Characterization of the continuous skin fibroblastoid cell line, WE‐skin11f, from walleye ( Sander vitreus )

Wang

et al. 2023

Aquaculture Research

With the growing sophistication of cloning technology, rescuing cell resources is of great significance for the protection of endangered animals. The Yangtze sturgeon (Acipenser dabryanus), one of the three Acipenseriformes species in the Yangtze River, is critically endangered. Natural reproduction of the Yangtze sturgeon has not been detected since 2000. Less than 20 wild individuals are kept in husbandry, and all are too old to breed. Therefore, it is urgent to rescue the genetic resources of every wild Yangtze sturgeon. Here, we isolated and preserved viable cells from critically endangered postmortem Yangtze sturgeon for the first time. Attempts to rescue and preserve cell resources were carried out from 8 tissues, brain, kidney, gonad, fin, liver, skin, spleen, and muscle of an over 35-year-old female wild Yangtze sturgeon between 11 and 14 hours after death at 19.8°C in an outdoor concrete pond, and only muscle tissue cells could be successfully subcultured and preserved. Furthermore, the cultured cells showed high post-thaw cell viability and normal growth with a population doubling time of 52.98 h. Moreover, they were fibronectin- and desmin-positive, characterizing them as fibroblastic confirmed muscle cells with fibroblastic properties and myogenic origin. Tests for microbial contamination of the cell lines were negative. Chromosome analysis demonstrated that muscle cells possess a modal polyploid chromosome number of 264. The mitochondrial sequence data of COI genes and 12S rRNA confirmed that the developed cell line originated from A. dabryanus. Furthermore, transfection results indicate that muscle cells could be used for gene manipulation and functional studies. These results suggest that viable muscle cells could also be successfully isolated and cryopreserved from the wild Yangtze sturgeon in a short time after death. This report is not only of great significance for the germplasm rescue of critically endangered Yangtze sturgeon but also provides some scientific reference for the germplasm preservation of other endangered fish.

Section: Discussionsupporting

confidence: 57%

Germplasm Rescue of Postmortem Critically Endangered Yangtze Sturgeon (Acipenser dabryanus) by Cell Preservation

Wang

et al. 2023

Aquaculture Research

“…The passage number was restricted to 25th since biological characteristics of cells preserved as germplasm resources may be adversely affected by more passages and repeated digestion of trypsin (Yang et A homogeneous population of broblasts cells was observed after the 8th generation. Furthermore, the immuno uorescence staining results supported the opinion that the WYSM cells were broblastic in nature due to its strong immunoreaction to broblastic marker ( bronectin) and a negative reaction to epithelial marker (pancytokeratin), which was different from the positive expression of mesenchymally-derived cell marker (vimentin) in some other broblasts reported (Vo et al 2019). Myosatellite cell is a kind of muscle derived stem cells that is able to self-renew and have the potential to differentiate, however, it makes up less and less of a sh's body weight as it ages (Koumans et al 1991;Wang et al 2012).…”

Section: Discussionmentioning

confidence: 52%

Germplasm rescue of post-mortem critically endangered Yangtze Sturgeon by cell cryopreservation

Wang

et al. 2022

Preprint

With the growing sophistication of cloning technology, rescuing cell resources is of great significance for the protection of endangered animals. The Yangtze sturgeon (Acipenser dabryanus), one of the three Acipenseriformes species in the Yangtze River, is critically endangered. Natural reproduction of the Yangtze sturgeon has not been detected since 2000. Less than 20 wild individuals are kept in husbandry and all are too old to breed. Therefore, it is urgently to rescue the genetic resources of every wild Yangtze sturgeon. Here, we isolated and preserved viable cells from the post-mortem critically endangered Yangtze Sturgeon for the first time. Attempt of rescuing and preserving cell resources were carried out from 8 tissues, brain, kidney, heart, fin, liver, skin, spleen and muscle of over-35-years-old female wild Yangtze sturgeon between 11 and 14 hours after death at 19.8℃ in outdoor concrete pond, and only muscle tissue cells could be successfully sub-cultured and preserved. Furthermore, the cultured cells were assessed by population doubling time, immunofluorescence analysis, microorganism detection, karyotyping and origin identification, and no abnormality was observed in the morphology, structure, growth and ploidy. These results suggest that viable cells could also be successfully isolated and cryopreserved from the wild Yangtze sturgeon in a short time after death, and muscle tissues had more potential to separate living cells than other tissues. This report is not only of great significance to the germplasm rescue of critically endangered Yangtze sturgeon, but also provides some scientific reference for the germplasm preservation of other endangered fish.

Derivation and characterization of new cell line from intestine of turbot (Scophthalmus maximus)

In Vitro Cell.Dev.Biol.-Animal

et al. 2023

A continuous intestine cell line from turbot (Scophthalmus maximus) designated as SMI was established utilizing the tissue explant technique. Primary SMI cell was cultured at 24 °C in a medium with 20% fetal bovine serum (FBS), then subcultured in 10% FBS after 10 passages. Impacts of medium or temperature on the growth of SMI were examined and the results indicated it grew well in DMEM supplemented with 10% FBS at 24 °C. The SMI cell line was subcultured more than 60 times. Karyotyping, chromosome number, and ribosomal RNA genotyping analysis revealed that SMI had a modal diploid chromosome number of 44 and originated from turbot. After being transfected with pEGFP-N1 and FAM-siRNA, a large number of green fluorescence signals were observed in SMI, indicating that SMI could be used as an ideal platform to explore gene function in vitro. In addition, the expression of epithelium-associated genes such as itga6, itgb4, gja1, claudin1, zo-1, and E-cadherin in SMI suggested the SMI had some characteristics of epidermal cells. The upregulation of immune-associated genes such as TNF-β, NF-κB, and IL-1β in SMI after stimulation with pathogen-associated molecular patterns suggested the SMI might exhibit immune functions similar to the intestinal epithelium in vivo.