Impaired Cardiovascular Function Caused by Different Stressors Elicits a Common Pathological and Transcriptional Response in Zebrafish Embryos

Abstract

Zebrafish embryos have been widely used to study the genes and processes needed for normal vertebrate heart development. We recently observed that exposure to 2,3,7,8-tetra-chlorodibenzo-p-dioxin (TCDD) or retinoic acid (RA) produces very similar signs of heart failure in developing zebrafish via divergent molecular pathways. The fact that diverse stressors and mutations cause severe pericardial edema and circulatory collapse in developing zebrafish has been largely unexplored. We hypothesized that unrelated chemicals can trigger a common pathological response leading to the same end-stage heart failure. To test this hypothesis, we compared the effects of TCDD, RA, carbaryl, valproic acid, and morpholino oligonucleotide (MO) knockdown of TBX5 on the developing heart in zebrafish embryos. These model stressors have all been previously reported to affect zebrafish heart development, and elicited very similar signs of embryonic heart failure. Microarray analysis showed that one cluster of 92 transcripts affected by these different treatments was significantly downregulated by all treatments. This gene cluster is composed of transcripts required for chromosome assembly, DNA replication, and cell cycle progression. We refer to this cluster as the cell cycle gene cluster (CCGC). Immunohistochemistry revealed that downregulation of the CCGC precedes a halt in cardiomyocyte proliferation in the hearts of zebrafish exposed to any of the treatments. Previous work has shown that the initial response to TCDD is a decrease in cardiac output. Since this precedes the signs of edema, heart failure, and fall in CCGC expression, we postulated that any factor that decreases cardiac output will produce the same syndrome of heart failure responses. To test this, we used MO knockdown of cardiac troponin T2 (TNNT2) to specifically block contractility. The TNNT2-MO produced exactly the same signs of cardiotoxicity as the other treatments, including downregulation of the signature CCGC. Our results indicate that agents altering cardiac output can have amplified consequences during specific periods in development.

Comparative toxicological assessment of PAMAM and thiophosphoryl dendrimers using embryonic zebrafish

“…Pericardial edema is not necessarily a predicator of any specific pathway being triggered but rather an early stage stressor. 34 Pericardial edema in zebrafish has been associated with exposure to a wide variety of toxicants including polycyclic aromatic hydrocarbons (PAHs), 35 dioxins, 36 and gasoline oxygenates. 37 The PAMAM dendrimers had relatively high toxicity in comparison to the thiophosphoryl dendrimers, indicating higher risk for the combination of surface chemistry and generation studied here.…”

Zebrafish as a Vertebrate Model System to Evaluate Effects of Environmental Toxicants on Cardiac Development and Function

“…Heart development is regulated by molecular, cellular, and environmental factors [10,17,18]. Anatomically, the zebrafish heart is different from the human heart [10,19].…”

“…In recent decades, an ample number of transgenic zebrafish lines have been created that express fluorescence proteins labeling myocardium, endocardium, different cardiac lineages, blood, and the vascular system [22,23,37,38,39,40,41,42,43,44] (Table 1). There are also transgenic lines reporting different signaling pathways in cardiogenesis and providing over-expression or blockage of cardiac regulatory genes [17,18]. Together, transgenic zebrafish facilitate precise understanding of the effects of substances on cardiogenesis and cardiac functions.…”

“…There have been great efforts to establish zebrafish as a toxicology model organism [13,14,15,16]. Cardiovascular defects in response to various stressors and toxicants have been characterized using zebrafish, illustrating the potential utility of the zebrafish model [17,18]. However, the full utility of zebrafish as a toxicology model is still being realized.…”

Zebrafish: A Marvel of High-Throughput Biology for 21st Century Toxicology

“…The zebrafish embryo model has been deployed successfully to screen for cardiotoxic effects elicited by carbaryl and valproic acid (78); atypical antipsychotic drugs (79); kinase inhibitors (80); human cardiotoxic drugs (81); and chemotherapeutics (82), thus illustrating the value of zebrafish as a preclinical testing paradigm to predict cardiotoxic effects elicited by drugs under development. Recently, in vivo analysis of proliferating cardiomyocytes (the functional unit of cardiac muscle) in zebrafish embryos was leveraged to identify chemicals that could promote cardiomyocyte proliferation, thus opening a window into the possible therapeutic replacement of infarct-damaged cardiac tissue in humans (83).…”