The anticoagulant treatment and clinical efficacy of heparin in sepsis remains controversial. We conducted a meta-analysis to estimate the clinical efficacy of unfractionated heparin (UFH) in adult septic patients.A systematic review of Medline, Cochrane Library, PubMed, Embase, WEIPU database, CNKI database, WANFANG database was performed from inception to January 2021. We included Randomized controlled trials (RCTs) and the main outcome was 28 d mortality. Data analysis was performed with Review Manager (RevMan) version 5.3 software. The meta-analysis included 2617 patients from 15 RCTs.P > 0.05) failed to reach statistical significance. UFH decreased multiple organ dysfunction syndrome (MODS) incidence , length of stay (LOS) in ICU  and ventilation time . And UFH had no adverse impact on bleeding .Comparing to control group, UFH could reduce 28 d mortality  especially for patient with Acute Physiology and Chronic Health Evaluation II (APACHE II) > 15, . In UFH group, the platelet (PLT)  was higher, the activated partial thromboplastin time (APTT) was shorter  and the prothrombin time (PT) results (This meta-analysis suggests that UFH may reduce 28 d mortality and improve the clinical efficacy in sepsis patients without bleeding adverse effect.The online version contains supplementary material available at 10.1186/s12871-021-01545-w. Sepsis is defined as life-threatening organ dysfunction by a dysregulated host response to infection . The patMany research studies focused on the possibility that anticoagulant therapy could improve the mortality of sepsis patients. However, there is a considerable controversy regarding the treatment of coagulopathy in sepsis. There is positive results in Yamakawa et al’s group which meant anticoagulant therapy was associated with better outcome according to the deterioration of both DIC and disease severity for septic patients . WhereasHeparin is a widely used anticoagulant which can prevent venous thromboembolism (VTE). In 2016, Surviving Sepsis Campaign (SSC) guideline first evaluated the use of heparin for the treatment of sepsis and septic shock . The JapHeparin contains UFH and low molecular weight heparin (LMWH). The mechanisms of action for UFH and LMWH are different in sepsis, so their effects are various. UFH mainly inhibits the activity of thrombin and factor (F) Xa by binding to antithrombin (AT). The inhibition of thrombin requires a heparin chain comprising of at least 18 saccharide units. Therefore, LMWH can only exert anticoagulant effect by inhibiting the action of FXa. Furthermore, recent research studies proved that UFH possesses various biological properties, such as anti-inflammatory and immunomodulatory effects .UFH seems to be more promising in the treatment of sepsis because of its multiple biological activities. Our previous studies have shown that UFH could protect endothelial cells, improve endothelial barrier dysfunction, and inhibit inflammatory response to preserve organ function and improve the prognosis of septic patients , 12.TherThe research was in accordance with the Preferred Reporting Items for the Systematic Reviews and Meta-Analyses (PRISMA) guidelines . We condFor example, “Sepsis” [MeSH], “DIC” [MeSH], “Heparin” [MeSH] and corresponding keywords were used for search with various combinations of the operators “AND” and “OR” in PubMed. The search was slightly adjusted according to the requirements of the various databases. The PubMed strategy details were presented in the Supplementary Table Studies that were included met the following criteria:1. RCTs; 2. Studies that aimed to assess the clinical efficacy of UFH treatment on mortality, the incidence of bleeding complications and the coagulation indicators, such as the platelet (PLT), prothrombin time (PT) and activated partial thromboplastin time (APTT); 3. Participants in studies were diagnosed with sepsis, septic shock or DIC; 4. The administration of low-dose UFH, given at any route or frequency.11/L; 6.Patients with severe brain trauma, cerebral aneurysm, arteriovenous malformation and gastrointestinal bleeding history; 7.Renal failure required hemodialysis  or hemofiltration (hemofiltration); 8.Both the experimental and control group received UFH.1.Quasi-randomized controlled trials, for example randomization according to hospital number; 2.Review, repeated literature reports, or animal studies; 3. Studies involving pediatric patients, pregnant or lactating patients; 4.Ex-transplant recipients, patients who had been treated with anticoagulant drugs 48 h ago and had coagulation disorders or history of abnormal coagulation; 5.Patients’ PLT < 30 × 10Two researchers independently extracted the data from each article that met the inclusion criteria but not exclusion criteria. If the opinion of two reviewers conflicted in the process, another reviewer evaluated the studies independently. We then finalized the decisions through group discussion. The following data were extracted: the first author’s name, year of publication, number of study sites, number of patients, population, age, Acute Physiology and Chronic Health Evaluation II (APACHE II) score, intervention (dose and duration), control treatment, the 28 d mortality and incidence of bleeding complications.. Multiple organ dysfunction syndrome (MODS) incidence, length of stay (LOS) and the duration of ventilation in ICU were recorded as the secondary outcomes. We graphically displayed the outcomes by forest plots and visually inspected the potential publication bias with a funnel plot [The primary outcome was 28 d mortality. We made reference to APACHE II score to perform subgroup analysis. Also, PLT, PT and APTT were regarded as the coagulation indicatorsnel plot .The Cochrane Collaboration Risk Of Bias tool (ROB) was utilized to assess the quality of all analyzed trials by two authors . DisagreThe meta-analysis was performed using the Review Manager (RevMan) version 5.3 software. Reference to Cochrane Handbook of Systematic Reviews, 28 d Mortality and bleeding complication (Binary variables) were expressed as relative risk (RR) with 95% confidence intervals (CIs), a weighted pooled RR was calculated using the Mantel-Haenszel method . The con2 test was used to measure statistical heterogeneity. When I2 = 0, there was no heterogeneity. When the I2 < 40% or I2 between 30 and 60%, there was considered as low or moderate statistical heterogeneity in the studies, the fixed effect model was used for analysis [2 > 50% [Ianalysis . Random [2 > 50% . All resTwo researchers graded the evidence for each outcome based on the following six domains: study design, risk of bias, inconsistency, indirectness, imprecision, and publication bias. If there were discrepancies, another author would take part in this discussion. The strength of evidence could be classified as “high,” “moderate,” or “low” .Of the 2326 citations identified from electronic and hand searches, after duplicate checking and screening the titles and abstracts of all searched articles, we selected 52 studies for full text review. Through the full-text screening, this meta-analysis contained 15 studies –36, as tThe characteristics of the included studies were shown in Tables Moreover, after calculation, the result showed that the median value of APACHE II score could be used to estimate its mean in Jaimes et al’ trial. In Supplementary Fig. Main reasons for bias for individual studies were shown in Fig. P = 0.009 < 0.05), indicating a statistically significant reduction in 28 d mortality in UFH-treated patients with sepsis. There was no evidence of heterogeneity between studies (I2 = 0.0%).The forest plot for 28 d mortality was shown in Fig. P = 0.02 < 0.05; I2 = 0.0%. And for Sepsis 2.0, with the RR = 0.58; 95% CI = 0.35 to 0.94; P = 0.03 < 0.05; I2 =0.0%. The 28 d mortality of patients treated with UFH was statistically significant in both sepsis 1.0 group and sepsis 2.0 group. There was no evidence of heterogeneity between studies (I2 = 0.0%). There was high heterogeneity among subgroup studies (I2 = 54.1%). The finding from this subgroup analysis further reflects the benefit of UFH for treating septic patients. In clinical practice, APACHE II score was widely used to indicate the disease severity and predict the prognosis of critically ill patients. To date, there is no unified international rules relating to APACHE II cut-off value. In this trial, the mean of APACHE II score ranged from 9 to 23.8 for patients in UFH group, 13, 15, 17 and 19 were selected as APACHE II cut-off values respectively. On the basis of various APACHE II scores, subgroup analysis was performed  in UFH group was higher than that in control group. The PT results  failed to reach statistical significance, although the considerable heterogeneity, the APTT in UFH group was shorter than that in control group .There were eight studies , 32–36 rP = 0.002 < 0.05; I2 = 0%).UFH reduced the incidence of MODS and the LOS in ICU. Four studies , 30, 34 P < 0.00001 < 0.05; I2 = 66%). But the heterogeneity was considerable.Five researches , 30, 33 P < 0.00001 < 0.05; I2 = 0%).Four studies , 30, 34 P = 0.80 > 0.05; I2 = 0%). There were no statistically significant differences. UFH had no effect on bleeding events in patients with sepsis.Four studies reported the bleeding complications , 27, 32 In this study, we demonstrated that UFH was an effective treatment for sepsis. The 28 d mortality was relatively reduced by 16% in the UFH group. What’s more, the 28 d mortality of sepsis patients with APACHE II > 15 was relatively reduced by 17% in the UFH group. There were no significant bleeding complications in UFH group, which indicated the safety of UFH. To our knowledge, our study is the first meta-analysis to focus on the effect of UFH in sepsis.Almost all the sepsis patients experienced coagulation abnormalities, ranging from minor changes that could only be detected by extremely sensitive tests to DIC . Many anThe role of UFH in sepsis is much more than just an anticoagulant . Our teaThe conclusion of this meta-analysis was consistent with previous reports. Multiple lines of evidence suggested that UFH may improve clinical outcomes in sepsis. UFH is more suitable in China as well because it is widely available and inexpensive. Higher quality evidence is needed to guide clinical practice .To date, there were three meta-analyses on the role of heparin in sepsis published in English. Wang et al’s meta-analysis published in 2014 concluded that heparin therapy reduced 28 d mortality in adult severe sepsis patients . There wSubsequently, Zarychanski et al’s meta-analysis was published in 2015, which showed that heparin was associated with decreased mortality in patients with sepsis, septic shock and DIC . The disThe third meta-analysis was about the efficacy and safety of LMWH in patients with sepsis . There aThere are several advantages in this article. First, we enrolled studies both in English and in Chinese in this research, which meant the generalization of the current findings. Second, to our knowledge, our study is the first meta-analysis to focus on the effect of UFH in sepsis. Last, the large number of subjects included and the diversity in patients’ characteristics provided diverse information.The study also has several limitations. First, as shown in the forest plot, the participants in Levi et al’s study took up more than half of the total number and all of them received recombinant activated protein C, which has been withdrawal from the market. Second, due to the various publication years, the definitions of sepsis, septic shock and DIC had changed. In 2016, the Sepsis-3 Task Force updated the clinical criteria for sepsis, excluding the need for systemic inflammatory response syndrome (SIRS) and the concept of severe sepsis . Third, UFH may reduce 28 d mortality and improve the clinical efficacy for sepsis patients without bleeding complications. We call for large multicenter RCTs to evaluate the clinical value of UFH in sepsis patients because of the moderate quality of this body of evidence.Additional file 1 : Figure S1. The funnel plot of 28 d mortality.Additional file 2 : Figure S2. The forest plot of PLT.Additional file 3 : Figure S3. The forest plot of PT.Additional file 4 : Figure S4. The forest plot of APTT.Additional file 5 : Figure S5. The forest plot of MODS incidence.Additional file 6 Figure S6. The forest plot of LOS in ICU.Additional file 7 : Figure S7. The forest plot of the duration of Ventilation.Additional file 8 : Figure S8. The forest plot of bleeding complication.Additional file 9 : Table S1. The PubMed strategy.Additional file 10 : Table S2. Excluded literature information.Additional file 11 : Table S3. Evidence assessment.


