Resveratrol and miR-222 Synergistically Regulate Apoptosis of Porcine (Sus scrofa) Ovarian Granulosa Cells
ZHANG Hui-Bin, ZHOU Ren, XU Li-Ming, CHEN Yi-Ge, ZHANG Peng, YANG Ming-Yue, YANG Min, DING Yue-Yun, YIN Zong-Jun, ZHANG Xiao-Dong*
College of Animal Science and Technology, Anhui Agricultural University/Anhui Provincial Key Laboratory of Genetic Resources Protection and Biological Breeding of Local Livestock and Poultry, Hefei 230036, China
Abstract：Apoptosis of ovarian granulosa cells affects follicle development and ovulation in animals. Resveratrol plays an important role in the apoptosis of ovarian granulosa cells. This research was aimed to provide data support for the application of resveratrol in pig (Sus scrofa) production by analyzing the mechanism of resveratrol and miR-222 regulate the apoptosis of porcine ovarian granulosa cells (pGCs). The primary pGCs were treated with different concentrations of resveratrol (0~100 μmol/L), the morphological changes of pGCs were observed under an inverted microscope, the apoptosis rate and cell cycle distribution were detected by flow cytometry, and the expression of CyclinD, CyclinB, p21 and miR-222 were detected by the qRT-PCR method. The pGCs apoptosis change was also detected in the miR-222 overexpression condition. The results showed that when the concentration of resveratrol reached 100 μmol/L, the morphology of pGCs was changed, the apoptosis of pGCs was decreased and the cell cycle of pGCs was inhibited. The expression level of miR-222 was affected by different concentrations of resveratrol and reached the lowest level at 75 μmol/L. The pGCs apoptosis was significantly decreased when miR-222 overexpressed in 75 μmol/L resveratrol. These results indicated that resveratrol (100 μmol/L) could inhibit pGCs proliferation, and regulate miR-222 expression depend on the concentration. Overexpression of miR-222 could decrease pGCs apoptosis with 75 μmol/L resveratrol treatment. The results of the study provide more reference data for resveratrol combined miRNA to regulate the biological functions of pGCs.
[1] Allegri L, Rosignolo F, Mio C, et al.2018. Effects of nutraceuticals on anaplastic thyroid cancer cells[J]. Journal of Cancer Research and Clinical Oncology, 144(2): 285-294. [2] Babu D, Leclercq G, Goossens V, et al.2015. Antioxidant potential of CORM-A1 and resveratrol during TNF-alpha/cycloheximide-induced oxidative stress and apoptosis in murine intestinal epithelial MODE-K cells[J]. Toxicology and Applied Pharmacology, 288(2): 161-178. [3] Bartel D P.2004. MicroRNAs: Genomics, biogenesis, mechanism, and function[J]. Cell, 116(2): 281-297. [4] Basini G, Tringali C, Baioni L, et al.2010. Biological effects on granulosa cells of hydroxylated and methylated resveratrol analogues[J]. Molecular Nutrition & Food Research, 54(Suppl 2): S236-243. [5] Bezerra M E S, Gouveia B B, Barberino R S, et al.2018. Resveratrol promotes in vitro activation of ovine primordial follicles by reducing DNA damage and enhancing granulosa cell proliferation via phosphatidylinositol 3-kinase pathway[J]. Reproduction In Domestic Animals, 53(6): 1298-1305. [6] Brennecke J, Hipfner D R, Stark A, et al.2003. Bantam encodes a developmentally regulated microRNA that controls cell proliferation and regulates the proapoptotic gene hid in Drosophila[J]. Cell, 113(1): 25-36. [7] Chen C Z, Li L, Lodish H F, et al.2004. MicroRNAs modulate hematopoietic lineage differentiation[J]. Science, 303(5654): 83-86. [8] Gatouillat G, Balasse E, Joseph-Pietras D, et al.2010. Resveratrol induces cell-cycle disruption and apoptosis in chemoresistant B16 melanoma[J]. Journal of Cellular Biochemistry, 110(4): 893-902. [9] Gusman J, Malonne H, Atassi G.2001. A reappraisal