Skip to main content
SpringerLink
Log in

Single-cell RNA sequencing reveals the transcriptional heterogeneity of Tbx18-positive cardiac cells during heart development

  • Original Article
  • Published:
Functional & Integrative Genomics Aims and scope Submit manuscript

Abstract

The T-box family transcription factor 18 (Tbx18) has been found to play a critical role in regulating the development of the mammalian heart during the primary stages of embryonic development while the cellular heterogeneity and landscape of Tbx18-positive (Tbx18+) cardiac cells remain incompletely characterized. Here, we analyzed prior published single-cell RNA sequencing (scRNA-seq) mouse heart data to explore the heterogeneity of Tbx18+ cardiac cell subpopulations and provide a comprehensive transcriptional landscape of Tbx18+ cardiac cells during their development. Bioinformatic analysis methods were utilized to identify the heterogeneity between cell groups. Based on the gene expression characteristics, Tbx18+ cardiac cells can be classified into a minimum of two distinct cell populations, namely fibroblast-like cells and cardiomyocytes. In terms of temporal heterogeneity, these cells exhibit three developmental stages, namely the MEM stage, ML_P0 stage, and P stage Tbx18+ cardiac cells. Furthermore, Tbx18+ cardiac cells encompass several cell types, including cardiac progenitor-like cells, cardiomyocytes, and epicardial/stromal cells, as determined by specific transcriptional regulatory networks. The scRNA-seq results revealed the involvement of extracellular matrix (ECM) signals and epicardial epithelial-to-mesenchymal transition (EMT) in the development of Tbx18+ cardiac cells. The utilization of a lineage-tracing model served to validate the crucial function of Tbx18 in the differentiation of cardiac cells. Consequently, these findings offer a comprehensive depiction of the cellular heterogeneity within Tbx18+ cardiac cells.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

Availability of data and materials

The datasets used and analyzed during the current study are available from the corresponding author Jianlin Du.

Abbreviations

Tbx18+ :

Tbx18-positive

E:

Embryonic

P:

Postnatal

ScRNA-seq:

Single-cell RNA sequencing

ECM:

Extracellular matrix

EMT:

Epithelial-to-mesenchymal transition

CPC:

Cardiac progenitor cell

TF:

Transcription factor

SCENIC:

Single-cell regulatory network inference and clustering

PCR:

Polymerase chain reaction

GRN:

Gene regulatory network

t-SNE:

T-distributed stochastic neighbor embedding

DEGs:

Differentially expressed genes

GO:

Gene Ontology

KEGG:

Kyoto Encyclopedia of Genes and Genomes

EPDCs:

Epicardial-derived cells

hiPSCs:

Human-induced pluripotent stem cells

References

  • Aibar S, González-Blas CB, Moerman T, Huynh-Thu VA, Imrichova H, Hulselmans G et al (2017a) SCENIC: single-cell regulatory network inference and clustering. Nat Methods 14(11):1083–1086

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Aibar S, Gonzalez-Blas CB, Moerman T, Huynh-Thu VA, Imrichova H, Hulselmans G et al (2017b) SCENIC: single-cell regulatory network inference and clustering. Nat Methods 14(11):1083–1086

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Al-Hattab DS, Safi HA, Nagalingam RS, Bagchi RA, Stecy MT, Czubryt MP (2018) Scleraxis regulates Twist1 and Snai1 expression in the epithelial-to-mesenchymal transition. Am J Physiol Heart Circ Physiol 315(3):H658–H668

    Article  CAS  PubMed  Google Scholar 

  • Asp M, Giacomello S, Larsson L, Wu C, Furth D, Qian X et al (2019) A spatiotemporal organ-wide gene expression and cell atlas of the developing human heart. Cell 179(7):1647–6019

    Article  CAS  PubMed  Google Scholar 

  • Bak ST, Harvald EB, Ellman DG, Mathiesen SB, Chen T, Fang S et al (2023) Ploidy-stratified single cardiomyocyte transcriptomics map Zinc Finger E-Box Binding Homeobox 1 to underly cardiomyocyte proliferation before birth. Basic Res Cardiol 118(1):8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Beecroft SJ, Ayala M, McGillivray G, Nanda V, Agolini E, Novelli A et al (2021) Biallelic hypomorphic variants in ALDH1A2 cause a novel lethal human multiple congenital anomaly syndrome encompassing diaphragmatic, pulmonary, and cardiovascular defects. Hum Mutat 42(5):506–519

    Article  CAS  PubMed  Google Scholar 

  • Bellchambers HM, Ware SM (2021) Loss of Zic3 impairs planar cell polarity leading to abnormal left-right signaling, heart defects and neural tube defects. Hum Mol Genet 30(24):2402–2415

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bourajjaj M, Armand AS, da Costa Martins PA, Weijts B, van der Nagel R, Heeneman S et al (2008) NFATc2 is a necessary mediator of calcineurin-dependent cardiac hypertrophy and heart failure. J Biol Chem 283(32):22295–22303

