Generation of Organized Porcine Testicular Organoids in Solubilized Hydrogels from Decellularized Extracellular Matrix

This method describes the generation of porcine testicular organoids using piglet testicular cells seeded in a testicular extracellular matrix (tECM) hydrogel. To generate the solublized tECM hydrogel, porcine immature testicular tissues (ITTs) were dissected in small fragments and decellularized in a 0.01% sodium dodecyl sulfate solution followed by agitation in a 1% Triton X-100 solution before being lyophilized and digested in a solution of HCl/pepsin. To generate the organoids, testicular cell suspensions were isolated from the porcine ITT and seeded into the hydrogel to form each organoid (500 000 cells per hydrogel). The generated organoids were then cultured in vitro and showed both an architecture and endocrine function that are similar to that found in the native organ in vivo.