Cultured cardiac fibroblasts and myofibroblasts express Sushi Containing Domain 2 and assemble a unique fibronectin rich matrix

Schmuck, Eric G.; Roy, Sushmita; Dhillon, Anisa; Walker, Sydney; Spinali, Keith; Colevas, Sophia M.; Zhou, Tianhua; Chhabra, Gagan; Liu, Yuming; Sagar, Abdul Kader; Childs, Charlie J.; Kink, John A.; Eliceiri, Kevin W.; Hematti, Peiman; Raval, Amish N.

doi:10.1016/j.yexcr.2021.112489

Cited by 9 publications

(7 citation statements)

References 54 publications

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“…Immunofluorescence staining also showed similar overall expression of fibroblast specific protein 1 (FSP1), TE7, VIM, collagen type 1 alpha 1 chain (COL1A1, and FN in both populations ( Supplementary Figure S2 ). SUSD2 has been previously identified as enriched in left ventricular cardiac fibroblasts compared to dermal fibroblasts ( Schmuck et al, 2021 ). Interestingly, SUSD2 protein expression was only observed in the LVCFBs while minimal expression was observed in the VICs by Western blotting, suggesting that these two populations have unique molecular signatures and SUSD2 can be used for distinguishing between the two CFB populations ( Supplementary Figure S3 ).…”

Section: Resultsmentioning

confidence: 99%

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Identifying molecular and functional similarities and differences between human primary cardiac valve interstitial cells and ventricular fibroblasts

Floy

Shabnam

Givens

et al. 2023

Front. Bioeng. Biotechnol.

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Introduction: Fibroblasts are mesenchymal cells that predominantly produce and maintain the extracellular matrix (ECM) and are critical mediators of injury response. In the heart, valve interstitial cells (VICs) are a population of fibroblasts responsible for maintaining the structure and function of heart valves. These cells are regionally distinct from myocardial fibroblasts, including left ventricular cardiac fibroblasts (LVCFBs), which are located in the myocardium in close vicinity to cardiomyocytes. Here, we hypothesize these subpopulations of fibroblasts are transcriptionally and functionally distinct.Methods: To compare these fibroblast subtypes, we collected patient-matched samples of human primary VICs and LVCFBs and performed bulk RNA sequencing, extracellular matrix profiling, and functional contraction and calcification assays.Results: Here, we identified combined expression of SUSD2 on a protein-level, and MEOX2, EBF2 and RHOU at a transcript-level to be differentially expressed in VICs compared to LVCFBs and demonstrated that expression of these genes can be used to distinguish between the two subpopulations. We found both VICs and LVCFBs expressed similar activation and contraction potential in vitro, but VICs showed an increase in ALP activity when activated and higher expression in matricellular proteins, including cartilage oligomeric protein and alpha 2-Heremans-Schmid glycoprotein, both of which are reported to be linked to calcification, compared to LVCFBs.Conclusion: These comparative transcriptomic, proteomic, and functional studies shed novel insight into the similarities and differences between valve interstitial cells and left ventricular cardiac fibroblasts and will aid in understanding region-specific cardiac pathologies, distinguishing between primary subpopulations of fibroblasts, and generating region-specific stem-cell derived cardiac fibroblasts.

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Section: Resultsmentioning

confidence: 99%

“…We identified that the LVCFBs expressed SUSD2, a novel cultured CFB marker ( Schmuck et al, 2021 ), while the VICs did not. Furthermore, from our bulk RNA-sequencing results, we identified a panel of genes, RHOU , MEOX2 , and EBF2 to distinguish between VICs and LVCFBs at a transcript-level.…”

Section: Discussionmentioning

confidence: 99%

Identifying molecular and functional similarities and differences between human primary cardiac valve interstitial cells and ventricular fibroblasts

Floy

Shabnam

Givens

et al. 2023

Front. Bioeng. Biotechnol.

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“…The VWD domain is identified as one of the four domains of the von Willebrand factor gene, a multimeric glycoprotein that plays an important role in hemostasis (Ginsburg et al, 1985). The VWD domain is found in the ECM proteins such as vitellogenin (Sun et al, 2013), zonadhesin (Bi et al, 2003), otogelin (Avan et al, 2019), and Sushi domain containing 2 (Schmuck et al, 2021). Although the actual function of VWD motif has not been identified, one characteristic of the VWD domain is the RGD sequence.…”

Section: Discussionmentioning

confidence: 99%

von Willebrand factor D and EGF domains regulate ameloblast differentiation and enamel formation