    Article  CAS  PubMed  Google Scholar 

  • Buckingham M, Meilhac S, Zaffran S (2005) Building the mammalian heart from two sources of myocardial cells. Nat Rev Genet 6(11):826–835

    Article  CAS  PubMed  Google Scholar 

  • Butler A, Hoffman P, Smibert P, Papalexi E, Satija R (2018) Integrating single-cell transcriptomic data across different conditions, technologies, and species. Nat Biotechnol 36(5):411–420

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cai CL, Martin JC, Sun Y, Cui L, Wang L, Ouyang K et al (2008) A myocardial lineage derives from Tbx18 epicardial cells. Nature 454(7200):104–108

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Cao J, Spielmann M, Qiu X, Huang X, Ibrahim DM, Hill AJ et al (2019) The single-cell transcriptional landscape of mammalian organogenesis. Nature 566(7745):496–502

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Carter B, Zhao K (2020) The epigenetic basis of cellular heterogeneity. Nat Rev Genet 22(4):235–250

    Article  PubMed  Google Scholar 

  • Charron S, Roubertie F, Benoist D, Dubes V, Gilbert SH, Constantin M et al (2015) Identification of region-specific myocardial gene expression patterns in a chronic swine model of repaired tetralogy of fallot. PLoS ONE 10(8):e0134146

    Article  PubMed  PubMed Central  Google Scholar 

  • Chen Y, Wu X, Hu D, Wang W (2020) Importance of mitochondrial-related genes in dilated cardiomyopathy based on bioinformatics analysis. Cardiovasc Innov Appl 5(2):117–129

    Google Scholar 

  • Christoffels VM, Grieskamp T, Norden J, Mommersteeg MT, Rudat C, Kispert A (2009) Tbx18 and the fate of epicardial progenitors. Nature 458(7240):E8–9; discussion E-10

  • Chu M, Wang L, Wang H, Shen T, Yang Y, Sun Y et al (2014) A novel role of CDX1 in embryonic epicardial development. PLoS ONE 9(7):e103271

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Churko JM, Garg P, Treutlein B, Venkatasubramanian M, Wu H, Lee J et al (2018) Defining human cardiac transcription factor hierarchies using integrated single-cell heterogeneity analysis. Nat Commun 9(1):4906

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Cui Y, Zheng Y, Liu X, Yan L, Fan X, Yong J et al (2019) Single-cell transcriptome analysis maps the developmental track of the human heart. Cell Rep 26(7):1934–50 e5

    Article  CAS  PubMed  Google Scholar 

  • D’Amato G, Phansalkar R, Naftaly JA, Fan X, Amir ZA, Rios Coronado PE et al (2022) Endocardium-to-coronary artery differentiation during heart development and regeneration involves sequential roles of Bmp2 and Cxcl12/Cxcr4. Dev Cell 57(22):2517–32.e6

    Article  PubMed  PubMed Central  Google Scholar 

  • Dai W, Weber C (2018) Tbx18 sets the pace. J Physiol 596(24):6129–6130

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dang H, Ye Y, Zhao X, Zeng Y (2020) Identification of candidate genes in ischemic cardiomyopathy by gene expression omnibus database. BMC Cardiovasc Disord 20(1):320

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • de Soysa TY, Ranade SS, Okawa S, Ravichandran S, Huang Y, Salunga HT et al (2019) Single-cell analysis of cardiogenesis reveals basis for organ-level developmental defects. Nature 572(7767):120–124

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • DeLaughter DM, Bick AG, Wakimoto H, McKean D, Gorham JM, Kathiriya IS et al (2016) Single-cell resolution of temporal gene expression during heart development. Dev Cell 39(4):480–490

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dingar D, Konecny F, Zou J, Sun X, von Harsdorf R (2012) Anti-apoptotic function of the E2F transcription factor 4 (E2F4)/p130, a member of retinoblastoma gene family in cardiac myocytes. J Mol Cell Cardiol 53(6):820–828

    Article  CAS  PubMed  Google Scholar 

  • Dorn LE, Lawrence W, Petrosino JM, Xu X, Hund TJ, Whitson BA et al (2021) Microfibrillar-associated protein 4 regulates stress-induced cardiac remodeling. Circ Res 128(6):723–737

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Duran J, Troncoso M, Lagos D, Ramos S, Marin G, Estrada M (2018) GDF11 modulates Ca2+-dependent Smad2/3 signaling to prevent cardiomyocyte hypertrophy. Int J Mol Sci 19(5):1508

  • Farini A, Villa C, Di Silvestre D, Bella P, Tripodi L, Rossi R et al (2020) PTX3 Predicts Myocardial Damage And Fibrosis In Duchenne Muscular Dystrophy. Front Physiol 11:403

