Strategies for improving follicular culture efficiency in vitro: Importance of medium supplementation and study of epigenetic changes




Epigenetic changes; In vitro culture; Oxidative stress; Ovarian follicles.


The oxidative stress (OS) and epigenetic changes can impact folliculogenesis in different species. In vitro culture (IVC) of follicles efficiently will require a better understanding of these events. Therefore, the objective of this review is to address the main features and advances of IVC of ovarian follicles and exposing its limitations (i.e., OS and epigenetic changes) and alternatives (i.e., antioxidants and conditioned medium of mesenchymal stem cells - CM-MSCs).  In this review literature, 143 bibliographical references were used, dated from 1987 to 2021. As inclusion criteria, experimental and review literature articles and chapters were used, published in Portuguese, English and Spanish. Various researches revealed that the best results obtained by IVC of preantral follicles in mice than other species with the birth of healthy offspring. Currently, inadequate in vitro culture conditions are a potent obstacle to this biotechnology has been reported. These conditions are responsible for increased OS and changes in the epigenetic patterns of follicles (oocytes and somatic cells). Thus, to overcome this problem, different substances added into the IVC medium, such as antioxidants and media from the IVC of MSCs (i.e., CM).


Abedelahi, A., Salehnia, M., & Allameh, A. A. (2010). Sodium selenite improves the in vitro follicular development by reducing the reactive oxygen species level and increasing the total antioxidant capacity and glutathione peroxide activity. Human Reproduction, 25, 977-985.

Abir, R., Nitke, S., Ben-Haroush, A., & Fisch, B. (2006). In vitro maturation of human primordial ovarian follicles: Clinical significance, progress in mammals, and methods for growth evaluation: Review. Histology and Histopathology, 21, 887-898.

Agarwal, A., & Allamaneni, S. S. R. (2004). Role of free radicals in female reproductive diseases and assisted reproduction. Reproduction Biomedicine Online, 9, 338-347.

Aguiar, F. L. N., Gastal, G. D. A., Alves, K. A., Alves, B. G., Figueiredo, J. R., & Gastal, E. L. (2020). Supportive techniques to investigate in vitro culture and cryopreservation efficiencies of equine ovarian tissue: A review. Theriogenology, 156, 296-309.

Amarowicz, R., Pegg, R. B., Rahimi-Moghaddam, P., Barl, B., & Weil, J. A. (2004). Free-radical scavenging capacity and antioxidant activity of selected plant species from the Canadian prairies. Food Chemistry, 84(4), 551-562.

Amorin, B., Valim, V. S., Lemos, N. E., Júnior, L. M., Silva, A. M. P., Silva, M. A. L., & Silla, L. (2012). Células-tronco mesenquimais - caracterização, cultivo, propriedades imunológicas e aplicações clínicas. Revista HCPA, 32, 71-81.

Andrade, E. R., Van Den Hurk, R., Lisboa, L. A., Hertel, M. F., Melo-Sterza, F. A., Moreno, K., Bracarense, A. P., Landim-Alvarenga, F. C., Seneda, M. M., & Alfieri, A. A. (2012). Effects of ascorbic acid on in vitro culture of bovine preantral follicles. Zygote, 5, 1-10.

Ang, L., Haixia, C., Hongxia, L., Ruijiao, L., Xingping, G., & Huaixiu, W. (2021). Supplementation of c-type natriuretic peptide during in vitro growth period benefits the development of murine preantral follicles. Zygote, 29(2), 150-154.

Arunakumari, G., Shanmugasundaram, N., & Rao, V. H. (2010). Development of morulae from the oocytes of cultured sheep preantral follicles. Theriogenology, 74, 884-894.

Asgari, H. R., Akbari, M., Abassi, M., Ai, J., Korouji, M., Aliakbari, F., Babatunde, K. A., Aval, F. S., & Joghataei, M. T. (2015). Human Wharton’s jelly-derived mesenchymal stem cells express oocyte developmental genes during co-culture with placental cells. Iranian Journal of Basic Medical Sciences, 18(1), 22-29.

Asgharzadeh, S., Mirshokraei, P., Hassanpour, H., Ahmadi, E., & Nazari, H. (2015). The effect of mesenchymal stem cells as co-culture in in vitro nuclear maturation of ovine oocytes. Animal Science Papers and Reports, 33, 223-232.

Azari, O., Babaei, O., Derakhshanfar, A., Nematollahi-Mahani, S. N., Poursahebi, R., & Moshrefi, M. (2011). Effects of transplanted mesenchymal stem cells isolated from Wharton’s jelly of caprine umbilical cord on cutaneous wound healing, histopathological evaluation. Veterinary Research Communications, 35, 211-222.

Baumann, C., Olson, M., Wang, K., Fazleabas, A., De La Fuente, R. (2015). Arginine methyltransferases mediate an epigenetic ovarian response to endometriosis. Reproduction, 150(4), 297-310.

Barros, V. R., Cavalcante, A. Y. P., Macedo, T. J. S, Barberino, R. S., Lins, T. L. B., Menezes, V. G., Queiroz, M. A. A., Araújo, V. R., Palheta J. R. R. C., Leite, M. C. P., & Matos, M. H. T. (2013). Immunolocalization of Melatonin and Follicle‐Stimulating Hormone Receptors in Caprine Ovaries and their Effects During in vitro Development of Isolated Pre‐Antral Follicles. Reproduction in Domestic Animals, 48(6), 1025-1033.