Iwata

Kawarabayashi

Yoshizaki

et al. 2021

Journal Cellular Physiology

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Cell‐ and tissue‐specific extracellular matrix (ECM) composition plays an important role in organ development, including teeth, by regulating cell behaviors, such as cell proliferation and differentiation. Here, we demonstrate for the first time that von Willebrand factor D and epidermal growth factor (EGF) domains (Vwde), a previously uncharacterized ECM protein, is specifically expressed in teeth and regulates cell proliferation and differentiation in inner enamel epithelial cells (IEEs) and enamel formation. We identified the Vwde as a novel ECM protein through bioinformatics using the NCBI expressed sequence tag database for mice. Vwde complementary DNA encodes 1773 amino acids containing a signal peptide, a von Willebrand factor type D domain, and tandem calcium‐binding EGF‐like domains. Real‐time polymerase chain reaction demonstrated that Vwde is highly expressed in tooth tissue but not in other tissues including the brain, lung, heart, liver, kidney, and bone. In situ hybridization revealed that the IEEs expressed Vwde messenger RNA in developing teeth. Immunostaining showed that VWDE was localized at the proximal and the distal ends of the pericellular regions of the IEEs. Vwde was induced during the differentiation of mouse dental epithelium‐derived M3H1 cells. Vwde‐transfected M3H1 cells secreted VWDE protein into the culture medium and inhibited cell proliferation, whereas ameloblastic differentiation was promoted. Furthermore, Vwde increased the phosphorylation of extracellular signal‐regulated kinase 1/2 and protein kinase B and strongly induced the expression of the intercellular junction protein, N‐cadherin (Ncad). Interestingly, the suppression of endogenous Vwde inhibited the expression of Ncad. Finally, we created Vwde‐knockout mice using the CRISPR‐Cas9 system. Vwde‐null mice showed low mineral density, rough surface, and cracks in the enamel, indicating the enamel hypoplasia phenotype. Our findings suggest that Vwde assembling the matrix underneath the IEEs is essential for Ncad expression and enamel formation.

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“…Briefly, cECM (Tandem TM , Cellular Logistics Inc, Madison, WI) was produced from primary human cardiac fibroblasts cultured under high-density conditions followed by a decellularization procedure as previously described. 17, 18 The injectable cECM formulation was manufactured by milling cECM scaffolds into a fine particulate of approximately 80-micron diameter using a pharmaceutical bead mill. cECM particles were added to the cell suspension and the cells are allowed to bind at room temperature for 1 hour prior to injection.…”

Section: Methodsmentioning

confidence: 99%

“…Our group previously described a cardiac extracellular matrix (cECM) engineered biomaterial using high-density culture-expanded cardiac fibroblasts isolated from human cadaveric hearts. cECM is an acellular, uniquely fibronectin abundant, non-chemically cross-linked, non-hydrogel scaffold with immunomodulatory and angiogenic properties that can be formulated into particulates, reconstituted into solution, and administered as an injectable 17, 18 . cECM was shown to educate monocytes to a distinct population of macrophages that exhibit anti-inflammatory and pro-angiogenic features, 18 and can promote myocardial retention of mesenchymal stem cells in rodent heart disease models when co-delivered epicardially via an open chest.…”

Section: Introductionmentioning

confidence: 99%

Human iPSC-derived Committed Cardiac Progenitors Generate Cardiac Tissue Grafts in a Swine Ischemic Cardiomyopathy Model without Triggering Ventricular Arrhythmias

Raval,

Schmuck,

Roy

et al. 2024

Preprint

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Background: The adult human heart following a large myocardial infarction is unable to regenerate heart muscle and instead forms scar with the risk of progressive heart failure. Large animal studies have shown that intramyocardial injection of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) following a myocardial infarction result in cell grafts but also ventricular arrhythmias. We hypothesized that intramyocardial injection of committed cardiac progenitor cells (CCPs) derived from iPSCs, combined with cardiac fibroblast-derived extracellular matrix (cECM) to enhance cell retention will: i) form cardiomyocyte containing functional grafts, ii) be free of ventricular arrhythmias and iii) restore left ventricular contractility in a post-myocardial infarction (MI) cardiomyopathy swine model. Methods: hiPSCs were differentiated using bioreactors and small molecules to produce a population of committed cardiac progenitor cells (CCPs). MI was created using a coronary artery balloon occlusion and reperfusion model in Yucatan mini pigs. Four weeks later, epicardial needle injections of CCPs+cECM were performed in a small initial feasibility cohort, and then transendocardial injections (TEI) of CCPs+cECM, CCPs alone, cECM alone or vehicle control into the peri-infarct region in a larger randomized cohort. A 4-drug immunosuppression regimen was administered to prevent rejection of human CCPs. Arrhythmias were evaluated using implanted event recorders. Magnetic resonance imaging (MRI) and invasive pressure volume assessment were used to evaluate left ventricular anatomic and functional performance, including viability. Detailed histology was performed on the heart to detect human grafts. Results: A scalable biomanufacturing protocol was developed generating CCPs which can efficiently differentiate to cardiomyocytes or endothelial cells in vitro. Intramyocardial delivery of CCPs to post-MI porcine hearts resulted in engraftment and differentiation of CCPs to form ventricular cardiomyocyte rich grafts. There was no significant difference in cardiac MRI-based measured cardiac volumes or function between control, CCP and CCP+cECM groups; however, dobutamine stimulated functional reserve was improved in CCP and CCP+cECM groups. TEI delivery of CCPs with or without cECM did not result in tumors or trigger ventricular arrhythmias. Conclusions: CCPs are a promising cell source for post-MI heart repair using clinically relevant TEI with a low risk of engraftment ventricular arrhythmias.

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Cultured cardiac fibroblasts and myofibroblasts express Sushi Containing Domain 2 and assemble a unique fibronectin rich matrix

Cited by 9 publications

References 54 publications

Identifying molecular and functional similarities and differences between human primary cardiac valve interstitial cells and ventricular fibroblasts

Identifying molecular and functional similarities and differences between human primary cardiac valve interstitial cells and ventricular fibroblasts

von Willebrand factor D and EGF domains regulate ameloblast differentiation and enamel formation

Human iPSC-derived Committed Cardiac Progenitors Generate Cardiac Tissue Grafts in a Swine Ischemic Cardiomyopathy Model without Triggering Ventricular Arrhythmias

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