Obesity and type 2 diabetes are causally related, yet there is considerable heterogeneity in the consequences of both conditions and the mechanisms of action are poorly defined. Here we show a genetic-driven approach defining two obesity profiles that convey highly concordant and discordant diabetogenic effects. We annotate and then compare association signals for these profiles across clinical and molecular phenotypic layers. Key differences are identified in a wide range of traits, including cardiovascular mortality, fat distribution, liver metabolism, blood pressure, specific lipid fractions and blood levels of proteins involved in extracellular matrix remodelling. We find marginal differences in abundance of Bacteroidetes and Firmicutes bacteria in the gut. Instrumental analyses reveal prominent causal roles for waist-to-hip ratio, blood pressure and cholesterol content of high-density lipoprotein particles in the development of diabetes in obesity. We prioritize 17 genes from the discordant signature that convey protection against type 2 diabetes in obesity, which may represent logical targets for precision medicine approaches. Coral et al. characterize genetically determined discordance between obesity and type 2 diabetes, identifying discordant genes that may convey protection against type 2 diabetes in obesity. The prevalences of obesity and T2D have risen sharply over the past decades worldwide2, corresponding with a shift to sedentary lifestyles and poor diet3. Even though obesity and T2D often coincide, their relationship is complex and remains incompletely understood. Indeed, while more than 80% of people with T2D also have obesity, 10–30% of people with obesity appear metabolically healthy6. Conversely, metabolic abnormalities occur in ~30% of normal-weight individuals9. Likewise, despite weight loss improving glycaemic control in people with T2D10, when T2D occurs in people with normal weight, mortality rates are higher than those in people with overweight or obesity12. Here, we refer to these divergent features as ‘discordant diabesity’. We focus on this unusual phenotype because it helps leverage the independent roles of excess adiposity and T2D in life-threatening disease.Cardiometabolic diseases are the leading cause of death globally, with obesity and type 2 diabetes mellitus (T2D) accounting for a large proportion of this burden14. For instance, even when BMI is comparable, lean and fat mass distributions often vary from one person to the next15. Genetics has helped provide pathophysiological explanations for discordant diabesity, whereby, collectively, common variants affecting adipose distribution mimic monogenic syndromes such as familial lipodystrophies21. Expanding our knowledge of the phenotypic signature of discordant diabesity using the quantitative framework of genetics may help elucidate the mechanisms by which the broader health consequences of excess adiposity varies from one person to the next.To some extent, this discordance can be attributed to the imprecision with which body mass index (BMI), the conventional metric used to define obesity, characterizes adiposityHere, we characterize genetically determined discordant diabesity through a comparative analysis with its concordant counterpart . We used a range of machine learning methods to undertake phenome-wide scans to identify traits other than T2D that distinctively characterize these profiles. We concluded by undertaking robust causal inference analyses to determine the causal relationships underlying discordant diabesity with other features of health and disease.22 and T2D23 by cross-referencing publicly accessible genome-wide association study (GWAS) summary statistics, finding 67 relatively independent single nucleotide polymorphisms (SNPs) strongly associated with both conditions (P < 5 × 10−8). After alignment to the BMI-increasing allele, these variants were labelled as ‘concordant’ (48 SNPs) or ‘discordant’ (19 SNPs) according to the positive or negative sign of their coefficients for T2D, respectively : −0.001, 0.004), βD = −0.008 s.d. units per allele , pδ = 1.39 × 10−4). We also found differences in risk of coronary heart disease (CHD) and stroke, which were lower in the discordant compared to the concordant profile c = 1.01 per allele , ORD = 0.98 per allele , pδ = 1.3 × 10−6). The levels of biomarkers of liver function such as gamma-glutamyl transferase (GGT) and alanine aminotransferase (ALT) enzymes were lower in the discordant relative to the concordant profile , βD = −0.011 , pδ = 2.07 × 10−6). SHBG, a protein also produced in the liver, was higher in the discordant as opposed to the concordant profile , βD = 0.013 s.d. units per allele , pδ = 1.94 × 10−8). Additionally, the discordant profile was associated with higher mean corpuscular volume , βD = 0.006 s.d. units per allele , pδ = 8.76 × 10−4) and lower levels of urate , βD = −0.005 s.d. units per allele , pδ = 3 × 10−6) compared to the concordant profile. The odds of receiving treatment with alendronate was higher in the discordant than in the concordant profile, a drug indicated for osteoporosis , ORD = 1.001 per allele , pδ = 3.26 × 10−6).Among the clinical phenotypes, we found that concordant and discordant diabesity profiles differed predominantly in cardiometabolic features including high-density lipoprotein (HDL) cholesterol, waist-to-hip ratio (WHR), waist circumference, and blood pressure Fig. . GeneralGCKR, associated with higher SHBG and lower liver enzymes , P = 7.5 × 10−199) and a second near TOMM40 associated with higher HDL , P = 3.7 × 10−107). In the discordant profile, the last variant to be aggregated to the clustering tree  is located near SLC2A2 and, in contrast to the overall discordant estimates, was associated with higher levels of AST and GGT , P = 2.7 × 10−22).Further exploration of the molecular features of the discordant and concordant profiles revealed that some variants used to characterize these profiles deviated from the overall pattern of trait association for their respective SNP set. Using single-linkage clustering on the SNP–trait matrix  coefficients for concordant and discordant profiles and assessed their association with multiple phenotypes , ORD = 1.02 per allele , pδ = 1.6 × 10−3) and that the concordant and discordant profiles were positively and negatively associated with diabetes diagnosis, respectively , ORD = 0.95 per allele , pδ = 3.2 × 10−49). Both scores were associated with increased odds of bariatric surgery , ORD = 1.03 per allele , pδ = 0.24). We found divergent associations in multiple diseases directly related to the main traits : ORC = 1.014 per allele , ORD = 0.99 per allele , pδ = 1.2 × 10−6). We also observed differences for other disease outcomes such as chronic kidney disease , ORD = 0.98 per allele , pδ = 2.9 × 10−6) and osteoarthrosis , ORD = 1.02 per allele , pδ = 0.012). Because both scores were also strongly associated with type 1 diabetes , ORD = 0.96, , pδ = 4.8 × 10−17), we repeated the analyses excluding individuals with T1D. This attenuated the differences between concordant and discordant profiles for a small subset of traits including diabetic retinopathy and end-stage chronic kidney disease.We sought replication of the discoveries outlined above in an independent European-ancestry cohort from BioVU, a de-identified collection of electronic health records and a linked biobank including inpatient and outpatient data from Vanderbilt University Medical Center (VUMC), a tertiary-care centre in Nashville, Tennessee, USApes Fig. . We firsn > 68,000) and African American  descent , s.d. difference per discordant allele: −0.06 , pδ = 1.5 × 10−39). We confirmed the difference between the two profiles in HDL and observed differences in the other two main lipid fractions , s.d. difference per discordant allele: −0.04 , pδ = 1.5 × 10−14). The findings for red blood cell phenotypes were also replicated, and additional differences were found in leucocyte count, urea, creatinine, phosphate, C-reactive protein and parathyroid hormone (PTH), all of which were higher in carriers of concordant SNPs. Of the liver enzymes, only ALT values were available for comparison, whose levels were weakly associated with the concordant but not the discordant GRS , s.d. difference per discordant allele: 0.06 , pδ = 0.2). In individuals of African American descent, significant differences were found in HbA1c, glucose and urea levels in urine.We also assessed the association of each GRS to multiple laboratory measurements in individuals of European , P = 4.54 × 10−9 and Supplementary Table We examined the relationship of GRSs to mortality owing to cardiovascular events in >337,000 participants of European descent from the UK Biobank (mean follow-up of 11.8 years). Around 35,000 deaths were reported, of which approximately 20% were related to cardiovascular events. The concordant GRS was associated with higher mortality (hazard ratio (HR) per allele: 1.01 ), whereas the discordant GRS was not , pδ = 0.02). However, when assessing each SNP separately, we observed that the concordant variant near βC = −0.008 s.d. units per allele , βD = 0.008 s.d. units per allele , pδ = 3.09 × 10−10). Discordant diabesity also correlated with lower levels of branched-chain amino acids and aromatic amino acids, whereas in concordant diabesity they tended to be higher , βD = −0.008 s.d. units per allele , pδ = 1.46 × 10−6).Of the metabolites available, those related to lipid subfractions were the strongest discriminators of concordant and discordant profiles Fig. . DiscordP < 0.05) within either the concordant or the discordant profiles , all of which were less abundant in discordant relative to concordant diabesity , βD = −0.004 s.d. units per allele , pδ = 0.004). The remaining taxa belonged to the phylum Firmicutes, most of them members of the obligately anaerobic class Clostridia, which tended to be more abundant in the discordant profile compared to the concordant profile , βD = 0.006 s.d. units per allele , pδ = 0.006). The family Lactobacillaceae was also lower in the discordant compared to the concordant profile , βD = −0.006 s.d. units per allele , pδ = 0.02).There were no differences between pooled concordant and discordant estimates for bacterial abundance in the gut that were statistically significant after false discovery rate (FDR) correction. Across ten taxa, several were nominally associated , which was higher in discordant relative to concordant diabesity , βD = 0.03 s.d. units per allele , pδ = 7.52 × 10−7; Fig. trans is likely to be weaker than that of those in the gene encoding the protein. Thus, we also searched for strong cis effects (P < 5 × 10−8) in the discordant profile. We found one association between the discordant variant near PPARG and metalloproteinase inhibitor 4 , P = 5 × 10−14).We found a significant difference between concordant and discordant estimates after FDR correction in a single protein: heparan sulfate 6-28 tool to compare the enrichment for tissues and biological pathways in each profile  for adipose tissue in the discordant profile, which was not found in the concordant profile. We also found significant enrichment for adrenal glands, ileum and kidney in the discordant but not in the concordant profile. Conversely, there was significant enrichment for endocrine tissue and retina in the concordant but not the discordant profile. Tissues for which there was significant enrichment in both profiles included pancreas and myometrium.We used the Data-driven Expression Prioritized Integration for Complex Traits (DEPICT)29. In eQTLGen30, we found significant associations of concordant SNPs with expression of 493 genes. Discordant SNPs were associated with 94 genes. Around 46% of all the associations found in GTEx were replicated in eQTLGen .We found 506 genes whose expression/splicing was significantly influenced by concordant SNPs and 76 which were influenced by discordant SNPs across