This paper describes the rationale, intervention development, study design and results from the pilot feasibility study of the Keep On Running (KOR) trial. KOR aims to test a web-based brief theory-based intervention, targeting maintenance of recreational running behavior over time (i.e. relapse preventing).n=18) to measure intervention adherence and participant satisfaction in order to establish the feasibility and acceptability of the intervention toolkit. Furthermore, this pilot study was also used to test the feasibility and acceptability of the questionnaires selected to be part of the later RCT.Intervention development was based both on Self-Determination Theory and on Self-Regulation Theory. As part of it, a pilot study was implemented . The protocol of the RCT was adjusted to test the efficacy of the refined final version of the intervention, and the RCT that will test it, contributing to the understanding of recreational running sustainability, allowing the optimization of future interventions aimed at physical activity promotion. While benefits of regular physical activity (PA) and exercise are well established   are available world-wide for smartphones and represent a unique opportunity to reach a broad audience of users. The most popular apps are for exercise, diet, and weight management, and 500 million users use mobile health applications. Mobile health interventions have surged in popularity but their implementation varies widely, and evidence of effectiveness is mixed  trial. KOR aims to test a brief theory-based intervention, delivered through digital technology (web app), targeting maintenance of recreational running behavior over time (i.e. relapse prevention) after being voluntarily initiated in the previous 3 months. The pilot study was focused on intervention adherence and participant satisfaction in order to establish the feasibility and acceptability of the intervention toolkit. Furthermore, it was also used to test the feasibility and acceptability of the questionnaires selected to be part of the later RCT.The study's objective is to develop and present a new digital toolkit aiming at maintenance of recreational running behavior over time (i.e. relapse preventing), and evaluate whether it can be delivered and accepted by participants. We hypothesize that the built intervention can be delivered and will be accepted by the participants, and tested by the RCT.KOR is a Randomized controlled trial (RCT). It was developed based on a logic model , adaptinAs a result, it comprises four different groups : (i) a gThis approach was taken because (a) comparing runners belonging to PNMC with FRR (i.e. control group) allows examining the effect of the PNMC on behavior maintenance; (b) comparing FRR enrolled in the SRM intervention with FRR allows testing the effectiveness of a light-touch intervention on motivational and other psychological variables, and on behavior maintenance; and (c) comparing runners belonging to PNMC and receiving the SRM intervention with FRR receiving the SRM intervention will allows examining whether the SRM intervention has an additional effect on the study outcomes. Trial duration is one year with assessments at 6 weeks, and 6 and 12 months postbaseline .Figure Inclusion criteria: age between 18 and 65 years; running for no longer than 3 months; performing a minimum of 60 min of vigorous physical activity per week; being free from major orthopedic limitations ); and from clinical diagnosis with any condition that may interfere with performing MVPA; being free from untreated or major psychological disorders ; and possessing or willing to acquire a smartphone or computer with internet access.Exclusion criteria: inability to provide written informed consent; inability to follow written material or telephone conversations in Portuguese that would preclude completion of study questionnaires and use of the digital intervention ; intention of pregnancy in the next 12 months; planning to travel for more than 2 weeks in the next 12 months or to emigrate; suffering any injury in the course of the study.We developed a web-based intervention adapted from the motivational and self-regulation arm of the NoHoW tria , the questionnaires platform to design interactive features, and whiteboard drawings to convey some of the more theoretical constructs of the intervention (http://www.videoscribe.co/), were used to develop these implementations, supported by the team's previous experience in similar interventions.Access to a web-based intervention comprising 11 distinct sessions of 5–10 min each and 8 educational modules concerning motivation and self-regulation constructs, delivered through several technological implementations, including videos, animations, quizzes and exercises, as well as images, text and audio . SeveralBi-weekly text messages  will be recruited from the community . To recruit this sample, we will use online advertisement and running events list servers, reach out for running clubs, and partnerships with both state and private organizations.We will use a rolling recruitment strategy: participants will join the study as soon as they are considered eligible to facilitate management of available human resources. Each set of runners will be randomly assigned to the KoR intervention group or the control group general information group in a 1:1 ratio, stratified by group (PNMC and FRR). The method of randomized permuted blocks will be used, with random block lengths (4 or 6) Broglio, . BecauseData on demographics  and potential psychological moderators (i.e. index of autonomous functioning) will be collected only at baseline.The primary outcome of this study is running behavior, which will be measured at baseline, intervention's end and 6-month follow-up by self-report (frequency and minutes of running per week). At 12 months, participants will be asked by telephone whether they keep running (at least) at their initial level, or not.Secondary outcomes will include putative mediators and outcomes. Basic psychologic need satisfaction will be assessed via the Psychological Need Satisfaction in Exercise Scale , 6 months, and 12 months follow-up. The training and clarification session will be completed by telephone to ensure participants are equipped with the skills to access the intervention contents. Before the first, third and fourth assessment periods, eligibility will be checked against inclusion and exclusion criteria through a telephone screening/re-screening call, 1 week prior to each visit. Participant's timeline is shown in Assessment 1 (baseline): Clarification session about the study. If participants are still interested in enrolling in the study, eligibility will be checked, and they will be asked to fill out an informed consent form. Randomization and allocation to one of the four arms of the trial will be performed. Data on demographics and potential psychological moderators will be collected. A battery of psychological and behavioral questionnaires will be completed online using google forms (in one-week time) – running behavior (i.e. frequency and minutes per week), motivations, basic psychological needs, motives and gains, self-regulation skills, passion, flow, and vitality. This task is estimated to take approximately 1 h, but participants will be able to do breaks and return later to complete the psychometric evaluation during that week. Debrief on how to use the KOR web-based intervention content – the toolkit (approximately 1 h).Assessments 2 and 3 (intervention's end and 6-month follow-up). Email reminder prior to this evaluation period will be sent. The same battery of psychological and behavioral questionnaires will be completed online using google forms (in one-week time) – running behavior (i.e. frequency and minutes per week), motivations, basic psychological needs, motives and gains, self-regulation skills, passion, flow, and vitality –, and will take approximately 1 h.Assessment 4 (12-month follow-up). Running behavior maintenance will be assessed through a telephone call, asking participants whether they keep running at their initial level or higher, or not.p < 0.05.Statistical analysis will be performed using SPSS Statistics version 25.0 . Descriptive statistics will be calculated . Repeated measures with Bonferroni corrections for adjusted comparisons will be used to examine differences between each intervention group(PNMC or SRM) and controls , at all assessment points, in the primary and secondary outcomes measured. Statistical significance will be set at Mediation analysis, will test the psychological mechanisms underlying changes in running behavior from baseline to post program, 6 and 12 months. Multilevel modeling will be used to determine the SRM intervention effect on these motivational variables, with intervention group (SRM vs. Control group) as the between-subject factor and change in self-regulation and competence for running at baseline, 6–8 weeks, 6months and 12 months, serving as dependent variables. If necessary, multiple imputation methods will be implemented to provide robust results for primary and main secondary outcomes.d = 0.23) is expected, a value previously registered in studies with intrinsic motivation interventions  or 55 participants (p = 0.01). Concerning our secondary outcomes, we are expecting effects sizes ranging from 0.23 to 0.73, based on previous reports . Thus, recruiting 220 participants will ensure high statistical power for our primary analyses, extending its power to the secondary analyses.Two types of sample size calculations were made: (a) using the primary outcome – running minutes and (b) using secondary outcomes, in particular, changes on key psychosocial variables. Regarding our primary outcome, we have used vigorous physical activity for our calculations. A small effect size . Bearing in mind the rule of thumb for pilot sample size , between 25 and 54 years-old (average 42.1 yrs.), were generally self-perceived as healthy (94%), highly educated (78%), and lived mostly in the Lisbon area (83%). Participants reported to run an average of 27 km and 4.4 h per week. Running experience ranged from 1 month to 30 years (average 6.3 yrs.). Most participants (88.9%) wore a running monitoring device to keep track of their running, social sharing and coaching, and 94.4% included other types of exercise in their training regime, such as warm-up and stretching, weight training or calisthenics, exercise classes or biking. A total of 61.1% used social media for motivation, education, interaction and coaching purposes.Participants , five , and only three participants (16.7%) completed sessions from eight to eleven. During the course of the program 12 participants (66.7%) logged at least one race , and the average run lengthened 9.4 Km (4–14 Km).From the 18 participants allocated to the intervention, 17 (94.4%) logged at least once, 50% logged once, and 22.2% twice. Only 6 (33.3%) completed the first session. Participants’ session completion decreased at sessions two  found the questionnaire too long. Participants generally found the toolkit website to be attractive and user friendly, classifying their overall experience with the study as positive. Most of the interviewed participants already used a running monitoring app, so they found the program to be less intuitive, repetitive in some questions and missing out important aspects such as ‘time of day, speed/ pace, and altimetry’. Additionally, participants reported problems with platform adjustment that led to ‘information overlap when used on cellphone’, the fact that ‘it didn't keep data inserted in previous sessions’ and ‘errors in google docs that forced them to restart the questionnaire’.Three participants suffered injuries during the intervention period (25%). Two had muscular running-related injuries in the lower limbs and one had a foot trauma injury while playing football.This pilot study had limitations that need to be considered, Due to recruitment constraints, the final sample included long time runners, which may differ from beginners, targeted by the RCT protocol. The low sample size and the absence of follow-up hindered further interpretation about the efficacy of the toolkit as it is.The study protocol was approved by the Ethics Committee of the Faculty of Human Kinetics, University of Lisbon (CEFMH 3/2018) and will be conducted in accordance with the declaration of Helsinki for human studies  and secondary outcomes related to the quality of the running experience  will be used as the dependent variables.Additionally, the results of the pilot feasibility study, intended at testing intervention adherence, participant satisfaction, and questionnaire acceptance, were also presented. Overall, the pilot study indicated that the intervention has good acceptability, feedback was very positive, but toolkit interaction decreased after the first weeks, which is common in web-based interventions , to increase data detail and prevent ‘unnecessary’ data insertion by the participants. Features such as feedback and social sharing, added to the goal setting features already integrated in the toolkit, may also increase attractiveness (Hosseinpour & Terlutter, This paper describes the rationale, the process of developing, study design, methods, and the pilot study of the KOR intervention. Valuable lessons are taken from the pilot feasibility study and adjustments will be made in the intervention and in its delivery, in order to improve adherence and overall experience. Results from the RCT will test the efficacy of this approach, contribute to the understanding of recreational running sustainability, and can be used in the development and optimization of future interventions aimed at physical activity promotion.


