title: Effects of coculture with cumulus-derived somatic cells on in vitro maturation of porcine oocytes.
abstract: In the process of IVM, cumulus-oocyte complexes (COCs) separate from the follicular microenvironment, leading to the loss of endocrine interactions between follicular mural somatic cells and COCs. To restore the microenvironment, a coculture system was established using cumulus-derived somatic cells (CSCs) for IVM. The CSCs were cultured in Dulbecco's modified Eagle's medium for 48 hours with varying numbers of CSCs (0.0, 2.5 × 10(4), 5.0 × 10(4), and 10.0 × 10(4)) and then cultured in tissue culture medium 199 (TCM 199) for 4 hours before adding the oocytes. Cumulus-oocyte complexes from 3- to 6-mm follicles were matured in 500 μL of TCM 199 with eCG and hCG for 22 hours and then cultured in TCM 199 without hormones for 22 hours. After IVM, the group with 2.5 × 10(4) CSCs showed a significant increase in intracellular glutathione levels compared with the control group. In the evaluation of sperm penetration, efficient fertilization was increased in the groups with 2.5 × 10(4) and 5.0 × 10(4) CSCs compared with controls (44.9 and 46.5 vs. 32.1, respectively). The mRNA expression pattern analysis in matured COCs showed a significant upregulation of PCNA, COX-2, Has2, Ptx3, and Nrf2 in the 2.5 × 10(4) CSC group compared with controls. During COC maturation at 0, 11, 22, 33, and 44 hours, the 2.5 × 10(4) and 5.0 × 10(4) CSC groups showed a significantly altered mRNA expression of BMP15 and GDF9. The developmental competence of the matured oocytes in all groups was evaluated after IVF and parthenogenetic activation (PA). After IVF, the 2.5 × 10(4) CSC group showed significantly higher cleavage, blastocyst formation rate, and total cell numbers compared with controls (60.0%, 35.7%, and 127.3 vs. 43.2%, 21.1%, and 89.3, respectively). After PA, the 2.5 × 10(4) CSC group had significantly higher blastocyst formation rate and total cell number than the control group (52.0% and 120.4 vs. 35.4% and 90.9, respectively). In conclusion, these results suggest that the presence of a population of 2.5 × 10(4) CSCs during IVM synergistically improved the developmental potential of IVF- and PA-derived porcine embryos by increasing the intracellular glutathione level via changing of a specific gene expression pattern during oocyte maturation.
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
authors.forename: Junchul David
authors.initials: JD
authors.lastname: Yoon
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
authors.forename: Yubyeol
authors.initials: Y
authors.lastname: Jeon
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
authors.forename: Lian
authors.initials: L
authors.lastname: Cai
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
authors.forename: Seon-Ung
authors.initials: SU
authors.lastname: Hwang
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
authors.forename: Eunhye
authors.initials: E
authors.lastname: Kim
authors.affiliation: Laboratory of Theriogenology, College of Veterinary Medicine, Kangwon National University, Kangwon, Republic of Korea.
authors.forename: Eunsong
authors.initials: E
authors.lastname: Lee
authors.affiliation: Department of Life Science, College of BioNano Technology, Gachon University, Incheon, Republic of Korea. Electronic address: davekim@gachon.ac.kr.
authors.forename: Dae Y
authors.initials: DY
authors.lastname: Kim
authors.affiliation: Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea. Electronic address: shhyun@cbu.ac.kr.
authors.forename: Sang-Hwan
authors.initials: SH
authors.lastname: Hyun
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000502:physiology; D018920:Coculture Techniques; D054885:Cumulus Cells / Q000502:physiology*; D047108:Embryonic Development; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary; D015870:Gene Expression; D005978:Glutathione / Q000032:analysis; D059471:In Vitro Oocyte Maturation Techniques / Q000379:methods / Q000662:veterinary*; D009865:Oocytes / Q000737:chemistry / Q000502:physiology*; D010312:Parthenogenesis; D012333:RNA, Messenger / Q000032:analysis; D017382:Reactive Oxygen Species / Q000032:analysis; D034421:Sus scrofa
chemical_list: D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D005978:Glutathione
keywords: Coculture; In vitro maturation; Oocyte; Porcine; Somatic cell