P < 1.0 × 10−7). Expect for rs4483616, other three loci were validated by the TaqMan probe genotyping assay (P < 0.05) in another cohort including 250 ITP cases and 250 controls. And rs5998634 T allele was more sensitive to glucocorticoids for ITP patients . Moreover, we identified three overrepresented signaling pathways including the neuroactive ligand-receptor interaction, pathways in cancer, and the JAK-STAT pathway, which involved in the etiology of ITP. In conclusion, our results revealed four novel loci and three pathways related to ITP and provided new clues to explore the pathogenesis of ITP.Immune thrombocytopenia (ITP) is an acquired bleeding disease due to immune-mediated destruction of antilogous platelets and ineffective thrombopoiesis. Although the etiology of ITP remains unknown, genetic variants are thought to predispose individuals to the disease. Several candidate gene analyses have identified several loci that increased ITP susceptibility, but no systematic genetic analysis on a genome-wide scope. To extend the genetic evidence and to identify novel candidates of ITP, we performed a pooling genome-wide association study (GWAS) by IlluminaHumanOmniZhongHua-8 combining pathway analysis in 200 ITP cases and 200 controls from Chinese Han population (CHP). The results revealed that 4 novel loci  were strongly associated with ITP ( Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by low platelet count . The incGenome-wide association studies (GWAS) are a powerful tool in searching for gene variants of complex diseases by comparing single-nucleotide polymorphisms (SNPs) . A numbeP = 0.0019). Moreover, we analyzed the relationship of loci and clinical therapy and found rs5998634 had a positive association with response to glucocorticoids , suggesting that this SNP may have predictive value for the response to steroid treatment.To extend the present genetic data and to identify the novel genetic and biological functional evidence of ITP, we firstly performed a pooling GWAS in 200 ITP patients and 200 control subjects from CHP using an IlluminaHumanOmniZhongHua-8 array scanning 862,620 SNPs across the autosomal region. By SNP-Map (single-nucleotide polymorphism microarrays and pools) analysis, our scanning revealed 4 novel loci  were strongly associated with ITP from CHP. Furthermore, we validated the relationship between rs117503120, rs5998634, and rs16866133 and ITP by the TaqMan probe genotyping assay  pathway analysis based on the GWAS data. The most potential candidate pathways associated with ITP were the neuroactive ligand-receptor interaction, the pathways in cancer, and the JAK-STAT pathway.In conclusion, our results suggest that these significantly associated loci, genes, and pathways may provide novel insights into the genetic etiology of ITP and novel clues for investigating the pathogenesis of ITP.This study was carried out in accordance with the principles of the 1964 Declaration of Helsinki and its later amendments or comparable ethical standards and was approved by the Ethics Committee of the Second Affiliated Hospital of Nanchang University (No. Review [2010] No. (002)). All participants signed a written informed consent. A total of 450 adult ITP patients, who met the diagnostic criteria of consensus of Chinese experts on diagnosis and treatment of adult primary immune thrombocytopenia , white blood cell (WBC), red blood cell (RBC), and hemoglobin (HB) were collected. The flowchart of two-stage sample collection was shown in Supplementary 9/L, or with a partial response, which was defined as PLT ranging from 30–100 × 109/L or at least doubling of the baseline count. The nonresponse group only contained patients without a response, which was defined as PLT lower than 30 × 109/L or less than doubling of the baseline count. The criteria for complete response, response, and nonresponse were judged according to previous criteria [A total of 183 inpatients with ITP in the second stage were treated with glucocorticoids, including high-dose dexamethasone (HD-DEX) 40 mg daily for 4 days every 4 weeks and prednisone 1.0 mg/kg daily, which was then tapered. The enrolled patients were classified into two groups according to their response to glucocorticoid treatment: glucocorticoid response group (120 cases) and nonresponse group (63 cases). The response group included patients with complete response, which was defined as PLT higher than 100 × 10μL using DNase-free water. The compared pools consisted of 200 cases and 200 control subjects, respectively. DNA (5 μL) of the case group was added to the case pool in equivalent molar amounts, and the same operation was applied for the control group. The concentration and purity of each pool were measured again. At last, eligible pools were processed for labeling and hybridization on IlluminaHumanOmniZhongHua-8 arrays according to the manufacturer's instructions .Genomic DNA was extracted from peripheral blood leukocytes from 200 cases and 200 controls using the Qiagen DNA Isolation kit  according to the manufacturer's protocol. Samples of intact genomic DNA showing no evidence of contamination by RNA and DNA degradation were selected for further analysis. DNA concentration and purity were calculated with a Nano Drop ND-2000 spectrophotometer . For SNP-MaP scanning, the DNA concentration of each sample was quantified and adjusted to 5 ± 0.5 ng/X, Y, and mitochondria), so the loci on the extraordinary chromosomes are not considered. Differences in allelic frequency between the sample pools were evaluated by combined Z-test, as follows:The hybridization intensities of two probes for each SNP allele were derived from the raw scanning files. The frequencies of autosomal SNPs were averaged over three replicate case and control arrays. Before analyzing the chip data, the quality control should be conducted, as mentioned in the following points: (1) due to the phenomenon of hybridization failure of a small number of sites will occur in the process of gene chip hybridization, the failed sites should be deleted; (2) considering that too few SNP detection experiments will lead to unreliable experimental results, it is necessary to remove the sites with less than 3 repetitions; and (3) the DNA pooling method mixes the whole genomic DNA of male and female patients, resulting in uneven loci on the extraordinary chromosomes  in cases and controls accounting for sampling variance:Chi-square statistic, f = Graw/(Graw + Rraw) and represents the approximation of allele A frequencies for each replicate, averaged over the number of replicates in each pool . nk pool replicates, Nk represents the number of controls and cases . Z-statistics for testing the differences in mean allele frequencies between cases and controls:(b)The statistic combinesrelative allele frequency (RAF) of these two DNA pools was calculated using a method described previously [P value corresponding to the odds ratio (OR) scores [Then, the eviously . For loc) scores .https://fuma. ctglab.nl/) [The other SNP in linkage disequilibrium (LD) to the leading SNPs was analyzed using the functional mapping and annotation of genome-wide association studies (FUMA GWAS) tool (lab.nl/) .μL using DNase-free water. Genotyping was performed using TaqMan SNP Genotyping Assays , TaqMan Genotyping Master Mix , and an Applied Biosystems ViiA™ 7 Real-Time PCR System  in a 96-well format. The selected SNPs were genotyped with TaqMan® SNP Genotyping Assays: AHCTBKM for rs5998634, AHQJQ0L for rs17503120, and C_32336830_10 for rs16866133, except for rs4483616 due to the failure probe synthesis. Each reaction (10 μL) contained 5.0 μL TaqMan Genotyping master mix, 0.25 μL primers and probes, 3.75 μL DNase-free water, and 1.0 μL DNA (20 ng/μL). Thermal cycling conditions were 95°C for 10 min, followed by 40 cycles of 95°C for 15 seconds and 60°C for 1 min.The concentration of DNA samples from 250 cases and 250 controls was adjusted to 20 ng/http://www.ncbi.nim.nih.gov/entrez/). Using the information of chromosome and position, we located SNPs to the genes within a window 20 Kbp upstream and downstream. The annotations for human genome assembly version 37 , which was downloaded from the UCSC genome annotation databases (http://hgdownload.cse.ucsc.edu/), was used to map SNPs to genes. In our study, if multiple SNPs were mapped to the same gene, only the gene with the lowest P value was selected for further analysis. If no gene was found in a +/-20Kbp window of the SNP, the nearest gene on each side of the SNP was included.To scan for the genetic factors related to ITP, all identified SNPs were mapped to genes utilizing the EntrezGene database  to conduct functional enrichment analysis for genes in our study. All identified overrepresented pathways in our study derived from the KEGG pathway database (http://www.genome.ad.jp/kegg), which integrates genomic, chemical, and systemic functional information [P < 1.0 × 10−7, it was difficult to conduct a pathway analysis with such a few numbers of genes. Thus, we selected 287 genes by selecting P value less than 10−5 as a gene set to analyze. The term “hsapiens” was selected as the organism and “hsapiens_gene_symbol” as the gene ID type when uploading both the interesting gene list and the reference gene set. Then, “hsapiens_genome” was selected as the reference set, and P < 0.05 was considered as significant when the hypergeometric method was used for statistical analysis.To provide further insight into the molecular function of identified associated variants, we utilized the WebGestalt . Chi-square tests were used to detect whether the genotype distributions for the studied SNPs fit Hardy-Weinberg equilibrium (HWE), and P > 0.05 was considered to be consistent with the HWE equilibrium. Finally, 2 × 3 contingency tables were applied to compare genotype frequencies between cases and controls, and P < 0.05 was considered statistically significant.In the clinical characteristics analysis, the continuous data were presented as mean ± standard deviation (SD), and the differences between cases and controls were evaluated using Student P < 0.001); however, WBC in ITP patients was significantly higher than in controls (P < 0.001). The detailed clinical information of all the subjects is shown in All of the recruited ITP patients demonstrated similar typical symptoms and signs. Two-hundred cases  and 200 controls were recruited in the first stage. The mean age at the time of ITP onset was 40.36 ± 16.39 years, and the ratio of males and females was 1 : 2.1. A replicate cohort containing 250 cases  and 250 unrelated controls was collected for the TaqMan probe genotyping assay. The mean age at the time of ITP onset was 40.85 ± 12.21 years, and the ratio of male to female was 1 : 2.04. Our data displayed a significant gender disparity. In both cohorts, PLT, RBC, and HB were significantly lower in the ITP group than in the control group (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE76744). The averaged relative allele frequencies (RAF) of cases and controls presented a very high Pearson correlation  located within or near GBE1/LINC02027, TENM4, SYN3/TIMP3, and RBM45/OSBPL6 genes reached a statistical significance in association with ITP in the genome-wide scale  to the top four loci were shown in Supplementary For SNP-Map scanning, 862,620 autosomal SNPs were screened by the IlluminaHumanOmniZhongHua-8 arrays. The raw data were deposited in the Gene Expression Omnibus as data set GSE76744 . Both the distributions of rs117503120 G/A and rs16866133 T/G in ITP patients were significantly lower than in controls .To confirm the association between significant SNPs and ITP, we performed a TaqMan PCR genotyping assay. Genotypic frequencies in patients and controls were in the Hardy-Weinberg equilibrium  (P < 0.05).We compared the genotype and allele frequencies of rs117503120, rs5998634, and rs16866133 between the response and nonresponse group for glucocorticoid therapy. The results showed that only the rs5998634 minor allele T was significantly associated with a favorable response to glucocorticoid treatment among ITP patients ( = 0.03) . In termP = 0.0054), neuroactive ligand-receptor interaction (Adj − P = 0.0054), and pathways in cancer (Adj − P = 0.0099) , TIMP3/SYN3 (rs5998634), TENM4 (rs4483616), and RBM45/OSBPL6 (rs16866133) were strongly associated with ITP from CHP.The genetic factor plays a nonnegligible role in modulating the course of ITP. Due to the low incidence of ITP, genetic analysis with large sample sizes can be challenging. Traditional candidate gene approaches have identified the relationship of genes such as IL-10, IL-3, and IFN-me scale , 16. To GBE1/LINC02027 gene at chromosome 3q12.3. The glucan  branching enzyme 1 (GBE1) was reported strongly associated with glycogen storage disease in previous research [GBE1 was significantly associated with ITP (P = 6.45 × 10−9), and the TaqMan probe genotyping