Diagnóstico oftalmológico del síndrome de linfedema-distiquiasis a través de la mutación FOXC2 / Ophthalmologic diagnosis of lymphedema-distichiasis syndrome through the FOXC2 mutation

El síndrome de linfedema-distiquiasis es uno de los fenotipos más frecuentes de aparición de linfedema primario; aun así, su prevalencia es baja.Este síndrome cursa con la aparición en la infancia o en la pubertad de pestañas aberrantes y otras formas de distiquiasis que pueden disminuir la calidad de vida, especialmente en pacientes en edades tan tempranas. La valoración clínica de este tipo de signos debe hacernos tener en mente este grupo de síndromes, ya que contamos, en este caso, con el diagnóstico de certeza gracias al análisis genético en suero del gen FOXC2, como ocurre en el caso que presentamos.De esta manera podemos prevenir, diagnosticar y tratar tanto los síntomas oftalmológicos como el resto de los síntomas sistémicos de forma precoz y más efectiva, aumentando así la calidad de vida de nuestros pacientes. (AU)

Lymphedema-distichiasis syndrome is one of the most frequent phenotypes of primary lymphedema, even so, its prevalence is still low.This syndrome courses with the appearance of abnormal eyelashes and distichiasis during childhood or puberty. This can cause a notable discomfort on our patients, especially at such an early age. The clinic evaluation of this signs must make us have in mind this group of syndromes, because in the case of lymphedema distichiasis syndrome, we can certainly diagnose it with the genetic analysis of the FOXC2 gen on patient's serum.With this we could prevent, diagnose and treat the ophthalmologic syndrome alongside the rest of systemic symptoms of this syndrome in a more effective way, giving our patients a higher quality of life. (AU)

Variants in FOXC1 and FOXC2 identified in patients with conotruncal heart defects.

Conotruncal heart defects (CTD), subtypes of congenital heart disease, result from abnormal cardiac outflow tract development (OFT). FOXC1 and FOXC2 are closely related members of the forkhead transcription factor family and play essential roles in the development of OFT. We confirmed their expression pattern in mouse and human embryos, identifying four variants in FOXC1 and three in FOXC2 by screening these two genes in 605 patients with sporadic CTD. Western blot demonstrated expression levels, while Dual-luciferase reporter assay revealed affected transcriptional abilities for TBX1 enhancer in two FOXC1 variants and three FOXC2 variants. This might result from the altered DNA-binding abilities of mutant proteins. These results indicate that functionally impaired FOXC1 and FOXC2 variants may contribute to the occurrence of CTD.

Ophtalmologic diagnosis of lymphedema-distichiasis syndrome through the FOXC2 mutation.

Lymphedema distichiasis syndrome is one of the most frequent phenotypes of primary lymphedema, even so, its prevalence is still low. This syndrome courses with the appearance of abnormal eyelashes and distichiasis during childhood or puberty. This can cause a notable discomfort on our patients, especially at such an early age. The clinic evaluation of this signs must make us have in mind this group of syndromes, because in the case of lymphedema distichiasis syndrome, we can certainly diagnose it with the genetic analysis of the FOXC2 gen on patient's serum. With this we could prevent, diagnose and treat the ophthalmologic syndrome alongside the rest of systemic symptoms of this syndrome in a more effective way, giving our patients a higher quality of life.

Association between pathological infiltrative tumor growth pattern and prognosis in patients with resected lung squamous cell carcinoma.

INTRODUCTION: Lung squamous cell carcinoma (LUSC) usually shows expansive growth with large tumor nests; few reports on invasive growth patterns (INF) in LUSC have been associated with poor prognosis in gastrointestinal and urothelial cancers. In this study, we examine the association between INF and the prognosis of LUSC. MATERIALS AND METHODS: We analyzed INF as a potential prognostic factor in 254 consecutive patients with LUSC who underwent complete surgical resection at our hospital between 2008 and 2017. INF was classified into 3 categories based on the structure of the tumor other than the large round solid nest of tumor cells. RESULTS: INF was categorized as INFa in 59 patients (23 %) with only well-demarcated large solid tumor cell nests, INFb in 89 patients (35 %) with medium to small, alongside large solid nests, and INFc in 98 patients (39 %) with cord-like/small nests or isolated cells plus large or medium solid nests. No significant lymph node metastasis differences were observed between INFc and INFa/b tumors. However, in patients with p-stage I, INFc had a poorer prognosis with regard to recurrence-free survival (RFS), with a 5-year RFS rate of 53.3 %, compared to 74.9 % for INFa/b (p = 0.010). CONCLUSION: Our study highlights a novel pathological concept of INF in LUSC, and contributed to the proposal that it is a factor indicating an unfavorable prognosis in patients with early-stage LUSC. A prospective multicenter study is warranted for INFc patients, as careful follow-up and adjuvant chemotherapy might lead to the early detection and prevention of recurrence.

