半滑舌鳎雌性个体垂体细胞系CSFPI的建立、鉴定及其特征分析OA

Cynoglossus semilaevis is a high-value marine commercial fish species endemic to northern China,which exhibits a remarkable sexual size dimorphism,with females growing 2-4 times faster than males.The pituitary,as the core endocrine tissue that synthesizes and secretes growth hormone and other pivotal regulatory hormones,represents an ideal target for dissecting the molecular mechanism underlying this sex-dependent growth disparity.While several cell lines derived from the ovary,testis and other somatic tissues of C.semilaevis have been successfully established previously,long-term stable in vitro culture of pituitary cells has long been hindered by the extremely small anatomical size of the pituitary and the high heterogeneity of its constituent cell populations.In the present study,we overcame this long-standing technical bottleneck for the first time,and successfully established a continuous cell line designated as CSFPI from the female pituitary of C.semilaevis using an explant primary culture protocol.To date,the CSFPI cell line has been stably subcultured for over 35 passages within a 10-month culture period,and retains robust proliferative activity.Karyotype analysis verified that CSFPI cells maintain a normal diploid karyotype of 2n=42t.Further authentication using mitochondrial COI barcoding and female-specific SCAR markers confirmed that the cell line originates exclusively from the pituitary tissue of female C.semilaevis.Transfection assays demonstrated that the delivery efficiency of small interfering RNAs(siRNAs)into CSFPI cells reached approximately 90%,while the transfection efficiency of the pSKD-Tol2 transposon plasmid was approximately 20%,indicating its suitability for exogenous gene manipulation.Transcriptomic profiling revealed that 1918 genes in CSFPI cells showed an expression level with FPKM>40,and these highly expressed genes were significantly enriched in pathways including ribosome biogenesis,endoplasmic reticulum protein processing,and oxidative phosphorylation.Based on the abundant expression of signature marker genes including clu,ptprf,col6a2,luzp2,nedd4l,mdk and glul,we further inferred that the CSFPI cell line is predominantly composed of PIT-1 lineage cells and pituitary progenitor cells.The successful establishment of the CSFPI cell line greatly expands the available in vitro pituitary research models for marine teleosts.This cell line not only provides a highly accessible cellular platform for investigating the neuroendocrine regulatory network governing fish growth and reproduction,but also lays a critical experimental foundation for the genetic improvement and selective breeding of C.semilaevis and other fish species.