Gremlin1 repression-mediated mitochondrial hyperfunction contributes to TCE-induced zebrafish cardiac defects

Trichloroethylene (TCE) is a ubiquitous pollutant with potential capacity to induce congenital heart disease (CHD). However, the mechanisms underlying TCE-induced CHD is largely unraveled. Herein, we found that early-stage exposure to TCE induced significant cardiac defects characterized by elongated SV-BV distance, thinned myocardium and attenuated contractility. Gremlin1 encoding gene, grem1a, was screened out as a putative target of TCE and was expressed in heart since its appearance. Further, grem1a knockdown in zebrafish induced phenotypes generally similar to that of TCE-treated group, accompanying with the disarrangement of myofibril structure. Single-cell RNA sequencing depicted that mitochondria respiration in grem1a-repressed cardiomyocytes were greatly enhanced, ultimately leading to an abnormal branch from normal developmental trajectory. Accordingly, the results from AC16 and human pluripotent stem cell-derived cardiomyocytes showed that GREM1 repression increased mitochondrial content, ATP production, mitochondrial reactive oxygen species and mitochondrial membrane potential, which probably disrupted the process of myofibril expansion. Altogether, these results suggested that TCE-induced gremlin1 repression could prematurely stimulate mitochondrial hyperfunction, thereby hamper cardiomyocyte development and cause cardiac defects in zebrafish embryos. This study provided a novel insight of gremlin1 in the associations between TCE exposure burden and CHD risk and a better understanding of the etiology of environmental stressor-caused CHD. Cells from zebrafish whole body at 12 hpf were subjected to bulk RNA-seq for detection of responding genes to TCE treatment. After screening out grem1a as candidate target genes of TCE, we collected cells from zebrafish rostral half at 48 hpf for singl-cell RNA-seq in order to illustrate the function of grem1a in cardiomyocyte dysplasia induced by TCE.