Barros, V. R. P., Monte, A. P. O., Santos, J. M. S., Lins, T. L. B. G., Cavalcante, A. Y. P., Gouveia, B. B., Muller, M. C., Junior, J. L. O., Donfack, N. J., Araújo, V. R., & Matos, M. H. T. (2020a). Melatonin improves development, mitochondrial function and promotes the meiotic resumption of sheep oocytes from in vitro grown secondary follicles. Theriogenology, 144, 67-73.

Barros, V. R. P., Monte, A. P. O., Santos, J. M. S., Lins, T. L. B. G., Cavalcante, A. Y. P., Gouveia, B. B., Muller, M. C., Junior, J. L. O., Barberino, R. S., Donfack, N. J., Araújo, V. R., & Matos, M. H. T. (2020b). Effects of melatonin on the in vitro growth of early antral follicles and maturation of ovine oocytes. Domestic Animal Endocrinology, 71, 106386.

Bennemann, J., Grothmann, H., & Wrenzycki, C. (2018). Reduced oxygen concentration during in vitro oocyte maturation alters global DNA methylation in the maternal pronucleus of subsequent zygotes in cattle. Molecular Reproduction and Development, 85(11), 849-857.

Bezerra, M. E. S., Monte, A. P. O., Barberino, R. S., Lins, T. L. B. G., Junior, J. L. O., Santos, J. M. S., Bezerra, D. O., Neves, C. A., Silva, G. S., Carvalho, M. A. M., & Matos, M. H. T. (2019). Conditioned medium of ovine Wharton’s jelly-derived mesenchymal stem cells improves growth and reduces ROS generation of isolated secondary follicles after short-term in vitro culture. Theriogenology, 125, 56-63.

Bhardwaj, R., Ansari, M. M., Parmar, M. S., Vikash, C., & Sharma, G. T. (2016). Stem Cell Conditioned Media Contains Important Growth Factors and Improves In vitro Buffalo Embryo Production. Animal Biotechnology, 27, 118-125.

Blau, H. M., Brazelton, T. R., & Weimann, J. M. (2001). The evolving concept of a stem cell: entity or function? Cell, 105(7), 829-841.

Bomfim, M. M., Andrade, G. M., Collado, M. D., Sangalli, J. R., Fontes, P. K., Nogueira, M. F. G., Meirelles, F. V., Silveira, J. C., & Perecin, F. (2017). Antioxidant responses and deregulation of epigenetic writers and erasers link oxidative stress and DNA methylation in bovine blastocysts. Molecular Reproduction and Development, 84(12), 1296-1305.

Bongso, A., & Fong, C.Y. (2013). The therapeutic potential, challenges and future clinical directions of stem cells from the Wharton’s jelly of the human umbilical cord. Stem Cell Reviews and Reports, 9(2), 226-240.

Brasileiro, A. M M. (2013) Manual de produção de textos acadêmicos e científicos. São Paulo: Atlas. 47 páginas.

Bunkar, N., Pathak, N., Lohiya, N. K., & Mishra, P. K. (2016). Epigenetics: A key paradigm in reproductive health. Clinical and Experimental Reproductive Medicine, 43(2), 59.

Bydlowski, S. P., Debes, A. A., Maselli, L. M. F., & Janz, F. L. (2009) Características biológicas das células-tronco mesenquimais. Revista Brasileira de Hematologia e Hemoterapia, 31(1), 25-35.

Cadenas, J., Leiva-Revilla, J., Vieira, L. A., Apolloni, L. B., Aguiar, F. L. N., Alves, B. G., Lobo, C. H., Rodrigues, A. P. R., Apgar, G. A., Smitz, J., Figueiredo, J. R., & Maside, C. (2017). Caprine ovarian follicle requirements differ between preantral and early antral stages after IVC in medium supplemented with GH and VEGF alone or in combination. Theriogenology, 87, 321-332.

Cadenas, J., Maside, C., Ferreira, A. C. A., Vieira, L. A., Leiva-Revilla, J., Paes, V., Alves, B. G., Brandão, F. Z., Rodrigues, A. P. R., Wheeler, M. B., & Figueiredo, J. R. (2018). Relationship between follicular dynamics and oocyte maturation during in vitro culture as a non-invasive sign of caprine oocyte meiotic competence. Theriogenology, 107, 95-103.

Cavalcante, A.Y. P., Gouveia, B. B., Barberino, R. S., Lins, T. L., Santos, L. P., Gonçalves, R. J., Celestino, J. J., & Matos, M. H. (2015). Kit ligand promotes the transition from primordial to primary follicles after in vitro culture of ovine ovarian tissue. Zygote, 24, 578-582.

Cavalcante, B. N., Matos‐Brito, B. G., Paulino, L. R., Silva, B. R., Aguiar, A. W. M., Almeida, E. F. M., Souza, A. L. P., Vasconcelos, G. L., Ernando, I. T. A., Silva, A. W. B., & Silva, J. R. B. (2019). Effects of melatonin on morphology and development of primordial follicles during in vitro culture of bovine ovarian tissue. Reproduction in Domestic Animals, 54(12), 1567-1573.

Chen, Z., LI, S., Subramaniam, S., Shyy, J. Y. J., & Chien, S. (2017). Epigenetic regulation: a new frontier for biomedical engineers. Annual Review of Biomedical Engineering, 19, 195-219.

Chelenga, M., Sakaguchi, K., Abdel-Ghani, M. A., Yanagawa, Y., Katagiri, S., & Nagano, M. (2020). Effect of increased oxygen availability and astaxanthin supplementation on the growth, maturation and developmental competence of bovine oocytes derived from early antral follicles. Theriogenology, 157, 341-349.

Cloos, P. A., Christensen, J., Agger, K., Maiolica, A., Rappsilber, J., Antal, T., Hansen, K. H., & Helin, K. (2006). Histone demethylation by a family of JmjC domain-containing proteins. Nature, 439(7078), 811-816.

