Volume 13 Issue 8
Aug.  2023
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Article Navigation >  Journal of Pharmaceutical Analysis  > 2023  >  13(8): 894-907
Jianlin Du, Xin Yuan, Haijun Deng, Rongzhong Huang, Bin Liu, Tianhua Xiong, Xianglin Long, Ling Zhang, Yingrui Li, Qiang She. Single-cell and spatial heterogeneity landscapes of mature epicardial cells[J]. Journal of Pharmaceutical Analysis, 2023, 13(8): 894-907. doi: 10.1016/j.jpha.2023.07.011
Citation: Jianlin Du, Xin Yuan, Haijun Deng, Rongzhong Huang, Bin Liu, Tianhua Xiong, Xianglin Long, Ling Zhang, Yingrui Li, Qiang She. Single-cell and spatial heterogeneity landscapes of mature epicardial cells[J]. Journal of Pharmaceutical Analysis, 2023, 13(8): 894-907. doi: 10.1016/j.jpha.2023.07.011
shu

Single-cell and spatial heterogeneity landscapes of mature epicardial cells

doi: 10.1016/j.jpha.2023.07.011

  • a. Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China;

  • b. Department of Nephrology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China;

  • c. Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education, Institute for Viral Hepatitis, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China;

  • d. Precision Medicine Center, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China;

  • e. Basic Medicine Research and Innovation Center for Novel Target and Therapeutic Intervention, Ministry of Education, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China

Funds:

Tbx18: Cre mice were donated from Professor Sylvia M. Evans. We thank Dr. Andreas Kispert from Hannover Medical School (MHH) for providing probe plasmids of Tbx18. The computational work in this paper received partial support from the Supercomputing Center of Chongqing Medical University. This work was supported by grants from the National Natural Science Foundation of China (Grant No.: 82270281), Chongqing Medical University Program for Youth Innovation in Future Medicine (Grant No.: W0133), Senior Medical Talents Program of Chongqing for Young and Middle-aged, China (Program No.: JianlinDu [2022]), Postdoctoral Research Funding of the Second Affiliated Hospital of Chongqing Medical University, China (Grant No.: rsc-postdoctor114), and Kuanren Talents Program of the Second Affiliated Hospital of Chongqing Medical University, China (Program No.: kryc-gg-2102).