Response of lymphatic endothelial cells to combined spatial and temporal variations in fluid flow.

One-way valves within lymphatic vessels are required for the efficient drainage of lymphatic fluids. Fluid flow is proposed to be a key cue in regulating both the formation and maintenance of lymphatic valves. However, to our knowledge, no previous study has systematically examined the response of LECs to the complex combination of spatially and temporally varying fluid flows that occur at lymphatic valves in vivo. We built an in vitro microfluidic device that reproduces key aspects of the flow environment found at lymphatic valves. Using this device, we found that a combination of spatially and temporally varying wall shear stresses (WSSs) led to upregulated transcription of PROX1 and FOXC2. In addition, we observed that combined spatial and temporal variations in WSS-modulated Ca2+ signaling and led to increased cellular levels of NFATc1. These observations suggest that the physical cues generated by the flow environment present within lymphatic valves may act to activate key regulatory pathways that contribute to valve maintenance.

FOXC1 and FOXC2 maintain mitral valve endothelial cell junctions, extracellular matrix, and lymphatic vessels to prevent myxomatous degeneration.

Background: Mitral valve (MV) disease including myxomatous degeneration is the most common form of valvular heart disease with an age-dependent frequency. Genetic evidence indicates mutations of the transcription factor FOXC1 are associated with MV defects, including mitral valve regurgitation. In this study, we sought to determine whether murine Foxc1 and its closely related factor, Foxc2, are required in valvular endothelial cells (VECs) for the maintenance of MV leaflets, including VEC junctions and the stratified trilaminar extracellular matrix (ECM). Methods: Adult mice carrying tamoxifen-inducible, endothelial cell (EC)-specific, compound Foxc1;Foxc2 mutations (i.e., EC-Foxc-DKO mice) were used to study the function of Foxc1 and Foxc2 in the maintenance of mitral valves. The EC-mutations of Foxc1/c2 were induced at 7 - 8 weeks of age by tamoxifen treatment, and abnormalities in the MVs of EC-Foxc-DKO mice were assessed via whole-mount immunostaining, immunohistochemistry, and Movat pentachrome/Masson's Trichrome staining. Results: EC-deletions of Foxc1 and Foxc2 in mice resulted in abnormally extended and thicker mitral valves by causing defects in regulation of ECM organization with increased proteoglycan and decreased collagen. Notably, reticular adherens junctions were found in VECs of control MV leaflets, and these reticular structures were severely disrupted in EC-Foxc1/c2 mutant mice. PROX1, a key regulator in a subset of VECs on the fibrosa side of MVs, was downregulated in EC-Foxc1/c2 mutant VECs. Furthermore, we determined the precise location of lymphatic vessels in murine MVs, and these lymphatic vessels were aberrantly expanded in EC-Foxc1/c2 mutant mitral valves. Conclusions: Our results indicate that Foxc1 and Foxc2 are required for maintaining the integrity of the MV, including VEC junctions, ECM organization, and lymphatic vessels to prevent myxomatous mitral valve degeneration.

FOXC2-AS1/FOXC2 axis mediates matrix stiffness-induced trans-differentiation of hepatic stellate cells into fibrosis-promoting myofibroblasts.