multiple tissues in GTExP < 5 × 10−8). We followed the SMR & HEIDI approach31, which utilizes the strongest instrument for gene expression/splicing within the cis region of the corresponding gene (±500 Mb from the transcription start site) to calculate an estimate of the pleiotropic association across gene expression, BMI and T2D risk. This approach then determines if the association found reflects true pleiotropy rather than mere linkage by testing for heterogeneity of the estimates of SNPs in linkage disequilibrium (LD) with the lead SNP. We found 17 genes with robust expression signals for obesity and T2D whose directions of effect were in contrast  the proportion of median expression (in transcripts per million) across specific tissue types catalogued in GTEx and (ii) evidence of promoter/enhancer histone marks surrounding the genetic instruments, derived from multiple cell lines classified anatomically by the RoadMap Epigenomics Project33 that could be mapped to tissue samples in GTEx. For each gene, we sorted tissues where we found pleiotropic links according to its specificity score and presence of promoter/enhancer signals for the genetic instrument. This allowed us to prioritize potential main action sites of relevance to discordant diabesity, for example, LYPLAL1 in adipose tissue and JAZF1 in vasculature and pancreas, while confirming the widespread effects of SLC22A3 across multiple organs.To identify genes most likely involved in the molecular mechanisms leading to discordant diabesity, we chose genetic instruments for the 76 genes whose expression was influenced by discordant SNPs in the corresponding tissues in GTEx, and for the 94 genes in eQTLGen. A prerequisite for these instruments was that they are strongly associated with BMI  and metformin in both databases. SLC22A3 interacted with various quinoline derivatives , SarCNU , derivatives of the alpha blocker phenoxybenzamine, corticosterone and colchicine. There is also evidence of potent inhibition of GLUT2, the protein product of SLC2A2, by a specific class of pyrazolopyrimidines. SLC38A11, MAU2 and FBXO46 are classified in the ‘Tdark’ level of target development in the PHAROS database, composed of understudied targets, while the remaining genes fall under the ‘Tbio’ level, which includes targets with no known interactions yet satisfying other conditions, such as having functional annotations based on experimental evidence, repeated mentions in publications indexed in PubMed, and 50 or more available commercial antibodies.We performed a lookup of the genes identified previously in the comprehensive public access databases DGIdbP < 5 × 10−8) and decomposed these instruments into two groups based on their direction of effect on the trait of interest after alignment to the BMI-increasing allele. We then constructed two GRSs, one for each group of variants, and calculated the T2D risk conferred by each GRS using summary data from the DIAGRAM consortium; we focused on GRSs that confer protection from T2D.To quantify the potential impact of traits that emerged from the previous steps on offsetting the diabetogenic effect of obesity, we derived genetic instruments for each of these traits using SNPs that were also robustly associated with BMI (P = 6.71 × 10−5). Some traits in the clinical phenotypes required instruments to be in cis with the gene encoding the corresponding protein , to prevent confounding due to pleiotropy. We found two such instruments for ApoA1 and SHBG, respectively, which were not associated with T2D risk  in the cis region of the corresponding genes. We could only derive a valid instrument for TIMP4. Using the SMR & HEIDI method, we found a significant pleiotropic effect  of the A allele of rs1530559 ) with higher BMI and lower abundance of the order Bifidobacteriales. This variant was not associated with T2D risk (P = 0.76).No instruments for the microbial taxa where we found nominally significant differences reached the significance threshold required for BMI. Extending the exploration to other taxa revealed a single significant association (2), we focused on this subgroup in UK Biobank who localized to the top decile of one of the two profile GRSs37. Consistent with a binomial distribution, 18% of individuals with obesity were present in the two groups of extreme GRSs. The health characteristics of these individuals differed from all others with obesity (Supplementary Table P = 5.94 × 10−12). Conversely, individuals with obesity and an extreme discordant GRS had significantly lower HbA1c compared to the rest of individuals with obesity (P = 2.71 × 10−42). Persons with obesity at both extreme GRSs are also distinguished from the rest by the main clinical features identified in our previous analyses such as SBP, HDL and ALT. WHR did not adequately separate the concordant or discordant extreme GRS from the wider group of people with obesity. However, because the initial phenome scan revealed a gender-specific difference in WHR between concordant and discordant profiles, .To determine the relevance of concordant and discordant profiles in people with obesity  and Latin American (53%) ancestries, enriched for T2D cases (>80% had a prior diagnosis).We also sought replication of a finding from Pigeyre et al.21. Fourteen of the 19 discordant SNPs identified in our analysis (78%) are among or in LD with the 62 SNPs identified by Huang et al  were in the subcluster characterized mainly by higher BMI and lower fasting glucose and risk of T2D (enrichment P = 1.6 × 10−3) and 4  in the subcluster that conveyed an apparent protective effect on multiple cardiometabolic traits via peripheral adipose distribution . Four concordant variants  were also among the 62 SNPs described by Huang et al. As opposed to other variants in the concordant set, the variant in AKAP6 was associated with lower SBP , P = 1.25 × 10−4) and the variant near PPP1R3B-TNKS-MSRA was associated with higher HDL , P = 1.72 × 10−6). As we found and discussed in our analyses, TOMM40 and GCKR deviate from the concordant set owing to their favourable associations with lipids and liver enzymes that resemble the discordant set, a pattern that was also reported by Huang et al.Finally, we searched for the SNPs comprising the concordant and discordant profiles described above in the cluster analysis of discordant SNPs performed by Huang et al.Obesity conveys heterogenous effects in cardiometabolic health, making disease prevention and management challenging. Here we used genetics to deconstruct the obesity phenotype into concordant and discordant diabesity, with strikingly different health characteristics beyond diabetes and obesity. Through transcriptomic, metabolomic and metagenomic analyses, we identified biomarkers that shed light on mechanisms of action and may aid risk stratification. Further analyses identified potential targets for drug development and drug repurposing.41 and lower abdominal region around organs such as the liver, which might underlie the difference seen in biomarkers of liver failure42, play important and independent roles in genetically determined diabesity discordance.Obesity and T2D often coalesce, owing largely to the mediating effect of peripheral insulin resistance caused by excess adiposity. The trait discordances described here reflect mechanisms involved in uncoupling obesity risk from T2D risk, thereby exposing diabetes-independent pathways through which obesity affects disease risk, for example, through adipose distribution. It is likely that both a higher capacity to expand adipose tissue in the gluteo-femoral compartment44, affecting pancreas, muscle and adipose tissue45. For instance, capillary recruitment and permeability are key determinants of whole-body glucose uptake and glycaemic variation46.Another key phenotypic distinction between concordant and discordant profiles concerns blood pressure. Although T2D often causes vascular dysfunction, changes in the vascular bed may also precede metabolic perturbations through nutrient and hormonal flux47, possibly because they enhance glucose homeostasis in obesity by improving cross-talk between peripheral tissue and the liver48.Our findings relating to lipid metabolites support the use of more refined profiling of lipid subfractions to help determine risk in people with obesity. The cholesterol content of HDL particles and BCAA levels appear especially informative biomarkersDespite the contrasting health consequences of the two diabesity profiles, bariatric surgery was equally likely, which may predispose one group to health benefits following surgery, whereas the other may not benefit in this way.50. We found only one robust pleiotropic effect for discordant diabesity at TIMP4, which is proximal to PPARG, the likely causal gene. Moreover, PPARG activator medication inhibits matrix metalloproteinases52. TIMP4 has been associated with adipogenesis, possibly through its effect on the adipose tissue extracellular matrix in obesity53.We found a significant difference between concordant and discordant profiles in levels of HS6ST2, a protein expressed in brain, kidney and ovaries, which in animal knock-out models shows a strong association with increased body weight and insulin resistance, possibly owing to enhanced adipocyte differentiationSLC22A3, but also for other potential targets such as LYPLAL1, whose differential expression in both adipose tissue and adrenal glands appears linked to discordant diabesity. Moreover, three of the genes with pleiotropic links to T2D risk  interact with metformin. This suggests a potential effect of metformin in shifting individuals with obesity from a concordant to a discordant phenotype. SLC2A2 encodes GLUT2, which is part of the glucose sensor apparatus in pancreas and liver and is involved in intestinal glucose absorption in the gut54. Variants in this gene have been associated with preference for sugary foods55 and modified response to metformin57. SLC22A3 encodes OCT3, a protein widely expressed across tissues that aids adipocyte beiging58 and perivascular adipose tissue remodelling59. KCNJ11 encodes the Kir6.2 subunit of the ATP-sensitive potassium channel. As this is the target of sulphonylureas, this group of drugs may also harbor potential candidates for the phenotype shift to discordance in diabesity. The other ligands identified in the lookup may also constitute potential therapeutic agents to prevent cardiometabolic complications in obesity. For the rest of the genes, especially those in the ‘Tdark’ level in PHAROS, follow-up functional experiments in the tissues indicated by the lead genetic instruments and its corresponding epigenetic annotations are warranted.The colocalization analyses underscore the importance of tissue pleiotropy and tissue cross-talk in the molecular mechanisms of diabesity discordance. This is especially evident for TOMM40 increases T2D risk but, unlike other SNPs in the same profile, is associated with a better lipid profile and lower cardiovascular disease mortality. This gene and others in its proximity (APOC1 and APOE) have been consistently implicated in lipid metabolism60. In the discordant profile, the variant SLC2A2 conveys protection against T2D risk despite being associated with heavier weight and higher blood pressure, and worse liver function and dyslipidaemia. The opposite pattern was observed in the concordant variant in GCKR, which encodes a regulatory protein that inhibits glucokinase. This reflects disparate phenotypic effects of modulating the glucose sensor apparatus at different levels54. Deeper characterization of these mechanisms can further improve obesity stratification.Certain SNPs deviate from the overall association pattern of the profile within which they reside. In the concordant profile, the BMI-increasing allele of the variant near 61. Our results indicate higher Firmicutes and lower Bacteroidetes abundance in discordant diabesity, which may result in enhanced production of