Establishment of a cell line derived from the gills of Gymnocypris przewalskii, an endemic Schizothoracine fish from Qinghai Lake of Tibet Plateau.

Gymnocypris przewalskii (Naked carp), a native teleost, plays an important role in maintaining the ecological balance of Lake Qinghai (altitude, 3.2 km), the largest saline lake in China. In this study, a new gill cell line from G. przewalskii was developed using the explant technique and named as GPG. This cell line was maintained in Dulbecco's Modified Eagle Medium (DMEM) (high glucose), supplemented with 15% fetal bovine serum (FBS), and was successfully subcultured up to 32 passages. Meanwhile, this cell line was also authenticated by sequencing the mitochondrial cytochrome C oxidase subunit I (COI) and 16S rRNA genes and by chromosome analysis. With the Cytomegalovirus (CMV) promoter, the GPG cell line could express green fluorescent protein (GFP) at about 5% transfection efficiency. MTT test showed that Clostridium botulinum toxin (BTX) was toxic to the cell line. After cryopreservation with 10% dimethyl sulfoxide (DMSO), this cell line could be successfully revived at an efficiency over 70%. This study revealed that the GPG cell line could be used as materials for physio-chemical investigation of G. przewalskii and also provided a tool for gene function study and toxicological reaction in vitro.

Cell line derived from muscle of Gymnocypris przewalskii, a species of Schizothoracinae in Qinghai Lake, Qinghai-Tibet Plateau.

Gymnocypris przewalskii (naked carp), a native teleost, plays an important role in the ecosystem of Qinghai Lake (altitude, 3.2 km) on the Qinghai-Tibet Plateau in China. We developed a new cell line from the muscle of G. przewalskii using the explant technique and named the cell line GPM. This cell line was maintained in DMEM medium (high glucose) supplemented with 15% fetal bovine serum (FBS). The cell line was successfully subcultured up to 32 passages and was authenticated by immunofluorescence assay, sequencing the mitochondrial cytochrome C oxidase subunit I (COI) and 16S rRNA genes, and by chromosome analysis. In the medium containing 15% FBS, the cell line could be passaged stably at 25 °C. The GPM cell line could express green fluorescent protein (GFP) with a CMV promoter with about 5% transfection efficiency. MTT tests showed that Clostridium botulinum toxin (BTX) was toxic to the cell line. The cell line could be successfully cryopreserved in liquid nitrogen with a revival efficiency of over 70%. This study demonstrated that the GPM cell line can be used as an important tool for understanding the physiological characteristics of G. przewalskii, and it can provide a resource for studying gene function and toxicological reactions in vitro.

Transcriptomic and proteomic analyses provide insights into the adaptive responses to the combined impact of salinity and alkalinity in Gymnocypris przewalskii.

Gymnocypris przewalskii is the only high-land endemic teleost living in Qinghai Lake, the largest saline-alkaline lake in China. Its osmoregulatory physiology remains elusive due to a lack of precise identification of the response proteins. In the present study, DIA/SWATH was used to identify differentially expressed proteins (DEPs) under alkaline (pH = 10.1, carbonate buffer), saline (12‰, sodium chloride), and saline-alkaline [carbonate buffer (pH = 10.1) plus 11‰ sodium chloride] stresses. A total of 66,056 unique peptides representing 7,150 proteins and 230 DEPs [the false discovery rate (FDR) ≤ 0.05, fold change (FC) ≥ 1.5] were identified under different stresses. Comparative analyses of the proteome and transcriptome indicated that over 86% of DEPs did not show consistent trends with mRNA. In addition to consistent enrichment results under different stresses, the specific DEPs involved in saline-alkaline adaptation were primarily enriched in functions of homeostasis, hormone synthesis and reactions of defense response, complement activation and reproductive development. Meanwhile, a protein-protein interaction (PPI) network analysis of these specific DEPs indicated that the hub genes were ITGAX, MMP9, C3, F2, CD74, BTK, ANXA1, NCKAP1L, and CASP8. This study accurately isolated the genes that respond to stress, and the results could be helpful for understanding the physiological regulation mechanisms regarding salinity, alkalinity, and salinity-alkalinity interactions.