Costa, S. L., Costa, E. P., Pereira, E. C. M., Benjamin, L. A., Rodrigues, M. T., Mendes, V. R. A., & Silva, T. F. (2014). Influence of Insulin-like Growth Factor I (IGF-I) on the survival and the in vitro development of caprine preantral follicles. Pesquisa Veterinária Brasileira, 34, 1037-1044.

Csaki, C., Matis, U., Mobasheri, A., Ye, H., & Shakibaei, M. (2007). Chondrogenesis, osteogenesis and adipogenesis of canine mesenchymal stem cells: a biochemical, morphological and ultrastructural study. Histochemistry and Cell Biology, 128, 507-520.

Covarrubias, L., Hernández-García, D., Schnabel, D., Salas-Vidal, E., & Castro-Obregón, S. (2008). Function of reactive oxygen species during animal development: passive or active? Developmental Biology, 320(1), 1-11.

Dahl, J. A., Jung, I., Aanes, H., Greggains, G. D., Manaf, A., Lerdrup, M., Li, G., Kuan, S., Li, B., Lee, A. Y., Preissl, S., Jermstad, I., Haugen, M. H., Suganthan, R., Bjoras, M., Hansen, K., Dalen, K. T., Fedorcsak, P., Bing, R., & Klungland, A. (2016). Broad histone H3K4me3 domains in mouse oocytes modulate maternal-to-zygotic transition. Nature, 537(7621), 548-552.

Devine, P. J., Pereeault, S. D., & Luderer, U. (2012). Roles of reactive oxygen species and antioxidants in ovarian toxicity. Biology Reproduction, 86(2), 1-10.

Dominici, M., Le Blanc, K., Mueller, I., Slaper-Cortenbach, I., Marini, F. C., Krause, D. S., Deans, R. J., Keating, A., Prockop, D. J., & Horwitz, E. M. (2006). Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy, 8(4), 315-317.

Du Plessis, S. S., Makker, K., Desai, N. R., & Agarwal, A. (2008). Impact of oxidative stress on IVF. Expert Review of Obstetrics & Gynecology, 3, 539-554.

Durlej, M. G., Duda, M., Knapcyk, K., & Słomczyńska, M. (2008). Effects of transferrin on aromatase activity in porcine granulosa cells in vitro. Folia Histochemica Et Cytobiologica, 46, 423-428.

Edson, M. A., Nagaraja, A. K., & Matzuk, M. M. (2009). The mammalian ovary from genesis to revelation. Endocrine Reviews, 30(6), 624-712.

Elfayomy, A. K., Almasry, S. M., El-Tarhouny, S. A., & Eldomiaty, M. A. (2016). Human umbilical cord blood-mesenchymal stem cells transplantation renovates the ovarian surface epithelium in a rat model of premature ovarian failure: Possible direct and indirect effects. Tissue and Cell, 48, 370-382.

Eppig, J. J., & Downs, S. M. (1987). The effect of hypoxanthine on mouse oocyte growth and development in vitro: maintenance of meiotic arrest and gonadotropin-induced oocyte maturation. Developmental Biology, 199, 313-321.

Eppig, J. J., & O'brien, M. J. (1996). Development in vitro of mouse oocytes from primordial follicles. Biology of Reproduction, 54, 197-207.

Faita, T., Silva, V. N., Sattin, W. R., Pinheiro, A. O., & Ambrósio, C. E. (2016). Membrana amniótica: fonte alternativa de células-tronco mesenquimais em diversas espécies animais. Pesquisa Veterinária Brasileira, 36(6), 520-525.

Fang, Z., Yang, S., & Wu, G. (2002). Radicais livres, antioxidantes e nutrição. Nutrição, 18, 872- 879.

Ferreira, A. C. A., Maside, C., Sá, N. A. R., Guerreiro, D. D., Correia, H. H. V., Leiva-Revilla, J., Lobo, C. H., Araújo, V. R., Apgar, G. A., Brandão, F. Z., Figueiredo, J. R., & Campello, C. C. (2016). Balance of insulin and FSH concentrations improves the in vitro development of isolated goat preantral follicles in medium containing GH. Animal Reproduction Science, 165, 1-10.

Ferreira, A. C. A., Sá, N. A., Cadenas, J., Correia, H. H., Guerreiro, D. D., Alves, B. G., Lima, L. F., Celestino, J. J. H., Rodrigues, A. P. R., Gastal, E. L., & Figueiredo, J. R. (2020). Pituitary porcine FSH, and recombinant bovine and human FSH differentially affect growth and relative abundances of mRNA transcripts of preantral and early developing antral follicles in goats. Animal Reproduction Science, 219, 106461.

Figueiredo, J. R., Hulshof, S. C., Van Den Hurk, R., Nusgens, B., Bevers, M. M., Ectors, F. J., & Beckers, J. F. (1994). Preservation of oocyte and granulosa cell morphology in bovine preantral follicles cultured in vitro. Theriogenology, 41, 1333-1346.

Figueiredo, J. R., Rodrigues, A. P. R., Amorim, C. A., & Silva, J. R. V. Manipulação de oócitos inclusos em folículos ovarianos pré-natais, In: Gonçalves PBD, Figueiredo JR, Freitas VJF (Editors). (2008). Biotécnicas Aplicadas a Reprodução animal. São Paulo: Roca, 261-291.

Figueiredo, J. R., & Lima, L. F. (2017). Tecnologia do ovário artificial: aplicações, estado da arte, limitações e perspectivas. Revista Brasileira de Reprodução Animal, 41(1), 248-253.