Altered DNA damage response (DDR) has emerged as an important mechanism for the development of aggressive prostate cancer among men of European ancestry but not other ancestry groups. Because common mechanisms for aggressive disease are expected, we explored a large panel of DDR genes and pathways to demonstrate that DDR alterations contribute to development of aggressive prostate cancer in both African American and European American men.We performed a case-case study of 764 African American and European American men with lethal or indolent prostate cancer treated at 4 US hospitals. We calculated carrier frequencies of germline pathogenic or likely pathogenic sequence variants within 306 DDR genes, summarized by DDR pathway, and compared lethal cases against indolent cases using 2-sided Fisher’s exact tests. Secondary analysis examined if carrier frequencies differed by ancestry.P = 4.30 × 10−4), even after excluding BRCA2 . The carrier frequency was similar among lethal cases of African (16.7% including and 15.8% excluding BRCA2) and lethal cases of European (19.3% including and 14.2% excluding BRCA2) ancestry. Three DDR pathways were statistically significantly associated with lethal disease: homologous recombination (P = .003), Fanconi anemia (P = .002), and checkpoint factor (P = .02).Lethal cases were more likely to carry a pathogenic sequence variant in a DDR gene compared with indolent cases (18.5% vs 9.6%, Our findings suggest that altered DDR is an important mechanism for aggressive prostate cancer not only in men of European but also of African ancestry. Therefore, interrogation of entire DDR pathways is needed to fully characterize and better define genetic risk of lethal disease. BRCA2, among men with advanced prostate cancer  has emerged as an important mechanism for the development of aggressive prostate cancer. Several studies have reported enrichment of rare germline pathogenic variants within DDR genes, particularly e cancer . Pritchae cancer  examinednd CHEK2 ,6.Less is known regarding the role of DDR genes in the development of aggressive prostate cancer among men of other ancestry groups. This is particularly important in men of African ancestry. African American men have higher prostate cancer incidence and mortality rates than European American men , and genWith the ultimate goal of identifying genetic markers that can predict disease course and personalize treatment for men of all ancestries, we sought to identify the DDR pathways associated with lethal prostate cancer in African American and European American men. We hypothesized that underlying mechanisms for aggressive disease are similar and that DDR gene alterations in general contribute to development of aggressive prostate cancer irrespective of ancestry.We conducted an unmatched case-case study of African American and European American men with lethal or indolent prostate cancer who had previously provided a blood or buccal swab tissue sample and had been treated at one of the following hospitals up until 2016: Dana-Farber Cancer Institute and Brigham and Women’s Hospital, the Hospital of the University of Pennsylvania, Tulane University, and Wayne State University. Information on self-reported race and ethnicity was extracted from medical records and subsequently verified using ancestral markers as described below. Lethal cases included men diagnosed with metastatic disease (at diagnosis or during follow-up) or who had died from prostate cancer. Indolent cases were selected based on their clinical characteristics and included men treated with curative intent or undergoing active surveillance, with Gleason score 6 tumors, and without evidence of recurrence or additional treatment at 5 years. Of the 837 men originally included in the sample, 24 men who did not fulfil the inclusion criteria for lethal disease (n = 1) or indolent cancer (n = 23) were removed following a medical chart review. Fourteen control men who had biopsy Gleason 6 cancer but were upgraded at radical prostatectomy to Gleason 3 with a small quantity of Gleason 4 were included. Of the 813 men remaining, 764 (94.0%) were included for analysis after successful sequencing and quality control as outlined below.All patients provided written informed consent, and the project was approved by the institutional review board committees at each site. The project as a whole was approved by the Brigham and Women’s Hospital Institutional Review Board (2015P002118).Targeted sequencing of 306 preselected DDR genes was done at the McDonnell Genome Institute at Washington University in St. Louis   and postEU = 102, NAF = 108), and their loci overlapped with the targeted regions. The genotypes of these locations were collected into a single matrix, and the principal components (PCs) of the dataset were identified using a singular value decomposition approach. We found that the first 2 eigenvectors separated the 2 ancestries almost perfectly. Outliers were identified using a k-nearest neighbors (k = 15) approach, measured in the PC1-PC2 plane, and were subsequently removed (n = 12).To ensure that the self-reported information on race and ethnicity corresponded to either African or European ancestry groups, 210 single-nucleotide polymorphisms were collected from the ExAC database  that werBased on literature and database searches ,15, geneP less than .05 considered as statistically significant. Given the sample size and rarity of variants, hypothesis testing and P value calculation were limited to pathway-based analysis only. In secondary analyses, comparisons were made stratified by ancestry and age at diagnosis. We further used logistic regression to calculate odds ratios (ORs) and 95% confidence intervals (CIs) for the association between carrying any PSV within a DDR gene and lethal prostate cancer, adjusting for age at diagnosis and PCs 1-3. A model including an interaction term with ancestry was fitted to test for effect modification by ancestry. Variance of unknown clinical significance (VUS) was examined in a sensitivity analysis that included the most likely pathogenic variants among the VUS  were African American and 576 (75.4%) European American. Among the 410 lethal cases, the median age at diagnosis was 62.7 years and the median age at lethal event (metastatic disease or prostate cancer–specific death) was 66.9 years . The mosWithin the 306 sequenced DDR genes, 95 unique PSVs were identified . Most meP = 4.30 × 10−4) (Pinteraction = .73). In adjusted analyses, the odds ratio was 2.30 (95% CI = 1.44 to 3.66) for the combined group, 1.81 (95% CI = 0.67 to 4.86) for the African ancestry group, and 2.49 (95% CI = 1.47 to 4.22) for the European ancestry group. Analyses stratified by age at diagnosis demonstrated an even higher carrier frequency in lethal cases diagnosed before age 65 years (19.8%)    Table 3 (19.8%) . This paBRCA2 is well known to be associated with aggressive prostate cancer. To determine if additional DDR alterations were associated with risk, we repeated the analysis excluding PSVs in BRCA2. The carrier frequency of having at least 1 PSV in a DDR gene was 14.6% in lethal cases overall compared with 9.6% in indolent cases (P = .04)  compared with lethal cases of European ancestry (5.1%)  and European (14.2%) ancestry. Using the gene panel from Pritchard et al. , with the highest carrier frequency among lethal African American cases (P = .03), suggesting that not all pathogenic sequence variants are captured by current classification. In addition to VUS within the checkpoint factor pathway, we observed a possible enrichment of VUS among lethal cases within BRCA1, SETX, and DCLRE1C  compared with indolent cases  and the MMR pathway (which includes MLH3 and MSH2) may be of special interest for men of African ancestry given the observation of PSVs among lethal cases only, for whom carrier frequencies of 2.6% were observed. ATM and the MMR pathway have previously been pointed out as potentially relevant for prostate cancer etiology and aggressiveness among men of African ancestry  is involved in cell cycle regulation and has been associated with DNA damage-induced apoptosis  and indolent cases (9.5% for African and 9.6% for European indolent cases). However, our analysis is based on a variant classification of genes that relies on data from European ancestry groups, and it is possible that VUS matters for African ancestry groups. Data from both our study and others .Role of the funder: The funder had no role in the design of the study; the collection, analysis, and interpretation of the data; the writing of the manuscript; and the decision to submit the manuscript for publication.Disclosures: The authors report no conflict of interest.Author contributions: AP: conceptualization, data curation, formal analysis, investigation, methodology, software, visualization, writing—original draft. MK: conceptualization, data curation, formal analysis, investigation, methodology, software, visualization, writing—review and editing. ZS: conceptualization, investigation, methodology, validation, writing—review and editing. OS: conceptualization, investigation, resources, validation, writing—review and editing. JS: conceptualization, investigation, resources, validation, writing—review and editing. IJP: conceptualization, investigation, resources, validation, writing—review and editing. TRR: conceptualization, investigation, resources, validation, writing—review and editing. KLP: conceptualization, investigation, validation, writing—review and editing. LAM: conceptualization, investigation, validation, writing—review and editing. MMP: conceptualization, investigation, resources, validation, writing—review and editing. ASK: conceptualization, funding acquisition, methodology, project administration, supervision, investigation, resources, validation, writing—review and editing.Prior presentations: Preliminary result from this study has been presented at the Annual Dana-Farber/Harvard Cancer Center Celebration of Early Investigators in Cancer Research on January 21, 2021.The data underlying this article cannot be shared publicly due to the privacy of individuals that participated in the study. The data will be shared on reasonable request to the corresponding author.pkab097_Supplementary_DataClick here for additional data file.