Ferroptosis, an iron-dependent form of regulated cell death, has drawn an increasing amount of attention since it was first mentioned in 2012 and is found to play a significant role in the treatment of certain diseases. Our study is aimed at analysing the scientific output of ferroptosis research and at driving future research into novel publications. Publications focused on ferroptosis were retrieved from the SCI-EXPANDED database of the Web of Science Core Collection and were screened according to inclusion criteria. CiteSpace V and Microsoft Excel 2016 were used to evaluate and visualize the results, including generating network maps and analysing annual publications, country, category, references and cocited references, and keywords. As of October 1, 2021, a total of 1690 original articles related to ferroptosis were included, and the overall trend of the number of publications rapidly increased. Among the common categories in the field of ferroptosis, the most common category was biochemistry and molecular biology. Worldwide, China and the United States were the leading countries for research production. The retrieved 1690 publications received 44,650 citations, with an average of 26.42 citations per paper . By citation analysis, Scott J Dixon's article in 2012 was the most symbolic reference and the earliest publication in the field of ferroptosis, with the highest citation rate (2709 times). Among the most common keywords, most were related to the mechanisms and regulatory networks of ferroptosis. Furthermore, with accumulating evidence demonstrating the role of ferroptosis in cancers and other diseases, inducing ferroptosis in clinical treatment is becoming a new research focus that should be closely monitored. Ferroptosis is a newly identified form of nonapoptotic regulated cell death characterized by iron-dependent accumulation of lipid peroxides , 2. FerrBibliometric sciences can not only offer a quantitative and statistical analysis of publications in specific areas but can also accurately uncover the most representative studies –18. In aIn this study, we chose the Science Citation Index Expanded (SCI-EXPANDED) index of the Web of Science Core Collection (WoSCC) as the source database for data retrieval. The WoS Core Collection is a canonical online database providing a standardized and up-to-date dataset of reference for scientific research and analysis, among which SCI-EXPANDED is considered to be the most appropriate database for bibliometric analysis , 21. Datg articles received (together) at least g2 citations [The retrieved data were downloaded and exported into different formats for further analysis. Two authors performed the data extraction process independently. Information including titles, authors, institutions, abstracts, countries, source journals, total number of citations, publication year, keywords, and g-index was identified and extracted. The g-index is introduced as an improvement in the h-index, both of which are measures of productivity and the impact of a researcher . If a seitations , 24.Visualization plays a significant role in bibliometric analysis because the collaborative networks, research hotspots, and trends of a new field can be intuitively observed through visualization maps. In this study, data analysis was performed using Microsoft Office Excel 2016 and CiteSpace v5.7.R2. Microsoft Office Excel 2016 was applied to manage the database and analyse the annual publication output and trend of ferroptosis publications. CiteSpace is a free software for visualizing data and analysing trends and patterns of the identified scientific literature as well as exploring the significant achievements, key research subjects, and outstanding researchers, institutions, and countries .k = 25), pruning (pathfinder + pruning sliced network + pruning the merged network); and the term source and node types are flexible parameters according to specific needs. Analyses of the methods included cooccurrence, clustering, and burst. The visualization diagram types included histograms, cluster views, timelines, and collinearity. All the clusters were labelled by keywords, and the log-likelihood rate (LLR) was used as the clustering algorithm. Synonyms were combined in the obtained analysis map according to the actual situation.The retrieved data were put into the CiteSpace software to produce visualized maps to analyse the annual output counts, countries, top-cited references, keywords, and burst detection. Moreover, cocitations and coreferences were visualized to study cooperative relationships between identified publications. The parameters of CiteSpace were set as follows: time slicing (from 2012-01-01 to 2021-10-01), years per slice (1), links ; selection criteria .The retrieved 1690 documents received 44,650 citations in total, with an average of 26.42 citations per paper and an h-index of 100 (October 8th). The top 10 research articles related to ferroptosis citations are shown in decreasing order in Supplementary Table Cocitation analysis can help researchers find the common knowledge basis efficiently and conveniently among many studies , 28. We Cell by Wan Seok Yang et al. in 2014, followed by Scott J Dixon, 2012 (strength = 72.44), and Jose Pedro Friedmann Angeli, 2014 (strength = 65.71).Furthermore, we explored the cocited references with strong citation bursts by using the CiteSpace software. References with citation bursts are defined as those cited frequently over a timespan. The top 10 cocited references with the strongest citation bursts are listed in ascending order according to the starting year of burst in Supplementary Table Q score is 0.8884, while the mean silhouette value is 0.972, indicating that the clustering structure is stable and highly convincing. Notably, the Q/S value of the cocitation analysis is even higher than that of the coauthor analysis. The cluster nomenclature reflects the study frontiers in a certain field. In this study, the clusters were autogenerated and labelled by the log-likelihood ratio (LLR) algorithm. The largest cluster was #0, labelled “mitochondria,” followed by “IncRNA” (cluster #1), “prognosis” (cluster #2), and “heme oxygenase-1” (cluster #3). Other important clusters were “autophagy,” “hepcidin,” and “intracerebral haemorrhage,” which may represent a turn in the significance.We also constructed a visualized map for clustering the cocited references with the aid of the CiteSpace clustering function. The retrieved documents were divided into 19 clusters through clustering analysis, as shown in k = 25 to k = 5 to obtain a better view of the distribution of keywords. As a result, a total of 100 keywords extracted from the title and author keywords of all 1690 publications were visualized and analysed with 110 links. As shown in Keywords are the core content of a document . Using tCluster analysis can demonstrate the knowledge structure of a certain field . ClusterKeyword burst detection refers to detecting keywords with a high frequency of appearance in a certain period of time, which helps researchers analyse the evolution of ferroptosis research. In this paper, the time period was set from 2012-01 (since the concept of ferroptosis was coined in 2012) to the date when we performed the analysis. Through the burst test, the seven keywords identified with the strongest citation burst are shown in Supplementary Table In general cancer cells had the strongest burst strength (strength = 5.18) followed by iron (strength = 3.58) and ferroptotic cell death (strength = 3.52). Dixon et al. introduced the concept of ferroptosis in 2012. Since then, ferroptosis has drawn increasing interest from scientific researchers. Iron dyshomeostasis is one the characteristics of ferroptosis. As an important inducing factor of ferroptosis, iron (strength = 3.58) was considered from 2015 to 2018. Cancer cells (strength = 5.18) were also consistently considered at almost the same time (from 2016 to 2019). The term ferroptotic cell death (strength = 3.52) had a citation burst during a short period from 2016 to 2017, while glutathione peroxidase (strength = 2.56) had a citation burst from 2016 to 2018. Cell death (strength = 2.56) became a burst word from 2017 until now. In recent years, oxidative stress (strength = 3.17) and ferroptosis induction (strength = 2.46) have become novel burst words, with citation burst years starting from 2019 to the present. These words will likely continue in the near future.The term “ferroptosis” was originally mentioned by Stockwell's group in 2012; thus, the literature was retrieved from WoS Core Collection from 2012 until now . Their sAmong the common categories in the field of ferroptosis, the most important category was biochemistry and molecular biology. With the second-highest centrality and the second-highest frequency, the cross-domain of cell biology was exceedingly extensive, suggesting that it was a breakthrough in interdisciplinary research on ferroptosis. Moreover, with relatively high centrality, pathology, oncology, biotechnology, and applied microbiology were also breakthrough points. Since 2019, chemistry/applied food science and technology have gradually appeared in the scope of scholarly research and showed a high correlation with ferroptosis. For the dual-map overlay, Among the 62 countries and regions, only four of them have issued more than 100 papers, and it is worth noting that studies published by Chinese researchers accounted for more than half of the publications. However, the earliest research on ferroptosis was initially conducted by American scholars, Dixon and Stockwell's group, who conducted correlated research earlier than scholars from other regions of the world , 31, 32.Highly cited references are classic studies that mainly address the mechanisms and potential pathways of ferroptosis. However, as research continues, new studies focusing on the regulation and manipulation of ferroptosis in human diseases are inevitably