angulations. The maximum intra-operator error was very low (0.336 mm), closely followed by the acquisition error (0.496 mm). The maximum inter-operator error was 0.532 mm, and the highest degree of error was found in reproducibility (0.890 mm). Anthropometric measurements from alternative acquisition conditions resulted in significantly equivalent TOST, with the exception of Zygion (l)–Tragion (l) and Cheek (l)–Tragion (l) distances measured with pitch angulation compared to no rotation position. Face Shape 3D Maxiline has sufficient accuracy for orthodontic and surgical use. Precision was not altered by head orientation, making the acquisition simpler and not constrained to a critical precision as in 2D photographs. The quantitative analysis of the human face has always received large attention from both scientists and artists . QualitaAnthropometric analysis of soft tissues is an integral part of orthodontic diagnosis along with therapeutic planning, and two-dimensional (2D) photography has been, for years, one of the main devices for the analysis of facial measurements. Despite this, such a technique carries considerable limitations. It can reproduce reality but only in two dimensions, leaving out valuable information about the depth and the transversal dimension of a face. Moreover, in 2D the position of the patient inside the framework of the image is critical. Any error in the vertical, horizontal, or rotational positioning of the face during the shot creates distortions, making images practically useless. A 3D photograph, on the contrary, can be shot even in the presence of less than ideal positioning, which can then be easily corrected on the computer. The 3D image, therefore, is not only a general improvement of 2D, but a means that provides a completely new concept of the image, by being able to perform measurements and overlaps and, thus, making objective what was previously only a clinical impression. In addition, stereophotogrammetry provides an accurate assessment of the aesthetic outcome of the therapy, which, being not invasive, can also be repeated with frequent surveys over time. Moreover, even if clinicians and researchers are still discussing how it may be possible to reach a reproducible natural head position (NHP), the smoothest variations on the positioning of the patient’s head during a 2D photography session could unavoidably alter results within a single assessment or time-to-time comparisons. Interest in overcoming the limitations of direct measurements and of 2D photogrammetry has led to the development of numerous non-invasive methods for capturing and quantifying craniofacial surface morphology . Today, we have sophisticated digital scanning devices that complete our evaluations with third-dimension (3D) data, and even more recently the fourth dimension (4D), that includes movement ,7,8. In Chung et al.  give an The goals of this study are (1) to validate the present digital 3D photogrammetric device in terms of measurement error; (2) to compare craniofacial measurements obtained from 3D images which were generated from alternative captures of a mannequin head with different degree of yaw, pitch, and roll rotations. The hypothesis is that anthropometric measures recorded in different conditions of head orientation are equivalent to each other. The scanner we used was Face Shape 3D Maxi Line, developed by Polishape 3D Srl . This photogrammetric system counted six Canon EOS 1100D  reflex cameras, each one fixed on rigid support with a specific inclination. Two lateral and external professional flashes were used to minimize any possible external light distortions and to obtain uniform light on the surface of the object. ® .After the required calibration procedure, the subject could be shot and the corresponding 3D image created by a specific software by a 3D rendering function; in this study, we used Viewbox 0.48 median: Glabella (Gb), Nasion (N), Pronasale (Prn), Subnasale (Sn), Labiale superius (Ls), Labiale inferius (Li), Sublabiale (Sl), and Pogonion (Pg);7 bilateral: Frontotemporalis (Ft r/l), Zygion (Zy r/l), Tragion (Tr r/l), Gonion (Gn r/l), Cheek (Ch r/l), Cheilion (Chel r/l), and Orbitale inferius (Or inf r/l).We used a mannequin head as a subject, which is ideal for in vitro experimentation due to immobility and absence of facial mimicry. To improve image acquisition, the texture of the dummy was faded and opaque to reduce light reflection, and, as suggested by the literature data , the folThe dummy was placed on external support equipped with a graduated scale that allowed the operator to orientate the dummy on the three planes of the space with extreme control on specific angular rotational values during the pitch, roll, and yaw movements .To investigate the accuracy and precision of the system, a series of captures was taken with the mannequin head with no rotation (reference position) and with various pitch, roll, and yaw angulations. The protocol of image acquisition is reported in x, y, and