Skip to main content
ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Animal Biosciences & Biotechnology Laboratory » Research » Publications at this Location » Publication #408847
Research Project: Development of New Technologies and Methods to Enhance the Fertility, Utilization, and Long-Term Storage of Poultry and Swine Germplasm

Location: Animal Biosciences & Biotechnology Laboratory

Title: Gene expression patterns as a screening tool for assessing turkey ovarian tissue vitrification protocols

Author
item HALL, GEORGE - US Department Of Agriculture (USDA)
item Shannon, Amy
item Hildenberger, Diane
item Brady, Kristen
item Long, Julie

Submitted to: Cryobiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/5/2023
Publication Date: 3/9/2024
Citation: Hall, G.B., Shannon, A.E., Hildenberger, D.M., Diehl, K.M., Long, J.A. 2024. Gene expression patterns as a screening tool for assessing turkey ovarian tissue vitrification protocols. Cryobiology. 114 (2024) 104837. https://doi.org/10.1016/j.cryobiol.2023.104837.
DOI: https://doi.org/10.1016/j.cryobiol.2023.104837

Interpretive Summary: Storing reproductive tissues and cells indefinitely at low temperatures, known as biobanking, is a way to preserve genetic material of agriculturally important species, such as the turkey. One method for preserving ovarian tissue is a process known as vitrification, where rapid freezing rates are combined with specialized cryoprotectants to minimize cellular damage. For turkey ovarian tissue, one standard vitrification protocol has been used without determining the impact on the ability of the tissue to resume reproductive function, and the effectiveness of the protocol has only been assessed after surgical transplantation of the thawed ovary into a recipient bird. The development of a screening tool that permits a more rapid assessment of vitrification protocols and does not rely on transplantation would be of great benefit to biobanking. Here we assessed a short-term culture method using chicken embryos to evaluate reproductive function of vitrified turkey ovarian tissue and discovered that follicular density and the expression of two genes, heat shock protein A2 (HSPA2) and growth determining factor 9 (GDF9), may serve as a screening tool.

Technical Abstract: Biobanking of vitrified turkey ovarian tissue will play a crucial part in preserving female turkey genetics. To date, ovarian tissue has only been vitrified using a standard vitrification protocol, and only analyzed immediately after warming, and so long-term cryoinjury has not been explored. Here this was investigated using an in-ovo culture system, where fresh, standardly vitrified (StV) and poorly vitrified (PV) ovarian tissue were assessed for cellular morphological changes and mRNA gene expression immediately after warming or after in-ovo culture for 2, 4, or 6 days. After 6 days of in-ovo culture, cortical follicle density was the highest in fresh tissue (4,701 ± 950 #/mm3), followed by the StV tissue (1,601 ± 300 #/mm3), and lowest in PV (172 ± 145 #/mm3). This demonstrated that, while more follicles survived and grew in StV tissue verses PV tissue, a considerable number were lost or didn’t develop compared to non-vitrified tissue. Immediately after warming, the gene expression levels of heat shock protein A2 (HSPA2) were lowest in the fresh tissue (1.0 ± 0.1), higher in the StV tissue (6.4 ± 2.3), and highest in the PV tissue (19.8 ± 3.5). After 6 days of in-ovo culture, growth determining factor 9 (GDF9) expression was highest for the fresh tissue (1.8 ± 0.5) compared to the StV (0.4 ± 0.1) and PV (0.1 ± 0.1) treatments. This suggests that there is room to improve the standard turkey ovarian tissue vitrification protocol, and that HSPA2 and GDF9 gene expression, along with follicle density, could serve as screening tools to assess vitrification protocols.