Clostridium perfringens is a spore-forming anaerobic pathogen responsible for a variety of histotoxic and intestinal infections in humans and animals. High-resolution genotyping aiming to identify bacteria at strain level has become increasingly important in modern microbiology to understand pathogen transmission pathways and to tackle infection sources. This study aimed at establishing a publicly available genome-wide multilocus sequence-typing (MLST) scheme for C. perfringens. A total of 1,431 highly conserved core genes  were indexed for a core genome-based MLST (cgMLST) scheme for C. perfringens. The scheme was applied to 282 ecologically and geographically diverse genomes, showing that the genotyping results of cgMLST were highly congruent with the core genome-based single-nucleotide-polymorphism typing in terms of resolution and tree topology. In addition, the cgMLST provided a greater discrimination than classical MLST methods for C. perfringens. The usability of the scheme for outbreak analysis was confirmed by reinvestigating published outbreaks of C. perfringens-associated infections in the United States and the United Kingdom. In summary, a publicly available scheme and an allele nomenclature database for genomic typing of C. perfringens have been established and can be used for broad-based and standardized epidemiological studies.IMPORTANCE Global epidemiological surveillance of bacterial pathogens is enhanced by the availability of standard tools and sharing of typing data. The use of whole-genome sequencing has opened the possibility for high-resolution characterization of bacterial strains down to the clonal and subclonal levels. Core genome multilocus sequence typing is a robust system that uses highly conserved core genes for deep genotyping. The method has been successfully and widely used to describe the epidemiology of various bacterial species. Nevertheless, a cgMLST typing scheme for Clostridium perfringens is currently not publicly available. In this study, we (i) developed a cgMLST typing scheme for C. perfringens, (ii) evaluated the performance of the scheme on different sets of C. perfringens genomes from different hosts and geographic regions as well as from different outbreak situations, and, finally, (iii) made this scheme publicly available supported by an allele nomenclature database for global and standard genomic typing. Clostridium perfringens is a Gram-positive anaerobic bacterium that is widely distributed in the soil and feces of humans and animals   in poultry  . The tox(A to G) . Of the gression . Type A gression , 6. In cin lambs , 4, whil poultry .Bacterial strain typing is essential for outbreak investigations, epidemiological surveillance, and evaluation of control measures . MultiloC. perfringens, a cgMLST system was first utilized to describe the clonal relationship between netF-positive strains from enteritis cases in foals and dogs  in the reference genome ATCC 13124 . The core target genes cover 41.2% (1.34 megabases) of the full genome size and were distributed unevenly across the genome, with a higher representation toward one chromosomal replichore . This distribution is consistent with previous studies that reported skewed presence of the species core genome . The cgMLST typing results showed that at least 95% of the cgMLST target genes were present in 273 of the 282 (96%) genomes examined , with a mean ± SD of 99.08 ± 1.3% of the 1,431 target genes detected per genome for all genomes (Table S1a). The nontypeable target genes averaged 13 ± 20 genes  (Table S1a). This includes (i) genes that were completely absent from the genome, i.e., with BLAST hits below the defined thresholds , and (ii) genes without allele assignment due to frameshifts, internal stop codons, or absence of a single start/stop codon  (Table S1b).In order to evaluate the novel scheme, an independent set of 282 P < 0.05) (Table S1b).The average number of alleles reported for each cgMLST target gene was 67 ± 28 alleles  . The number of distinct cgMLST allelic profiles was 259 for the 282 genomes (missing data ignored in pairwise comparisons). The Simpson's index of the cgMLST was 0.999  . Of inten = 259), indicating that cgMLST and core genome SNP provided comparable resolution for the 282 genomes . A total of 151,338 variable sites (11.3%) were identified in an alignment concatemer of the 1,431 genes (1.34 Mb). Of these, 60,587 polymorphic sites were predicted with Gubbins to be outside recombinant regions, including 42,344 sites (3.1%) that were parsimony informative. The number of distinct genotypes defined with recombination-unfiltered single nucleotide polymorphisms (SNPs) was 261  and with recombination-free SNPs was 244 , nearly equivalent to the distinct profiles identified with cgMLST ( genomes .C. perfringens (n = 219) belonged to phylogroup III, followed by phylogroups I (n = 45), II (n = 11), IV (n = 1), and V (n = 6)  . As desc (n = 6) , phylogr (n = 6) .Furthermore, calculation of the number of differing alleles and nucleotides (SNPs) for each pair of genomes showed direct correlation , except for large genetic differences between strains, where cgMLST distances became saturated and less informative .sigK gene of the Deguchi scheme and the recA gene of the Jost and Hibberd schemes were included with reduced lengths . In total, the Hibberd scheme had 11 loci of size 5,420 bp, the Deguchi scheme had 8 loci of size 5,090 bp, and the Jost scheme had 8 loci of size 2,833 bp. The sequence types (ST) defined with the modified schemes included 166 STs , 180 STs , and 191 STs  for Jost’s, Hibberd’s, and Deguchi’s schemes, respectively  genomes required minor modifications; the ectively . As expeectively . The treectively .de novo processed, revealing genomes with average size of 3.08 ± 0.21 Mb and average sequencing depth of 85.7 ± 18.6-fold (Table S1a).We applied the scheme to published outbreaks strains described by Carey and colleagues  and Kiu The typeability percentage of the 52 genomes with cgMLST was 99.1% ± 0.4% on average , except one genome with low assembly quality having 91.3% of cgMLST targets. The 52 genomes were indexed into 45 distinct cgMLST allele profiles, with missing data ignored in pairwise comparisons  tree of the Roary-calculated core gene alignment (ERR3377387) because of poor assembly metrics . The other 103 genomes comprise 71 from 13 foodborne outbreaks and 32 from 10 care homes outbreaks. The genomes had a mean genome size of 3.31 ± 0.18 Mb with a mean sequencing depth of 86.03 ± 15.9-fold. On average, 99.1%  of cgMLST targets were present in the genomes , with all genomes represented by 85 distinct cgMLST allele profiles.Kiu and colleagues  studied lignment . We recoWe first used cgMLST to reexamine the foodborne outbreaks   for use with SeqSphere (https://pubmlst.org/) as an open-source platform that provides web-accessible analyses for comparative genomics  reference for the data set under study, which is important for reference-based SNP typing methods. The allele nomenclature databases of cgMLST are easily expandable with new typing data that enhance global epidemiologic surveillance. On the other hand, because cgMLST uses assembled genomes for typing, the results are likely to be affected by sequencing errors and assembly artifacts that are difficult to distinguish from true variants , and a further 38 genomes were selected as representative of the five major phylogroups of the species  were allowed up to three codons per gene relative to the reference genes. The typeability percentage was estimated in this pilot set of genomes (Table S1a), and we excluded 71 genes that were missing or have not been typed in more than 5% of these genomes. The final set of core genes served as targets for cgMLST (Table S1b). In addition, 1,365 genes were compiled from the filtered genes as accessory targets  database and recent studies  was performed using nucleotide sequences of all cgMLST genes from the evaluation set (282 genomes) . For each gene, multiple sequence alignment was done with MAFFT v7.30 , 37. TheC. perfringens were used to compare with cgMLST  genomes. For Hibberd’s and Jost’s schemes, we also included 208 bp of recA, as 16 genomes had this gene fragmented at contig breaks or had the gene split by an insertion of an ∼2,860-bp DNA region. In addition, 5 genomes were excluded because of missing MLST loci; thus, 277 genomes were left for comparison. Extracted MLST loci of each genome were aligned using MAFFT v7.30 . The genome sequences from these outbreaks were recently analyzed using SNP approaches (de novo processed using FastQC (To evaluate the applicability of the proaches , 17 and g FastQC , shovillg FastQC , FastANIg FastQC , Quast (g FastQC , and cheg FastQC  as menti


Blood-brain barrier (BBB) injury is prevalent in patients with HIV-associated dementia (HAD) and is a frequent feature of HIV encephalitis. Signs of BBB damage are also sometimes found in neuroasymptomatic HIV-infected individuals without antiretroviral therapy (ART). The aim of this study was to investigate the integrity of the BBB before and after initiation of ART in both neuroasymptomatic HIV infection and in patients with HAD.We determined BBB integrity by measuring cerebrospinal fluid (CSF)/plasma albumin ratios in archived CSF samples prior to and after initiation of ART in longitudinally-followed neuroasymptomatic HIV-1-infected individuals and patients with HAD. We also analyzed HIV RNA in blood and CSF, IgG Index, CSF WBC counts, and CSF concentrations of β2-micoglobulin, neopterin, and neurofilament light chain protein (NfL).We included 159 HIV-infected participants; 82 neuroasymptomatic individuals and 77 with HAD. All neuroasymptomatic individuals (82/82), and 10/77 individuals with HAD, were longitudinally followed with a median  follow-up of 758 (230–1752) days for the neuroasymptomatic individuals, and a median (IQR) follow-up of 241 (50–994) days for the individuals with HAD. Twelve percent (10/82) of the neuroasymptomatic individuals and 80% (8/10) of the longitudinally-followed individuals with HAD had elevated albumin ratios at baseline. At the last follow-up, 9% (7/82) of the neuroasymptomatic individuals and 20% (2/10) of the individuals with HAD had elevated albumin ratios. ART significantly decreased albumin ratios in both neuroasymptomatic individuals and in patients with HAD.These findings indicate that ART improves and possibly normalizes BBB integrity in both neuroasymptomatic HIV-infected individuals and in patients with HAD. Human immunodeficiency virus type 1 (HIV-1) invades the central nervous system (CNS) early in primary infection . IntrathPrior to the introduction of modern antiretroviral therapy (ART), approximately 30 to 40% of patients with AIDS developed HIV-associated dementia (HAD) , 11. BBBART drastically reduces CSF HIV RNA levels  and CSF The effect of ART on the BBB has only been studied in a few case reports, where patients with HAD showed normalized BBB function after initiation of ART , 22. TheIn this retrospective longitudinal study, we analyzed archived CSF samples from HIV-infected adult (≥ 18 years) individuals with no neurological symptoms, and from individuals with HAD, both before and after initiation of ART, which included three active agents: two nucleoside reverse transcriptase inhibitors plus one non-nucleoside reverse transcriptase inhibitors, protease inhibitor, or