increasing in this field, for example, Kagan et al.'s publication and Sun et al.'s study , 34.In Supplementary Table Furthermore, the cocited reference cluster analysis in Based on the existing knowledge, ferroptosis is a novel cell death pathway of nonapoptotic regulated necrosis . The mosAccumulating evidence has demonstrated the contribution of ferroptosis to many organ injuries and neurodegenerative disorders. The most common type of organ injury is ischaemia/reperfusion injury of the kidney and heart –40. The As shown in the burst detection results, oxidative stress and ferroptosis induction are the latest burst terms in recent years, which suggest that these are the current research hotspots in this field. In recent years, studies of ferroptosis have extended to various aspects. In the first ten months in 2021, there have already been over 200 studies published in well-known journals. However, it did not become a burst term of ferroptosis in the study, similar to many other subfields in the ferroptosis study. Two possible reasons may contribute to the result: on the one hand, the research network has been broadened by researchers to a variety of research areas instead of several specific aspects; on the other hand, new publications still need time to be found and cited to form a new hotspot.Combined with the results of keyword cooccurrence analysis and burst word detection, early studies mainly focused on the mechanism of ferroptosis, while in recent years, its clinical applications have become increasingly attractive. Thus, we can conclude that exploration of mechanisms has always been the mainstream of ferroptosis research, interpretation of ferroptosis in human diseases, and the application in tumour suppression are the hotspots and newly emerging research directions.In recent years, the involvement of ferroptosis has been implicated in the occurrence and development of multiple human diseases; however, correlated studies are still very limited . FerroptIn summary, ferroptosis research is in a rapid development stage. Annual publications on ferroptosis have been continually increasing since 2012, especially in the last three years. The most common category correlated with ferroptosis is biochemistry and molecular biology. China has made the most contribution to this field, while Germany has also played a significant role, with high centrality. Since ferroptosis is a newly discovered cell death modality, a majority of the keywords with the highest frequency in the field of ferroptosis are related to the mechanisms or pathways of ferroptosis. “Cancer cell” is the term with the strongest citation bursts, while “oxidative stress” and “inducing ferroptosis” are the latest two burst terms in recent years. With accumulating evidence demonstrating ferroptosis-related genes in the pathogenesis of multiple diseases, manipulation of ferroptosis-related genes (otherwise known as gene therapy) in clinical cancer treatment may soon become a new hotspot that should be closely monitored. Overall, compared to traditional reviews, we believe that the results of this study will provide objective insight into further research.


Sirenomelia, also called mermaid syndrome, is a rare lethal multi-system congenital deformity with an incidence of one in 60,000–70,000 pregnancies. Sirenomelia is mainly characterized by the fusion of lower limbs and is widely associated with severe urogenital and gastrointestinal malformations. The presence of a single umbilical artery derived from the vitelline artery is the main anatomical feature distinguishing sirenomelia from caudal regression syndrome. First-trimester diagnosis of this disorder and induced abortion may be the safest medical option. In this report, two cases of sirenomelia that occurred in an white family will be discussed.We report two white cases of sirenomelia occurring in a 31-year-old multigravid pregnant woman. In the first pregnancy (18 weeks of gestation) abortion was performed, but in the third pregnancy (32 weeks) the stillborn baby was delivered by spontaneous vaginal birth. In the second and fourth pregnancies, however, she gave birth to normal babies. Three-dimensional ultrasound imaging showed fusion of the lower limbs. Neither she nor any member of her family had a history of diabetes. In terms of other risk factors, she had no history of exposure to teratogenic agents during her pregnancy. Also, her marriage was non-consanguineous.This report suggests the existence of a genetic background in this mother with a Mendelian inheritance pattern of 50% second-generation incidence in her offspring. Sirenomelia is a rare congenital anomaly in which the most noticeable feature is fusion of the lower limbs, resulting in the appearance of a mermaid's tail, and thus the name mermaid syndrome. This syndrome is characterized by severe urogenital abnormalities and the presence of a singular umbilical cord blood vessel. Sirenomelia or sirenomelia sequence was originally described by Rocheus in 1542 [Sirenomelia is a lethal disorder with a rare chance of live birth, varying from 0.1:10,000 to 0.47:10,000 based on different reports . BecauseSirenomelia can be classified into several categories based on the wide variety of limb malformation phenotypes. Saint-Hilaire and Forster classified sirenomelia based on the number of feet. The other widely used classification is the Stocker and Heifetz method, which has seven types (I–VII) and is based on the presence or absence of the femur, tibia, and fibula Fig.  2, 5, 6, 65, 6. The etiology of sirenomelia remains unclear, and most cases are sporadic, with very little chance of incidence in subsequent pregnancies. However, the case of this study is unique as it reports two occurrences of sirenomelia in one family.A multigravid 31-year-old white woman and her 35-year-old white husband were admitted for pre-pregnancy consulting for their fourth pregnancy. In their first pregnancy, no fetal movement was detected by sonography at the 18th gestational week. Subsequently, three-dimensional ultrasound imaging showed a fetus with cystic kidneys and fusion of the lower limbs, so the pregnancy was terminated by induced abortion. The woman later gave birth to a normal baby at gravida 2, but at 32 weeks of her gravida 3, vaginal delivery following intrauterine fetal death (IUFD) occurred. The physical examination of the stillborn baby showed typical Potter facies and normal upper part of the body with fused lower limbs and two feet. The two feet were fused, and the femur and tibiae with four toes were discriminable. The external genitalia and anal opening were absent, as shown in Fig. Eventually, the follow-up to her fourth pregnancy indicated that she gave birth to a normal baby.Although the etiology of sirenomelia is still unclear, there are two chief pathogenetic hypotheses. One of these is defective blastogenesis, which involves deficient mesodermal generation and subsequent impairment in the formation of caudal structures. Based on this theory, sirenomelia could be considered a severe form of CD .Another hypothesis for its etiology is an abnormal vasculature pattern. In most cases, fetuses with sirenomelia exhibit a single umbilical artery (SUA) with an abnormal origin, derived from the vitelline artery. A normal fetus, on the other hand, has two umbilical arteries  with a net al. reported that the risk of sirenomelia increases in cases with mothers younger than 20 years of age. Nevertheless, there is a strong relationship between maternal diabetes or twin pregnancy and higher maternal age in cases of sirenomelia [Risk factors such as maternal diabetes, heavy metal exposure for sirenomelia, and CD are well established in both experimental models and humans, although only 0.5–3.7% of reported cases had diabetic mothers . The freenomelia . An assoenomelia .Exposure to extrinsic elements such as teratogenic agents can lead to sirenomelia. In this regard, some studies have found a relationship between sirenomelia and certain teratogenic drugs such as methylergonovine maleate , phenobaUnlike in humans, the genetic basis in the animal model is well established. A sirenomelia-like phenotype in animal models has been illustrated by genetic modifications through induction of loss-of-function (LOF) mutations in bone morphogenetic protein (Bmp) or gain-of-function (GOF) mutations in RA signaling. The genetic disruption of the key enzyme that degrades RA, Cyp26a1, reportedly induced sirenomelia in mice . The admBmp signaling plays a vital role in the remodeling of the vasculature . There ide novo balanced translocation involving chromosome 16 and triploid mosaicism was reported [Unlike mutant mice with sirenomelia, such as Bmp7 on 20q13.31, Bmp4 on 14q22.2, and Cyp26B1 on 2p13.3, very few chromosomal conditions have been reported in humans. Recently, a reported .et al. [Orioli et al.  reportedet al. , 22. In et al. . In our Diagnosis of sirenomelia is more feasible in the first trimester than later in pregnancy because there is still a sufficient amount of amniotic fluid that is less affected by fetal urinary production . OligohyUltrasound examination is the gold standard method for sirenomelia diagnosis. Color and power Doppler ultrasound is helpful in the diagnosis of the single umbilical artery, especially in oligohydramnios cases . A firstThe roles of maternal diabetes, exposure to heavy metals, and RA have been acknowledged in the incidence of sirenomelia, and have been described as important environmental risk factors. Given its sporadic nature in humans, it is possible that sirenomelia occurs due to an autosomal-dominant genetic background; however, it is more likely that a combination of genetic and environmental components are responsible. The limitations of this study including a lack of ultrasound images and postnatal investigations prevent us from reaching an ultimate conclusion on the disease etiology, but the pedigree of this family could add value to the literature.


