Current progress, challenges, and future prospects of testis organoids†

Abstract

Spermatogenic failure is believed to be a major cause of male infertility. The establishment of a testis organoid model would facilitate the study of such pathological mechanisms and open the possibility of male fertility preservation. Due to the complex structures and cellular events occurring within the testis, the establishment of a compartmentalized testis organoid with a complete spermatogenic cycle remains a challenge in all species. Since the early 20th century, a great variety of scaffold-based and scaffold-free testis cell culture systems have been established to recapitulate de novo testis organogenesis and in vitro spermatogenesis. The utilization of the hydrogel scaffolds provides a 3D microenvironment for testis cell growth and development, facilitating the reconstruction of de novo testis tissue-like structures and spermatogenic differentiation. Using a combination of different strategies, including the use of various scaffolding biomaterials, the incorporation of the living cells with high self-assembling capacity, and the integration of the advanced fabrication techniques, a scaffold-based testis organoid with a compartmentalized structure that supports in vitro spermatogenesis may be achieved. This article briefly reviews the current progress in the development of scaffold-based testis organoids while focusing on the scaffolding biomaterials (hydrogels), cell sources, and scaffolding approaches. Key challenges in current organoid studies are also discussed along with recommendations for future research.

Generation of a Highly Biomimetic Organoid, Including Vasculature, Resembling the Native Immature Testis Tissue

“…Our organoid system has therefore fulfilled the three main criteria for organoid formation, including (a) testis cell reassembly, (b) the compartmentalized architectures, and (c) the inclusion of major testis cell types. Since not all testis organoid studies reported have performed a complete analysis according to the above criteria for organoid formation, it is difficult to fully compare all testis organoid studies [ 3 ]. For example, some studies did not perform immunostaining of Leydig cells and PTMCs, leading to incomplete interpretation of the overall architecture, cell types, and spatial orientation in their systems [ 19 , 38 , 57 , 58 , 59 ].…”

“…The goal of an ideal testis organoid system is to reconstruct an artificial tissue that is structurally and functionally similar to intact testis tissue. Three criteria have been proposed by Edmonds and Woodruff (2020) to evaluate the testis organoid formation in a given culture system [3,15]. These criteria include (a) testis cell reassembly, (b) the presence of a compartmentalized architecture, and (c) the inclusion of major testis cell types (Sertoli, Leydig, germ, and PTMCs).…”

“…The establishment of an in vitro testis organoid system from dissociated testis cells has become a hot topic for research in regenerative medicine and reproductive biotechnology [ 1 , 2 , 3 ]. Testis organoids have the potential to be used as an important experimental model for biological investigations or pharmaco-toxicology testing of various factors for their effects on testis development and function [ 4 , 5 ].…”

“…Since the late 20th century, a wide range of testis organoid culture systems have been introduced to recapitulate de novo testis organogenesis [ 2 , 3 , 16 ]. Despite tremendous efforts, limitations including the lack of non-rodent models, progressive decrease in the number of germ cells, missing key testis cell types, improper cellular orientation, and incomplete de novo testis organogenesis were reported in these earlier culture systems [ 15 , 16 , 17 , 18 , 19 , 20 , 21 ].…”

Generation and Characterization of Bovine Testicular Organoids Derived from Primary Somatic Cell Populations

“…This protocol differs from those commonly used in 3D testicular models, where testicular cells are obtained through enzymatic digestion and the remotion of connective tissue [ 6 , 8 , 30 , 32 , 34 , 48 ]. These cell isolation protocols may unintentionally provide other cell types to the testicular cell mix, including endothelial-like or uncharacterized cell types that may interfere with further experiments [ 8 , 30 , 31 , 32 , 49 ]. Following our testicular cell isolation method, we obtained cultures with populations of 21–37% of Leydig cells, 65–77% of Sertoli cells and 10–16% of peritubular myoid cells.…”