Matrix stiffness is a central modulator of hepatic stellate cells (HSCs) activation and hepatic fibrogenesis. However, the long non-coding RNAs (lncRNAs)-regulated transcriptional factors linking matrix stiffness to alterations in HSCs phenotype are not completely understood. In this study, we investigated the effects of matrix stiffness on HSCs activation and its potential mechanism. Through analysis the RNA-seq data with human primary HSCs cultured on 0.4 kPa and 25.6 kPa hydrogel, we identified that forkhead box protein C2 (FOXC2) and its antisense lncRNA FXOC2-AS1 as the new mechanosensing transcriptional regulators that coordinate HSCs responses to the matrix stiffness, moreover, FOXC2 and FOXC2-AS1 expression were also elevated in human fibrosis and cirrhosis tissues. The matrix stiffness was sufficient to activate HSCs into myofibroblasts, resulting in nuclear accumulation of FOXC2. Disrupting FOXC2 and FOXC2-AS1 level abrogated stiffness-induced activation of HSCs. Further mechanistic studies displayed that stiffness-upregulated lncRNA FOXC2-AS1 had no influence on transcription of FOXC2. FOXC2-AS1 exerted its biological function through maintaining the RNA stability of FOXC2, and protecting FOXC2 mRNA from degradation by RNA exosome complex. Additionally, rescue assays confirmed that reintroduction of FOXC2 in FOXC2-AS1-depleted HSCs reversed the repression of FOXC2-AS1 knockdown on stiffness-induced HSCs activation. In AAV6-treated mice fibrotic models, targeting FOXC2 in vivo lead to a reduced degree of liver fibrosis. In sum, our study uncovers a reciprocal crosstalk between matrix stiffness and FOXC2-AS1/FOXC2 axis leading to modulation of HSCs mechanoactivation and liver fibrosis, and present AAV6 shRNA as an effective strategy that targets FOXC2 leading to the resolution of liver fibrosis.

Identification of quiescent FOXC2+ spermatogonial stem cells in adult mammals.

In adult mammals, spermatogenesis embodies the complex developmental process from spermatogonial stem cells (SSCs) to spermatozoa. At the top of this developmental hierarchy lie a series of SSC subpopulations. Their individual identities as well as the relationships with each other, however, remain largely elusive. Using single-cell analysis and lineage tracing, we discovered both in mice and humans the quiescent adult SSC subpopulation marked specifically by forkhead box protein C2 (FOXC2). All spermatogenic progenies can be derived from FOXC2+ SSCs and the ablation of FOXC2+ SSCs led to the depletion of the undifferentiated spermatogonia pool. During germline regeneration, FOXC2+ SSCs were activated and able to completely restore the process. Germ cell-specific Foxc2 knockout resulted in an accelerated exhaustion of SSCs and eventually led to male infertility. Furthermore, FOXC2 prompts the expressions of negative regulators of cell cycle thereby ensures the SSCs reside in quiescence. Thus, this work proposes that the quiescent FOXC2+ SSCs are essential for maintaining the homeostasis and regeneration of spermatogenesis in adult mammals.

All eyes on FOXC2.

New evidence in mice suggests that cells expressing the transcription factor FOXC2 may form a reservoir of quiescent stem cells that contributes to sperm formation.

Foxc1 and Foxc2 function in osteochondral progenitors for the progression through chondrocyte hypertrophy and mineralization of the primary ossification center.

The forkhead box transcription factor genes Foxc1 and Foxc2 are expressed in the condensing mesenchyme of the developing skeleton prior to the onset of chondrocyte differentiation. To determine the roles of these transcription factors in limb development we deleted both Foxc1 and Foxc2 in lateral plate mesoderm using the Prx1-cre mouse line. Resulting compound homozygous mice died shortly after birth with exencephaly, and malformations to this sternum and limb skeleton. Notably distal limb structures were preferentially affected, with the autopods displaying reduced or absent mineralization. The radius and tibia bowed and the ulna and fibula were reduced to an unmineralized rudimentary structure. Molecular analysis revealed reduced expression of Ihh leading to reduced proliferation and delayed chondrocyte hypertrophy at E14.5. At later ages, Prx1-cre;Foxc1Δ/ Δ;Foxc2 Δ / Δ embryos exhibited restored Ihh expression and an expanded COLX-positive hypertrophic chondrocyte region, indicating a delayed exit and impaired remodeling of the hypertrophic chondrocytes. Osteoblast differentiation and mineralization were disrupted at the osteochondral junction and in the primary ossification center (POC). Levels of OSTEOPONTIN were elevated in the POC of compound homozygous mutants, while expression of Phex was reduced, indicating that impaired OPN processing by PHEX may underlie the mineralization defect we observe. Together our findings suggest that Foxc1 and Foxc2 act at different stages of endochondral ossification. Initially these genes act during the onset of chondrogenesis leading to the formation of hypertrophic chondrocytes. At later stages Foxc1 and Foxc2 are required for remodeling of HC and for Phex expression required for mineralization of the POC.