Background: Although mental health is an important part of health and wellbeing, very little is known about the impact of the COVID-19 pandemic on the mental health of marginalized communities like urban slum dwellers. Our study estimated the prevalence of generalized anxiety disorder and insomnia among the residents of the informal settlements of Dhaka, Bangladesh, during the COVID-19 pandemic.Methods: A cross-sectional phone-based survey was conducted from October to November 2020 among adult residents of five informal settlements of Dhaka city randomly chosen from an existing Urban Health and Demographic Surveillance Systems (UHDSS) run by icddr,b. Data on Generalized Anxiety Disorder-7 (GAD-7) and Insomnia Severity Index (ISI) were collected. A multinomial logistic regression was performed to assess the associated factors of anxiety and insomnia.Results: Of the total 586 participants, the prevalence of mild to severe anxiety and insomnia were 53% and 43%, respectively. As per the multinomial regression analysis, participants with mild anxiety were significantly more likely to be older (>50 years) and afraid of COVID-19 infection. Likewise, participants with moderate/severe anxiety were significantly more likely to share less household facilities  , to have difficulties in food availability , to be afraid of self , and to worry about the family members  getting infected. Participants with mild insomnia were significantly more likely to share fewer household facilities and be afraid of being infected with COVID-19 infection. Moreover, participants with moderate/severe insomnia were significantly more likely to be female , to receive food aid , to be afraid of self , and to worry about someone like friends or neighbors  getting infected with COVID-19.Conclusions: We found elevated prevalence of both anxiety and insomnia among the urban poor of Bangladesh in the context of COVID-19. This indicates the importance of integrating mental health in the mitigation and recovery efforts related to similar crises for the urban poor in the future. Globally, the urban population has increased rapidly from 751 million in 1950 to 4.2 billion in 2018, and it is projected to increase by 68% within 2050 –7. The aResearchers in many low- and middle-income countries (LMICs) have reported that disease outbreaks such as Ebola and influenza have led to a rise in unemployment, malnutrition, gender violence, and reduced access to healthcare services , 19, 20.During the COVID-19 pandemic, there are a few studies that focused on the mental health of urban poor in Bangladesh –27. MostA cross-sectional survey was conducted among slum dwellers in Dhaka city, Bangladesh, under the Urban Health and Demographic Surveillance System (UHDSS) of icddr,b. It is a large surveillance system of icddr,b since 2015 covering 31,577 households and 121,912 individuals in five slums of the Korail, Mirpur, Shampur, Dholpur, and Gazipur areas of Dhaka North and South city corporations. Households were selected randomly with a probability proportional to size (PPS) so that the population represents the urban slums in general. From each household, one adult (≥ 18 years of age) with phone number registered with UHDSS was selected for the study interview ensuring that the male–female ratio remained similar. The residents who had migrated permanently out of the slum were excluded from the study.As no similar studies among slum dwellers during the pandemic were available at the designing of the study, we assumed the prevalence of anxiety to be 50%. The minimum sample size (384) was estimated with an assumption of 50% poor mental health conditions with a 5% precision. Finally, a total of (384 × 1.5) 578 households were calculated with the design effect (1.5), and taking 20% non-response into consideration, ~695 households were estimated. Afterward, we determined the number of the respondents from each slum by using PPS methodology. Initially, a total of 1,519 adult slum residents (one from each household) were approached to take part in the survey. Finally, 586 participated and completed the survey, whereas 65 respondents refused to participate and 902 residents were unable to reach by phone, generating a non-response rate of 61.4%.To avoid moving around and face-to-face contact during the government lockdown , a mobile phone-based survey was conducted. Trained research assistants who were already working within the selected communities and obtained data through phone calls after getting verbal consent by using a Bangla pretested questionnaire from the respondents. The study protocol was reviewed and approved by the Institutional Review Board (IRB) of icddr,b.A semi-structured questionnaire was used for data collection that included sections on socio-demographics including questions on age, sex, years of schooling, occupation, and income. Other sections included COVID-19 impact on household characteristics, food environment, and emotional wellbeing characteristics. Under the section household characteristics, we have assessed crowding index (number of people per room) and shared facilities . We collected questions about food availability, accessibility, and food aid under the section of food environment. Emotional wellbeing had some variables such as fear of contracting the COVID-19 infection for self, family members, and other close network; in addition, this section also measured anxiety and insomnia using two psychometric scales that have been used previously in the Bangladeshi population: the Generalized Anxiety Disorder-7 (GAD-7) scale and the Insomnia Severity Index (ISI) , 29.The Generalized Anxiety Disorder (GAD-7) includes seven questions about symptoms of anxiety experienced over the last 2 weeks. Each question is scored on a four-point Likert scale . The total score ranges from 0 to 21 and is obtained by summing the raw scores of the seven items. A higher score is indicative of greater anxiety. In the present study, the following GAD-7 cutoffs were considered for assessing anxiety: > 15 for “severe anxiety”; 10–14 for “moderate anxiety”; 5–9 for “mild anxiety”; and < 5 considered was “minimal anxiety” . The CroThe Insomnia Severity Index (ISI) scale consists of seven items related to symptoms of insomnia over the past 2 weeks scored on a five-point Likert scale . The total score is obtained by summing the item scores. Individual scores range from 0 to 28, with higher scores reflecting more severe insomnia. The following ISI cutoffs were used for assessing insomnia; 22–28 for “severe insomnia”; 15–21 for “moderate insomnia”; and 8–14 for “mild” or “subthreshold insomnia”—and < 8 was considered as “minimal insomnia” . The Crop < 0.05 was considered as a statistically significant association. SPSS software (version 25) was used to analyze data.The descriptive statistics were computed for each variable. The bivariate analysis  were computed to assess the association between independent variables and outcome variables . Multinomial logistic regression analysis was performed involving all examined variables together to determine the associated factors of anxiety and insomnia categories (mild and moderate/severe). During regression analysis, “minimal” categories were kept as reference. Before performing the multinomial logistic regression models, multi-collinearity was checked. The multi-collinearity was absent as all values of the tolerance and variance inflation factor (VIF) were > 0.1 and < 2, respectively . All theOut of the 586 study participants, 50.9% were males; 48.6% were of 31–50 years old; 50.9% had 5–10 years of schooling; and 35.2% were housewives or students followed by 30.2% unemployed or daily wager. About 93.7% of respondents reported that COVID-19 had adversely affected their household income .Among the 586 study participants, 53.0% had mild to severe anxiety, and 43.0% had mild to severe insomnia . About aThe distributions of minimal, mild, and moderate/severe anxiety and insomnia are presented in p < 0.05) and to be afraid of being infected by COVID-19 , to have difficulties in food availability , to be afraid of being infected by COVID-19 , and to be worried about family members to be infected .A multinomial logistic regression evaluated the associated factors of anxiety and insomnia categories (mild and moderate/severe). The reference group was minimal anxiety and insomnia, respectively. Participants with mild anxiety were significantly more likely to be of older age (> 50 years)  and to be afraid of being infected by COVID-19 , to receive food aid , to be afraid of being infected by COVID-19 , and to be worried about someone like friends or neighbors getting infected .Participants with mild insomnia were significantly more likely to share fewer household facilities  have a significantly greater risk of anxiety than younger people. Prior to the pandemic, evidence from Bangladesh and India also documented that older people are disproportionately affected by mental illness due to the underlying long-term chronic illness, dependency on their family members, loneliness, etc. , 46. Sev years haIn this current study, insomnia was found significantly higher among females compared to males during this pandemic. The global prevalence of mental illness is also found to be higher among women (24.5%) compared to men (16.3%) , 52. SevThe significant association between increased number of shared facilities with decreased anxiety and insomnia is important to note. Although during the COVID-19 pandemic the public health officials have discouraged social interaction, our evidence suggests that sharing household facilities with other families, which is probably associated with interacting and socializing with neighbors, was beneficial for mental health as reported by researchers from India as well . This fiAmong our respondents during the COVID-19 pandemic, stressors like difficulties with food availability were significantly associated with increased anxiety. Lack of availability and accessibility of food can lead to food insecurity which is a stressful experience that can result in frustration, feeling of hopelessness, alienation, and feelings of shame that may trigger new or amplify preexisting psychosocial stress . In prevAccording to our study, participants who received food aid were significantly associated with insomnia. Over the past few years, different support programs are in place to support the residents