    Article  PubMed  PubMed Central  Google Scholar 

  • Filosa A, Sawamiphak S (2023) Heart development and regeneration—a multi-organ effort. FEBS J 290(4):913–930

    Article  CAS  PubMed  Google Scholar 

  • Fujii M, Sakaguchi A, Kamata R, Nagao M, Kikuchi Y, Evans SM et al (2017) Sfrp5 identifies murine cardiac progenitors for all myocardial structures except for the right ventricle. Nat Commun 8:14664

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Galdos FX, Wu SM (2019) Single-cell delineation of who’s on first and second heart fields during development. Circ Res 125(4):411–413

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Galindo CL, Nguyen VT, Hill B, Easterday E, Cleator JH, Sawyer DB (2022) Neuregulin (NRG-1beta) is pro-myogenic and anti-cachectic in respiratory muscles of post-myocardial infarcted Swine. Biology (Basel) 11(5):682

  • Gancz D, Perlmoter G, Yaniv K (2020) Formation and growth of cardiac lymphatics during embryonic development, heart regeneration, and disease. Cold Spring Harb Perspect Biol. 12(6):a037176

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gao Y, Qi GX, Guo L, Sun YX (2016) Bioinformatics analyses of differentially expressed genes associated with acute myocardial infarction. Cardiovasc Ther 34(2):67–75

    Article  CAS  PubMed  Google Scholar 

  • Gharibeh L, Yamak A, Whitcomb J, Lu A, Joyal M, Komati H et al (2021) GATA6 is a regulator of sinus node development and heart rhythm. Proc Natl Acad Sci U S A 118(1):e2007322118

  • Gibb N, Lazic S, Yuan X, Deshwar AR, Leslie M, Wilson MD et al (2018) Hey2 regulates the size of the cardiac progenitor pool during vertebrate heart development. Development 145(22):dev167510

  • Gorabi AM, Hajighasemi S, Tafti HA, Atashi A, Soleimani M, Aghdami N et al (2019) TBX18 transcription factor overexpression in human-induced pluripotent stem cells increases their differentiation into pacemaker-like cells. J Cell Physiol 234(2):1534–1546

    Article  CAS  PubMed  Google Scholar 

  • Gu Y, Zhou Y, Ju S, Liu X, Zhang Z, Guo J et al (2022) Multi-omics profiling visualizes dynamics of cardiac development and functions. Cell Rep 41(13):111891

    Article  CAS  PubMed  Google Scholar 

  • Guo Y, Cao Y, Jardin BD, Sethi I, Ma Q, Moghadaszadeh B et al (2020) Sarcomeres regulate murine cardiomyocyte maturation through MRTF-SRF signaling. Proc Natl Acad Sci 118(2):e2008861118

  • Han Z, Yu Y, Cai B, Xu Z, Bao Z, Zhang Y et al (2020) YAP/TEAD3 signal mediates cardiac lineage commitment of human-induced pluripotent stem cells. J Cell Physiol 235(3):2753–2760

    Article  CAS  PubMed  Google Scholar 

  • Han C, Li H, Ma Z, Dong G, Wang Q, Wang S et al (2021) MIR99AHG is a noncoding tumor suppressor gene in lung adenocarcinoma. Cell Death Dis 12(5):424

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hong L, Li N, Gasque V, Mehta S, Ye L, Wu Y et al (2022) Prdm6 controls heart development by regulating neural crest cell differentiation and migration. JCI Insight 7(4):e156046

  • Huang GN, Thatcher JE, McAnally J, Kong Y, Qi X, Tan W et al (2012) C/EBP transcription factors mediate epicardial activation during heart development and injury. Science 338(6114):1599–1603

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Islas JF, Liu Y, Weng K-C, Robertson MJ, Zhang S, Prejusa A et al (2012) Transcription factors ETS2 and MESP1 transdifferentiate human dermal fibroblasts into cardiac progenitors. Proc Natl Acad Sci 109(32):13016–13021

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Jia G, Preussner J, Chen X, Guenther S, Yuan X, Yekelchyk M et al (2018) Single cell RNA-seq and ATAC-seq analysis of cardiac progenitor cell transition states and lineage settlement. Nat Commun 9(1):4877

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Jia Y, Chang Y, Guo Z, Li H (2019) Transcription factor Tbx5 promotes cardiomyogenic differentiation of cardiac fibroblasts treated with 5-azacytidine. J Cell Biochem 120(10):16503–16515

    Article  CAS  PubMed  Google Scholar 

  • Jiang D-S, Liu Y, Zhou H, Zhang Y, Zhang X-D, Zhang X-F et al (2014) Interferon regulatory factor 7 functions as a novel negative regulator of pathological cardiac hypertrophy. Hypertension 63(4):713–722

    Article  CAS  PubMed  Google Scholar 

  • Jiang Z, Feng T, Lu Z, Wei Y, Meng J, Lin C-P et al (2021a) PDGFRb mesenchymal cells, but not NG2 mural cells, contribute to cardiac fat. Cell Rep 34(5):108697