FOXC2-induced circCASK aggravates colorectal cancer progression by upregulating SIX1 expression.

Colorectal cancer (CRC) ranks as the most common gastrointestinal solid carcinoma globally. Substantial evidence has established a pivotal role for circular RNAs (circRNAs) in CRC progression. In this study, differentially expressed circRNAs were analyzed based on a public dataset (GSE126094) and elevated expression of circCASK (hsa_circ_0001917) was validated in CRC. Moreover, increased circCASK was also confirmed in CRC patients. Functionally, circCASK knockdown led to a significant decrease in CRC cell growth and attenuated cell migration and invasion. Similarly, circCASK knockdown markedly attenuated tumor growth in vivo. Mechanistically, circCASK sponged miR-1271-5p and enhanced sine oculis homeobox homolog 1 (SIX1) expression. More importantly, both SIX1 overexpression and miR-1271-5p knockdown could reverse the cellular behavior inhibition induced by circCASK knockdown. Furthermore, SIX1 was most strongly and positively linked with Wnt/ß-catenin signaling pathways, circCASK triggered Wnt/ß-catenin signaling through the miR-1271-5p/SIX1 axis, and FOXC2 transcriptionally induced circCASK expression. In conclusion, circCASK induced by FOXC2 accelerated CRC progression through the miR-1271-5p/SIX1 axis, thus providing an interesting insight into CRC tumorigenesis.

The FOXC2 Transcription Factor: A Master Regulator of Chemoresistance in Cancer.

FOXC2, a member of the forkhead box family of transcription factors, is an emerging oncogene that has been linked to several hallmarks of cancer progression. Among its many oncogenic functions is the promotion of drug resistance, with evidence supporting roles for FOXC2 in escape from broad classes of chemotherapeutics across an array of cancer types. In this Mini-Review, we highlight the current understanding of the mechanisms by which FOXC2 drives cancer chemoresistance, including its roles in the promotion of epithelial-mesenchymal transition, induction of multidrug transporters, activation of the oxidative stress response, and deregulation of cell survival signaling pathways. We discuss the clinical implications of these findings, including strategies for modulating FOXC2-associated chemoresistance in cancer. Particular attention is given to ways in which FOXC2 and its downstream gene products and pathways can be targeted to restore chemosensitivity in cancer cells. In addition, the utility of FOXC2 expression as a predictor of patient response to chemotherapy is also highlighted, with emphasis on the value of FOXC2 as a novel biomarker that can be used to guide therapeutic choice towards regimens most likely to achieve clinical benefit during frontline therapy.

Haploinsufficiencies of FOXF1, FOXC2 and FOXL1 genes originated from deleted 16q24.1q24.2 fragment related with alveolar capillary dysplasia with misalignment of pulmonary veins and lymphedema-distichiasis syndrome: relationship to phenotype.

OBJECTIVE: We describe a fetus with a 2.12-Mb terminal deleted fragment in 16q associated with alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV) and lymphedema-distichiasis syndrome (LDS) and intend to provide a comprehensive prenatal management strategy for the fetuses with ACDMPV and LDS through reviewing other similar published studies. METHODS: The fetus presented a series of diverse structural malformations including congenital cardiovascular, genitourinary and gastro-intestinal anomalies in ultrasound at 23 + 5 weeks of gestation (GA). Amniocentesis was conducted for karyotype analysis and copy number variation sequencing (CNV-seq) after informed consent. RESULTS: The fetal karyotype was 46,XX, however the result of CNV-seq showed an approximately 2.12-Mb deletion in 16q24.1q24.2 (85220000-87340000) × 1 indicating pathogenicity. CONCLUSION: Genomic testing should be recommend as a first line diagnostic tool for suspected ACDMPV and/or LDS or other genetic syndromes for the fetuses with structural abnormalities in clinical practice.

FOXC2 as a prognostic marker and a potential molecular target in patients with human solid tumors.

