Spontaneous reporting of adverse drug reactions (ADRs) is a method of monitoring the safety of drugs and is the basic strategy for the post-marketing surveillance of the suspected drugs. Despite its importance, there is very little reporting of ADRs by healthcare professionals. The present study has evaluated the knowledge, attitude and practices of health care professionals (HCPs) regarding pharmacovigilance activities in Lahore, Pakistan.A cross-sectional questionnaire-based survey was employed, and a convenience sampling was opted to collect the data among physicians, pharmacists and nurses working in tertiary care public hospitals of Lahore, Pakistan from September 2018 to January 2019.P < 0.05), 70.2% pharmacists and 76.1% nurses showed a positive attitude that they are the most important HCPs to report an ADR. About 64.3% of physicians (P < 0.05) emphasized that consulting other colleagues is important before reporting an ADR. Of all, 77.7% physicians, 75.7% pharmacists and 68% of nurses had positive attitude that ADR reporting is a professional obligation and 67.6% of the pharmacists stated that they have reported ADRs in their workplace and 77.2% nurses have verbally reported ADRs to the concerned personnel or department.Of the 384 questionnaires distributed, 346 health care professionals responded to the questionnaire (90.10% response rate). Most participants had good knowledge about ADR reporting, but pharmacist had comparatively better knowledge than other HCPs regarding ADR (89.18%) pharmacovigilance system (81.08%), its centres (72.97%) and function (91.89%). Most of the participants exhibited positive attitude regarding ADR reporting, such as 49.1% of physicians (Among all HCPs, pharmacists had better knowledge about ADR reporting and pharmacovigilance. All HCPs had positive attitude and inclination towards ADR reporting. The discrepancies were observed in the practices related to ADR reporting, whereas most of the participants including physicians and nurses did not report any ADR. Based on the above, strategies are needed to educate, train, and empower the HCPs in the domain of pharmacovigilance. The World Health Organization (WHO) defines an adverse drug reaction (ADR) as, “a response which is noxious and unintended, and which occurs at doses normally used in humans for the prophylaxis, diagnosis, or therapy of disease, or for the modification of physiological function . ADRs poSpontaneous or voluntary reporting of ADRs refers to the passive reporting of ADRs by healthcare professionals (HCPs) or patients and is the basic strategy for the post-marketing surveillance of the suspected drugs . These sHealthcare professionals are responsible for the identification, documentation, and reporting of ADRs and their contribution is essential to the early detection and reporting of an ADR . HoweverConcerning the scarcity of literature about the evaluation of pharmacovigilance activities in Pakistan, assessment of the knowledge, attitude and practices are critical to study. Thus, keeping in view the available literature, this quantitative survey has been developed to evaluate the trends of pharmacovigilance activities in the city of Lahore, Pakistan.This survey has adopted a triangulation of qualitative and quantitative methods, such as the findings from the qualitative study were used to design the questionnaire to assess the current scenario of knowledge, attitude and practices of pharmacovigilance activities in Lahore, Pakistan –21. By cA cross-sectional study design was selected and the sampling strategy that was considered appropriate for a cross-sectional survey was convenience sampling. In convenience sampling, participants were selected according to their convenient accessibility and proximity .To calculate an appropriate representative sample from the targeted population, the Cochran formula was used and a sample size of 384 HCPs was calculated to demonstrate a statistically significant result for the study .Participants were registered with the relevant provincial/national council.Participants provided a written consent to the researcher.The selected participants were HCPs including physicians, pharmacists and, nurses working full time, as permanent employee in tertiary care public hospitals of Lahore. The participants were selected only, if:The study was approved by the Humans Ethics Committee (HEC), University College of Pharmacy, University of the Punjab, Lahore, Pakistan with reference no. HEC/PUCP/1943.The survey questionnaire was developed from the literature review based on the knowledge, attitude, and practices of HCPs in relation to the adverse drug reaction reporting and the understanding of pharmacovigilance activities , 24–31.The questionnaire was comprised of five main sections with 35 items covering the following components: The first part of the questionnaire consisted of 18 items, which were divided into three main sections including knowledge about ADR reporting, types of ADRs to be reported and, knowledge about pharmacovigilance. The response for this part of the questionnaire was provided in the form of ‘yes’, ‘no’ and ‘don’t know’. The second part was comprised of 7 items exploring the attitudes of the HCPs and the factors that cause a positive or negative change in the attitude of these HCPs. These items were provided in the form of statements and HCPs were provided with a 5-point Likert scale  to indicate their disagreement or agreement. Moreover, the response  was labelled as negative attitude and (4 = ‘agree’ and 5 = ‘strongly agree’) as a positive attitude. The third part of the questionnaire covered 10 items based on the practices of HCPs towards ADR reporting. It was comprised of 4 items for reporting practices and 6 items related to the modes of reporting. Whereas the response was provided in the form of ‘yes’, and ‘no’.Before the survey was implemented, the questionnaire was tested for its face and content validity. Two academicians from the School of Pharmaceutical Sciences, Universiti Sains Malaysia were asked to review the questionnaire in terms of its clarity, relevance, and ease to understand the questions. The response from these reviewers was considered before finalizing the questionnaire.Before the implementation of the actual survey, the questionnaire was pilot tested on 30 HCPs, who were excluded from the actual study. This helped in the reliability assessment of the questionnaire by calculating Cronbach’s coefficient alpha. The internal consistency of the questionnaire was 0.72.The survey-based data was collected from the HCPs working in tertiary care public hospitals of Lahore, Pakistan during September 2018 to January 2019. The self-administered questionnaire was distributed among HCPs. A statement about the research project and consent form was also distributed. It was later on collected together with the filled questionnaire from the participants. The response from the participants was entirely anonymous and voluntary.p value < 0.05 was considered as significant [The data from the questionnaire was added directly to the Statistical Package for the Social Sciences (SPSS) version 24 (IBM 2013) and rechecked for any incorrect entry. To describe demographic variables, descriptive statistics were used, percentages and frequencies were used to express categorical variables. Data that emerged from domains using the Likert scale were found non-parametric. The association between the demographics of the respondents and knowledge, attitude, practices were calculated using the Kruskal Wallis test, whereas the nificant .n = 384) questionnaires were distributed to HCPs (based on inclusion criteria). The returned questionnaires were collected by the researcher with a response rate of 90.10% (n = 346) for all participating HCPs. The majority of the HCPs who contributed for this survey were nurses, 56.9% (n = 197), followed by physicians, 32.4% (n = 112) and pharmacists, 10.7% (n = 37). Most participating HCPs were females, i.e., 82.9% (n = 287), while only 17.1% (n = 59) of males responded to the survey. About 56.9% (n = 197) of participants belong to the age group of 31–35, while only 1.2% (n = 4) were from age above 45 with an overall mean age of 32.30 years. About 70.8% (n = 245) participants were graduates and only 19.7% (n = 68) had specialization in their field. Majority of participating HCPs, i.e., 51.4% (n = 178), had a job experience of 5 years, while 3.2% (n = 11) had an experience of more than 20 years, with an overall mean experience of 4.31 years. The demographics of the participants are given in Table A total of 384 (n = 95), pharmacists, 89.18% (n = 33) and nurses, 82.23% (n = 162) gave correct response of basic definition of ADR. Most of the physicians and nurses gave incorrect responses regarding whether if the serious ADRs are known before the marketing of a new drug. While most pharmacists responded correctly, and the association was found significant with a p value < 0.005. About 60.40% nurses, (n = 119) gave correct answer that ADRs should be reported only about the suspected drugs, while 57.14% physicians (n = 64) and 48.64% pharmacists (n = 18) responded otherwise, and a negative association was found (p < 0.005) with physicians to the response (Table n = 68), 64.86% pharmacists (n = 24) and 63.45% nurses (n = 125) responded incorrectly, that previously reported ADRs by manufacturers should not be reported  were agreed, that minor reactions should also be reported. Most participants considered that reactions to old drugs, to newly introduced drugs in the market or reaction in the special population should also be reported, see Fig. A large majority of the physicians, pharmacists, and nurses were of the view that suspected reactions (suspected drug is uncertain), reaction causing hospitalization, persistent disability or death should be reported Table . The majn = 30) responded correctly to the definition. The majority of physician 70.53% (n = 79), pharmacists 91.89% (n = 34), and nurses 60.40% (n = 119) responded with the right answer that pharmacovigilance ensures the safety of drugs, as given in Table n = 27) were aware that DRAP is responsible for the monitoring of ADRs in Pakistan, see Fig. When participants were asked about the definition of pharmacovigilance, among all, majority of the pharmacists 81.08% , 56.7% pharmacists  and 41.6% nurses (n = 82) agreed that ADR reporting related information can be better learnt during the internship/training/clinical posting. When participants were asked, if their educational background has provided enough information about ADR reporting, 42.9% of physicians  showed disagreement over it, but 54.3% nurses (n = 107) and 48.6% pharmacists (n = 18) agreed with the statement. About 49.1% of physicians , 70.2% pharmacists (n = 26) and 76.1% nurses (n = 150) showed a positive attitude that they are the most important HCPs to report an ADR. Majority participants, i.e., 64.3% of physicians  emphasized that consulting other colleagues is important before reporting an ADR. Of all, 77.7% physicians (n = 87), 75.7% pharmacists (n = 28) and 68% of nurses (n = 134) had a positive attitude that ADR reporting is a professional obligation, as given in Table n = 55), 51.4% pharmacists (n = 19) and 41.1% nurses (n = 81) had a positive inclination that the reporting of an ADR should be mandatory for all HCPs. Similarly, 41.1% physicians (n = 46) and 48.6% pharmacists (n = 18) and 38.1% nurses (n = 75) had positive attitude that ADR should be confirmed before reporting to the hospital management. A large majority of the participants, i.e., 84.9% physicians (n = 95), 81.1% pharmacists (n = 30) and 81.7% nurses (n = 161) positively agreed that workplace should encourage HCPs about ADR reporting, as illustrated in Fig. The majority participants considered ADR reporting as a way to improve the safety of medicines. About 35.7% physicians n = 0, 56.7% n = 89, p < 0.005) and 58.4% nurses (n = 115) stated that they did not report any ADR, while 67.6% of the pharmacists (n = 25) stated that they reported ADRs in their workplace. Similarly, only 18.8% physicians (n = 21) and 23.4% nurses (n = 46) reported ADRs in the last 12 months, contrary to this, 59.5% of pharmacists (n = 22) reported ADRs in the last 12 months. About 83.9% physicians (n = 94), 62.2% pharmacists (n = 23) and 69.5% nurses (n = 137) never kept records of ADR. Only 9.8% physicians (n = 11), 18.9% pharmacists (n = 7) and 15.7% nurses (n = 31) sent a suspected ADR reports to the DRAP or to the drug manufacturer, see Table When respondents were asked about their practices regarding ADR reporting in hospitals, 79.5% physicians (n = 75), 77.2% nurses (n = 152), reported the ADRs verbally, during the discussions/meetings, while 56.8% pharmacists (n = 21) did not verbally report about ADRs. About 56.3% physicians (n = 63) and 91.9% pharmacists (n = 34) reported an ADR on the ADR reporting forms, while 50.8% nurses (n = 100) did not report on ADR forms, available in the hospitals. Among all participants 67.9% physicians (n = 76), 81.1% pharmacists (n = 30) and 67.5% nurses (n = 133) reported to hospital pharmacy about an ADR. Similarly, 72.3% physicians (n = 81), 62.2% pharmacists (n = 23) and 70.1% nurses (n = 138) reported ADRs directly to the hospital management. Moreover, 58% physicians (n = 65) and 64.9% pharmacists (n = 24) reported ADRs to the relevant manufacturer, while 55.3% nurses (n = 109) did not report to the manufacturer. Similar trend was observed in the case of 64.3% physicians (n = 72), 64.9% pharmacists (n = 24) and 73.6% nurses (n = 145) who did not reported ADR through the online portal, as shown in Table When respondents were asked about the modes of ADR reporting in hospital, 67% physicians  by Uppsala Monitoring Centre (UMC). In many member countries, the hospitals have a pharmacovigilance center responsible for the monitoring and recording of ADR. These centers then communicate about ADRs to the national pharmacovigilance center and ultimately to the UMC in Sweden receives the ADR reports. In our study, among all HCPs, only the pharmacists had good knowledge about pharmacovigilance system, recently established under the umbrella of DRAP  and wereThe attitudes of HCPs are considered pivotal for the reporting of an ADR, thus a positive attitude may encourage the prompt reporting of an ADR. The current study explored the attitudes of all HCPs towards pharmacovigilance activities and showed that overall, all HCPs had a positive attitude towards pharmacovigilance activities in general and ADRs reporting in particular. Although the majority of the participants did not report ADRs in their work setting, they expressed their willingness to report, provided, they are motivated to perform such activities . Among aMany low- and middle-income countries are facing the challenge of low ADR reporting, as low ADR reporting generates minimum signals and thus lacks the pharmacovigilance data . The parParticipants in our study identified different modes of ADR reporting, including both verbal and non-verbal. It was seen in our survey that the majority of the participants including physicians and nurses have verbally reported ADRs to the concerned person or hospital pharmacy or hospital management, while some physicians and majority pharmacists reported an ADR on the ADR reporting forms. A study conducted in Northern Nigeria has found that healthcare professionals working in a tertiary care setting had similar practices of reporting an ADR . It was Though all HCPs help in improving ADR reporting, but hospital pharmacists can play a crucial role in this regard, as the majority of ADRs occur in a hospital or could lead to hospitalization . Being aThe study was carried out in Lahore; therefore, the results of the present study cannot be generalized to represent the other provinces in the country. However, Punjab is the most developed province and Lahore with a population of over 15 million people (highly populated city of Punjab province) it is the second biggest city in the country. Hence it is expected that the results in the other parts would not be very different.The findings from the present study suggest that medicines safety related knowledge and ADR reporting-based training should be provided to all HCPs. To do so, a system of hands-on training and workshops regarding dealing with ADR events must be established at hospital level. Besides, collaboration between academic institutes, drug manufacturers, drug regulatory authority and HCPs should be increased to sensitize ADR reporting related practices.The current study has shown that among all HCPs, pharmacists had better knowledge about ADR reporting and pharmacovigilance. All HCPs had a positive attitude and inclination towards reporting an ADR, but the discrepancies were observed in the practices related to ADR reporting. Based on the above, strategies are needed to educate, train and empower the HCPs in the area of pharmacovigilance.


Staphylococcus aureus, explore the anti‐biofilm effects of LL‐37, and compare the effects of the cationic peptide LL‐37 and Cefalexin in treatment of postoperative infection of femoral fracture in vivo.Antimicrobial peptides are widely present in nature, with many of the antimicrobial peptides having antimicrobial activity against Gram‐positive and Gram‐negative bacteria, fungi, parasites, and even coated viruses. Internal fixation of fractures is a reliable technique. However, the fracture is difficult to heal and internal fixation is not easy to maintain after infection. This study aims to verify the antibacterial effect of cationic peptide LL‐37 on Staphylococcus aureus was clinically isolated from one patient with clinical infection after the fracture fixation at Wuxi 9th People's Hospital. The cationic peptide LL‐37 was synthesized by Shanghai Apeptide Co. Ltd. To compare the effects of the cationic peptide LL‐37 and Cefalexin in the treatment of postoperative infection of femoral fracture in vivo, 63 rabbits with internal fixation of femoral fractures were inoculated intravenously with clinically isolated pathogenic bacteria suspensions. Rabbits in the treatment groups were treated with peptide LL‐37 and Cefalexin after surgery. Rabbits in the control groups were treated with physiological saline after surgery. The biofilms on internal fixtures were harvested from euthanized rabbits 1 h, 12 h, 1 day, 2 days, and 7 days after injection of LL‐37, Cefalexin, or saline and calculated by colony count. The biofilms from treatment and control groups at 7 days were analyzed by fluorescence microscopy. Blood samples were collected at 1 h, 12 h, 1 day, 2 days, and 7 days following peptide LL‐37 and Cefalexin injection.The t‐test or two‐way analysis of variance (ANOVA). Cationic peptide LL‐37 showed significant inhibitory effects on clinically isolated Staphylococcus aureus (P < 0.05) compared with Cefalexin and control group at 1 day (P = 0.021), 2 days (P = 0.019), and 7 days (P = 0.025). Fluorescent images of the biofilm reveal that the numbers of cells on biofilms are far less than those in the Cefalexin and control groups at 7 days. The levels of Interleukin‐6 (IL‐6), tumor necrosis factor‐α (TNF‐α) and C‐reactive protein (CRP) reached a maximum at 2 days following the operation. After the injection of LL‐37, there was an increase in the serum IL‐6, TNF‐α, and CRP contents in rabbits in both groups, however from 1 day postoperative the level of IL‐6 (P = 0.034), TNF‐α (P = 0.043), and CRP (P = 0.039) decreased significantly compared to the Cefalexin and control group. At 7 days postoperative, the level of IL‐6 (P = 0.029), TNF‐α (P = 0.033), and CRP (P = 0.027) had reverted to normal levels in LL‐37 groups.The results were compared statistically using Student's in vivo.Cationic peptide LL‐37 may be a promising agent to control internal femoral fracture fixation infection  Fracture is a disruption or break in the continuity of the structure of a bone. Fracture occurs mainly in the elderly and children. Most patients can restore the original function of bone with timely and appropriate treatment; however, some patients will have potentially different outcomes. The application of internal fixation technique for fractures in clinical settings is very common, and although advanced equipment and necessary aseptic conditions have been developed, postoperative infection of the fractures undergoing internal fixation is still an inevitable complicationInternal fixation of fractures is a reliable techniqueStaphylococcus aureus is the important pathogen of nosocomial infection and the leading cause of biofilm infection. Staphylococcus aureus easily adheres to the surface of orthopaedic implants, therefore, seeking an effective drug to resist biofilm in the treatment of orthopaedic infection is really urgent.In recent years, while the use of antibiotics in large doses has reduced infection rates, it has also promoted development of drug resistance of pathogenic microorganismsin vitroin vivo. Therefore, to explore the cationic peptide LL‐37 in the control of internal femoral fracture fixation infection in vivo seems really meaningful in the orthopaedic treatment of clinical infection.In finding an effective weapon against these infectious diseases, the scientists have focused on the exploration of antibacterial peptides. Antibacterial peptides are one kind of natural protein, which can kill bacteria, viruses and fungi, and researchers have been trying to use antibacterial peptides to replace traditional antibiotics. Natural antibacterial peptides usually consist of 20 kinds of amino