Blattabacterium), and a rich gut microbiota. The transmission mode of Blattabacterium is vertical, while the gut microbiota of a new generation is mainly formed by bacterial species present in feces. We have carried out a metagenomic analysis of Blattella germanica populations, treated and non-treated with two antibiotics (vancomycin and ampicillin) over two generations to (1) determine the core of bacterial communities and potential functions of the gut microbiota and (2) to gain insights into the mechanisms of resistance and resilience of the gut microbiota. Our results indicate that the composition and functions of the bacteria were affected by treatment, more severely in the case of vancomycin. Further results demonstrated that in an untreated second-generation population that comes from antibiotic-treated first-generation, the microbiota is not yet stabilized at nymphal stages but can fully recover in adults when feces of a control population were added to the diet. This signifies the existence of a stable core in either composition and functions in lab-reared populations. The high microbiota diversity as well as the observed functional redundancy point toward the microbiota of cockroach hindguts as a robust ecosystem that can recover from perturbations, with recovery being faster when feces are added to the diet.Symbiosis between prokaryotes and eukaryotes is a widespread phenomenon that has contributed to the evolution of eukaryotes. In cockroaches, two types of symbionts coexist: an endosymbiont in the fat body ( Drosophila  relationship with the hosts, living intracellularly in specialized host cells, and functionally complementing the host in many ways  in the fat body, that plays an essential role in nitrogen metabolism (Cockroaches (Blattodea) are a particularly intriguing case since both, endosymbiont and microbiota, coexist in each individual host. On one hand, the obligate endosymbiont tabolism . On the tabolism .Fusobacterium strain FuSL in the gnotobiotic cockroach Shelfordella lateralis is dependent on the presence of other microbial species, indicating that the order in which species colonize the gut could determine community structure  or transmitted vertically through the eggs or the offspring  was reared in climatic chambers at the Cavanilles Institute of Biodiversity and Evolutionary Biology  in plastic jars with aeration at 26 °C, 65% humidity and 12L:12D photoperiod. Cockroaches were fed dog-food pellets  and water was supplied ad libitum. The antibiotics vancomycin  and ampicillin  were supplied with the water at 0.02% (w/v) in two different experiments . Vancomycin is a glycopeptide antibiotic, acting against Gram-positive bacteria by binding to D-alanyl-D-alanine portion of the bacterial cell wall precursors, and prevents binding of this portion with the PBP (penicillin-binding protein). Ampicillin is a beta-lactam antibiotic acting against Gram-positive and some Gram-negative bacteria, which binds to PBPs, inactivating them and interfering with the cross-linkage of peptidoglycan chain.0). Then, the population was divided into three subpopulations: One was never treated with antibiotics and was used as control (C population), and the other two were treated with vancomycin (V population) or ampicillin (A population). Samples were taken at 10 and 30 days in the three G1 populations. When the ootheca were fully formed, adults from each antibiotic population were divided into two groups, with or without antibiotic, to generate the second generation (G2) populations. Newly hatched nymphs’ population from the groups maintained on antibiotic initiated the antibiotic treatment populations (VV and AA populations). The nymphs hatched in antibiotic-free environment were immediately divided into two populations, one without any further treatment giving rise to the VC and AC populations, and the other one supplemented with feces obtained from a control population never treated with antibiotics, yielding the VF and AF populations. At G2, we collected samples at 22 and 34 days that correspond approximately to the nymphal stages n3 and n4 and at 0, 10, and 30 days after ecdysis. The AC30a sampling time point was not taken because none of the individuals reached this age. Overall, 113 samples were analyzed, 52 in the vancomycin experiment , 49 in the ampicillin experiment  and 12 in control. We analyzed the microbiota composition and functions in the hindgut by metagenomics, and quantified the Blattabacterium population in the fat body by qPCR.We started with a synchronized adult population, composed of individuals collected between 0–48 h after adult ecdysis   following the manufacturer’s recommendations, adding lysozyme (20 mg/ml) to the cell lysis buffer to break Gram-positive bacterial cell wall, and used for metagenomic sequencing using the Illumina MiSeq (2 × 300 bp) technology at the FISABIO . Total DNA from the fat body was extracted following the protocol described in From the raw sequencing data, the adaptors were removed by using Cutadapt . To remoTaxonomic assignments of the metagenomes were carried out through Kaiju and the non-redundant bacteria database , a progrTo assign functional annotations, the reads were previously assembled into contigs via Ray (v2.3.1) . Then, wTaxonomic and functional annotations provide bacterial species and gene compositional abundance matrices, respectively. The taxonomy was based on the non-redundant bacteria database and the functional annotation on the TIGRFAMs protein family database. We used these abundance tables to carry out comparative analyses. The analyses were performed at different classification levels: family in the case of taxonomy and gene and biological process (main role and sub role levels) in the case of functions. The analyses were performed with in-house R scripts (R version: v3.1) .Alpha diversity  of the different sample points was based on the Shannon and Chao1 indexes and statistically tested with the Wilcoxon signed-rank test implemented in the R library vegan . We usedp-value below 0.05 was used and 90% confidence intervals for the relative abundances of the two groups could not overlap.Univariate comparative analyses were performed  to identify both taxonomic and functional shifts between groups of samples. To determine shifts, each feature  was tested for exhibiting statistically significant differences between two groups according to Wilcoxon test. Moreover, in order to reduce false positives, a false discovery rate (FDR) adjusted Blattabacterium population was analyzed by qPCR using ArialMx Real-Time PCR System . The genes ureC (accession number NC_013454.1) and actin5C (accession number AJ861721.1) were used as specific of the endosymbiont population and the host control, respectively. The primers UC1F: 5′-GTCCAGCAACTGGAACTATAGCCA-3′ and UC1R: 5′-CCTCCTGCACCTGCTTCTATTTGT-3′ were used for the ureC gene, and ActinF: 5′-CACATACAACTCCATTATGAAGTGCGA-3′ and ActinR: 5′-TGTCGGCAATTCCAGGGTACATG-3′ were used for the actin5C gene, as previously described .To check if the antibiotics have an effect on the endosymbiont and therefore in the host–microbiota interaction, the escribed . StatistA total of 113 individuals corresponding to 36 time points of the vancomycin and ampicillin experiments were analyzed by metagenomic sequencing . The avep-value = 0.006 and A, p-value = 0.031). In the vancomycin experiment, the antibiotic treatment affected the microbial richness (based on the Chao1) that significantly decreased at G1 compared to the controls , and that remained lower than the control at G2 , including the VC samples . For the ampicillin experiment the richness estimator Chao1 was also significantly lower than the control only at G1 . Interestingly, the VF and AF diversity and richness did not show significant differences compared to the controls in any case, and even reached higher values.The Shannon index  and the p-value = 0.21). Therefore, for the comparisons, we have considered C0a, C10a, and C30a as a homogeneous population (named Ca). In the vancomycin experiment . In the ampicillin experiment . The results indicate that antibiotics have a strong effect on the microbiota, but there is a resilience capacity since the diversity of the control microbiota could be restored (at least in part) by removing the antibiotic from the diet. Clearly, this recovery was faster, when the diet was supplemented with feces, with adults at 10 and 30 days at G2 achieving a similar composition to adults in the control population.We carried out a CCA of microbial community composition from all individuals analyzed in each experiment . The 12 periment , the firperiment , a similTo further investigate the results obtained, we focused on the taxa with abundance levels above 0.1% in at least one of the 36 time point samples, both at phylum and family levels see , S4.The outcome of the antibiotic treatment on the gut microbiota composition was different in both experiments  and in adults . However, when comparing Ca vs. VF only the differences at 22 day nymphal stage  and at adults day 0 was significant . Thus, the results indicated that there was no total recovery of the vancomycin treatment with adding feces until adults of 10 days. For the ampicillin experiment, in the comparisons Ca vs. VC only one significant difference was found at 0 days adults . When adding feces (Ca vs. AF), non-significant differences were obtained in nymphal and adult stage. Respect to the families affected by the treatment at G1, they tended, in general, to reach abundance values close to control population, recovery being faster when feces were added to the diet. VF and AF populations reached abundance values similar to controls and only a few taxa  showed significant differences, mainly in nymphal stages , and with feces added to the diet (VF). For the vancomycin experiment, significant differences (Adonis test) were observed in the comparisons Ca l stages , giving l stages , we obseB. germanica adults (The results obtained showing similar taxa composition in the second-generation adults (at 10 and 30 days) in both experiments when feces were added to the diet, and similar to that of control samples, indicate the existence of a resident gut microbiota. Therefore, we decided to analyze the taxa present in C10a, C30a, VF10a, VF30a, AF10a, and AF30a in all the samples and with an abundance value higher than 0.1% as indicative of the phyla and families that form the core of the gut