integrase inhibitor. The samples were collected from 23 September 1986 until 1 June 2017 at three locations: Sahlgrenska University Hospital, Gothenburg, Sweden; San Francisco General Hospital, San Francisco, CA, USA, and St. Vincent’s Hospital, Sydney, Australia. To meet the inclusion criteria, participants showing no neurological symptoms had to be treatment-naïve, or off treatment for a minimum of 6 months prior to initiation of ART. To study the effect of efficient ART, they had to reach viral suppression within 7 months and remain virally suppressed for more than 6 months. Viral suppression within 7 months was chosen to allow individuals who had a reduction in plasma HIV RNA levels as expected on treatment, but where the lumbar puncture was not performed exactly at 6 months. Blips with a maximum of 500 HIV RNA copies/mL at the time of lumbar puncture during follow-up were considered acceptable. For individuals who had > 1 valid sample date according to the inclusion criteria, the most recent sample before initiation of ART was included in the study. Neuroasymptomatic participants were clinically assessed for neurocognitive and neurologic symptoms. Those who had neurologic symptoms or complaints at baseline, or developed during follow up, were excluded in order to more clearly evaluate the BBB and the associations with other biomarkers. No routine neurocognitive testing was performed on neuroasymptomatic patients.Participants without neurologic signs or complaints were designated neuroasymptomatic. The diagnosis of HAD was based on the Centers for Disease Control and Prevention and American Academy of Neurology Task Force criteria, using standard laboratory, neuropsychological, and clinical evaluations , 24. TreInclusion in the HAD cohort required a lumbar puncture sample both prior to and while on ART. Participants had to be treatment-naïve or off treatment for 6 months or more, and must have reached viral suppression. In accordance with the inclusion criteria participants with HAD who recovered on treatment were not excluded. Since HAD is a rare condition and few individuals had been followed longitudinally prior to and during ART, we also included individuals with HAD who had a lumbar puncture either prior to initiation of ART or while on ART, and who had been virally suppressed for 6 months or more.Lumbar punctures during follow up in all participants were performed according to different schedules depending on the research site. In Gothenburg, lumbar punctures after initiation of ART were made after 3 months, 12 months, and yearly thereafter. In San Francisco lumbar punctures were made frequently during the first 3 months and thereafter with a few months to 1 year’s interval. From Sydney, only cross-sectional data on patients with or without ART was provided.CSF samples were obtained in accordance with research protocols approved by the institutional review boards of each of the study sites, and in compliance with the Helsinki Declaration, either within the context of studies of the natural history of HIV infection , or during diagnostic evaluations (for patients with HAD). All participants gave their informed consent or, if their capacity to provide consent was questioned, consent was also obtained from those with power of attorney.Quantitative determinations of CSF and plasma albumin was performed with nephelometry . Albumin ratio was calculated as CSF albumin (mg/L)/plasma albumin (g/L) and used to evaluate BBB function. Reference values were < 6.8 for individuals younger than 45 years and <  10.2 for individuals ≥45 years of age .+ T-cell counts, CSF leukocytes (WBC), IgG index, and CSF/blood albumin measurements were performed at the local clinical laboratories. IgG index as a marker of intrathecal IgG synthesis was defined as [CSF IgG (mg/L)/serum IgG (g/L)]/[CSF albumin (mg/L)/serum albumin (g/L)] [CSF neopterin was analyzed using a commercially available immunoassay  according to the manufacturer’s instructions. The normal upper reference value for CSF neopterin is 5.8 nmol/L.  CSF NfL n (g/L)] . The refn (g/L)] ; and for10 transformed to reduce skewness.Descriptive statistics were performed using Prism, version 9.0  or SPSS, version 27  or R, version 4.0, software. All continuous variables except age and leukocyte count were logAssociations were analyzed with Pearson correlation coefficients. Independent Student’s t-test was used for comparison between two groups, and Wilcoxon signed rank test was used for evaluation over time at 6 months and 3 years with interpolated values using LOCF (last observation carried forward) if all values were prior to these time points.n = 47) or on ART (n = 20); for these individuals only cross-sectional data were available (Table p < 0.0001), albumin ratio (p < 0.0001), CSF neopterin (p < 0.0001), and CSF NfL (p < 0.0001), and significantly lower CD4+ T-cell count (p < 0.0001) and CD4 nadir (p < 0.0001), compared to neuroasymptomatic participants. There was no significant difference in age between the groups.In total, we included 159 HIV-infected participants; 82 neuroasymptomatic individuals, and 77 with HAD. All of the neuroasymptomatic individuals and 10 of those with HAD were followed longitudinally with lumbar punctures both pretreatment and during ART. Sixty-seven of the participants with HAD had lumbar punctures either only pre-ART  had an elevated albumin ratio at baseline. Baseline characteristics for these ten did not differ compared to the others in the neuroasymptomatic cohort. At the last follow-up 9 % (7/82) had elevated albumin ratios.p < 0.05) and 3 years (p < 0.01) , CSF neopterin , CSF β2-microglobulin , WBC , and CSF NfL  also decreased significantly. There was a trend towards a significant decrease in the IgG index at 6 months (p = 0.06), and a significant decrease after 3 years of treatment (p < 0.001) p < 0.05)p < 0.001) and a significant correlation with age . No correlations were found between albumin ratio and CSF NfL, inflammatory markers such as CSF neopterin, β2-microglobulin, or WBC; nor with plasma or CSF HIV RNA, CD4+ T-cell count, or CD4 nadir.At baseline, there was an inverse correlation between albumin ratio and IgG Index . At the same defined time points, there was a significant reduction of CSF HIV RNA  and CSF neopterin (p < 0.01), p < 0.01),  showed elevated albumin ratios. In the cross-sectional HAD cohort of those with only one lumbar puncture who were on ART, 25% (5/20) had elevated albumin ratios. In the longitudinal HAD cohort, 20% (2/10) had elevated albumin ratios at their last follow-up. For these ten individuals with HAD who were longitudinally followed, the albumin ratio declined, although not significantly, at 6 months; and significantly after 3 years .In the 57 untreated individuals with HAD , no correlation was found between albumin ratio and plasma HIV RNA, CSF HIV RNA, CD4In all longitudinally followed individuals (82 neuroasymptomatic and 10 with HAD), the decrease in CSF viral load and neopterin occurred early after initiation of ART, whereas the decrease in CSF β2-microglobulin and IgG index occurred later. Despite the normalization of CSF HIV RNA, levels of inflammatory markers in CSF remained slightly elevated. In individuals with HAD, the plateau of elevated CSF neopterin was higher than for neuroasymptomatic individuals Fig. .Fig. 3Tin = 631), where 16% of untreated neuroasymptomatic individuals showed elevated albumin ratios. It also is in accordance with other studies [In our study we found that BBB integrity improved after initiation of ART in both neuroasymptomatic individuals and in those with HAD. A small proportion 12%) of neuroasymptomatic individuals had elevated albumin ratios before initiation of ART, as opposed to 80% of the individuals with HAD. This is in the same range as in one of our previous cross-sectional studies n = 61, where % of neur studies , 28. How studies , 20. The studies –31. In o studies . The remSince the majority of the neuroasymptomatic individuals we examined had albumin ratios within the normal range, it is not surprising that the ratios remained fairly stable over time after initiation of ART. The neuroasymptomatic individuals whose BBB was impaired at baseline were similar in immune status and other baseline data. There was a slight, although not significant, difference in age: a median age of 40 years in those with elevated albumin ratios compared to 38 years for those with a normal albumin ratio, which would not explain the discrepancy between baseline levels of albumin ratios. The duration of HIV disease could not be assessed, this may be a cause for the discrepancy. Another possible explanation may be that the BBB impairment is not related to the HIV-infection. We have previously found that neuroasymptomatic HIV-infected individuals have albumin ratios in the same range in as those of HIV negative controls , 30, andIn agreement with the findings of earlier research, the neuroasymptomatic individuals in our study had a significant reduction of CSF viral load , 36 and The albumin ratio in individuals with HAD also declined, but was not statistically significant until the second time point, at 3 years, indicating that improvement of the BBB takes longer in individuals with HAD. Viral replication in individuals with HAD primarily takes place in cells in brain tissue or in tissue macrophages rather than in CSF, as for neuroasymptomatic individuals. This may be reflected in the slower reduction of HIV RNA after ART initiation. Nevertheless, a majority (75%) of those with elevated albumin ratios at baseline normalized this following the initiation of ART.n = 415) [An intact BBB is vital to maintain homeostasis in the CNS and protect the brain from peripheral, potentially neurotoxic, substances. It is composed of a semipermeable layer of a highly specialized capillary bed that surrounds the CNS. The endothelial cells are fused together with tight junctions . Viral pn = 415) , 3, 6. ACSF/plasma albumin ratio is a reliable marker of BBB impairment , 27, andOur study has several limitations. First, the sample size was fairly small, especially for the HAD cohort. As HAD is rare nowadays it is difficult to include many such individuals, although, to the best of our knowledge, ours is at present the largest study. Second, the number of samples during follow-up differs considerably between individuals, some having been followed for many years, and others only a few months. Also, some individuals had a very short interval before their first sample was taken, whereas others had been on ART for several years before their first follow-up sample. Due to a lack of some data in individuals with HAD, important biomarkers, such as IgG index, CSF β2-microglubulin, and CSF NfL, could not be analyzed during follow-up. An additional consequence of the currently rare HAD diagnosis is that data had to be collected over a very long period of time, during which ART has continued to be developed, something which also might have influenced results.In conclusion, albumin ratios declined significantly after initiation of ART in neuroasymptomatic HIV-infected individuals, as well as in patients with HAD, although the decline took longer for the latter group, indicating that healing may be slower in individuals with a more pronounced BBB injury. This would confirm the central role of BBB in the process of neuroinflammation and neuronal injury.


