The eye is an organ formed by the protrusion of a part of the brain during development. Two protrusions arise from the front of the primitive brain vesicle, the precursor of the brain, forming the primary optic vesicles. The neural retina and the brain both develop from the neuroectoderm, truly making the eye a part of the brain.
Eye development begins with gastrulation. The blastula transforms into a gastrula, forming the three germ layers: endoderm, mesoderm, and ectoderm. By the third week of development, the three germ layers form a trilaminar embryonic disc.
Immediately after gastrulation, neurulation occurs. The neural plate folds inward to form the neural tube, and around day 22 of gestation, the optic groove appears on the neural fold. By day 25, the optic groove develops into the optic vesicle.
When the distal end of the optic vesicle approaches the surface ectoderm, the surface ectoderm thickens to form the lens placode. The anterior wall of the optic vesicle invaginates toward the posterior wall, forming a double-walled cup-shaped structure, the optic cup.
The tubular part connecting the optic cup and the brain ventricle is called the optic stalk. The optic stalk eventually becomes the optic nerve.
The eye and its adnexa are composed of the following four tissue lineages:
Surface ectoderm
Corneal epithelium: Differentiates after lens vesicle separation
Lens: Formed from invagination of surface ectoderm
Eyelid epithelium and conjunctival epithelium: Derived from surface ectoderm
Lacrimal gland and Meibomian gland: Develop from conjunctival epithelium
Neuroectoderm
Retina and retinal pigment epithelium: Differentiate from the inner and outer layers of the optic cup
Iris epithelium and ciliary epithelium: Derived from the anterior rim of the optic cup
Optic nerve: Formed by axons of retinal ganglion cells
Vitreous body: Largest contribution by volume
Mesoderm
Extraocular muscles: Formed from preotic myotomes
Orbital fat and connective tissue: Derived from mesoderm
Choroidal vascular network: Induced from mesoderm surrounding the optic cup
Periciliary muscle tissue: Contribution from mesoderm
Neural crest cells (fourth germ layer)
Corneal stroma and endothelium: Formed by migration of neural crest cells
Iris stroma: Melanocyte concentration determines iris color
Sclera and trabecular meshwork: Derived from neural crest
Orbital bones: Mainly ossify from neural crest lineage
The neural crest is a transient structure formed between the surface ectoderm and neural plate during neural tube formation in vertebrates. Due to its importance, it is called the “fourth germ layer.” Neural crest cells undergo delamination and epithelial-mesenchymal transition, migrating to various sites in the embryo, and play a crucial role in eye development.
Eye development begins around the 3rd week of gestation (approximately day 22 of pregnancy) with the appearance of the optic sulcus on the neural plate. By day 25, it progresses to the optic vesicle, followed by the formation of the optic cup and differentiation into various tissues. The basic structure of the eyeball is formed during the fetal period, and the completion of the macula continues until 16 weeks after birth.
Normal eye development is precisely regulated by multiple genes and molecular signals. Abnormalities in these processes can lead to congenital eye diseases.
| Gene | Function and Associated Diseases |
|---|---|
| PAX6 | Master gene for eye formation. Mutations cause aniridia, coloboma, microphthalmia, and Peters anomaly |
| SHH | Divides the single eye field into two eyes. Mutations cause cyclopia |
| PAX2 | Essential for optic stalk formation and closure of the embryonic fissure |
The PAX6 gene is a master control gene essential for eye formation, discovered through developmental studies in fruit flies. In humans, it is identified as the causative gene for aniridia. The PAX6 gene is adjacent to the tumor suppressor gene WT1 on chromosome 11p13, and deletion of both results in WAGR syndrome (Wilms tumor, aniridia, genitourinary abnormalities, and intellectual disability).
Retinoic acid (RA) is an essential signaling molecule for eye development 1). Retinol (vitamin A) is converted to retinal by RDH10, and then to RA by ALDH1A1, ALDH1A2, and ALDH1A3 1).
In humans, mutations in four RA signaling pathway genes—RBP4, STRA6, ALDH1A3, RARB—and in PITX2 and FOXC1, which are regulated by RA, are associated with anophthalmia and microphthalmia 1).
PITX2 mutations cause Axenfeld-Rieger syndrome, and FOXC1 mutations cause anterior segment dysgenesis 1).
Coloboma (uveal coloboma) is a congenital anomaly resulting from incomplete closure of the embryonic fissure. The embryonic fissure begins to close from the central portion around the 6th week of gestation and is completed by the 7th week. If closure is impeded, a fissure extending downward from the pupil remains, leading to iris coloboma, choroidal coloboma, giant coloboma, etc. It is often accompanied by microphthalmia.
The PAX6 gene is a master control gene for eye formation. Mutations cause aniridia, coloboma, microphthalmia, Peters anomaly, macular hypoplasia, etc. Additionally, simultaneous deletion with the adjacent WT1 gene leads to WAGR syndrome (Wilms tumor, aniridia, genitourinary abnormalities, and intellectual disability).
