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Animals 2019, 9(3), 106; https://doi.org/10.3390/ani9030106

Optimization of In Vitro Culture Conditions of Sturgeon Germ Cells for Purpose of Surrogate Production

1
Key Laboratory of Freshwater Biodiversity Conservation, Ministry of Agriculture of China, Yangtze River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Wuhan 430223, China
2
Research Institute of Fish Culture and Hydrobiology, University of South Bohemia in České Budějovice, Faculty of Fisheries and Protection of Waters, South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses, Zátiší 728/II, 38925 Vodňany, Czech Republic
3
Marine College, Shandong Universit, Weihai 264209, China
4
Sino-Czech Joint Laboratory for Fish Conservation and Biotechnology, Yangtze River Fisheries Institute, Chinese Academy of Fishery Sciences, Wuhan 430223, China
*
Author to whom correspondence should be addressed.
Xuan Xie and Ping Li should be regarded as joint First Authors.
Received: 11 February 2019 / Accepted: 2 March 2019 / Published: 21 March 2019
(This article belongs to the Section Aquatic Animals)
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Simple Summary

The sturgeon is among the most ancient of actinopterygian fishes. Most species of sturgeon are listed as critically endangered due to habitat alteration caused by damming of rivers, pollution and overharvesting. Germ cell transplant is a useful tool to save these endangered species. To expand germ cell populations and sustain the supply for long periods for transplant, we established basal culture conditions for sturgeon germ cells. Germ cell mitotic activity has been enhanced by eliminating gonad somatic cells, supplementing with growth factor and using an alternative to fetal bovine serum. The optimal condition identified was purified germ cells cultured in serum-free medium supplemented with leukemia inhibitory factor (LIF) and glial cell line-derived neurotrophic factor (GDNF) at 21 °C. Cultured sterlet germ cells showed development after transplant into Russian sturgeon. The study provided useful information for sturgeon germ cell culture.

Abstract

To expand germ cell populations and provide a consistent supply for transplantation, we established basal culture conditions for sturgeon germ cells and subsequently increased their mitotic activity by eliminating gonad somatic cells, supplementing with growth factor, and replacing fetal bovine serum (FBS). The initial basal culture conditions were Leibovitz’s L-15 medium (pH 8.0) supplemented with 5% FBS (p < 0.001) at 21 °C. Proliferation of germ cells was significantly enhanced and maintained for longer periods by elimination of gonad somatic cells and culture under feeder-cell free conditions, with addition of leukemia inhibitory factor and glial-cell-derived neurotrophic factor (p < 0.001). A serum-free culture medium improved germ cell proliferation compared to the L-15 with FBS (p < 0.05). Morphology remained similar to that of fresh germ cells for at least 40 d culture. Germline-specific gene expression analysis revealed no significant changes to germ cells before and after culture. Sterlet Acipenser ruthenus germ cells cultured more than 40 days showed development after transplant into Russian sturgeon Acipenser gueldenstaedtii. Polymerase chain reaction showed 33.3% of recipient gonads to contain sterlet cells after four months. This study developed optimal culture condition for sturgeon germ cells. Germ cells after 40 d culture developed in recipient gonads. This study provided useful information for culture of sturgeon germ cells. View Full-Text
Keywords: feeder cells; germ cell culture; glial-cell-derived neurotrophic factor; sturgeon; transplantation feeder cells; germ cell culture; glial-cell-derived neurotrophic factor; sturgeon; transplantation
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MDPI and ACS Style

Xie, X.; Li, P.; Pšenička, M.; Ye, H.; Steinbach, C.; Li, C.; Wei, Q. Optimization of In Vitro Culture Conditions of Sturgeon Germ Cells for Purpose of Surrogate Production. Animals 2019, 9, 106.

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