of the informal settlements including food aids by different organizations , 61–64. Respondents who were afraid of contracting COVID-19 were more likely to suffer from anxiety and insomnia. Similarly, the respondents who were worried about someone with a close network getting infected had greater odds of insomnia. These findings were also in line with other Western and South Asian countries , 76–80. Although unprecedented in scale, the experience of the COVID-19 pandemic has demonstrated the gaps in the existing social safety nets for the urban poor. In Dhaka city, 4 million of the urban population live in 5,000 slums and they are a very important part of different sectors such as RMG (readymade garments), transportation, construction, domestic workforce, and part of other local businesses , 73. TheThere were a few limitations of our study. The study was cross-sectional which means the findings presented were applicable to that point in time only and did not provide us with the insights over time. As the study relied on self-report, this might have resulted in some under- or overestimation due to recall bias or social desirability bias. Due to the restriction of the pandemic, the data were collected over phone which might cause some bias in the assessment of the mental health conditions of the respondents. Slum dwellers were not interviewed who did not have mobile phones or who could not be reached, which may lead to a kind of selection bias. Further, we do not have pre–post data of the same population to assess the additional contribution of COVID-19 pandemic in terms of mental health outcomes.This study used the PPS methodology, which meant that the sample represents the residents of the five major informal settlements of Dhaka and Gazipur districts. We applied standard validated scales to assess the anxiety disorder and insomnia among the respondents. Furthermore, we have collected all the information just after the initial lockdown was over, which means that the findings represent the effect of the lockdown on mental health.The findings indicate that anxiety and insomnia were prevalent and higher than previously observed. This study emphasizes the need to integrate the consideration of mental wellbeing in the infection control mechanism for the present pandemic as well as for similar situations in the future. Our study also recommends that during emergencies such as one presented by COVID-19, it is important that the health and wellbeing of the urban poor are safeguarded through a well-designed comprehensive social safety net program. Special effort could be made to educate communities about how to be safe while having social interactions. Primary care facilities could be equipped to deal with identification and referral of mental health conditions, and local opinion leaders could be engaged in talking about the importance of mental health. Finally, in the context of severe shortage of trained staff who are equipped to deal with mental health conditions, it is important that telemedicine and other online counseling facilities are made accessible and affordable for the urban poor so that their wellbeing can be ensured.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the Institutional Review Board (IRB) of icddr,b [PR-20075]. The participants provided their verbal consent through phone calls to participate in this study.KK and SR: conceptualization. KK, SR, FK, SM, SH, and DR: methodology. MI, KK, MK, and SR: formal analysis and investigation. KK, MK, and MI: writing—original draft preparation. SM, SH, DR, FK, and SR: writing—review and editing. SR and DR: resources. SR: supervision. All authors have read and approved the final manuscript.This study was funded by the Swedish International Development Cooperation Agency (SIDA), and the Grant Number was GR-01455. No other specific grant from any funding agency or from any other non-profit sectors was received for this research.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.


A sense of belonging is a significant predictor of mental health and well-being in later life. A sense of belonging in childhood and adolescence contributes to a number of adult behavioural and psychological outcomes. A high sense of belonging has been associated with better health, longevity, psychological well-being, and disease recovery.In this study, the Persian version of the Sense of Belonging Instrument (SOBI) for older adults in Iran was evaluated psychometrically to develop an accurate measure for belonging. Participants in the study were 302 older adults, 60 years old and above, living independently in Iran and chosen through convenience sampling.An exploratory factor analysis indicated that the four-factor structure, which included 16 items, accounted for 54.12% of the total variance, and was characterized by strong factor loadings, with values ranging from .50 to .87. Thereafter, a confirmatory factor analysis confirmed the four-factor latent structure of the SOBI, providing adequate data-model fit statistics. All latent structures were characterized by adequate-to-strong latent construct (H) internal reliability (α) coefficients.The Persian version of the SOBI is a useful tool in understanding older adult patients’ sense of belonging when living independently within the community. The implications for practice and research are discussed. Theory of Human Relatedness (1993) [fit, or are congruent with others, groups, organizations, environments, or spiritual dimensions [According to Hagerty et al. 1992), a sense of belonging occurs when people feel like they are an integral part of a system or environment [992, a sedness 199 [fit, ormensions –7. Howevmensions . For examensions . It is tmensions , 11–14.The outcomes of a sense of belonging, according to Hagerty et al. 1992), include meaningful engagement at the psychological, social, spiritual, or physical level. A low sense of belonging is a significant predictor of mental health problems  and is related to stress, loneliness, and suicide –18. In o92, incluThe benefits of belonging include adaptive physiological and psychological outcomes such as lowered heart rate and improved social self-esteem . In oldeThe maintenance and promotion of mental health and well-being in later life are an important, yet an under-researched, area of knowledge –28. A seSocial groups have been found to be a protective factor for premature death in older adults who have retired . Older aBelonging has been identified as enhancing the mental health of older residents in long-term care . BelongiFrom a research perspective, our understanding of belonging has been weighted towards literature from first world, Western countries. However, the experience of belonging is highly contextual and interplays with notions of culture, race, religion, language, and socio-historical backgrounds . For insDespite the reported importance of belonging in the literature, there are relatively few measures available to assess this construct, especially in applied settings such as clinics and aged care homes. Several belonging measures rely on a scale designed to be used only for a single study  . FurtherThe Sense of Belonging Instrument (SOBI) is one of the most utilised measures of belonging available. The 27-item self-report measure developed by Hagerty and Patusky (1995) consists of two distinct scales: the SOBI-P , which measures sense of belonging in terms of valued involvement and fit in relationships, and the SOBI-A (antecedents), which examines the antecedents to a sense of belonging . While tr = .42) and negatively correlated with loneliness (r = −.76). Unlike other data-driven measures of belonging, the SOBI-P was developed from the Theory of Human Relatedness [The 1995 study by Hagerty and Patusky confirmed that the SOBI was positively correlated with social support , 0.98 (sample of Thai women), and 0.84 (sample of adolescents) , 53. HowFor the SOBI to be used in Iran, research is needed to evaluate the Persian version’s psychometrics as no previous studies have examined the psychometric properties of the SOBI in an Iranian context previously. However, there has been sufficient research in Persian countries on constructs related to belonging to assume that it is a construct of relevance and value , 25, 54.The current study aims to psychometrically evaluate a Persian version of the SOBI for older adults in Iran. There are several potential implications for the findings of this study. First, an empirically rigorous tool for measuring belonging has utility for psychologists and other healthcare professionals who work with older adults. Second, a context-specific measurement tool may also provide researchers with the ability to measure belonging and devise interventions more accurately. In responding to the call for examining the effect of cultural factors in explaining belonging  and its This study’s sample consisted of 302 older adults aged 60 years or older living independently in the community. Data were collected from public places in two urban provinces in Iran: Tehran and Qazvin . Some of the participants were classified as illiterate , so the research team explained the study's purpose in a simple and clear language. Data collection took place in 2018. Older adults were eligible if they were willing to participate, were age 60 years or over, were fully oriented to time and place, and had the ability to communicate and answer the questionnaire’s items.Participants completed the questionnaire that collected data on SOBI as well as demographic information. The demographic information contained participants’ age, gender, educational and socioeconomic status, marital status, and main income sources.The Sense of Belonging Instrument (SOBI)  consistsWritten permission was obtained from Professor Hagerty, the author of SOBI to use the measure in the present study. Then, using The World Health Organization translation protocol, the SOBI was translated into Persian, with a forward-backward technique . Two EngThe factor structure of the SOBI was examined using exploratory and confirmatory factor analysis. Exploratory factor analysis was first performed to explore the factor structure of the measure. The Kaiser–Meyer–Olkin (KMO) and Bartlett’s test of sphericity were used to check the sample’s appropriateness to conduct the factor analysis. Factor extraction was based on: (i) eigenvalues > 1; (ii) communalities > .3; and (iii) scree plot . The EFAH), and composite reliability (CR) estimates were used to evaluate the reliability of the Persian version of SOBI, and the coefficient ≥ .70 was considered adequate [After examining the factor structure of the measure, reliability analyses were performed. The internal (α), latent construct  were male. The age of older adults who participated in this study was 67.75 ± 6.59. The majority  of participants were married. Table Most of the participants n = 82; 60.3%χ2 = 4108.89, df = 351, p < .001), supporting an exploratory factor analysis. Results of the first-factor analysis revealed seven factors with eigenvalues > 1, and these constructs explained approximately 56% of the variance. However, the pattern matrix results indicated five cross-loading items (λ > .32) and five non-loading items (λ ≤ .40), and the scree plot and parallel analysis results suggested that a four-factor latent structure would be a better fit to the data. After excluding cross and non-loading items, the factor analysis was re-run, demonstrating the four-factor structure , which included 16 items accounting for 54.12% of the total variance, with eigenvalues of these four factors ranging from 1.21 to 5.54. The first component of the measure comprised four items related to fitting in. The second included four items measuring social esteem. The third consisted of four items assessing collective self. The final component comprised four items related to self-awareness of older adults, see Table Following the assumption that the normality outcomes and factors would be inversely correlated, exploratory factor analysis was conducted using principal axis factoring with a promax rotation to explore the latent factor structure of the SOBI. Preliminary analysis results showed the 27 SOBI items had adequate KMO sampling adequacy (.80) and lack of singularity (Bartlett’s χ2) value was significant , and the χ2/df ratio was 3.16. The root mean square error of approximation and the standardized root mean square residual values were adequate . In addition, comparative fit index and non-normed fit index values were .90 and higher, suggesting an adequate data-model fit . All latent structures were also characterized by strong latent construct reliability coefficients (H range = .82–.85). Factor loadings of the scales were adequate-to-strong, ranging from .47 to .85 , and corrected item-total correlation coefficients, which are the Pearson’s correlation between each item, ranged between moderate and large .43 to .73). The scale also provided adequate–to–strong internal consistency reliability coefficients , which included 16 items accounting for 54.12% of the total variance. Factor loadings of these latent constructs were strong, with values ranging from .50 to .87. The findings contrast the findings from the original two factor-based scales . After eWhen interpreting the results of this study, the characteristics of the study participants should be a consideration. As the study of the older Iranian adults was a convenience sample, and the participants were living independently, results cannot be considered representative of those that may be found for the population of all Iranian adults over 60 years. The lived experience of older Iranians in residential care would likely differ from those residing in the community. Future research should consider evaluating the SOBI with older adults living in residential care, especially given the risk for loneliness in this demographic group. Additional validity of other belonging measures requires the support of the discriminant and convergent validity of the Persian form of the SOBI. In terms of preventative care, it is plausible to assume that the early identification of low belonging in younger age groups with an appropriate measure may help prevent loneliness in older adulthood, assuming the appropriate interventions can be implemented. Finally, we conducted CFA and EFA within the same sample, which is considered an additional limitation of the present study. There is a need to confirm the factor structure of the SOBI using CFA with different samples in future studies.The current study evaluated the psychometric properties of a Persian version of the SOBI. The findings indicated that the four-factor Persian version of the SOBI is a valid and reliable tool for measuring belonging in older adults over 60 living independently. The findings of this study have implications for how belonging is measured in older Iranian adults. Pending further investigation into its psychometric properties within the Iranian context, The Persian version of the SOBI offers the promise of utility for psychologists and health care professionals working with older adults living in the community.


The medial (MEC) and lateral entorhinal cortex (LEC), widely studied in rodents, are well defined and characterized. In humans, however, the exact locations of their homologues remain uncertain. Previous functional magnetic resonance imaging (fMRI) studies have subdivided the human EC into posteromedial (pmEC) and anterolateral  parts, but uncertainty remains about the choice of imaging modality and seed regions, in particular in light of a substantial revision of the classical model of EC connectivity based on novel insights from rodent anatomy. Here, we used structural, not functional imaging, namely diffusion tensor imaging (DTI) and probabilistic tractography to segment the human EC based on differential connectivity to other brain regions known to project selectively to MEC or LEC. We defined MEC as more strongly connected with presubiculum and retrosplenial cortex (RSC), and LEC as more strongly connected with distal CA1 and proximal subiculum (dCA1pSub) and lateral orbitofrontal cortex (OFC). Although our DTI segmentation had a larger medial-lateral component than in the previous fMRI studies, our results show that the human MEC and LEC homologues have a border oriented both towards the posterior-anterior and medial-lateral axes, supporting the differentiation between pmEC and alEC. It is cWhile comprehensive entorhinal delineations based on cytoarchitectonic analyses exist , we cannfMRI studies have indeed shown that certain properties of the rodent and non-human primate EC also apply to the human EC . Based oIn addition to the neuroimaging modality, the second reason for a re-evaluation has gained additional importance since the assumption about EC connectivity on which parts of the previous fMRI studies  were basIn order to identify the human homologues of MEC and LEC, we should take advantage of known unique connections to each subregion. For example, in rodents the presubiculum projects almost exclusively to MEC, whereas distal CA1 and proximal subiculum  project most strongly to LEC , 2021. MThe objective of this study is therefore to identify the human homologues of the rodent MEC and LEC using DTI, incorporating the novel insights from rodent anatomy. To achieve this, we performed probabilistic tractography on high-quality DTI data acquired by the Human Connectome Project . We iden22.1https://ida.loni.usc.edu, http://db.humanconnectome.org), in line with the MGH-USC HCP Data Agreement. All participants provided written informed consent, and the experiments were approved by the institutional review board of Partners Healthcare (2 (64 directions), 3000 s/mm2 (64 directions), 5000 s/mm2 (128 directions) and 10,000 s/mm2 (256 directions). One non-diffusion-weighted (b = 0) image was collected every 14 image volumes.Publicly available structural and diffusion MRI data from 35 healthy adults were obtained from the MGH-USC Human Connectome Project database  functions recon-all and dt-recon on the participant's structural and diffusion images, respectively (http://fsl.fmrib.ox.ac.uk/fsl/) function BET  based on mutual information . To create RSC and OFC ROIs, respectively, the FreeSurfer parcellations named “isthmus cingulate” and “lateral orbitofrontal” were used as a starting point. The final RSC ROI was obtained by tailoring the isthmus cingulate and removing the excess superior areas, while the final OFC ROI was obtained by extracting the posterolateral quadrant of the lateral orbitofrontal area. All resulting ROIs are shown in Supplementary Fig. 1. The ROIs were registered to the participants’ individual spaces by applying the calculated transformations from ANTs. To increase the anatomical precision of the ROIs, the registered ROIs were then masked by respective participant-specific FreeSurfer parcellations.Regions of interest (ROIs) including the EC, presubiculum, CA1 and subiculum were extracted from the automated cortical and subcortical parcellation obtained from running FreeSurfer's template . The EC template . Since t2.3bedpostx on the diffusion data, using the zeppelin deconvolution model, a Rician noise model, and burn-in period 3000  were computed by running the FSL function iod 3000 . Probabihe fODFs , 2003b. he fODFs . For tra2.4find_the_biggest  and medial-lateral (ML) axes, respectively. This was performed by first calculating the centers of gravity of the differently defined MECs and LECs, and the vector between these centers of gravity. Next, the angle between this vector and a pure PA or ML vector was determined. We defined the PA axis as the long axis of the hippocampus. The degree of PA- or ML-oriented border was then defined between 0 and 100% such that an angle of 0° to the PA or ML vector means that the border is 100% oriented along the PA or ML vector, respectively. Correspondingly, an angle of 90° would mean that the border is 0% oriented along the respective axis, i.e. it is orthogonal to that axis. In addition, the different segmentations were compared with respect to the sizes of the resulting MECs and LECs, and the size ratios between these were calculated. All these segmentation comparisons were also carried out on the two fMRI-based segmentations of pmEC and alEC available for download from earlier studies .3To qualitatively visualize the connectivity paths between the EC and the regions hypothesized to be connected with its subregions, we ran probabilistic tractography between the regions. By seeding paths from all voxels in the EC, presubiculum, dCA1pSub, RSC and OFC ROIs, maps of the connectivity paths between the EC and the other ROIs were created. The resulting group averaged paths are shown in Because we wanted to segment the EC into the