acids and the antibacterial peptides can exert a specific function through the adjustment of the order of amino acids. Antimicrobial peptides are widely present in nature, with many of the antimicrobial peptides having antimicrobial activity against Gram‐positive and Gram‐negative bacteria, fungi, parasites, and even coated virusesStaphylococcus aureus; and (ii) compare the effects of the cationic peptide LL‐37 and Cefalexin in the treatment of postoperative infection of femoral fracture in vivo and (iii) invest the improvement of inflammatory factor after the injection of cationic peptide LL‐37, Cefalexin, and physiological saline. In order to verify these effects we isolated one S. aureus strain from one patient with clinical infection after the fracture fixation, analyzed its biofilm formed by clinically isolated S. aureus in infected femoral fracture of rabbit model, and measured the serum concentrations of IL‐6, TNF‐α, and CRP in rabbits with S. aureus infections.We aim to: (i) verify the antibacterial effect of cationic peptide LL‐37 in clinically isolated The project was approved by the Ethical Committee of Wuxi 9th People's Hospital affiliated to Soochow University. One patient with clinical infection after fracture fixation at Wuxi 9th People's Hospital was selected to extract the clinical pathogenic bacteria. Bacterial specimens were obtained after washing the surface of the wound vigorously with saline, followed by debridement of superficial exudates, and were cultured under microaerophilic conditions for 7 days.The isolated stock culture strains were subcultured onto MacConkey agar plates to check their purity. The turbidity of the bacterial suspensions was adjusted with a densitometer to a McFarland 0.5 standard in 0.45% sterile sodium chloride solution. The suspensions were then transferred to VITEK 2 ID‐GNB cards and AST‐No. 12 cards and manually loaded into the VITEK 2 system. The data was analyzed by the VITEK 2 system and the results were reported automatically.Cationic peptide LL‐37 (LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES) was synthesized by Shanghai Apeptide Co. Ltd. . LL‐37 was purified by high‐performance liquid chromatography and the identity was verified by SDS‐PAGE. The purity of LL‐37 (>95%) and mass were confirmed by electrospray ionization mass spectrometry.A total of 63 New Zealand white rabbits (weighing 2.0 to 2.5 kg) were used in this study. Twenty‐one rabbits in the control groups were treated with physiological saline after surgery, and 21 rabbits in the treatment groups were treated with LL‐37 and Cefalexin after surgery.via inhalation of isoflurane (2%). The surgical procedure was as described in the previous work with few modificationsRabbits were anesthetized S. aureus (1 × 105 cfu/mL) in 2 mL of normal saline was pipetted into the femur space containing the cut end of the implantAn inoculum of clinically isolated To evaluate biofilm formation on the femoral fracture internal fixation in our rabbit model, the biofilms on internal fixtures were harvested from euthanized rabbits 1 h, 12 h, 1 day, 2 days, and 7 days after injection of LL‐37, Cefalexin, or saline. Each sample was washed in 10 mL of phosphate‐buffered saline (PBS) and then 0.1 mL of the suspension were inoculated and counted for the colonies on Columbia Blood Agar Base.After 7 days, internal fixtures were extracted and fixed with 2.5% glutaraldehyde at 4°C for 1 h. After fixation, the internal fixtures were stained using BacLight Live/Dead staining kit  and observed under a Nikon 80i fluorescence microscope at an excitation wavelength of 488 nm.Blood samples were collected 1 h, 12 h, 1 day, 2 days, and 7 days after injection of peptide LL‐37, Cefalexin, or application of saline. The samples were centrifuged at 4000 r/min for 3 min and stored at −70°C. The serum interleukin‐6 (IL‐6), tumor necrosis factor‐α (TNF‐α), and C‐reactive protein (CRP) levels were analyzed using ELISA and expressed as pg./mg proteint‐test or two‐way analysis of variance (ANOVA). Values were considered statistically significant at P < 0.05.All tests were repeated three times for consistency of results. SPSS 14.0 software for Windows was used for data analysis. The results were compared statistically using Student's S. aureus by using VITEK 2 fluorescent system. At 1 day following injection with peptide LL‐37, the colony count of the biofilm began to show the significant changes as compared to the Cefalexin and saline groups , TNF‐α (P = 0.043), and CRP (P = 0.039) decreased significantly compared to the Cefalexin and control group , TNF‐α (P = 0.033), and CRP (P = 0.027) has reverted to normal levels in LL‐37 groups.The levels of IL‐6, TNF‐α, and CRP reached a maximum at 2 days following the operation. After the injection of LL‐37, there was an increase in the serum IL‐6, TNF‐α, and CRP contents in rabbits in both groups; however, from 1 day the level of serum IL‐6 P = 0.03, TNF‐α (Staphylococcus aureus and S. pyogenesOrthopaedic clinical infection presents a difficult problemS. aureus often leads to sepsis, colonizes catheters, causes pneumonia and wound infections, and is still one of the most common pathogenic bacteria in fracture infectionsS. aureus were found in 58% of the infections after internal fixationStaphylococcus can exist in the internal fixation of the surface for a long time, especially in the power screw in the large surface area, which can get implanted in bone tissue and gradually breed until they give rise to clinical symptoms. Although the incidence of orthopaedic clinical infections has been significantly reduced by using antibiotics, multidrug resistant bacteria pose a significant problemS. aureus infections may be complicatedS. aureus is sensitive, is necessary.Bioglass is one of the most important inorganic biomedical materials and has been used in orthopaedic treatment as non‐antibiotic approachesCytokines are small molecule polypeptide proteins produced by the body's immune or non‐immune cells, which are closely associated with hematopoietic, inflammatory, and immune reactionsin vivo.In our study, we isolated the pathogenic bacteria from only one person, however, the etiology of infection of femoral fracture after internal fixation is so complicated that the result is affected by a lot of factors on the surface of the material, and needs further research. Meanwhile, our results just showed biofilm formation and serum levels; more importantly, histological sectioning as well as radiological findings are needed. In the future, we hope to consider all aspects of different factors when conducting experiments in vivo.Cationic peptide LL‐37 could significantly reduce biofilm formation, modify biofilm composition and levels of inflammatory factors. These results indicate that LL‐37 may be one of the promising compounds in the control of femoral fracture infection after internal fixation


N = 547). Specifically, multimodal magnetic resonance imaging features extracted from voxel‐wise amplitude of low‐frequency fluctuations, gray matter volume, and fractional anisotropy were used to build an individual VWM capacity prediction model through a machine learning pipeline, including the steps of feature selection, relevance vector regression, cross‐validation, and model fusion. The resulting model exhibited promising predictive performance on VWM , and identified features within the subcortical‐cerebellum network, default mode network, motor network, corpus callosum, anterior corona radiata, and external capsule as significant predictors. The main results were then compared with those obtained on emotional regulation and fluid intelligence using the same pipeline, confirming the specificity of our findings. Moreover, the main results maintained well under different cross‐validation regimes and preprocess strategies. These findings, while providing richer evidence for the importance of multimodality in understanding cognitive functions, offer a solid and general foundation for comprehensively understanding the VWM process from the top down.The indispensability of visual working memory (VWM) in human daily life suggests its importance in higher cognitive functions and neurological diseases. However, despite the extensive research efforts, most findings on the neural basis of VWM are limited to a unimodal context (either structure or function) and have low generalization. To address the above issues, this study proposed the usage of multimodal neuroimaging in combination with machine learning to reveal the neural mechanism of VWM across a large cohort , and identified features within the subcortical‐cerebellum network, default mode network, motor network, corpus callosum, anterior corona radiata, and external capsule as significant predictors. Our findings, while providing richer evidence for the importance of multimodality in understanding cognitive functions, offer a solid and general foundation for comprehensively understanding the VWM process from the top down.We used multimodal neuroimaging in combination with machine learning to reveal the neural mechanism of visual working memory (VWM) across a large cohort. The resulting model exhibited promising predictive performance on VWM ( It is fundamental in human daily life for being indispensable from simple behaviors like direction identification and cash payment to more complex ones like web search and test taking. Besides, VWM is strongly correlated with cognitive ability . In particular, ALFF measures the spontaneous fluctuations in brain gray matter and can reflect the functional activity of brain. GMV and FA quantify the volume of gray matter and the integrity of white matter, respectively, and thus offer descriptions of voxel‐wise attributes of brain structure. Second, as stated before, there has been abundant evidences supporting the strong associations of these three indicators with VWM. Specifically, it has been found that the ALFF values of the relevant brain regions  may have distinct roles in VWM processes . The R‐fMRI, T1 weighted image, DWI and VWM score data from 625 healthy adults were used in the current study. The data and the ethical approval for this study were obtained by the Cambridgeshire 2  Research Ethics Committee and written informed consent was obtained from each participant.All participants were from Stage 2 Cam‐CAN study  3D images were acquired using the following parameters: TR/TE = 2,250 ms/2.99 ms; inversion time (TI) = 900 ms; flip angle = 9°; FOV = 256 × 240 × 192 mm3; voxel size = 1 × 1 × 1 mm3; and GRAPPA acceleration factor = 2. The acquisition lasted for 4 min and 32 s. The twice‐refocused spin‐echo axial DWI were acquired using following parameters: 30 diffusion gradient directions for each of two b‐values: 1,000 and 2,000 s/mm2, along with three images acquired