This section details the developmental processes of each ocular tissue in chronological order.
At the beginning of the third week of gestation, the optic sulcus appears in the central part of the neural plate. This marks the beginning of visual organ development. At the end of the third week, both sides of the optic pit expand into vesicles, forming the optic vesicle.
At the fourth week of gestation, the distal anterior wall of the optic vesicle approaches the surface ectoderm, forming the lens placode. Subsequently, the anterior wall of the optic vesicle invaginates to become the optic cup, and the lens placode thickens and invaginates to form the lens vesicle, which separates into the optic cup by the fifth week.
A fissure (the optic cup fissure) appears in the lower part of the optic cup, and a fissure (the optic stalk fissure) also appears in the lower wall of the optic stalk. Together, these are called the embryonic fissure. The hyaloid artery, branching from the dorsal ophthalmic artery, enters the optic cup through the embryonic fissure. Closure begins around the central part at the sixth week and is completed by the seventh week.
Initially, both the inner and outer layers of the optic cup are pseudostratified columnar epithelium, but they later follow different fates.
The inner layer thickens due to active cell division and differentiates into the sensory retina (neural retina). However, near the pupillary margin, it does not thicken and remains as simple cuboidal epithelium, forming the epithelial parts of the ciliary body and iris.
The outer layer becomes thinner as the optic cup expands, and melanin granules appear in the late fifth week, differentiating into the retinal pigment epithelium (RPE). Notably, the retinal pigment epithelium is the only pigment tissue in the body that does not originate from neural crest cells.
The area where the inner layer folds over the outer layer forms a round opening toward the front, which will become the pupil.
When the lens vesicle separates from the surface ectoderm and is enveloped by the anterior part of the optic cup, the basement membrane of the simple epithelial cells becomes the lens capsule. The cells of the anterior wall remain as a single layer of lens epithelium, while the cells of the posterior wall extend forward as primary lens fibers.
At 6–7 weeks of gestation, the lumen of the lens vesicle disappears, forming the embryonic nucleus. Cells at the equator divide and proliferate to form the fetal nucleus, and further outward, secondary lens fibers are added successively. Secondary lens fibers continue to develop throughout life.
The lens originates from ectodermal epithelium, and mesenchymal tissue does not participate in its formation. During the fetal period, nutrition is supplied by the tunica vasculosa lentis (derived from the hyaloid artery).
From the inner layer of the optic cup, the neural retina, iris epithelium, and ciliary non-pigmented epithelium are formed; from the outer layer, the retinal pigment epithelium, ciliary pigmented epithelium, and iris pupillary muscles are formed.
Differentiation of the neural retina proceeds in two stages.
Stage 1 (vertical gradient differentiation): The neuroblastic layer differentiates into inner and outer neuroblastic layers. From the inner neuroblastic layer, ganglion cells differentiate first, followed by Müller cells, bipolar cells, amacrine cells, and horizontal cells. From the outer neuroblastic layer, photoreceptors differentiate. Cones appear at 3 months of gestation, and rods at 4 months.
Stage 2 (horizontal gradient differentiation): Differentiation progresses from the posterior pole to the periphery. Except for the macula, retinal development is nearly complete by 9 months of gestation. Macular differentiation begins at 6 months of gestation, foveal formation starts at 7 months, and histogenesis continues until 16 weeks after birth.
The vitreous body forms through three stages.
| Stage | Timing | Characteristics |
|---|---|---|
| Primary vitreous | From 6 weeks of gestation | Contains the hyaloid artery. After regression, Cloquet’s canal remains. |
| Secondary vitreous | From 9 weeks of gestation | Acellular network. Constitutes most of the mature vitreous. |
| Tertiary vitreous | Late gestation | Forms the ciliary zonule (Zinn’s zonule). |
In the late fetal period, when the hyaloid artery degenerates and disappears, the primary vitreous also disappears. The branches running along the surface of the inner layer of the optic cup remain as the central retinal artery and vein.
At 6 weeks of gestation, retinal ganglion cells appear. Their axons pass through the innermost layer of the retina, penetrate the inner layer of the optic cup at the optic disc, and extend into the optic stalk. By 7 weeks of gestation, they reach the optic chiasm, and then extend to the occipital lobe via the lateral geniculate body.
At 3 months of gestation, the pia mater is formed from neural crest cells around the optic stalk. The dura mater appears by 5 months of gestation, and the arachnoid mater differentiates at 6 months. Myelination begins at the lateral geniculate body at 5 months of gestation and progresses toward the retina.
At 4 weeks of gestation, after the separation of the lens vesicle, the surface ectoderm differentiates into the corneal epithelium. At 6 weeks, neural crest cells migrate between the corneal epithelium and the lens to form Bowman’s layer and the corneal endothelium. Then, neural crest cells migrate again to form the corneal stroma.
