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Determination of Medium Condition Effective to Cryopreservation of Primary Spermatogonial Stem Cells Derived from Porcine Neonatal Testes

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BACKGROUND: The superior genetic resources of breeding pigs have been preserved for use through freezing the sperm or semen. However, because there is no way to collect their sperm or semen after depletion, the generation of sperm via the differentiation of porcine spermatogonial stem cells (SSCs) can be an alternative. To date, there have been no reports of techniques customized to in-vitro culture and differentiation into sperm in porcine SSCs. Accordingly, it is important to preserve porcine SSCs with outstanding genetic backgrounds until these technologies are developed. Unfortunately, a protocol for the long-term preservation of porcine SSCs has yet to be reported. OBJECTIVE: We tried to develop a cryopreservation medium to preserve the characteristics of undifferentiated porcine SSCs for long-term cryopreservation. MATERIALS AND METHODS: SSCs retrieved from porcine testes were freeze-cryopreserved in StemPro-34 medium supplemented with various concentrations of fetal bovine serum (FBS), dimethyl sulfoxide (DMSO), and trehalose; then, after 7 days, the viability and alkaline phosphatase (AP) activity was measured in thawed porcine SSCs. Additionally, we investigated the use of hypotaurine and/or glutathione as antioxidants in the optimized freezing medium for maintaining the viability and AP activity of porcine SSCs during the freezing- cryopreservation-thawing process. RESULTS: Porcine SSCs frozen-cryopreserved-thawed in StemPro-34 medium supplemented with 10% (v/v) FBS, 10% (v/v) DMSO, 200 mM trehalose, 5 mM hypotaurine, and 5 mM glutathione showed the highest viability and AP activity. CONCLUSION: We optimized a cryopreservation medium that inhibits the loss of viability and the increases differentiation post-thawing of the frozen porcine SSCs.
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Keywords: CRYOPRESERVATION; MEDIUM; PIGS; SPERMATOGONIAL STEM CELLS; UNDIFFERENTIATION

Document Type: Research Article

Publication date: November 1, 2019

More about this publication?
  • CryoLetters is a bimonthly international journal for low temperature sciences, including cryobiology, cryopreservation or vitrification of cells and tissues, chemical and physical aspects of freezing and drying, and studies involving ecology of cold environments, and cold adaptation

    The journal publishes original research reports, authoritative reviews, technical developments and commissioned book reviews of studies of the effects produced by low temperatures on a wide variety of scientific and technical processes, or those involving low temperature techniques in the investigation of physical, chemical, biological and ecological problems.

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