z coordinates were collected from the software and saved in an Excel spreadsheet. Next, all shots were processed, and the anthropometric points previously marked on the dummy were also marked on the three-dimensional reconstructions obtained. Their 4 median: Chel(r)-Chel(l), N-Prn, Sn-Pg, and N-Pg;5 bilateral: Glab-Ch(r), Glab-Ch(l), Glab-Ft(r), Glab-Ft(l), Zy(l)-Tr(l), Zy(r)-Tr(r), Ch(r)-Tr(r), Ch(l)-Tr(l), Ch(r)-Gn(r), and Ch(l)-Gn(l).The following 14 linear distances have been calculated : 4 mediaThe 14 measurements were chosen to cover various facial regions, having different size and orientation on the transverse, frontal, and sagittal plane. The Euclidean distance between two landmarks has been calculated as the square root of the sum of the differences in the three dimensions of the space, as indicated in the following formula: articles ,14. Errors may be introduced during imaging acquisition, placement of landmarks on the images, or calculation of distances. To estimate the relative contributions of these sources of errors, the precision of the system was investigated in terms of repeatability  and reproducibility . Notably, repeatability included intra-operator, inter-operator, and acquisition errors.All the investigators were orthodontists, with at least 20 years of clinical experience in recognizing anthropometric points.To assess intra-operator error, the same investigator repeated (10 times) the placement of anthropometric landmarks and measurements of linear distances on the same 3D reconstruction taken in the reference position. All parameters were measured again by a second investigator on the previous acquisition and compared (inter-operator error). Acquisition error was assessed by measuring, using the same operator, the selected parameters of 5 different 3D image captures in reference position. To investigate reproducibility error, we compared measurements performed by the two operators on different acquisitions. To avoid recall bias, a minimum of 24 h was allowed to elapse between measurement sessions.To assess how the head position might affect the accuracy of anthropometric measurements, each set of linear measurements of yaw, roll, and pitch was compared with data obtained from the reference position, and the mean value of the linear measurements derived from each set of 3D images taken with yaw, roll, and pitch angles was compared to the mean of those obtained in the reference position. Statistical analysis was performed using various software, among them the open-source statistical software Jamovi , which iTo quantify measurement error magnitude, the following measurement error indices were calculated: mean error, maximum error, standard deviation (of errors), standard error (of errors), and coefficient of variation (CV).p < 0.05). Bonferroni’s correction was applied for multiple comparisons.To demonstrate the equivalence between measurements obtained in alternative head positions, the mean difference between repeated measures was calculated, and the Westlake–Schuirmann two one-sided test (TOST) was used ,18. The The mean of standard errors was 0.041 mm, with a minimum value of 0.023 mm in the Glabella–Frontotemporal (l) measurement and a maximum value of 0.081 mm in the Nasion–Pogonion measurement; such a measurement also reached the highest maximum error (0.336 mm). The inter-operator standard error was higher than the intra-operator value with a mean value of 0.064 mm, a minimum value of 0.04 mm in the Chelion–Gonion (r) measurement, and a maximum value of 0.111 mm in the Nasion–Pogonion measurement, whose maximum error was found to be 0.532 mm. The mean standard error of acquisition was 0.103 mm with a minimum value of 0.066 mm for Glabella–Frontotemporale (r), and a maximum value of 0.177 mm for Zygion (r)– Tragion (r). The highest maximum error was found in the Cheek (l)–Tragion (l) measurement of 0.496 mm. The mean standard error was 0.086 mm with a distribution of values between 0.032 mm for Glabella–Frontotemporale (r) and 0.164 mm for Cheek (l)–Tragion (l). The latter is also characterized by the largest value of the maximum recorded error of 0.890 mm. The error magnitude statistics are reported in The higher degree of maximum error was found in reproducibility (0.890 mm), which combined both the error due to digitization and the imaging system.The statistical equivalence between anthropometric measurements coming from the alternative acquisition conditions was calculated considering an equivalence limit of 0.89 mm, which represented the maximum error of the measurement system at the reference position.The mean difference between linear measures and equivalence data is reported in p <0.01). The highest mean difference was 0.422 mm for the Cheek (l)–Tragion (l) measurement.Yaw versus reference position: the two sets of measurements were found to be statistically equivalent . The maximum difference between the averages was 0.543 mm calculated for Zygion (l)–Tragion (l).Roll versus reference position: the two series of measurements were found to be statistically equivalent  with the exception of two distances, Zygion (l)–Tragion (l) (p = 0.510) and Cheek (l)–Tragion (l) (p = 0.166). The maximum difference between the averages was 0.814 mm calculated for Zygion (l)–Tragion (l).Pitch versus reference position: the two sets of measurements were statistically equivalent .Intra-operator maximum error was very low (0.334 mm), and acquisition error and inter-operator error were very similar (0.496 mm/0.532 mm). As expected, the maximum error of reproducibility was the highest (0.89 mm). Reproducibility refers to the variation of measurements made under changing conditions, which in the present experiment were due to measurements being made by two operators on acquisitions with different head angulations; thus, such a value combines various types of errors owing both to digitization and to the imaging system itself. Lateral regions, along with interoperability variability, were sources of greater acquisition and reproducibility errors, especially in measurements involving the Tragion area.In the context of medical facial treatment, a patient’s photographs represent an extremely important datum both for the follow-up and simple clinical documentation. So far, in 2D photographs it has always been the practice to search the NHP of the patient’s head as a reference position. Despite that, among the scientific community, there is still discussion on how to reach the NHP and whether it can be reproducible.Cassi et al. , in a reOn the other hand, although the NHP has less variability than intracranial reference lines, it is also influenced by balance, vision, and proprioception from joints and muscle involved in maintaining erect posture. Therefore, it depends on the subject’s neuromuscular condition as well, and it may be difficult to obtain in some patients, especially children, and subjects with neuromuscular disorders, vertebral column deformity, and alterations in eye muscles balancing .In the literature, some protocols for obtaining the NHP might influence reproducibility, and there is also some evidence that the success might depend on the operator . To someIn light of this, since the facial scanning system is continuously being developed as a valid alternative to 2D photography, we wanted to investigate if head rotations on the three spatial planes could represent any critical aspects in terms of reproducibility and precision, with notable implications on the success of the results.We can accordingly conclude that the position of the dummy does not influence the precision, accuracy, and repeatability of anthropometric measurements, at least within 16 degrees on right and left for yaw and roll, and within 11 degrees upward and downward for pitch.These results agree with data from studies conducted with other stereophotogrammetric systems. In fact, Lubber et al. stated that by progressively changing the spatial orientation of a dummy , the mean error of measurements on the corresponding 3D image remained very low and steady within the central range of movement (TRE: 0.195 mm), which then showed very little increases along with large spatial variations of the dummy . MoreoveIn conclusion, we can state that the Face Shape 3D Maxi Line has sufficient accuracy for orthodontic and surgical use, especially in the median areas of the face. Despite the presence of areas of non-equivalence , the differences are clinically acceptable. Considering the magnitude of the intra-operator and inter-operator errors, which represent a significant proportion of the total error, we might suppose also that acting on them and the learning curve might reduce the system error.Based on the results, we might assert that such precision is not altered by the rotation of the head on three planes of space, making the acquisition process even simpler and less constrained to a predetermined position, which is not always easy to obtain with all patients. This improves the comparison of acquisitions obtained at different times and conditions, facilitating the clinician during long-term treatment.The mannequin head represented an ideal object, as it did not move or perform facial expressions. This allowed it to reach a very high degree of precision, without the influence of human variability on the photographed subject. On the other hand, the estimates of precision and accuracy might be inflated by the experimental setting, and the lack of a human sample is a limitation to the study, namely not being able to assess the effect of the stretching of the soft tissues in the precision of the measurement.Further studies are needed to confirm the results obtained in vitro by repeating the study in vivo. Living subjects, as opposed to inanimate mannequin heads, may be affected by motion artefacts such as breathing or swallowing, and thickness and soft tissue characteristics, also related to different age ranges, might influence the measurement error of 3D surface imaging systems.