using b‐value of zero; TR/TE = 9,100/104 ms; voxel size = 2 × 2 × 2 mm3; FOV = 192 × 192 mm2; slices = 66; number of averages = 1. This acquisition lasted for 10 min and 2 s.All participants were scanned at the Medical Research Council Cognition and Brain Sciences Unit on a 3 T Siemens TIM Trio System with a 32‐channel head coil. The R‐fMRI was collected axially using a gradient‐echo echo‐planar imaging sequence. The image parameters were as follows: repetition time (TR)/echo time (TE) = 1,970 ms/30 ms; flip angle = 78°; field of view (FOV) = 192 × 192 mmAll the participants finished a modified version of a previous experiment , Resting‐State fMRI Data Analysis Toolkit   coregistration to the individual average functional image, (b) segmentation into gray matter, white matter, and cerebrospinal fluid, (c) modulating the resulting normalized gray matter images with Jacobian determinants of the deformation parameters, (d) smoothing GMV image with an 8 mm FWHM Gaussian kernel, and (e) resampling to 3 mm isotropic voxels to match the same voxel size as preprocessed R‐fMRI data. Thus, for each participant, there was a GMV map.https://fsl.fmrib.ox.ac.uk/fsl/fslwiki) with the standard preprocessing procedure , there were 547 participants  left for subsequent analyses. The demographics and behavioral data of the participants are summarized in Table 2.4Nvoxels = 57,806) generated by a threshold of 0.2 on a priori gray matter probability map in SPM and the white matter mask  generated by a predefined FA skeleton mask in FSL (FMRIB58_FA‐skeleton_1mm.nii). Then, for each participant, the ALFF values were extracted from the ALFF map and the GMV values were extracted from the GMV map, both of which were constrained within the gray matter mask. The FA values were extracted from the FA map, which was constrained within the FA skeleton mask. Hence, for each participant, there were 57,806 ALFF features; 57,806 GMV features; and 137,832 FA features. Second, the RVR method  by using the z‐score approach on each modality separately. Then the Pearson correlations of the normalized features with the VWM scores were computed within the training sample. For each modality, the features with correlation significance beyond a corrected threshold  were selected. Using the selected features in one modality, the RVR method was applied to train a mono‐modality model that predicted the VWM score. Three mono‐modality models, namely the ALFF, GMV, and FA models, were thus obtained. We then built four multimodality models by bagging the three mono‐modality models into three bimodality models  and one tri‐modality model . Using the bagging strategy of model averaging  were normalized by the z‐score approach using means and standard deviations acquired from the training set, as suggested in previous studies  was performed. In each permutation, the above modeling procedure was repeated with the association between the features and the observed VWM scores randomized  in Steiger , which is a simple yet effective machine learning technique and is widely used to build baseline model  between the observed and the predicted VWM scores of all the 547 participants. Specifically, the MAE was calculated as the mean of absolute differences between the observed and the predicted VWM scores. To validate the significance of the correlation, a permutation test , the SPM12 toolbox (http://www.fil.ion.ucl.ac.uk/spm/), and our in‐house MATLAB code.All the above analyses were implemented with the Pattern Recognition for Neuroimaging Toolbox (2.5N = 100) were used to avoid potential overoptimistic bias induced by LOOCV. In terms of preprocessing strategies, we considered the following three aspects. (a) Global signal regression. Previous studies have suggested that whether to remove the global signal or not might significantly alter the research results , the tri‐modality model ALFF + GMV + FA , and the mono‐modality model GMV  achieved the best predictive power, while the other models achieved prediction performance with the empirical‐predicted correlation r∈ (p < .001) and MAE∈. The Steiger's method , but was significantly better than the other four models (p < .05). Further, when tested in the repeated 5‐ and 10‐fold cross‐validation frameworks (N = 100), the tri‐modality model achieved the highest predictive power with the smallest variance across repetitions ; 36,366 GMV features ; and 19,918 FA features  were used. Figure In the ultimate prediction model, 118 ALFF features p < .0/57,806; r = .404 ± 0.013, p < .001, MAE = 0.602 ± 0.005, Supplementary Table r = .408 ± 0.010, p < .001, MAE = 0.601 ± 0.004, Supplementary Table r ≤ .414, p < .001, 0.599 ≤ MAE ≤0.605) given that the global signal was not removed or the TBSS strategy was used. It was only in the case of motion scrubbing could the mono‐modality model GMV  slightly outperform the tri‐modality model , while the bi‐modality model GMV + FV remained to be the best one . In the aspect of feature analyses, with the repeated 5‐ and 10‐fold cross‐validation, the number of selected features for GMV and FA dropped to around 400 and 200, respectively  and preprocessing strategies . Regarding the two different cross‐validation regimes, we found that the advantage of multimodality models against mono‐modality models maintained, with the tri‐modality model achieving the best performance in both cases , and that between the predicted and the actual FI scores was 0.656 (p < .001). Most importantly, the discriminative features drawn from the prediction models for the three cognitive functions were totally different  than all the other models except for the two involving the GMV features . This phenomenon indicates that GMV is a prominent indicator for distinguishing individual VWM capacity, which is in line with the results of former studies . Moreover, we identified several key predictors to individual VWM capacity from both functional and structural perspectives, which can serve as potential neuromarkers of VWM and might provide further implications for future related research.In summary, we combined multimodal imaging data of healthy participants to investigate individual VWM capacity using a well‐designed machine learning pipeline that included feature selection, RVR, cross‐validation, and model fusion. Using voxel‐wise features extracted from the whole brain in three modalities, we can predict individual VWM capacity with correlation at 0.402 (The authors declare no conflict of interest.All study data were obtained from repositories of publicly available data. These data were originally acquired with written informed consent of all studied participants.AppendixS1. Supporting Information.Click here for additional data file.


Psychopathology is prevalent in Williams (WS), fragile X (FXS) and Prader–Willi (PWS) syndromes. However, little is known about the potential correlates of psychopathology in these groups. A questionnaire study was completed by 111 caregivers of individuals with WS (n = 35); FXS (n = 50) and PWS (n = 26). Mean age was 26 years (range 12–57 years); 74 (67%) were male. Multiple regression analyses indicated that higher rates of health problems and sensory impairments predicted higher psychopathology in WS (p < .0001). In PWS, poorer adaptive ability predicted higher overall psychiatric disturbance (p = .001), generalised anxiety (p = .006) and hyperactivity (p = .003). There were no significant predictors in FXS. This study highlights dissociations in the potential risk markers of psychopathology between genetic syndromes. Implications for intervention are discussed.The online version of this article (10.1007/s10803-019-04317-1) contains supplementary material, which is available to authorized users. The prevalence of psychopathology in individuals with intellectual disabilities (ID) is estimated as being four to five times higher than in the general population (Matson and Shoemaker In this study we examine the nature and correlates of psychopathology in three syndromes that are broadly comparable for degree of ID: Williams (WS), fragile X (FXS) and Prader–Willi syndromes (PWS). WS is caused by a microdeletion of approximately 28 genes on chromosome 7q11.23, with an estimated prevalence of up to 1 in 7500 births , health problems and auditory sensory processing. Our focus was specifically on adolescents and adults because of the high rates of mental health problems in these age groups.To describe the profile of psychopathology in individuals with WS, FXS and PWS and identify differences in profiles between the syndromes.The aims of the study were:2)To assess whether age, adaptive ability, health difficulties and auditory sensory processing impairments are associated with psychological disturbance within syndromes.We hypothesised that social anxiety would be significantly higher in FXS; generalised anxiety would be higher in FXS and WS compared to PWS; obsessive–compulsive behaviour would be higher in PWS.We hypothesised that health difficulties and auditory sensory processing impairments would be positively associated with psychopathology for all groups.www.williams-syndrome.org/registry), administrated by the Williams Syndrome Association, USA.Participants were recruited from the Cerebra Centre for Neurodevelopmental Disorders (CCND) participant database . Participants were also recruited from the Fragile X Society, the Williams Syndrome Foundation in the UK, the Canadian Association for Williams Syndrome and the Williams Syndrome Registry . There were 35 participants in the WS sample , 49 in the FXS sample  and 26 in the PWS group .A total of 132 families participated in the survey. Participants were included if they had completed at least 25% of the questionnaire survey  and the person they cared for was over the age of 12 (2 excluded). Inclusion criteria were a confirmed genetic diagnosis from a clinician  or a positive result from prior genetic testing. Individuals with mobility difficulties (n = 3) and limited verbal ability  were excluded to ensure better group matching. Due to the tendency for higher cognitive functioning in females with FXS . and DBC intensity score  (full table of correlations available in Online Resource B).In this study, the ADAMS subscales were highly correlated with each other for every syndrome group, indicating high interrelatedness between categorical constructs (Online Resource A). Correlations between the ADAMS subscales and the DBC mean item and intensity score were also assessed. Overall, the measures were highly correlated, with the ADAMS total score significantly correlating with DBC mean item score  and epsilon squared  = 43.02, p = <.0001, Cramer’s V = .63, with the FXS group including more males (100%) than the WS (40%) and PWS (42%) groups. Auditory sensory processing scores were significantly