results also showed that the MAF of rs117503120 in ITP cases was significantly lower than controls, suggesting that the rs117503120 minor allele may be protective for ITP. Thus, GBE1 may have a potential vital association with ITP from CHP, although the mechanism is yet to be investigated. In addition, the LINC02027 (long intergenic nonprotein coding RNA 2027) has been reported to be highly expressed in liver and kidney tissues from the 95 human individuals [Rs117503120 is located on the ividuals , but theTIMP3/SYN3 at chromosome 22q12.3, which was strongly associated with ITP in our study (P = 8.06 × 10−8). The tissue inhibitor of metalloproteinase-3 (TIMP3) is an inhibiting matrix metalloproteinase protein [γ in ITP patients, suggesting that this protein may lead to Th1/Th2 polarization via affected antigen-presenting cells and contribute to the occurrence and development of autoimmune disease [χ2 = 12.5, P = 0.0019). The MAF of rs5998634 CT genotype in ITP patients (0.144) was significantly higher than in controls (0.045), suggesting that the T allele is a major genetic risk factor to ITP from CHP. Furthermore, we also evaluated the change difference of platelet count from the 183 ITP patients treated with glucocorticoid based on the different genotypes of rs5998634. Interestingly, the mean platelet count of 35 ITP patients with the rs5998634 CT or TT genotype was significantly higher than in 148 patients with the CC genotype after the fifth day of glucocorticoid treatment (P < 0.05). These results shown that the patients with ITP who carry the rs5998634 T allele may be more sensitive to glucocorticoids than patients with the C allele. The limited sample size and lack of replication resulted in low statistical efficiency, and follow-up studies with large sample sizes need to be carried out. Synapsin III (SYN3) mainly involved in the development of brain or neurons disease, such as Parkinson disease [Rs5998634 is located on  protein . Researc disease . Importa disease , but theRBM45/OSBPL6 at chromosome 2q31.2 and also had a strong association with ITP in our study (P = 9.39 × 10−8). The association of RNA-binding motif protein 45 (RBM45) with ITP or other autoimmune diseases is unknown. The oxysterol-binding protein-like 6 (OSBPL6) gene encodes the oxysterol-binding protein-like 6 receptor, which associated with multiple sclerosis (P = 4.64 × 10−4) in the United Kingdom (UK) population [OSBPL6 may participate in the pathogenesis of ITP in terms of immune regulation. Also, the TaqMan probe genotyping results showed a significantly lower MAF of rs16866133 TG genotype in ITP patients compared with the controls (0.005 vs 0.060), suggesting that rs16866133 G was a protective allele for ITP.Rs16866133 is located on pulation . It is nTo provide further insight into the molecular function of identified associated variants, the pathway analysis converged GWAS datasets was conducted. In this study, the JAK-STAT signaling pathway, neuroactive ligand-receptor interaction, and pathways in cancer were proposed to be the most potentially associated with ITP from CHP. Among them, the JAK-STAT signaling pathway has been previously reported to participate in the etiological mechanism of pediatric ITP using the gene expression profile analysis methods . It also


Chronic obstructive pulmonary disease (COPD) is the third leading cause of death in the United States with no effective treatment. The current diagnostic method, spirometry, does not accurately reflect the severity of COPD disease status. Therefore, there is a pressing unmet medical need to develop noninvasive methods and reliable biomarkers to detect early stages of COPD. Lipids are the fundamental components of cell membranes, and dysregulation of lipids was proven to be associated with COPD. Lipidomics is a comprehensive approach to all the pathways and networks of cellular lipids in biological systems. It is widely used for disease diagnosis, biomarker identification, and pathology disorders detection relating to lipid metabolism.In the current study, a total of 25 serum samples were collected from 5 normal control subjects and 20 patients with different stages of COPD according to the global initiative for chronic obstructive lung disease (GOLD) . After metabolite extraction, lipidomic analysis was performed using electrospray ionization mass spectrometry (ESI-MS) to detect the serum lipid species. Later, the comparisons of individual lipids were performed between controls and patients with COPD. Orthogonal projections to latent structures discriminant analysis (OPLS-DA) and receiver operating characteristic (ROC) analysis were utilized to test the potential biomarkers. Finally, correlations between the validated lipidomic biomarkers and disease stages, age, FEV1% pack years and BMI were evaluated.p < 0.05) with a two-fold change. In addition, lipid ratios between every two lipid species were also evaluated as potential biomarkers. Further multivariate data analysis and receiver operating characteristic (ROC: 0.83 ~ 0.99) analysis suggest that four lipid species (AUC:0.86 ~ 0.95) and ten lipid ratios could be potential biomarkers for COPD (AUC:0.94 ~ 1) with higher sensitivity and specificity. Further correlation analyses indicate these potential biomarkers were not affected age, BMI, stages and FEV1%, but were associated with smoking pack years.Our results indicate that a panel of 50 lipid metabolites including phospholipids, sphingolipids, glycerolipids, and cholesterol esters can be used to differentiate the presence of COPD. Among them, 10 individual lipid species showed significance (Using lipidomics and statistical methods, we identified unique lipid signatures as potential biomarkers for diagnosis of COPD. Further validation studies of these potential biomarkers with large population may elucidate their roles in the development of COPD. PG and LPG were determined by the same diacyl standards with other PC and PE species, but no response factors were reported.All scans were performed in a positive mode. Lipid species with a common head group fragment were determined by the precursor ion and neutral loss scans of the lipid extracts. Lipid species were detected with the following scans: CE, [M + NH4]Deconvolution is applied to the data for overlapping of isotopic peaks. Each spectrum was subtracted by background. Later, the signal normalized by internal standards was used for relative quantification. Since molecular species in the same class would have similar ionization efficiency, the same class of internal standards was applied to quantification analysis. Quality controls were made by pooling from all 25 samples. After the first 6 quality controls, the mass spectra of internal standard mixture only, one quality control, and normally 10 patients’ samples were acquired successively. The average molar amount of lipid species detected in “internal standards only” was used for background subtract. At last, the data were normalized by the analyzed sample amount to generate data with the concentration unit of nmol/μL.p < 0.05; 4) fold changes in concentration in serum lipid is > 2 (up or down). Later, Orthogonal Projections to Latent Structures (OPLS) and receiver operating characteristic curve (ROC) were applied to evaluate the biomarker candidates. In MetaboanalystR, the OPLS-DA model was constructed, and ROC analysis was performed based on the selected biomarker candidates [p-values were reported in the figures with values of p < 0.05 considered as statistically significant.Four criteria were applied to select lipid biomarkers candidates. Species below the detection of limit, species with excessive missing values, species with insignificant difference between patient and control groups, and species with insignificant fold change would be removed. Criteria for retention were: 1) mean of serum lipid with concentration was higher than 0.0007 nmol/μL; 2) lipid species with less than 50% missing value; 3) difference between control and patient groups was statistically significant ndidates . All thePrevious data demonstrated that patients with COPD exhibit systematic dysregulation in lipid metabolism, in which sphingolipid pathway is higher in smokers with COPD compared with smokers without COPD . Other sUsing electrospray ionization mass spectrometry (ESI-MS), 377 individual lipid species were identified using previous published method . Later, the Log2 value of fold change was used as the cut-off value to compare the lipid species from disease and from control groups (Log2Fc > 1 or < − 1). Of interest, 43 lipids were obtained to have distinct concentration values of lipid species , 7 lipid species had statistically significant difference but fold change < 2 (blue bar), and 10 lipid species met both criteria as fold change > 2 and p < 0.05 (orange bar). These results demonstrate a total of 10 lipid species have satisfied both p-value and fold change cut-off and could be selected as potential biomarker candidates.In order to quantify the individual lipid species, the concentrations of individual lipid species with more than 50% detection values were retained for further analysis . The remaining missing values were replaced by the half of the minimum positive value in original data. Log transformation was performed before statistics analysis. Fold changes and Log2Fc >  or < − 1p-value) were chosen as lipid ratio candidates for further biomarker analysis. In Table Previous data suggest that the ratios of the concentrations between two relative metabolites could convey more information compared to the two individual metabolites . In addiTo validate the diagnostic potential of those biomarker candidates, a supervised multivariate data analysis (OPLS-DA) was performed to discriminate the biomarker candidates between COPD patients and control . The disNext, we confirmed the feasibility of using lipid ratio as COPD diagnostic markers with receiver operating characteristic curve (ROC) plot analysis. The area under the ROC curve was also utilized to assess the efficacy of the selected biomarker candidates. The results of the area under the curve (AUC), the sensitivity (true-positive rate) and specificity  was calculated by MetaboAnalystR in R . AccordiFinally, we explored the ability of lipid ratio biomarkers to determine different stages of COPD. The lipid ratio PI 38:4)/C16:1 CE is shown in the spots plot in Fig. /C16:1 CE2 = 0.35), and this correlation is significantly affected by age and the presence of hypertension [Due to the prevalence of COPD, there is a demanding for reliable non-invasive biomarkers in diagnosing and treating patients. Blood biomarkers are relatively non-invasive and easy to obtain for this purpose . Among artension . Other srtension . In addirtension . Among trtension . Therefortension , 48. In In single lipids level, 10 lipid species were selected as biomarker candidates, including six species of phospholipids, two species of glycerolipids, one species of sphingolipids, and one species of sterol lipid. Among them, three phospholipids (PC(32:1), PI(36:6), PI(44:6)), one glycerolipid TAG(54:6) 22:6/16:0/16:0, one sphingolipid (SM(22:0)), and one sterol lipid (C16:1 CE) were higher in patients with COPD comparing with the control. On the other hand, one glycerolipid TAG(54:5) 18:2/18:1/18:2, and four other phospholipids (ePE(34:2), ePS(38:3), LPE(20:2), and PS(34:0)) were lower in patients with COPD.Lipid ratios have been proven to be a useful biomarker for several diseases, including cardiovascular disease, Alzheimer’s disease, and diabetes –51. In aIn the ROC analysis, the AUC of all biomarker candidates was calculated. Generally, an AUC of 0.5 suggests no diagnostic ability to distinguish patients with and without the disease, 0.7 to 0.8 is an acceptable level, 0.8 to 0.9 is an excellent level, and more than 0.9 is an outstanding level . Based oGlycerolphospholipids, which are the main lipid class identified in this study, are highly enriched within the pulmonary airway surfactant. A surfactant disorder was reported to be related to the pathogenesis of COPD . PI consHowever, PE was reported to increase in COPD patients’ serum or sputum by previous lipidomics study , 58, whiSphingolipids play an essential role in the development of COPD , 60. SmoThe high hydrophobicity of cholesterol ester (CE) leads the esterified cholesterol moving from the surface into the center within discoidal high-density lipoprotein (HDL), which will transform discoidal HDL into spherical HDL . This trFinally, correlation analysis was performed to explore the possible effects of age, BMI, stages, FEV1% and smoking pack years, on the identified biomarkers. All the correlation analysis were shown as scatter plot in supporting information .Additional file 3: Supplemental Figure 3–12. Correlations between the lipid ratios and FEV1, Smoking Pack Years, Age and BMI.Additional file 4.Additional file 5.