microbiota of a adults . We decia adults . NeverthWe have carried out the functional analysis of the 113 metagenomes by comparison against the TIGRFAM database, obtaining a hierarchical classification for the main-roles , sub roles (more specific metabolic functions for each one of the roles), and genes (metabolic functions) . In the vancomycin experiment  ; pathogenesis (cellular processes); carbohydrates, organic alcohols and acids, anions and nucleosides, purines and pyrimidines (transport and binding proteins); and two decrease their abundance: sporulation and germination (cellular processes) and tRNA aminoacylation (protein synthesis). Furthermore, five sub roles were overrepresented  in some samples: toxin production and resistance (cellular processes); nitrogen metabolism and sulfur metabolism ; anaerobic and Entner-Doudoroff (energy metabolism) and only one, DNA replication, recombination and repair (DNA metabolism) was underrepresented in one comparison.The only main role showing a slight increase for both antibiotic treatments is vs. VVa) . Only setransport and binding proteins. At sub role level, three changed significantly in the first generation as a direct response to the antibiotic (Ca vs. Aa), and seven in nymphs in the second generation (Ca vs. AAn). However, there were no consistent changes in the two treated generations. Moreover, non-significant changes were found in treated second-generation-adults (Ca vs. AAa)  .vs. VC) and without feces (Ca vs. VF) were not significant. In the case of the nymphs, the comparisons were significant in samples without added feces  but not significant in the samples with added feces. Then, we compared the relative abundance of the subroles in nymphs and adults  with the control (Ca) (vs. VFn) and adults (Ca vs. VFa), only one difference in the pyruvate family (amino acid biosynthesis) was found in nymphs (vs. VCn) highlighted 27 significantly different sub roles, whereas in adults (Ca vs. VCa) recovery was complete with only one exception in the two component systems , which suggests that the control functions can be achieved in adults without fecal supplementation. In the ampicillin experiment, the Adonis test was non-significant in any of the comparisons, both adults and nymphs samples with or without feces. Non-significant differences were found in adult stage in the non-treated samples with and without fecal supplementation (Ca vs. AFa and Ca vs. ACa) (vs. AFn) showed seven differences, and those of the non-supplemented diet population (Ca vs. ACn) lacked significant differences. This result should be taken with caution as despite the AC population was funded as the other ones, all individuals of ACn and ACa came from the same female, and no individual reached the 30 days adult stage, as it was indicated in section “Materials and Methods.”An important finding of the taxonomic analysis was the almost total recovery of the main taxa in adults at G2 when feces were added to the diet. As we did for the bacterial composition, we compared the functions (gene abundance) between groups with the Adonis test. The vancomycin experiment showed that at functional level all the comparisons of the control with the adult samples with (Ca rol (Ca) . When con nymphs . An unexvs. ACa) . HoweverB. germanica. We decided to determine the core functions of adults , protein synthesis, DNA metabolism, protein fate, and regulatory functions. Furthermore, pathways related to the synthesis of vitamins were identified, such as biosynthesis of vitamin B9 (folic acid), B1 (thiamin), B12 , K2 (menaquinone), or biotin. Nitrogen fixation and metabolism was also identified in the metagenomes. It is worth mentioning that one of the most abundant sub roles was related to mobile and extrachromosomal element functions.Overall, the results observed showed functional redundancy in the microbiota of f adults , based oacrB, bacA, mexB, and tetM could confer resistance to different antibiotics like aminoglycoside, glycylcycline, β-lactam, macrolide, acriflavin, bacitracin, tigecycline, fluoroquinolone, and tetracycline.In order to find whether there was any change in the abundance of antibiotic resistance genes (ARGs) as a consequence of antibiotic treatment, we have compared all reads against the ARDB in the control and in vancomycin and ampicillin treated samples. For this preliminary survey, we considered those genes present in at least two of the biological replicates in at least two of each grouped sampling time as genes representative of controls and antibiotic-treated samples . Ten diftetM. In nymphal stage VVn (VV22n and VV34n samples), in addition to the three ARGs detected in Va and in the control, four new ARGs were detected: macB, mdtK, rosA, and smeE, which presumably confer the same resistance profile as the control condition besides enoxacin, norfloxacin, and fosmidomycin. However, in adults (VVa) the genes macB and mdtK were not detected. In the ampicillin treated first-generation samples Aa (A10a and A30a), only the gene bacA conferring resistance to bacitracin was detected. However, in the second generation we observed an increase in the number of ARGs. In nymphal stages (AAn) we detected four genes acrB, bacA, mexB also detected in control, and ksgA, which confer resistance to kasugamycin. In the adult stage (AAa), six ARGs were detected, two (macB and rosB) more than in the control condition.The vancomycin treated samples did not harbor a resistance gene for vancomycin, in neither the first (Va) or in the second generation (VVn and VVa). The genes detected in controls were also detected in Va (V10a and V30a), with the exception of tetM gene, assigned to Bacteria_uc or Firmicutes. Most of ARGs in antibiotic treated samples are multidrug resistance efflux pumps. Furthermore, the ARGs detected seemed not to respond to the selective pressure produced by the antibiotic treatment administrated, but reflected taxonomic level changes, therefore the surviving bacteria. Further studies and analyses will be necessary to deep on this important topic.When the ARGs were assigned to a putative phylum, all of them were detected in the Proteobacteria, with one exception, ureC gene from Blattabacterium at all time points at G1 and G2. Non-significant differences were obtained, independently of whether or not samples were treated with vancomycin or ampicillin. The average copy number of the actin5C gene from the host, used as a control, did not show significant changes .We estimated the average number of copies of  changes . Thus, BB. germanica is an omnivorous insect in which two symbiotic systems coexist in each individual, Blattabacterium in the fat body and the microbiota in the hindgut. Blattabacterium has a role in the provision of essential amino acids and participation in the nitrogen metabolism , and rifampicin appears to affect the bacterial community most drastically . Similar disturbances in this function have been described in the gut microbiota of mice and humans treated with antibiotics  were the most abundant in a metagenome analysis of the human gut microbiota  were the ones described in previous studies of gut microbiota composition in B. germanica   , and witteralis) . On the teralis) , whereasteralis) .B. germanica gut microbiota transmission and equilibrium at adult stage is even more evident if we look at the functional profiles. Adults in the second antibiotic-free generation with a feces-supplemented diet (VFa and AFa) proved similar to control-population adults (Ca) than to the antibiotic affected insects. Moreover, nymphs also tended to have similar functions, at least in one population (VFn), with only significant differences in amino acid biosynthesis. This could indicate that in similar environments, like that of the lab-reared cockroaches, the bacterial community of B. germanica is assembled to reach a characteristic composition, and functions, some of which might be those required by the host. The convergence for essential functions has been described in the hindgut paunch of Amitermes wheeleri collected from cow dung and in Nasutitermes corniger feeding on sound wood. In this study, the authors found community divergence in both species but similar functions were performed, such as hydrolytic enzymes, homoacetogenesis and cell motility and chemotaxis  would help us to disentangle the role of the microbiota and if there is a crosstalk between the two symbiotic systems.The importance of feces in emotaxis . Future B. germanica were conserved even in the antibiotic treated-samples, suggesting a buffering capacity of the microbiota to perform essential functions. Functional redundancy must be essential to guarantee the function of the system even in the face of a disturbance, since other microorganisms can take over the functions carried out by those affected by the perturbation  belonging to the Bacteroidetes phylum. This protein participates in the metabolism of proteins and carbohydrates  and the MATE efflux family protein, both systems related to antibiotic and drug resistance. Moreover, processes related to mobile and extrachromosomal element functions, including prophage and plasmid functions are also abundant in the gut microbiota of this animal. Thus, much like humans, cockroaches harbor a repertoire of resistance genes and spreading mechanisms in their gut microbiota.The main roles of gut microbiota in urbation . This coial bees , since mhydrates , possiblhydrates . A previtabolism . Here, wB. germanica microbiota harbors ARGs in laboratory control conditions, and thus we can postulate that natural populations also harbor a resistance gene repertoire (resistome) in their microbiota. It is well known that one factor inducing the spread of antibiotic resistant bacteria is the systematic used of antibiotic in humans that can have transformed the gut microbiota into a reservoir of ARGs  rather than to ARGs .All the sequences of gut metagenomes in this study were submitted to the European Bioinformatics Institute (EBI), EMBL Nucleotide Archive under the study accession number RD-S, AM, CG-F, and AL design the experiment. RD-S, AP-C, AM, CG-F, and AL wrote the manuscript. RD-S carried out the laboratory work. CG-F and IT contributed to the laboratory work. AA, AP-C, RD-S, and AL analyzed the data. RD-S, AP-C, JC, and AM contributed to the data analysis. All authors edited the manuscript and approved the final draft.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.