Background: Forkhead Box Protein C2 (FOXC2) belongs to the Forkhead/Wing-helix family. The regulatory role of this transcription factor in physiological function and carcinogenic activity has been proven in subsequent investigations. However, there is still scarcity of evidence on the relationship between FOXC2 expression and prognosis in human solid tumors. We conducted this meta-analysis to evaluate the role of FOXC2 as a prognosis factor and a possible target marker in human solid tumors. Methods: PubMed, Web of Science, Embase, and the Cochrane library database were all searched methodically. Eligible publications on FOXC2 in human solid tumors were gathered and reviewed. The effect sizes were calculated using pooled hazard ratios (HRs) or odds ratios (ORs) with the corresponding 95% confidence interval (CI). Statistical analysis was conducted with Stata SE12.0. Results: This meta-analysis comprised 3,267 patients from 20 studies covering a variety of solid tumors. Increased FOXC2 expression was related to shorter overall survival (OS) (HR = 2.05, 95% CI: 1.73-2.42). High expression of FOXC2 is associated with lymph node metastases (OR = 3.33, 95% CI: 2.65-4.19), TNM stage (OR = 3.09, 95% CI: 2.00-4.78), and age (OR = 1.26, 95% CI: 1.06-1.50), according to the pooled ORs. However, no significant association was observed between the high expression of FOXC2 and sex, tumor size or tumor differentiation. Conclusion: Increased expression of FOXC2 is associated with unfavored OS, lymph node metastases, TNM stage, and age. FOXC2 is a promising prognostic marker and a novel target marker in human solid tumors.

FOXC2 Promotes Vasculogenic Mimicry in Ovarian Cancer.

FOXC2 is a forkhead family transcription factor that plays a critical role in specifying mesenchymal cell fate during embryogenesis. FOXC2 expression is associated with increased metastasis and poor survival in various solid malignancies. Using in vitro and in vivo assays in mouse ovarian cancer cell lines, we confirmed the previously reported mechanisms by which FOXC2 could promote cancer growth, metastasis, and drug resistance, including epithelial-mesenchymal transition, stem cell-like differentiation, and resistance to anoikis. In addition, we showed that FOXC2 expression is associated with vasculogenic mimicry in mouse and human ovarian cancers. FOXC2 overexpression increased the ability of human ovarian cancer cells to form vascular-like structures in vitro, while inhibition of FOXC2 had the opposite effect. Thus, we present a novel mechanism by which FOXC2 might contribute to cancer aggressiveness and poor patient survival.

Foxc1 and Foxc2 are indispensable for the maintenance of nephron and stromal progenitors in the developing kidney.

Nephron development proceeds with reciprocal interactions among three layers: nephron progenitors (NPs), ureteric buds and stromal progenitors (SPs). We found that Foxc1 and Foxc2 (Foxc1/2) are expressed in NPs and SPs. Systemic deletion of Foxc1/2 2 days after the onset of metanephros development (embryonic day 13.5) resulted in the epithelialization of NPs and ectopic formation of renal vesicles. NP-specific deletion did not cause these phenotypes, indicating that Foxc1/2 in other cells (likely in SPs) contributed to the maintenance of NPs. Single-cell RNA-sequencing analysis revealed the existence of NP and SP subpopulations, the border between committed NPs and renewing NPs, and similarity between the cortical interstitium and vascular smooth muscle type cells. Integrated analysis of the control and Foxc1/2 knockout data indicated transformation of some NPs to strange cells expressing markers of the vascular endothelium, reduced numbers of self-renewing NP and SP populations, and downregulation of crucial genes for kidney development, such as Fgf20 and Frem1 in NPs, and Foxd1 and Sall1 in SPs. It also revealed upregulation of genes that were not usually expressed in NPs and SPs. Thus, Foxc1/2 maintain NPs and SPs by regulating the expression of multiple genes.

Oncogenic functions of the FOXC2 transcription factor: a hallmarks of cancer perspective.

Epigenetic regulation of gene expression is a fundamental determinant of molecular and cellular function, and epigenetic reprogramming in the context of cancer has emerged as one of the key enabling characteristics associated with acquisition of the core hallmarks of this disease. As such, there has been renewed interest in studying the role of transcription factors as epigenetic regulators of gene expression in cancer. In this review, we discuss the current state of knowledge surrounding the oncogenic functions of FOXC2, a transcription factor that frequently becomes dysregulated in a variety of cancer types. In addition to highlighting the clinical impact of aberrant FOXC2 activity in cancer, we discuss mechanisms by which this transcription factor becomes dysregulated in both tumor and tumor-associated cells, placing particular emphasis on the ways in which FOXC2 promotes key hallmarks of cancer progression. Finally, we bring attention to important issues related to the oncogenic dysregulation of FOXC2 that must be addressed going forward in order to improve our understanding of FOXC2-mediated cancer progression and to guide prognostic and therapeutic applications of this knowledge in clinical settings.