Figueiredo, J. R., Cadenas, J., Lima, L. F., & Santos, R. R. (2019). Advances in in vitro folliculogenesis in domestic ruminants. Animal Reproduction (AR), 16(1), 52-65.

Fortune, J. E., Rivera, G. M., Evans, A. C. O., & Turzillo, A. M. (2001). Differentiation of dominant versus subordinate follicles in cattle. Biology Reproduction, 65, 648-645.

Fujii, J., Iuchi, Y., & Okada, F. (2005). Fundamental roles of reactive oxygen species and protective mechanisms in the female reproductive system. Reproduction Biology Endocrinology, 3, 43.

Glanzner, W. G., Wachter, A., Coutinho, A. R. S., Albornoz, M. S., Duggavathi, R., Gonçalves, P. B., & Bordignon, V. (2017). Altered expression of BRG1 and histone demethylases, and aberrant H3K4 methylation in less developmentally competent embryos at the time of embryonic genome activation. Molecular Reproduction and Development, 84(1), 19-29.

Glanzner, W. G., Rissi, V. B., Macedo, M. P., Mujica, L. K. S., Gutierrez, K., Bridi, A., Souza, J. R. M., Gonçalves, P. B. D., & Bordignon, V. (2018). Histone 3 lysine 4, 9, and 27 demethylases expression profile in fertilized and cloned bovine and porcine embryos. Biology of Reproduction, 98(6), 742-751.

Green, L. J., Zhou, H., & Shikanov, A. (2016). Utilization of adipose derived stem cells for the in vitro maturation of primary and early secondary ovarian follicles. Fertility and Sterility, 106, e49.

Gupta, P. S. P., Ramesh, H. S., Manjunatha, B. M., Nandi, S., & Ravindra, J. P. (2008). Production of buffalo embryos using oocytes from in vitro grown preantral follicles. Zygote, 16, 57-63.

Hammadeh, N., Coomarasamy, A., Ola, B., Papaioannou, S., Afnan, M., & Sharif, K. (2008). Ultrasound-guided hydrosalpinx aspiration during oocyte collection improves outcome in IVF: a randomized controlled trial. Human Reproduction, 23, 1113- 1117.

Hiyun, K., Jeon, J., Park, K., & Kim, J. (2017). Writing, erasing and reading histone lysine methylations. Experimental & Molecular Medicine, 49, 324.

Huang, W., Nagano, M., Kang, S. S., Yanagawa, Y., & Takahashi, Y. (2013). Effects of in vitro growth culture duration and prematuration culture on maturational and developmental competences of bovine oocytes derived from early antral follicles. Theriogenology, 80(7), 793-799.

Huang, J., Zhang, H., Wang, X., Dobbs, K. B., Yao, J., Qin, G., Whitworth, K., Walters, E. M., Prather, R. S., & Zhao, J. (2015). Impairment of preimplantation porcine embryo development by histone demethylase KDM5B knockdown through disturbance of bivalent H3K4me3-H3K27me3 modifications. Biology of Reproduction, 92(3).

Idelchik, M. D. P. S., Begley, U., Begley, T. J., & Melendez, J. A. (2017). Seminars in Cancer Biology. Academic Press, 57-66.

Itoh, T., Kacchi, M., Abe, H., Sendai, Y., & Hoshi, H. (2002). Growth, Antrum Formation, and Estradiol Production of Bovine Preantral Follicles Cultured in a Serum-Free Medium. Biology of Reproduction, 67, 1099-1105.

Juengel, J. L., Sawyer, H. R., Smith, P. R., Quirke, L. D., Heath, D. A., Lun, S., Wakefield, S. J., Mcnatty, K. P. (2002). Origins of follicular cells and ontogeny of steroidogenesis in ovine fetal ovaries. Molecular and Cellular Endocrinology, 191, 1-10.

Kaipia, A., & Hsueh, A. J. (1997). Regulation of ovarian follicle atresia. Annual Review of Physiology, 59(1), 349-363.

Klose, R. J., Yamane, K., Bae, Y., Zhang, D., Erdjument-Bromage, H., Tempst, P., Wong, J., & Zhang, Y. (2006). The transcriptional repressor JHDM3A demethylates trimethyl histone H3 lysine 9 and lysine 36. Nature, 442(7100), 312-316.

Laguerre, M., Lecomte, J., & Villeneuve, P. (2007). Evaluation of the ability of antioxidants to counteract lipid oxidation: Existing methods, new trends and challenges. Progress in Lipid Research, 46(5), p. 244-282.

La Rocca, G. (2011). Connecting the dots: The promises of wharton’s jelly stem cells for tissue repairand regeneration. The Open Tissue Engineering and Regenerative Medicine Journal, 4, 3-5.

Lee, S. E., Moon, J. J. M., Kim, E. Y., & Park, S. P. (2015). Stem cell–derived bioactive materials accelerate development of porcine in vitro–fertilized embryos. Cellular Reprogramming (Formerly" Cloning and Stem Cells"), 17(3), 181-190.

Leitão, C. C. F., Costa, J. J. N., Brito, I. R., Magalhães-Padilha, D. M., Almeida, A. P., Figueiredo, J. R., Van Den Huk, R., & Silva, J. R. V. (2014). Effects of GDF-9 and FSH on mRNA expression for FSH-R, GDF-9 and BMPs in in vitro cultured goat preantral follicles. Brazilian Archives of Biology and Technology, 57(2), 200-208.

