Abstract
The creation of a testis organoid (artificial testis tissue) with sufficient resemblance to the complex form and function of the innate testis remains challenging, especially using non-rodent donor cells. Here, we report the generation of an organoid culture system with striking biomimicry of the native immature testis tissue, including vasculature. Using piglet testis cells as starting material, we optimized conditions for the formation of cell spheroids, followed by long-term culture in an air–liquid interface system. Both fresh and frozen-thawed cells were fully capable of self-reassembly into stable testis organoids consisting of tubular and interstitial compartments, with all major cell types and structural details expected in normal testis tissue. Surprisingly, our organoids also developed vascular structures; a phenomenon that has not been reported in any other culture system. In addition, germ cells do not decline over time, and Leydig cells release testosterone, hence providing a robust, tunable system for diverse basic and applied applications.
Highlights
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]
We established a new testis organoid culture system using neonatal porcine testis cells by first forming cell spheroids in low attachment culture wells followed by long-term culture in an air–liquid interface system
Both fresh and cryopreserved testis cells were fully capable of forming testis organoids comprising testis cord-like structures, which could be maintained for at least 4 weeks
Summary
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]. Testicular organogenesis occurs during embryonic/fetal stages and as a result of a complex folding of the germinal layers as well as the formation, migration, and assembly of both germ cells and somatic cells [6]. These events lead to compartmentalization of the tissue into two morphologically and functionally distinct tubular and interstitial components. The tubular compartment is constructed primarily by two somatic cell types, peritubular myoid cells (PTMCs) and Sertoli cells, which harbor germ cells of various types undergoing development [10]. Disorders of the normal testis development and interferences in the production/action of androgens do occur [13], and can predispose the individual to early or long-term complications such as cryptorchid testis, testicular germ cell tumors, or impaired spermatogenesis [14]
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