Abstract

The Chinese sturgeon ( Acipenser sinensis ) is a first-class protected wildlife species in China and a living fossil, often referred to as the “Giant Panda of the waters”. Currently, research on A. sinensis primarily focuses on morphology and reproductive biology, while studies in cellular biology remain relatively limited. This study established the first A. sinensis embryonic cell line (ASE48h) derived from 48-h neurula stage embryos. The cell line has been successfully sub-cultured to the 80th passage. ASE48h cells are maintained in L-15 medium supplemented with 10% fetal bovine serum (FBS) at 24 °C. After cryopreservation and recovery, approximately 80% of cells resumed normal growth. Cell proliferation reaches its maximum rate in medium containing 20% FBS, with a population doubling time of 87.34 h. Chromosomal analysis revealed a variable chromosome number ranging from 190 to 276, with 69% cells containing 220–270 chromosomes. The cells demonstrated successful transfection capability with cy3-siRNA using Lipo8000 transfection reagent. Furthermore, lipopolysaccharide (LPS) and Poly (I:C) were employed to examine the immune responses to bacteria and virus stimulation, separately. The subsequent transcriptomic analysis revealed that LPS stimulation triggered 2210 differentially expressed genes, which significantly enriched in Cytokine-cytokine receptor interaction, TNF signaling pathway, NF-kappa B signaling pathways and others. On the other hand, Poly (I:C) treatment induced 1095 differentially expressed genes involved in ECM-receptor interaction, Focal adhesion, Human papillomavirus infection and etc. Alkaline phosphatase (AP) staining detected pluripotency in about 20% of ASE48h cells. Subsequently, functional enrichment analysis of genes including id1 , myc , nes , and wdr43 with FPKM >50 in ASE48h cells confirmed that this cell line predominantly consisted of neural stem/progenitor-like cells. The establishment of the ASE48h cell line provides a va