Uveal melanoma (UM) is a rare but aggressive cancer, which is the most common primary intraocular malignancy in adults. We aimed to develop and validate a competing risk nomogram to predict cancer-specific survival (CSS) of patients with UM, as well as compare its prognostic value with that of the American Joint Committee on Cancer (AJCC) staging system.Data of patients diagnosed with UM from 2010 to 2015 were identified from the Surveillance, Epidemiology, and End Results (SEER) database. We extracted and integrated significant prognostic factors based on competing risk regression to build a nomogram. The nomogram with an online prediction version was also created. The performance of the nomogram was evaluated using Harrell’s concordance index (C-index) and calibration plots. Receiver operating characteristic (ROC) curve was carried out to estimate clinical applicability of the model. Improvements in the predictive accuracy of our new model compared with AJCC staging system were estimated by calculating the relative integrated discrimination improvement (IDI) and the net reclassification improvement (NRI).P = 0.076) and − 0.045 (P = 0.004), respectively.A total of 839 eligible patients with primary UM were randomly assigned to a training cohort  and a validation cohort . Age, histological type, T stage and M stage were independent prognostic factors to predict CSS of UM and were incorporated in the nomogram. The calibration plots indicated that the 3- and 5-year CSS probabilities were consistent between the nomogram prediction and the actual observation. The C-index for this model was 0.778 (95% CI:0.756–0.800) and 0.786 (95% CI: 0.749–0.816) in the training cohort and validation cohort. Areas under the curve (AUCs) were 0.814, 0.771, and 0.792 in the training cohort, 0.788, 0.781 and 0.804 in the validation cohort, respectively. The NRI value in AJCC staging system was − 0.153 (95% CI -0.29 – − 0.041) for 3 years of follow-up and − 0.276 (95% CI -0.415 – − 0.132) for 5 years of follow-up. The IDI values for 3 and 5 years of follow-up in the AJCC staging system were − 0.021 (We have developed and validated a competing risk nomogram to reliably predict cancer-specific survival of patients with UM. This convenient tool may be useful for evaluating cancer-specific prognosis.The online version contains supplementary material available at 10.1186/s12886-021-01968-6. Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, with the mean age-adjusted incidence of 5.1 per million in the USA . UM is aAlthough there have been a few articles reporting the incidence, treatment and survival of UM, the clinicopathological characteristics and prognosis of UM still need further study because of its low prevalence rate and limited cases in previous studies. In addition, no ideal prediction model for prognosis of UM was yet identified, especially for the cancer-specific survival (CSS). A nomogram is a convenient tool to predict and quantify the probability of a patient developing a certain clinical event, which was valuable for clinical decision making and risk stratification. On the other hand, the staging system for UM was changed to American Joint Committee on Cancer (AJCC) seventh edition since 2010, incorporating significant changes into the T-staging , 6. TherThe SEER database consists of 18 population-based registries covering nearly 28% of the US population. SEER*Stat software version 8.3.6 was used to select patients from the Incidence-SEER 18 Regs Research database based on our application on November, 2020 to build the cohort. The inclusion criteria were as follows: 1) diagnosed from 2010 to 2015; 2) pathologically confirmed UM; 3) the International Classification of Diseases for Oncology-3 (ICD-3) histology code (morphology code 8720–8790); 4) ICD-O-3 site code of C69.3 (choroid) and C69.4 (ciliary body and iris); 5) only one malignant primary tumor of UM; 6) patients with active follow up and age of 18 or more than 18. We excluded patients with incomplete survival data or survival time of fewer than 1 month, unknown AJCC stage (according to the 7th edition of AJCC TNM classification), unknown laterality and race. The flow chart of patient selection was indicated in Fig. Several variables were extracted: baseline demographics , tumor features , therapy , and survival variables . CSS was the study endpoint and it was defined as the time from diagnosis to death attributed to uveal melanoma.n = 588) and validation cohort (n = 251), respectively, which was consistent with the assignment in some previous studies [In pursuit of full use of our data for constructing the predictive model, as well as keeping a considerable number of patients for its validation, we randomly allocated 70 and 30% of the enrolled eligible patients to the training cohort n = 58 and valThe optimal cutoff values for age range were determined using the X-tile software in survival analyses. Patient characteristics were analyzed by descriptive methods, with standard summary statistics including mean (S.D.), median, interquartile range (IQR), and proportions. We performed the Student’s t test for differences for continuous, normally distributed data; continuous, non-normally distributed data were analyzed by the Mann-Whitney test. Categorical variables were processed by X2 or Fisher’s exact tests.P-value < .05 identified in univariable analyses to develop multivariable regression models. Competing-risk regression was used to estimate the subhazard ratio (SHR) and evaluate the association between variables and risk of CCS. Death due to other causes was considered a competing event [Fine and Gray’s competing risks regression model was applied to identify predictors of CSS. We incorporated factors with ng event . Cumulatng event .A nomogram was constructed based on the result of the multivariable analysis. The nomogram was subjected to 500 bootstrap resamples for internal validation with the training cohort and external validation with the validation cohort. Performance of the nomogram to predict CSS was evaluated with discrimination and calibration. The Harrell’s concordance index (C-index) was calculated to measure the discrimination ability , which rP < 0.05 was considered to be statistically significant.We constructed and validated the nomogram by RStudio software (V. 1.2.5001). R project’s mstate package and rms package were applied to build a nomogram, and risk Regression package was used to evaluate the performance of the nomogram. The nomogram with an online prediction version was created by DynNom package. Besides, we performed all statistical analyses by Stata/SE 15.0 , except for the development and validation of nomogram. All statistical tests were two-tailed. A total of 839 eligible patients with primary UM were identified from the SEER database and included in the analysis. They were randomly assigned to a training cohort  and a validation cohort 251, 30%). The demographic, clinicopathological characteristics and treatment information of each cohort were shown in Table , 30%. ThThe characteristics did not significantly differ between the training and validation cohorts. The follow-up time from diagnosis to CSS was 32 (IQR: 20–51) and 32 (IQR: 18–53) months in the two groups, respectively. In the training cohort, there were 104 cancer-specific deaths (17.7%); in the validation cohorts, 55 cancer-specific deaths (21.9%) were identified.P = 0.001), primary site (P = 0.026), histological type (P < 0.001), T stage (P < 0.001), N stage (P < 0.001), M stage (P < 0.001), surgery (P < 0.001), radiotherapy (P < 0.001) and chemotherapy (P < 0.001) were found to be significant prognostic factors. Multivariate analysis revealed that age (P = 0.003), histological type (P = 0.006), T stage (P < 0.001) and M stage (P < 0.001) were independent prognostic factors to predict CSS of UM.To identify the prognostic factors associated with CSS in the training cohort, we used the univariate and multivariate analysis based on competing-risk models Table . In univAll independent prognostic factors were included to develop a nomogram to calculate the 3-year and 5-year CSS probabilities Fig. . From thhttps://zengqiaozhu123.shinyapps.io/UM_prognosis/.Our nomogram was tested by 500 bootstrap resamples for the internal validation with the training cohort, as well as the external validation with the validation cohort. Calibration plot for predicting 3- and 5-year CSS of UM demonstrated a good agreement between the predicted and actual CSS probabilities in both the internal and external validations Fig. . The nomn = 388, 66.0%, score < 108) and high risk  groups. Distinct cumulative incidence curves of UM cancer-specific mortality were depicted in the training and validation cohorts . For radiation, there was no difference in cancer specific mortality between radiation