The aim was to evaluate the prognostic factors, clinicopathological characteristics, and surgical outcomes after hepatectomy in very elderly patients with hepatocellular carcinoma (HCC).n = 49); patients younger than 80 years old and aged 65 years or older were classified into the elderly group , and patients younger than 65 years old were classified into the young group . We investigated the prognoses, clinicopathological characteristics, and surgical outcomes after hepatectomy.We analyzed 796 patients with HCC from 2000 to 2017. Patients aged 80 years or older were classified into the very elderly group  < 80% and PIVKA-II ≥ 400 mAU/ml were unfavorable factors for survival, and PIVKA-II ≥ 400 mAU/ml and the presence of portal venous invasion (PVI), hepatic venous invasion, and fibrosis were unfavorable factors for recurrence. In group E, ChE < 180 IU/l, AFP ≥ 20 ng/ml, tumor size ≥ 10 cm, and the presence of multiple tumors, PVI, and hepatic venous invasion (HVI) were unfavorable factors for survival, and ChE < 180 IU/l, tumor size ≥ 10 cm, and the presence of multiple tumors, PVI, and HVI were unfavorable factors for recurrence. In group Y, AFP ≥ 20 ng/ml, the presence of multiple tumors, poor differentiation, PVI, HVI, and blood loss ≥ 400 ml were unfavorable factors for survival, and PA < 80%, albumin < 3.5 g/dl, AFP ≥ 20 ng/ml, tumor size ≥ 10 cm, and the presence of multiple tumors, poor differentiation, and PVI were unfavorable factors for recurrence.Tumor factors might have limited influence on the prognosis of very elderly patients, and liver function reserve might be important for the long-term survival of very elderly patients. Hepatectomy can be performed safely, even in very elderly patients. Hepatectomy should not be avoided in very elderly patients with HCC if patients have a good general status because these patients have the same prognoses as nonelderly individuals. It is estimated that 18.1 million new cancer cases and 9.6 million cancer deaths occurred in 2018 worldwide . AccordiThe size of the aging population is increasing worldwide. According to a report by the Ministry of Health, Labour and Welfare, the number of people aged 80 years or older was 10.35 million in 2016, which represented 8.3% of the entire Japanese population at the time [In general, treatment options for HCC include surgical resection; ablation, such as radiofrequency ablation; transcatheter arterial chemoembolization (TACE); hepatic arterial infusion (HAI); liver transplantation; and molecular targeting drugs, such as sorafenib, lenvatinib, regorafenib, and ramucirumab. Furthermore, immunotherapy, i.e., dendritic cell vaccination, may play a critical role as an alternative option in the treatment of advanced HCC patients in whom these traditional therapeutic modalities cannot be applied , 8. HoweIn this study, we evaluated the prognostic factors and clinicopathological characteristics of patients with HCC aged 80 years or older and compared these patients to those younger than 80 years old after hepatectomy. We also investigated surgical outcomes after hepatectomy.n = 49, 6.2%), patients younger than 80 years old and aged 65 years or older into the elderly group , and patients younger than 65 years old into the young group . We compared the prognoses in terms of survival and recurrence, clinicopathological characteristics, and surgical outcomes after hepatectomy between these groups. We defined HBs-Ag positive as HBV and HCV-Ab positive as HCV.Between January 2000 and December 2017, 796 consecutive patients with HCC underwent primary liver resection at the Gastroenterological Surgery I Unit of the Hokkaido University Hospital in Sapporo, Japan. We divided the patients into three groups: patients aged 80 years or older were classified into the very elderly group  and was performed in accordance with the Helsinki Declaration guidelines. Informed consent was obtained in the opt-out form on the website of Hokkaido University Hospital.The indications for hepatic resection were as follows: patients with a performance status score between 0 and 2, patients with an American Society of Anesthesiologists (ASA) grade between 1 and 3, patients who were not senile, and patients whose comorbidities were controlled. Patients with or suspected to have ischemic heart disease or cardiac failure were assessed by cardiologists. The type of surgical procedure was usually determined based on the patients’ liver function reserve, i.e., according to the results of the indocyanine green retention test at 15 min (ICGR15) . AnatomiPostoperative morbidity was assessed using the validated Clavien–Dindo classification system . SeriousPatients were followed up at 3-month intervals. Patients underwent physical examination and serological examination, including alpha-fetoprotein (AFP) level, protein induced by vitamin K absence-II (PIVKA-II), and liver function. In addition, radiological examinations, including contrast-enhanced computed tomography (CT) scans and/or ultrasound sonography (US) or contrast-enhanced magnetic resonance imaging (MRI), were performed. Follow-up using these modalities after curative treatment at 3- or 4-month intervals was recommended by the Clinical Practice Guidelines for Hepatocellular Carcinoma 2017 .U test for continuous variables or chi-square test for noncontinuous variables. Kaplan-Meier survival curves were compared by using the log-rank test. Overall survival (OS) and relapse-free survival (RFS) were evaluated. Prognostic factors were evaluated by these univariate analyses and multivariate analyses using the Cox proportional hazard model. We used JMP Pro 14.0.0 for Windows  for statistical analyses. A p value of < 0.05 was considered significant.Differences in characteristic factors were evaluated by the Mann-Whitney We divided the period between 2000 and 2017 and designated the period from 2000 to 2008 as the early period and the period from 2009 to 2017 as the late period. In the early period, group VE included 8 patients . In contrast, this group had 41 patients  in the late period.In this cohort, the median survival time (MST) and 5-year OS rate in our 796 study patients were 103 months and 63%, respectively. The median RFS time was 20 months. The median length of hospital stay was 24 (9–386) days. The number of cases with HBV and HCV was 277 and 217, respectively. There were 15 patients with both HBV and HCV. There were 287 cases of NBNC (NonBNonC)-HCC. The median follow-up period of the whole cohort was 39 months. These were 24 months, 38 months, and 45 months (VE vs. E vs. Y), respectively.Univariate analysis showed that the proportion of patients who were HCV positive, without HBV and HCV (NBNC), was significantly increased in group VE compared with group Y Table . In contTable Regarding recurrence, 24 patients experienced recurrence (49%) with a median recurrence time of 11 months 3–68) in group VE. In group E, 231 patients experienced recurrence (64%) with a median recurrence time of 11 months (0.4–111). In group Y, 261 patients experienced recurrence (68%) with a median recurrence time of 9 months (0.2–197). The initial recurrence sites were not significantly different among the three groups  patients died; 237 (82%) were cancer-related deaths. Five (1.7%) patients with controlled HCC experienced liver failure-related deaths. In total, 1 (0.3%) death was related to the surgical procedure (posthepatectomy liver failure); 46 (16%) deaths were classified as other disease-related deaths.p = 0.41, 0.01); liver failure not related to cancer, 0 (0%) vs. 3 (2.5%) vs. 2 (1%) ; and others, 4 (36%) vs. 27 (22.5%) vs. 15 (10%) . Group VE showed significantly fewer cancer-related deaths and more non-cancer-related deaths than group Y. No significant differences in the cause of death were noted between groups VE and E.There were 11 (22%) deaths in group VE, 120 (33%) deaths in group E, and 157 (41%) deaths in group Y. The cause of death and the breakdown among the three groups were as follows (group VE vs. group E vs. group Y): cancer-related, 7 (64%) vs. 90 (75%) vs. 140 (89%) (p = 0.86). The median RFS times of groups VE, E, and Y were 22 months, 21 months, and 17 months, respectively (p = 0.65). Both the OS and RFS rates were not significantly different among the three groups . On the other hand, there was no significant difference in recurrence between patients with an FRLR ≥ 50% and FRLR < 50%  in group VE. Patients with an FRLR ≥ 50% showed significantly more favorable survival than patients with an FRLR < 50%  and that there were an increased proportion of patients with HCV and patients without HBV and HCV.The incidence of PVI and fibrosis was lower in group VE compared with group Y. The number of surgical procedures was not significantly different between groups. Group VE showed significantly fewer cancer-related deaths and more non-cancer-related deaths than group Y. The surgical outcomes and morbidities of group VE were almost the same as those of the other two groups. These three groups showed similar OS and RFS results. Regarding prognostic factors, tumor factors, such as tumor size and tumor number, less influenced the prognosis of patients in group VE compared with patients in groups E and Y.According to the nationwide survey of HCC patients in Japan, the rate of nonviral HCC was 32.5% in 2015 . In thisLiver function reserve might be more important in group VE compared with groups E or Y. Interestingly, patients with an FRLR ≥ 50% exhibited significantly more favorable survival than patients with an FRLR < 50% in group VE according to the univariate analysis. Furthermore, no significant differences in recurrence were noted. During hepatectomy for very elderly patients, surgeons might have to make a maximum effort to preserve the remnant liver as much as possible.The prognoses of very elderly patients with HCC are under investigation. Many reports have claimed that the prognoses after hepatectomy do not differ between elderly patients and nonelderly patients , 27, 33.Hepatic resection is the main therapeutic method for HCC even in elderly patients. However, hepatectomy is a highly invasive surgical procedure with a high morbidity rate . TherefoRegarding treatment after recurrence, rehepatectomy was not performed in group VE in this cohort. This strategy was attributed to conservative patient selection. However, laparoscopic surgery might be a useful tool for rehepatectomy in very elderly patients.n = 49). Second, elderly patients had a possibility of selection bias when they were referred from internal medicine.The limitations of the study are as follows. First, the number of patients aged 80 years or older was small (Tumor factors may less influence the prognoses of very elderly patients with HCC compared with patients younger than 80 years old, and liver function reserve might be important for the long-term survival of these elderly patients. Hepatectomy can be safely performed, even in very elderly patients, using a close evaluation. Hepatectomy should not be avoided in very elderly patients with HCC if patients have a good general status because these elderly patients with HCC have the same prognoses as nonelderly individuals with HCC.