Lima-Verde, I. B., Matos, M. H. T., Bruno, J. B., Martins, F. S., Santos, R. R., Báo, S. N., Luque, M. C. A., Vieira, G. A. B., Silveira, E. R., Rodrigues, A. P. R., Figueiredo, J. R., Oliveira, M. A. L., & Lima, P. F. (2009). Effects of α-tocopherol and ternatin antioxidants on morphology and activation of goat preantral follicles in vitro cultured. Arquivo Brasileiro de Medicina Veterinária e Zootecnia, 61(1), 57-65.

Lins, T. L. B. G., Cavalcante, A. Y. P., Santos, J. M. S., Menezes, V. G., Barros, V. R. P., Barberino, R. S., Bezerra, M. E. S., Macedo, T. J. S., & Matos, M. H. T. (2017). Rutin can replace the use of three other antioxidants in the culture medium, maintaining the viability of sheep isolated secondary follicles. Theriogenology, 89, 263-270.

Liu, W., Liu, X., Wang, C., Gao, Y., Gao, R., Kou, X., Zhao, Y., Li, J., Wu, Y., Xiu, W., Wang, S., Yin, J., Liu, W., Cai, T., Wang, H., Zhang, Y., & Gao, S. (2016). Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing. Cell Discovery, 2(1), 1-15.

Liu, J., Zhang, W., Wu, Z., Dai, L., & Koji, T. (2018). Changes in DNA methylation of oocytes and granulosa cells assessed by HELMET during folliculogenesis in mouse ovary. Acta Histochemica et Cytochemica, 17039.

Liu, Z., Zhang, G., Deng, M., Yang, H., Pang, J., Cai, Y., Wan, Y., Wang, F. (2020). Inhibition of lysine-specific histone demethylase 1A results in meiotic aberration during oocyte maturation in vitro goats. Theriogenology, 143, 168-178.

Macedo, T. J. S., Barros, V. R. P., Monte, A. P. O., Gouveia, B. B., & Bezerra, M. É. S. (2017). Resveratrol has dose-dependent effects on DNA fragmentation and mitochondrial activity of ovine secondary follicles cultured in vitro. Zygote, 25(4), 434.

Macedo, T. J. S., Santos, J. M. S., Bezerra, M. É. S., Menezes, V. G., Gouveia, B. B., Barbosa, L. M. R., Lins, T. L. B. G., Monte, A. P. O., Barberino, R. S., Bastista, A. M., Barros, V. R. P., Wischral, A., Queiroz, M. A. A., Araújo, G. G. L., & Matos, M. H. T. (2019). Immunolocalization of leptin and its receptor in the sheep ovary and in vitro effect of leptin on follicular development and oocyte maturation. Molecular and Cellular Endocrinology, 495, 110506.

Magalhães, D. M., Duarte, A. B. G., Araújo, V. R., Brito, I. R., Soares, T. G., Lima, I. M. T., Lopes, C. A. P., Campello, C. C., Rodrigues, A. P. R., & Figueiredo, J. R. (2011). In vitro production of a caprine embryo from a preantral follicle cultured in media supplemented with growth hormone. Theriogenology, 75, 182-188.

Magalhães-Padilha, D. M., Geisler-Lee, J., Wischral, A., Gastal, M. O., Fonseca, G. R., Eloy, Y. R. G., Geisler, M., Figueiredo, J. R., & Gastal, E. L. (2013). Gene expression during early folliculogenesis in goats using microarray analysis. Biology of Reproduction, 89, 1-12.

Manabe, N., Goto, Y., Matsuda-Minehata, F., Inoue, N., Maeda, A., Sakamaki, K., & Miyano, T. (2004). Regulation mechanism of selective atresia in porcine follicles: regulation of granulosa cell apoptosis during atresia. Journal of Reproduction and Development, 50(5), 493-514.

Matsuda, F., Inoue, N., Goto, Y., Maeda, A., Cheng, Y., Sakamaki, K., & Manabe, N. (2008). cFLIP regulates death receptor-mediated apoptosis in an ovarian granulosa cell line by inhibiting procaspase-8 cleavage. Journal of Reproduction and Development, 54(5), 314-320.

Martins, G. R., Teixeira, M. F. S., Junior, R. Q. B., Dias, R. P., Aguiar, T. D. F., Marinho, R. C., & Pinheiro, A. R. A. (2014). Células-tronco mesenquimais: características, cultivo e uso na Medicina Veterinária. Revista Brasileira de Higiene e Sanidade Animal, 8, 181-202.

Mcgee, E. A., & Hsueh, A. J. (2000). Initial and cyclic recruitment of ovarian follicles. Endocrine Reviews, 21(2), 200-214.

Mclaughlin, E. A., & Mciver, S. C. (2009). Awakening the oocyte: controlling primordial follicle development. Reproduction, 137, 1-11.

Mclaughlin, M., & Telfer, E. E. (2010). Oocyte development in bovine primordial follicles is promoted by activin and FSH within a two-step serum-free culture system. Reproduction, 139, 971-978.

Mcnatty, K. P., Fidler, A. E., Juengel, J. L., Quirke, L. D., Smith, P. R., Heath, D. A., Lundy, T., O’connell, A., & Tisdall, D. J. (2000). Growth and paracrine factors regulating folicular formation and cellular function. Molecular and Cellular Endocrinology, 163, 11-20.

Metere, A., & Graves, C. E. (2020). Factors Influencing Epigenetic Mechanisms: Is There A Role for Bariatric Surgery? High-Throughput, 9(1), 6.

Menezes, V. G. (2017). Efeito do ácido protocatecuico sobre o desenvolvimento in vitro de folículos secundários ovinos isolados. Dissertação (Mestrado em Ciência Animal). Universidade Federal do Vale do São Francisco, Campus de Ciências Agrárias, Petrolina - PE, 81f.

