Gross Anatomy of the Eye...6 Graw (2010) in Seminars in Developmental Biology 90: 343.11 Primary Eye Field in Anterior Neural Plate Eyes develop from cells in anterior neural plate

1

PHOP Core Fall 2018

Intro to Genetics and Ocular Development

D. C. Otteson PhD UHCO 1

Gross Anatomy of the Eye

Graw, Eye Development. (2010) in

Current Topics in Developmental Biology (Volume 90) p. 343. 2

2

From the National Eye Institute

http://www.nei.nih.gov/photo/ 3

Body Axis Terminology

Caudal

(posterior)

Ventral

Dorsal

Medial

Lateral

Rostral

(anterior)

4

3

Eye-specific Axis Terminology

Dorsal / Superior

Ventral / Inferior

Posterior Anterior

Nasal

Temporal

5

http://www.becomehealthynow.com/article/bodyembryo/789/1

6

4

GastrulationInduction and cell migration forms three germ layers

Ectoderm

Mesoderm

Endoderm

Gastrulation initiates at posterior end

Figure from Developmental Biology by Scott Gilbert. Sinauer

Associates, Inc. 7

(From Martin, Neuroanatomy

Text and Atlas, Elsevier press)

Neural Crest

8

5

Neural tube segmentation: brain vesicles

(From Martin, Neuroanatomy Text and

Atlas, Elsevier press)

Mouse: day 8 post fertilization day 10-11 post fertilization

Human: week 4 week 5

9

Growth of

cranial nerves

Dekaban, A.S. and Sadowsky, D, Ann. Neurology, 4:345-356, 1978

3-vesicle

stage5-vesicle stage

Telencephalon

grows

Cortical maturation

and expansion

NOT TO SCALE !!!!

EYE

10

6

Graw (2010)

in Seminars in Developmental Biology 90: 343.

11

Primary Eye Field in Anterior Neural Plate

Eyes develop from cells in

anterior neural plate

Eye field contains cells

competent to form eye

structures

Not all cells will actually

contribute to eye structures

Early genes important in

specification of eye fields:

Pax6, Rax/Rx, Six3,

Tbx3, Six6, Otx2, Lhx2,

TllSpecies: Mouse E7 ~Human E17

http://www.med.unc.edu/embryo_images/12

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=dbio&part=A2960#A3077


7

Neurulation: Folding of Neural Plate

Neural plate grows rapidly

Eye fields in rostral neural

plate expand

Eye fields separate at

midline

Secreted sonic hedgehog

(SHH) from notocord

specifies ventral midline

http://www.med.unc.edu/embryo_images/

Species: Mouse E8 ~ Human E21

View: Frontal13

PAX2 specifies ventral optic cup and optic stalk

Mutations: loss of ventral diencephalon, optic chiasm,

ventral optic nerve coloboma, kidney and ear defects

Homozygous: lethal: no chiasm, no uritogenital tract

SHH secreted from notocord early embryo

induces ventral and midline structures in neural plate

required for separation of eye fields

Mutations: holoprosencephaly, cyclopia, midline facial

defects, microcephaly

PAX6 specifies dorsal and lateral structuresneural retina, RPE, lens, cornea (+ non-ocular structures)

Mutations: heterozygous: aniridia, glaucoma

homozygous: (lethal) anophthalmia, failure to form nasal

passage, brain defects

Separation of Eye Fields

SHH represses PAX6, induces PAX2

14

http://www.biolcell.org/boc/097/0321/boc0970321f03.htm


8

Cells in Eye Fields Invaginate Forming Optic

Grooves (a.k.a. optic sulci, optic pits)

Mouse E8.5 ~ Human E24

Fronto-LateralMouse E8 ~ Human E22

View: Frontal

http://www.med.unc.edu/embryo_images/ 15

Neural tube, rat development

showing optic vesicles

Zhang, Fu, Barnstable (2002)

Molec. Neurobiol. 26:137-152.

Rat embryo, E11.

Rostral view of neural tube

Head ectoderm removed

Arrows show

anterior neuropore

ov=optic vesicles

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9

Species: Mouse E8.5 to 9 (~Human E25)


Optic vesicle/Lens induction

17

Mouse E 9 (~Human E28) Mouse E10 (~ Human E29)


18







10

Species: Mouse E11 (~Human E36)

View: Coronal

Lens placode invaginates to form

lens vesicle

Invagination of optic vesicle forms

bilayered optic cup

Lens vesicle pinches off surface

ectoderm

Overlying ectoderm becomes

cornea


Lens Vesicle and Optic Cup

19

Species: Mouse E 11

(~Human E36)

View: Coronal Cut


Retina/RPE

Pax6 (+Mitf in RPE)

Lens vesicle

Pax6, Sox2/3

Optic Stalk

Pax2

Irido-pupillary

Membrane

From neural crest

20



11

Lens DevelopmentPrimary

Fiber elongation

Secondary

Fiber elongation

Graw (2010)

in Seminars in Developmental Biology 90: 343.

