Studies On The Mechanism Of Scoliosis Caused By Ccdc57 Gene Knockout In Zebrafish

Scoliosis affects around 3% of children worldwide,even can lead to physical deformity,thoracic deformity and cardiopulmonary dysfunction when it's severe.It's essential to establish more animal models to improve research in this area,in the case of limited understanding of their pathogenesis and limited treatment with adjuvant therapy with in vitro devices.Zebrafish is selected as model in the study because the water resistance on the spine of zebrafish is similar to the gravity on the spine of human walking upright.In this study,two ccdc57 knockout lines,ten and four base pair deletion,are obtained using CRISPR/Cas9 gene editing technology and both homozygous mutants show typical scoliosis.Based on that the homozygote in F3 become curvature at the end of the spine at 72 hours post fertilization(hpf),rescue experiment is carried out by WT_ccdc57 mRNA micro injection and proved to be effective,which demonstrates Ccdc57 deficiency exactly lead to scoliosis.However,Ccdc57 seems not to regulate bone development from the results that no difference is identified in spine ossification and spine bone mineral density in ccdc57 knockout zebrafish and wild-type zebrafish through Calcein staining and micro CT scanning,even no ccdc57 expression is detected in the spine and other skeleton in embryos when the curvature has occurred.There are studies suggesting that cerebrospinal fluid flow(CSF)disorders result from the cilia defect can cause scoliosis.Then the cilia in brain ventricle is examined under scanning electron microscopy and CSF is labeled with dextran conjugated to Texas Red.The cilia in mutant turns out to be sparse obviously and dye transport in mutant is substantially impaired,which prove Ccdc57 control CSF through affecting cilia development in brain ventricle.But how the CSF affects body-axis.According to the latest study,we examine the model of Catecholamines-urotensin signal-muscle contraction.The curvature at the end of the spine at 72 hpf can be rescued through epinephrine treatment.The expression of urotensins and urotensin receptors are down regulated in mutant compared to wild-type zebrafish while treated with epinephrine Immersion or WT_ccdc57 mRNA injection can induce the expression of urotensins and urotensin receptors in mutants,suggesting Ccdc57 controls body-axis through epinephrine and urotensin signals.This study demonstrates Ccdc57 is associated with cilia development and affects the CSF to control the body axis formation through epinephrine and urotensin signals.This study firstly reveals the role of Ccdc57 in cilia and body-axis formation and provides an experimental basis for further explaining the mechanism of scoliosis.