Two times higher blood viscosity was observed in large aneurysms compared with small aneurysms. Lower WSS was found in large aneurysms (1.38 ± 1.36 Pa) than in small aneurysms (3.53 ± 1.22 Pa). All the differences in hemodynamic factors between small and large aneurysms were statistically significant. Conclusions: Large aneurysms tended to have complex flow patterns, low shear rate, high blood viscosity, and low WSS. The hemodynamic factors that we analyzed might be useful for decision making before surgical treatment of aneurysms.Purpose: To analyze size-dependent hemodynamic factors  in unruptured cerebral aneurysms using computational fluid dynamics (CFD) based on the measured non-Newtonian model of viscosity. Methods: Twenty-one patients with unruptured aneurysms formed the study cohort. Patient-specific geometric models were reconstructed for CFD analyses. Aneurysms were divided into small and large groups based on a cutoff size of 5 mm. For comparison between small and large aneurysms, 5 morphologic variables were measured. Patient-specific non-Newtonian blood viscosity was applied for more detailed CFD simulation. Quantitative and qualitative analyses of velocity, shear rate, blood viscosity, and WSS were conducted to compare small and large aneurysms. Results: Complex flow patterns were found in large aneurysms. Large aneurysms had a significantly lower shear rate (235 ± 341 s Detection of asymptomatic unruptured aneurysms has become feasible with the improvements in neuroimaging methods . After dThe risk of aneurysm rupture can also be predicted using hemodynamic factors . Since aOne of the important hemodynamic factors, shear rate, is defined as the rate of change of velocity per distance. Previous studies have reported that the shear rate is dependent upon the geometry and size of the aneurysm. In the dome of the aneurysm, an almost zero shear rate can be observed, but regions of high shear rates also exist . TherefoThus, the purpose of this study was to conduct CFD simulations using patient-specific measurements of non-Newtonian viscosity to accurately assess size-dependent hemodynamic factors within the dome of an unruptured aneurysm. Morphological variables were measured. Quantitative and qualitative comparison of the shear rate, blood viscosity, and wall shear stress (WSS) was carried out between small and large aneurysms.This study was approved by our Institutional Review Board, which waived informed consent. Twenty-one patients (8 men and 13 women) with unruptured cerebral aneurysms were enrolled to study size-dependent distributions of hemodynamic factors. All patients were diagnosed with three-dimensional rotational angiography (3DRA). They underwent venous sampling before infusion with physiologic (0.9%) saline when they visited our hospital.The common locations of cerebral aneurysms in the anterior circulation were identified: 5 aneurysms at an anterior communicating artery, 10 aneurysms at a posterior communicating artery, and 6 aneurysms at a middle cerebral artery. There are various cutoff values  for small and large aneurysms, and we used the cutoff value of 5 mm for this study. Patients with an aneurysm size ≤5 mm and >5 mm were divided into “small” and “large” groups, respectively [All source images were obtained using 3DRA  at total 126 frame during 8.34 s as 1.5° rotation interval and 1024 × 1024 matrix. The obtained 3DRA data was saved in Digital Imaging and Communications in Medicine (DICOM) format for each patient. Using Materialise Mimics v20.0 , the obtained data in 2D DICOM format were converted into a patient-specific 3D model in STL format. By controlling and regulating the threshold method, cropping operation, and edit mask in 3D tools, a 3D model was reconstructed for CFD analyses. Unnecessary branches in the 3D aneurysm model were removed before morphology measurements.The height, width, and ostium diameter were measured for each patient. The detailed information for illustrating the measurement method is shown in −1 to 1000 s−1 were obtained automatically as shown in After venous sampling of all patients, whole blood viscosity was measured using a scanning capillary tube viscometer . Values for whole blood viscosity at shear rate range from 1 su. In Equation (2), p and The reconstructed 3D model for each patient was imported into COMSOL Multiphysics v5.2a  for CFD analyses. Fully-coupled method and iterative solver ) were applied for numerical calculation. We set 0.01 as relative tolerance and 50 as maximum number of iteration. The continuity equation and Navier–Stokes equation were used to carry out simulations of blood flow (Equations (1) and (2), respectively). In both equations, flow velocity was represented as ectively ,18:(1)∇·Blood flow was considered to be a laminar and incompressible non-Newtonian fluid . For bloIn the Casson model (Equation (3)), ear rate . The walear rate ,21. Usinear rate ,23. Tracear rate . The CFDear rate .WSS is the wall shear stress, and iWSS is the instantaneous wall shear stress, and T denotes the duration of one cardiac cycle [Based on CFD simulations, the following calculated hemodynamic factors were obtained: average shear rate; average blood viscosity; minimal, time-averaged, and maximal WSS. The shear rate is defined as the rate of change of velocity per distance, and this term is crucial for assessment of shear stress . The sheWSS=1T∫0TFlow patterns between small and large aneurysms were analyzed. The flow pattern can be determined by flow complexity and inflow jets. Flow complexity is divided into “simple” and “complex” depending on the number of flow recirculation areas. Simple flow has 1 recirculation area, and complex flow has 2 or more recirculation areas . The inft-test  or Mann–Whitney rank sum test  were used. p < 0.05 was considered as statistically significant.All the obtained data for small and large aneurysms were expressed with mean and standard deviation value (mean ± SD). Statistical analyses were undertaken using PASW v18 . To assess significant differences between small and large aneurysm groups, the independent sample The average height (aneurysm size), width, ostium diameter, cross-sectional area of the ostium, and surface area of small and large aneurysms are shown in The distributions of blood flow velocity are shown in Blood flow patterns (flow complexity and inflow jet) were compared between small and large aneurysms, and the recirculation zone was marked by arrows. In small aneurysms, mostly simple flow and diffused inflow were observed. For example, in case B, one recirculation zone was found, and blood flowed along with the wall of the aneurysm dome. In large aneurysms, complex blood flow and diffused inflow were observed. For example, in case D, 2 recirculation zones were observed in the aneurysm dome with diffused inflow.−1). Regardless of aneurysm size, a higher shear rate was found in the parent artery. Due to an increase in aneurysm area, a reduced range of shear rate appeared in the aneurysm dome (1–200 s−1). In small aneurysms, a low shear rate was observed at the aneurysm tip, whereas an enlarged region of decreased shear rate was found in the dome of large aneurysms at peak-systole. At end-diastole, a region of higher shear rate was observed around the aneurysm neck for all small and large aneurysms. Compared with the shear rate at peak-systole, a lower shear rate was observed in the parent artery at end-diastole. In large aneurysms, most of the dome showed a lower shear rate (1–100 s−1).Distributions of shear rate at peak-systole and end-diastole are shown in −1, whereas a lower average shear rate value at peak-systole was observed in large aneurysms (235 ± 341 s−1), and this difference was significant (p = 0.008). Compared with the average shear rate of large aneurysms at end-diastole (112 ± 123 s−1), 3 times higher average shear rate of small aneurysms was found (339 ± 258 s−1), and this difference was significant (p = 0.011).The average shear rate at peak-systole and end-diastole was calculated in small and large aneurysms . Overall−1)  . In smalp = 0.001) and end-diastole (p = 0.006).Comparison of blood viscosity between small and large aneurysms is shown in −1. The minimum and maximum values of WSS are represented by arrows in Distributions of WSS varied depending on aneurysm size . The scap = 0.011). The mean time-averaged WSS on the aneurysm was 3.53 ± 1.22 Pa for small aneurysms and 1.38 ± 1.36 Pa for large aneurysms, and this difference was significant (p = 0.004). In addition, the average maximal WSS was 9.90 ± 3.92 Pa for small aneurysms and 2.90 ± 3.21 Pa for large aneurysms, which was 3.4 times greater, and the difference was significant (p = 0.004).The average minimal, time-averaged, and maximal WSS were calculated at the surface of the aneurysm dome. Overall, small aneurysms tended to have higher WSS than large aneurysms . The aveIn this study, measured non-Newtonian viscosity was used for a patient-specific CFD study to analyze hemodynamic factors such as velocity, shear rate, blood viscosity, and WSS according to aneurysm size. Based on size-dependent CFD analysis of aneurysms, we were able to identify the different hemodynamic characteristics of small and large aneurysms. Large aneurysms tended to have significantly lower velocity, shear rate, and WSS than those of small aneurysms. In addition, large aneurysms had significantly greater area with increased blood viscosity compared with those of small aneurysms.Cebral et al.  reportedIn the present study, 21 patients with unruptured aneurysms were assessed. A diffused inflow jet was observed in all unruptured cases, but flow complexity was different according to size . As the CFD studies have been undertaken based on the assumption of Newtonian blood flow ,10,28, b−1; In the present study, hemodynamic factors were analyzed with measured non-Newtonian model for more precise results. As a result, a zone of decreased shear rate was captured at the aneurysm dome  by inaccurate thresholding or removal of unnecessary branches of aneurysms. Therefore, we attempted to obtain a source image of the aneurysm of optimal quality using 3DRA (not using a 2D image obtained through computed tomography or magnetic resonance imaging) to minimize these potential errors.The final limitation was that we applied commonly used boundary conditions due to the difficulty of measuring patient-specific data. The inlet boundary condition was calculated based on the flow rate in the literature. We also used the wall properties of “rigid” and “no-slip” condition. For more accurate analyses, measurement of patient-specific blood viscosity was undertaken instead of applying a Newtonian model for blood. In future studies, we will attempt to measure the inlet velocity using phase-contrast magnetic resonance imaging.By applying measured patient-specific non-Newtonian viscosity to CFD analysis, hemodynamic factors according to aneurysm size could be thoroughly compared. Large aneurysms had lower velocity, shear rate, and WSS in the aneurysm dome. Diffused inflow jets were found in small and large aneurysms. Small aneurysms had simple flow patterns, whereas large aneurysms had complex flow patterns. Blood viscosity was higher in the domes of large aneurysms. All analyzed hemodynamic factors showed statistically significant differences between small and large aneurysms. The hemodynamic factors that we analyzed could help to predict the rupture risk of aneurysms and aid decision-making before surgical planning.