  • Received Date: Nov. 28, 2022
  • Rev Recd Date: Jul. 13, 2023
    Abstract
  • Tbx18, Wt1, and Tcf21 have been identified as epicardial markers during the early embryonic stage. However, the gene markers of mature epicardial cells remain unclear. Single-cell transcriptomic analysis was performed with the Seurat, Monocle, and CellphoneDB packages in R software with standard procedures. Spatial transcriptomics was performed on chilled Visium Tissue Optimization Slides (10x Genomics) and Visium Spatial Gene Expression Slides (10x Genomics). Spatial transcriptomics analysis was performed with Space Ranger software and R software. Immunofluorescence, whole-mount RNA in situ hybridization and X-gal staining were performed to validate the analysis results. Spatial transcriptomics analysis revealed distinct transcriptional profiles and functions between epicardial tissue and non-epicardial tissue. Several gene markers specific to postnatal epicardial tissue were identified, including Msln, C3, Efemp1, and Upk3b. Single-cell transcriptomic analysis revealed that cardiac cells from wildtype mouse hearts (from embryonic day 9.5 to postnatal day 9) could be categorized into six major cell types, which included epicardial cells. Throughout epicardial development, Wt1, Tbx18, and Upk3b were consistently expressed, whereas genes including Msln, C3, and Efemp1 exhibited increased expression during the mature stages of development. Pseudotime analysis further revealed two epicardial cell fates during maturation. Moreover, Upk3b, Msln, Efemp1, and C3 positive epicardial cells were enriched in extracellular matrix signaling. Our results suggested Upk3b, Efemp1, Msln, C3, and other genes were mature epicardium markers. Extracellular matrix signaling was found to play a critical role in the mature epicardium, thus suggesting potential therapeutic targets for heart regeneration in future clinical practice.
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  • E.J. Benjamin, M.J. Blaha, S.E. Chiuve et al., Heart disease and stroke statistics - 2017 update: report from the American heart association, Circulation 135 (2017) e146-e603.
    L. He, N.B. Nguyen, R. Ardehali, et al., Heart regeneration by endogenous stem cells and cardiomyocyte proliferation: ontroversy, fallacy, and progress, Circulation 142 (2020) 275-291.
    P. Quijada, M.A. Trembley, E.M. Small, The role of the epicardium during heart development and repair, Circ. Res. 126 (2020) 377-394.
    W. Ding, J. Li, F. Zheng, et al., The challenges of treating acute myocardial infarction due to variant angina: esson from an interesting case, 5 (2021) 213-218.
    J. Cao, K.D. Poss, The epicardium as a hub for heart regeneration, Nat. Rev. Cardiol. 15 (2018) 631-647.
    R. Marin-Juez, H. El-Sammak, C.S.M. Helker, et al., Coronary revascularization during heart regeneration is regulated by epicardial and endocardial cues and forms a scaffold for cardiomyocyte repopulation, Dev. Cell 51 (2019) 503-515.e4.
    B. Zhou, L.B. Honor, Q. Ma, et al., Thymosin beta 4 treatment after myocardial infarction does not reprogram epicardial cells into cardiomyocytes, J. Mol. Cell. Cardiol. 52 (2012) 43-47.
    L. Zangi, K.O. Lui, A. von Gise, et al., Modified mRNA directs the fate of heart progenitor cells and induces vascular regeneration after myocardial infarction, Nat. Biotechnol. 31 (2013) 898-907.
    Y. Li, Y. Li, X. Jing, et al., Sphingosine 1-phosphate induces epicardial progenitor cell differentiation into smooth muscle-like cells, Acta Biochim. Biophys. Sin. 51 (2019) 402-410.
    T. Moore-Morris, P. Cattaneo, M. Puceat, et al., Origins of cardiac fibroblasts, J. Mol. Cell. Cardiol. 91 (2016) 1-5.
    R. Carmona, S. Barrena, A.J. Lopez Gambero, et al., Epicardial cell lineages and the origin of the coronary endothelium, FASEB J. 34 (2020) 5223-5239.