and no radiation in the high-risk or low-risk groups. In terms of chemotherapy, there was also no difference in cancer specific mortality between chemotherapy and no chemotherapy in the high-risk or low-risk groups, indicating that the nomogram appears to be able to differentiate patients with a different risk. In addition, we compared the cancer specific mortality of patients with different AJCC stage of 2 and 3 in different risk groups. In low-risk or high-risk group, there was no difference in mortality between patients with AJCC stage 2 and 3; while high-risk group presented with higher cancer specific mortality than low-risk group regardless of the AJCC stage  and − 0.045 (P = 0.004), respectively.The NRI value in AJCC staging system was − 0.153 (95% CI -0.29 – − 0.041) for 3 years of follow-up and − 0.276 (95% CI -0.415 – − 0.132) for 5 years of follow-up. These results indicate that our new model exhibited markedly superior predictive performance compared to the AJCC model. Similarly, the IDI values for 3 and 5 years of follow-up in the AJCC staging system were − 0.021 , histopathological features , cytogenetic features  and etc. . Xu et aBased on our nomogram, we developed a risk classification system that excellently classified patients with UM into high- and low-risk of cancer-specific death. In low-risk group, surgery could statistically reduce the mortality, suggesting the significant positive impact of surgery on patients with low risk. For radiation/chemotherapy, no difference in cancer specific mortality was found between radiation/chemotherapy and no radiation/chemotherapy whether in the high-risk or low-risk groups, indicating that patients were categorized into distinct risk groups regardless of the treatment. With limited information on detailed treatment from SEER database, we should cautiously interpret the influence of different therapies on prognosis. Management of UM mostly depends on the site, size of tumor and local extension , TreatmeRegardless of the AJCC stage, high-risk group presented with higher cancer specific mortality than low-risk group, indicating that our model appears to be able to efficiently differentiate patients with a different risk. Moreover, we introduced two indexes of NRI and IDI. NRI reflects how the nomogram reclassifies the risk probabilities better than the AJCC staging system, and IDI indicates the improvement in the ability of the nomogram to distinguish between AJCC stages. Compared with AJCC staging system, our new model exhibited markedly superior predictive performance. Considering our model couldn’t incorporate all prognostic factors for clinical decision, more clinical researches were needed to validate its applicability.Meanwhile, there are several limitations in our study. First, the retrospective nature of SEER-based study may bring about biases. Some crucial data, including lymphovascular invasion, detailed information of the tumor, detailed treatment approaches, some relevant molecular or cytogenetic factors and information of the 8th edition of AJCC staging system were not available, restricting further analysis. Cytogenetic studies are currently at the forefront of uveal melanoma management. In our clinical practice, it is not easy to obtain cytogenetic data, which requires more investment and support. We will take this indicator and more other factors into consideration in our further research, to construct a more advanced prediction model. Additionally, we could not neglect the geographical bias of the nonuniform distribution of the SEER registries, including the race heterogeneity. Second, because of the low disease prevalence, the cohort was relatively small. Larger sample sizes are necessary for further independent validation and prognostic stratification analysis. Third, we only included patients with complete information, which would have introduced selection bias. Finally, although our nomogram and risk classification system were constructed and validated in two independent subgroups from the SEER database, external validation with other populations is still warranted.In conclusion, we have developed and validated a competing risk nomogram to reliably predict cancer-specific survival of patients with UM, of which the prognostic value was better than that of the AJCC staging system alone. This convenient tool may be useful for evaluating cancer-specific prognosis, identifying patients with high-risk of cancer-specific death and clinical decision making. Future external validation and larger prospective studies are still required.Additional file 1: Supplementary Table 1. The compare and contrast table of different studies about the UM prognosis based on SEER database.


Nephronophthisis (NPHP) is a chronic tubular interstitial disorder that exhibits an autosomal recessive genetic form and causes progressive renal failure in children. Patients with NPHP rarely show urinary abnormalities, edema, or hypertension. Thus, NPHP is often detected only when renal failure becomes advanced. NPHP can be divided into three types based on the age of end-stage renal failure, i.e., infant type (approximately 5 years old), juvenile type (approximately 13–14 years old), and adolescent type (approximately 19 years old). Here, we report a case of NPHP diagnosed by genetic analysis at 26 years of age with atypical histological abnormalities.2). Urine tests indicated low specific gravity of urine, but not proteinuria or microscopic hematuria. Urinary β2-microglobulin was high (805 μg/L), and renal biopsy was performed for definitive diagnosis. Histological findings showed no significant findings in glomeruli. However, moderate fibrosis was observed in the interstitial area, and moderate atrophy was observed in the tubules. There were no significant findings in immunofluorescence analysis, and no electron dense deposits were detected by electron microscopy. Although cyst-like expansion of the tubules was unclear, tubular atrophy was dominantly found in the distal tubule by cytokeratin 7 staining. Genetic analysis of the NPHP1 gene showed complete deletion of this gene, leading to a definitive diagnosis of NPHP.A 26-year-old woman showed no growth disorders or urinary abnormalities in annual school physical examinations. However, at a check-up at 26 years old, she exhibited renal dysfunction (eGFR 26 mL/min/1.73 mNPHP1 gene, which is responsible for the onset of NPHP. In patients with renal failure with tubular interstitial disease dominantly in the distal tubules, it is necessary to discriminate NPHP, even in adult cases.NPHP is not merely a pediatric disease and is relatively high incidence in patients with adult onset end-stage of renal disease. In this case, typical histological abnormalities, such as cyst-like expansion of the tubular lesion, were not observed, and diagnosis was achieved by genetic analysis of the  Nephronophthisis (NPHP) is a chronic tubular interstitial injuries and is the most common genetic causes of renal failure in children and young adults . The estNPHP1, which is identified in 20% of patients with NPHP harboring NPHP gene mutations [NPHP is divided into three types based on the onset of ESRD, viz., infant type (approximately 5 years old), juvenile type (approximately 13–14 years old), and adolescent type (approximately 19 years old) . It is nutations , 8. OverIn the current report, we describe a case of NPHP diagnosed by genetic analysis at 26 years of age in a patient with atypical histological abnormalities.2, and urine tests indicated low specific gravity of urine (1.004); however, proteinuria and microscopic hematuria were not detected. Urinary β2-microglobulin (β2-MG) was high . The estimated glomerular filtration rate (eGFR) was 26 mL/min/1.73 mHistological findings showed global sclerotic lesions in approximately half of glomeruli Fig. a. ModeraNPHP1 gene, the most common gene responsible for NPHP. No exons of NPHP1  were amplified  being the most common gene [NPHP is the most common genetic cause of kidney failure in children and young adults , 10, witmon gene , 16, 17.mon gene . Genome-mon gene . Thus, Nmon gene , 20.The findings of this case report emphasize the importance of CD10 and CK7 staining in cases in which the cause of renal dysfunction is unclear. Irregular splitting TBM in tubules is an important finding suggesting autosomal dominant tubulointerstitial kidney disease . In patiIn summary, NPHP often progresses to ESRD at an average age of 13–14 years old. However, NPHP is not merely a pediatric disease and is relatively high incidence in patients with adult onset ESRD. Even in cases without typical histological abnormalities, such as cyst-like expansion of tubular lesions, differential diagnosis of NPHP is critical. Thus, wider application of genetic testing is recommended to patients with unidentified ESRD.