Predictors of problematic smartphone use have been found mainly in studies on elementary and high school students. Few studies have focused on predictors related to social network and messaging apps or smartphone model. Thus, the objective of our study was to identify predictors of problematic smartphone use related to demographic characteristics, loneliness, social app use, and smartphone model among university students. This cross-sectional study involved 257 Brazilian university students who answered a smartphone addiction scale, a questionnaire about smartphone usage patterns, and the Brazilian version of the UCLA-R loneliness scale. Women, iPhone owners, and users of Instagram and Snapchat had significantly higher smartphone addiction scores. We found correlations between scores for the Brazilian version of smartphone addiction scale and the importance attributed to WhatsApp, Facebook, Instagram, and Snapchat, and the Brazilian version of the UCLA-R loneliness scale. Our hierarchical regression model predicted 32.2% of the scores of the Brazilian version of the smartphone addiction scale, with the greatest increase in predictive capability by the step that added smartphone social app importance, followed by the step that added loneliness. Adding the smartphone model produced the smallest increase in predictive capability. The theoretical and practical implications of these results are discussed. Technological progress is one of the hallmarks of the new millennium. The use of smartphones has increased substantially over time, in part due to the many apps that these devices can support Andrew, . AlthougCurrently, the accuracy of the term “smartphone addiction” is being questioned. Panova and Carbonell  explaineMontag, Wegmann, Sariyska, Demetrovics, and Brand  proposedA model was created to explain the interaction between several individual characteristics with Internet use disorder. The model is called “I-PACE” and is an acrogram for Interaction of Person-Affect-Cognition-Execution. This model accounts for biopsychological and social features of the person , affect and cognition , and executive functions  have also been investigated, with conflicting results. While Shaw, Ellis, Kendrick, Ziegler, and Wiseman  reportedThere has also been research on the social networks accessed by smartphone users because of concerns about their problematic use of networks , the two concepts probably overlap within the problematic Internet use, in which case it is necessary to raise more evidence, especially in cultures that escape the rich and industrialized context, as is the case in most research reported. Furthermore, it is important to understand how the use of smartphones can be healthy and productive.Age was negatively correlated with smartphone use  .We used the Brazilian Portuguese version Busin,  of the SThis self-responsive, multiple-choice questionnaire was developed by the authors to assess characteristics of the participants and their smartphones, and their usage of social apps  through the device. The first part of this questionnaire collected data on participants’ age, gender, family monthly income, university major, smartphone model/system, and the approximate value of their device. The second part of the questionnaire inquired about specific social apps  they accessed through their smartphone and if the participants used their smartphone for work purposes. They then answered a question asking what degree of importance (ranging from 1 = “not important” to 5 = “very important”) they attributed to each social app and to the use of the device for work purposes.We assessed participants’ loneliness using the Portuguese Brazilian version of the UCLA Loneliness Scale  and one-way analysis of variance (ANOVA) (for items with three or more possible answers) to assess differences in mean smartphone usage scores (SAS-BR) according to participant characteristics. Analysis of covariance (ANCOVA) was conducted to investigate if age and sex could be mediators of the analysis made with t tests and ANOVAS. We used Spearman’s rank correlation coefficient to assess the correlation between participants’ smartphone use scores (SAS-BR) and the importance they attributed to social apps. To assess the correlation between smartphone use scores (SAS-BR) and loneliness (UCLA-BR) scores, we used Pearson’s correlation coefficient. Lastly, we conducted a hierarchical multiple regression to investigate the influence of demographic characteristics, loneliness, smartphone social apps, and smartphone model on problematic smartphone use. R , percentages, and minimum and maximum values. We used Student’s Because some of the predictors can be categorical variables, we transformed them into dummy variables ; loneliness scores were entered in the second step (loneliness); Facebook importance, WhatsApp importance, and Instagram importance were entered in the third step ; and the model of the smartphone was entered in the fourth and final step (smartphone model). All standardized coefficients were reported in each step for each variable, and the collinearity was reported for the last model.p = .107, Skewness = 0.272, Skewness SE = 0.152, Kurtosis = − 0.042, Kurtosis SE = 0.303). Table n = 13) either did not answer this question or did not know the price of their device.The SAS-BR score was normally distributed  of the participants had minimal loneliness, 24.5% (n = 63) had mild loneliness, 9.7% (n = 25) had moderate loneliness, and 2.7% (n = 7) had intense loneliness scores.The participants’ mean (SD) smartphone use score (SAS-BR) was 98.00 (26.73), out of a maximum of 198 points, with a range from 40 to 183. Their mean (SD) loneliness score (UCLA-BR) was 19.49 (12.50), out of a maximum of 60 points, with a range from 0 and 59. Nearly 63% , university major , and smartphone usage for work  did not predict problematic usage; however, smartphone model , Facebook usage , Instagram usage , and Snapchat usage  did. No results that were statistically significant in Table To ensure that effects found in Table p < 0.001) as well as importance attributed to WhatsApp (p < 0.001), Facebook (p < 0.001), Instagram (p < 0.001), and Snapchat (p = 0.012). We found no correlation between SAS-BR scores and the importance attributed to use of the smartphone for work purposes (p = 0.155).Several correlations were conducted between SAS-BR, UCLA-BR, and importance attributed to smartphone-related activities. These correlations are shown in Table R = 0.57)—with demographic characteristics, loneliness, social apps importance, and smartphone model—predicted 32.2% of the SAS-BR variance.To examine the association of these variables with the SAS-BR score, a hierarchical multiple linear regression analysis was conducted. The regression predictors, standardized and unstandardized coefficients, standard error, tolerance, and prediction percentage are shown in Table We found a positive correlation between problematic smartphone use in Brazilian university students and the importance they attributed to social media as well as between SAS-BR scores and loneliness scores. The magnitude of this effect was moderate. Although some studies  had higher SAS-BR scores. On the other hand, we did not detect differences in smartphone use scores according to the students’ university major, which is consistent with research by Sanal and Ozer .p < 0.10): users in the lowest (< 2000 BRL) and highest  family income strata had higher SAS-BR scores, indicating a greater likelihood of problematic smartphone use. Similarly, several other studies  and not necessarily related to the importance they attributed to these apps. Therefore, it seems that the use of additional social apps is perhaps more relevant to problematic smartphone use than the importance the users attribute to these social apps. In addition, the importance of the apps may change over time. For example, we hypothesize that users attribute more importance to the social apps in which they have the most friends or followers. As a result, the importance attributed to these social apps will change over time based on changes in numbers of users of these apps. These factors should be considered in future research examining social apps. In our study, the most important social network was Facebook, which was also the network with the most users in the study period  were predictors of problematic smartphone use. Specifically, being young, being female, and having a family monthly income higher than 10,000 BRL were related to more problematic smartphone use. These results are consistent with findings in previous studies  increase the likelihood of problematic smartphone use. Our results can be useful for clinical psychiatrists in the management of patients struggling with smartphone use and to identify high-risk groups. Finally, our results shed new light on some of the predictors of problematic smartphone use; we found that demographics characteristics such as age, sex, and family monthly income, loneliness, and social app importance were predictors of problematic smartphone use, but the smartphone model was less important in this model.In sum, this study produced interesting findings for problematic smartphone use. First, one of the biggest predictors of problematic smartphone use was the importance attributed to social network apps, even when loneliness was controlled. This indicates that the relationship between problematic smartphone use and social network use was the most important relationship we examined. Additionally, the results suggest that the smartphone model (iPhone/Samsung/other) is not particularly relevant in predicting problematic smartphone use. Furthermore, this study was carried in Brazil and is one of the first studies from South America on smartphone usage. Lastly, this manuscript contributes to advancing the models contained in other scientific articles. Most prior models included different variables, with some common ground. In this study, the common variables used in other studies were examined as well as additional measures. Future studies should consider including socioeconomic measures, loneliness measures, and social network usage/importance measures as well as adding new variables, such as personality traits.


A second stepwise multiple linear regression analysis was performed after excluding MT and SCAT thickness from the independent variables. Sex and age for Japanese participants  and lean body mass and body mass index for Brazilian participants  were identified as significant independent variables. The present results suggest that MT is closely correlated with muscle quality in Japanese and Brazilian older individuals. Increases in muscle size may induce decreases in intramuscular adipose tissue and/or connective tissues, which are beneficial for reducing the risks of metabolic impairments in Japanese and Brazilian older individuals.Muscle quality is well-known to decrease with aging and is a risk factor for metabolic abnormalities. However, there is a lack of information on race-associated differences in muscle quality and other neuromuscular features related to functional performance. This study aimed to compare muscle quality, function, and morphological characteristics in Japanese and Brazilian older individuals. Eighty-four participants aged 65–87 years were enrolled in the study . Echo intensity (EI) and muscle thickness (MT) of the quadriceps femoris were measured using B-mode ultrasonography. A stepwise multiple linear regression analysis with EI as a dependent variable revealed that MT was a significant variable for Japanese participants (R Aging is an irreversible physiological phenomenon that causes functional impairments and morphological changes in skeletal muscle. In a cross-sectional study carried out by Lexell et al. , the redMuscle echo intensity (EI) has been widely employed for muscle quality evaluation using B-mode ultrasonography. EI is considered to provide important physiological information on the quantity of adipose tissue and/or connective tissue within skeletal muscle –9. In pa2) and women (25.7 kg/m2) aged >18 years, while Japan is ranked 166th for BMI in men (22.6 kg/m2) and women (21.7 kg/m2) [While the effect of aging on muscle quality is well-documented, there is limited evidence on race-associated differences in muscle quality. Melvin et al.  comparedThe purpose of this study was to compare muscle quality (EI), functional capacity, and morphological characteristics in Japanese and Brazilian older individuals. We hypothesized that Japanese participants would have lower EI, higher functional capacity, and better morphological characteristics compared with Brazilian participants.The study had a cross-sectional design. Japanese