Evidence suggests that body composition has impact on arterial stiffness. However, evidence in Chinese are limited, and results remain controversial. The aim of our study is to investigate whether skeletal muscle mass is associated with arterial stiffness in Chinese community-dwelling men and women aged 45 years and older.In this cross-sectional study, 20,477 participants  were included in the analysis. Brachial-ankle pulse wave velocity (baPWV), an indicator of arterial stiffness was measured using a waveform device. Total muscle mass and muscle mass of arm, leg and trunk were measured by bioelectrical impedance analysis. Height and weight were measured and appendicular skeletal muscle mass index (ASMI) was calculated as appendicular skeletal muscle mass (sum of arm and leg muscle mass) divided by height square.p < 0.0001; for women: − 0.245 (0.012), p < 0.0001]. High ASMI was a protective factor for the presence of arterial stiffness (defined as baPWV) . Similar associations were found between quantity of muscle mass  and arterial stiffness in men and women after further adjustment for height .After adjustment for age, body fat percentage, systolic blood pressure and diastolic blood pressure, ASMI was negatively associated with baPWV [β (SE) for men: − 0.208 (0.016), Low skeletal muscle mass is associated with increased risk of arterial stiffness in Chinese community-dwelling adults aged 45 years and older. Cardiovascular disease (CVD) remains to be the top cause of death in China and brings out huge medical and financial burden . ArteriaEvidence suggests that body composition has impact on the prevalence and development of arterial stiffness –14. MorePossible pathways for the association between low muscle mass and arterial stiffness might lie in insulin resistance and chronic inflammation. Loss of muscle mass was considered as a cause of insulin resistance by impairing glucose metabolism , and eviThe aim of this study was to explore the associations between muscle mass indices  and arterial stiffness (defined as baPWV) in community-dwelling Chinese men and women aged 45 years and older.Data of the participants were from the Screening of Chronic Diseases in Chinese Adults (SCDCA) study, a cross-sectional study started in 2017, which aims to screen four chronic diseases  and explore the influencing factors among Chinese community-dwelling adults in urban areas from 24 administrative regions. According to the population size and cooperation, we included several cities/areas in each administrative region. In each city/town, the communities for screening were enrolled using cluster random sampling. The research staff were divided into five groups, each group was equipped with mobile health check-up vehicle, a specially designed truck carried research instruments to designated sampling cities. Eligible participants in this study were men and women: 1) aged above 18 years old, 2) who have resided locally for more than 5 years, 3) who were cooperative and volunteered. The following participants were excluded: 1) pregnant and lactating women, 2) participants with major organ diseases, including heart, liver or kidney disease, 3) participants with mental diseases, 4) unable to carry out body examination due to disabilities. In the present analysis, participants from March 2017 to May 2018 were enrolled and the following were further excluded: 1) aged under 45 years, 1) participants with peripheral artery disease whose ankle-brachial index (ABI) ≤ 0.9 , 2) part2).Height and weight were measured by trained medical workers according to standard procedures, with subjects wearing light indoor clothing and barefoot. Height was measured to the nearest 0.1 cm and weight was measured to the nearest 0.1 kg. BMI (body mass index) was calculated as weight (kg) divided by height squared (m2) was calculated as ASM divided by height square.The quantity of total skeletal muscle mass (SMM), muscle mass of the left arm (LAMM), right arm (RAMM), left leg (LLMM), right leg (RLMM) and trunk (TMM) and the percentage of body fat (BFP) were measured using a dual bioelectrical impedance analyzer . Bioelectrical impedance analysis (BIA) is widely used in the detection of body composition and this instrument were used in several studies , 27. AppIn this study, the baPWV, mean arterial pressure (MAP), pulse pressure (PP), upstroke time (UT) as well as the brachial and ankle blood pressures on the left and right sides of the subjects were measured with a plethysmography apparatus  after rested in a supine position for 5 min. Evidence showed that the validity and reproducibility of this instrument is considerably high and suitable for screening vascular damages in a large population . baPWV, A standard questionnaire was used to collect each participant’s age, gender, education level, medical history, and habits of smoking, drinking and exercise. Participants filled the questionnaire on a computer/smartphone under the supervision of trained research staff.2, 8.53 ± 0.22 kg/m2, 9.78 ± 0.73 kg/m2; women: 6.77 ± 0.34 kg/m2, 7.51 ± 0.18 kg/m2, 8.41 ± 0.57 kg/m2). Differences between groups were tested using Student’s t-test or one-way ANOVA analysis for normally distributed continuous variables and Kruskal–Wallis test for continuous variables with non-normal distribution. Pearson’s correlation analysis was used to access the correlation between muscle mass indices  and baPWV. The association of muscle mass indices with baPWV and the association of mass indices with arterial stiffness  were carried out by multivariate linear regression analysis and multiple logistic regression analysis, respectively. The regression models testing ASMI with baPWV / arterial stiffness were performed after adjusting for age, body fat, SBP and DBP. The models testing quantity of muscle mass  with baPWV / arterial stiffness were further adjusted for body height. All analyses were conducted using SAS version 9.1 . A p value < 0.05 was considered statistically significant.Categorical variables were described as number and percentage. Continuous variables with normal distribution were expressed using mean with standard deviation (SD), and non-normally distributed continuous variables were presented as median with 25th and 75th percentile. The participants were stratified into three groups by tertiles of the ASMI (men: 7.71 ± 0.37 kg/mp < 0.05) , 68.79% were women. Compared with women, men were older and had higher BMI, SMM, AMM, LAMM, RAMM, LLMM, RLMM, TMM, baPWV, SBP, DBP and ABI, however, their BFP, PP, MAP and UT were lower than that of women. Compared with women, the prevalence of diabetes and hypertension, the percentage of high education level, current smoker, current alcohol user and regular exercise were higher in men . Both among men and women, higher ASMI was observed in participants who were younger and those with lower baPWV and MAP . In contrast, participants with higher ASMI had higher BMI, ASM, SMM, LAMM, RAMM, LLMM, RLMM, TMM, SBP, DBP, PP, ABI and UT . The highest BFP was found in median ASMI group (T2) for men and in high ASMI group (T3) for women.Characteristics of body composition and artery stiffness in men and women are presented in Table p < 0.0001).Pearson’s correlations between muscle mass indices and baPWV are shown in Table p < 0.0001; women: − 0.245 (0.012), p < 0.0001] adjusted for age, body fat percentage, systolic blood pressure and diastolic blood pressure. Similar associations were found between quantity of muscle mass  and baPWV in men and women after further adjustment for height  (Table The results of the multiple linear regression showed that ASMI was negatively associated with baPWV . Similar associations were found between quantity of muscle mass  and arterial stiffness in men and women after further adjustment for height   for men: 0.730 , %CI for mThe present study found that ASMI and quantity of muscle mass  were negatively associated with baPWV. High levels of these muscle mass indices were protective factors for the presence of arterial stiffness in Chinese community-dwelling men and women aged 45 years and older.Recently, epidemiological data showed that the middle-aged population in North China had a high prevalence of arterial stiffness (high baPWV) . CompareThe negative associations between SMM / ASM / ASMI and baPWV were found in previous studies , 17, 20.Several studies further explored the impact of regional muscle mass quantity on pulse wave velocity which showed controversial results. A study among multi-race American elderly reported that high cfPWV  was associated with lower leg and arm mass in all men and lower leg mass in white women . However2, the risks of developing arterial stiffness decreased by 27.0 and 36.6% in men and women, respectively. The indices of muscle mass quantity increased by 1 SD, the risks of developing arterial stiffness decreased from 5.9% (SMM) to 52.0% (LAMM) in men and from 10.0% (SMM) to 65.2% (LAMM) in women. The effect of arm muscle mass quantity on risk of arterial stiffness was more pronounced.Although previous studies reported liner association between muscle mass indices and baPWV, to our knowledge, this was the first study performed multiple logistic regression analysis to examine the potential effect of muscle mass indices on arterial stiffness (baPWV as the dependent variable). Since arterial stiffness is a predictor for future cardiovascular disease , this anIt is well known that CVD brings out huge medical and financial burden because of its high mortality . ArteriaSeveral mechanisms may involve in the association between muscle mass and arterial stiffness. Firstly, loss of muscle mass was considered as a cause of insulin resistance by impairing glucose metabolism . EvidencThere are some limitations in the present study. First, due to the cross-sectional design of our study, we cannot make inferences about causality which warrants further prospective and interventional researches. Secondly, bioelectrical impedance analysis (BIA) but not Dual energy X-ray absorptiometry  was used to measure body composition. However, evidence have shown that body composition measured by BIA were strongly correlated with that measured by DEXA  and haveIn conclusion, our study indicated that low muscle mass was associated with increased risk of arterial stiffness in Chinese community-dwelling adults aged 45 years and older. Further longitudinal researches are needed to confirm the potential effects of muscle mass on arterial stiffness observed in this study for the prevention of cardiovascular disease.