MEC and LEC homologues based on the connectivity with other regions, a voxel-by-voxel measure of connectivity probability was needed. We therefore also ran the tractography only seeding from the EC ROIs. Then, for each voxel in the ROI, we counted how many of the seeded paths reached the other ROIs. These connectivity counts were normalized to a probability, providing connectivity maps for the EC with the other four ROIs. The resulting smoothed and thresholded group averaged connectivity maps are shown in For segmentation into the MEC and LEC homologues, the main hypothesis was that these regions could be identified based on connectivity with presubiculum vs. dCA1pSub, respectively. The actual segmentation was performed on a voxel-by-voxel level in the EC determining with which of the other two regions the connection probability was highest, using the connectivity maps described in the previous paragraph. For comparison, the MEC-LEC segmentation was also performed based on connectivity with RSC vs. OFC, respectively. This was first performed individually for all participants, and inter-participant segmentation variability maps for the presubiculum vs. dCA1pSub and RSC vs. OFC segmentation approaches are shown in The same connectivity-based MEC-LEC segmentation was performed on a group level using the group averaged connectivity maps from In a next step, since the borders of the segmentations from different approaches showed slightly different orientations along the posterior-anterior (PA) and medial-lateral (ML) axes, we wanted to quantify this directional difference by calculating the “degree” of PA- and ML-orientation of the borders. This was defined as a percentage from 0 to 100%, dependent on the angle between the MEC-LEC center of gravity vector and a pure PA or ML vector. Finally, we wanted to compare the resulting sizes of the MEC and LEC homologues from all the different segmentation approaches, and these are shown in 4In this study, we used DTI and probabilistic tractography in 35 healthy adults to segment the human EC into homologues of what in other mammals have been functionally, connectionally, and cytoarchitectonically defined as MEC and LEC. We based the segmentation on EC connectivity with four brain regions known to selectively project to either of the EC subregions in multiple species. Different combinations of these four regions all showed both a posterior-anterior (PA) and a medial-lateral (ML)-oriented border between the human homologues of MEC and LEC. This orientation of the thus defined border is similar to that defined in previous fMRI studies resulting in the definition of the two subregions as pmEC and alEC . Note hoThe results from our study substantiate the pmEC and alEC subdivision of the human EC suggested in previous fMRI studies . AlthougThere are several possible explanations as to why our DTI study showed slightly different segmentation results than the fMRI studies. First, DTI and fMRI are two different imaging modalities with inherently different mechanisms of connectivity. While DTI exploits the diffusion of water molecules in order to trace the structural paths of connectivity between brain regions , fMRI idInterestingly, using different seed regions to identify MEC and LEC resulted in varying degrees of PA- and ML-orientation of the border between them. It is unclear whether this is inherently linked to the DTI method, or due to an actual connectivity difference between the regions. Using presubiculum and dCA1pSub as the seed regions, which are situated medially and laterally with respect to each other, respectively, resulted in a border with higher degree of ML- than PA-orientation. On the other hand, using RSC and OFC, which are situated posteriorly and anteriorly in the brain, respectively, resulted in a border with higher degree of PA- than ML-orientation. Although it is not unnatural to assume that the brain is organized such that connected regions are situated more closely to each other, this could also be an effect of using probabilistic tractography, where the apparent connectivity probability depends on e.g. the length of the path and the size of the ROIs . In otheIn order to determine and compare the connectivities between the EC and the other ROIs, we normalized the connectivity maps by dividing them by the maximum probability of each map. This could introduce a bias in the results. By doing this, we intrinsically assume that the maximum connectivity strength to each of the other ROIs are equal, and the segmentation process does not take into account that the MEC connections might be stronger than the LEC connections, or vice versa. However, little is known about the strength of connectivities at this level of detail, particularly since it is not straightforward to examine or even define connectivity strength. Connectivity strength surely depends on axonal density, but other factors like synaptic density and efficacy are other important variables. Nevertheless, even if we were to know that some of the connections are stronger than the others, probabilistic tractography provides a relative instead of an absolute measure of connectivity and is also dependent on path lengths, ROI sizes and the number of possible path directions in a voxel. Normalizing the connectivity maps based on different connectivity strengths would therefore be a highly complex task. Therefore, we did not impose any further assumptions about connectivity strengths in our analyses.ex vivo DTI data , FMRIB Software Library's , and Advanced Neuroimaging Toolbox .The following, freely available code/toolboxes were used in this study: FreeSurfer (version 7.1.1, Ingrid Framås Syversen: Conceptualization, Methodology, Software, Formal analysis, Investigation, Writing – original draft, Writing – review & editing, Visualization. Menno P. Witter: Methodology, Validation, Writing – review & editing. Asgeir Kobro-Flatmoen: Validation, Writing – review & editing. Pål Erik Goa: Conceptualization, Methodology, Resources, Writing – review & editing, Supervision, Funding acquisition. Tobias Navarro Schröder: Conceptualization, Methodology, Validation, Writing – review & editing, Supervision. Christian F. Doeller: Conceptualization, Validation, Writing – review & editing, Supervision, Funding acquisition.The authors declare no competing interests.


Post reperfusion syndrome (PRS) is a relatively common and life-threatening complication during orthotopic liver transplantation (OLT). It is associated with poor patient and transplanted liver outcomes.This study aimed to compare the risk factors of PRS during OLT.Clinical-epidemiological observational retrospective study.We gathered the records of patients who underwent OLT in 3 years, from May 22, 2016, to May 22, 2019, in Namazi and Bu-Ali Sina organ transplantation hospitals.In this study, we assessed 1182 patients who underwent OLT. Patients were divided into two groups based on the presence or absence of PRS.Diagnosing the predictors of PRS was the primary outcome of this study.Results showed that age > 60 years, Child-Pugh scores C, higher Model End Stage liver disease score, and preoperative sodium < 130 mmol/l (parameters of the liver recipient), increase in cold ischemic time (the donors’ parameters), and the classical technique  were the strong predictors of PRS.The results indicated that underlying liver disease was not the predictor of PRS in the presence of other risk factors; therefore, clinicians have to consider these risk factors in patients undergoing OLT.The online version contains supplementary material available at 10.1186/s12871-022-01635-3. Liver transplantation is a life-saving procedure and the treatment of choice for severe chronic liver disease and acute liver failure patients.Post reperfusion syndrome is a relatively common but life-threatening complication during liver transplantation.This study showed that 33% of patients who underwent liver transplantation at this center had PRS.Age > 60 years, CTP class C, higher MELD score, preoperative sodium < 130 mmol/l, CIT, and classical technique were PRS predictors.Orthotopic Liver transplantation (OLT) is a life-saving procedure and the treatment of choice for severe chronic liver disease and acute liver failure , 2. PostOLT is major surgery with a high occurrence of fluid shift and is often performed in patients with hemodynamic changes associated with the pathology of the underlying liver disease. On the other hand, reperfusion syndrome is a severe complication during surgery following portal vein declamp. Despite the improvement of surgical and anesthesia techniques, it is associated with severe metabolic and cardiovascular disorders that can affect mortality and morbidity of patients , 15. PosThis clinical-epidemiological observational retrospective study extracted data from May 22, 2019, to November 22, 2019. We gathered the records of patients who underwent OLT in 3 years, from May 22, 2016, to May 22, 2019, in Namazi and Bu-Ali Sina organ transplantation hospitals. The Ethical Committee of the Vice-chancellor of Research (Chairperson: Dr. Mohammad Javad Ashraf) at Shiraz University of Medical Sciences approved this study . We conducted this study to determine the predictors of PRS, including the type of underlying liver disease leading to transplantation and other risk factors.Inclusion criteria included adult patients (age ≥ 18 years) with end-stage liver disease who underwent deceased-donor OLT.Exclusion criteria included OLT from a living donor, split liver graft, simultaneous liver and kidney transplantation, patients with acute liver failure, incomplete information recorded before and during surgery, and the occurrence of intraoperative severe adverse events before the reperfusion phase.Upon admission to the operating room and after appropriate venous line placement for all patients, we performed non-invasive blood pressure monitoring, electrocardiogram, and pulse oximetry. Besides, we placed the arterial line for invasive pressure monitoring, the central venous line under ultrasonography for central pressure, temperature probe, and capnography immediately after induction of anesthesia. PICCO monitoring and, if needed, TEE was used to improve patient management in patients with high MELD (model for end-stage liver disease), heart problems, or aortopulmonary shunt.We performed induction of anesthesia with the injection of 0.02 mg. Kg-1 