short-chain fatty acid species such as butyrate, which is involved in glucose-lowering and anti-inflammatory mechanisms62.Although no statistically robust differences were observed in gut microbiota between the two diabesity profiles, possibly owing to low statistical power, nominal differences emerged in taxa belonging to the Bacteroidetes and Firmicutes phyla, which together constitute 90% of the human intestinal flora21. Our phenome-wide approach consisted of leveraging the wealth of genetic associations harvested to date to dissect the phenotypic structure relevant for discordant diabesity, having three main advantages: (1) variables defining the differential phenotypic structure of each profile are selected a in data-driven manner across many phenotypic layers; (2) leveraging genetic data across multiple datasets enhances power and minimizes cohort-specific biases that would be anticipated if analyses were performed in a single cohort; and (3) although concordant and discordant diabesity profiles may be driven by molecular mechanisms that are independent of DNA variation, using germline DNA variants helps mitigate reverse causality and other sources of confounding that hamper the interpretation of associations for most other types of biological variation and phenotypes. An example of this is the analysis of epigenetic factors, which has led to identification of obesity sub-phenotypes even in the context of genetic homogeneity, as found in monozygotic twins that are discordant for adiposity traits63. However, these findings might be driven by variations in environmental exposures and behaviours that exist within and between twin pairs, as well as confounded by factors such as age, which differed between twin pairs in the reported analyses.Previous strategies to characterize the discordance between BMI and metabolic risk have been based on predefined sets of phenotypes traditionally linked with metabolic statusIn conclusion, obesity profiles with either diabetogenic or antidiabetogenic proclivities reveal distinctive aetiological subtypes, with key differences in fat distribution, blood pressure and cholesterol content in HDL particles. We identified 1 protein (TIMP4) and 17 genes potentially involved in the molecular mechanisms leading to diabesity discordance, involving pleiotropic effects across multiple tissues.24 and case, control and exclusion criteria are defined. Two codes on different visit days were required to instantiate a case for each phecode. The biobank was launched in 2007 and comprises excess blood samples that their donors had consented for use in biomedical research. Details of programme operations, ethical considerations, continuing oversight and patient engagement are published elsewhere25. DNA samples were analysed using genome-wide genotyping platforms including Illumina multi-ethnic genotyping array. After quality assessment, the genotype data were then imputed to the Haplotype Reference Consortium reference panel at the Michigan imputation server. Populations of African American and European descent were identified by projecting individuals onto the major principal-component space derived from 1000 Genomes reference panel.Collection of electronic health records in BioVU was established in 1990 and includes data on billing codes from the International Classification of Diseases, 9th and 10th editions (ICD-9 and ICD-10). Disease phenotypes (‘phecodes’) are derived from these billing codes as described previously64. It has collected comprehensive genetic and phenotypic information, biochemical assays and longitudinal health outcomes through health records, such as hospitalization and mortality. The genotypes were assayed using the UK Biobank Lung Exome Variant Evaluation and the Applied Biosystems UK Biobank Axiom Array, and imputed to the Haplotype Reference Consortium panel. Population structure was also assessed using principal-component analysis. We excluded individuals with inconsistency between their reported and genetic sex, had sex chromosome aneuploidy or were outliers for heterozygosity or missingness. Only individuals who were included in the calculation of genetic principal components were included, which ensures minimal genetic kinship with other participants.The UK Biobank is an ongoing prospective study of approximately 500,000 adults. Initial enrolment took place from 2006 to 2010 and included individuals aged 40–69 years across the United Kingdomr2 < 0.01 over a 500-kb window) to identify nearly independent SNPs that were strongly associated with both conditions (P < 5 × 10−8). The directions of the effect of these SNPs on T2D were consistent in a second independent set of GWAS summary statistics extracted from the FinnGen database65  > 1%). Insertions, deletions and potentially ambiguous palindromic SNPs  were excluded. Because both GWAS were conducted predominantly in populations of European descent, we used 1000 Genomes EUR reference panel for clumping :We extracted association data for concordant and discordant SNPs from a variety of sources. From the curated repository Open GWASquations  and 2)  66, whicWe aligned all the estimates from these scans to the BMI-increasing alleles, so that they represent phenotypic variations associated with higher BMI in both profiles.69 and 3,282 proteins in plasma39. Associations with several bacterial taxa in the gut were obtained from the MiBioGen consortium70. Association with expression and splicing of nearby genes in multiple tissue samples and in whole blood were obtained from data generated by GTEx and eQTLGen consortia, respectively.We also obtained data for 657 blood metabolitesβC) and discordant (βD) SNPs separately using a random-effects meta-analysis with the Paule–Mandel estimator of between-SNP variance τ2 . We then calculated their difference δ = |βC − βD| and computed its standard error as 73. We excluded from these analyses T2D traits  to this matrix, which attempted to classify SNPs in their correct category, and used the Boruta algorithm74 to identify which traits were relevant to distinguish discordant from concordant SNPs. Briefly, this algorithm creates randomly shuffled copies of all traits in the SNP–trait matrix, and then evaluates for each trait if its contribution to the accuracy of decision trees in the Random Forest is higher than its corresponding random set.We then compared the effects of discordant versus concordant SNPs for every trait in two stages: we first obtained the combined effect of concordant 24. We kept phecodes with at least 200 cases67 and identified those associated with either of the GRS coefficients and a significant difference between the estimated effects after a 5% FDR correction.Where In the UK Biobank, we examined the relationship of GRSs to mortality due to cardiovascular events in individuals followed up to the latest censor date (30th September 2021) using Cox regression. Primary cause of death was ascertained using ICD-10 codes reported in death certificates . We retained signals where we found evidence of true pleiotropy for both BMI and T2D.The SMR method consists of identifying for a protein or gene the strongest association signal, which is used as a genetic instrument to test for its pleiotropic effect on an outcome. The HEIDI method consists of calculating the heterogeneity in the estimates of SNPs in LD with the lead SNP used in SMR. A higher pThe scoring method to identify the most likely tissue of action assumes that if a genetic instrument for the expression of a gene in a certain tissue where it is highly expressed  is in or close (in LD) to a promoter/enhancer region in the same tissue, and this genetic instrument is also associated with an outcome, then it is likely that the pleiotropic association on the outcome is due to perturbation of gene activity in that tissue. Promoter/enhancer signals were obtained by querying the RoadMap Epigenomics Project through the ‘haploR’ package in R.The lookups in DGIdb and PHAROS were performed using the web-based tool. DGIdb assigns an interaction score to the drug–gene interactions, which is the result of combining publication count, source count, relative drug specificity and relative gene specificity. The PHAROS database classifies targets into four ‘Target Development Levels’, according to the evidence of drug interactions available: ‘Tdark’ contains understudied targets, ‘Tbio’ contains highly studied targets but without interaction with compounds, ‘Tchem’ includes targets that bind to small molecules, and ‘Tclin’ interact with approved drugs.76.All analyses were done using packages within the R environment (v4.1.2)Further information on research design is available in the Supplementary InformationSupplementary Figs. 1–6Reporting SummarySupplementary Tables 1–18


Cancer antigen 125 (CA125) is clinically used to monitor response to therapy in ovarian cancer and has been proposed for use in detecting ovarian cancer. This population-based study examines how demographic characteristics, gynecologic/reproductive history, chronic non-malignant medical conditions, history of non-ovarian cancer, lifestyle practices, and biomarkers of inflammation correlate with serum CA125 in both premenopausal and postmenopausal women without ovarian cancer across the United States.Participants were identified from the National Health and Nutrition Examination Survey 2001–2002. Linear and logistic regression models were applied.N = 1157), current smoking was associated with lower CA125 , and history of non-ovarian cancer was associated with higher CA125  by multivariable linear regression; both current smoking (odds ratio (OR) = 0.42, p = 0.043) and oral contraceptive pill (OCP) use of 5–10 years  were less likely to be associated with having CA125 level ≥ 35 U/ml by multivariable logistic regression. In postmenopausal women (N = 1116), coronary artery disease (CAD) history was associated with higher CA125  by multivariable linear regression; history of CAD , history of breastfeeding , and increased CRP level  were more likely to be associated with having CA125 level ≥ 35 U/ml by multivariable logistic regression.Higher CA125 levels were found to correlate with younger age, Non-Hispanic White race/ethnicity, and lower body mass index. In premenopausal women (Results suggest CA125 is lower in premenopausal women who are current smokers and OCP users of moderately longer duration but higher in those with non-ovarian cancer. CA125 is higher in those postmenopausal women with CAD, history of breastfeeding and elevated CRP level. These associations can inform clinical interpretation of individual patients’ CA125 levels.The online version contains supplementary material available at 10.1186/s12885-022-09637-7. MUC16 and recognized by the murine monoclonal antibody OC125 , p = 0.043). Compared to a short duration of OCP use (less than 2 years), OCP use of 5–10 years (N = 178) in premenopausal women was less likely to be associated with CA125 level ≥ 35 U/ml .Univariable logistic regressions showed that among premenopausal women, current smoking, OCP use for 2–5 years and for 5–10 years, CRP level, and history of endometriosis demonstrated possible associations (1) Table . Of noteN = 1116), alcohol consumption of more than two drinks per day, caffeine intake, CRP, ferritin level, history of hysterectomy, oophorectomy, endometriosis, fibroids, and CAD demonstrated possible associations with CA125 concentration (p < 0.25) via linear regression models (Table N = 58) have higher CA125 levels than those without this history .For postmenopausal women (N = 41) (p < 0.25) (Table N = 23 out of 556), and no postmenopausal women with endometriosis had CA125 level ≥ 35 U/ml, no crude logistic regression was performed with respect to alcohol consumption or endometriosis. After performing multivariable logistic regression, breastfeeding history, CRP, and CAD remained significant. History of breastfeeding in postmenopausal women (N = 561) was more likely