participants were recruited through the Nagoya City Health Promotion Bureau from 2017 to 2019 and Brazilian participants were recruited using a database for participants involved in previous studies at the Exercise Research Laboratory (LAPEX), Federal University of Rio Grande do Sul from 2018 to 2019. All participants had no history of brain and heart disease, were able to do light–intensity exercise without problems, and had no anxiety about their lower back.2; 19 women: age, 71.7 ± 6.1 years; height, 152.3 ± 4.2 cm; weight, 50.4 ± 7.4 kg; BMI, 21.7 ± 3.0 kg/m2) and 42 Brazilian older individuals  participated in the study. All participants in the Japanese group were Japanese. The participants in the Brazilian group comprised 30 white, 6 black, 5 multiracial, and 1 Japanese ancestry. The participants were healthy, habitually exercised for a few days per week, and were free of neurological, cardiovascular, and lower-extremity diseases. Before enrolment, all procedures, purposes, risks, and benefits associated with the study were explained and written consent was obtained. The study was approved by the Institutional Review Board of the Research Center of Health, Physical Fitness & Sports at Nagoya University (30–09) and the Federal University of Rio Grande do Sul (3.809.300), and was conducted in accordance with the guidelines of the Declaration of Helsinki.Forty-two Japanese older individuals  was used to measure body weight, %body fat, and lean body mass (LBM) for Japanese participants essentially as described previously . Eight eA whole-body DXA machine  was used to measure body weight, %body fat, and LBM for Brazilian participants. The participants removed all metal objects and other items that could interfere with the scan, and were positioned in the center of the scanning table with their arms placed palm-down at their sides. Age, height, weight, sex, and ethnicity were entered into the computer, and the default software of the device was used to determine indices of body composition, including body weight, %body fat, and LBM. DXA is considered the gold standard methodology for measurement of body composition. Regarding precision errors, the coefficient of variation (CV) values were 0.82% for total fat mass and 0.86% for %body fat in 52 men and women aged 34.8 ± 8.4  years .Real-time B-mode ultrasonography  with an 8–12-MHz linear-array probe  was used to measure the muscle thickness (MT), SCAT thickness, and EI at the anterior and lateral regions of the right mid-thigh essentially as described previously , 20, 21.The image files stored in the ultrasound device were subsequently transferred to a personal computer  for storage. The SCAT thickness, MT, and EI of the target muscles were analyzed using ImageJ software . MT of the RF and VL was defined as the distance between the superior border of the subcutaneous fascia and the deep aponeurosis. In the VI, MT was defined as the distance between the inferior border of the superficial aponeurosis and the superior border of the femur , 9.The MT and SCAT thickness of the QF were calculated according to the following equations:The first step in the analysis of EI involved smoothing to convert the original grayscale ultrasound image to a 256-value grayscale image using the “smooth function” in ImageJ. The region of interest, which included as much muscle as possible but avoided visible bone and fascia, was determined using polygon selections to calculate the EI of each target muscle , 22. TheThe EI of the QF was calculated using the following equation:The participants performed four functional tests in a gymnasium or laboratory  as described previously , 23. ForThe short version of the International Physical Activity Questionnaire (IPAQ) was used to obtain information on physical activity levels of the participants. We calculated and evaluated data on vigorous activity (days/week and min/day), moderate activity (days/week and min/day), and sedentary time (min/day) in this study.d was calculated to evaluate effect size between the two groups. The Pearson product-moment correlation coefficient (r) was used to determine the association between variables. A partial correlation analysis was carried out with a control factor of SCAT thickness to estimate the effect of SCAT thickness on EI attenuation. A stepwise multiple regression analysis was performed by entering variables into a forward stepwise regression analysis if they made a significant contribution to the explained variance  on two occasions. For the first step in the analysis, the dependent variable was EI of the QF and the independent variables were age, grip strength, sit-to-stand, MT of the QF, and SCAT thickness of the QF, with their selection based on our previous studies [All values are reported as mean and standard deviation. An unpaired-sample Student’s t-test was used to compare variables for demographic, functional, and morphological parameters between the two groups. Cohn’s  studies , 5 and s2; Brazilians: 27.4 ± 2.9 kg/m2; P = 0.001), abdominal girth , %body fat , and LBM  than Brazilian participants.Japanese participants had significantly lower frequency of vigorous activity  and moderate activity  than Brazilian participants.Overall, functional test performance was better in Japanese participants than in Brazilian participants. For example, there were significant differences between Japanese and Brazilian participants in the sit-up  and supine-up  tests. There was no significant difference in grip strength between the two groups .There were no significant differences in any MT parameters between Japanese and Brazilian participants. Meanwhile, anterior SCAT thickness , lateral SCAT thickness , and QF SCAT thickness  were significantly lower in Japanese participants compared with Brazilian participants.The EIs of the RF, VL, and QF in the participants are shown in There were several significant relationships between EIs and functional parameters. In Japanese participants, EI of the RF was significantly correlated with supine-up , sit-to-stand , and grip strength , and EI of the QF was significantly correlated with grip strength . In Brazilian participants, EIs of the VL and QF were significantly related with grip strength .To assess the effect of SCAT thickness on the relationships between EIs and MTs, we performed at partial correlation analysis with control factor of SCAT thickness for each measurement site. The partial correlation coefficients (Pearson’s r) between EIs and MTs of the RF, VL, and QF were –0.625 (–0.615), –0.563 (–0.564), and –0.650 (–0.651)  in Japanese participants, respectively, and –0.722 (–0.667), –0.734 (–0.718), and –0.698 (–0.651)  in Brazilian participants, respectively.2 = 0.424) and by MT and SCAT thickness of the QF in Brazilian participants .A stepwise multiple linear regression analysis was performed with EI of the QF as the dependent variable, and age, grip strength, MT and SCAT thickness of the QF, and sit-to-stand as independent variables . In the 2 = 0.381) and by LBM and BMI in Brazilian participants .The second step of the stepwise multiple linear regression analysis was performed with EI of the QF as a dependent variable and exclusion of independent variables that showed statistical significance in the first step of the analysis (MT and SCAT thickness of the QF). In the second step of the analysis, EI of the QF was explained by sex and age in Japanese participants  in the study. Furthermore, relative fat consumed in total energy intake was relatively higher in second-generation immigrants than the first-generation immigrants [The Brazilian participants had poor results for body composition and physical function. A reason for these findings may be the changes in physical characteristics of Japanese immigrants whose dietary habits have changed from Japanese to Brazilian style. Ferreira et al.  showed smigrants , suggestInterestingly, there were no significant difference in MTs of any regions between the two groups, even though Japanese participants had significantly smaller LBM than Brazilian participants. This result implies that relative muscle mass to the thigh was larger in Japanese participants than in Brazilian participants. Similar results were reported by Visser et al. , who demContrary to our expectations, the EI values did not satisfactorily explain the functional capacity in Japanese and Brazilian participants . These rThe EIs of the RF, VL, and QF in the present study were inversely correlated with the MTs , consist2, which was slightly higher than the mean BMI in Japanese male participants (23.0 kg/m2). His grip strength (37.8 kg), MT (33.9 mm), and EI of the QF (62.1 a.u.) were similar to the mean values in Japanese male participants. Furthermore, he had hyperlipidemia and diabetes, but did not have hypertension, which would be a normal physical condition considering his age. Therefore, this Brazilian male participant with Japanese ancestry had a similar body profile to that of Japanese male participants.We compared a Brazilian male participant who was a third-generation Japanese-Brazilian (66 years of age) and Japanese male participants. This makes it possible to investigate the effects of lifestyle-related differences on body composition and muscle quality. His height and weight were 163.0 cm and 65.7 kg, respectively, and thus his BMI was 24.7 kg/mIn the stepwise regression analysis, MT of the QF was extracted to predict EI of the QF in both Japanese and Brazilian participants. The same relationship was found in a simple regression analysis in the present study , and sim1H-magnetic resonance spectroscopy [According to a study based on spin-spin relaxation time (T2) using magnetic resonance imaging, there was a significant positive relationship between T2 and age in the gastrocnemius muscle of humans , reflecttroscopy . Importatroscopy , suggestIn Brazilian participants, SCAT thickness of the QF was extracted as another independent variable to explain EI in the first step of the stepwise regression analysis. This result was unexpected, because there were no significant correlations between EIs and SCAT thickness at any regions of the QF . AlthougIn the second step of the stepwise regression analysis, two completely different independent variables were selected between Japanese and Brazilian participants. For Japanese participants, sex and age were extracted to explain the EIs. Age is known as an index that affects muscle quality , 35, 36.For Brazilian participants, LBM and BMI were selected as independent variables to explain the EIs. Rech et al.  reportedThere were a few limitations to this study. The first limitation is that two different devices were used to measure the body compositions of Japanese and Brazilian participants. This was an unavoidable situation because of the difficulties associated with bringing BIA or DXA systems to another country. The agreement between BIA and DXA findings was examined in many previous studies. For example, BIA and DXA were found to be interchangeable methodologies  and agreThe second limitation is that attenuation of EIs may have been induced due to differences in SCAT thickness, particularly in Brazilian participants. SCAT thickness was significantly lower in Japanese participants compared with Brazilian participants and this could have affected the EIs. However, as shown in the results, the Pearson’s r values and partial correlation coefficients controlled by SCAT thickness were sufficiently small that they could be ignored for both groups. We concluded that the effect of the SCAT thickness difference between the groups on the EIs was negligibly small.Finally, the results for the EIs of the RF and VL were not consistent between the Japanese and Brazilian participants. This inconsistency is unlikely to arise from the effects of the SCAT thickness difference between the races. In our previous study on Japanese older men and women, the EIs of the RF and VL were approximately 70 a.u. and 60 a.u., respectively , 9, and In conclusion, the results of the present study revealed that MT, sex, and age were factors that explained the EIs in Japanese older individuals, while MT, SCAT thickness, LBM, and BMI were factors that explained the EIs in Brazilian older individuals. For Japanese participants, MT was the only changeable factor according to lifestyle, while all four variables are changeable by lifestyle in Brazilian participants. For Brazilian older individuals, the amounts of muscle and adipose tissue would have strong effects on muscle quality. These results suggest that muscle size is the most important factor to maintain higher muscle quality in both Japanese and Brazilian older individuals.S1 Data(XLSX)Click here for additional data file.