    Article  CAS  PubMed  Google Scholar 

  • Jiang Z, Feng T, Lu Z, Wei Y, Meng J, Lin C-P et al (2021b) PDGFRb+ mesenchymal cells, but not NG2+ mural cells, contribute to cardiac fat. Cell Rep 34(5):108697

    Article  CAS  PubMed  Google Scholar 

  • Jing X, Gao Y, Xiao S, Qin Q, Wei X, Yan Y et al (2016) Hypoxia induced the differentiation of Tbx18-positive epicardial cells to CoSMCs. Sci Rep 6:30468

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Jing Y, Gao B, Han Z, Xia L, Xin S (2021) The protective effect of HOXA5 on carotid atherosclerosis occurs by modulating the vascular smooth muscle cell phenotype. Mol Cell Endocrinol 534:111366

    Article  CAS  PubMed  Google Scholar 

  • Kern CB, Wessels A, McGarity J, Dixon LJ, Alston E, Argraves WS et al (2010) Reduced versican cleavage due to Adamts9 haploinsufficiency is associated with cardiac and aortic anomalies. Matrix Biol 29(4):304–316

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Koenig AL, Shchukina I, Amrute J, Andhey PS, Zaitsev K, Lai L et al (2022) Single-cell transcriptomics reveals cell-type-specific diversification in human heart failure. Nat Cardiovasc Res 1(3):263–280

    Article  PubMed  PubMed Central  Google Scholar 

  • Kraus F, Haenig B, Kispert A (2001) Cloning and expression analysis of the mouse T-box gene Tbx18. Mech Dev 100(1):83–86

    Article  CAS  PubMed  Google Scholar 

  • Kumar S, Wang G, Zheng N, Cheng W, Ouyang K, Lin H et al (2019) HIMF (hypoxia-induced mitogenic factor)-IL (interleukin)-6 signaling mediates cardiomyocyte-fibroblast crosstalk to promote cardiac hypertrophy and fibrosis. Hypertension 73(5):1058–1070

    Article  CAS  PubMed  Google Scholar 

  • Kwon T, Kwon O-S, Cha H-J, Sung BJ (2019) Stochastic and heterogeneous cancer cell migration: experiment and theory. Sci Rep 9(1):16297

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Lalit PA, Salick MR, Nelson DO, Squirrell JM, Shafer CM, Patel NG et al (2016) Lineage reprogramming of fibroblasts into proliferative induced cardiac progenitor cells by defined factors. Cell Stem Cell 18(3):354–367

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Le T, Chong J (2016) Cardiac progenitor cells for heart repair. Cell Death Discov 2:16052

    Article  PubMed  PubMed Central  Google Scholar 

  • Lescroart F, Wang X, Lin X, Swedlund B, Gargouri S, Sanchez-Danes A et al (2018) Defining the earliest step of cardiovascular lineage segregation by single-cell RNA-seq. Science 359(6380):1177–1181

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Li G, Xu A, Sim S, Priest JR, Tian X, Khan T et al (2016) Transcriptomic profiling maps anatomically patterned subpopulations among single embryonic cardiac cells. Dev Cell 39(4):491–507

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li X, Poire A, Jeong KJ, Zhang D, Chen G, Sun C et al (2023a) Single-cell trajectory analysis reveals a CD9 positive state to contribute to exit from stem cell-like and embryonic diapause states and transit to drug-resistant states. Cell Death Discov 9(1):285

    Article  PubMed  PubMed Central  Google Scholar 

  • Li J, Wang CQ, Xiao WC, Chen Y, Tu J, Wan F et al (2023b) TRAF family member 4 promotes cardiac hypertrophy through the activation of the AKT pathway. J Am Heart Assoc 12(17):e028185

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Litvinukova M, Talavera-Lopez C, Maatz H, Reichart D, Worth CL, Lindberg EL et al (2020) Cells of the adult human heart. Nature 588(7838):466–472

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu F, Tan A, Yang R, Xue Y, Zhang M, Chen L et al (2017) C1ql1/Ctrp14 and C1ql4/Ctrp11 promote angiogenesis of endothelial cells through activation of ERK1/2 signal pathway. Mol Cell Biochem 424(1–2):57–67

    Article  CAS  PubMed  Google Scholar 

  • Liu X, Chen W, Li W, Li Y, Priest JR, Zhou B et al (2019) Single-cell RNA-seq of the developing cardiac outflow tract reveals convergent development of the vascular smooth muscle cells. Cell Rep 28(5):1346–61 e4

    Article  CAS  PubMed  Google Scholar 

  • Long X, Yuan X, Du J (2023) Single-cell and spatial transcriptomics: advances in heart development and disease applications. Comput Struct Biotechnol J 21:2717–2731