to be associated with CA125 level ≥ 35 U/ml , as was history of CAD (N = 58) . Additionally, for a one-unit increase in CRP level, there was 41.10% of increase in the odds of having CA125 ≥ 35 U/ml .Univariable logistic regressions showed that among postmenopausal women, breastfeeding history, CRP level, CAD, and non-ovarian cancer demonstrated possible associations with CA125 levels ≥ 35 U/ml (5) Table . Of noteThis study analyzed associations between various demographic, clinical, and laboratory variables and CA125 levels in both premenopausal and postmenopausal women without history of OC in a large nationally representative sample. CA125 level has been reported to be higher in pre- versus postmenopausal women and decreases with advancing age , 26. Ourp = 0.05), which is consistent with the results of prior studies suggesting that hysterectomy history may be associated with lower CA125 level [N = 23 who had CA125 level ≥ 35 U/ml in postmenopausal subgroup) or the choice of cut-off value for CA125.Of the reproductive/gynecologic history variables examined, hysterectomy history was not significantly associated with CA125 levels in the multivariable analyses in postmenopausal women, though the crude linear regression did show an inverse relationship  and showed similar trend in postmenopausal women via both univariable linear regression  and logistic regression . This finding was consistent with previously reported positive correlation between bone mineral density and CA125 level in both pre- and postmenopausal women [. For postmenopausal women, Akinwunmi et al. had demonstrated that both osteoporosis and osteoarthritis were associated with lower levels of CA125, [N = 76 for osteoporosis and N = 225 for osteoarthritis in Akinwunmi’s study, versus N = 203 for osteoporosis and N = 175 for osteoarthritis in this study).In terms of bone health, osteoporosis was significantly associated with lower CA125 levels in premenopausal women by crude linear regression . This finding contradicts Akinwunmi et al’s study in which CA125 was reported to be higher in premenopausal women with CAD based on a very small number of CAD cases (N = 2 versus N = 10 in present study) [p = 0.047) and more likely to be associated with CA125 level ≥ 35 U/ml in multivariable logistic regression ; these findings are in agreement with prior literature [Regarding cardiovascular comorbidities, this study showed decreased levels of CA125 in premenopausal women with CAD in crude linear regression  were included in this study. As a protein produced by coelomic epithelial cells lining the pleura, peritoneum and ovaries in the setting of inflammatory stress, CA125 level was hypothesized to demonstrate positive association with inflammatory biomarkers , 38. TheN = 7, effect estimate = 0.63, p = 0.082) but did not reach any statistical significance in postmenopausal women .History of non-ovarian cancer was found to be associated with higher CA125 levels in premenopausal women. In postmenopausal women, those with non-ovarian cancer history tended toward a lower likelihood of having CA125 levels ≥ 35 U/ml. Funston et al. have reported in a UK population-based study that 12.3% of those with CA125 level ≥ 35 U/ml were diagnosed with a non-ovarian cancer, and non-ovarian cancer diagnosis was found in 20.4% of women with CA125 level ≥ 35 U/ml who were ≥ 50 years of age .The resuCurrent smoking was associated with lower CA125 level among premenopausal women in both multivariable linear and logistic regressions, consistent with previous findings , 18. SmoThis study has several unique strengths. First, study participants comprised a nationally representative sample from across the United States. Results from this data are more generalizable compared to results from control groups in previous studies, which were more restricted in terms of the ethnicities, ages and/or geographies represented. Furthermore, those study enrollees from long-term cancer screening or prevention clinical trials tended to be generally healthier compared to the general population . AnotherBRCA mutation status and family history of OC could not be assessed, although increased CA125 level has been associated with BRCA mutations and familial OC [We acknowledge several limitations of this study. The cross-sectional design did not allow for longitudinal measurements of CA125 and precluded interpretation of CA125 trends over time. Study participants provided only a snapshot of their clinical history and lifestyle practices at one particular point in time via self-report without further validation. The clinical data obtained regarding participants' medical conditions, especially with respect to the non-ovarian cancer diagnosis, may not have reflected the actual status of such medical conditions. Whether a participant was in remission, receiving treatment, or progressing at the moment when CA125 level was measured, and how long ago the treatment was given, cannot be confirmed. As such, this analysis was unable to eliminate the confounding effect related to the status of non-ovarian cancer diagnoses and other medical conditions, and the real associations might be underestimated. It is interesting to note, however, that Akinwunmi et al. previously found no evidence that treatments associated with a particular medical condition changed the association between the medical condition and CA125 level . Additioilial OC , 41–43. ilial OC . By perfIn summary, this is a large analysis of factors associated with CA125 levels among women without OC, and a broad population-level study of CA125 levels in both pre- and postmenopausal women across the United States. In premenopausal women, CA125 was found to be lower in current smokers and OCP users of moderately long duration, but higher in those with non-ovarian cancer. In postmenopausal women, CA125 was found to be higher in those with CAD, breastfeeding history, and elevated CRP level. Such associations should be evaluated and weighed in routine clinical interpretation of CA125 levels. Future studies examining underlying biologic and pathologic mechanisms of the observed associations are warranted. Studies with larger sample sizes are needed to improve statistical power, distill more robust findings and refine a strategy for CA125 interpretation in individual patients.Additional file 1.


Colorectal cancer (CRC) is the third most common cancer and the second leading cause of cancer‐related death worldwide. Alterations in proteins of the p53‐family are a common event in CRC. ΔNp73, a p53‐family member, shows oncogenic properties and its effectors are largely unknown. We performed an in‐depth proteomics characterization of transcriptional control by ∆Np73 of the secretome of human colon cancer cells and validated its clinical potential. The secretome was analyzed using high‐density antibody microarrays and stable isotopic metabolic labeling. Validation was performed by semiquantitative PCR, ELISA, dot‐blot and western blot analysis. Evaluation of selected effectors was carried out using 60 plasma samples from CRC patients, individuals carrying premalignant colorectal lesions and colonoscopy‐negative controls. In total, 51 dysregulated proteins were observed showing at least 1.5‐foldchange in expression. We found an important association between the overexpression of ∆Np73 and effectors related to lymphangiogenesis, vasculogenesis and metastasis, such as brain‐derived neurotrophic factor (BDNF) and the putative aminoacyl tRNA synthase complex‐interacting multifunctional protein 1 (EMAP‐II)–vascular endothelial growth factor C–vascular endothelial growth factor receptor 3 axis. We further demonstrated the usefulness of BDNF as a potential CRC biomarker able to discriminate between CRC patients and premalignant individuals from controls with high sensitivity and specificity. High‐density antibody microarrays and stable isotopic metabolic labeling allowed the identification of secreted dysregulated effectors by the ectopic ∆Np73 expression in HCT116 colorectal cancer (CRC) cells. After validating the dysregulation of the effectors by orthogonal techniques, an important association with lymphangiogenesis, vasculogenesis and CRC metastasis was found. Brain‐derived neurotrophic factor was validated by ELISA as potential biomarker of CRC. ACNacetonitrileATCCAmerican Type Culture CollectionAUCarea under the curveBDNFbrain‐derived neurotrophic factorCRCcolorectal cancerDMEMDulbecco's modified Eagle's mediumECMendothelial cell mediumEMAP‐IIendothelial‐monocyte‐activating polypeptide IIGFRgrowth factor reducedHLEChuman lymphatic endothelial cellsHUVEChuman umbilical vein endothelial cellsISCIIIInstituto de Salud Carlos IIIMSmass spectrometryMTT3‐‐2,5‐diphenyl‐2H‐tetrazolium bromideP/Spenicillin/streptomycinqPCRquantitative PCRROCreceiver operating characteristicRT‐PCRreverse transcriptase‐PCRSDHAsuccinate dehydrogenase complex subunit ASILACstable isotopic labeling with amino acids in cell culturesiRNAsmall interfering RNATrptryptophanVEGFCvascular endothelial growth factor CVEGFR3vascular endothelial growth factor receptor 3WBwestern blot1Colorectal cancer (CRC) is the second most common cancer in Europe and the second leading cause of cancer‐related death following lung cancer. CRC appears sporadically with only a small percentage of the cases due to inherited predisposition . Events p53 family member, TP73, is translated into different isoforms ‐l‐lysine and [12C6]‐l‐arginine or heavy [13C6]‐l‐lysine and [13C6]‐l‐arginine  supplemented with 10% dialyzed FBS, 100 units·mL−1 of penicillin/streptomycin at 37 °C in 5% CO2. At least eight duplications were performed to achieve > 98% incorporation of the heavy amino acids, which was determined as previously described . In. Inproteone Fig. .After normalization, by using a permutation‐based statistical test, we fixed a fold‐change of ≥ 1.5 (mean of two experiments) as significant for identification of dysregulated proteins , 15. In According to Gene Ontology analysis, among the top five enriched fractions, extracellular exosome and proteins from the extracellular space represented the 64.7% of the total number of quantified dysregulated proteins  and two proteins (APEX1 and CRABP2) described as transcription factors, not previously described to be altered by ∆Np73. We also found several proteins  whose dysregulation has been associated to the tumorigenesis process, prognosis of patients and resistance to treatment , 28, 29.To identify altered biological functions that might play a role in CRC, differentially secreted proteins due to ∆Np73 were analyzed using STRING  Fig. S2S2. UsingIn summary, ∆Np73 produces the dysregulation of cell adhesion proteins, transcription factors, receptors, and previously published proteins related to cancer or cancer metastasis, with most of them not previously identified as modulated by ∆Np73. All of them are producing important changes affecting cellular adhesion, cell growth, cell proliferation and cell death and survival, increasing the tumorigenesis and metastatic properties of CRC cells , 32.3.4We performed an initial validation of ∆Np73‐dysregulated proteins by PCR, WB, dot‐blot and ELISA Fig. . First, We next evaluated at protein level the dysregulation of BDNF, VEGFR3, EMAP‐II, and VEGFC by dot‐blot, ELISA and/or WB analyses Fig. . BDNF, V−/−‐∆Np73 and mock control cells were analyzed. First, ∆Np73 overexpression was confirmed in HCT116 p53−/−‐∆Np73 in comparison to mock control cells. Then, selected markers were analyzed by PCR. All of them  showed dysregulation at similar extents in HCT116 and HCT116 p53−/− cells upon ∆Np73 overexpression , and EMAP‐II , significantly discriminated patients from controls samples for BDNF and EMAP‐II   and subjects with premalignant lesions (n = 17), obtaining a potential correlation for BDNF  and for EMAP‐II (r = −0.813 for patients with cancer). On the other hand, values for VEGFC and VEGFR3 were not significant , levels in plasma for BDNF  