Development of tg(Uas:Sec‐hsa.anxa5‐yfp,myl7:Rfp); casper(roy−/−,nacre−/−) transparent transgenic in vivo zebrafish model to study the cardiomyocyte function

Abstract

The zebrafish provided an excellent platform to study the genetic and molecular approach of cellular phenotype‐based cardiac research. We designed a novel protocol to develop the transparent transgenic zebrafish model to study annexin‐5 activity in the cardiovascular function by gen-erating homozygous transparent skin Casper (roy−/−,nacre−/−); myl7:RFP; annexin‐5:YFP transgenic zebrafish. The skin pigmentation background of any vertebrate model organism is a major obstruc-tion for in vivo confocal imaging to study the transgenic cellular phenotype‐based study. By developing Casper (roy−/−,nacre−/− ); myl7; annexin‐5 transparent transgenic zebrafish strain, we estab-lished time‐lapse in vivo confocal microscopy to study cellular phenotype/pathologies of cardiomy-ocytes over time to quantify changes in cardiomyocyte morphology and function over time, com-paring control and cardiac injury and cardio‐oncology. Casper contributes to the study by integrat-ing a transparent characteristic in adult zebrafish that allows for simpler transparent visualization and observation. The Casper (roy−/−,nacre−/− ) transgenic progenies developed through cross‐breed-ing with the transgenic strain of Tg (UAS:SEC‐Hsa.ANXA5‐YFP,myl7:RFP). Confocal and fluorescent microscopy were being used to obtain accurate, precise imaging and to determine fluorescent protein being activated. This study protocol was conducted under two sections; 1.1: Generation of homozygous Tg (UAS:SEC‐Hsa.ANXA5‐YFP,myl7:RFP); Casper (roy−/−,nacre−/− ) zebrafish (generation F01‐F06) and 1.2: Screening and sorting the transparent transgenic progeny and in vivo imaging to validate cardiac morphology through in vivo confocal imaging. We coined the newly developed strain as Tg (UAS:SEC‐Hsa.ANXA5‐YFP,myl7:RFP); Casper (roy−/−,nacre−/−)gmc1. Thus, the newly developed strain maintains transparency of the skin throughout the entire life of zebrafish and is ca-pable of application of a non‐invasive in vivo imaging process. These novel results provide an in vivo whole organism‐based platform to design high‐throughput screening and establish a new hori-zon for drug discovery in cardiac cell death and cardio‐oncology therapeutics and treatment.