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lu F, Langenbacher A, Chen JN (2017) Tbx20 drives cardiac progenitor formation and cardiomyocyte proliferation in zebrafish. Dev Biol 421(2):139–148

    Article  CAS  PubMed  Google Scholar 

  • Luo H, Li Q, Pramanik J, Luo J, Guo Z (2014) Nanog expression in heart tissues induced by acute myocardial infarction. Histol Histopathol 29(10):1287–1293

    CAS  PubMed  Google Scholar 

  • Lupu IE, Redpath AN, Smart N (2020) Spatiotemporal analysis reveals overlap of key proepicardial markers in the developing murine heart. Stem Cell Rep 14(5):770–787

    Article  CAS  Google Scholar 

  • Ma L, Li J, Liu Y, Pang S, Huang W, Yan B (2013) Novel and functional variants within the TBX18 gene promoter in ventricular septal defects. Mol Cell Biochem 382(1–2):121–126

    Article  CAS  PubMed  Google Scholar 

  • Mahmoud AI, Kocabas F, Muralidhar SA, Kimura W, Koura AS, Thet S et al (2013) Meis1 regulates postnatal cardiomyocyte cell cycle arrest. Nature 497(7448):249–253

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Martucciello S, Turturo MG, Bilio M, Cioffi S, Chen L, Baldini A et al (2020) A dual role for Tbx1 in cardiac lymphangiogenesis through genetic interaction with Vegfr3. FASEB J 34(11):15062–15079

    Article  CAS  PubMed  Google Scholar 

  • Onizuka T, Yuasa S, Kusumoto D, Shimoji K, Egashira T, Ohno Y et al (2012) Wnt2 accelerates cardiac myocyte differentiation from ES-cell derived mesodermal cells via non-canonical pathway. J Mol Cell Cardiol 52(3):650–659

    Article  CAS  PubMed  Google Scholar 

  • Osorio D, Cai JJ (2021) Systematic determination of the mitochondrial proportion in human and mice tissues for single-cell RNA-sequencing data quality control. Bioinformatics 37(7):963–967

    Article  CAS  PubMed  Google Scholar 

  • Palomer X, Román-Azcona MS, Pizarro-Delgado J, Planavila A, Villarroya F, Valenzuela-Alcaraz B et al (2020) SIRT3-mediated inhibition of FOS through histone H3 deacetylation prevents cardiac fibrosis and inflammation. Signal Transduct Target Ther 5(1):14

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Qin G, Bian Z, Liao H, Zhang Y, Wu Q, Zhou H et al (2014) Never in mitosis gene A related kinase-6 attenuates pressure overload-induced activation of the protein kinase B pathway and cardiac hypertrophy. PLoS ONE 9(4):e96095

    Article  ADS  Google Scholar 

  • Qiu X, Mao Q, Tang Y, Wang L, Chawla R, Pliner HA et al (2017a) Reversed graph embedding resolves complex single-cell trajectories. Nat Methods 14(10):979–982

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Qiu X, Hill A, Packer J, Lin D, Ma YA, Trapnell C (2017b) Single-cell mRNA quantification and differential analysis with Census. Nat Methods 14(3):309–315

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Qiu XB, Qu XK, Li RG, Liu H, Xu YJ, Zhang M et al (2017c) CASZ1 loss-of-function mutation contributes to familial dilated cardiomyopathy. Clin Chem Lab Med 55(9):1417–1425

    Article  CAS  PubMed  Google Scholar 

  • Qu X, Liu Y, Cao D, Chen J, Liu Z, Ji H et al (2019) BMP10 preserves cardiac function through its dual activation of SMAD-mediated and STAT3-mediated pathways. J Biol Chem 294(52):19877–19888

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sahara M, Santoro F, Sohlmer J, Zhou C, Witman N, Leung CY et al (2019) Population and single-cell analysis of human cardiogenesis reveals unique LGR5 ventricular progenitors in embryonic outflow tract. Dev Cell. 48(4):475–90 e7

    Article  CAS  PubMed  Google Scholar 

  • Sanchez-Fernandez C, Rodriguez-Outeirino L, Matias-Valiente L, Ramirez de Acuna F, Hernandez-Torres F, Lozano-Velasco E et al (2022) Regulation of epicardial cell fate during cardiac development and disease: an overview. Int J Mol Sci. 23(6):3220

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Seidenberg J, Stellato M, Hukara A, Ludewig B, Klingel K, Distler O et al (2021) The AP-1 transcription factor Fosl-2 regulates autophagy in cardiac fibroblasts during myocardial fibrogenesis. Int J Mol Sci 22(4):1861

  • Singh MK, Li Y, Li S, Cobb RM, Zhou D, Lu MM et al (2010) Gata4 and Gata5 cooperatively regulate cardiac myocyte proliferation in mice. J Biol Chem 285(3):1765–1772