different between the groups (F(2107) = 19.67, ηp2 = .27, p < .0001). Pairwise comparisons revealed that individuals with WS (mean = 18.69) and FXS (17.69) had higher auditory processing impairments than individuals with PWS (13.19). Group differences were also found for overall lifetime health (χ2(2) = 23.90, ε2 = .22, p < .0001), with WS (74.74) scoring higher for health difficulties compared with FXS (40.52). For overall current health problems, significantly more health problems were found in WS (66.80) and PWS (62.85) compared with FXS (42.15) (χ2(2) = 15.15, ε2 = .14, p = .001).Prior to other analyses, group differences in auditory sensory processing scores, health, age, gender, adaptive ability and psychopathology were examinedTable . There w2(2) = 9.61, ε2 = .09, p = .008) with FXS (60.29) and WS (61.34) scoring higher than PWS (38.62), 2) manic/hyperactive (χ2(2) = 10.16, ε2 = .09, p = .006) with FXS (65.87) scoring higher than PWS (43.02), and 3) social avoidance (χ2(2) = 37.13, ε2 = .34, p < .0001) with FXS (76.09) scoring higher than WS (39.17) and PWS (38.67). On the DBC-A, only the social relating subscale differed between groups (χ2(2) = 26.96, ε2 = .32, p < .0001). Individuals with FXS (58.14) scored higher for social relating difficulties than individuals with PWS (34.93) and WS (27.28). There were no other differences between the groups on either measure.To examine the profile of psychopathology in WS, FXS and PWS, group differences were assessed using the ADAMS and DBC assessment measures. Based on the ADAMS, general anxiety was the highest scoring domain for individuals with WS , social avoidance for individuals with FXS  and depressed mood for those with PWS  . Table To assess whether the full model of individual variables considered together was associated with psychological disturbance within syndromes, hierarchical regressions were conducted. The ANOVA models for the hierarchical regressions are presented in Table 2 = .28). For all other subscales, age and adaptive ability in Step 1 did not produce significant results, though the addition of health and sensory processing to the model in Step 2 led to statistically significant increases in the predictive ability of the model, which accounted for an additional 31–48% of the variance.In WS, the overall model of predictors was significant for every outcome variable, except for the obsessive–compulsive subscale  = 2.20, p = .081, RThe influence of the individual predictors  on the psychopathology scales is illustrated in Fig. For PWS, none of the overall models were significant and the addition of the predictors in Step 2 accounted for an additional 11–28% of the variance. For the individual predictors, higher adaptive ability was the only predictor of psychopathology and significantly predicted a lower total score , lowered generalised anxiety  and manic/hyperactivity  on the ADAMS.2 = .38). The addition of health and sensory processing in Step 2 accounted for an additional 3–20% of the variance across the outcome variables. The predictors for FXS differed from those for WS and PWS, with no associations identified for age, adaptive ability, health and auditory sensory processing.Only the overall model for the DBC mean item score for FXS was significant,  = 5.01, p = .001, RWith regard to overall level of psychopathology, auditory sensory processing impairments and a higher number of health problems were associated with general psychiatric problems in individuals with WS; poorer adaptive ability was associated with psychopathology for individuals with PWS. No associations were identified between any of the individual predictors and psychopathology for individuals with FXS.This study is the first to examine differences in internalising psychopathology and potential predictors of general psychopathology and specific symptoms in individuals with WS, FXS and PWS. These syndromes were chosen for comparison as they are broadly comparable for degree of ID. Exploratory analyses (p < .01) focused on associations with age, adaptive ability, health and auditory sensory processing. To examine the impact of methodological assessment on the results obtained, both dimensional and categorical measures were used. The study identified cross-syndrome differences in: (1) health and auditory sensory processing, (2) psychopathology generally and (3) correlates of psychopathology .Profiles of psychopathology and associated factors differed across groups. Higher rates of auditory sensory processing impairments were found for individuals with WS and FXS compared with PWS; health difficulties were more prevalent in WS and PWS compared with FXS . Consistent with our hypotheses and past research, generalised anxiety was significantly higher in FXS and WS compared with PWS, and both ADAMS and DBC social subscales were elevated in FXS compared with WS and PWS, although effect sizes were small. Our hypothesis that individuals with PWS would score higher on the compulsive behaviour subscale of the ADAMS was not supported. The high proportion of adults in this study (80% aged over 18) may account for this non-significant finding, as there is some evidence that compulsive behaviour decreases with age in PWS Dykens .Cross-syndrome comparisons of correlates of psychopathology were also examined. Age was not predictive of psychopathology for any of the syndrome groups, consistent with several studies in the literature Below is the link to the electronic supplementary material.


Atherogenic lipoprotein profile of plasma is an important risk factor for atherosclerosis. The atherogenic index of plasma (AIP) has been suggested as a novel marker for atherosclerosis.AIP is a useful marker of advanced subclinical coronary artery disease (CAD) in subjects without overt renal dysfunction.2 evaluated by coronary computed tomography angiography (CCTA) for health check‐up were included. The relation of AIP to advanced CAD  was evaluated.A total of 6928 subjects with estimated glomerular filtration rate > 60 mL/minutes/1.73 mP < .001) increased with elevating AIP quartiles. Higher AIP (per 0.1 unit increase) was associated with an increased risk of CACS >100  1.010 to 1.106, P = .017; relative risk (RR) 1.048, 95% CI 1.009‐1.089, and P = .015) and OCP  after adjusting for age > 60 years, male sex, hypertension, diabetes mellitus, dyslipidaemia, obesity, and proteinuria.All participants were stratified into four groups based on AIP quartiles. The prevalence of CACS >100 (group I [lowest] 4.7% vs group II 7.0% vs group III 8.8% vs group IV 10.0%) and OCP (group I 3.7% vs group II 6.4% vs group III 8.8% vs group IV 10.9%) (all AIP is independently associated with advanced subclinical CAD beyond traditional risk factors. The atherogenic index of plasma (AIP) has been suggested as a marker of plasma atherogenicity based on its strong and positive association with cholesterol esterification rates, lipoprotein particle size, and remnant lipoproteinaemia.Coronary artery calcium score (CACS) has been regarded as a useful marker of coronary atherosclerosis because it represents the degree of atheromatous plaque burden.22.12 (n = 958), and (c) insufficient medical records for traditional risk factor and AIP calculation or poor image quality (n = 693). As a result, 6928 participants were finally included in the present study. The study protocol was approved by the ethics committee of severance CV Hospital, and informed consent for the procedure was obtained from each participant. Patients or the public were not involved in the design, conduct, reporting, or dissemination of our research.This observational, retrospective, single‐center registry consisted of 8648 consecutive participants evaluated by CCTA for health check‐up with 64‐slice multidetector computed tomography between January 2004 and April 2009 at Severance CV hospital.2 based on the cutoffs for Asian population. Proteinuria was defined based on the sex‐specific urinary albumin/creatinine ratio.Medical history of hypertension, diabetes mellitus, and dyslipidaemia was systematically acquired. Height, body weight, and blood pressure (BP) were measured during visits. Height and weight measurements were obtained while the subjects wore light clothing without shoes. Body mass index (BMI) was calculated as weight in kilograms divided by the square of height in meters. After the participants rested for ≥5 minutes, the BP was measured at the right arm using an automatic manometer with an appropriate cuff size. Serum levels of total cholesterol, triglycerides, high‐density lipoprotein cholesterol (HDL‐C), low‐density lipoprotein cholesterol (LDL‐C), glucose, and creatinine were measured after a minimum of 12 hours fasting period. The AIP was calculated as the base 10 logarithm of the ratio of the triglycerides to HDL‐C concentrations.2.22 within or adjacent to the vessel lumen, which were clearly distinguishable from the lumen and the surrounding pericardial tissue. Plaques without calcium were classified as noncalcified, those with calcified tissue involving ≥50% of the plaque area (density > 130 HU) were classified as calcified, and those with <50% calcium were classified as mixed plaques.During CCTA examination, subjects with an initial heart rate > 65 beats/minutes received a single oral dose of 50 mg metoprolol tartrate  1 to 2 hours before the examination if beta blocking agents were not contraindicated. A 64‐slice scanner  was used for examination. First, a nonenhanced prospective electrocardiogram (ECG)‐gated scan was obtained for the measurement of CACS with the following parameters: rotation time of 330 ms, slice width of 3.0 mm, slice collimation of 0.6 mm, tube current of 50 mA, tube voltage of 120 kV, and table feed/scan of 18 mm. Subsequently, CCTA was performed using retrospective ECG gating with the following scan parameters: rotation time of 330 ms, slice collimation of 64 × 0.6 mm, tube voltage of 100 to 120 kV, tube current of 600 to 800 mA depending on patient size, table feed/scan of 3.8 mm, and pitch factor of 0.2. ECG‐based tube current modulation was applied to 65% of the R‐R interval. A real‐time bolus‐tracking technique was used to trigger the initiation of the scan. The total estimated average radiation dose for the multislice protocol was 10.9 ± 1.9 mSv. Contrast enhancement was achieved with 60 mL iopamidol  injected at 5 mL/s, followed by an injection of 30 mL diluted contrast, and then 30 mL saline at 5 mL/s using an injector . The images were evaluated by two experienced cardiac radiologists. CACS, which was measured using the scoring system described by Agatston et al,2.32‐test or Fisher's exact test was used for categorical variables, as appropriate. Univariate logistic regression analysis was performed to identify the association between clinical variables and