The anticipated serum LPS dose-dependent response was not observed. Therefore, the low, mid, and high LPS groups were combined for analysis. Compared to the placebo group, endpoint serum LPS was elevated in both males (p < 0.05) and females (p < 0.05) when all LPS treatment groups were combined. However, there was no significant change in trabecular or cortical bone outcomes in the combined LPS groups compared to the placebo following the 12-week LPS intervention for either sex. This suggests that although serum LPS was elevated following the 12-week LPS intervention, the dosages administered using the osmotic pumps was not sufficient to negatively impact trabecular or cortical bone outcomes in either male or female CD-1 mice.Chronic low-grade inflammation has been identified as an underlying cause of many diseases including osteoporosis. Lipopolysaccharide (LPS) is a potent inducer of the inflammatory response that can negatively affect bone outcomes by upregulating bone resorption and inhibiting bone formation. The objective of this study was to assess the longitudinal response of trabecular and cortical bone structure and bone mineral density to LPS continuously administered for 12 weeks in male and female CD-1 mice. Mice were assigned to one of four LPS groups at 8-weeks of age: placebo (0.0 μg/d), low (0.9 μg/d), mid (3.6 μg/d) and high (14.4 μg/d) dose. Trabecular and cortical bone outcomes were measured at 8, 12, 16, and 20 weeks of age using  Inflammation is an important response to injury or infection that signals the immune system to repair damaged tissues or protect the body from foreign invaders. Whereas, chronic low-grade inflammation is often characterized by low concentrations of circulating pro-inflammatory cytokines . ChronicBacteroides, Prevotella, Escherichia). LPS can be released from these bacteria, with limited amounts passing through the gut epithelium into circulation under normal conditions. However, with increased intestinal permeability, which has been associated with obesity and even aging [Lipopolysaccharide (LPS) is a component of the outer membrane of Gram-negative bacteria such as those found in the gut microbiota , Harlan Teklad, Mississauga, ON) and water.All procedures that involved animals were in compliance with the Canadian Council on Animal Care and approved by the Brock University Animal Care Committee (AUP #17-06-01). Both male and female 7-week old CD-1 mice  were ordered from Charles Rivers (Canada) and allowed to acclimatize for 5 days in the Comparative Bioscience Facility. Females were housed 4–5 per cage, while males were housed individually. All mice were kept on a standard 12-hour light: 12-hour dark cycle and had Escherichia coli Serotype O55:B5 by phenol extraction; Sigma, St. Louis, MO) was mixed in sterile PBS at a concentration of 4 μg/μL for the high-dose (14.4 μg/d) and serially diluted to 1 μg/μL for the mid-dose (3.6 μg/d) and 0.25μg/μL for the low-dose (0.9 μg/d). These concentrations were selected based on a previous study by Droke et al. that used slow-release pellets which delivered 0, 0.133, 1.33, and 13.3 μg LPS/d [Lipopolysaccharide , with a release rate of 0.15 μL/hour designed to provide a consistent dose for 42 days (model 2006), was subcutaneously implanted in the dorsal region of the neck. This was for a total of three pump implantations over the course of the study and occurred immediately after a μCT scan while mice were still anesthetized. Metacam was subcutaneously injected to assist with any discomfort and bupivacaine was applied to the incision site as a nerve blocker. In short, an incision was made on the dorsal region of the neck, hemostat forceps were used to create a subcutaneous pocket, the pump was inserted, and the incision was closed. Initially, surgical glue was used to close the incisions but was switched to surgical staples to minimize the reopening of incisions. Mice were monitored daily to ensure proper incision healing. If an incision reopened, mice were anaesthetized, and the incision was closed with either surgical staples or sutures.The microcomputed tomography (μCT) scanning parameters and analysis have previously been published  for the in vivo scans isoflurane anesthetic was used since the flow rate could be tightly controlled and allowed mice to remain still throughout the scan, essential for obtaining a clear scan. In short, mice were anaesthetized in an induction chamber with isoflurane at a constant flow rate of 2–5% and transitioned to a nose cone housed inside the μCT for maintenance of the anesthetic. Ophthalmic gel was applied to the eyes to prevent dryness. Scans of the hindlimb were performed with the mouse positioned on its back with the scanning leg extended and non-scanning leg and tail secured away from scanning region, as previously described [Scans were completed using μCT  and host software  as previously described –28. To eescribed . Scans wescribed   and NRecon Reconstruction  were used to reconstruct cross-sectional images . The samProximal tibia trabecular bone outcomes assessed using 3D analysis included: bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), trabecular separation (Tb.Sp), degree of anisotropy (DA), structural model index (SMI), and volumetric BMD (vBMD) . Cortica®, Millipore Sigma, St. Louis, MO) was added for two minutes and then mixed with 100 μL dH2O for one minute followed by an additional 100 μL of dH2O for two minutes. Excess liquid on the slide was discarded. The blood smears were viewed on a Leica HC microscope at 20x magnification. Because the amount of blood applied to a slide varied, we averaged the number of leukocytes per field of view–for a total of 10 fields of view—instead of quantifying the entire slide. Remaining blood was centrifuged to collect serum for determination of circulating LPS concentrations. Serum LPS was analyzed using a mouse LPS ELISA kit  according to manufacturer instructions. This kit has been used to analyze serum LPS in previous studies [At the end of the study, mice were anesthetized, and blood was collected via cardiac puncture. Leukocytes were counted via manual microscopy of blood smears. Briefly, a small drop of blood was smeared across a slide and allowed to air dry. Following which 100 μL of the Wright stain . For serum LPS a one-way ANOVA was used to compare all groups. Due to the overall similarity in serum LPS among LPS treatment groups, the low, mid, and high LPS groups were combined for analysis. A two-tailed unpaired t-test with Welch’s correction was used to compare LPS treatment to the placebo group for both the serum LPS and leukocyte count. Trabecular and cortical bone outcomes were analyzed using a two-way ANOVA (LPS treatment x time) with Bonferroni correction for multiple comparisons. All statistical analysis was done using GraphPad Prism (version 8.3.0) with statistical significance set to p > 0.05; p < 0.05; p < 0.05; p < 0.01; p > 0.05; p > 0.05; Endpoint serum LPS concentrations did not differ between any of the treatment groups for the males , Tt.Ar , and TMD  and Ma.Ar . In addition, there was a significant LPS x time interaction for Ps.Pm  for 4 weeks . More spex vivo using either histomorphometric analysis, dual energy X-ray absorptiometry, or μCT. In the present study, we used in vivo analysis to track bone outcomes longitudinally to better understand how bone is altered with LPS exposure and over time. Performing multiple scans allowed us to determine that the placebo and LPS groups did not differ over the course of the 12-week LPS treatment at any of the scanning timepoints–at 8, 12, 16, or 20 weeks of age. However with time, outcomes of both trabecular and cortical bones were altered and the direction of these changes were expected based on previous studies by our lab [Altered bone formation and resorption can translate to alterations in trabecular bone structure, as demonstrated in Sprague-Dawley rats, implanted with LPS slow-release pellets. LPS caused an up-regulation of proinflammatory cytokines, IL-1 and TNF-α, resulting in a reduced BV/TV, Tb.N, and an increased Tb.Sp, which translated to impaired biomechanical strength . Similar our lab ,34 and o our lab ,36. The  our lab , these r our lab –15 and tMultiple injections to deliver LPS longitudinally are not preferable since the half-life of LPS is suggested to be around 12 hours . AlthougS1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S5 Table(DOCX)Click here for additional data file.


The availability of reliable point-of-care tests for malaria has heralded a paradigm shift in the management of febrile illnesses away from presumptive antimalarial therapy. In the absence of a definitive diagnosis, health care providers are more likely to prescribe empirical antimicrobials to those who test negative for malaria. To improve management and guide further test development, better understanding is needed of the true causative agents and their geographic variability.A systematic review of published literature was undertaken to characterise the spectrum of pathogens causing non-malaria febrile illness in Africa (1980–2015). Literature searches were conducted in English and French languages in six databases: MEDLINE, EMBASE, Global Health (CABI), WHO Global Health Library, PASCAL, and Bulletin de la Société Française de Parasitologie (BDSP). Selection criteria included reporting on an infection or infections with a confirmed diagnosis, defined as pathogens detected in or cultured from samples from normally sterile sites, or serological evidence of current or past infection. A number of published articles  reporting a given pathogen were presented.Staphylococcus aureus (18.2%), non-typhoidal Salmonella (17.3%), and Escherichia coli (15.4%) were the commonly reported bacterial infections whereas Rift Valley fever virus (7.4%), yellow fever virus (7.0%), and Ebola virus (6.7%) were the most commonly reported viral infections. Dengue virus infection, previously not thought to be widespread in Africa, was reported in 54 (5.1%) of articles.A total of 16,523 records from 48 African countries were screened, of which 1065 (6.4%) met selection criteria. Bacterial infections were reported in 564 (53.0%) records, viral infections in 374 (35.1%), parasitic infections in 47 (4.4%), fungal infections in nine (0.8%), and 71 (6.7%) publications reported more than one pathogen group. Age range of the study population was not specified in 233 (21.9%) publications. This review summarises the published reports of non-malaria pathogens that may cause febrile illness in Africa. As the threat of antimicrobial resistance looms, knowledge of the distribution of infectious agents causing fever should facilitate priority setting in the development of new diagnostic tools and improved antimicrobial stewardship.CRD42016049281PROSPERO,  The introduction of malaria antigen-based rapid diagnostic tests (RDTs) has heralded a paradigm shift in the management of febrile illnesses in malaria-endemic countries. Historically, malaria was considered by default the principal cause of fever, and presumptive antimalarial therapy was widespread policy and practice . In 2010The “test before treat” approach has demonstrated that misclassification of much febrile illness had previously led to an overestimation of malaria incidence . Since tWhere reliable diagnostics are lacking, a knowledge of pathogen distributions may inform judicious empiric prescription of antimicrobials . CurrentThe systematic review followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines , restricTitles and abstracts, as well as full texts when the abstract did not provide sufficient information, were first screened for compliance with the inclusion and exclusion criteria Table . One autData were extracted from selected articles for pre-defined variables on study design, study location, and pathogens  or serological evidence of current or past infection. Where additional assays were done , these were noted in the database but not used in case definitions due to heterogeneous reporting standards.case series, which included individual case reports or series of patients with the same infection; (ii) fever series, where a group of febrile patients was tested for a number of causative agents and where the total population tested (denominator) was reported; and (iii) seroprevalence studies, where serum samples were tested for one pathogen or a panel of pathogens simultaneously. Details regarding the timing of sample collection  were not extracted for the seroprevalence studies.Studies were categorised into (i) pracma package [maps package [Countries were classified by sub-region according to United Nations designations . To asse package  in R sof package . The coo package , and theSalmonella except for Typhi, Paratyphi A, Paratyphi B, and Paratyphi C were defined as non-typhoidal Salmonella (NTS). Details regarding the categorisation of the infections are presented in Additional file Infections were categorised