    B. Liu, D. Wang, T. Xiong, et al., Inhibition of Notch signaling promotes the differentiation of epicardial progenitor cells into adipocytes, Stem Cells Int. 2021 (2021), 8859071.
    C.M. Braitsch, M.D. Combs, S.E. Quaggin, et al., Pod, Dev. Biol. 368 (2012) 345-357.
    J. Hesse, C. Owenier, T. Lautwein, et al., Single-cell transcriptomics defines heterogeneity of epicardial cells and fibroblasts within the infarcted murine heart, 10 (2021), e65921.
    C. Cai, J.C. Martin, Y. Sun, et al., A myocardial lineage derives from Tbx18 epicardial cells, Nature 454 (2008) 104-108.
    P. Soriano, Generalized lacZ expression with the ROSA26 Cre reporter strain, Nat. Genet. 21 (1999) 70-71.
    S. Srinivas, T. Watanabe, C.S. Lin, et al., Cre reporter strains produced by targeted insertion of EYFP and ECFP into the ROSA26 locus, BMC Dev. Biol. 1 (2001), 4.
    F. Kraus, B. Haenig, A. Kispert, Cloning and expression analysis of the mouse T-box gene Tbx18, Mech. Dev. 100 (2001) 83-86.
    D. Piette, M. Hendrickx, E. Willems, et al., An optimized procedure for whole-mount in situ hybridization on mouse embryos and embryoid bodies, Nat. Protoc. 3 (2008) 1194-1201.
    X. Jing, Y. Gao, S. Xiao, et al., Hypoxia induced the differentiation of Tbx18-positive epicardial cells to CoSMCs, Sci. Rep. 6 (2016), 30468.
    C. Hafemeister, R. Satija, Normalization and variance stabilization of single-cell RNA-seq data using regularized negative binomial regression, Genome Biol. 20 (2019), 296.
    W. Feng, A. Bais, H. He, et al., Single-cell transcriptomic analysis identifies murine heart molecular features at embryonic and neonatal stages, Nat. Commun. 13 (2022), 7960.
    T. Stuart, A. Butler, P. Hoffman, et al., Comprehensive integration of single-cell data, Cell 177 (2019) 1888-1902.e21.
    X. Qiu, A. Hill, J. Packer, et al., Single-cell mRNA quantification and differential analysis with Census, Nat. Methods 14 (2017) 309-315.
    G. Yu, L. Wang, Y. Han, et al., clusterProfiler: n R package for comparing biological themes among gene clusters, OMICS 16 (2012) 284-287.
    M. Efremova, M. Vento-Tormo, S.A. Teichmann, et al., : nferring cell-cell communication from combined expression of multi-subunit ligand-receptor complexes, Nat. Protoc. 15 (2020) 1484-1506.
    S. Durinck, P.T. Spellman, E. Birney, et al., Mapping identifiers for the integration of genomic datasets with the R/Bioconductor package biomaRt, Nat. Protoc. 4 (2009) 1184-1191.
    S. Jin, C.F. Guerrero-Juarez, L. Zhang, et al., Inference and analysis of cell-cell communication using , Nat. Commun. 12 (2021), 1088.
    Zenodo, zktuong/ktplots: v1.2.3. https://doi.org/10.5281/zenodo.7699617. (Accessed 28 June 2023).
    X. Yuan, L. Zhang, J. Du, Tbx18-positive cells-derived myofibroblasts contribute to renal interstitial fibrosis via transforming growth factor-β signaling, Exp. Cell Res. 405 (2021), 112682.
    D. Pu, J. Du, J. Zhang, et al., An economical and practical method for whole-mount in situ hybridization for mouse embryos and organs, Biotech. Histochem. 88 (2013) 27-37.
    T. Grieskamp, C. Rudat, T.H. Ludtke, et al., Notch signaling regulates smooth muscle differentiation of epicardium-derived cells, Circ. Res. 108 (2011) 813-823.
    J. Niderla-BieliNska, E. Jankowska-Steifer, A. Flaht-Zabost, et al., : urrent understanding of its structure, induction, and fate, Anat. Rec. 302 (2019) 893-903.
    V.R. Knight-Schrijver, H. Davaapil, S. Bayraktar, et al., A single-cell comparison of adult and fetal human epicardium defines the age-associated changes in epicardial activity, 1 (2022) 1215-1229.
    B. Zeng, X. Ren, F. Cao, et al., Developmental patterns and characteristics of epicardial cell markers Tbx18 and Wt1 in murine embryonic heart, J. Biomed. Sci. 18 (2011), 67.
    P. Tandon, Y.V. Miteva, L.M. Kuchenbrod, et al., Tcf21 regulates the specification and maturation of proepicardial cells, 140 (2013) 2409-2421.