coronary atherosclerotic parameters. Multiple logistic regression analysis was used to identify the predictive value of AIP for advanced CAD. The forced entry method was used to enter independent variables into the multivariate regression analysis model. Receiver operating characteristic (ROC) curve analysis with Youden index was conducted to determine an optimal cutoff of AIP for predicting advanced CAD. We then compared the predictive significance of the optimal AIP cutoff for advanced CAD after considering traditional risk factors. All statistical analyses were performed with the Statistical Package for the Social Sciences version 19  and SAS version 9.1.3 . A P‐value <.05 was considered significant for all analyses.Continuous variables were expressed as mean ± SD. Categorical variables were presented as absolute values and proportions. To compare the characteristics among the AIP quartiles, one‐way analysis of variance was used for continuous variables, and the χ33.1The mean age of participants was 52.0 ± 9.7 years and 3977 (57.4%) of them were men. The prevalence of hypertension, diabetes mellitus, dyslipidaemia, obesity, and proteinuria was 28.8%, 11.2%, 53.8%, 38.0%, and 6.6%, respectively. The CACS >100 and OCP was observed in 7.6% and 7.4% of subjects, respectively. All participants were categorized into four‐groups based on the AIP quartiles. The ranges of the AIP in stratified groups of I (lowest), II, III, and IV (highest) were 0.60 to 0.14, 0.15 to 0.35, 0.36 to 0.55, and 0.56 to 1.79, respectively. Systolic and diastolic BP values, serum levels of total cholesterol, triglycerides, glucose, and creatinine, albumin/creatinine ratio, and the prevalence of male sex, hypertension, diabetes mellitus, dyslipidaemia, obesity, and proteinuria significantly increased with increasing AIP quartiles. In contrast, the mean level of HDL‐C significantly decreased with increasing AIP quartiles  and 0.33  0.593‐0.641, P < .001), respectively  and OCP  incrementally improved  0.581, 95% CI 0.556‐0.606, 3.4P = .017; relative risk (RR) 1.048, 95% CI 1.009‐1.089, P = .015) and OCP  after adjusting for traditional risk factors  was associated with an increased risk of CACS >100  1.010‐1.106, 4In the present study, a significant association was observed between AIP and the risk of advanced subclinical CAD in subjects beyond traditional risk factors. Recently, CCTA has been widely used for the comprehensive evaluation of coronary atherosclerosis, including lesion location, severity, and plaque characteristics in clinical practice.High levels of the triglycerides‐to‐HDL‐C ratio have been associated with obesity and metabolic syndrome.Recently, several studies demonstrated that AIP is closely related to the coronary atherosclerosis in specific clinical conditions.Several limitations to the study should be acknowledged. First, this was a retrospective, cross‐sectional, and observational study. Thus, selection bias might be present. Second, potential roles of other environmental risk factors and the diet or exercise in subclinical atherosclerosis were not evaluated. Finally, this study included only the Korean adults, which may limit the generalization of obtained results to other populations. Despite these limitations, this study is unique in that an independent association was identified between AIP and advanced subclinical coronary atherosclerosis beyond traditional risk factors in an Asian population. Furthermore, the present study investigated the association of AIP with coronary plaque subtypes.In conclusion, elevated AIP levels are significantly associated with a higher risk of advanced subclinical CAD as revealed by CCTA in Korean adults with near‐normal renal function even after adjusting for traditional risk factors. With consideration of traditional risk factors together, AIP may be an effective predictive marker for advanced coronary atherosclerosis in general population without overt renal dysfunction.The authors declare no potential conflict of interest.All authors have made substantial contributions. Ki‐Bum Won and Eun Ji Park performed the statistical analysis. Hyuk‐Jae Chang contributed to the data acquisition. Ki‐Bum Won, Mi‐Hee Jang, Eun Ji Park, Hyung‐Bok Park, Ran Heo, Donghee Han, JHL, and Hyuk‐Jae Chang contributed to the data interpretation. Ki‐Bum Won and Mi‐Hee Jang drafted the manuscript. Hyuk‐Jae Chang critically revised the manuscript. All authors read and approved the final manuscript. All authors also gave final approval and agree to be accountable for all aspects of work ensuring integrity and accuracy.Supplementary figure 1 Prevalence of coronary plaque subtypes according to AIP quartilesAIP, atherogenic index of plasmaClick here for additional data file.Supplementary figure 2 Optimal cutoff of AIP for CACS >100 and OCPAIP, atherogenic index of plasma; AUC, area under curve; CACS, coronary artery calcium score; CI, confidence interval; OCP, obstructive coronary plaqueClick here for additional data file.Supplementary figure 3 Comparison of ROC models related to the cutoffs of AIP for predicting advanced subclinical CADAIP, atherogenic index of plasma; AUC, area under curve; CACS, coronary artery calcium score; CAD, coronary artery disease; OCP, obstructive coronary plaque; ROC, receiver operating characteristicClick here for additional data file.Supplementary table 1 Association of individual components of AIP with CACS >100 and OCP beyond traditional risk factorsClick here for additional data file.


Although gender plays a pivotal role in the psychological adaptation of immigrant youth, its association with acculturation strategy and mental health among 1st generation immigrant adolescents are still scarce and inconsistent. Therefore, the purpose of the current study was to investigate gender-related differences in acculturation patterns and their association with mental health .N = 440) aged between 12 and 19 years  was collected in a representative German school survey. Fifty-one percent of the sample were female (n = 224). Almost half of the sample was born in the Former Soviet Union, followed by Poland (9.3%). Sociodemographic variables, acculturation strategies, and internalizing as well as externalizing problems were assessed by questionnaires.Self-reported data of immigrant adolescents . Whereas girls more often showed an integration pattern, boys scored higher on the separation and marginalization scale. After adjusting for age and educational level, regression analyses revealed for both gender that marginalization was associated with more internalizing problems. Separation was related to more externalizing problems.1st generation adolescents experiencing a lack of belongingness to German society, socio-economic and educational disadvantages might be particularly vulnerable to mental distress. Findings are discussed in terms of gender-related differential socialization processes in context of immigration. In many societies worldwide, the proportion of immigrant youth is steadily growing. According to Federal census statistics , the proIntegration describes a strong involvement in both the culture of origin and the host culture. Assimilation refers to a strong orientation to the culture of the receiving society combined with low maintenance of the heritage culture. Separation is characterized by strong maintenance of the heritage culture and limited intercultural contact to the host society. Marginalization is defined by low involvement in both cultures. Previous research demonstrated that immigrant youths’ mental health is related to their acculturation strategy [Mage= 16.07, SDage= 2.90; 71% immigrants) Kupper et al. [In order to cope with acculturative stress, different acculturation patterns, describing beliefs, attitudes and behaviors related to the culture of origin and the host culture, are applied. On the basis of the bi-dimensional model, Berry  proposedstrategy . Integrastrategy , 15, 16.r et al.  found hir et al. . In immir et al. . Regardir et al. , 18. In r et al. . Studiesr et al.  or schoor et al. , 20. As r et al. , 22 and The purpose of the current study is to deepen the understanding of the mechanism underlying our previous empirical finding which indicated reduced mental health of 1st generation immigrants . Our dat1. How do 1st generation immigrant girls differ from 1st generation immigrant boys in regard to their acculturation strategy?2. How are acculturation strategies associated with mental health  among girls and boys after adjusting for age and educational level?Based on theoretical assumptions and empirical evidence from previous research, we assumed that separation and marginalization are associated with reduced mental health, while integration is a protective factor against distress. Due to previous inconsistent findings, we examined differences between girls and boys regarding acculturation strategy at an exploratory level. It was hypothesized that the strength of the relationship between acculturation strategy and mental health may differ between girls and boys, with a stronger impact of acculturation difficulties on boys’ mental health.The current study is based on a subsample of 1st generation immigrants participating in a representative school survey. Self-reported data were collected in randomly selected schools stratified by school type , school grade and area selected by number of inhabitants  in North Rhine-Westphalia and Rhineland Palatinate in 2012. Each participant who provided informed consent completed the questionnaires in their schools in the presence of a trained investigator. For participants aged under 14 years an additional informed consent of their parents was required.The study design, procedure and applied questionnaires were approved by the institutional ethics review boards. Three positive ethical votes were required and received. The relevant ethical chambers of Rhineland-Palatinate  and both ethical chambers for North Rhine-Westphalia, specifically the ethical chamber Westfalen-Lippe  and the ethical chamber of North Rhine  voted positively.N = 440 1st generation immigrants were analyzed in the current study. Participants with missing data on the acculturation scale (n = 57) were not considered in the analyses. Dropout analyses revealed that participants with missing values did not differ from the analyzed sample regarding gender, age, country of origin and mental health. 