Menezes, V. G., Monte, A. P. O., Gouveia, B. B., Lins, T. L. B. G., Donfack, N. J., Macedo, T. J. S., Barberino, R. S., Santos, J. M., Matos, M. H. T., & Batista, A. M., Wischral, A. (2019). Effects of leptin on the follicular development and mitochondrial activity of ovine isolated early antral follicles cultured in vitro. Theriogenology, 138, 77-83.

Morais, M. L. G. S. (2018). Efeitos da suplementação de anetol ou robinina na vitrificação e incubação in vitro do tecido ovariano ovino. Dissertação (Mestrado em Ciências Morfofuncionais) - Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, 69 f.

Morohaku, K., Tanimoto, R., Sasaki, K., Kawahara-Miki, R., Kono, T., Hayashi, K., Hirao, Y., & Obata, Y. (2016). Complete in vitro generation of fertile oocytes from mouse primordial germ cells. Proccedings of the National Academy of Sciences of the United States, 113, 9021-9026.

Nacev, B. A., Feng, L., Lemiesz, A. E., Gao, J., Soshnev, A. A., Kundra, R., Schultz, N., Muir, T. W., & Allis, C. D. (2019). The expanding landscape of ‘oncohistone’mutations in human cancers. Nature, 567(7749), 473-478.

Neves, C. A., Silva, L. S., Carvalho, C. E. S., Carvalho, M. S., Sarmento, J. L. R., Cavalcante, T. V., Arrivabene, M., Neves, T. A., Bezerra, M. E. S., Júnior, A. L. G., Silva, C. M. G., & Carvalho, A. C. M. (2020). Culture of goat preantral follicles in situ associated with mesenchymal stem cell from bone marrow. Zygote, 28(1), 65-71.

O'brien, M. J., Pendola, J. K., & Eppig, J. J. (2003). A revised protocol for in vitro development of mouse oocytes from primordial follicles dramatically improves their developmental competence. Biology of Reproduction, 68, 1682-1686.

Okediran, B. S., Biobaku, K. T., Olaifa, F. H., & Atata, A. J. (2017). Haematological and antioxidant enzyme response to Lead toxicity in male Wistar rats. Ceylon Journal of Science, 46(2), 31-37.

Oktem, O., & Oktay, K. (2008). The ovary: anatomy and function throughout human life. Annals of the New York Academy of Sciences, 1127(1), 1-9.

Paes, V. M., Lima, L. F., Ferreira, A. C. A., Lobo, C. H., Alves, B. G., Rodrigues, A. P. R., Oliveira, A. C., Figueiredo, J. R., & Feugang, J. M. (2020). The subtle balance of insulin and thyroxine on survival and development of in vitro cultured caprine preantral follicles enclosed in ovarian tissue. Theriogenology, 147, 10-17.

Panta, A. M. T., Silva, A. F. B., Padilha, R. T., Correia, H. H. V., Rondina, D., Figueiredo, J. R., & Magalhães-Padilha, D. D. M. (2019). Evaluation of in vitro culture systems for goat preantral follicles using reused ovaries from reproductive biotechniques: An alternative to maximize the potential of reproduction. Reproduction in Domestic Animals, 54(3), 480-485.

Park, K. S., Kim, Y. S., Kim, J. H., Choi, B., Kim, S. H., Tan, A. H., Lee, M. S., Lee, M. K., Kwon, C. H., Joh, J. W., Kim, S. J., & Kim, K.W. (2010) Trophic molecules derived from human mesenchymal stem cells enhance survival, function, and angiogenesis of isolated islets after transplantation. Transplantation, 15, 509-517.

Pasini, D., Hansen, K. H., Christensen, J., Agger, K., Cloos, P. A., & Helin, K. (2008). Coordinated regulation of transcriptional repression by the RBP2 H3K4 demethylase and Polycomb-Repressive Complex 2. Genes & Development, 22(10), 1345-1355.

Pflum, M. K., Tong, J. K., Lane, W. S., & Schreiber, S. L. (2001). Histone deacetylase 1 phosphorylation promotes enzymatic activity and complex formation. Journal of Biological Chemistry, 276(50), 47733-47741.

Picton, H. M., Harris, S. E., Muruvi, W., & Chambers, E. L. (2008). The in vitro growth and maturation of follicles. Reproduction, 136(6), 703-715.

Polzin, G. M., Stanfill, S. B., Brown, C. R., Ashley, D. L., & Watson, C. H. (2007). Determination of eugenol, anethole, and coumarin in the mainstream cigarette smoke of Indonesian clove cigarettes. Food and Chemical Toxicology, 45, 1948-1953.

Qingming, Y., Xianhui, P., Weibao, K., Hong, Y., Yidan, S., Li, Z., Yanan, Z., Yuling, Y., Lan, D., & Guoan, L. (2010). Antioxidant activities of malt extract from barley (Hordeum vulgare L.) toward various oxidative stress in vitro and in vivo. Food Chemistry, 118, (1), 84-89.

Richards, J. S., & Pangas, S. A. (2016). The ovary: basic biology and clinical implications. Journal of Clinical Investigation, 120, 963-972.

Rocha, R. M., Lima, L. F., Alves, A. M., Celestino, J. J., Matos, M. H., Lima-Verde, I. B., Bernuci, M. P., Lopes, C. A., Báo, S. N., Campello, C. C., Rodrigues, A. P., & Figueiredo, J. R. (2013). Interaction between melatonin and follicle-stimulating hormone promotes in vitro development of caprine preantral follicles. Domestic Animal Endocrinology, 44, 1-9.

Rodrigues, P., Limback, D., Mcginnis, L. K., Plancha, C. E., & Albertini, D. F. (2008). Oogenesis: prospects and challenges for the future. Journal of Cellular Physiology, 216(2), 355-365.