    Article  CAS  PubMed  Google Scholar 

  • Song SE, Kim YW, Kim JY, Lee DH, Kim JR, Park SY (2013) IGFBP5 mediates high glucose-induced cardiac fibroblast activation. J Mol Endocrinol 50(3):291–303

    Article  CAS  PubMed  Google Scholar 

  • Srinivas S, Watanabe T, Lin CS, William CM, Tanabe Y, Jessell TM et al (2001) Cre reporter strains produced by targeted insertion of EYFP and ECFP into the ROSA26 locus. BMC Dev Biol 1:4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Srivastava D (2006) Making or breaking the heart: from lineage determination to morphogenesis. Cell 126(6):1037–1048

    Article  CAS  PubMed  Google Scholar 

  • Stefanovic S, Laforest B, Desvignes JP, Lescroart F, Argiro L, Maurel-Zaffran C et al (2020) Hox-dependent coordination of mouse cardiac progenitor cell patterning and differentiation. Elife 9:e55124

  • Steimle JD, Rankin SA, Slagle CE, Bekeny J, Rydeen AB, Chan SS et al (2018) Evolutionarily conserved Tbx5-Wnt2/2b pathway orchestrates cardiopulmonary development. Proc Natl Acad Sci U S A 115(45):E10615–E10624

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Stephen LJ, Fawkes AL, Verhoeve A, Lemke G, Brown A (2007) A critical role for the EphA3 receptor tyrosine kinase in heart development. Dev Biol 302(1):66–79

    Article  CAS  PubMed  Google Scholar 

  • Stuart T, Butler A, Hoffman P, Hafemeister C, Papalexi E, Mauck WM 3rd et al (2019) Comprehensive integration of single-cell data. Cell 177(7):1888–902 e21

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sun LY, Zhao JC, Ge XM, Zhang H, Wang CM, Bie ZD (2020) Circ_LAS1L regulates cardiac fibroblast activation, growth, and migration through miR-125b/SFRP5 pathway. Cell Biochem Funct 38(4):443–450

    Article  CAS  PubMed  Google Scholar 

  • Sun J, Guo X, Yu P, Liang J, Mo Z, Zhang M et al (2022a) Vasorin deficiency leads to cardiac hypertrophy by targeting MYL7 in young mice. J Cell Mol Med 26(1):88–98

    Article  CAS  PubMed  Google Scholar 

  • Sun J, Peterson EA, Wang AZ, Ou J, Smith KE, Poss KD et al (2022) hapln1 defines an epicardial cell subpopulation required for cardiomyocyte expansion during heart morphogenesis and regeneration. Circulation. https://doi.org/10.1161/CIRCULATIONAHA.121.055468

  • Teittinen KJ, Grönroos T, Parikka M, Junttila S, Uusimäki A, Laiho A et al (2012) SAP30L (Sin3A-associated protein 30-like) is involved in regulation of cardiac development and hematopoiesis in zebrafish embryos. J Cell Biochem 113(12):3843–3852

    Article  CAS  PubMed  Google Scholar 

  • Teng L, Huang Y, Guo J, Li B, Lin J, Ma L et al (2020) Cardiac fibroblast miR-27a may function as an endogenous anti-fibrotic by negatively regulating early growth response protein 3 (EGR3). J Cell Mol Med 25(1):73–83

    Article  PubMed  PubMed Central  Google Scholar 

  • Toran JL, Lopez JA, Gomes-Alves P, Aguilar S, Torroja C, Trevisan-Herraz M et al (2019) Definition of a cell surface signature for human cardiac progenitor cells after comprehensive comparative transcriptomic and proteomic characterization. Sci Rep 9(1):4647

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  • Trivedi CM, Zhu W, Wang Q, Jia C, Kee HJ, Li L et al (2010) Hopx and Hdac2 interact to modulate Gata4 acetylation and embryonic cardiac myocyte proliferation. Dev Cell 19(3):450–459

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Van de Sande B, Flerin C, Davie K, De Waegeneer M, Hulselmans G, Aibar S et al (2020) A scalable SCENIC workflow for single-cell gene regulatory network analysis. Nat Protoc 15(7):2247–2276

    Article  PubMed  Google Scholar 

  • van den Berg ME, Warren HR, Cabrera CP, Verweij N, Mifsud B, Haessler J et al (2017) Discovery of novel heart rate-associated loci using the Exome Chip. Hum Mol Genet 26(12):2346–2363

    Article  PubMed  PubMed Central  Google Scholar 

  • Varma E, Burghaus J, Schwarzl T, Sekaran T, Gupta P, Górska AA et al (2023) Translational control of Ybx1 expression regulates cardiac function in response to pressure overload in vivo. Basic Res Cardiol 118(1):25

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vincentz JW, McWhirter JR, Murre C, Baldini A, Furuta Y (2005) Fgf15 is required for proper morphogenesis of the mouse cardiac outflow tract. Genesis 41(4):192–201