21

A model of a genetic switch composed of SOX2/SOX3 and Pax6, which regulates initiation of lens development.

Kamachi Y et al. Genes Dev. 2001;15:1272-1286

©2001 by Cold Spring Harbor Laboratory Press 22

12

Lens maturation

Proliferation of cells at the equator

Elongation of cells at bow

Fiber cells

nuclei are gone in central lens

cells remain connected by gap junctions

Species: Human 8 Weeks


Lens Fiber Cells

Song et al J Clin Invest. 2009;119(7):1837–1848

Scanning electron micrographs of bovine lens fiber cells

24



13

induced by lens

• outer epithelial layer forms from surface

ectoderm

• inner layers primarily from neural crest

cells

Image of cornea from: T. Caceci (2001)

Anatomy and Physiology of the Eye V2.0


Cornea

25

RPE: Outer pigmented layer becomes relatively thinner

Single cell layer

RPE cells express PAX6 and MITF. Expression of MITF helps

specify RPE identity and this transcription factor directly regulates genes

that are responsible for pigment formation in RPE.

RETINA: Inner, neural portion thickens

Pseudostratefied epithelium

Differentiation begins at ~E11.5

Mouse Day E14 (~ Human 7 weeks) http://www.med.unc.edu/embryo_images/

RPE

Ventricle/

Sub-retinal

Space

Retina

26



14

The tunica vasculosa lentis

From human fetus

(a) Hyaloid artery

(b) Posterior ciliary artery

From Duanes Ophthalmology 2006

Chapter 15: Lens, Kleiman and Worgul

Vasculature

Mouse(Gerhardt et al., 2003)

Superficial Plexus

Deep Plexus

27

Iris and Ciliary Body Formation

Human ~15 Weeks


Human 8 Weeks

28

15

Ittner et al. Journal of Biology 2005 4:11

Neural Crest Derivatives in Eye

29

Neural Crest

Ittner et al. Journal of Biology 2005 4:11 30

16

Human: 8 Weeks

Eyelids

•begin to form at end of embryonic period

•fuse at the start of 2nd trimester

•reopen at the beginning of the 3rd trimester

Human: 10 Weeks


Loci for Inherited Retinal Disease

RetNet; Stephen P Daiger PhD

The University of Texas Health Science Center, Houston, Texas

http://www.sph.uth.tmc.edu/RetNet

32



17

genetics

DNA Replication and Inheritance

The basis of

perpetuation of life and

transmission of

traits/genes

8-34

Inherited vs. Non-inherited Mutations

• Germ-line mutations lead to inherited mutations– Occurs in germ line tissue

– If mutation passes into gametes (egg, sperm), it will be passed

on to next generation. • e.g. sickle cell anaemia

• Retinitis pigmentosa,

• Keratoconus

• Somatic mutations occur in somatic tissues – Population of identical cells derived from a single mutated

somatic cell is a clone

– Often results in a patch of phenotypically mutant cells

– Not inherited

– Many tumours result from somatic mutation

– Two-hit theory of cancer e.g. retinoblastoma

18

8-35

Heritable Genetic Changes

Single gene mutation

Change in allele of a gene

• Protein not expressed or

• Protein non-functional or

• Protein acquires novel/harmful/beneficial functions

• Contributes to evolution

Chromosome mutation: multiple genes

Changes in segments of chromosome

Deletion, duplication, inversion, translocation

Loss/duplication of whole chromosome

Epigenetic Changes

DNA modifications that can permanently change gene

expression

Genes + Environment= Phenotype

genes environmentphenotype

Contributions from genotype/environment varies with disease

Some diseases have both genetic and environmental components

e.g. macular degeneration; cataract; heart disease

36

19

8-37

Classifying Mutations : Phenotypic Aspects

• Morphological mutations– Visible/measurable change in phenotype– e.g. eye color; ocular coloboma

• Lethal mutations– Heterozygous: normal– Homozygotes: do not survive– e.g. Cystic Fibrosis= mutation in CFTR gene–