2nd generation immigrants were prior excluded from the study due to anticipated different socialization processes.Migration status was defined according to the German micro census (1). Participants who had migrated themselves to Germany and those who had at least one parent born abroad were considered as 1st generation immigrants. The country of origin of their mothers and fathers was also assessed.The Strengths and Difficulties Questionnaire  is a briScale for Measuring Applied Acculturation Strategy [The Strategy  was appl2-tests for categorical variables were calculated. To analyze gender differences an ANCOVA was used with internalizing problems and externalizing problems as dependent variables, respectively a MANCOVA was calculated with assimilation, integration, separation and marginalization as dependent variables. Before analyzing the acculturation styles, a confirmatory factor analysis (CFA) was calculated first evaluating the suggested four-factor solution of the acculturation scale. The model was estimated with the maximum likelihood method approach. Model fit was evaluated by using following model fit indices [f2= .02) with a statistical power level of .08. Hence, the current sample size was sufficient to carry out the analyses. All analyses considered age and higher education (i.e. grammar school) as covariates. Data analyses were performed by SPSS Version 21.0 and AMOS© 21.0.For group comparisons between girls and boys χ indices : Chi squ indices . A minimM = 16.2; SD = 1.6). 50.9% of the sample were female (n = 224). About half of the sample were from countries of the Former Soviet Union (52.5%), followed by Poland (8.4%), Arabic-Islamic countries (7.7%), Turkey (6.6%), Former Yugoslavia (5.9%) and countries in Middle- and West Europe (4.5%). 14.3% were from other countries. The four largest groups of country of origin were compared regarding mental health and acculturation controlling for age and education. There were no group differences regarding internalizing problems  = 1.36; p = .27) and acculturation pattern  = 1.09; p = .36). However, the groups differed with respect to externalizing problems  = 3.19; p = .024; η2= .03). The highest levels of externalizing problems were found among participants from Turkey , followed by participants from Arabic-Islamic countries , Poland  and Former Soviet Union .Participants were aged between 12 and 19 years  = 4.89; p = .03; η2= .01) and was correlated positively with internalizing problems. The ANCOVA with externalizing problems as outcome variable revealed no gender differences, but both covariates were significant  = 12.43; p < .001; η2 = .03; Fgrammar school = 6.93; p = .009; η2 = .02). The results are shown in Table The distribution of the countries of origin was comparable among girls and boys. No differences emerged for age and grade repetition, but for school type. Whereas more girls attended grammar school, more boys attended vocational school. After controlling for the covariates, girls reported higher levels of internalizing problems than boys. Age was a significate covariate  = 115.5, p < .001). Two CFA separated for boys and girls were calculated  = 86.13, p < .001); CFI = .94, TLI = .92, RMSEA = .06; girls:  = 113.5, p < .000); CFI = .83, TLI = .86, RMSEA = .07). The fit indices suggested a slightly better model fit within the boys’ group. Within the girls’ group, only the RMSEA suggested an acceptable model fit. Detailed psychometric characteristics of the acculturation scale, separated for girls and boys are provided in Confirmatory factor analysis revealed that both incremental fit indexes  and absolute measures of fit indexes were good (RMSEA = .05) supporting the four- dimensional model  = 3.79; p = .005; η2 = .03). Girls more often showed an integration pattern  = 7.23; p < .007; η2= .02) than boys. Boys scored higher on the separation scale  = 4.88; p = .028; η2= .01) and on the marginalization scale  = 6.04; p = .014; η2= .01) than girls. The covariate age was significant  = 7.11; p < .000; η2= .06). Older adolescents reported higher values of assimilation  = 6.84; p = .009; η2= .02) and integration  = 12.39; p < .000; η2= .03), but lower values of separation  = 8.08; p = .005; η2= .02). There was a main effect of school type  = 4.71; p = .001; η2= .04) with adolescents attending a grammar school reporting lower scores for the separation scale  = 9.51; p = .002; η2= .02).Figure F = 6.54; p = .000; R2= .12) and the model with externalizing problems as dependent variable  = 5.35; p = .000; R2= .10). Internalizing problems were associated with female gender and higher age, whereas externalizing problems were associated with younger age and not attending grammar school. Regarding acculturation style, marginalization was related to higher scores in internalizing problems. Higher levels of separation were associated with more externalizing problems. Interaction effects between gender and acculturation styles were not found either for internalizing problems nor externalizing problems.The multiple linear regression analyses Table  yielded The current study aimed to deepen the understanding of former findings suggesting elevated prevalence of internalizing problems and externalizing problems among adolescent 1st generation immigrants by exploring the role of acculturation strategies considering potential gender differences. The findings of this school-based survey suggested that girls and boys differed in the prevalence of internalizing problems and their acculturation pattern. Regardless of gender, marginalization was associated with more internalizing problems, whereas a separated acculturation pattern was linked to more externalizing problems.The results of the confirmatory factor analyses supported the suggested model by Berry  with fouAs expected the findings of the current study revealed a significant correlation between separation and marginalization and reduced mental health. Regarding gender differences, more girls reported internalizing symptoms compared to boys. Higher prevalence of depression and anxiety among adolescent girls has often been demonstrated in research on adolescent’s psychopathology , 36 and Separation as an acculturation style was related to externalizing problems like hyperactivity and conduct problems. This finding contradicts the hypothesis that strong identification with the heritage culture might create a feeling of belongingness and buffer the harmful effect of perceived discrimination with positive impact on individual’s mental well-being . The assIn contrast to our hypotheses and previous research, we could not identify integration as protective acculturation pattern. Nguyen and Benet-Martínez  concludeFinally, several limitations of the current study need to be considered while interpreting the results. The data obtained was based on self-report, which can lead to response bias. In regard to the sample, the large proportion of adolescents from the Former Soviet Union is probably explained by historical reasons as the majority of these immigrants are very likely so-called “Spätaussiedler” (a German minority who had been living in the former Soviet Union and other former eastern bloc states and re-migrated to Germany). Despite the applied procedure of random selection, the proportions of the immigrants’ countries of origin is not according to representative data of the population and might reflect local clustering . While we took effort to recruit a representative sample, we cannot preclude that certain groups of migrants (e.g. Turkish immigrants) were underrepresented. Importantly, after the migration movements starting in 2015, the proportion of immigrants from Arabic-Islamic countries could be higher today compared to 2012 when the survey was conducted.A further significant concern is participation bias given that the sample was limited to immigrants with sufficient language skills as only German questionnaires were utilized. Language skills determine the intensity of interpersonal contact in the receiving society  and are Moreover, education level was used as an indicator for socioeconomic status. Despite its significant correlation with migration status, socioeconomic status is difficult to assess in youth samples due to lacking information from parents. However, the representative sampling in the current study promoted variation in socioeconomic status across the sample. As mentioned above, the cross-sectional study design does not allow to draw causal conclusions. Longitudinal studies are required to examine developmental trajectories of acculturation pattern and mental health across adolescents. Since we did not assess age-salient developmental tasks in adolescence like developing independence from parents, we could not disentangle the impact of acculturation stress and general challenges in puberty on immigrated adolescents’ mental health. To develop a full picture of immigrant youth adaptation, more studies will be needed that additionally take family-related factors into account like parental support cf.  and parest generation adolescents experiencing socio-economic hardship, educational disadvantage and a lack of belongingness to the host society might be especially vulnerable to mental distress. The results have practical applications suggesting that e.g. promoting school engagement in immigrant pupils by teachers and through means of policy can be useful to reduce separation and marginalization as risk factors for distress. Although we were not able to examine the mechanisms underlying the association between acculturation styles and mental health, differentiating internalizing and externalizing problems might be useful for future studies in order to identify possibly unique patterns shaping different syndromes in adolescence.Consistent with previous findings of this school-based survey, gender-related considerations within the acculturative process are pivotal to understand young immigrants’ psychological adaption. 1