Rodrigues, G. Q., Silva, C. M. G., Faustino, L. R., Bruno, J. B., Magalhães, D. M., Campello, C. C., & Figueiredo, J. R. (2010). Bovine serum albumin improves in vitro development of caprine preantral follicles. Animal Reproduction, 7, 382-388.

Rossetto, R., Lima-Verde, I. B., Matos, M. H. T., Saraiva, M. V. A., Martins, F. S., Faustino, L. R., Araújo, V. R., Silva, C. M. V., Name, K. P. O., Báo, S. N., Campello, C. C., Figueiredo, J. R., & Blume, H. (2009). Interaction between ascorbic acid and follicle-stimulating hormone maintains follicular viability after long-term in vitro culture of caprine preantral follicles. Domestic Animal Endocrinology, 37, (2), 112-123.

Sá, N. A. R., Araújo, V. R., Correia, H. H. V., Ferreira, A. C. A., Guerreiro, D. D., Sampaio, A. M., Escobar, E., Santos, F. W., Moura, A. A., Lôbo, C. H., Ceccatto, V. M., Campello, C. C., Rodrigues, A. P. R., Leal-Cardoso, J. H., & Figueiredo, J. R. (2017). Anethole improves the in vitro development of isolated caprine secondary follicles. Theriogenology, 89, 226-234.

Sá, N. A. R., Bruno, J. B., Guerreiro, D. D., Cadenas, J., Alves, B. G., Cibin, F. W. S., Leal-Cardoso, J. H., Gastal, E. L., & Figueiredo, J. R. (2018). Anethole reduces oxidative stress and improves in vitro survival and activation of primordial follicles. Brazilian Journal of Medical and Biological Research, 51(8).

Sá, N. A., Ferreira, A. C. A., Sousa, F. G. C., Duarte, A. B. G., Paes, V. M., Cadenas, J., Anjos, J. C., Fernandes, C. C. L., Rosseto, R., Cibin, F. W. S., Alves, B. G., Rodrigues, A. P. R., Rondina, D., Gastal, E. L., & Figueiredo, J. R. (2020). First pregnancy after in vitro culture of early antral follicles in goats: Positive effects of anethole on follicle development and steroidogenesis. Molecular Reproduction and Development, 87(9), 966-977.

Saeedabadi, S., Abazari-Kia, A. H., Rajabi, H., Parivar, K., & Salehi, M. (2018). Melatonin improves the developmental competence of goat oocytes. International Journal of Fertility & Sterility, 12(2), p. 157.

Sahebkar, A., Serban, M. C., Ursoniu, S., & Banach, M. (2015). Effect of curcuminoids on oxidative stress: A systematic review and meta-analysis of randomized controlled trials. Journal of Functional Foods, 18, 898-909.

Sales, A. D., Duarte, A. B. G., Rodrigues, G. Q., Lima, L. F., Silva, G. M., Carvalho, A. A., Brito, I. R., Maranguape, R. M. S., Lobo, C. H., Aragão, J. A. S., Moura, A. A., Figueiredo, J. R., & Rodrigues, A. P. R. (2015). Steady-state level of messenger RNA and immunolocalization of aquaporins 3, 7, and 9 during in vitro growth of ovine preantral follicles. Theriogenology, 84(1), 1-10.

Santos, L. P., Barros, V. R., Cavalcante, A. Y., Menezes, V. G., Macedo, T. J., Santos, J. M., Araujo, V. R., Queiroz, M. A., & Matos, M. H. (2014a). Protein localization of epidermal growth factor in sheep ovaries and improvement of follicle survival and antrum formation in vitro. Reproduction in Domestic Animals, 49, 783-789.

Santos, J. M. S., Menezes, V. G., Barberino, R. S., Macedo, T. J., Lins, T. L., Gouveia, B. B., Barros, V. R., Santos, L. P., Gonçalves, R. J., & Matos, M. H. (2014b). Immunohistochemical Localization of Fibroblast Growth Factor-2 in the Sheep Ovary and its Effects on Pre-antral Follicle Apoptosis and Development In vitro. Reproduction in Domestic Animals, 49, 522-528.

Santos, J. M. S., Monte, A. P. O., Lins, T. L. B. G., Barberino, R. S., Menezes, V. G., Gouveia, B. B., Macedo, T. J. S., Júnior, J. L. O., Donfack, N. J., & Matos, M. H. T. (2019). Kaempferol can be used as the single antioxidant in the in vitro culture medium, stimulating sheep secondary follicle development through the phosphatidylinositol 3-kinase signaling pathway. Theriogenology, 136, 86-94.

Schafer, F., & Buettner, G. R. (2001). Redox environment of the cell as viewed through the redox state of the glutathione disulfide/glutathione couple. Free Radical Biology and Medicine, 11, 1191-1212.

Schipani, E., Kronenberg, H. M. (2009). Adult mesenchymal stem cells. In: Harvard Stem Cell Institute. Stembook, Cambridge, MA: The Stem Cell Research Community, 1-7.

Seneda, M. M., Godmann, M., Murphy, B. D., Kimmins, S., & Bordignon, V. (2008). Developmental regulation of histone H3 methylation at lysine 4 in the porcine ovary. Reproduction, 135(6), 829-838.

Sha, Q. Q., Jiang, Y., Yu, C., Xiang, Y., Dai, X. X., Jiang, J. C., Ou, X. H., Fan, H. Y. (2020). CFP1-dependent histone H3K4 trimethylation in murine oocytes facilitates ovarian follicle recruitment and ovulation in a cell-nonautonomous manner. Cellular and Molecular Life Sciences, 77(15), 2997-3012.