At 7 weeks of gestation, neural crest cells migrate between the corneal endothelium and the lens to form the pupillary membrane and iris stroma. At 3–4 months of gestation, Schlemm’s canal forms, the anterior chamber appears, and the trabecular meshwork is also formed from neural crest cells.
Iris: At 3 months of gestation, the anterior and posterior layers of the iris epithelium are formed from the anterior margin of the optic cup. The pupillary sphincter muscle begins to differentiate at 4 months and completes by 8 months. The pupillary dilator muscle begins differentiation at 6 months and completes after birth. The intrinsic muscles of the iris are derived from the neuroectoderm.
Ciliary body: At 3 months of gestation, folds appear in the inner and outer layers of the optic cup, forming the ciliary processes. The ciliary stroma and ciliary muscle are formed from neural crest cells.
Choroid: At 5 weeks of gestation, melanin granules appear in the retinal pigment epithelium, and a capillary network is induced from the mesodermal tissue around the optic cup. At 4 months of gestation, the choroidal vascular network is formed.
Sclera: Formation begins at 7 weeks of gestation from neural crest cells at the anterior margin of the optic cup, extends posteriorly, and reaches the posterior pole by 5 months of gestation.
Eyelids: At 6 weeks of gestation, two folds form above and below the eye. They fuse temporarily at 3 months, begin to separate at 6 months, and open by 7 months. The conjunctival epithelium, eyelashes, and various glands (Moll glands, Zeis glands, Meibomian glands) develop from the surface ectoderm, while the orbicularis oculi muscle and tarsal plate develop from the mesoderm.
Lacrimal gland: At 10 weeks of gestation, basal cells of the conjunctival epithelium in the temporal superior fornix invaginate into the mesodermal tissue to form the gland. Reflex tearing may not begin until 1–3 weeks after birth.
Extraocular muscles: At 4 weeks of gestation, mesenchymal tissue around the optic cup condenses to form the primordium. At 8 weeks, the four rectus muscles and two oblique muscles differentiate, and the levator palpebrae superioris separates from the superior rectus muscle.
Orbit: The orbital bones are mainly derived from the neural crest, and membranous ossification begins at 6 weeks of gestation. The sphenoid and ethmoid bones develop through endochondral ossification.
Retinoic acid (RA) controls two critical stages in eye development1).
Stage 1: Optic cup formation (equivalent to mouse E8.5–E10.5) RA is essential for optic cup formation via invagination (folding) of the optic vesicle1). In particular, ventral optic vesicle invagination is impaired in RA deficiency1). Aldh1a2 produces RA in the periocular mesenchyme at E8.5–E9.5, and loss of RA synthesis at this stage leads to optic cup malformation1).
Stage 2: Anterior segment morphogenesis (mouse E10.5 onward) RA is produced in the dorsal (Aldh1a1) and ventral (Aldh1a3) retina and diffuses into the neural crest-derived periocular mesenchyme outside the optic cup1). Loss of RA causes excessive proliferation of the mesenchyme, leading to microphthalmia, corneal malformation, and eyelid malformation1).
RA activates Pitx2 in the periocular mesenchyme, and Pitx2 induces Dkk2 (a WNT antagonist), thereby suppressing WNT signaling and limiting excessive mesenchymal proliferation1).
Retinoic acid (RA) is an active metabolite of vitamin A and controls two stages in eye development: optic cup formation and anterior segment morphogenesis. RA is produced in the retina, diffuses into the surrounding neural crest-derived mesenchyme, and suppresses WNT signaling via the Pitx2-Dkk2 pathway. Genetic mutations in the RA signaling pathway cause congenital eye diseases such as anophthalmia and microphthalmia.
RA signaling functions through nuclear RA receptors (RARs) binding to RA response elements (RAREs) to regulate transcription1). However, direct target genes of RA in eye development have not yet been identified1). Because thousands of RAREs exist in the mouse and human genomes and thousands of genes change expression upon RA loss, identifying direct targets is not straightforward1).
Recent studies have developed methods to narrow down direct target genes by detecting RA-dependent deposition of H3K27ac (gene activation mark) and H3K27me3 (gene repression mark) using ChIP-seq and integrating with RNA-seq data1). Applying this approach, demonstrated in trunk tissues, to eye development is expected to comprehensively identify RA target genes1).
RDH10 is the only enzyme responsible for the first step of RA synthesis (retinol to retinal conversion), and Rdh10 knockout mice survive until E10.5, show no RA activity in the optic field, and exhibit optic cup malformation 1). It is easier to manipulate experimentally than Aldh1a1/Aldh1a2/Aldh1a3 triple knockouts, making it a useful model for elucidating the mechanism of optic cup formation in the future 1).