as bacterial, fungal, parasitic, or viral and were sub-categorised using an epidemiological definition based on their principal mode of transmission as contact , vector-borne, air-borne, and food- and/or water-borne. Infections caused by all serotypes of Study populations were grouped into four mutually exclusive categories: neonates (aged < 28 days), infants (1–12 months), children (1–12 years), and older (≥ 13 years). If a study reported any participants from each age group, then they were grouped as participants of “all ages”.An on-line database enabled multiple users  in different locations to work on the review database simultaneously. Each study site location was geo-coded onto an on-line interactive map (surveyor), searchable by country, pathogen, year, and patient age group, and hosted by the Infectious Diseases Data Observatory (IDDO) .The unit of analysis was “published article”. Articles reporting a given pathogen were categorised by geographic region, patient age group, pathogen category, and predominant epidemiologic mode of transmission. In fever series studies that reported the number of individuals tested for a given pathogen (the denominator), the median (range) proportion testing positive was presented by article. The heterogeneity of study design, pathogens sought, laboratory methods, reporting, and limitations in data extraction precluded meta-analysis or estimation of pathogen prevalence.The currently available tools for assessing the quality and risk of bias were not applicable to our review design , 22. We The database search identified 16,523 records, of which 14,777 (89.4%) were in English and 1746 (10.6%) in French. Additional 80 records were identified from the bibliographies of other reports and through the personal knowledge of the co-authors, bringing the total number of articles identified to 16,603. Of the records resulting from the initial search, 703 (4.2%) were identified as duplicates and removed. Of the remaining 15,900 unique records, a further 9715 (61.1%) were excluded after title and abstract screening because they did not fit the selection criteria, leaving 6185 papers for full-text screening. Additional 5120 publications were excluded after full-text assessment bringing the total number of articles included in the review to 1065 , Nigeria , and Tunisia  contributed the most reports. There were five or fewer articles from Benin, Burundi, Libya, Namibia, Botswana, Comoros, Djibouti, Niger, Equatorial Guinea, and Swaziland , with 585 (72.5%) of 806 study sites being within a radius of 100 km and 392 (48.6%) of 806 sites being within a radius of 20 km of the nearest major city  published articles, infants in 12 (1.1%), children aged 1 to < 13 years in 146 (13.7%), and older children and adults (≥ 13 years) in 224 (21.0%). Of all reports, 386 (36.2%) included all ages, while age was not specified in 233 (21.9%) reports. The age distribution for each geographical region is provided in Additional file Blood was the main specimen analysed in 886 (83.2%) published reports, reflecting our selection criteria. Cerebrospinal fluid (CSF) samples were reported in 79 (7.4%) articles; a combination of CSF and blood culture in 40 (3.8%); bone marrow, joint, or liver aspirates in 13 (1.2%); multiple sample sources in 44 (4.1%); and the source was not stated in three (0.3%) articles  reports and serological assays in 111 (19.7%) reports. For viruses, 297 (79.4%) articles reported serological testing and 70 (18.7%) used PCR  published articles, viral infections in 374 (35.1%), parasitic infections in 47 (4.4%), and fungal infections in nine (0.8%). Of the 71 (6.7%) articles reporting multiple groups, 46 (64.8%) reported bacteria and fungi; 21 (29.6%) reported bacteria and viruses; two (2.8%) reported bacteria, viruses, and parasites; and two (2.8%) reported bacteria, viruses, and fungi. The median (range) number of pathogens reported in a study was one (1–31) with 827 (77.7%) studies reporting four or fewer pathogens and 82 (7.8%) reporting ≥ 10 pathogens. A list of pathogens reported by country is presented in Additional file Staphylococcus aureus (n = 194 reports), non-typhoidal Salmonella (NTS) (n = 184), Escherichia coli (n = 164), and Streptococcus pneumoniae (n = 149) , NTS (n = 49), and Staphylococcus aureus (n = 46) were the most reported isolates. Among infants, non-typhoidal Salmonella (n = 7), E. coli (n = 7), Streptococcus agalactiae (n = 6), Staphylococcus aureus (n = 5), and Streptococcus pneumoniae (n = 5) were the five most commonly reported. Among neonates, Klebsiella spp. including Klebsiella pneumoniae (n = 60), Escherichia coli (n = 48), Staphylococcus aureus (n = 41), and Streptococcus agalactiae (n = 25) were most reported. The overall distribution of articles by patient age and predominant mode of transmission is presented in Additional file Among 636 published reports of bacterial infections, the most commonly reported were due to us n = 19 reports,n = 69 published reports) and tick- and louse-borne relapsing fevers caused by Borrelia spp. (n = 32) were the most commonly reported vector-borne infections , the cause of Mediterranean spotted fever, which is transmitted by the dog tick and appears to have a widespread distribution throughout the African continent. There were five reports of Borrelia recurrentis, which is associated with relapsing fever, all from Ethiopia. Among neonates, there was a single report of Borrelia caucasica from Rwanda. There were no reports of vector-borne infections among infants.Rickettsial infections  was the leading cause of food- and water-borne bacterial infections followed by typhoidal Salmonella (n = 76), Brucella spp. (n = 21), Leptospira spp. (n = 14), and Shigella spp. (n = 14). Infections due to Listeria spp., Aeromonas spp., Cholera spp., and Campylobacter spp. were infrequent and reported in fewer than 10 articles , Streptococcus pneumoniae (n = 149), Klebsiella spp. (n = 92), Klebsiella pneumonia (n = 80), and Enterobacter spp. (n = 76) were the most commonly reported bacterial infections in this category. E. coli (n = 48) was the most commonly reported bacterium among neonates (n = 48) and infants (n = 7), and Streptococcus pneumoniae (n = 50) among children aged 1 to < 13 years.Coxiella spp. (n = 35 published reports) of which 31 were due to Coxiella burnetii, Mycobacterium spp. (n = 2), and Legionella spp. (n = 1) were the only predominantly air-borne bacterial infections reported.Among 415 published reports of viral infections, there were none in infants, and only three in neonates attributed to cytomegalovirus, enterovirus, and herpes simplex virus once each  was the most commonly reported arbovirus. Dengue virus (DENV) serotypes 1 to 4 were reported in 54 articles from 20 countries . Chikungunya virus was reported in 37 articles, the majority from studies conducted in Eastern Africa  , Ghana (n = 1), Morocco (n = 2), South Africa (n = 1), and Ivory Coast (n = 1). Hepatitis E was reported in Egypt in three studies and in one study each from Ethiopia, Ghana, Kenya, Somalia, and Sudan , Sudan (n = 3), Ghana (n = 1), and Somalia (n = 1). Hantaan virus was reported in seven articles from Cameroon, Central African Republic, Chad, Egypt, Guinea, Equatorial Guinea, Gabon, Mauritania, and Nigeria. Respiratory syncytial virus  was reported in an article from Egypt. No other air-borne viruses were reported.Measles virus was reported in eight articles in Nigeria (n = 79 published reports) was the most common viral infection reported in this category. Ebola virus disease (n = 71 reports) was the second most common viral infection reported in this category, of which 18 (25.3%) were published during the 2014–2016 West African outbreak. The other most frequently reported viruses were Lassa virus (n = 35), Marburg virus (n = 25), hepatitis B virus (n = 20), and hepatitis C virus (n = 14)  and children (n = 7). Cryptococcus spp. (n = 7) were commonly reported among adults and were reported in studies from Botswana, Malawi, and South Africa; of these, two studies included some HIV-infected patients.A list of all reported fungal infections, from 57 published articles, is presented in Additional file n = 28 reports) among both children (n = 7) and adults (n = 8) . Trypanosoma spp. (n = 4), Mansonella perstans (n = 2), Wuchereria bancrofti (n = 2), and Babesia spp. (n = 1) were the pathogens of the other vector-borne parasitic infections. Fasciola hepatica, the common liver fluke, was reported in three articles from Egypt, Schistosoma spp. in three articles from Egypt, Taenia spp. in an article each from Egypt and South Africa, and Loa loa in an article from Nigeria , moderate in 322 (30.2%), and low in 305 (28.6%). In the remaining 12 (1.1%) articles, the risk of bias was unclear  for neonatal sepsis in many LMICs. Other studies over the last decade have suggested that Klebsiella is being diagnosed more frequently than before in early onset neonatal sepsis [Most of the bacterial pathogens reported are well known and common in countries at all income levels Table . Howevern Africa , a resuln Africa , 33. Takl sepsis , 35.TabThe literature on viral infections was heavily biased toward the more lethal viral haemorrhagic fevers reflecting the attention that outbreaks of these diseases attract globally. The 2014–2016 West African outbreak was associated with a spike in the number of publications on Ebola virus disease, accounting for nearly a quarter of the Ebola virus disease reports published during the period of this review. There were only five reports of Enteroviruses, specifically coxsackie B3 virus, from the whole of Africa which contrasts with multiple descriptions of Enterovirus-71 outbreaks reported in Asia . Dengue This review has several limitations. First, as stated, the data presented here reflect neither overall incidence nor the prevalence of the microorganisms and should not be over-interpreted. That a pathogen was not reported from a country or region cannot be taken as definitive evidence of its absence, because this review did not capture records of an organism being sought but not found. Researchers are likely to look for pathogens that have been previously recognised or reported, or in order to answer a specific research question, such as on vaccine protective efficacy for a particular pathogen, or management of outbreaks (e.g. Ebola virus). It is worth noting that while malaria was reported across the entire African continent in the early 1980s, some northern and far southern regions have now eliminated the disease; this review did not exclude malaria-free countries. Reports on the presence of newsworthy pathogens, such as Ebola virus and Zika virus, are also more likely to be published, and thus, the clinical literature is susceptible to substantial reporting biases. Under-reporting is further compounded by the fact that some patients might have taken antimicrobials prior to hospital presentation, masking some bacterial pathogens. Nucleic acid amplification tests can also lack sensitivity for some infections, such as when timing of collection is after the bacteraemia or viremia has peaked. Routine data from clinical specimens tested in hospitals and private laboratories would provide a more representative picture of the leading causes of non-malaria febrile illness, but these are published infrequently. Increased electronic laboratory and clinical data capture has the potential to transform routine disease surveillance and information sharing in the future. Restricting the search to pathogens detected in blood and CSF meant that some important or common infections such as helminths and respiratory and sexually transmitted infections were omitted. However, restriction to organisms isolated from normally sterile sites limits the potential for misinterpretation of pathogen data in terms of distinguishing colonisation from true infection.The studies included in this review exhibited large heterogeneity of study design, patient population, and diagnostic assays used. It was not feasible to comprehensively assess the heterogeneity of case definitions, or the quality of various study elements, because of inconsistent reporting and varied laboratory and diagnostic techniques. Just over two-third of the studies included in this review were considered to be at moderate to high risk of bias (Additional file Candida spp. and some Gram-negative bacteria such as Acinetobacter baumannii. The latter was reported from all regions except Western Africa; of note, this predominantly nosocomial infection has been associated with a high case fatality risk of 41% in hospitalised patients in Thailand [Certain pathogens are more likely to be hospital-acquired, such as Thailand . The GloThailand . This reThis review provides a comprehensive summary of published reports on potential causes of non-malarial febrile illnesses in Africa. Pathogens and diseases previously under-recognised or not thought to be endemic on the continent are clearly present, including dengue virus and melioidosis. Testing and reporting of febrile disease aetiology in Africa is patchy, with probably substantial under-reporting of and under-consideration of important pathogens in many regions. Reporting of microbiology data needs to improve, including classifying bacterial infections as community- or hospital-acquired. This concern has been noted by other authors and is an impediment to current assessments of the burden of drug-resistant infections in LMICs . GuideliAdditional file 1. Further results Additional file 2. List of studies includedAdditional file 3. Assessment of risk of bias