of the potential chemopreventive and chemotherapeutic properties of resveratrol[J]. Carcinogenesis, 22(8): 1111-1117. [10] Hossein H A, Leila K, Azita A, et al.2017. Association of miR-146a rs2910164 and miR-222 rs2858060 polymorphisms with the risk of polycystic ovary syndrome in Iranian women: A case-control study[J]. Taiwanese Journal of Obstetrics & Gynecology, 56(5): 652-656. [11] Huang X, She L, Luo X, et al.2019. MiR-222 promotes the progression of polycystic ovary syndrome by targeting p27 Kip1[J]. Pathology, Research and Practice, 215(5): 918-923. [12] Jin X, Wang K, Liu H, et al.2016. Protection of bovine mammary epithelial cells from hydrogen peroxide-induced oxidative cell damage by resveratrol[J]. Oxidative Medicine and Cellular Longevity, 1: 2572175. [13] Joe A K, Liu H, Suzui M, et al.2002. Resveratrol induces growth inhibition, S-phase arrest, apoptosis, and changes in biomarker expression in several human cancer cell lines[J]. Clinical Cancer Research, 8(8): 893-903. [14] Kong X X, Fu Y C, Xu J J, et al.2011. Resveratrol, an effective regulator of ovarian development and oocyte apoptosis[J]. Journal of Endocrinological Investigation, 34(11): e374-381. [15] Larrosa M, Tomas-Barberan F A, Espin J C.2003. Grape polyphenol resveratrol and the related molecule 4-hydroxystilbene induce growth inhibition, apoptosis, S-phase arrest, and upregulation of cyclins A, E, and B1 in human SK-Mel-28 melanoma cells[J]. Journal of Agricultural and Food Chemistry, 51(16): 4576-4584. [16] Lee E J, Min H Y, Joo Park H, et al.2004. G2/M cell cycle arrest and induction of apoptosis by a stilbenoid, 3,4,5-trimethoxy-4'-bromo-cis-stilbene, in human lung cancer cells[J]. Life Sciences, 75(23): 2829-2839. [17] Liu J, Yao W, Yao Y, et al.2014. MiR-92a inhibits porcine ovarian granulosa cell apoptosis by targeting Smad7 gene[J]. FEBS Letters, 588(23): 4497-4503. [18] Manna S K, Mukhopadhyay A, Aggarwal B B.2000. Resveratrol suppresses TNF-induced activation of nuclear transcription factors NF-kappa B, activator protein-1, and apoptosis: Potential role of reactive oxygen intermediates and lipid peroxidation[J]. Journal of Immunology, 164(12): 6509-6519. [19] Mukhopadhyay P, Mukherjee S, Ahsan K, et al.2010. Restoration of altered microRNA expression in the ischemic heart with resveratrol[J]. PLOS ONE, 5(12): e15705. [20] Poy M N, Eliasson L, Krutzfeldt J, et al.2004. A pancreatic islet-specific microRNA regulates insulin secretion[J]. Nature, 432(7014): 226-230. [21] Salilew-Wondim D, Ahmad I, Gebremedhn S, et al.2014. The expression pattern of microRNAs in granulosa cells of subordinate and dominant follicles during the early luteal phase of the bovine estrous cycle[J]. PLOS ONE, 9(9): e106795. [22] Singh S K, Banerjee S, Acosta E P, et al.2017. Resveratrol induces cell cycle arrest and apoptosis with docetaxel in prostate cancer cells via a p53/p21WAF1/CIP1 and p27KIP1 pathway[J]. Oncotarget, 8(10): 17216-17228. [23] Wong D H, Villanueva J A, Cress A B, et al.2010. Effects of resveratrol on proliferation and apoptosis in rat ovarian theca-interstitial cells[J]. Molecular Human Reproduction, 16(4): 251-259. [24] Ye H, Liu X-J, Hui Y, et al.2020. Downregulation of microRNA-222 reduces insulin resistance in rats with PCOS by inhibiting activation of the MAPK/ERK pathway via pten[J]. Molecular Therapy Nucleic acids, 22: 733-741. [25] Zhao F, Zhao W, Ren S, et al.2014. Roles of SIRT1 in granulosa cell apoptosis during the process of follicular atresia in porcine ovary[J]. Animal Reproduction Science, 151(1/2): 34-41. [26] Zhu W, Yang M, Shang J, et al.2019. MiR-222 inhibits apoptosis in porcine follicular granulosa cells by targeting the THBS1 gene[J]. Animal Science Journal, 90(6): 719-727.