    Article  CAS  PubMed  Google Scholar 

  • Wagner N, Wagner K-D (2021) Every beat you take—the Wilms′ tumor suppressor WT1 and the heart. Int J Mol Sci 22(14):7675

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang G, Xiong Z, Yang F, Zheng X, Zong W, Li R et al (2022) Identification of COVID-19-associated DNA methylation variations by integrating methylation array and scRNA-seq data at cell-type resolution. Genes (basel). 13(7):1109

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang F, Zhao H, Yin L, Zhang W, Tang Y, Wang X et al (2022) The paired-related homeobox protein 1 promotes cardiac fibrosis via the Twist1-Prrx1-tenascin-C loop. Cell Biol Int 47(1):167–177

    Article  PubMed  Google Scholar 

  • Wu Y, Liu X, Zheng H, Zhu H, Mai W, Huang X et al (2020) Multiple roles of sFRP2 in cardiac development and cardiovascular disease. Int J Biol Sci 16(5):730–738

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wu HY, Zhou YM, Liao ZQ, Zhong JW, Liu YB, Zhao H et al (2021a) Fosl1 is vital to heart regeneration upon apex resection in adult Xenopus tropicalis. NPJ Regen Med 6(1):36

    Article  PubMed  PubMed Central  Google Scholar 

  • Wu Q, Chen Q, Wang J, Fan D, Zhou H, Yuan Y et al (2021b) Long non-coding RNA Pvt1 modulates the pathological cardiac hypertrophy via miR-196b-mediated OSMR regulation. Cell Signal 86:110077

    Article  CAS  PubMed  Google Scholar 

  • Wu F, Huang W, Tan Q, Guo Y, Cao Y, Shang J et al (2021c) ZFP36L2 regulates myocardial ischemia/reperfusion injury and attenuates mitochondrial fusion and fission by LncRNA PVT1. Cell Death Dis 12(6):614

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xia Y, Duca S, Perder B, Dündar F, Zumbo P, Qiu M et al (2022) Activation of a transient progenitor state in the epicardium is required for zebrafish heart regeneration. Nat Commun 13(1):7704

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Xia M, Luo W, Jin H, Yang Z (2019) HAND2-mediated epithelial maintenance and integrity in cardiac outflow tract morphogenesis. Development 146(13):dev177477

  • Xiao L, Peng H, Yan M, Chen S (2021) Silencing ACTG1 expression induces prostate cancer epithelial mesenchymal transition through MAPK/ERK signaling pathway. DNA Cell Biol 40(11):1445–1455

    Article  CAS  PubMed  Google Scholar 

  • Xiong H, He A (2020) Single-cell transcriptomic analysis of cardiac progenitor differentiation. Curr Cardiol Rep 22(6):38

    Article  PubMed  Google Scholar 

  • Xiong H, Luo Y, Yue Y, Zhang J, Ai S, Li X et al (2019) Single-cell transcriptomics reveals chemotaxis-mediated intraorgan crosstalk during cardiogenesis. Circ Res 125(4):398–410

    Article  CAS  PubMed  Google Scholar 

  • Xu Y, Lv X, Cai R, Ren Y, He S, Zhang W et al (2022) Possible implication of miR-142-3p in coronary microembolization induced myocardial injury via ATXN1L/HDAC3/NOL3 axis. J Mol Med (berl) 100(5):763–780

    Article  CAS  PubMed  Google Scholar 

  • Yan S, Jiao K (2016) Functions of miRNAs during mammalian heart development. Int J Mol Sci. 17(5):789

    Article  PubMed  PubMed Central  Google Scholar 

  • Yang J, Hou Y, Zhou M, Wen S, Zhou J, Xu L et al (2016) Twist induces epithelial-mesenchymal transition and cell motility in breast cancer via ITGB1-FAK/ILK signaling axis and its associated downstream network. Int J Biochem Cell Biol 71:62–71

    Article  CAS  PubMed  Google Scholar 

  • Yoshitomi Y, Ikeda T, Saito-Takatsuji H, Yonekura H (2021) Emerging role of AP-1 transcription factor JunB in angiogenesis and vascular development. Int J Mol Sci 22(6):2804

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yu X, Chen X, Amrute-Nayak M, Allgeyer E, Zhao A, Chenoweth H et al (2021) MARK4 controls ischaemic heart failure through microtubule detyrosination. Nature 594(7864):560–565

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Yuan X, Zhang L, Du J (2021) Tbx18-positive cells-derived myofibroblasts contribute to renal interstitial fibrosis via transforming growth factor-β signaling. Exp Cell Res 405(2):112682

    Article  CAS  PubMed  Google Scholar 

  • Zhang S-M, Zhu L-H, Chen H-Z, Zhang R, Zhang P, Jiang D-S et al (2014) Interferon regulatory factor 9 is critical for neointima formation following vascular injury. Nat Commun 5(1):5160