midazolam, 2 μg. Kg-1 fentanyl, 3-5 mg. Kg-1 sodium thiopental and 0.2 mg. Kg − 1 pancuronium. The anesthesia maintenance was done using isoflurane with MAC of less than 1% in combination with oxygen/air.After intubation, we provided mechanical ventilation with a tidal volume of 8-10 ml / kg, respiratory rate of 10–16 breaths/min, and 5cmh2o PEEP to maintain end-tidal CO2 partial pressure (PETCO2) of 35–45 mmHg for the patient. Albumin and normal saline were used for fluid therapy based on hemodynamic parameters and central venous pressure. Patients were monitored to maintain systolic blood pressure and heart rate at 20% of baseline during surgery. We performed necessary interventions in case of hemodynamic changes using anesthetics, cardiovascular drugs, and fluids.Based on the patient’s condition and the surgeon’s decision, we used piggyback or classical techniques during surgery for OLT. We did not use venovenous bypass during the surgery. Before reperfusion, the graft was rinsed with 1 ml / kg normal saline through the portal vein.We extracted the following donor’s data from the records:Age, sex, length of stay in the intensive care unit (ICU), the blood concentration of alanine transferase (ALT), aspartate transferase (AST), alkaline phosphatase, bilirubin, sodium, and potassium levels, and preoperative international normalized ratio (INR), as well as warm ischemia time (WIT) graft, and cold ischemia time (CIT) graft.We collected the recipient’s data from patients’ records. It included age, sex, weight, type of underlying liver disease leading to transplantation, preoperative sodium level, MELD , , and ChiFollowing adverse events occurring in ICU were evaluated as well: primary graft nonfunction (PGNF: retransplantation or death within 7 days), need for renal replacement therapy (RRT), major cardiovascular events , need for surgical revision , central nervous system events , length of stay in ICU and hospital, and finally the outcome of patients before discharge and within 2 years of follow-up after discharge.PRS was noted by the decreased mean arterial pressure of more than 30% of baseline values within the first 5 min after PRS of the donor’s liver with the recipient’s blood during the transition from the anhepathic phase to neohepathic phase, which lasts for at least 1 min. We used norepinephrine as a bolus and infusion to maintain mean arterial pressure (MAP) for this complication. In case of hypotension along with severe bradycardia, we used epinephrine bolus and infusion. Finally, we used vasopressin infusion in case of the need for a high dose of norepinephrine or its ineffectiveness.After extracting information from the records, based on the occurrence or non-occurrence of this complication, patients were compared in groups 1 (PRS +) and 2 (PRS -).P-value< 0.05 indicated statistical significance.Data were analyzed in IBM SPSS Statistics for Windows, version 19 . The continuous variables were reported by median (IQR) and tested by the Mann-Whitney U test. The categorical variables were reported by number (percent) and tested by Chi-square or Fisher exact. Univariate and multivariable logistic regression models were used to determine risk factors for predicting PRS. In logistic regression models, we have to consider one category of categorical variables as the reference. In this study, we chose the variable with the highest risk of RPS as the reference. A total of 1590 patients underwent OLT within 3 years of the study . In terms of age distribution, there was a significant difference between groups. The highest incidence of PRS was in patients aged more than 60 years, and the lowest incidence was in patients under 20 years . But in terms of weight and sex, there was no significant difference. Regarding disease severity distribution between the two groups, a higher incidence of PRS was noted in patients with higher MELD and CTP scores. The highest incidence of PRS occurred in patients with CTP C compared to A . Also, assessing the effect of preoperative sodium on PRS showed that patients with sodium less than 130 compared to sodium more than 130 had a higher incidence of this complication   had PRS. There was a significant difference between groups regarding underlying disease leading to transplantation. We found the highest incidence of PRS in the viral hepatitis group and the lowest in the biliary disease group . However, there was no significant difference between the two groups regarding sex and test levels of K, AST, ALT, ALKP, Bilirubin, and INR before donation (P > 0.05). Only in donors with higher sodium levels, more PRS was observed (P = 0.03). Donor organs also had a longer CIT for patients with PRS (P = < 0.001). But in terms of WIT, no statistically significant difference was observed between the groups (P = 0.132) . Among the surgical parameters, the classical technique  was the strong predictor of PRS compared to the type of underlying liver disease, donors’ age, and sodium level. In the case of underlying liver disease and through univariate analysis, the odds ratio of PRS in biliary and autoimmune diseases was significantly lower than in viral hepatitis. However, in multivariate analysis, this factor was not indicated as a predictor of PRS Table .P < 0.001). Patients with postoperative PRS also experienced more delirium  and major cardiovascular event . The need for postoperative surgical revision was higher in the PRS group than in the other group, of which bleeding was the most common cause of surgical revision. The length of hospitalization in the ICU as well as the hospital in the group with PRS was significantly longer than the other group . There was a significant difference between the groups in the rate and causes of death in the postoperative period. The mortality rate in the PRS group was higher than the other group  and PGNF, multiple organ failure (MOF) / sepsis and bleeding were the most common causes of death in the initial period after LT, respectively. Also in the two-year follow-up after discharge, it was found that the mortality rate in the PRS group was significantly higher than other group  and MOF / sepsis and chronic rejection were the most common causes of death in both groups, respectively , increase in cold ischemic time (the donors’ parameters), and the classical technique .The incidence of PRS varies from 3.6–81% among studies . The mosAssessing the distribution of liver disease leading to OLT, our results showed that biliary disease, viral hepatitis, and metabolic diseases were the most common causes of OLT from 2016 to 2019. In a previous Iranian study examining the status of OLT before 2016, the most common indications of OLT were cirrhosis caused by viral hepatitis, cryptogenic, and biliary disease . The difAlthough the exact pathophysiological mechanism of PRS is not fully understood, several studies have attempted to provide a list of possible risk factors for PRS to improve prevention and treatment strategies. Based on the results of studies, these risk factors were divided into three categories: donor/organ-related, recipient-related, and procedure-related .Most of the risk factors were related to liver graft recipients . Findingp-value < 0.05). However, when we assessed this parameter in the multivariate analysis and other risk factors for PRS, no significant difference was observed between the underlying liver diseases in terms of PRS, so the underlying liver disease was not recognized as the PRS predictor. This result indicated the greater importance of other risk factors for PRS than the recipient’s underlying liver disease. The cause of higher incidence of PRS in patients with viral hepatitis than biliary and autoimmune diseases can be attributed to older age and higher severity of underlying diseases such as higher MELD and CTP scores in these patients. In an abstract examining the risk factors of PRS, there was no significant correlation between PRS and the etiology of liver disease [In terms of the incidence of PRS based on the underlying liver disease leading to OLT, so far, the studies compared the incidence of PRS between the two groups of patients with acute and chronic hepatitis. To the best of our knowledge, few studies investigated the incidence of this complication among chronic liver diseases. The univariate analysis on the incidence of PRS based on chronic underlying liver disease showed that the highest incidence of PRS occurred in patients with viral hepatitis (42%) and the lowest in patients with biliary (23%) and autoimmune diseases (30%). We observed a significant difference between the groups ( disease .In terms of age, studies have shown that the recipients’ age > 60 years is a risk factor for PRS . In thisThe most important risk factors related to donor and liver graft resulting from studies were the age of the donor over 60 years, long CIT , and DonOne notable finding of our study was the association between the occurrence of PRS with postoperative poor liver graft and patient’s outcome. The incidence of PGNF and mortality in the PRS group was significantly higher than the other group. In the study of Siniscalchi et al., PRS was identified as an independent risk factor for PGNF and mortality . HoweverSo far, many studies have been performed to diagnose PRS risk factors. Still, according to this search, few studies have been conducted to investigate the incidence of PRS based on underlying liver disease leading to OLT. On the other hand, the sample size of this study was large, and so far, no study with this size has been conducted to evaluate the incidence of PRS, which can be the strengths of this study. As we performed a retrospective single-center study, further prospective mutlicenter studies can berecommened.The study showed that among the PRS risk factors related to the organ recipient, age > 60 years, CTP class C, higher MELD score, preoperative sodium < 130 mmol/l, and among the PRS risk factors related to the organ donor, only CIT and among PRS risk factors related to surgery technique, classical technique were PRS predictors. Despite the significant differences between liver diseases leading to OLT in PRS, it was not the OLT predictor. This result indicated the greater importance of other risk factors for PRS than the recipients’ underlying liver disease. Therefore, clinicians have to consider these risk factors in patients undergoing OLT.Additional file 1.Additional file 2.