• Conditional mutations – Phenotype only presents under specific conditions – e.g. predisposition for developing disease: diabetes, heart

disease, cancer: can change life-style and reduce risk

• Biochemical mutations– Change in enzyme, biochemical pathway components– e.g. Albinism (inability to synthesize melanin) mutations in

Tyrosinase and other enzymes of melanin biosynthesis

8-38

Mutations: Mechanistic Aspects

• Silent mutations/Polymorphisms

• Change in DNA sequence:

• doesn’t change sequence of protein (synonymous)

– or

• occurs in non-critical region of gene

– Used for:

– DNA “fingerprinting”

– paternity analysis

– genetic mapping (linkage analysis)

20

Consequences of point

mutations

39

8-40

21

Gametes get one copy of each chromosome

Gene R is on a separate chromosome from A and B and segregates independently of A and B

All allelic combinations are found: AB +R AB + r ab + R ab + r

Therefore, R is not linked to the A or B genes

Genes A and B are on the same chromosome

Parental allelic combinations inherited together (AB or ab)

A and B genes are linked

Independent assortment of

chromosomes during meiosis

41

Predicting Patterns of Inheritance: Punnett Square

Genotype:

B/B B/b b/b

1 : 2 : 1

Phenotype:

Black brown

3 : 1

diploid

diploid

haploid

Shown: two alleles of a single gene

42

22

8-43

Mendelian Non-Mendelian

Autosomal Epigenetic

Sex-linked Mitochondrial

Recessive Imprinting

Dominant Multifactorial

Monogenic

Other including complex

Syndromic

Semi-dominant/co-dominant

Sporadic/spontaneous

Digenic

Patterns of Inheritance

• Autosomal inheritance

– Based on the variation of single genes on regular chromosomes

(autosomes)

• Sex-linked inheritance

– Based on the variation of single genes on sex-determining

chromosomes

• Cytoplasmic inheritance

– Based on the variation of single genes on organelle’s

chromosomes (e.g. Mitochondrial)

Patterns of Inheritance

44

23

Patterns of Inheritance: Recessive

Appearance of macula in Usher syndrome

Fundus pigmentation in Usher

syndrome

Usher Syndrome I (human) USHIB

mutations in MYO7A (Myosin7A)

• Heterozygous (-/+) normal

• Homozygous (-/-)

• profound congenital hearing

impairment

• unintelligible speech

• early retinitis pigmentosa (<10 yrs

• vestibular dysfunction

• defects in cilia (photoreceptor

connecting cilium; hair cells)


•How to detect this pattern in

patients?

•Can you distinguish this from

spontaneous mutation?

46

http://disorders.eyes.arizona.edu/sites/disorders.eyes.arizona.edu/files/disorder_images/Copy of slidescan_053.jpg




http://www.glowm.com/resources/glowm/graphics/figures/v5/0740/004f.gif

http://www.glowm.com/resources/glowm/graphics/figures/v5/0740/004f.gif

24

Parents:

heterozygous

(+/-)

normal

+

CSNB

-

normal

+

CSNB

-Genotypes: +/+ +/- -/-

Ratios (genotype)

Phenotypes

Ratios (phenotypes)


47

Parents:

heterozygous

(+/-)

normal

+

CSNB

-

normal

++/+ -/+

CSNB

-+/- -/-

Genotypes: +/+ +/- -/-

Ratios (genotype)

Phenotypes

Ratios (phenotypes)

1 2 1

Normal Normal mutant

13


48

25

Parents:

Heterozyous (+/-)

Homozygous (+/+)+ -

+

+


Ratios (genotype)

Phenotypes

Ratios (phenotypes)


49

Parents:

Heterozyous (+/-)

Homozygous (+/+)+ -

+ +/+ -/+

+ +/+ -/+


Ratios (genotype)

Phenotypes

Ratios (phenotypes)

1 1 0

Normal Normal

1 0


50

26

• Recessive genes are not major contributors in

human genetic disease unless:

• highly consanguineous group

• frequency of mutant allele in general population is

high

• Difficult to determine if this is inherited or

spontaneous if only one affected individual


51

Patterns of Inheritance: Dominant

• Dominant: Mutant allele is fully expressed and masks

the expression of the allele.

• With true dominant, individuals heterozygous and

homozygous for the mutated allele show the same

phenotype.

