Establishment And Application Of Zebrafish Slc22a23 Gene Knockout Model

Solute Carrier(SLC)superfamily is an important membrane transporter family on the cell membrane,and it mediates the transport of various solutes between the inner cell and external environment.Studies have shown that abnormal expression or dysfunction of SLC protein are closely related to depression,diabetes,coronary heart disease and other major diseases.The SLC22A23 protein encoded by the SLC22A23(solute carrier family 22 member 23)gene belongs to the SLC family.As a member of the SLC superfamily,people have found that the Slc22a23gene was expressed in the outflow tract of heart tissue at E8.5 of mouse embryos through in situ hybridization.This study indicated that Slc22a23gene may play an important role in mouse heart development,but the specific function of this gene has not yet been reported in the literature.According to the gene expression data analysis of normal tissues,SLC22A23 gene was found to be expressed in human,mouse,zebrafish and other vertebrate brain,eye,heart and other tissuses,and SLC22A23protien is highly conserved evolutionistically.This study has found that slc22a23 expressed in the heart of zebrafish at 48 hpf by in situ hybridization.We successfully established a 59 bp deletion zebrafish slc22a23 gene knockout model using CRISPR/Cas9 gene editing technology.Also,we established an EGFP transgenic fluorescence reporter in slc22a23 gene knockout line by crossing the knockout model with the NPPA:EGFP line.Phenotype analysis of heart development showed that no significant deformity was found in the slc22a23^-/-animals.Morphological analyses of the heart tissues of slc22a23^-/-adults were carried out.The results showed that the ventricles of slc22a23^-/-adults were slightly larger than those of wild type control;HE staining analysis showed that the ventricular wall tissuse of slc22a23^-/-adults heart was loosely arranged.Ultrastructural analysis showed that the subcellular structure of the heart tissue of slc22a23^-/-adult fish was disordered,and the number of mitochondria and autophagy lysosomes were increased.ECG monitoring of adult fish treated with quetiapine fumarate showed QT prolongation in 22%of wild-type zebrafish and only 8%of slc22a23^-/-fish,suggesting that loss of slc22a23 may affect quetiapine transport.In summary,the knockout of slc22a23 gene may lead to abnormal zebrafish heart structure,and the zebrafish slc22a23 gene knockout model is used to clarify the function of this gene and provides a new research model for reducing the risk of adverse reactions to quetiapine drugs.