Hemiplegic gait is the most common sequela of stroke. Patients with hemiplegic gait are at a risk of falling because of poor balance. The theory of cognitive-motor networks paved the way for a new field of research. However, the mechanism of the relationship of cognition with gait or posture control networks is unclear because of the dynamic characteristics of walking and changing postures. To explore differences in the balance function and fall risk between patients with and without cognitive impairment after stroke, we utilized the Berg balance scale, Timed “Up and Go” test, and 10 m walking test. Patients were divided into two groups: the observation group , comprising patients with cognitive impairment after stroke, and the control group , comprising patients without cognitive impairment after stroke. We found that patients with cognitive impairment had worse balance function and a higher risk of falls. They needed a longer time to turn around or sit down. Our findings indicated that posture control in turning around and sitting down required more cognitive resources in daily life. Approximately 60% to 80% of patients after stroke cannot ambulate independently after completion of rehabilitation, and many of them have hemiplegic gait, which limits motor function . DecreasIn earlier reports, most researchers thought that our actions were controlled by the motor system alone. However, recently, many researchers have found that a sensory-cognitive-motor network system is necessary to ascertain the accuracy of an action –8. The sA retrospective case-control study design was used. We collected the data of 32 patients with hemiplegic gait after stroke. Patients were divided into two groups: the observation group, comprising patients with cognitive impairment (CI), and the control group, comprising patients without CI. From the department of rehabilitation from May 2019 to January 2020, we randomly selected 16 patients, including ten men and six women, with a Montreal Cognitive Assessment (MoCA) score of 15–25, and 16 patients, including nine men and seven women, with an MoCA score ≥ 26. Inclusion criteria were (1) the definitive diagnosis of first-ever stroke, (2) localization of stroke in the basal ganglia region of the right or left hemisphere, (3) age of 40–60 years, (4) ≤3 months poststroke, (5) junior high school education, (6) lower-limb Fugl-Meyer motor assessment score of 20–30, and (7) independent walking for several meters. Exclusion criteria were (1) prestroke vascular dementia, (2) aphasia, (3) traumatic hemorrhage, (4) sensory impairment, (5) medical instability hindering participation, and (6) visual or hearing impairment.MoCA was first developed by Nasreddine et al. . It is aThe Berg balance scale (BBS) was used to assess balance function for activity limitations . It is aThe fall risk was assessed with the Timed “Up and Go” test (TUGT) , constitThe 10 m walking test (10MWT) was used to assess the walking function. Patients walked from the taped line on the floor to an invisible line drawn 10 m away, which they were unaware of. Testers recorded the time with a stopwatch between the patient's first step crossing the line and the patient's first leg crossing the invisible line .t-test was performed to evaluate between- and within-group differences before and after the treatment of patients in the observation and control groups. A P value < 0.05 was considered statistically significant.The statistical analysis and graphing were performed using GraphPad Prism 8.0 . Continuous variables are expressed as the mean ± standard deviation. The unpaired P = 0.039; P = 0.005; P = 0.18; WT, P = 0.19). TGUG, TT, and ST were significantly longer in the observation group than in the control group . The 10MWT scores did not differ between the two groups  requires a complex control of the center of body weight. The TUGT, which detects the risk of falls in individuals with stroke, includes several parts of autonomous posture control, such as the movement from sitting to standing (which requires the participation of expected posture adjustment ability), continuous walking (which requires dynamic control ability), and turning and sitting  . The preFalls after stroke are common . There aAn inappropriate preparation can cause a fall. However, studies are limited on accurate postural assessment, balance assessment, or resource allocation of cognitive function after stroke. Mechanisms mediating the preparation of coordinated motor function have not been clearly elucidated. It is important to understand the potential preparatory mechanisms for coordinated actions. Motor mechanisms in the brain are viewed as a slave system under the dictate of cognition. Accordingly, modality-specific sensory modules channel information to the central systems for attention, memory, language, concepts, and decisions, which, in turn, drive the motor output . PreviouThe simple view of a balance system that one or a few balance centers in the central nervous system were responsible to control balance is limiting and can partially account for our limited abilities to assess risks of falling accurately, to improve balance, or to reduce falls. Cognitive dysfunction, particularly executive dysfunction, is obscured by other sequelae after stroke. Mild CI may often be overlooked during evaluation in clinical settings . With fuOur study had some limitations. First, it was a retrospective study, which would provide a lower level of evidence than a randomized controlled trial. Second, we grouped patients only based on MoCA as the screening tool. In our subsequent studies, different cognitive domains were evaluated to understand the influence of different cognitive disorders on postural control. Finally, we did not collect the executive function, spatial orientation, and attention separately to clarify the effects of different cognitive functions on posture control and gait. We will continue to explore the characteristics of postural control disorders with CI after stroke. At the same time, in our further research , functional magnetic resonance imaging will be used to scan the brain of patients with or without CI after stoke. We will analyze the data, observe the activation of the regions of interest, and finally explore the cognitive-postural control brain network to find new therapeutic targets.Patients with poststroke CI had poorer balance and a higher risk of falls. In daily activities, the risk of falls is significant during posture control, including turning and sitting. Our study suggested that posture control in turning around and sitting down required more cognitive resources in daily life. In physical therapy of balance and motor control after stroke, the assessment of cognitive function should not be neglected.


Background: The published data on the long-term outcomes of glycogen storage disease (GSD) patients is sparse in the literature. The aim of this study was to analyze the long-term (over 20 years) follow-up of patients with hepatic types of GSD-I, III, VI, and IX—from childhood to adulthood, managed by one referral center. Patients and methods: Thirty adult patients with hepatic GSD were included in the study. A retrospective chart review of patients’ medical records has been performed. Results: During the long-term follow-up, the most frequent complications observed in a group of 14 GSD I patients were nephropathy with blood hypertension (10/14), hyperuricemia (8/14), and development of hepatocellular adenomas . All individuals but four presented with normal height. Two patients with GSD Ib suffered from inflammatory bowel disease (IBD). Nine (64%) GSD I patients were in balanced metabolic condition at the age of 18. Regarding GSD III/VI/IX, the most frequent complication was short stature observed in 5 out of 16 patients. All patients but one with GSD VI were in balanced metabolic condition at the age of 18. Conclusion: The long-term outcomes of patients with GSD depend mainly on proper (adjusted to each type of GSD) dietary management and patient compliance. However, in GSD type I, even proper management does not eliminate all long-term complications in adulthood. Glycogen storage disorders (GSD) are rare inborn errors of carbohydrate metabolism; there are eight liver types: Ia and b, III, IV, VI, IX, XI, and 0, respectively. Each type is a distinct entity with a distinct metabolic block (deficiency of a different enzyme). GSD type I is caused by the deficiency of glucose-6-phosphase (G6P in subtype Ia) or G6P transporter/translocase through the mitochondrial membrane (G6PT in subtype Ib) . That isSince maintaining a stable blood glucose concentration is a goal of the therapy, uncooked cornstarch (CS) is a mainstay of nutritional treatment in type I, in which hypoglycemia occurs faster than in types III/VI/IX, and these episodes are more severe because ketogenesis has not the time to be triggered, the fasting tolerance is being too short . HoweverThese are so-called ketotic types since ketone production during hypoglycemia is higher than in type I. GSD type III is caused by decreased/lack of activity of debranching enzyme; type VI is caused by glycogen phosphorylase inactivity; and in type IX phosphorylase kinase (pK) is dysfunctional. These GSDs are characterized by hepatomegaly and episodes of hypoglycemia, which are less severe than in type I and their frequency decreases with age ,9. In type III, cardiomegaly and cardiomyopathy may occur, and creatine kinase (CK) level is usually increased. Type VI is clinically the mildest one, and some adults are almost asymptomatic. GSD IX includes four subtypes and the most common one is an X-linked form , accounting for 75% of all cases. The symptoms include hepatomegaly, mild psychomotor retardation, short stature, and mild muscle hypotonia. The patients’ symptoms become milder with age . In GSD The diagnosis of GSD is established based on the patient’s clinical phenotype and the results of biochemical analyses, and is finally confirmed (if available) by molecular analyses.Typical symptoms common for all GSD types but one (type 0) are hepatomegaly (not present in type 0), hypoglycemia, short stature, and frequently, due to the dietary management with CS—obesity  ,8,9.Biochemical abnormalities include hypoglycemia, hyperlipidermia, hyperuricemia (in type I), elevated serum transaminases, increased LA level (type I), increased CK level , and neutropenia in type Ib ,10.Prolonged fasting tests are not usually performed and are not useful (and even dangerous) for differentiating between type I and type III. Enzymatic assays are no longer performed, and they have been replaced by molecular analyses. Furthermore, abdominal ultrasonography and transient elastography by FibroScan (if available) are performed to assess hepatomegaly and nephromegaly; the liver’s echostructure and its steatosis/fibrosis state; and the presence of any liver lesions. Various diagnostic tools are used to detect GSD’s complications. In our center, anthropometric assessment and calorimetry are available and are performed to analyze each patients’ nutritional status. Densytometry is provided to assess each patient’s skeletal condition. In order to detect HCA/HCC, both CT and MRI scans are used, and AFP (Alpha-fetoprotein) level is regularly checked.The mainstay of medical management in GSD is to prevent episodes of hypoglycemia and to maintain stable blood glucose level; therefore, dietary treatment is the basic therapy. Specific dietary interventions for each type are mentioned above. Nonetheless, the disease complications, if present, need some additional management, including allopurinol in patients who develop hyperuricemia, ACE-inhibitors in patients with blood hypertension, 5-ASA in GSD Ib patients who develop inflammatory bowel disease (IBD), G-CSF in GSD Ib patients with neutropenia, and hypolipemic agents in patients with hyperlipidemia.The aim of this study was to analyze the long-term outcomes (over 20 years) of patients with hepatic types of GSD-I, III, VI, and IX—from childhood to adulthood, managed by one referral center. This is a unique opportunity, since our group is large for a rare disease, and the monitoring and follow-up have been provided by one site only.Thirty adult patients with hepatic GSDs, including 8 with GSD Ia, 6 with GSD Ib, 8 with GSD III, 7 with GSD VI, and 1 with GSD IX, were included in the study. The observation period was between 1982 and 2018. Among 30 patients diagnosed and treated since infancy, 25 (83%) have still followed-up with and been monitored by our Institute  within the outpatient clinic since the age of 18.Based on medical charts of patients, the following parameters were analyzed: signs and symptoms at time of diagnosis; age at diagnosis; dietary and medical management; anthropological and biochemical parameters, including body mass index (BMI) and serum aspartate aminotransferase (AST), serum alanine aminotransferase (ALT), serum triglyceride (TG), serum total cholesterol (TC), lactate (LA), and uric acid (UA) levels; and long-term complications.Ethical approval  of the study protocol was obtained from the Children’s Memorial Health Institute Bioethical Committee, Warsaw, Poland. There were 14 patients with GSD type I, including eight individuals with type Ia and six with type Ib. The majority (5/8) of patients with type Ia were diagnosed within their first year of life , and the mean age of GSD Ib diagnosis was approximately 3 (age range: 6 months old to 7 years old) . All patients had dietary management  introduced once the diagnosis was established, and CS consumption commenced since the age of 6–8 months . Only one GSD Ia patient had nightly glucose infusions for his first year of medical management, which were then substituted with CS. Over half of the patients (9/14) were in balanced metabolic condition at the age of 18 . Serum ASerum AST and ALT were within normal laboratory values in all GSD Ib patients. Elevated TG levels were present in 3/5 patients, and TC levels were within normal values in all of them. Serum lactate was elevated in 3/4 patients, and uric acid in 1/5 patients.The majority of GSD I patients have been followed at our Metabolic Outpatient Clinic, where the diet was supervised and biochemical parameters monitored—in most cases both the compliance and laboratory results were satisfactory . During Almost all patients with GSD type I who developed hyperuricemia were given allopurinol to prevent arthritis. We tried to manage hyperlipidemia with dietary interventions; however, if TG level was above 500 mg/dl, which according to literature  is assocThe majority of those complications developed either during childhood or adolescence, although the proper diet had been introduced since each diagnosis had been established. Two women with GSD Ia (patient 4 and 5) had menorrhagia; one gave birth to a healthy child without any complications. One patient with GSD Ib (patient 10) developed hearing problems . One GSD Ia patient (patient 2) presented with bleeding problems (excessive epistaxis). Two patients with GSD Ib (patient 11 and 12) suffered from IBD, and patient 12 had arthritis. Both patients had active IBD at the age of 18 and have experienced a lot of flare-ups during adulthood. There were 16 patients diagnosed with GSD III/VI/IX: 8, 7, and 1, respectively. In comparison to patients with GSD I, the diagnosis was established later—the mean age at diagnosis was approximately 3 (age range: 4 months old to 8 years old).All patients but one had balanced metabolic conditions at the age of 18 . Serum ADuring the long-term follow-up the most common complication in these patients was short stature, present in 5/16 patients; in three of them it was a considerably short stature . One patient with type VI (patient 11) developed diabetes mellitus (DM) type 1 at the age of 4 and was treated with insulin. The manuscript presents data from a 20-year follow-up of 30 patients with hepatic GSDs. So far, data published on the long-term outcomes of hepatic GSD patients is sparse. According to the natural course of GSD I, complications develop with time , and include short stature, renal disorders , and increased risk for HCA and HCC development ,14,15. NIn our cohort, all patients had proper dietary management  introduced as soon as the diagnosis of GSD was established. In GSD I patients, CS was introduced at the age of at least 6 months old to prevent diarrhea. Patient compliance was generally satisfactory. Moreover, all of them closely and regularly followed-up, even as adults. Due to that, the patients’ adherence improved (they are taught how to manage the diet and control themselves), and by transition  the vast majority of them had balanced metabolic homeostasis. Such nutritional management prevents hypoglycemia, and thus leads to normalization of biochemical markers  .In our study, nephropathy with hypertension was the most common complication in GSD I patients. The European study for GSD reported in 2002 that all GSD Ia patients developed microalbuminuria or proteinuria by 24 years of age . Once miIn the reported cohort, HCA was diagnosed in five patients with GSD Ia and was one of the most common complications in these patients. In all of those patients, they developed it at adolescence, and it was associated with high serum levels of TG (>500 mg/dL), which is consistent with the literature. HCA formation was one of the first complications reported in GSD type I, but also in type III. Initial studies on GSD type I adults have demonstrated that HCA developed in over 70% of those patients . The lesOne GSD Ia patient presented bleeding problems (excessive epistaxis), which may occur in GSD type I. The impaired coagulation in type I GSD is due to dysfunctional platelets function.Inflammatory bowel disease (IBD) due to neutropenia and impaired neutrophils function is a common complication in GSD Ib . The treThe summary of GSD Ia’s outcome in adults by Weinstein et al. has demonstrated that complications in GSD Ia can be delayed or prevented with a properly balanced diet . Our resShort stature was observed both in type I and in ketotic types, but it was not as a common complication as we may expect, which only confirmed the proper management of these patients by our site.The important, although rather obvious observation, is that there are significant differences in both short and long-term complications and outcomes between GSD type I and types III/VI/IX. In our group, no patient with type IIIa developed cardiomyopathy, and the only significant complication in these patients was short stature. Peripheral myopathy may occur in GSD III. Although few patients in our group had increased CPK levels, none of them developed miopatic disorders. Patients with GSD III with normal CPK levels were almost asymptomatic.The majority of individuals with ketotic GSD types, especially with type VI, were asymptomatic, and they did not even consider themselves to be ill when becoming adults. However, most patients followed their diets once educated at infancy; they stuck to the management they got used to, unlike patients with type I. Although the proper diet was introduced, some complications could not be prevented. In our cohort, the majority of GSD I complications developed either during childhood or adolescence. In some cases, it was due to unsatisfied compliance, but in the majority of the patients it was due to the natural course of the disease. This observation demonstrates that type I is much more severe than types III/VI/IX, and unlike them, diet, though it improves the outcome, does not eliminate some complications.The advantages of our work are a significant number of patients, long-term follow-up, and proper, up-to-date clinical management provided by a single-referral center.1. Long-term outcomes of patients with GSD depend mainly on proper (adjusted to each type of GSD) dietary management and patient compliance. However, in GSD type I, even proper management does not eliminate all long-term complications in adulthood.2. Nephropathy with hypertension was the most common complication in our cohort of GSD I patients.3. HCA was diagnosed in 5 out of 14 patients with GSD Ia, and it was one of the most common complications in these patients, associated with high levels of serum TG (>500 mg/dL).4. Regarding GSD III/VI/IX, the quite common complication was short stature, present in 5/16 patients. All patients but one with type VI were in balanced metabolic condition at the age of 18.


Low back pain (LBP) is a major contributor to chronic pain and disability. The purpose of this study was to evaluate health-care access and utilization among patients with LBP in Iran. We also sought to study the pattern and characteristics of care-utilization behavior in these patients.Data from the Community Oriented Program for Control of Rheumatic Diseases (COPCORD) were used for this study. Three cities  were selected to represent the Iranian population, with different socioeconomic status and ethnic, cultural, and religious background. Demographic data, acute or chronic LBP, disability index, and utilizing care from conventional medicine (CM), allied health providers (AHP), and complementary and alternative medicine (CAM) providers were recorded.Of 9101 patients, 38.6% reported LBP. Only 3.3% did not utilize care of any kind, 66.7% referred to CM providers, 20.8% to AHP, and 9.2% to CAM care. Health-care utilization was higher in female patients, older age, higher education, and higher disability index.The findings of this study indicate a high rate of health-care utilization among patients with LBP in Iran. CM is the most prevalent health-care resource sought by patients. These findings could be used as a framework in developing more efficient health-care programs according to the needs of specific populations. Low back pain (LBP) is one of the most common symptoms experienced by people of all ages globally . A speciWhile the high prevalence and costs of LBP have encouraged clinicians of many medical disciplines to develop and study the disease, an ideal prevention or treatment protocol is as elusive as ever. A variety of non-pharmacologic and pharmacologic therapies have been studied by researchers around the world, with varying results . This haThe Community Oriented Program for Control of Rheumatic Diseases, or COPCORD, was an initiative started in 1983 by the world health organization (WHO) and international league of associations for rheumatology (ILAR) with the same concepts in mind. As a ‘community program for control of rheumatic diseases’, this program was created for recognition, prevention, and control of rheumatic disorders in the developing countries. So far, 21 countries have participated in COPCORD, and multiple publications have elucidated different aspects of rheumatologic disorders in the developing world [The results of the COPCORD initiative have shaped our understanding of the global epidemiology of LBP in the developing world. A recent study combining the Latin-American COPCORD data shows that LBP affects 6.59% of the population, with a high impact on the quality of life of young adults. Also, LBP has a significant association with social variables including gender inequality index, human development index, and income inequality , 11.We sought to determine the epidemiology of LBP and the disability caused by the symptoms, as well as access and utilization of different health-care resources available to the individuals. This study is based on the results of Stage I, Phase III of the Iranian COPCORD study, which evaluated rheumatologic disorders in three cities.Three cities from the urban phase of the Iranian COPCORD were used for this study. Zahedan, which is situated in south-eastern Iran, with an estimated population of 681,460. Residents of Zahedan are mainly from the Balouch or Fars ethnicities, both considered Caucasian. This city is an underdeveloped region in Iran, and the socioeconomic status is very low . The secIn general, the Iranian health system consists of a universal health-care system which relies on a hierarchical network starting from rural medical centers to urban, tertiary referral centers. While only medical doctors  are licensed to practice medicine, allied health professionals (AHPs), including nurse practitioners, midwives, and physical therapists can visit and treat patients under the supervision of a medical doctor. Complementary and alternative medicine generally consists of unlicensed individuals treating patients with herbal and unconventional therapies.The sampling plan for all three cities were similar and consisted of cluster sampling based on the population of neighborhoods in the city. Districts and neighborhoods were extracted from the national postal service zip code bank. All interviews were performed by trained physicians or nurse practitioners. The interviewers were local to each city to know the local culture and inhabitants. Interviews were conducted on Fridays (the weekend in Iran) each week on a door-to-door manner, to ensure the presence of as many household members as available. The interviewing team went back to the same cluster on 3 consecutive Fridays to collect data from those who were absent the previous Friday. Every interviewer started data collection by introducing him/herself and showing an identification card, and then continued to fill in the CCQ. After completing the CCQs for each family, questionnaires were submitted to the team head. The head would check and confirm the CCQ.Data collection for each city was independent, and due to the limited resources, had to be performed sequentially. Therefore, the complete dataset was only ready when the last city was surveyed. Data from Zahedan was surveyed from October 2008 to September 2009. Sanandaj COPCORD data collection was performed between July 2011 and June 2012, and Yazd study was conducted Between July 2014 and July 