calculating its individual sensitivity and specificity by means of ROC curves. Individual AUC values for BDNF for discriminating premalignant individuals from controls were 96.9% (sensitivity 83.3% and specificity 100%), 87.5% (sensitivity 90% and specificity 65.0%) for CRC patients from controls, and 92% (sensitivity 68.4% and specificity 100%) for all patients from controls Fig. . RegardiCollectively, these results confirm the predictive value of ∆Np73 effector BDNF as CRC biomarker in plasma of patients.3.6P = 0.1; by the Fisher's exact test). In contrast, none control mice exhibited metastasis in any of the evaluated organs. A representative ex vivo lung metastasis is shown  and mock cells (HCT116‐mock) by tail vein injection. After 3 weeks post‐injection mice were sacrificed and lungs, liver and kidney were extracted and examined. Three out of 10 mice injected with HCT116‐∆Np73 presented lung metastasis versus none of the control mice, showing a statistical trend . Optimal glucose supply to glycolytic cancer cells is a common hallmark of cancer. Among the upregulated effectors, LDHB and PYGL are associated to this supply , 48. Thein vivo approach  , 53, and5In conclusion, this unbiased proteome‐wide approach of the secretome of ΔNp73‐ and Mock‐stably transfected cells identified a large number of novel effectors of the ΔNp73 oncogenic modulator in CRC that seem to play a major role in tumor formation, progression and metastasis of CRC, and exhibit potential as diagnostic biomarkers of the disease. Further studies must be performed to determine whether the identified effectors might become new targets for therapeutic intervention in CRC.The authors declare no conflict of interest.MG‐A is involved in formal analysis, investigation, methodology, writing, review and editing manuscript. JR‐C is involved in formal analysis, investigation, methodology, writing, review and editing manuscript. CSM is involved in formal analysis and methodology. CP is involved in data curation, resources and review and editing manuscript. MJF‐A is involved in data curation, resources and review and editing manuscript. DP‐M is involved in formal analysis, methodology and resources. DV is involved in data curation, resources and review and editing manuscript. AM‐C is involved in formal analysis, methodology and resources. GS‐F is involved in methodology and resources. M‐ÁC is involved in data curation, resources and review and editing manuscript. MG‐C is involved in methodology and resources. NR is involved in data curation, resources and review and editing manuscript. AG‐A is involved in formal analysis, methodology, resources and review and editing manuscript. RB is involved in conceptualization, data curation, formal analysis, investigation, methodology, resources, validation, writing, review and editing manuscript. GD is involved in conceptualization, data curation, formal analysis, investigation, methodology, resources, validation, writing, review and editing manuscript.https://publons.com/publon/10.1002/1878‐0261.13228.The peer review history for this article is available at Fig. S1. Schematic design of the in‐depth proteomic analysis of ΔNp73 secretome effectors.Click here for additional data file.Fig. S2. Identification of protein interactions between the identified dysregulated proteins by the database STRING.Click here for additional data file.Fig. S3. Western blot of BDNF and EMAP‐II and ELISAs of VEGFC and VEGFR‐3.Click here for additional data file.Fig. S4. Metastasic potential of ΔNp73.Click here for additional data file.Table S1. Antibodies and primers used for experimental work.Click here for additional data file.Table S2. Clinical and pathological information of patient's serum samples used in ELISA experiments.Click here for additional data file.Table S3. Identified and quantified proteins and peptides in the SILAC experiment.Click here for additional data file.


Parathyroid carcinoma (PC) and atypical parathyroid tumor (APT) are rare parathyroid disorders carrying the risk of recurrence of varying degrees. This study aims to explore the value of 2D-shear wave elastography (SWE) in the discrimination of PC/APT among suspicious parathyroid lesions.In this prospective study, patients with primary hyperparathyroidism and suspicious parathyroid lesions on ultrasonography (US) were recruited. All the lesions were assessed by SWE before surgery. The velocity (m/s), Young’s modulus (Kpa), and elastogram of SWE were compared between pathologically proven parathyroid carcinoma or atypical parathyroid tumor (Group1) and parathyroid adenoma (Group2). All the SWE parameters were displayed at the setting of 50 or 70 kPa. Correlations between SWE and the lesion size as well as biochemical parameters were analyzed.p <  0.001, p <  0.001; respectively). In contrast, the max SWV and Young’s modulus showed a better sensitivity of 0.75 and 0.81, respectively. The “colored lesion” and “stiff rim” patterns on the elastogram are more indicated in parathyroid carcinoma and atypical parathyroid tumor, whereas the negative elastogram prevails in parathyroid adenoma. The SWV and Young’s modulus of the parathyroid lesions were independent of the tumor size, but the max SWV and Young’s modulus slightly correlated with serum parathyroid hormone concentration (PTH) .36 target lesions were enrolled for analysis. The mean shear wave velocity (SWV) between the two groups was 2.4 m/s vs 1.9 m/s, respectively, while the mean Young’s modulus was 11.1 kPa vs 18.2 kPa, respectively. The cut-off values are 2.35 m/s and 17.05 kPa correspondingly. The sensitivity and specificity of the selecting cut-off values were 0.56 vs 0.63 and 0.95 vs 1.0 : 0.669–0.956 vs 0.720–0.983]; 2D-shear wave elastography plays a useful role in the preoperative assessment of parathyroid lesions with suspicious malignancy. The mean SWV and Young’s modulus are advised as the favored diagnostic parameter with the best AUC and excellent specificities, while the max SWV and Young’s modulus are more sensitive to distinguish the PC and APT compared with other parameters. Primary hyperparathyroidism (PHPT) is a biochemical diagnosis characterized by an abnormally elevated level of serum parathyroid hormone, with hypercalcemia or normocalcemia . The ent. Eligible patients diagnosed with primary hyperparathyroidism between July 2018 and July 2022 were enrolled in the cohort. All of them have parathyroid tumors with size> 1.5 cm on ultrasonography. In addition, the candidates must equip at least one of the following suspicious malignant features: (1) serum iPTH > 500 pg/mL or serum calcium> 2.9 mmol/L; (2) lesions with irregular shape or infiltrative border on ultrasonography; (3) newly-onset lesions after previous parathyroid surgery (suspicious of recurrent parathyroid carcinoma). Patients younger than 18 years old, without preoperative US imaging for target lesions confirmed by pathology or choosing the conservative management were excluded from this study. All of them received surgery after a multimodal neck ultrasound evaluation. Preoperative biochemical parameters including serum intact parathyroid hormone , serum calcium , serum phosphorus , and alkaline phosphatase  were analyzed. Symptoms in participants were classified into 3 categories by severity. Grade 0 referred to those without any complaint but discovered by physical examination. Grade 1 represented classical symptoms like palpable neck mass, kidney stone, fatigue, bone pain, and constipation. Grade 2 stood for symptoms requiring hospitalization, including but not limited to osteitis fibrosa cystica, fractures, pancreatitis, intractable vomiting, and hypercalcemic crisis. The definitive pathology diagnosis of parathyroid neoplasm phenotypes is based on the criteria set out by World Health Organization 2017. The patients were divided into two groups according to the pathology-Group1: parathyroid carcinoma or atypical parathyroid adenoma; Group2: parathyroid adenoma.This research adhered to the Declaration of Helsinki and was approved by the Institutional Review Board (No: S-K1518). Written informed consent was obtained from each participantAll the patients were performed with B-mode ultrasound to locate the target lesions responsible for primary hyperparathyroidism while in the supine position. The candidate lesions were selected according to the concordant imaging results of the MIBI scan and US. Features detected by ultrasound including tumor sizes , diameters’ ratio (DR), echogenic texture, shape, boundary, calcification, and cyst change were recorded. DR was the ratio between the lesion’s maximum diameter and minimum diameter measured in all the dimensions. Echogenic texture findings comprise “homogeneous” and “heterogeneous”. Shape information contains “round or oval” and “irregular shape”. The boundary was described as “clear” or “infiltrative or blurred”. Three specialists with over 5 years of expertise in thyroid and parathyroid ultrasonography imaging defined the feature of each lesion through the best-of-three method.A senior sonographer with systemic training in SWE imaging performed the examination using an Aixplorer US system  with a linear probe SL10–2). A guideline stated by WFUMB has been complied with during operation. The elasticity scale range was set in between 0 and 50 kPa in an evaluation firstly. For few cases whose max SWE parameters exceed the upper limit of 50 kPa, the scale of 0-70 kPa was applied. Region of interest (ROI) was traced conforming to the shape of lesions. As for large lesions with complex composition, ROI was placed inside the lesion and avoid the areas with cystic change and calcification. At least three measurements were conducted for each lesion. The median number of the mean SWE values in all the measurements for a single lesion was summarized for calculation. Parathyroid tumor stiffness was presented as shear wave velocity in meters per second and Young’s modulus in Kpa. For qualitative SWE pattern classification, the color-coded SWE elastograms were divided into four patterns: negative, stiff rim, void center, and colored lesion  and STATA 15.0 . Continuous variables are summarized as mean ± standard deviation and median ± interquartile range according to normality verified by Shapiro–Wilk test. Mann–Whitney U test was used for comparison of biochemical parameters. Age of patients, diameters on US and SWE variables were compared by independent samples Student’s t-test. Categorical variables including symptom grades were compared using the chi-square test or Fisher’s exact test and were shown as N and percent. Correlations between SWE and the lesion size as well as biochemical parameters were analyzed by the Spearman test. The receiver operating characteristic (ROC) curve was established and the area under the curve (AUC) was calculated to obtain a cutoff value for related variables of SWE. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated. A two-tailed n = 8) and APT(n = 8), while 20 lesions were benign adenomas. No age or gender preponderance was observed between the two groups’ patients (p = 0.178). Serum PTH, Ca and ALP levels were markedly higher in Group1 compared with Group2 . Serum P level was significantly lower in Group1 relative to Group2 (p <  0.017). More severe symptoms presented in Group1 compared with Group2 (p <  0.001).The demographic and clinical characteristics of eligible patients are reported in Table p = 0.600). A significant difference in the tumor DR was found between the groups, with the PC/APT group having a lower DR value (p = 0.018). The location of the tumor was not different between the two groups (p = 0.053). The irregular shape was more prevalent in the PC/APT group than in the PA group (p <  0.001). Concerning echogenicity, heterogeneous mass was more frequent than homogeneous nodules in the PC/APT group (p = 0.009). The infiltrative border was highly prevalent in parathyroid carcinoma and atypical parathyroid tumor (p = 0.002). Intra-lesion cystic change was present in 56.3% (n = 9) of PC/APT group and 30% (n = 6) of PA group, showing no significant group differences (p = 0.650). The B-mode ultrasound and shear wave elastography findings of atypical parathyroid tumors and parathyroid carcinomas were also summarized in the Table The sonographic features are shown in Table  p = 0.00. Intra-le p = 0.08. The lo p = 0.05. The irrp <  0.001; AUC = 0.795, p = 0.006; AUC = 0.852,p <  0.001; AUC = 0.794, p = 0.009; respectively). The min SWV and Young’s modulus were not useful in diagnosing PC/APT. No difference was observed between mean SWV/Young’s modulus and max SWV/Young’s modulus in recognizing PC/APT(p = 0.8582). The best cut-off value of mean SWV in predicting PC/APT was 2.35 m/s. The mean Young’s modulus showed an equally diagnostic prospect with the best cut-off value of 17.05 kPa. Meanwhile, the best cut-off values for max SWV and max Young’s modulus were 2.70 m/s and 21.90 kPa, respectively. For the qualitative SWE pattern classification, the negative pattern was predominant in the parathyroid adenoma group(p <  0.001), while the “rim of stiffness” and “colored lesion” pattern were representative in the parathyroid carcinoma/atypical parathyroid tumor group . Mean SWV was not associated with iPTH , but Max SWV and Young’s modulus slightly correlate with iPTH .No correlations were identified between the lesion size and mean SWV, mean Young’s modulus, max SWV, max Young’s modulus  –10, fromFour studies using the same Supersonic Imagine Aixplorer System to evaluate the parathyroid lesions were identified in the PubMed database. In a study carried out by Golu et al., the authors found that the mean elasticity indexes of parathyroid lesions were 10.2 kPa, which is comparable to our measurement of 10.89 kPa for parathyroid adenoma . StangieThe elastogram of a parathyroid lesion reflects the tumor stiffness more comprehensively by containing the minimum, mean and maximum value of SWE information all together in a color-coded image. It is expected that the negative pattern was more common in the parathyroid adenoma group, which may due to the homogeneous and soft nature of parathyroid adenomas. The components of this benign parathyroid neoplasm are relatively simple, with high tumor cellularity, low amounts of connective tissue, fibrosis and necrosis. The “Stiff rim” sign was emphasized by Zhou et al. in differentiating between benign and malignant breast lesions . The mecThis study has some limitations. Firstly, the small scale of cases restricts further statistical analysis for exploring independent risk factors of SWE parameters. The improved diagnostic performance of adding SWE to conventional US and clinical features could not be estimated too. Secondly, quantitative values of SWE are believed not possible to extrapolate between two different devices. Thirdly, the interobserver variability in the evaluation of parathyroid lesions using 2D-SWE was not assessed, which requires further research.Atypical parathyroid tumor or parathyroid carcinoma is often observed in colored and stiff rim pattern on elastogram and showing the mean SWE value > 2.35 m/s or the mean Young’s modulus > 17.05 kPa, whereas a negative elastogram with a less SWE value is characteristic of parathyroid adenoma. The mean SWV and Young’s modulus are good at recognizing the parathyroid adenoma with excellent specificities, while the max SWV and Young’s modulus are more sensitive to distinguish the PC and APT compared with other parameters.


TH. Two presentations are described. Type A, milder, presents after 12 months of age with progressive hypokinesis and rigidity. Type B presents before 12 months as a progressive complex encephalopathy. We report a girl with mild THD who had recurrent episodes of neurological decompensations. Before the first episode, she had normal development except for mild head tremor. Episodes occurred at 12, 19, and 25 months of age. After viral infections or vaccination, she developed lethargy, worsened tremor, language, and motor regression including severe axial hypotonia, recuperating over several weeks of intensive rehabilitation but with residual tremor and mild lower limb spasticity. Basal ganglia imaging was normal. Exome sequencing revealed two missense variants of uncertain significance in TH: c.1147G>T and c.1084G>A. Both have low gnomAD allele frequencies and in silico, are predicted to be deleterious. Cerebrospinal fluid analysis showed low homovanillic acid  and low HVA/5‐hydroxyindolacetic acid molar ratio . She responded rapidly to L‐Dopa/carbidopa without further episodes. Literature review revealed four other THD patients who had a total of seven episodes of marked hypotonia and motor regression following infections, occurring between ages 12 months and 6 years. All improved with L‐Dopa/carbidopa treatment. Intermittent THD is treatable, important for genetic counseling, and should be considered after even a single episode of marked hypotonia with recuperation over weeks, especially in patients with preexisting tremor, dystonia, or rigidity.Tyrosine hydroxylase deficiency (THD) is a treatable inborn error of dopamine biosynthesis caused by mutations in  Pathogenic variants decrease TH enzymatic function.Tyrosine hydroxylase deficiency  is a rare treatable inborn error of dopamine biosynthesis. It is an autosomal recessive neurometabolic disorder due to mutations in the l‐tyrosine) to levodopa (L‐Dopa), the first step of the biosynthesis of catecholamine neurotransmitters . TH is mainly expressed in dopaminergic and noradrenergic neuronsTH.TH catalyzes the conversion of tyrosine ; length, 55 cm (93th centile); and head circumference, 34.5 cm (24th centile). She received phototherapy for 24 hours for neonatal jaundice. Prior to the first episode, she had normal development except for mild head tremor. By 12 months, she walked with support, was stable in the sitting position, crawled up stairs, and had a pincer grasp and a vocabulary of several words.The first episode of deterioration occurred at 12 months of age. Two weeks earlier she had been vaccinated, followed by intermittent fever. She developed marked fatigue, loss of walking/sitting, tremor (appendicular and worsened head tremor), and severe axial hypotonia. She was hospitalized for 3 days and recuperated completely over several days.Episodes 2 (19 months) and 3 (25 months) each followed viral infections by a few days. Both presented as lethargy, axial hypotonia, appendicular rigidity, and worsening tremor with motor and language regression. After each, she required several weeks of intensive rehabilitation.Between episodes, examination revealed mild axial tremor, mild lower limb spasticity, and axial hypotonia. She regained her developmental milestones but had a mild motor and language delay after the second episode. At the age of 3 years, neurodevelopmental evaluation revealed a 6 month delay.SUCLA2, SUCLG1, and molecular testing for Aicardi‐Goutières syndrome were normal. Cerebral magnetic resonance imaging (MRI) at 19 months , inherited from the father and c.1084G>A (p.Glu362Lys), inherited from the mother. Both variants have allele frequencies of <1/50 000 . Both are predicted to be deleterious by SIFT, PolyPhen and MutationTaster. Both are located in the catalytic domain of TH.Exome sequencing was performed and analyzed at CHU Sainte‐Justine using a standard Illumina® workflow, an in‐house bioinformatics pipeline based on GATK and Annovar. This revealed biallelic missense variants of unknown significance in Lumbar puncture at 2 years and 11 months revealed that the dopamine metabolite homovanillic acid (HVA) was 160 nmol/L, 32% below the lower reference limit for age . 5‐Hydroxyindolacetic acid (5‐HIAA) was normal  and the HVA/5‐HIAA molar ratio was low ,Dopa/carbidopa treatment (1.7 mg/kg/d of L‐Dopa), initiated immediately after the lumbar puncture, was followed within days by clinical improvement and the patient has had no further episodes. She walked independently at the age of 3 years 3 months. She is now 5 years old and receives 2.7 mg/kg/day of L‐Dopa, divided into four doses. Dyskinesia was detected when doses were increased, needing very slow and gradual dose adjustments. Parents and caregivers have noted some “on and off” phenomena at the end of dosing intervals, which is treated by a MAO inhibitor . We chose this approach instead of further increasing the L‐dopa dose, because of concerns of potential dopamine supplementation‐related dyskinesia. At 4 years of age, her general physical examination was normal. Neurologically, her pattern of running was still somewhat immature, but she was stable without falls. She had mild axial hypotonia. Deep tendon reflexes were normal and symmetrical. She could heel‐ and toe‐walk. She had no dystonia or abnormal posturing. She had mild tremor on finger‐to‐nose testing and minor brief dyskinetic movements that did not interfere with daily functioning.33.1We reviewed the medical literature for THD patients with episodic neurologic decompensations. We last searched PubMed on 30/08/2020 for English‐language articles with the following interrogation: “Dopa‐responsive dystonia” OR “Infantile hypokinetic rigid Parkinsonism” OR “Tyrosine hydroxylase deficiency” OR “Segawa's disease.” From these articles and their references, we attempted to identify other articles. We summarized the clinical, genetic, and biochemical features of patients with episodic decompensations or crises Table .3.2Including our patient, we identified five individuals with mild THD who presented at least one acute neurologic decompensation Table . A totalEach crisis was accompanied by marked motor regression, often with loss of walking, standing and/or sitting. Loss or reduction of speech was frequent. The principal neurologic symptoms were tremor (4/5), hypo/bradykinesia (4/5), hypotonia (4/5), and lethargy (4/5). Complete or partial recuperation occurred in all cases after episodes.Two patients were diagnosed during infancy . Three were diagnosed between 16 and 31 years of age. Biochemical findings in the CSF are summarized on Table TH gene variants, except that c.1400A>G was present in patients 1 and 3. All variants except one were missense. Patient 2 had a frameshift variant on one allele. The locations and type of TH variants in intermittent THD do not stand out clearly from those reported in Types A and B patients with chronic progressive courses , and Type B . Future clinical series should reveal whether these three categories are distinct or merge into a continuum.TH variants, c.1147G>T and c.1084G>A.Neurologic decompensation with normal systemic metabolic testing results can occur in many neurologic diseases, including several inborn errors of metabolism.For completeness and for future clinical series, we note that patient 5 has a hemodynamically insignificant form of the rare heart malformation, cor triatriatum. This finding has no obvious relationship to THD.There is practical importance in expanding the clinical classification of THD to include an intermittent form that is mild in comparison to types A and B, because all five such patients identified to date responded to L‐Dopa treatment. For patients with intermittent presentations, the diagnosis of THD might not be considered from current descriptions of this condition in chapters and reviews. Identification of an intermittent form of THD will therefore facilitate early diagnosis for patients with intermittent manifestations. By extension, because of the L‐Dopa responsiveness of the intermittent THD patients known to date, this may help to reduce the risk of neurologic disability in affected individuals.The authors declare that there is no conflict of interest.All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation  and with the Helsinki Declaration of 1975, as revised in 2000 (5). Informed consent was obtained from all patients for being included in the study.