Silva, G. M., Araujo, V. R., Duarte, A. B. G., Lopes, C. A. P., & Figueiredo, J. R. (2011a). Papel dos antioxidantes no cultivo in vitro de células ovarianas. Revista Brasiliera de Reprodução Animal, 35(3), 315-326.

Silva, G. M., Araújo, V. R., Duarte, A. B. G., Chaves, R. N., Silva, C. M. G., Lobo, C. H., Almeida, A. P., Matos, M. H. T., Tavares, L. M. T., Campelo, C. C., & Figueiredo, J.R. (2011b). Ascorbic acid improves the survival and in vitro growth of isolated caprine preantral follicles. Animal Reproduction, 8, 14-24.

Silva, C. T. D., & Jasiulionis, M. G. (2014). Relação entre estresse oxidativo, alterações epigenéticas e câncer. Ciência e Cultura, 66(1), 38-42.

Souza-Cáceres, M. B., & Melo-Sterza, F. A. (2017). Metilação de histonas em oócitos de embriões mamíferos. Revista Brasileira de Reprodução Animal, 4(2), 620-627.

Sousa, R. P., Duarte, A. B. G., Pinto, Y., Sá, N. A. R., Alves, B. G., Cibin, F. W. S., Silva, G. C., Carvalho, C. E. S., Neto, N. M. A., Rodrigues, A. P. R., Silva, C. M. G., Figueiredo, J. R., & Carvalho, M. A. M. (2021). In vitro activation and development of goat preantral follicles enclosed in ovarian tissue co-cultured with mesenchymal stem cells. Reproductive Sciences.

Spitschak, M., & Hoeflich, A. (2018). Potential functions of IGFBP-2 for ovarian folliculogenesis and steroidogenesis. Frontiers in Endocrinology, 9, 119.

Takahashi, K., Tanabe, K., Ohnuki, M., Narita, M., Ichisaka, T., Tomoda, K., & Yamanaka, S. (2007). Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell, 131(5), 861-872.

Telfer, E. E. (1998). In vitro models for oocyte development. Theriogenology, 15(492), 451-60.

Thomas, F. H., Leask, R., Srsen, V., Riley, S. C., Spears, N., & Telfer, E. E. (2001). Effect of ascorbic acid on health and morphology of bovine preantral follicles during long-term culture. Reproduction-Cambridge, 122(3), 487-495.

Torres-Osorio, V., Urrego, R., Echeverri-Zuluaga, J. J., & López-Herrera, A. (2019). Oxidative stress and antioxidant use during in vitro mammal embryo production. Review. Revista Mexicana de Ciencias Pecuarias, 10(2), 433-459.

Tsukada, Y. I., Fang, J., Erdjument-Bromage, H., Warren, M. E., Borchers, C. H., Tempst, P., & Zhang, Y. (2006). Histone demethylation by a family of JmjC domain-containing proteins. Nature, 439(7078), 811-816.

Tyndall, A., & Pistoia, V. (2009). Mesenchymal stem cells combat sepsis. Nature Medicine, 15(1), 18-20.

Vaanholt, L. M., Milne, A., Zheng, Y., Hambly, C., Mitchell, S. E., Valencak, T. G., Allison, D. B., & Speakman, J. R. (2016). Oxidative costs of reproduction: Oxidative stress in mice fed standard and low antioxidant diets. Physiology and Behavior, 154, 1-7.

Van Den Hurk, R., & Zhao, J. (2005). Formation of mammalian oocytes and their growth, differentiation and maturation within ovarian follicles. Theriogenology, 63, 1717-1751.

Vasconcelos, E. M., Costa, F. C., Azevedo, A. V. N., Barroso, P. A. A., Assis, E. I. T., Paulino, L. R. F. M., Silva, B. R., Silva, A. W. B., Souza, A. L. P., & Silva, J. R. V. (2021). Eugenol influences the expression of messenger RNAs for superoxide dismutase and glutathione peroxidase 1 in bovine secondary follicles cultured in vitro. Zygote, 1-6.

Wrenzycki, C., & Niemann, H. (2003). Epigenetic reprogramming in early embryonic development: Effects of in-vitro production and somatic nuclear transfer. Reproductive Biomedicine Online, 7, 649-656.

Yamamoto, K., Otoi, T., Koyama, N., Horikita, N., Tachikawa, S., & Miyano, T. (1999). Development to live young from bovine small oocytes after growth, maturation and fertilization in vitro. Theriogenology, 52(1), 81-89.

Young, I. S., & Woodside, J. V. (2001). Antioxidants in health and disease. Antioxidants in health and disease. Journal of Clinical Pathology, 54(3), 176-186.

Yu, X. X., Liu, Y. H., Liu, X. M., Wang, P. C., Liu, S., Miao, J. K., Du, Z. Q., & Yang, C. X. (2018). Ascorbic acid induces global epigenetic reprogramming to promote meiotic maturation and developmental competence of porcine oocytes. Scientific Reports, 8(1), 6132.

Zhou, J., Peng, X., & Mei, S. (2019). Autophagy in ovarian follicular development and atresia. International Journal of Biological Sciences, 15(4), 726.



How to Cite

SILVA, A. F. B. da .; LIMA, L. F. de .; FIGUEIREDO, J. R. de. Strategies for improving follicular culture efficiency in vitro: Importance of medium supplementation and study of epigenetic changes. Research, Society and Development, [S. l.], v. 10, n. 9, p. e22910918022, 2021. DOI: 10.33448/rsd-v10i9.18022. Disponível em: Acesso em: 23 sep. 2021.



Agrarian and Biological Sciences