2015. All data from the three cities were combined after the completion of data collection, and eligible individuals were then queried within this database.The Farsi translation of the COPCORD Core Questionnaire (CCQ) was used to screen subjects for low back pain. The Farsi version of the CCQ has been previously validated and has been shown to be a reliable and reproducible translation , 14. TheIn this study, we included individuals who answered positive to any questions in the C2j and C9j sections, which asks about pain/stiffness/tenderness in their back during the last week or the past. Section E, which is the modified Stanford Health Assessment Questionnaire – Disability Index (HAQ-DI), was used to measure the degree of disability due to LBP. Generally, HAQ-DI scores of 0 to 1 are considered no to mild disability, 1 to 2 as moderate, and > 2 as severe disability. Health-care access and utilization are surveyed in sections G2a and G2b. A positive answer to any of the questions in the G2a subsection was considered the utilization of conventional medicine (CM) care. A positive answer to G2b1  or G2b7 (chiropractor) was considered the utilization of an allied health professional (AHP). A positive answer to other questions in the subsection G2b was considered utilization of complementary and alternative medicine (CAM) providers. Demographic data, including age, gender, ethnicity, and marital status, were collected for each patient. Also, the level of education of each patient was recorded.t-tests were used to test for differences between groups. The significance level was considered alpha < 0.05 for all tests.Statistical analysis was performed using IBM SPSS Statistics for Windows, version 24 . Descriptive data were analyzed by survey data analysis methods. Student’s In total, 9101 patients were interviewed and completed the CCQ in the three cities of Zahedan , Sanandaj (5001), and Yazd (2000). The prevalence of LBP was 38.6% among all participants . Sanandaj had the highest prevalence (55.3% of participants), followed by Zahedan (38.8%), and Yazd (17.1%). In total, 71.7% were female and 28.3% were male. The educational level of the participant was as follows: 33.4% no high-school education, 34% some high-school, 19.2% high-school diploma, and 13% had higher education. In patients with LBP, 13.2% were single, 78.5% married, and 8.2% divorced/widowed.Only 3.3% of patients had not sought any form of care for their LBP, with 66.7% of patients referring to CM providers, 20.8% to AHP, and 9.2% CAM care utilization.P = 0.03). This association was not observed with AHP (P = 0.443) or CAM (P = 0.718) utilization.Table P < 0.001). Moreover, using CM (P = 0.03) and AHP (P = 0.004) care was significantly higher in patients with a high-school diploma, but using CAM care and education level did not have a significant association (P = 0.092).Also, older age was significantly correlated with CM, AHP, or CAM care utilization . CM and AHP care utilization were significantly higher among divorced/widowed patients . There was no association between CAM care utilization and being divorced/widowed (P = 0.468).CM, AHP, and CAM care utilization was significantly higher in patients with Zoroastrian ethnicity compared to all other ethnicities  and AHP care (P < 0.001), but not CAM (P = 0.21). Also, patients with a high-school diploma were significantly more likely to utilize CM (P = 0.003) and AHP care (P = 0.004), but not CAM (P = 0.092).Table While access is an indicator of the quality of a health-care system, utilization of the provided health-care depends on various socio-economic and cultural factors, in addition to the health system itself. Knowledge about the patterns of health-care utilization is vital in developing and modifying health-care policies, especially in low- and middle-income countries. To our knowledge, this is the first study to describe the epidemiology of LBP and the utilization of health-care in the Iranian population. Also, differences in characteristics of patients utilizing a variety of health-care resources were studies for the first time in three cities among individuals with a range of socioeconomic status, educational level, access to health-care, and ethnicities.LBP, as one of the most common musculoskeletal complaints, affects all ages and groups. The chronicity, pain and disability range from a days-long mild pain to a life-long, debilitating pain that leads to early retirement from work, with a multitude of physical, emotional, and psychological disorders that may ensue . The preOur results show a mean prevalence of 38.6% for LBP among the cities surveyed. This rate is one of the highest reported in the COPCORD studies. The only study with similar results is the Chinese study, performed in Beijing, which showed a 35% prevalence of LBP . More reSeveral demographic, socioeconomic, medical, and cultural factors have been associated with LBP. Medical comorbidities, unhealthy habits, low educational level, living in rural areas, obesity, other musculoskeletal complaint/disorders, female gender, and anxiety have been highly associated with LBP in different studies –11, 19. We found a high rate of utilization of health-care resources among COPCORD interviewees with LBP, with the vast majority (96.7%) of patients utilizing care. Iran has a universal health-care system with a vast network of public-sector health-care resources, which mitigates the financial impediments to health-care access. While we were unable to find studies reporting LBP-specific health-care utilization behavior, but this rate is in the range of middle- to high-income countries with a universal health-care system –22. The Female individuals in this study were more likely to utilize CM care, but not AHP or CAM, which is in line with some studies, which have reported the association between female gender and utilization of health-care , 24. HowThe association between ethnicity and health-care utilization has been demonstrated before. Being of an indigenous ethnicity was a risk factor for a lower rate of health-care access and utilization, as well as a higher burden of symptoms of the quality of life , 26. In Higher education and higher levels of disability were correlated with utilization of health-care resources in this study. These findings have been reported consistently in previous studies, which shows a tendency for higher educated patients to utilize care irrespective of financial status , 24, 27.Older age was also correlated with utilization of CM and AHP care in this study. Previous studies have also reported similar results in countries with a universal health-care system . This maThis study has some limitations. First, our study population may not be representative of the whole Iranian population. We tried to mitigate this by including data from three cities with different ethnic, cultural, and socioeconomic situations. Second, some individuals refused to answer some questions, especially about ethnicity and educational level. Also, we had to exclude disability index data for some individuals, due to incomplete forms, which reduces the power of this study. We have provided the subtotal number of complete data in the tables. Third, we had no way of quantifying the use of health-care resources, specifically whether the patient was satisfied with the provided care and how likely they are to utilize care again. Finally, due to the heterogeneity in data collection pertaining to the individual’s job and activity level, we did not include them in the analysis, which are potentially influential factors in the prevalence of LBP. This study also has some strengths. The COPCORD study is a well-established program, with standardized evaluations which produces results that are comparable globally. Additionally, the high number of individuals in this study, with an array of socioeconomic and cultural background also adds to the power of this study.In conclusion, this study provides meaningful insight on the current pattern of healthcare utilization behavior of the Iranian population, and the factors associated with the utilization of health-care resources in patients with low back pain. The results of this study would be a valuable addition to the literature used in developing health-care programs to combat the increasing incidence of low back pain in the developing world, in hopes of decreasing the pain and disability associated with the disease.


The sixth meeting of the International Society for Zinc Biology (ISZB-2019) was held on September 9–13, 2019 in Kyoto, Japan. The meeting attracted 215 participants, had four plenary speakers, ten scientific symposia, two oral sessions, and one poster discussion session. In this chapter, we describe the outcomes and events of this very successful meeting. The International Society for Zinc Biology (ISZB) was launched in 2007 with the goal of providing a deep understanding of the roles of zinc in life . Since tThe 6th conference was held in Kyoto and the academic theme was “The New Sunrise of Zinc Biology” . During Bulleted lists look like this: We invited four plenary lecturers who brought their fascinating research outcomes that have greatly impressed attendees. Tom O’Halloran  gave his talk entitled, “Zinc Fluxes in Control of Gamete Function: Mechanistic Insights into the Oocyte to Egg Transition and the Sperm-Egg Interaction.” Nickolas Tonks  gave a lecture titled, “From Phosphatase-Based Therapeutics to the Discovery of a Novel Metal Chelator. Eric Skaar  addressed the intersection of nutrition and infection at the host-pathogen interface. Ananda Prasad  lectured about zinc deficiency, and the talk was entitled “Discovery and Impact of Zinc on Health: Biomarkers of Zinc Deficiency”. All lectures enhanced our understanding of zinc biology. We organized ten symposia with session topics covering almost all the life science fields which stimulated attendees’ discussions. Covered topics and chairs of each symposium (SP) were as follows: SP1 (zinc sensor), M. Merkx and C. Fahrni; SP2 (neurology 1), A. Bush, E. Aizenman; SP3 (physiology and genetics), M. Hershfinkel, S. Kelleher; SP4 (young investigators), T. Fukada, T. Kambe; SP5 (immunology and inflammation), L. Rink, D. Knoell; SP6 (biochemistry), W. Maret, D. Fu; SP7 (cell biology and cancer), K. Taylor, I. Sekler; SP8 (diabetes), H. Yasui, L. Huang; SP9 (neurology 2), A. Takeda, JY. Koh; and SP10 (zinc therapy), H. Kodama, M. Moriyama. SP10 was co-organized by the Japanese Society for Zinc Nutritional Therapy (JZNT) [In addition to symposia, oral session speakers selected by the organizers presented a broad range of new discoveries and approaches using various models and exploring the role of zinc homeostasis in cell physiology and molecular biology.We had 81 poster presentations, which were exhibited throughout the four days of the conference allowing attendees to read them at their convenience, sharing new data and ideas regarding the role of zinc in a large number of biological systems. Poster presentations covered a variety of aspects of zinc biology, as in the case of the symposia, including zinc in health and disease; zinc proteins, transporters, receptors, channels, and compounds; and zinc signaling. A diverse poster presentation session provided a stimulating environment for the interaction among young researchers, students, and established scientists, representing a good training opportunity for the next generation of zinc biologists. Five young researchers were awarded the Metallomics Poster Prize .This year the Frederickson Prize was awarded for the first time as the official prize of the ISZB and was open to all disciplines related to zinc research. The prize is named after Christopher F. Frederickson, one of the founders of the zinc signal conferences, which finally became ISZB. In ISZB-2019, we awarded the prize to Wolfgang Maret (King’s College of London), a former president and one of the founders of the ISZB. On the final day, we also awarded Chris Frederickson an honorary ISZB membership.Not only the scientific events, but also the social events including an excursion, helped us to extensively interact with one another. The information about all the programs is available on the meeting homepage .After the conclusion of the ISZB-2019, we administered a survey to learn about the impression and thoughts that the attendees had during and after the conference. The majority of the responses were very positive, especially for the use of the same venue for the conference and accommodation, which provided attendees many opportunities to interact with one another. Based on the results of the survey, we are proud to conclude that the ISZB-2019 was a very successful meeting (See group photo in the meeting home page ).Finally, many thanks to all the attendees and the generous funding from academic foundations and companies who supported ISZB-2019. Without their support the conference would not have been as successful.