To explore altered regional neuronal activity in patients with nonarteritic anterior ischemic optic neuropathy (NAION) and its correlation with clinical performances using the regional homogeneity (ReHo) method, which is based on resting-state functional magnetic resonance imaging (fMRI).  Thirty-one patients with NAION  and 31 age- and sex-matched normal controls (NCs)  were enrolled in the study. All patients underwent ophthalmic examination, including eyesight, intraocular pressure measurement, optimal coherence tomography (OCT), visual field analysis, and fMRI scans. After ReHo was calculated, we investigated group differences in results between the patients and NCs. We analyzed the relationship between ReHo values for different brain regions in patients with NAION and intraocular pressure, visual field analysis, and OCT. A receiver operating characteristic (ROC) curve was used to assess the diagnostic ability of the ReHo method. r = −0.462, P = 0.01). The area under the ROC curve for each brain region indicated that the ReHo method is a credible means of diagnosing patient with NAION.  Compared with NCs, patients with NAION exhibited higher ReHo values in the left middle frontal gyrus, left middle cingulate gyrus, left superior temporal gyrus, and left inferior parietal lobule. Additionally, they exhibited lower ReHo values in the right lingual gyrus, left putamen/lentiform nucleus, and left superior parietal lobule. ReHo values in the left superior parietal lobule were negatively correlated with right retinal nerve fiber layer values ( NAION was primarily associated with dysfunction in the default mode network, which may reflect its underlying neural mechanisms. Nonarteritic anterior ischemic optic neuropathy (NAION) typically presents in patients older than 50 years, with an estimated annual incidence of 2.3–10.2 cases per 100,000 people . NAION iRecently, optical coherence tomography (OCT) has been used to successfully detect NAION. OCT is a noninvasive technology that can detect microstructure changes in retinal layers, which is useful information for diagnosing NAION. Importantly, OCT can reveal changes in the integrity of the intermediate line and thickness of the outer nuclear layer of the retina. However, OCT only focuses on abnormal changes in the eyes. The visual system extends beyond the eye and retina to the optic nerve and the visual cortex of the brain. Whether NAION leads to dysfunction in these parts of the visual system remains unknown.Based on the blood oxygen level-dependent (BOLD) contrast technique, functional magnetic resonance imaging (fMRI) is an ideal choice for assessing cortical structural/functional abnormalities in NAION because of its high spatial resolution, noninvasiveness, and ability to reflect neuronal activity. A previous study using task-based fMRI found that activation in bilateral occipital cortex was lower in patients with NAION than in healthy controls . In contThirty-one patients with NAION were recruited from the ophthalmology department at Dongfang Hospital affiliated with Beijing University of Chinese Medicine. NAION was diagnosed by an experienced ophthalmologist with 5 years of experience. Among the 31 patients, 16 had unilateral NAION  and 15 had bilateral NAION, either sequentially or simultaneously. The following diagnostic criteria were applied: (1) a typical clinical history of sudden, painless, and monocular visual loss or successive bilateral visual loss; (2) received standardized treatment and evaluation in our hospital; and (3) no history of coronary artery disease, hypertension, sleep disorders, or drug addiction. Exclusion criteria included the following: (1) systemic features suggesting optic neuritis, giant cell arteritis, or posterior ischemic optic neuropathy, or a history of optic tumor or other ocular diseases; (2) symptoms of neurological disorders, mental disorders, or the inability or unwillingness to cooperate; and (3) abnormal function in the liver or kidney.Thirty-one normal controls (NCs) from the university (students) and hospital (staff) were also enrolled in the study. All the NCs were age and sex matched and met the following criteria: (1) no history of ocular disease or symptoms of neurological disease and (2) unaided eyesight > 1.0 on the vision chart.No participant had any contraindication for MRI scanning, such as claustrophobia or irremovable MRI-incompatible metal in the body. All participants underwent a vision acuity test, intraocular pressure measurement (IOP), visual field (VF) test, OCT to measure retinal nerve fiber layer (RNFL) thickness, and MRI scanning.The study was approved by the medical research ethics committee and institutional review board of Dongfang Hospital affiliated with Beijing University of Chinese Medicine, Beijing, China. All participants were asked to wear sponge earplugs and a black blinder during MRI scanning. In addition, the methods, potential risks, and purpose of the study were explained to each participant and all provided written informed consent.MR imaging was performed on a 1.5 Tesla Philips Intera Achieva system  with an eight-channel head coil. During the scan, participants were asked to keep their eyes open, remain motionless, and not to think about anything during the functional scans.General sagittal and axial T1-weighted turbo spin-echo (TSE) images, T2-weighted fast field-echo (FFE) images, and short T2-inversion recovery (STIR) images were acquired.Resting-state fMRI was obtained using an echo planar imaging (EPI) pulse sequence with each scan. Thirty-five axial slices were acquired with the following parameters: repetition time = 3000 ms, echo time = 30 ms, flip angle = 90, field of view = 220 mm × 220 mm, matrix = 64 × 64, thickness = 3.6 mm, and gap = 0.72 mm. Total scan time was 5.06 minutes with 30.36 seconds for each slice. Further, high-resolution structural images (3D BRAVO) were acquired with the following parameters: matrix = 256 × 256, field of view = 256 mm × 256 mm, thickness = 1.0 mm, number of excitation (NEX) = 2, repetition time = 6.5 ms, echo time = 3.2 ms, and flip angle = 8. Each T1 3D-BRAVO contained 161 images.http://www.rfmri.org/DPARSF_V2_3), which is based on SPM8  and REST  (http://restfmri.net/forum/REST_V1.8), and was implemented in MATLAB 2014a . Preprocessing comprised the following steps: (1) transform EPI DICOM files into NIFTI files, (2) remove first 10 volumes for signal equilibrium and participant adaptation to the scan environment, (3) slice timing correction, (4) head motion correction: to estimate translation and rotation for each participant  or whose rotation motion was greater than 1.5° in any direction was dismissed), and (5) fMRI images were spatially normalized and resampled to a standard stereotactic Montreal Neurological Institute (MNI) space using the echo planar-imaging template and coregistered at a resolution of 3 mm × 3 mm × 3 mm.Functional data processing was conducted using DPARSF (Data Processing Assistant for Resting State fMRI)  to reduce the effect of physiological high-frequency respiration, low-frequency drift, and cardiac noise. Then, individual ReHo maps were generated for each participant based on Kendall's coefficient of concordance (KCC) of the given voxel time series with its nearest 26 neighboring voxels. In addition, we made normalized ReHo maps because the averaged KCC for the whole brain can divide the KCC among each voxel. Finally, the remaining images were smoothed with a Gaussian kernel with a full-width-at-half-maximum of 4 mm × 4 mm × 4 mm.t-tests were performed using SPSS 17.0 software  to compare clinical data between patients with NAION and NCs. Pearson's linear correlation analyses were used to explore the correlation between the patient ReHo values and the clinical parameters with a statistical significance threshold of P < 0.05. The receiver operating characteristic (ROC) curves and the area under the curves (AUC) were used to analyze the ReHo values in the different brain regions, which were compared between the two groups of participants.Independent sample https://www.alivelearn.net/xjview) and REST software. REST was used for statistical analysis. Two-sample t-tests were used to evaluate the differences in ReHo values between patients and NCs with sex, age, and duration of disease as covariates of no interest. Voxels with a P < 0.05  corrected threshold of P < 0.05) and cluster size > 23 voxels indicated a significant difference between patients and NCs.The final functional MRI results were presented by xjview toolbox (P < 0.05). Furthermore, patients had significantly higher IOP and mean deficiency (MD) . Details are presented in We found no significant differences in age or sex between patients with NAION and the NCs. Compared with the NCs, patients with NAION had significantly worse visual acuity, thinner RNFLs, and lower mean sensitivity (MS)   . We did We speculated that ReHo values may be useful diagnostic markers for NAION. Thus, we performed the ROC curve analysis to determine the mean ReHo values for each brain region that differed between the groups. An area under the curve (AUC) above 0.8 indicates that NAION can be diagnosed accurately. We found individual AUCs for the left middle frontal gyrus (0.790), left middle cingulated gyrus (0.797), left superior temporal gyrus (0.869), left inferior parietal lobule (0.745), right lingual gyrus (0.846), left superior parietal lobule (0.843), and left putamen/lentiform nucleus (0.832). The combined AUC for the regions that exhibited lower ReHo values in patients  was 0.983. The combined AUC for the regions that exhibited higher ReHo values in patients  was 0.954 .During the resting-state condition, the default mode network (DMN) is continuously activated . The DMNThe lingual gyrus, located in area V2 of the visual cortex, is a key part of the visual cortex that receives feedforward connections from V1. Additionally, V2 plays a critical role in object and shape visual processing  and sterThe middle frontal gyrus (MFG), lying between the inferior and superior frontal gyri, has been widely reported to be involved in contingency awareness  and cognThe superior temporal gyrus (STG) is the secondary auditory area and plays an important role in auditory processing  and audiThe inferior parietal lobule (IPL) plays an important role in visual word recognition . DysfuncThe superior parietal lobule (SPL) is another part of the visual pathway, which plays a critical role in visuomotor coordination . It is aAn initially unexpected—and very interesting—finding was that locations with increased ReHo values were all located in the left hemisphere. Although this could be a coincidence, it might be related to the very important roles that the left cerebral hemisphere plays in visual-word recognition, which have been suggested by neuroimaging and neuropsychological studies , 46. IndIn the present study, ROC analysis was applied to determine the reliability of using ReHo values to diagnose patients. Several neuroimaging studies focusing on ocular diseases have successfully applied ROC analyses to discriminate those with ocular disease from NCs , 49, 50.The current study has several limitations that should be considered. First, NAION typically occurs unilaterally. Involvement of the other eye can occur years after the first eye is affected. However, patients typically visit different clinics because treatment is ineffective. When patients arrive at our hospital, some already exhibit NAION in both eyes. Thus, it was difficult to recruit patients with only one affected eye. Second, patients in the present study did not undertake neuropsychological tests though a few people complained insomnia or irritability. Further research is required to examine this issue in more detail. This limitation could be resolved using larger patient samples.The present study revealed that patients with NAION exhibit an abnormal spontaneous brain activity, including a negative correlation with contralateral RNFL. The abnormal spontaneous activity demonstrated that patients with NAION had undergone neural remodeling. The findings may be related to functional brain networks and the DMN, as well as to visual compensation. The current results provide important information that improves our understanding of the inherent neural mechanisms underlying NAION.