Method of producing in vitro testicular constructs and uses thereof

1. A cell composition in the form of a three-dimensional artificial testicular construct, the composition comprising, in combination:

(a) spermatogonial stem cells, wherein said spermatogonial stem cells are not immortalized,

(b) immortalized Sertoli cells, and

(c) immortalized Leydig cells,

wherein said three-dimensional construct is in the form of an organoid and has a diameter of 100 to 300 microns, and

wherein said spermatogonial stem cells, immortalized Sertoli cells and immortalized Leydig cells are human cells.

2. The cell composition of claim 1 , wherein:

(i) said spermatogonial stem cells are included in an amount by number of cells of from 70 to 90 percent;

(ii) said immortalized Sertoli cells are included in an amount by number of cells of from 5 to 20 percent; and

(iii) said immortalized Leydig cells are included in an amount by number of cells from 5 to 20 percent.

3. The cell composition of claim 1 , wherein the ratio of (a):(b):(c) is 8:1:1.

4. The cell composition of claim 1 , further comprising peritubular cells.

5. A culture composition comprising the cell composition of claim 1 in an aqueous culture medium, said culture media further comprising at least one extracellular matrix (ECM) protein.

6. The culture composition of claim 5 , wherein said ECM protein comprises laminin, collagen type I, collagen type IV, fibronectin, elastin, or a combination thereof.

7. The culture composition of claim 5 , wherein said ECM protein comprises collagen type I.

8. The culture composition of claim 5 , wherein said at least one ECM protein is included in an amount of from 10 nanograms per milliliter to 1 milligram per milliliter.

9. The cell composition of claim 1 , wherein said construct is provided in a biocompatible device.

10. The cell composition of claim 9 , wherein the construct is produced via a scaffold-free platform.

11. The cell composition of claim 9 , wherein the total number of all cells in said construct is from 100 to 10,000.

12. The cell composition of claim 9 , characterized by:

(i) the production or expression of testosterone by said construct;

(ii) the expression of spermatogonial cell markers UCHL1, ITGA6, cKIT, DAZL, or a combination thereof;

(iii) the expression of Leydig and Sertoli cell markers HSD3B1, CYP11a1, FSHr, CYP19a1, cKIT, or a combination thereof;

(iv) the expression of meiotic and post-meiotic markers SYCP3, PRM1, ACROSIN, or a combination thereof; or

(v) a combination of (i)-(iv).

13. The cell composition of claim 9 , wherein said construct has an outer cell layer comprising filopodia.

14. The cell composition of claim 9 , wherein a central region of the construct comprises said spermatogonial stem cells.

15. A composition comprising:

(a) a hydrogel; and

(b) a plurality of cell compositions in the form of artificial three-dimensional testicular constructs according to claim 1 ,

wherein said plurality is provided in a biocompatible device,

wherein said cell compositions are provided in said hydrogel, and

wherein said hydrogel is a crosslinked hydrogel.

16. A biocompatible device, comprising

(a) a substrate having at least one chamber formed therein; and

(b) at least one artificial three-dimensional testicular construct provided in said chamber, said construct comprising a cell composition according to claim 1 ,

where said chamber has an inlet opening and outlet opening formed therein.

17. A method of screening a compound for pharmacological or toxicological activity, comprising:

(a) providing the device of claim 16 ,

(b) administering a compound to said construct; and

(c) detecting a pharmacological or toxicological response from at least one cell of said device.

18. The method of claim 17 , wherein said response comprises cell death; cell growth; absorption, distribution, metabolism, or excretion (ADME) of said compound; or upregulation or downregulation of production of a compound by said at least one cell.

19. The method of claim 18 , wherein said cell death comprises senescence or apoptosis.

20. The method of claim 18 , wherein said cell growth comprises benign or metastatic cell growth.

21. A biocompatible device, comprising

(a) a substrate having at least one chamber formed therein; and

(b) at least one artificial three-dimensional testicular construct provided in said chamber, each construct comprising a cell composition according to claim 1 ,

wherein said device is packaged in a container with a transient protective support medium in said chamber in gelled form.

22. A method of making a device, comprising:

(a) providing a substrate and;

(b) depositing at least one cell composition of claim 1 on said substrate.

23. The method of claim 22 , wherein said depositing step is carried out by bioprinting, pipetting, microinjection, or microfluidic deposition.