Endometrial cancer (EC) is a common tumor of the genital tract that affects the female reproductive system but with only limited treatment options. We aimed to discover new prognostic biomarkers for EC.  We used mRNA-seq data to detect differentially expressed genes (DEGs) between EC and control tissues. Detailed clinicopathological information was collected, and changes in the mRNA and protein levels of hub DEGs were analyzed in EC. Copy number variation (CNV) was also evaluated for its association with the pathogenesis of EC. Gene set enrichment analysis (GSEA) was conducted to enrich significant pathways driven by the hub genes. Cox regression analysis was used to select variables to create a nomogram. The nomogram was calibrated by applying the concordance index (C-index), and net benefits of the nomogram at different threshold probabilities were quantified using decision curve analysis (DCA).  Differential expression analysis identified 24 DEGs as potential risk factors for EC. Survival analysis revealed that TPX2 expression was related to worsening overall survival in patients with advanced EC. A high CNV was associated with the overexpression of TPX2; this suggested that modifications in the cell-cycle pathway might be crucial in the advancement of EC. Moreover, an individualized nomogram was developed for TPX2 incorporating clinical factors; this was also evaluated for its ability to predict EC. Calibration and DCA analyses confirmed the robustness and clinical usefulness of the nomogram.  We offer novel insights into the pathogenesis and molecular mechanisms of EC. The overexpression of TPX2 was related to a poorer prognosis and could serve as a biomarker for predicting prognostic outcomes in EC patients. Endometrial cancer (EC) is the sixth most frequent malignancy among women worldwide . In the via the PI3K/AKT/mTOR pathway [The targeting protein for Xenopus kinesin-like protein 2 (TPX2) is a key factor that ensures the correct assembly of the mitotic spindle and is located on chromosome 20q11.1 in humans . Althoug pathway . HoweverHere, we explored the effects of TPX2 expression on overall survival (OS) in EC patients and further investigated the molecular mechanisms underlying its differential expression. Following univariate and multivariate COX regression analyses, we constructed a TPX2 nomogram with independent prognostic factors, effectively predicting 1-, 3-, and 5-year overall survival (OS) in patients with EC. In summary, TPX2 is associated with the pathogenesis of EC and serves as a marker for the prognostic evaluation of EC.https://www.ncbi.nlm.nih.gov/gds/) to screen for candidate genes. GSE63678 featured 7 EC and 5 normal endometrial tissues. GSE17025 contained 91 EC and 12 normal endometrial tissues. Subsequently, the clinical data and expression profiles of the EC patients  were retrieved from The Cancer Genome Atlas (TCGA) database (https://cancergenome.nih.gov) so that we could evaluate prognostic markers.We retrieved messenger RNA expression datasets (GSE63678 and GSE17025) from the Gene Expression Omnibus (GEO) database . The median ages of patients with malignant and normal endometrial samples were 57.4 (41–75) and 54.9 (31–80) years, respectively. The patients had not received radiotherapy, chemotherapy, or hormone therapy, and all patients provided written and informed consent. Experiments were approved by the Clinical Research Ethics Committee of the Benxi Central Hospital of China Medical University .https://www.ncbi.nlm.nih.gov/geo/geo2r/) was used to screen for DEGs in both the GSE63678 and GSE17025 datasets. Differential expression analyses of mRNAs in EC and control tissues from the TCGA datasets were conducted using the Bioconductor Linear Model for Microarray Analysis (LIMMA) R package. Genes with a |log2(fold-change)| of >2.0 and p < 0.05 were considered as potential DEGs and were subjected to further analysis. The Venny v2.1 web-based tool (https://bioinfogp.cnb.csic.es/tools/venny/index.html) was utilized to identify candidate DEGs among the three datasets.The GEO2R tool (n = 552) were assigned to low- and high-expression groups based on the cutoff point . Kaplan–Meier survival curves were plotted using the survival R package. To further evaluate whether the candidate DEGs can serve as prognostic factors for EC, we performed univariate and multivariate Cox regression analyses. The mRNA expression levels of each DEG were verified with Oncomine (https://www.oncomine.org/) microarray data sheets according to the fold-change and threshold p value of 2 and 1 × 10−4, respectively.The TCGA-EC samples . n = 17) for differential expression analysis of hub genes. The experimental methodology used here was previously described by Lei et al. [Immunohistochemical staining was performed using tissue microarrays  containing EC and paracancerous tissue samples  was used to retrieve TCGA-EC copy number variation (CNV) data. Genes that were fully located in the significantly aberrant CNV regions were then identified by alignment with the genome.The Affymetrix SNP 6.0 platform on the Genomic Data Commons (GDC) Cancer Browser  were randomly divided into a training group (n = 276) and a verification group (n = 276). There were no significant differences between the training and verification groups in terms of age, tumor grade, FIGO stage, or histological type (Supplementary n = 552), we performed survival analysis to investigate the relationship between the mRNA expression of each hub gene and its corresponding CNV and clinical outcomes. Next, univariate and multivariate Cox regression analyses were conducted to identify whether the hub gene was independent risk factor.Prognostic information and clinicopathological data, such as age, tumor grade, Fédération Internationale de Gynécologie et d'Obstétrique (FIGO) stage, and the histological types of 552 patients were obtained from the TCGA-EC cohort. The TCGA-EC samples , which can range from 0.5 (no discrimination) to 1 (perfect discrimination). Furthermore, we used decision curve analysis (DCA) to compare a nomogram that included all independent prognostic factors with only one independent prognostic factor . The DCAt-test was applied to compare the mean values between the groups, and all data were checked for normal distribution and homogeneity of variance using the Shapiro–Wilk test and the Levene test, respectively. Fisher's exact test was employed to determine the association between hub gene expression and clinicopathological characteristics in TMA-EC samples. The optimal cutoff age for patients with TCGA-EC was calculated with X-tile software v3.6.1 according to the survival status [p < 0.05.Perl scripting tool v5.26.3, R software v3.5.3, and R Studio v1.1.463 were employed for statistical analysis. The Student's l status . The effp = 6.07E − 13) and testis-specific Y-encoded-like protein 5 (TSPYL5)  were independent risk factors of EC pathogenesis (p < 0.001) : 1.035; 95% confidence interval (CI): 1.025-1.045; ogenesis . Oncominogenesis . Furtherogenesis . Moreove< 0.001) . These f< 0.001) . Compare< 0.001) .p < 0.001), there was only an effect on OS when the mRNA expression of TPX2 was increased in patients with advanced stage III-IV EC , age , and FIGO stage  were independent risk factors and had better prognostic value  in TPX2 that confer increased susceptibility to EC need to be analyzed further. Our research group has been collecting serum samples from EC patients admitted to the Obstetrics and Gynecology Department of Benxi Central Hospital so that we can screen for high-risk SNPs. In addition, more cytological studies are warranted to investigate the roles of TPX2 in the pathogenesis of EC and to verify the association between TPX2 and EC progression.In summary, TPX2 overexpression was considerably associated with a poorer prognosis in patients with EC. CNV alterations in TPX2 might be a potential mechanism for its overexpression during the development and progression of EC. TPX2 can serve as a prognostic biomarker for predicting OS in EC patients and may facilitate the development of novel gene-targeted therapy for this disease.


To explore the usefulness of analyzing histograms and textures of apparent diffusion coefficient (ADC) maps and T2-weighted (T2W) images to differentiate prostatic cancer (PCa) from benign prostatic hyperplasia (BPH) using histopathology as the reference.Ninety patients with PCa and 112 patients with BPH were included in this retrospective study. Differences in whole-lesion histograms and texture parameters of ADC maps and T2W images between PCa and BPH patients were evaluated using the independent samples t-test. The diagnostic performance of ADC maps and T2W images in being able to differentiate PCa from BPH was assessed using receiver operating characteristic (ROC) curves.th, and 95th percentiles of ADC values in images from PCa patients were significantly lower than those from BPH patients (p < 0.05). Significant differences were observed in the means, standard deviations, medians, kurtosis, skewness, and 5th percentile values of T2W image between PCa and BPH patients (p < 0.05). The ADC5th showed the largest AUC (0.906) with a sensitivity of 83.3 % and specificity of 89.3 %. The diagnostic performance of the T2W image histogram and texture analysis was moderate and had the largest AUC of 0.634 for T2WKurtosis with a sensitivity and specificity of 48.9% and 79.5 %, respectively. The diagnostic performance of the combined ADC5th & T2WKurtosis parameters was also similar to that of the ADC5th & ADCDiff−Variance.The mean, median, 5Histogram and texture parameters derived from the ADC maps and T2W images for entire prostatic lesions could be used as imaging biomarkers to differentiate PCa and BPH biologic characteristics, however, histogram parameters outperformed texture parameters in the diagnostic performance. Prostate cancer (PCa) is the second most commonly diagnosed cancer in men worldwide . PatientWith the increasing application of multi-parametric magnetic resonance imaging (mpMRI), more focal PCa tumors can be detected and accurately localized, making early and precise PCa therapies possible . Diffusith percentile ADC, average ADC, T2-weighted skewness, and Ktrans [Image texture analysis can be used to estimate the heterogeneity of image signals by quantifying the roughness and regularity of grayscale pixel value spatial distributions in normal and pathologic tissues; the macroscopic heterogeneity of images might reflect microscopic heterogeneity at the level of histopathology , 13. Sevd Ktrans . HoweverTherefore, the study aimed to explore the usefulness of ADC map and T2W image histogram and texture analyses to distinguish PCa from BPH using histopathology as the reference.This retrospective study was approved by the local institutional review board (NO. M20140149), and individual consent for this retrospective analysis was waived. Between March 2015 and July 2017, consecutive patients with pathologically proven PCa or BPH were enrolled in this study. Patients who met the following criteria were included in the present study: (1) had pathologically proven prostatic hyperplasia (systemic biopsy) or cancer (prostatectomy); (2) had prostatic MRI examinations performed; (3) the interval between prostatic biopsy/radical resection and MRI was less than 3 months; and (4) no history of other malignant tumors. The exclusion criteria were as follows: (1) histopathology of lesion biopsies were confirmed to be positive but were negative on MRI; (2) pretreatments