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Micropattern platform promotes extracellular matrix remodeling by human PSC-derived cardiac fibroblasts and enhances contractility of co-cultured cardiomyocytes.

ABSTRACT: In native heart tissue, cardiac fibroblasts provide the structural framework of extracellular matrix (ECM) while also influencing the electrical and mechanical properties of cardiomyocytes. Recent advances in the field of stem cell differentiation have led to the availability of human pluripotent stem cell-derived cardiac fibroblasts (iPSC-CFs) in addition to cardiomyocytes (iPSC-CMs). Here we use a novel 2D in vitro micropatterned platform that provides control over ECM geometry and substrate stiffness. When cultured alone on soft micropatterned substrates, iPSC-CFs are confined to the micropatterned features and remodel the ECM into anisotropic fibers. Similar remodeling and ECM production occurs when cultured with iPSC-CMs in a co-culture model. In addition to modifications in the ECM, our results show that iPSC-CFs influence iPSC-CM function with accelerated Ca²⁺ transient rise-up time and greater contractile strains in the co-culture conditions compared to when iPSC-CMs are cultured alone. These combined observations highlight the important role cardiac fibroblasts play in vivo and the need for co-culture models like the one presented here to provide more representative in vitro cardiac constructs.

SUBMITTER: Napiwocki BN

PROVIDER: S-EPMC8496154 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND: Celiac disease (CD) is mainly characterised by villous atrophy and mucosal architectural rearrangement. The fibroblasts (FBs) are the most abundant mesenchymal cell type in the intestinal mucosa and are responsible for both the architectural arrangement of the villi and the formation of the extracellular matrix (ECM). This study aimed at the evaluation of both the intracellular distribution of different proteins involved in ECM and FBs characterisation, and the cellular displacement of primary FBs obtained from duodenal endoscopic biopsies of healthy subjects and celiac patients. METHODS: Primary healthy and celiac duodenal FBs were evaluated by means of immuno-fluorescence assay for collagen type I and IV, fibronectin, actin, alpha-Smooth Muscle Actin (alpha-SMA), Fibroblast Surface Protein (FSP) and transglutaminase type 2 (TG2). The geometric indexes of the fluorescence signals were investigated by image analysis software (Image J, NIH). Both morphology and kinetic were evaluated during a 72 hours time course movie. TG2 medium activity was evaluated by means of ELISA. RESULTS: All the cells examined were immunopositive for FSP, alpha-SMA, actin, collagen I, collagen IV and TG2. CD cells showed a signet collagen-I and collagen-IV pattern, as compared to the controls being characterised by a spindle geometry. Moreover, the collagen signals in CD FBs showed a significantly higher circularity index (major orthogonal diameter ratio) than the controls (p<0.0001), whereas the perimeter and area ratio were significantly lower (p<0.0001). The TG2 signal had a decreased area (p<0.05), but a two-fold increased medium activity. The time course highlighted a reduction of the displacement of CD FBs. CONCLUSIONS: The isolated primary CD FBs showed a different collagen and TG2 pattern of distribution associated with a different cellular displacement. The reasons for such CD cell peculiar characteristics are yet unknown but they might represent a factor in the progression of the intestinal damage.

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Micropattern platform promotes extracellular matrix remodeling by human PSC-derived cardiac fibroblasts and enhances contractility of co-cultured cardiomyocytes.

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