    Article  ADS  CAS  PubMed  Google Scholar 

  • Zhong X, Wang T, Xie Y, Wang M, Zhang W, Dai L et al (2021) Activated protein C ameliorates diabetic cardiomyopathy modulating OTUB1/YB-1/MEF2B axis. Front Cardiovasc Med 8:758158

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhou L, Liu J, Olson P, Zhang K, Wynne J, Xie L (2015) Tbx5 and Osr1 interact to regulate posterior second heart field cell cycle progression for cardiac septation. J Mol Cell Cardiol 85:1–12

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhu B, Li H, Zhang L, Chandra SS, Zhao H (2022) A Markov random field model-based approach for differentially expressed gene detection from single-cell RNA-seq data. Brief Bioinform 23(5):bbac166

Download references

Acknowledgements

We thank Haijun Deng for his assistance with bioinformatic analysis and the processing of single-cell RNA-seq data. And we thank the other members of the Tbx18-positive cardiac cell study group from the Second Affiliated Hospital of Chongqing Medical University for their assistance: Qiang She, Songbai Deng, Xiaodong Jing, and Jing Wang. Figure 1 was created with BioRender.com.

Funding

This work was supported by grants from the Natural Science Foundation of Chongqing Science and Technology Commission (cstc2020jcyj-msxmX0210), Future Medicine Youth Innovation Team Development Support Program of Chongqing Medical University (W0133), High-End Medical Talents Project of Middle-Aged and Young People in Chongqing (JianlinDu [2022]) and Kuanren Talents Program of the Second Affiliated Hospital of Chongqing Medical University.

Author information

Authors and Affiliations

  1. Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China

    Xianglin Long, Jiangjun Wei, Qinghua Fang & Jianlin Du

  2. Department of Nephrology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China

    Xin Yuan

Authors
  1. Xianglin Long
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jiangjun Wei
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Qinghua Fang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xin Yuan
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jianlin Du
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Xianglin Long: methodology, software, writing-original draft, and investigation. Jiangjun Wei: software and writing-original draft. Qinghua Fang: visualization and software. Xin Yuan: writing-review and editing and project administration. Jianlin Du: conceptualization, resources, writing-review and editing, supervision, project administration, and funding acquisition. All authors contributed to the article and approved the submitted version.

Corresponding authors

Correspondence to Xin Yuan or Jianlin Du.

Ethics declarations

Ethics approval and consent to participate

All animal experiments were maintained according to the Regulation for the Administration of Affairs Concerning Experimental Animals (State Council of China, 2017 Revision), and our study was approved by the Commission of Chongqing Medical University for the Ethics of Animal Experiments.

Competing interests

The authors declare no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

10142_2024_1290_MOESM1_ESM.jpg

Supplementary Fig. 1 Heatmap showing the row-scaled expression of the top DEGs per cluster for all Tbx18+ cardiac cells (JPG 12929 KB)

10142_2024_1290_MOESM2_ESM.jpg

Supplementary Fig. 2 (A) Feature plots displayed the expression of the ECM-related genes of C0 to C9 clusters. (B, C) The GO and KEGG enrichment analysis of the C1 cluster. (D, E) The GO and KEGG enrichment analysis of the C5 cluster. The “Rich_factor” is the ratio of differentially expressed gene numbers annotated in this pathway term to all gene numbers annotated in this pathway term. The greater the Rich factor, the greater the degree of pathway enrichment. The values of rich factors are expressed as percentages (%) (JPG 4263 KB)

10142_2024_1290_MOESM3_ESM.jpg

Supplementary Fig. 3 (A, B) The GO and KEGG enrichment analysis of the MEM stage cells. (C, D) The GO and KEGG enrichment analysis of the ML_P0 stage cells. (E, F) The GO and KEGG enrichment analysis of the P-stage cells. The “Rich_factor” is the ratio of differentially expressed gene numbers annotated in this pathway term to all gene numbers annotated in this pathway term. The greater the Rich factor, the greater the degree of pathway enrichment. The values of rich factors are expressed as percentages (%) (JPG 8687 KB)

Supplementary Fig. 4 (A-C) The feature plots of selected genes' expression constructed by Monocle3 (JPG 3326 KB)

10142_2024_1290_MOESM5_ESM.jpg

Supplementary Fig. 5 (A-D) The Regulon specificity score map showed each stage's top 5 specific regulons. (E–F) The TF Twist regulon expression and activities were quantified using AUCell (JPG 3765 KB)

Supplementary Fig. 6 (A-D) The TFs' regulon expression and activities were quantified using AUCell (JPG 4412 KB)

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Long, X., Wei, J., Fang, Q. et al. Single-cell RNA sequencing reveals the transcriptional heterogeneity of Tbx18-positive cardiac cells during heart development. Funct Integr Genomics 24, 18 (2024). https://doi.org/10.1007/s10142-024-01290-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s10142-024-01290-6

Keywords

Search

Navigation