This study explores how the human brain solves the challenge of flicker noise in motion processing. Despite providing no useful directional motion information, flicker is common in the visual environment and exhibits omnidirectional motion energy which is processed by low‐level motion detectors. Models of motion processing propose a mechanism called motion opponency that reduces flicker processing. Motion opponency involves the pooling of local motion signals to calculate an overall motion direction. A neural correlate of motion opponency has been observed in human area MT+/V5, whereby stimuli with perfectly balanced motion energy constructed from dots moving in counter‐phase elicit a weaker response than nonbalanced (in‐phase) motion stimuli. Building on this previous work, we used multivariate pattern analysis to examine whether the activation patterns elicited by motion opponent stimuli resemble that elicited by flicker noise across the human visual cortex. Robust multivariate signatures of opponency were observed in V5 and in V3A. Our results support the notion that V5 is centrally involved in motion opponency and in the reduction of flicker. Furthermore, these results demonstrate the utility of multivariate analysis methods in revealing the role of additional visual areas, such as V3A, in opponency and in motion processing more generally. Motion opponency is the process by which opposing motion signals in the same local area are cancelled during visual motion processing, resulting in the attenuation of non‐informative flicker noise. Using multivariate pattern analysis, we examined flicker processing and motion opponency throughout human visual cortex. Our results implicate area V3A as a contributor to motion opponency and affirm the central role of V5 in the suppression of flicker noise. However, the successful interpretation of directional motion information is complicated by the presence of flicker noise. Any abrupt change in the luminance of a visual scene, like a flickering light or a bright object appearing suddenly against a dark background, creates flicker noise: omnidirectional and uninformative signals which can be processed just as any true motion signal   as a powerful tool for understanding neural processing using fMRI  stimuli, counter‐phase (CP) stimuli, or nonmotion (NM) stimuli exhibiting incoherent onset and offset flicker but no smooth translational movement. Classifiers were trained to discriminate two of the three different stimuli and tested on both trained and untrained stimuli. We predicted the following pattern of results:As a secondary analysis, we explored the performance of classifiers trained using BOLD data from V1, V2, V3, V3A, and V4 to assess whether BOLD responses indicative of opponency were present throughout the human visual system.22.12).All experimental stimuli were programmed in Python using the Psychopy library Peirce, . Stimuli2) of size 3 pixels were presented. Each dot was initially placed no more than 8 pixels away from its paired partner along a common orientation, creating a Glass pattern , nonmotion (NM), or in‐phase (IP) stimuli. During CP blocks, the two dots in a pair traveled in opposite directions. CP pairs were initially separated by 8 pixels along the Glass pattern orientation and traveled toward one another, crossed, and were randomly replotted after again achieving a separation of 8 pixels. To temporally stagger the replotting of CP dots, each CP pair was initially plotted at a randomly selected point along its full trajectory.During IP blocks, both dots within a pair traveled in the same direction along the orientation of the Glass pattern. Each pair was independently assigned a random initial lifetime to temporally stagger the replotting of dot pairs, and each pair was independently assigned a random within‐pair distance between 0 and 8 pixels. Different IP pairs traveled in opposite directions along the Glass pattern orientation, creating a bidirectional stimulus. Because IP and CP dots all traveled 8 pixels during their 150 ms limited lifetime, the dot speed ranged from 2.3 to 2.6°/s, depending on viewing distance.NM dots behaved identically to in‐phase dots, except that there was no translational motion. Critically, in‐phase and counter‐phase stimuli contained the same number of left and right motion signals, and the Glass patterns of all three conditions were indistinguishable from one another. One experimental run contained 6 blocks of each paired‐dot condition, totaling 18 blocks per run. Each participant performed 8 runs, totaling 144 blocks (48 blocks per condition). Stimulus diagrams are presented in Figure 2.2Functional neuroimaging data were collected from five participants. All participants had normal or corrected‐to‐normal vision. Informed consent was obtained, and all participants were treated in accordance with the Code of Ethics of the World Medical Association (Declaration of Helsinki). For their participant, participants received CAN$200 (CAN$50 per scanning hour).2.3All scans took place in the Centre for Functional and Metabolic Mapping at the University of Western Ontario's Robarts Research Institute on the 7T Siemens Magnetom scanner. All functional scans used an 8‐channel transmit, 32‐channel receive coil optimized for the occipital pole and providing an unobstructed field‐of‐view to the visual stimulus. All anatomical scans used an 8‐channel transmit, 32‐channel whole‐head coil. For each participant, we collected an anatomical scan , two retinotopy scans, one with rotating wedge stimuli and one with expanding ring stimuli , one V5 localizer scan , and eight experimental scans .2.4www.neuroelf.net). All transformations were applied with sinc interpolation.The fMRI data preprocessing, the functional localizer analysis, and the univariate experimental analysis were conducted using BrainVoyager QX 2.8.4 . V5 was defined as significant clusters of voxels bilaterally located near the ascending limb, or the posterior continuation, of the inferior temporal sulcus or the posterior bank of the superior temporal sulcus . The final ROIs were defined as the significant voxels within the ROI borders.To identify areas V1–V4, a 3D brain surface model was constructed from the skull‐stripped and Talairach‐transformed anatomical scan in BrainVoyager. The surface was inflated, cut across the calcarine sulcus, flattened, and corrected for surface distortions. A whole‐brain, voxel‐wise cross‐correlation analysis was carried out and mapped onto the flattened brain surface, and the borders of V1–V4 were identified by observing the cross‐correlation polarity reversals running along the calcarine sulcus. One GLM was fit using all experimental data collected for the participant with one regressor for each stimulus condition and 2.6z‐scored head‐motion data as additional nuisance regressors. Each block was modeled as an individual box‐car, convolved with NeuroElf's default double‐gamma HRF and z‐score normalized.For visualization, model‐independent univariate BOLD time courses were extracted from the Talairach‐space transformed data for each stimulus condition and visual area. However, the data were transformed to native functional space for the main MPVA analysis. Within‐subject multivariate pattern classification analyses were carried out on fitted voxel‐wise GLM betas using support vector machines for each visual area. The GLMs for multivariate pattern classification were calculated using NeuroElf and were separate from the GLMs for ROI localization. One GLM per ROI was fitted with each individual block as a separate regressor .In all multivariate analyses, significance was established using within‐subject permutation tests in which the condition labels were randomly permuted within runs, ensuring that each run preserved the same number of each type of label  = 5.1, p = .038. Pairwise contrasts found a significant difference between IP (3.39) and CP (3.08), t(8) = 2.6, p = .032, and between IP and NM (3.04), t = 2.9, p = .020. There was no significant difference between CP and NM. Figure To test for significance in the V5 timeseries, an average % change was calculated separately for each participant and stimulus condition by averaging the values of 10 successive TRs  in the time series, beginning with the fourth TR (4.8 s after stimulus onset) to account for the hemodynamic response delay. The average IP, CP, and NM values were tested using a one‐way repeated‐measures ANOVA, finding a significant effect of stimulus condition 3.2p possible is 115,201=6.6×10−5 when the true value is more extreme than every null value. IP, CP, and NM blocks were used to train and test IP v. CP, IP v. NM, and CP v. NM classifiers to examine the separability of each condition with a one‐tailed permutation test. A Bonferroni correction for multiple comparisons was applied. This analysis contains three comparisons across six ROIs; therefore, a critical p of 0.0518=2.8×10−3 was set to determine better‐than‐chance accuracy. See Supporting Information Table For all MVPA analyses, significance was determined using a permutation test with 15,200 random permutations. Therefore, the minimum p = p = p = p = p = p = p = p = p = p = p = p = p = p = p = p = p = p = Every ROI achieved greater than chance performance when discriminating IP and CP. In increasing performance order: V1—57%, p = p = .026 and mixed results when discriminating NM from CP. Table Individual‐subject discrimination was also examined to assess whether individual‐subject trends were consistent with the group analysis. Areas V1–V4 did not exhibit consistent trends across participants. See Supporting Information p of 0.056=8.3×10−3 was set to determine better‐than‐chance performance.Because the group data from V3A and V5 achieved at least 70% accuracy in two comparisons, these data were submitted to a further analysis designed to probe for any systematic misclassification biases. Three classifiers were trained to discriminate between IP and CP, IP and NM, and CP and NM identically as in the previous analysis, but they were tested with the untrained condition to determine whether the untrained condition would elicit a consistent misclassification error. A two‐tailed permutation test was used to determine significance, and a Bonferoni correction for multiple comparisons was applied. This analysis contains three comparisons across two ROIs; therefore, a critical p = p = p = p = The classifier trained to discriminate CP and NM was tested with IP data, and there was no systematic misclassification of IP in either V3A or V5 . No dataset demonstrated IP misclassification bias.At Area V3A, all five individual‐subject datasets misclassified CP as NM more often than IP, and two datasets individually reached Bonferroni corrected statistical significance. The four high performing datasets also misclassified NM as CP more often than IP, with one reaching the Bonferroni significance cutoff and two reaching uncorrected significance .Table A Group‐level discrimination performance across all ROIs.Table B:Group‐level misclassification bias across all ROIs.Table C: Individual‐subject discrimination performance across all ROIs.Click here for additional data file.Video A Example video of IP, CP, and NM trials composing a full experimental stimulus block. During the experimental scanning sessions, a block contained six trials of the same paired‐dot stimulus condition, all exhibiting Glass patterns that were randomly oriented 15° clockwise or counterclockwise from the block's overall cardinal direction. Trials were 1.1 s long, and participants indicated whether the Glass pattern was clockwise or counterclockwise. In the video, three vertical IP trials are presented, oriented counterclockwise, counterclockwise, and clockwise, respectively. Three vertical CP trials are then presented, also oriented counterclockwise, counterclockwise, and clockwise, respectively. Finally, three vertical NM trials are presented, oriented clockwise, counterclockwise, and counterclockwise.Click here for additional data file.