were given to treat prostatic lesions, such as endocrine, chemotherapy, or radiotherapy; (3) poor image quality due to motion artifacts or severe susceptibility artifacts; and (4) incomplete imaging protocol, images of DWI or T2W were missed to perform histogram and texture analysis. Patient clinical characteristics were recorded, including age, PSA levels, lesion volumes, score of Prostate Imaging Reporting and Data System , and Gleason scores (in the case of lesions was confirmed to be PCa).2, matrix = 120 × 150, slices = 20, slice thickness = 4 mm, gap = 0 mm, acceleration factor = 2, b-values (number of averages) = 0 (1), 500(2), 800(4), 1000(5), 1500(6), 2000(8) s/mm2, diffusion gradients applied in three orthogonal directions, and acquisition times = 6 min 43 s. ADC maps were inline calculated using the mono-exponential model, S(b) = S(0) e− b*ADC, where S(b) is the signal intensity with a b-value > 0, and S(0) is the signal intensity with a b-value = 0. Parameters for axial T2-weighted turbo spin echo sequence were TR/TE = 5460/104 ms, FOV = 180 × 180 mm2, matrix = 384 × 384, slices = 24, slice thickness = 4 mm, gap = 0 mm, echo train length = 18, and acquisition time = 3 min 49 s.All imaging was performed on a 3 Tesla MRI system  using a standard 18-channel phased-array body coil and 32-channel integrated spine coil. The main parameters of axial DWI were repetition time/echo time (TR/TE) = 5100/89 ms, field of view (FOV) = 224 × 280 mmAll images were sent to a dedicated workstation for data processing and were independently assessed by two experienced radiologists (P.X. and Q.Y.) with 6 and 8 years of experience in pelvic radiology, respectively. The radiologists were blinded to the data and clinical information, and using a consensus, selected the largest lesions in patients with multicentric or multifocal tumors for further analysis.2, and ADC maps were loaded into the histogram and texture analysis software. (2) Region of interest (ROI) delineations were acquired. Foreground and background seed points were manually drawn inside and outside of lesions, respectively, on three reformatted planes of the DWI images. Then, they were automatically copied to ADC maps and T2W images. (3) Lesion segmentation. Segmentation of the whole lesion was performed based on seed points with a random walker algorithm [th and 95th percentiles, differential variance (diff-variance), differential entropy (diff-entropy), contrast, entropy, skewness, and kurtosis. Figure Whole-lesion histogram and texture analyses were performed on ADC maps and T2W images with the prototypic MR Multiparametric Analysis software  by the radiologists using the following steps: 1) Import data. T2W images, DWI images with b = 1500 s/mmlgorithm . Manual  Import dAll patients underwent transrectal ultrasound-guided biopsy or radical prostatectomy. Biopsies and prostatectomies were formalin-fixed and subjected to tissue sectioning. Tissue sections were stained with a hematoxylin & eosin stain and subjected to immunohistochemical analyses. A urologic pathologist with 8 years of experience, observed the histologic sections and determined lesion locations and boundaries. If the lesions were confirmed to be PCa, Gleason scores were determined according to the PCa grading guidelines formulated by the 2014 Consensus of the International Urological Pathology Association . For patst and 3rd quartile ranges) based on the normal distribution of the data, and the categorical variables are expressed as percentages. Differences in PI-RADS score, PSA levels, and lesion volumes between the PCa and BPH patients were compared using Mann-Whitney U tests. Statistical variable differences in age, histograms and texture analyses of the ADC maps and T2W images between PCa and BPH patients were evaluated using the independent-samples t-test. In addition, the diagnostic performance of histogram and texture parameters on ADC maps and T2W images in being able to differentiate PCa from BPH was assessed using receiver operating characteristic (ROC) curves, and the 95 % confidence interval (CI) for the area can be used to test the hypothesis that the theoretical area is 0.5. If the CI does not include the 0.5 value, then there is evidence that the laboratory test does have an ability to distinguish between the two groups [p-value < 0.05 was considered statistically significant.Statistical analyses were performed using SPSS software  and MedCalc . Quantitative variables are presented as the mean ± standard deviation or as the median , age , PSA concentrations  vs. 9.81  ng/ml, p < 0.001), and lesion volumes  vs. 1.05  cm3, p < 0.001)  were included in the final analysis. Ninety (44.5 %) patients were diagnosed with PCa, and 112 (55.5 %) were diagnosed with BPH. There were significant differences between PCa and BPH patients in median scores (quartile range) of PI-RADS  vs. 2 , th and 95th percentiles of ADC maps were significantly lower in PCa patients compared with those in BPH patients . For ADC maps, aside from kurtosis , standard deviation, diff-variance, diff-entropy, contrast, entropy, and skewness measures were significantly larger in PCa patients than in BPH patients . Significant differences in the means, standard deviations, medians, kurtosis and skewness values, and 5th percentiles of T2W images were found between PCa and BPH patients , while no significant differences were observed in the 5th percentile, diff-variance, diff-entropy, entropy or contrast parameters.The statistical results regarding differences in histogram and texture parameters of ADC maps and T2W images in patients with PCa and BPH are summarized in Table th percentile of the ADC maps showed the largest AUC (0.906) with a sensitivity and specificity of 83.3 and 89.3 %, respectively, while kurtosis had the lowest AUC (0.536) with a sensitivity and specificity of 53.3 and 57.1 %, respectively. Histogram and texture analysis workflow are shown with two representative PCa and BPH patients  & the kurtosis of T2W (T2WKurtosis) parameters was the same as that of the combined ADC5th & ADCDiff−Variance parameters, yielding AUCs of 0.906 , and sensitivities and specificities of 83.3 %, and 89.3 %, respectively. However, these combined parameters were not better than the ADC5th parameter alone in distinguishing PCa from BPH, which was also true of the T2W5th & T2WDiff − Variance combined parameters (Table Table ts Figs.  and 3. Rst AUC 0.36 with ath percentile of T2W images between PCa and BPH lesions.In the present study, we evaluated the whole-lesion histogram and texture analyses of ADC maps and T2W images to distinguish PCa and BPH using histopathologic diagnoses as the reference standard and found those parameters could serve as useful biologic characterizations of PCa. The results demonstrated that histogram and texture analyses of parameters from ADC and T2W images could be useful to differentiate PCa from BPH, however, histogram parameters outperformed texture parameters in the diagnostic performance. All histogram and texture parameters, except for kurtosis, were significantly different in ADC values between PCa and BPH patients. Significant differences were observed in the means, standard deviations, medians, kurtosis and skewness values, and the 5b values selected. Thus, the current guidelines do not recommend using a single quantitative ADC parameter to characterize lesions [DWI detects the Brownian motion of water molecules and closely reflects tumor microenvironments, such as tumor cell densities, water content, the amount of fibrous stroma, and cell membrane integrities . ADC map lesions . ADC val lesions , 35. A f lesions , 37.ADC map histogram and texture parameters showed good diagnostic capabilities in detecting and characterizing diseases and evaluating therapeutic responses. These parameters can determine the spatial variations of ADC values and provide additional information about tumor heterogeneity, which could better reflect tumor characteristics than simply averaging these differences with ROIs. In this study, we found that texture features extracted from ADC maps of prostatic MRI could be used as potential biomarkers to distinguish BPH from PCa tumors. Although this is a preliminary study, the imaging phenotype based on the whole-lesion histograms of MR multi-parametric maps might provide as a noninvasive tool to evaluate the biological characteristics and heterogeneity of PCa.T2WI has been used to show prostatic zonation anatomies to localize lesions, which has served as the key protocol for prostatic MRI since it was first described in the early 1980 s . Tan et th percentile Kapp values of diffusion kurtosis imaging, the 10th percentile ADC values correlated better with Gleason classifications, and was superior to other DWI parameters in being able to distinguish low-grade and high-grade tumors [Texture analysis has been used to diagnose, differentiate, and assesses the types and therapeutic effects of various tumors, including PCa. Studies have shown that histogram and texture analyses of ADCs contributed to the characterization of prostate tumors. Compared with the mean and median ADC values and 90e tumors , 44.th percentiles of ADC were the best predictor of chemotherapeutic responses in patients with metastatic ovarian cancer and primary peritoneal cancers [Limited whole-lesion histogram analysis has been used to evaluate PCa therapeutic responses . Kyriazi cancers . Xie et  cancers . Another cancers . Texture cancers . Meng et cancers .For patients with increased PSA levels or clinical indications of disease on digital rectal examinations, transrectal ultrasound-guided biopsies are currently the most accepted definitive diagnostic method; however, biopsy results have been markedly inconsistent with the histopathology of complete prostatic resections. In this study, we used whole-lesion histogram and texture analysis to overcome this issue.There were several limitations in this study. First, the sample size was relatively small and the diagnostic performance was not validated on an independent dataset in the current study. We will enlarge the sample size and validate it in the future study. Second, this study mainly focused on patients who had histopathological proved to be positive as well as had definitive lesions in MRI, which may introduce a selection bias for these patients. Third, the histopathologic results of most patients were obtained by transrectal ultrasound-guided biopsy; and therefore, a mismatch between the pathologic locations and delineated ROIs was present. In future studies, 3D MRI-guided biopsies might help to obtain accurate matching between MRIs and pathologic locations and improve the repeatability of prostate MR image interpretations using texture analysis results. Fourth, ROIs of the lesions were acquired on DWI images and transferred to ADC maps and T2W images. Finally, it was retrospective study and all cases came from a single center. Future studies will be prospective and have larger sample sizes. Multiple centers and MR vendors will also be assessed to confirm the current findings.In conclusion, parameters derived from whole-lesion histogram and texture analyses of ADC maps and T2W images could be used as imaging biomarkers to assess the biologic characteristics of PCa and BPH lesions, however, histogram parameters outperformed texture parameters in the diagnostic performance, which could help clinicians differentiate benign and malignant prostate nodules, providing efficient and accurate clinical decisions.