Effects of Storage and Passage of Luteal Endothelial Cells on Endothelin-1 and Prostaglandin F2α Production

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To establish a storage system for isolated bovine luteal endothelial cells (LECs), we investigated the basal and tumor necrosis factor (TNF) α-stimulated production of endothelin-1 (ET-1) and prostaglandin (PG) F2α in unfrozen and frozen-thawed LECs until passage 10. LECs were obtained from developing corpora lutea (CL; days 5-7 of the estrous cycle) using enzymatic digestion and magnetic beads coated with lectin BS-1. The LECs were frozen at -80 C or further cultured and/or passaged until passage 10 in DMEM/Hams F-12 supplemented with 10% calf serum. The hormonal productions of unfrozen and frozen/thawed LECs were compared through passages 2-10. When both the unfrozen and frozen/thawed cells reached confluence, the culture medium was replaced with fresh medium containing 0.1% BSA, and the cells were incubated with TNF α (50 ng/ml) for 12 h. The basal productions of ET-1 and PGF2α by the unfrozen and frozen/thawed LECs were similar at passage 2. The basal production of PGF2α by LECs was not altered by passage and storage at -80 C, whereas the basal production of ET-1 decreased from passage 2 and 3 to passage 4 in the unfrozen LECs and from passage 2 to passage 3 in the frozen/thawed LECs. However, production of ET-1 by the unfrozen and frozen/thawed LECs was similar between passages 4-10 and passages 3-10, respectively. Exposure of LECs to TNFα increased (P<0.05) ET-1 and PGF2α production by the unfrozen and frozen-thawed LECs in all passages examined. Thus, LECs obtained from developing CLs and stored until passage 10 can be used for study of the physiology of LECs in vitro.
日本繁殖生物学会の論文

Effects of Storage and Passage of Luteal Endothelial Cells on Endothelin-1 and Prostaglandin F2α Production

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