    I.E. Lupu, A.N. Redpath, N. Smart, Spatiotemporal analysis reveals overlap of key proepicardial markers in the developing murine heart, . 14 (2020) 770-787.
    L. Bochmann, P. Sarathchandra, F. Mori, et al., Revealing new mouse epicardial cell markers through transcriptomics, PLoS One 5 (2010), e11429.
    C. MacNeill, R. French, T. Evans, et al., Modular regulation of cGATA-5 gene expression in the developing heart and gut, Dev. Biol. 217 (2000) 62-76.
    S.K. Longo, M.G. Guo, A.L. Ji, et al., Integrating single-cell and spatial transcriptomics to elucidate intercellular tissue dynamics, Nat. Rev. Genet. 22 (2021) 627-644.
    L.S. Rodgers, S. Lalani, R.B. Runyan, et al., Differential growth and multicellular villi direct proepicardial translocation to the developing mouse heart, 237 (2008) 145-152.
    R. Carmona, J.A. Guadix, E. Cano, et al., The embryonic epicardium: n essential element of cardiac development, 14 (2010) 2066-2072.
    P. Quijada, M.A. Trembley, A. Misra, et al., Coordination of endothelial cell positioning and fate specification by the epicardium, Nat. Commun. 12 (2021), 4155.
    Y. Cao, S. Duca, J. Cao, Epicardium in heart development, 12 (2020), a037192.
    M. Weinberger, F.C. Simoes, R. Patient, et al., Functional heterogeneity within the developing zebrafish epicardium, Dev. Cell 52 (2020) 574-590.e6.
    L. Gambardella, S.A. McManus, V. Moignard, et al., BNC1 regulates cell heterogeneity in human pluripotent stem cell derived-epicardium, . 146 (2019), dev174441.
    D. Li, J. Sun, T.P. Zhong, Wnt signaling in heart development and regeneration, Curr. Cardiol. Rep. 24 (2022) 1425-1438.
    Y. Nakajima, K.I. Yoshida, New insights into the developmental mechanisms of coronary vessels and epicardium. , Vol. 303, Elsevier B.V., Amsterdam, 2013, pp. 263-317.
    K.P. Hanson, J.P. Jung, Q.A. Tran, et al., Spatial and temporal analysis of extracellular matrix proteins in the developing murine heart: blueprint for regeneration, Tissue Eng. 19 (2013) 1132-1143.
    A.C. Silva, C. Pereira, A.C.R.G. Fonseca, et al., Bearing my heart: he role of extracellular matrix on cardiac development, homeostasis, and injury response, Front. Cell Dev. Biol. 8 (2021), 621644.
    J. Jiao, Y. Dang, Y. Yang, et al., Promoting reprogramming by FGF2 reveals that the extracellular matrix is a barrier for reprogramming fibroblasts to pluripotency, 31 (2013) 729-740.
    X. Wu, X. Kong, A. Pellicer, et al., Uroplakins in urothelial biology, function, and disease, . 75 (2009) 1153-1165.
    F. Deng, F. Liang, L. Tu, et al., Uroplakin IIIb, a urothelial differentiation marker, dimerizes with uroplakin Ib as an early step of urothelial plaque assembly, J. Cell Biol. 159 (2002) 685-694.
    M.A. Matuszewski, K. Tupikowski, L. Dolowy, et al., Uroplakins and their potential applications in urology, 69 (2016) 252-257.
    A.R. Carpenter, M.B. Becknell, C.B. Ching, et al., Uroplakin 1b is critical in urinary tract development and urothelial differentiation and homeostasis, . 89 (2016) 612-624.
    S. Kuriyama, Y. Tamiya, M. Tanaka, Spatiotemporal expression of UPK3B and its promoter activity during embryogenesis and spermatogenesis, Histochem. Cell Biol. 147 (2017) 17-26.
    K. Norris, T. Hopes, J.L. Aspden, Ribosome heterogeneity and specialization in development, 12 (2021), e1644.
    E.S. Cenik, X. Meng, N.H. Tang, et al., Maternal ribosomes are sufficient for tissue diversification during embryonic development in C. elegans, Dev. Cell 48 (2019) 811-826.e6.
    I. Livingstone, V.N. Uversky, D. Furniss, et al., The pathophysiological significance of fibulin-3, Biomolecules 10 (2020), 1294.
    R. Giltay, R. Timpl, G. Kostka, Sequence, recombinant expression and tissue localization of two novel extracellular matrix proteins, fibulin-3 and fibulin-4, . 18 (1999) 469-480.