Multiple myeloma is a malignant plasma cell disorder characterized by abnormal proliferation of clonal plasma cells in the bone marrow microenvironment. Involvement of the central nervous system is a rare extramedullary manifestation of multiple myeloma, occurring in approximately 1% of patients and confers a poor prognosis with a median overall survival of 2‐4 months . While nA 35‐year‐old Caucasian female with no significant past medical history presented to her general practitioner in September 2012 with a 2‐month history of epistaxis, abdominal pain, menorrhagia, diarrhea, and malaise. Initial laboratory results showed anemia, thrombocytopenia, hypercalcemia, and mild renal impairment. Subsequent investigations by hematology service showed an IgG kappa M‐protein of 72 g/L with the presence of immunoparesis on serum protein electrophoresis (SPEP) and immunofixation, kappa light chain of 3580 g/L, lambda light chain of <0.1 mg/L, albumin 33 g/l, and β2 microglobulin 1.8 mg/L. She had 80% CD138 positive plasma cells infiltrate in her bone marrow, with a complex hypodiploid clone . Whole body MRI showed bone marrow packing in the axial and appendicular skeleton, while PET‐CT showed focal concentration of FDG avidity in the right sacral bone. She was diagnosed with ISS stage 2 symptomatic myeloma.In October 2012, she began induction therapy with six cycles of lenalidomide, bortezomib, and dexamethasone (RVD), followed by high‐dose melphalan and autologous stem cell transplantation (ASCT) in June 2013, achieving complete response. Subsequently, she was commenced on high‐risk maintenance therapy with lenalidomide and bortezomib.In January 2016, she had clinical and biochemical relapse, and was treated with pomalidomide, bortezomib, and low‐dose dexamethasone (PVD). Clonal evolution was evident in her plasma cells showing del(17p), t, 1q gain, and, monosomy 13, confirming aggressive disease at relapse. Despite a three‐drug combination treatment, her disease progressed 4 months later, and she was salvaged with daratumumab. In September 2016, she presented with new‐onset neurological symptoms of headache and dizziness. Neurological examination was remarkable for bilateral papilledema. MRI brain showed CNS infiltration with leptomeningeal enhancement and MRI whole spine revealed an abnormal signal in the conus medullaris . However, her neurological symptoms returned, associating with a persistent presence of malignant plasma cells in her CSF. Pomalidomide was restarted along with daratumumab for synergistic effect to control her systemic progression.2 on days 1, 8, 15, and dexamethasone 10 mg on days 1, 2, 8, 9, 15, and 16 on a compassionate use basis. Salvage therapy with marizomib was well tolerated with no adverse events and resulted in a sustained clinical and radiological response and a reduction in CNS malignant plasmacytosis. After five cycles of marizomib, the patient's neurological symptoms of headaches and dizziness resolved. By cycle 7, her MRI brain demonstrated radiological improvement with no abnormal findings and complete resolution of leptomeningeal enhancement. CSF cytology was negative for malignant plasma cells. She also achieved partial response with a plateauing M‐protein at 3 g/L. However, 10 months after starting marizomib, her disease rapidly progressed systemically with the recurrence of myelomatous infiltration in her CSF, and she died in December 2017 (Figure In November 2016, she was commenced on marizomib 0.7 mg/mIn this letter, we showed that marizomib, a novel third generation proteasome inhibitor with activity and specificity that is distinct from other proteasome inhibitors, has the potential to fulfil this unmet clinical need in patients with CNS myeloma. Marizomib has the unique advantage of being the first PI that can irreversibly inhibit all three proteasome enzymatic subunits and can penetrate the blood brain barrier. Several studies have shown that marizomib can be retained in the CNS and substantially inhibits proteasome activity in the brain , 7. A prHere, the administration of marizomib resulted in a rapid and sustained clinical and radiological improvement with resolution of neurological symptoms and leptomeningeal enhancement, respectively. Furthermore, it led to >90% reduction in CSF plasmacytosis with a survival benefit of 1 year, which is three times that of the reported median survival , 9. In t


Systemic lupus erythematosus (SLE), especially neuropsychiatric SLE (NPSLE), is a complex systemic autoimmune disease, characterized by variable course and multiple organ dysfunction. Our study aimed to identify crucial microRNA (miRNAs) in SLE and NPSLE.Totally 12 cases of serum specimens were collected from General Hospital of Ningxia Medical University . After miRNA sequencing, differential expression analysis, miRNA target prediction, and miRNA‐messenger RNA (mRNA) regulatory network construction were performed to identify the hub miRNAs. The expression of target gene was determined by quantitative reverse transcription‐polymerase chain reaction and Western blot.There were 79 and 59 differentially expressed miRNAs (DEmiRNAs) in NPSLE versus Control, and SLE versus Control, respectively. Among 35 overlapped DEmiRNAs, 5 upregulated miRNAs'  target genes were supported by at least six databases. The miRNA‐mRNA network indicated that core miRNA hsa‐miR‐762 regulated 1270 target genes. MiR‐762 was significantly upregulated in SLE and NPSLE, and over expression of miR‐762 significantly suppressed GIPC PDZ domain containing family member 3 (GIPC3) expression in SLE and NPSLE.Upregulation of hub miRNA miR‐762 can suppress the expression of GIPC3 in both SLE and NPSLE samples, which is probably involved in the development of SLE and NPSLE. Meanwhile, along with the development from SLE to NPSLE, miR‐762 exhibits higher expression. In this study, we found that miR‐762 was significantly upregulated in Systemic lupus erythematosus (SLE) and neuropsychiatric SLE (NPSLE), and overexpression of miR‐762 significantly suppressed GIPC3 expression. MiR‐762 potentially contributes to the development of SLE and NPSLE, besides higher hsa‐miR‐762 expression might promote the development from SLE to NPSLE via inhibiting GIPC3 expression. Our findings would contribute to better understanding of possible mechanisms in SLE and NPSLE, and might be helpful for the diagnosis of SLE and NPSLE.22.1In present study, we totally collected 12 cases of human serum specimens from General Hospital of Ningxia Medical University, comprising 4 NPSLE samples, 4 SLE samples, and 4 normal control samples. The NPSLE samples were collected from August 2016 to October 2018, and SLE and control samples were collected during March 2018−September 2018. The experiments were approved by ethics committee of hospital, in line with The Helsinki Declaration. Informed consents were obtained from all donors, and the detailed clinical information was displayed in Supporting Informaton: Table The human miRNA expression profile of all samples were detected using Agilent Human miRNA Microarray Kit Release 21.0, 8x60K chip.2.2All NPSLE participants accorded with American College of Rheumatology Criteria for SLE (1999). Meanwhile, the following patients were excluded: (1) Patients with neuropsychiatric symptoms before the diagnosis of SLE/NPSLE; (2) NPSLE patients caused by traumatic brain trauma, intracranial infection, hypoxemia, hepatic encephalopathy, uremia, tumors, or severe electrolyte imbalance; (3) Patients with genetic risk factors.The SLE patients were in accordance with American College of Rheumatology Criteria for SLE (1997), meanwhile there was no neuropsychiatric symptom.Normal participants all had free history of chronic diseases.2.3p value was calculated.The tiff format picture data after chip scanning was preprocessed utilizing Feature Extraction software, and the raw data files (.txt) were obtained. Then, the raw data was input in GeneSpring software, and parameter information such as group information was written. All samples' data was normalized and undergone quality control. DEmiRNAs were analyzed, and 2.4Regarding the DEmiRNAs, we have predicted their target genes in 12 software, including miRWalk, DIANA‐microTv4.0, miRanda‐rel2010, mirBridge, miRDB4.0, miRmap, miRNAMap, PicTar2, PITA, RNA22v2, RNAhybrid2.1, and Targetscan6.2. Target genes predicted in at least six software programs in the meantime were considered to be DEmiRNAs' final targets.2.5p adjust <.05.Additionally, the target genes were subjected to the functional enrichment analysis in clusterProfiler package of R,2.62 incubator.HT22 mouse hippocampal neuronal cell line was purchased from Jennio Biological Technology. The cells were cultured in high‐glucose Dulbecco's modified Eagle's medium , supplemented with 1% penicillin and 10% fetal bovine serum , in a 37°C and 5% CO2.7Serum miRNA was extracted using miRNeasy Mini Kit . After RNA extraction, agarose gel electrophoresis was used to detect the RNA integrity. The concentration and purity of RNA was determined by UV spectrophotometer, and 1.9 < OD260/280 < 2.1 and OD260/230 > 2.0 indicated qualified RNA purity and concentration, respectively. The miScript SYBR Green PCR Kit  and miScript II RT kit  were used for reverse transcription and qPCR, separately. External reference was cel‐miR‐39. The procedure was as below: 95°C for 15 min, 94°C for 15 s, 55°C for 30 s, 70°C for 30 s, 40 cycles.The miRNAs in cells were extracted with miRcute miRNA isolation kit . The reagents used in qPCR included miRcute Plus miRNA qPCR Kit  and FastKing RT Kit . Internal reference was U6. Following procedure was conducted: 95°C for 15 min, 40 cycles of 94°C for 20 s and 60°C for 34 s.2–ΔΔCT formula.Total RNA in cells was extracted using RNAsimple Total RNA kit . TIANScript RT kit  and SuperReal PreMix Plus (SYBR‐Green) kit  were utilized in qPCR assay, and qPCR was done on Bio‐Rad CFX Manager 3.0. Internal reference was GAPDH. All primer sequences were designed using software Primer5 and were shown in Table 2.8Total protein was extracted from all samples, and the concentration was determined with bicinchoninic acid protein concentration assay kit . The Western blot assay was done in the previous methods.33.1Firstly, we have identified the DEmiRNAs in NPSLE versus Control, SLE versus Control, respectively. Compared with normal samples, there were totally 79 DEmiRNAs in NPSLE specimens, including 14 upregulated miRNAs and 65 downregulated miRNAs Figure . ExpressMoreover, compare with normal samples, a total of 59 DEmiRNAs were identified in SLE specimens, comprising 23 upregulated miRNAs and 36 downregulated miRNAs Figure , and DEm3.2There were 79 and 59 DEmiRNAs in NPSLE versus Control and SLE versus Control, separately. After cross analysis of these DEmiRNAs, 35 overlapped miRNAs were obtained Figure . Among tp value. adjust <.05), including 623 Biological Process terms, 63 Molecular Function terms, and 101 Cellular Component terms, and the top 20 GO terms were displayed in Figure There were totally 2319 non‐repetitive target genes of the above 5 miRNAs, which were subjected to GO and KEGG enrichment analysis. Those 2319 genes were significantly enriched in a total of 787 GO terms  with more robust target genes were subjected to the subsequent analysis. The miRNA‐mRNA