Although a large number of studies have shown brain volumetric differences between men and women, only a few investigations have analyzed brain tissue volumes in representative samples of the general elderly population. We investigated differences in gray matter (GM) volumes, white matter (WM) volumes, and intracranial volumes (ICVs) between the sexes in individuals older than 66 years using structural magnetic resonance imaging (MRI).Using FreeSurfer version 5.3, we obtained the ICVs and GM and WM volumes from the MRI datasets of 84 men and 92 women. To correct for interindividual variations in ICV, GM and WM volumes were adjusted with a method using the residuals of a least-square-derived linear regression between raw volumes and ICVs. We then performed an analysis of covariance comparing men and women, including age and years of schooling as confounding factors.Women had a lower socioeconomic status overall and fewer years of schooling than men. The comparison of unadjusted brain volumes showed larger GM and WM volumes in men. After the ICV correction, the adjusted volumes of GM and WM were larger in women.After the ICV correction and taking into account differences in socioeconomic status and years of schooling, our results confirm previous findings of proportionally larger GM in women, as well as larger WM volumes. These results in an elderly population indicate that brain volumetric differences between sexes persist throughout the aging process. Additional studies combining MRI and other biomarkers to identify the hormonal and molecular bases influencing such differences are warranted. Several studies have documented the brain’s anatomical differences across various age groups and sexes and multiple theories aim to explain the cause of these disparities. An example is to attribute these differences to hormones, for instance, the action of estrogen on synaptogenesis and the effects of the absence of this hormone during menopause ,2. ThereThe estimated volume of the cranial cavity, called the intracranial volume (ICV), is one of the main variables studied in the neuroanatomical variations between men and women . OverallStudies that have compared brain volumes between sexes after ICV correction methods have yielded mixed results. There is a tendency for larger GM volumes in women, while the results regarding WM volumes have been more varied ,9,11-14.In the present study, neuroanatomical differences between sexes were investigated in a group of 176 individuals without neurodegenerative diseases (84 men and 92 women) residing in São Paulo, Brazil, and above the age of 66 years. We analyzed the values of the ICV, total GM volume, and WM volume without any adjustments. The volumes of GM and WM were then analyzed after using an ICV correction named as the residual method, which has been validated in recent studies ,10,19. Fa priori hypothesis. Finally, we predicted that fewer years of schooling would be associated with smaller brain volumes, both in men and women.According to the present literature, we expected to find larger non-adjusted brain volumes in men. However, after correction for ICV, we expected to find significantly larger volumes of GM in women. Regarding WM, due to the inconsistent findings in the literature, we did not establish an We analyzed MRI scans of 176 individuals (84 men and 92 women) between 66 and 80 years of age. The group of elderly individuals described herein was drawn from a population-based sample recruited for a morphometric MRI study evaluating the association between cardiovascular risk factors and GM volumes in elderly individuals aged 65-75 years from a circumscribed, socioeconomically disadvantaged area of São Paulo, Brazil . We also gathered information about the participants’ sex, socioeconomic status, and years of schooling.All participants were healthy, with no signs of dementia or other neurodegenerative diseases. Further details regarding inclusion and exclusion criteria as well as the clinical evaluation of patients, can be found in Squarzoni 2016  . The strhttp://surfer.nmr.mgh.harvard.edu/).From the SPGR sequence described above, we extracted the values of the ICV, GM volume, and WM volume. The data were analyzed with FreeSurfer version 5.3  for the WM and GM volumes, after which we used the following mathematical formula:adj is the ICV-corrected volume of the GM or WM, Vol is its original volume, without correction, β is the inclination of the linear regression, ICV is the intracranial volume of each individual, and  is the mean of the ICV of all participants  IBM Statistics for Windows version 25.0 . The significance threshold was set at 0.05.First, we ran an independent sample t-test to compare age and years of schooling between sexes. The qui-squared test was used to describe the differences in the socioeconomic status among men and women in the study sample.adj. This new volume (Voladj) was then used in the ANCOVA test as a covariable.Furthermore, we carried out an analysis of covariance (ANCOVA) comparing GM volume, WM volume, and ICV separately between sexes. Since we decided to use the residual ICV correction method, we needed to obtain the beta values for GM and WM volumes, both of which had very similar results . The GM and WM volumes were adjusted after we used the formula described in the previous section “ICV Correction Methods for the Total Volumes of WM and GM” with which we were able to generate the value of Voladj were again inserted into the ANCOVA as covariables, as we decided to use the residual ICV correction method for GM and WM volumes.Subsequently, we ran another ANCOVA, comparing the ICV, GM volume, and WM volume between sexes, after including age and years of schooling as confounding variables. The values of VolFinally, we ran Pearson’s correlation tests between ICVs, GM volumes, WM volumes, and each of the following demographic variables: age, years of schooling, and socioeconomic status. The GM and WM volume correlation analyses were repeated using partial correlation rates after employing the residual method.vs. 70.31±2.52 for men)  .vs. 5.42±4.13; p>0.01) and a higher socioeconomic status than women .Having more years of schooling was positively associated with a higher socioeconomic status , and white matter volume after the residual correction method (WMr) remained statistically significant.Even after the inclusion of age and years of schooling as covariables in a separate ANCOVA analysis, the difference between the values of intracranial volume (ICV), WM volumes, GM volumes, gray matter volume after the residual correction method (GMp=0.022) and socioeconomic status (p=0.002).Pearson’s correlation results are shown in p=0.022) and socioeconomic status (p=0.000); however, they were inversely associated with the participant’s age (p=0.042).The GM values were also directly associated with years of schooling (p=0.022)  .r and WMr) and age. However, there was still a significant difference, with a negative association between years of schooling and volumes of GMr (p=0.040) and WMr (p=0.020) and socioeconomic status, in addition to the values of GMr (p=0.008) and WMr (p=0.001)  .The present study demonstrated the presence of significant differences in neuroanatomical structure volumes between men and women in a population sample of participants above 66 years of age. Using raw volumes, we found larger GM and WM volumes and ICVs in men. However, after applying an ICV correction method based on partial brain volumes, we found larger relative volumes of GM and WM in women. Epidemiological data from the current study revealed a greater socioeconomic status and more years of schooling among the male population. This epidemiological difference was already expected in our participant sample, according to recent large population-based surveys in Brazil .Our results are consistent with previous findings of larger brain volumes in men than in women before any adjustments. Barnes et al.  reportedCurrently, there are few studies with large samples that have evaluated brain volume variations between sexes during the aging process and have specifically investigated participants above 50 years of age. In most of these studies, the unadjusted brain volumes of GM and WM were found to be larger in men, as reported by Greenberg et al. , VoevodsRegarding WM, the results of the volume differences between sexes are, so far, discrepant among studies in the literature ; therefoConversely, our findings of larger relative GM volumes in the female sample after the residual correction method was applied were consistent with several studies. The same result was observed by Pintzka et al. , Király It is important to highlight that the chosen method for ICV correction has a direct implication on the results , since tEven though the women in the current study had fewer years of schooling and a lower socioeconomic status overall, they had the largest volumes of GM after adjusting the data with the residual method. This result is consistent with many previous studies that have demonstrated a larger relative GM compartment volume in women than in men. Numerous studies point to a possible sex-dependent neuronal volume redistribution ,31. The Another possible hypothesis, according to the findings of Ryan et al. , would br and WMr), are more difficult to interpret and have little support from the previous literature . These negative results may be explained by the narrow age range of the present elderly sample. On the other hand, we found direct correlations between ICVs and years of schooling. This finding is relatively intriguing, given that previous studies have classified ICV as a measure of brain reserve rather than cognitive reserve  . A studyterature ,16,18. Mterature . We inteOne important limitation of the present study is the risk of systematic errors associated with the use of methods for automatic segmentation of brain volumes . AnalyseIn conclusion, in a population sample of 176 elderly individuals, including those with relatively low educational levels and without any neurodegenerative diseases, we demonstrated differences in overall brain volumes between men and women. This difference was maintained even after the correction of GM and WM volumes for the total ICV, which reveals that part of the dimorphism between sexes can be attributed to the characteristics of each sex and not merely to a difference in cephalic size. We have identified larger volumes of GM in women, even though this particular group has had the lowest levels of socioeconomic status and fewer years of education. Further studies evaluating hormonal and molecular genetic aspects are needed to explain why women, despite their smaller cranial size, present larger relative GM volumes than those of men, and to explain how this process occurs.Busatto GF conceived the study, participated in study design, interpreted the data and revised the manuscript. Buchpiguel M analyzed and interpreted the data, drafted and revised the manuscript. Squarzoni P, Tamashiro-Duran JH, Duran FL, Leite CC, Scazufca M, Lotufo PA, and Alves TC interpreted the data and revised the manuscript. All authors have read and approved the final version of the manuscript.