Genetic landscape of FOXC2 mutations in lymphedema-distichiasis syndrome: Different mechanism of pathogenicity for mutations in different domains.

Lymphedema-dissociated syndrome (LDS), of which the pathogenesis is not fully understood, afflicts many patients. In this study, we investigated the effect of FOXC2 gene loss-of-function on the development of LDS disease. Two Han Chinese families with LDS were recruited in this study, pathogenic mutations were identified by Sanger sequencing. Reverse-transcription PCR, subcellular localization, dual fluorescein enzymes, and other in vitro experiments were used to study the functional effects of eight FOXC2 mutations. Two pathogenic FOXC2 duplication mutations (c.930_936dup and c.931-937dup) were identified in the two families. Both mutations caused uneven distribution in the nucleus and a chromatin contraction phenotype, weakening the DNA binding activity and transcription activity. We then performed functional analysis on six additional mutations in different domains of FOXC2 that were reported to cause LDS. We found mutations located in the forkhead domain and central region dramatically reduced the transactivation ability, while mutations in activation domain-2 enhanced this ability. All 8 mutations down-regulated the transcription of ANGPT2 and affected the activity of the ERK-RAS pathway, which may cause abnormal formation of lymphatic vessels. Our findings also showed that all 8 mutations decreased the ability of interaction between FOXC2 and the Wnt4 promoter, suggesting mutations in FOXC2 may also affect the Wnt4-Frizzled-RYK signaling pathway, leading the abnormal differentiation of the meibomian glands into hair follicle cells during the embryonic period and causing distichiasis. This study expanded and revealed the potential pathogenesis mechanism.

Microdeletion of 16q24.1-q24.2-A unique etiology of Lymphedema-Distichiasis syndrome and neurodevelopmental disorder.

Interstitial deletions of 16q24.1-q24.2 are associated with alveolar capillary dysplasia, congenital renal malformations, neurodevelopmental disorders, and congenital abnormalities. Lymphedema-Distichiasis syndrome (LDS; OMIM # 153400) is a dominant condition caused by heterozygous pathogenic variants in FOXC2. Usually, lymphedema and distichiasis occur in puberty or later on, and affected individuals typically achieve normal developmental milestones. Here, we describe a boy with congenital lymphedema, distichiasis, bilateral hydronephrosis, and global developmental delay, with a de novo microdeletion of 894 kb at 16q24.1-q24.2. This report extends the phenotype of both 16q24.1-q24.2 microdeletion syndrome and of LDS. Interestingly, the deletion involves only the 3'-UTR part of FOXC2.

First genome-wide association study of esophageal atresia identifies three genetic risk loci at CTNNA3, FOXF1/FOXC2/FOXL1, and HNF1B.

Esophageal atresia with or without tracheoesophageal fistula (EA/TEF) is the most common congenital malformation of the upper digestive tract. This study represents the first genome-wide association study (GWAS) to identify risk loci for EA/TEF. We used a European case-control sample comprising 764 EA/TEF patients and 5,778 controls and observed genome-wide significant associations at three loci. On chromosome 10q21 within the gene CTNNA3 (p = 2.11 × 10-8; odds ratio [OR] = 3.94; 95% confidence interval [CI], 3.10-5.00), on chromosome 16q24 next to the FOX gene cluster (p = 2.25 × 10-10; OR = 1.47; 95% CI, 1.38-1.55) and on chromosome 17q12 next to the gene HNF1B (p = 3.35 × 10-16; OR = 1.75; 95% CI, 1.64-1.87). We next carried out an esophageal/tracheal transcriptome profiling in rat embryos at four selected embryonic time points. Based on these data and on already published data, the implicated genes at all three GWAS loci are promising candidates for EA/TEF development. We also analyzed the genetic EA/TEF architecture beyond the single marker level, which revealed an estimated single-nucleotide polymorphism (SNP)-based heritability of around 37% ± 14% standard deviation. In addition, we examined the polygenicity of EA/TEF and found that EA/TEF is less polygenic than other complex genetic diseases. In conclusion, the results of our study contribute to a better understanding on the underlying genetic architecture of ET/TEF with the identification of three risk loci and candidate genes.