    J. Ehlermann, S. Weber, P. Pfisterer, et al., Cloning, expression and characterization of the murine Efemp1, a gene mutated in -Honeycomb retinal dystrophy, Gene Expr. Patterns 3 (2003) 441-447.
    W. Li, X. Lou, Y. Zha, et al., Single-cell RNA-seq of heart reveals intercellular communication drivers of myocardial fibrosis in diabetic cardiomyopathy, elife 12 (2023), e80479.
    H.W. Koh, A.P. Pilbrow, S.H. Tan, et al., An integrated signature of extracellular matrix proteins and a diastolic function imaging parameter predicts post-MI long-term outcomes, 10 (2023), 1123682.
    K. Chang, I. Pastan, Molecular cloning of mesothelin, a differentiation antigen present on mesothelium, mesotheliomas, and ovarian cancers, Proc. Natl. Acad. Sci. U 93 (1996) 136-140.
    Z. Tang, M. Qian, M. Ho, The role of mesothelin in tumor progression and targeted therapy, Anti Cancer Agents Med. Chem. 13 (2013) 276-280.
    G. Giordano E. Ferioli, A. Tafuni, The role of mesothelin expression in serous ovarian carcinoma: mpacts on diagnosis, prognosis, and therapeutic targets, Cancers 14 (2022), 2283.
    M.A. Missinato, K. Tobita, N. Romano, et al., Extracellular component hyaluronic acid and its receptor are required for epicardial EMT during heart regeneration, Cardiovasc. Res. 107 (2015) 487-498.
    P. Allison, D. Espiritu, J.V. Barnett, et al., Type III TGFβ receptor and Src direct hyaluronan-mediated invasive cell motility, Cell. Signal. 27 (2015) 453-459.
    S. Kapuria, H. Bai, J. Fierros, et al., Heterogeneous pdgfrb+ cells regulate coronary vessel development and revascularization during heart regeneration, 149 (2022), dev199752.
    S. Deng, X. Jing, X. Wei, et al., Triiodothyronine promotes the proliferation of epicardial progenitor cells through the MAPK/ERK pathway, Biochem. Biophys. Res. Commun. 486 (2017) 372-377.
    A. Onat, G. Can, R. Rezvani, et al., Complement C3 and cleavage products in cardiometabolic risk, Clin. Chim. Acta 412 (2011) 1171-1179.
    A. Gorelik, T. Sapir, L. Ben-Reuven, et al., Complement C3 affects Rac, Front. Mol. Neurosci. 11 (2018), 150.
    H. Huang, X. Zhou, Y. Liu, et al., Histone deacetylase inhibitor alleviates liver fibrosis by regulating hepatic stellate cell activation, Mol. Med. Rep. 23 (2021), 305.
    L. Bao, J. Zhou, V.M. Holers, et al., Excessive matrix accumulation in the kidneys of MRL/lpr lupus mice is dependent on complement activation, J. Am. Soc. Nephrol. 14 (2003) 2516-2525.
    O. Saifi, B. Ghandour, D. Jaalouk, et al., Myocardial regeneration: ole of epicardium and implicated genes, Mol. Biol. Rep. 46 (2019) 6661-6674.
    A.N. Redpath, N. Smart, Recapturing embryonic potential in the adult epicardium: rospects for cardiac repair, Stem Cells Transl. 10 (2021) 511-521.
    N. Smart, S. Bollini, K.N. Dube, et al., De novo cardiomyocytes from within the activated adult heart after injury, Nature 474 (2011) 640-644.
    J.M. Vieira, S. Howard, C. Villa del Campo, et al., BRG1-SWI/SNF-dependent regulation of the Wt1 transcriptional landscape mediates epicardial activity during heart development and disease, Nat. Commun. 8 (2017), 16034.
    K. Wei, V. Serpooshan, C. Hurtado, et al., Epicardial FSTL1 reconstitution regenerates the adult mammalian heart, Nature 525 (2015) 479-485.
    J. Bargehr, L.P. Ong, M. Colzani, et al., Epicardial cells derived from human embryonic stem cells augment cardiomyocyte-driven heart regeneration, Nat. Biotechnol. 37 (2019) 895-906.
    C. Rudat, T. Grieskamp, C. Rohr, et al., Upk3b is dispensable for development and integrity of urothelium and mesothelium, PLoS One 9 (2014), e112112.
    B. Zhou, L.B. Honor, H. He, et al., Adult mouse epicardium modulates myocardial injury by secreting paracrine factors, J. Clin. Invest. 121 (2011) 1894-1904.
    M. Litvinukova, C. Talavera-Lopez, H. Maatz, et al., Cells of the adult human heart, Nature 588 (2020) 466-472.
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