Abstract Organoid is a tissue substitute that is structurally and functionally similar to the origin organ. At present, many organoid culture systems have been established, however testicular organoid culture is still in the exploratory stage. In this study, testicular tissues of Saanen dairy goats (Capra hircus) were taken, and testicular cells were isolated and cultured by mixed enzyme digestion. Testicular cells were cultured in three dimensions using a mixed medium containing low melting point agarose. The formed organoids were evaluated using light microscope and immunofluorescence detection techniques, and the culture effects of different sources of testicular cells, different numbers of testicular cells and different concentrations of low melting point agarose on testicular organoids were compared. Finally, the model function of the testicular organoid was verified by lipopolysaccharide (LPS) infection. The result showed that cells ranging from 2×105 to 3×105, in 1% low-melting-point agarose were more favorable for the formation of goat testicular organs in goats. The organoids formed possess seminiferous tubule-like structures, with spermatogonial stem cells and Sertoli cells evenly distributed within. Testicular organoid infected with LPS were smaller than normal organoids, exhibited cellular debris, and were accompanied by a significant increase in inflammatory factors, indicating that these organoids could respond to external stimuli and simulate the immune response of organs to a certain extent, making them a suitable model for the in vitro study of testicular diseases. This study not only promotes the development of testicular organoid culture technology, but also provides an important scientific basis and experimental platform for a variety of fields, including reproductive biology, disease modeling, drug screening, regenerative medicine, and personalized medicine.
XU Wen-Jing,WU Wen-Ping,YANG Yu-Mei, et al. Culture and Characterization of Testicular Organoid in Goats (Capra hircus)[J]. 农业生物技术学报, 2024, 32(9): 2173-2180.
[1] Alves-Lopes J P, Stukenborg J B.2018. Testicular organoids: A new model to study the testicular microenvironment in vitro?[J]. Human Reproduction Update, 24(2): 176-191. [2] Baert Y, De Kock J, Alves-Lopes J P, et al.2017. Primary human testicular cells self-organize into organoids with testicular properties[J]. Stem Cell Reports, 8(1): 30-38. [3] Chen Y W, Huang S X, de Carvalho A L R T, et al.2017. A three-dimensional model of human lung development and disease from pluripotent stem cells[J]. Nature Cell Biology, 19(5): 542-549. [4] Chu Z L, Liu C, Bai Y F, et al.2013. A three-dimensional (3D) environment to maintain the integrity of mouse testicular can cause the occurrence of meiosis[J]. Journal of Integrative Agriculture, 12(8): 1481-1488. [5] Edmonds M E, Woodruff T K.2020. Testicular organoid formation is a property of immature somatic cells, which self-assemble and exhibit long-term hormone-responsive endocrine function[J]. Biofabrication, 12(4): 045002. [6] Fatehullah A, Tan S H, Barker N2016. Organoids as an in vitro model of human development and disease[J]. Nature Cell Biology, 18(3): 246-254. [7] Gholami K, Pourmand G, Koruji M, et al.2018. Organ culture of seminiferous tubules using a modified soft agar culture system[J]. Stem Cell Research & Therapy, 9(1): 249. [8] Lee J H, Kim H J, Kim H, et al.2006. In vitro spermatogenesis by three-dimensional culture of rat testicular cells in collagen gel matrix[J]. Biomaterials, 27(14): 2845-2853. [9] Oliver E, Alves-Lopes J P, Harteveld F, et al.2021. Self-organising human gonads generated by a matrigel-based gradient system[J]. BMC Biology, 19(1): 212. [10] Quadrato G, Nguyen T, Macosko E Z, et al.2017. Cell diversity and network dynamics in photosensitive human brain organoids[J]. Nature, 545(7652): 48-53. [11] Ravi M, Paramesh V, Kaviya S R, et al.2015. 3D cell culture systems: Advantages and applications[J]. Journal of Cellular Physiology, 230(1): 16-26. [12] Richer G, Baert Y, Goossens E.2020. In-vitro spermatogenesis through testis modelling: Toward the generation of testicular organoids[J]. Andrology, 8(4): 879-891. [13] Rore H, Owen N, Piña-Aguilar R E, et al.2021. Testicular somatic cell-like cells derived from embryonic stem cells induce differentiation of epiblasts into germ cells[J]. Communications Biology, 4(1): 802. [14] Rossi G, Manfrin A, Lutolf M P.2018. Progress and potential in organoid research[J]. Nature Reviews Genetics, 19(11): 671-687. [15] Sakib S, Goldsmith T, Voigt A, et al.2020. Testicular organoids to study cell-cell interactions in the mammalian testis[J]. Andrology, 8(4): 835-841. [16] Sakib S, Uchida A, Valenzuela-Leon P, et al.2019. Formation of organotypic testicular organoids in microwell culture[J]. Biology of Reproduction, 100(6): 1648-1660. [17] Sato T, Vries R G, Snippert H J, et al.2009. Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche[J]. Nature, 459(7244): 262-U147. [18] Takebe T, Sekine K, Enomura M, et al.2013. Vascularized and functional human liver from an iPSC-derived organ bud transplant[J]. Nature, 499(7459): 481-484. [19] Vermeulen M, Del Vento F, Kanbar M, et al.2019. Generation of organized porcine testicular organoids in solubilized hydrogels from decellularized extracellular matrix[J]. International Journal of Molecular Sciences, 20(21): 5476. [20] Zhu H, Zheng L, Wang L, et al.2020. p53 inhibits the proliferation of male germline stem cells from dairy goat cultured on poly-L-lysine[J]. Reproduction in Domestic Animals, 55(3): 405-417.