regulatory network indicated that hsa‐miR‐762 was the core miRNA regulating more than half target genes. Moreover, hsa‐miR‐762 showed significantly higher expression in clinical NPSLE samples, compared with SLE or control samples. To the best of our knowledge, miR‐762 has been seldom reported in SLE or NPSLE at present. However, the role of miR‐762 has been explored in various cancers. In nasopharyngeal carcinoma, upregulation of miR‐762 was suggested to associate with progression by promoting tumor cells' proliferation and invasion.Subsequently, we mainly focused on miR‐762 and a key target gene GIPC3. Our data suggested that hsa‐miR‐762 expression was significantly upregulated in SLE and NPSLE, compared with control samples. Furthermore, significantly higher hsa‐miR‐762 expression was observed in NPSLE compared with SLE, which inspired us whether much higher hsa‐miR‐762 expression contributed to the development from SLE to NPSLE. Then, the over expression of miR‐762 evidenced that remarkably inhibited the GIPC3 expression. Thus, we suspected that miR‐762 contributed to the development from SLE to NPSLE via suppressing GIPC3 expression. GIPC is a Post‐synaptic density‐95, disks‐large and zonula occludens‐1‐domain containing adapter protein family regulating cell surface expression and endocytic trafficking of multiple components,Although we have revealed the pathogenic role of miR‐762 in SLE and NPSLE for the first time, some limitations were still existed in our study. Owing to few public mRNA data of SLE and NPSLE patients, GIPC3 related bioinformatics analysis was not conducted. Moreover, regarding miR‐762's influence on GIPC3, limited cell lines were employed, and there was a lack of further validation in SLE and NPSLE clinical samples. More detailed role of GIPC3 in SLE and NPSLE patients will be investigated integrating the transcriptome sequencing data in our future work.5In summary, via miRNA sequencing, miRNA‐mRNA regulatory network construction, and target gene validation, the significant pathogenic role of hub miRNA miR‐762 in SLE and NPSLE has been revealed for the first time. The upregulation of miR‐762 suppresses the expression of GIPC3 in both SLE and NPSLE samples, which is probably involved in the development of SLE and NPSLE. Furthermore, higher expression of miR‐762 in NPSLE compared with SLE is deserved to be investigated in near future.Jijuan Yang and Chun Li contributed to the study conception and design. Material preparation, data collection and analysis were performed by Shuhong Chi and Hongliang Wei. Jijuan Yang, Chun Li, and Wenqing Du conducted the experiments to validate the results of this study. The first draft of the manuscript was written by Jijuan Yang, Chun Li, Shuhong Chi, Hongliang Wei, and Wenqing Du. Qikuan Hu commented on previous versions of the manuscript. All authors read and approved the final manuscript.The authors declared no conflict of interest.This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by the Scientific Research Ethics Committee of General Hospital of Ningxia Medical University [No. 2019‐487]. Informed consent was obtained from all individual participants included in the study.Supporting information.Click here for additional data file.Supporting information.Click here for additional data file.Supporting information.Click here for additional data file.Supporting information.Click here for additional data file.


Maintaining ≥20 teeth is a public health goal worldwide. Healthy aging, which includes psychological and social well‐being, as well as physical indicators, has attracted a great deal of attention with the progression of aging societies. However, no studies have examined the association between the number of remaining teeth and healthy aging. This study aimed to investigate the association between the number of remaining teeth and healthy aging.This community‐based longitudinal cohort study included 8300 Japanese people aged ≥65 years who were free of disability and depression in the baseline survey in 2006. The participants were categorized into four groups according to the number of remaining teeth at baseline: 0–9, 10–19, 20–24 and ≥25. The primary outcome was healthy aging , as assessed by a questionnaire survey carried out in 2017. Multiple logistic regression was used to calculate the corresponding odds ratios and 95% confidence intervals.During about 11 years of follow‐up, 621 (7.5%) participants attained healthy aging. Participants with ≥20 remaining teeth showed a higher healthy aging rate. Compared with participants with 0–9 teeth, the multivariate‐adjusted odds ratios  for 10–19, 20–24 and ≥25 teeth were 0.98 (0.77–1.26), 1.28 (1.01–1.63) and 1.59 (1.24–2.03), respectively.Geriatr Gerontol Int 2022; 22: 68–74.These findings suggest that maintaining ≥20 teeth was associated with healthy aging.  With population aging, “successful aging” or “healthy aging” has attracted a great deal of attention worldwide, because people have begun to seek well‐being during aging rather than mere longevity and absence of disability.The concept of “successful aging” was first proposed by Rowe and Kahn, and they defined successful aging as, “low probability of disease and disease‐related disability and related risk factors,” “high cognitive and physical functional capacity” and “active engagement with life”.Good oral health has been shown to be associated with better physical or cognitive function.Therefore, the present study aimed to investigate the association between number of remaining teeth and healthy aging using data from a community‐based longitudinal cohort study on older people in Japan. We defined healthy aging as not only physical capability, but also a broad domain of well‐being, including free of depression, high HRQOL and high life satisfaction.n = 31 694). The baseline survey was carried out between 1 and 15 December 2006. A questionnaire was distributed to individual households by the heads of individual administrative districts and then collected by mail. Among the source population, 23 091 persons who provided valid responses formed the study cohort. We further excluded 6333 people who did not provide written consent for review of their long‐term care insurance (LTCI) information, 1979 who had already been certified as having a disability by the LTCI system before follow up, five who had died or moved out of the district before follow up and 4827 who had depressive symptoms or whose responses regarding depression were missing at baseline. Finally, 9947 participants were followed from 16 December 2006 to 30 November 2017 . The questionnaire included self‐report questions regarding depression, HRQOL and life satisfaction, which were the components used to assess healthy aging. We further excluded 651 participants who did not return a questionnaire or provide eligible responses, 464 whose responses on components assessing healthy aging were missing, and 227 whose answers on the number of remaining teeth were missing at baseline. Finally, 8300 participants, consisting of 2645 who died during follow up and 5655 who provided valid responses in the 2017 Survey, were included in the statistical analysis , most (25–27 teeth), moderate (20–24 teeth), about half (10–19 teeth), few (1–9 teeth) and none (0 teeth). Then, we divided the respondents into the following four groups according to quartiles of the categorization of the number of remaining teeth: 0–9 teeth, 10–19 teeth, 20–24 teeth and ≥25 teeth.We also asked the respondents whether they used dentures. Then, we combined the responses regarding number of teeth and denture use, and divided the respondents into the following six groups: 0–9 teeth, 10–19 teeth without dentures, 10–19 teeth with dentures, 20–24 teeth without dentures, 20–24 teeth with dentures and ≥25 teeth. The respondents who answered 0–9 teeth and ≥25 teeth were not classified based on whether they used dentures, because those who had 0–9 teeth mostly did (94.0%), and those who had ≥25 teeth mostly did not (86.1%).The primary outcome of the present study was healthy aging, which consisted of the following four components according to our previous studyIn the present study, we defined incident functional disability as certification under the LTCI system in Japan (Support Level 1 or higher), which uses a nationally uniform standard of functional disability.et al., is a brief screening instrument for depression and suicidal ideation for the aged.vs >1) produced satisfactory sensitivity and specificity in detecting depression , so we defined a Depression and Suicide Screen score of <2 as “free of depression”.Depression was measured both at baseline and in the 2017 Survey through the Depression and Suicide Screen. The Depression and Suicide Screen, which was developed in Japanese by Fujisawa et al.HRQOL was evaluated using the three‐level version of the European Quality of Life‐5 Dimensions (EQ‐5D‐3L) in the 2017 Survey.et al.Life satisfaction was assessed in the 2017 Survey using the Satisfaction With Life Scale developed by Diener 2).Body mass index was calculated as the self‐reported body weight (kg) divided by the square of the self‐reported body height  and 95% confidence intervals (CIs). Participants having 0–9 teeth were used as a reference category. Dummy variables were created for each multicategorical covariate for the corresponding model. For cases where values for a covariate were missing, we created a separate category for missing values and included this in the model. The following model was used to analyze the association between number of remaining teeth and healthy aging. The model was adjusted for sex, age, smoking status , drinking status , time spent walking , sleep duration , education level , history of disease , cognitive function score  and social participation , because the above variables were associated with health status.We also carried out two sets of sensitivity analyses. First, we replaced the exposure by combining the number of teeth and whether they used dentures. Second, we replaced the outcome with survival and four components of healthy aging using the same multiple logistical regression model, respectively.sas software package . All statistical tests were two‐sided, and P‐values <0.05 were considered significant.All statistical analyses were carried out using the We considered the return of a completed questionnaire to imply consent to participate in the study involving the baseline survey data, subsequent follow up for death and emigration, and the 2017 Survey data. The Ethics Committee of the Tohoku University Graduate School of Medicine  reviewed and approved the study protocols .In the present study, 8300 participants  were included in the statistical analysis. During approximately 11 years of follow up, 621 (7.5%) participants met all the criteria for healthy aging, and 5746 (69.2%) survived.Table Table Table This relationship did not differ when exposure was replaced with the combined number of teeth and use of dentures. The multivariate‐adjusted ORs (95% CIs) for 10–19 teeth without dentures, 10–19 teeth with dentures, 20–24 teeth without dentures, 20–24 teeth with dentures and ≥25 teeth were 0.97 (0.65–1.47), 0.98 (0.75–1.28), 1.18 (0.90–1.56), 1.46 (1.06–2.00) and 1.58 1.23–2.03), respectively .Click here for additional data file.