Method for Producing Retinal Tissue

1 . A method for producing a retinal cell or retinal tissue, comprising:

(A) suspension-culturing a pluripotent stem cell to form a cell aggregate of pluripotent stem cells; and

(B) suspension-culturing the cell aggregate obtained in the step (A) in the presence of a BMP signaling pathway agonist and a CHK1 signaling pathway inhibitor to obtain a cell aggregate containing a retinal cell.

2 . The production method according to claim 1 , wherein, in the step (B), the BMP signaling pathway agonist is present in such a concentration that an almost truly spherical cell aggregate is formed and differentiation into a retinal pigment epithelial cell is suppressed.

3 . The production method according to claim 1 , wherein the cell aggregate obtained in the step (B) is a sphere-like cell aggregate comprising a multilayer structure of an outer structure including a retinal pigment epithelial cell and an inner structure including a neural retina, and, in the step (B), the BMP signaling pathway agonist is present in such a concentration that the sphere-like cell aggregate comprising a multilayer structure is formed.

4 . The production method according to claim 1 , wherein the CHK1 signaling pathway inhibitor is PD407824.

5 . The method according to claim 1 , wherein the BMP signaling pathway agonist is one or more proteins selected from the group consisting of BMP2, BMP4, BMP7 and GDF7.

6 . The production method according to claim 1 , wherein, in the step (B), the BMP signaling pathway agonist is added to a culture medium between Day 2 and Day 9 after start of suspension culture in the step (A).

7 . The production method according to claim 1 , wherein, in the step (B), the CHK1 signaling pathway inhibitor is added to a culture medium simultaneously with the BMP signaling pathway agonist.

8 . The production method according to claim 1 , wherein a concentration of the CHK1 signaling pathway inhibitor is such a concentration that a CHK1 signaling pathway inhibitory effect comparable to a CHK1 signaling pathway inhibitory effect of 0.1 UM to 10 UM PD407824 is exerted.

9 . The production method according to claim 1 , further comprising dissecting retinal tissue of size required for transplantation from the cell aggregate obtained in the step (B).

10 . A sphere-like cell aggregate comprising:

a multilayer structure of an outer structure including a retinal pigment epithelial cell and an inner structure including a neural retina, wherein

(1) a neural retinal layer including at least a photoreceptor layer is formed in the neural retina in the inner structure, and the photoreceptor layer includes one or more cells selected from the group consisting of a photoreceptor cell, a photoreceptor progenitor cell and a retinal progenitor cell,

(2) the neural retina is present in a folded state in the inner structure,

(3) the retinal pigment epithelial cell in the outer structure is an RPE65-positive cell, an MITF-positive cell, or an RPE65-positive and MITF-positive cell, and

(4) the cell aggregate is free of a lens, a vitreous body, a cornea and a blood vessel.

11 . The sphere-like cell aggregate according to claim 10 , wherein, in at least a part of the sphere-like cell aggregate, a basal surface of the retinal pigment epithelial cell is facing the inner structure, and a basal surface of the neural retina is facing the outer structure.

12 . The sphere-like cell aggregate according to claim 10 , wherein the retinal pigment epithelial cell and the neural retina are further connecting to each other as an epithelium structure, and the sphere-like cell aggregate further includes a ciliary marginal zone-like structure between the retinal pigment epithelial cell and the neural retina.

13 . The sphere-like cell aggregate according to claim 12 , wherein the ciliary marginal zone-like structure includes an Rdh10-positive cell, an Otx1-positive cell, and/or a Zic1-positive cell.

14 . The sphere-like cell aggregate according to claim 10 , wherein 30% or more of the inner structure constitutes a neural retina.

15 . The sphere-like cell aggregate according to claim 10 , having a diameter of 0.2 mm to 2 mm.

16 . A method for producing the sphere-like cell aggregate according to claim 10 , comprising:

(A) suspension-culturing a pluripotent stem cell to form a cell aggregate of pluripotent stem cells; and

(B) suspension-culturing the cell aggregate obtained in the step (A) in the presence of a BMP signaling pathway agonist and a CHK1 signaling pathway inhibitor to obtain a sphere-like cell aggregate comprising a multilayer structure of an outer structure including a retinal pigment epithelial cell and an inner structure including a neural retina.

17 . A pharmaceutical composition comprising:

the sphere-like cell aggregate according to claim 10 or a part thereof.

18 . A method for treating a disease caused by damage of a retinal cell or retinal tissue or injury of retinal tissue, comprising:

transplanting the sphere-like cell aggregate according to claim 10 or a part thereof to a subject in need of transplantation.