Single-Cell RNA Sequencing Analysis Reveals a Crucial Role for CTHRC1 (Collagen Triple Helix Repeat Containing 1) Cardiac Fibroblasts After Myocardial Infarction

Abstract

Background: Cardiac fibroblasts (CF) have a central role in the ventricular remodeling process associated with different types of fibrosis. Recent studies have shown that fibroblasts do not respond homogeneously to heart injury. Due to the limited set of bona fide fibroblast markers, a proper characterization of fibroblast population heterogeneity in response to cardiac damage is still missing. The purpose of this study was to define the CF heterogeneity during ventricular remodeling and the underlying mechanisms that regulate their function. Methods: Collagen1α1-GFP + CF were characterized after myocardial infarction (MI) by single-cell and bulk RNA-seq, ATAC-seq and functional assays. Swine and patient samples were studied using bulk RNA-seq. Results: We identified and characterized a unique CF subpopulation that emerges after MI in mice. These activated fibroblasts exhibit a clear pro-fibrotic signature, express high levels of Collagen Triple Helix Repeat Containing 1 ( Cthrc1 ) and localize into the scar. Non-canonical TGF-β signaling and different transcription factors including SOX9 are important regulators mediating their response to cardiac injury. Moreover, the absence of CTHRC1 results in pronounced lethality due to ventricular rupture. Finally, a population of CF with a similar transcriptome was identified in a swine model of MI and in heart tissue from patients with MI and dilated cardiomyopathy. Conclusions: We report CF heterogeneity, their dynamics during the course of MI and redefine the CF that respond to cardiac injury and participate in myocardial remodeling. Our study identifies Cthrc1 as a novel regulator of the healing scar process, and as a target for future translational studies.

Comprehensive Integration of Single-Cell Transcriptional Profiling Reveals the Heterogeneities of Non-cardiomyocytes in Healthy and Ischemic Hearts

“…We then compared differentially expressed genes between Fibro_Myo and five other clusters and observed dramatic upregulation of Cthrc1, which has been confirmed to be a marker of myofibroblasts ( 32 ), and Ddah1 ( Figure 5E ). Some previous studies confirmed that Ddah1 plays a crucial role in the clearance of asymmetric dimethylarginine and monomethyl arginine, which are strongly associated with premature cardiovascular disease and death ( 33 ).…”

“…Additionally, GO analysis of Fibro_Myo with other FB clusters showed significant enrichment of prohealing pathways, such as actin filament organization, extracellular matrix (ECM) organization, and wound healing ( Figure 5D). We then compared differentially expressed genes between Fibro_Myo and five other clusters and observed dramatic upregulation of Cthrc1, which has been confirmed to be a marker of myofibroblasts (32), and Ddah1 ( Figure 5E). Some previous studies confirmed that Ddah1 plays a crucial role in the clearance of asymmetric dimethylarginine and monomethyl arginine, which are strongly associated with premature cardiovascular disease and death (33).…”

Identification of candidate biomarkers and therapeutic agents for heart failure by bioinformatics analysis

“…ITGB7, RASGRP1, CNR1, SLC2A1, SLC11A1, GPR84, SSTR5, KCNB1, GLUL (glutamate-ammonia ligase), BANK1, CACNA1E, LGR5, AQP3, SIGLEC7, SSTR2 and DNER (delta/notch like EGF repeat containing) could be an index for diabetes, but these genes might be responsible for progression of HF. Experiments show that expression of FAP (fibroblast activation protein alpha) [66], THBS4 [67], CD27 [68], LEF1 [69], CTHRC1 [70], ESR1 [71], CXCL9 [72], SERPINA3 [73], TRPC4 [74], F13A1 [75], PIK3C2A [76], KCNIP2 [77] and GPR4 [78] contributed to myocardial infarction. MFAP4 [79], ALOX15 [80], COL1A1 [81], APOA1 [82], PDE5A [83], CX3CR1 [84], THY1 [85], GREM1 [86], FMOD (fibromodulin) [87], NPPA (natriuretic peptide A) [88], LTBP2 [89], LUM (lumican) [90], IL34 [91], NRG1 [92], CXCL14 [93], CXCL10 [94], ACE (angiotensin I converting enzyme) [95], CFTR (ystic fibrosis transmembrane conductance regulator) [96], S100A8 [97], S100A9 [97], HP (haptoglobin) [98] [162], Zhang et al [163] and Chen et al [164] study indicated that the expression of CCL22, CCR1, FPR1, KNG1, CRISPLD2, CD38 and GPRC5A were linked with progression of ischemic heart disease.…”

“…ITGB7, RASGRP1, CNR1, SLC2A1, SLC11A1, GPR84, SSTR5, KCNB1, GLUL (glutamate-ammonia ligase), BANK1, CACNA1E, LGR5, AQP3, SIGLEC7, SSTR2 and DNER (delta/notch like EGF repeat containing) could be an index for diabetes, but these genes might be responsible for progression of HF. Experiments show that expression of FAP (fibroblast activation protein alpha) [66], THBS4 [67], CD27 [68], LEF1 [69], CTHRC1 [70], ESR1 [71], CXCL9 [72], SERPINA3 [73], TRPC4 [74], F13A1 [75], PIK3C2A [76], KCNIP2 [77] and GPR4 [78] contributed to myocardial infarction. MFAP4 [79], ALOX15 [80], COL1A1 [81], APOA1 [82], PDE5A [83] [154], OSMR (oncostatin M receptor) [155] and IL15RA [156] are involved in development of obesity, but these genes might be key for progression of HF.…”

Ex uno, plures–From One Tissue to Many Cells: A Review of Single-Cell Transcriptomics in Cardiovascular Biology

“…Additionally, the authors compared genetically diverse inbred mouse strains characterized by different reparative outcomes to highlight the cell composition and transcriptomic features associated with a higher frequency of cardiac rupture in the transition between the inflammatory and proliferative phases. Three studies selectively profiled the fibroblast component of the heart [ 116 , 117 , 118 ]. The first early study [ 116 ] used the Fluidigm C1 platform to profile several hundred CD31-CD45 cells isolated from the scar area of inducible reporter mice for Tcf21 or Postn , 7 days post-MI.…”