• Haplo-insufficient: Absence of one allele results in

intermediary phenotype; often referred to as semi-

dominant

• Co-dominant: both alleles are fully expressed

• e.g. blood groups (A, AB, B, O)

52

27

genetics53


Retinitis Pigmentosa

Phenotype:

• Constriction of the visual fields

• Night blindness

• Fundus changes

• “bone spicule” lumps of pigment

• Photoreceptor Degeneration

• Variable age of onset

Multiple patterns of inheritance

• Autosomal dominant

• Autosomal recessive

• X-linked

More than 3100 distinct mutations in

56 genes in have been identified(Daiger et al 2013 Clinical Genetics 84:132.)

From: www.stlukeseye.com/

genetics54

Male unaffected

Male affected

Female

unaffected

Female affected



28

genetics55

Genotypes: +/+ +/- -/-Ratios (genotype)

Phenotypes

Ratios (phenotypes)

Parents

Heterozyous (+/-)

Homozygous (+/+) + +

+

-



genetics56


Phenotypes

Ratios (phenotypes)

1 1 0

Normal Mutant Mutant

1 1 0

Parents

Heterozyous (+/-)

Homozygous (+/+) + +

+ +/+ +/+

- +/- +/-



29

genetics57

Patterns of Inheritance: Semi-Dominant Haploinsufficient

Aniridia

• Mutations in transcription factor Pax6

• Haploinsufficient

• Homozygous lethal

• Heterozygotes: anterior segment malformations:• aniridia

• corneal clouding with variable iridolenticulocorneal adhesions

• Peters anomaly (central corneal leukoma, absence of the posterior corneal stroma and Descemet membrane, and a variable degree of iris and lenticular attachments to the central aspect of the posterior cornea)

• foveal hypoplasia

• glaucoma

• autosomal dominant keratitis

A normal eye is pictured above.

Below is the eye of a child with

aniridia, a congenital eye disorder.

People born with the disease have

no iris and are generally legally

blind.

CREDIT ANIRIDIA FOUNDATION

genetics58


Phenotypes

Ratios (phenotypes)

Parents

Heterozyous (+/-)+ -

+

-

Patterns of Inheritance: Semi-Dominant / Haploinsufficient

30

genetics59


Phenotypes

Ratios (phenotypes)

1 2 1

Normal Mutant More severe

1 2 1

Parents

Heterozyous (+/-)+ -

+ +/+ +/-

- +/- -/-

Patterns of Inheritance: Semi-Dominant Haploinsufficient

genetics60

• Mutation on X chromosome

• Females: XX

• Males: XY

• Mutation/disease phenotype manifests in males

• Females are carriers, typically normal

– But may manifest mosaic defects

• Generation skipping pattern

Patterns of Inheritance: X-linked

31

genetics61

Examples of X-linked Retinal Diseases

OA1 X-linked ocular albinism

RP23,RP6, X-linked Retinitis Pimentosa

RS1 Retinoschisis

OPA2 X-linked optic atrophy

NDP Norrie disease; familial exudative

vitreoretinopathy

Colorblindness (mutations in red/green opsin genes):

OPN1SW protan (red deficient)

OPN1LW deutan (green deficient)

blue cone monochromacy (red & green deficient)

X-linked Diseases: X-inactivation

X chromosomes:

Males have 1

Females have 2

How to maintain same level of

transcription (RNA levels) from

X chromosome genes?

Random X-inactivation

(Lyonization) in females

after Mary Lyon

e.g. coat color in cats62

32

X-inactivation of B-gal

transgene in retina

http://mentor.lscf.ucsb.edu/course/winter/mcdb101b/x-inactivation/xinactivation.html

X-inactivation in the

cornea

John West

http://www.cip.ed.ac.uk/gallery/index.htm

http://mentor.lscf.ucsb.edu/course/winter/mcdb101b/x-inactivation/Figure 1.jpeg

http://mentor.lscf.ucsb.edu/course/winter/mcdb101b/x-inactivation/Figure 1.jpeg

http://mentor.lscf.ucsb.edu/course/winter/mcdb101b/x-inactivation/Figure2.jpeg

http://mentor.lscf.ucsb.edu/course/winter/mcdb101b/x-inactivation/Figure2.jpeg

33

Mutant phenotype present in males x* Y

Carrier females may show mosaic defects Xx*

Skips generations

carrier

XY Xx*

XX x*Y XY Xx* XY x*Y XX

XY Xx* Xx* Xx* Xx* XY x*Y XX XX x*Y Xx* Xx* XY


65

genetics66

Genotypes: X*X XX X*Y XY

Ratios (genotype)

Phenotypes

Ratios (phenotypes)Normal

*carrier

Normal Affected Normal

Parents

Father affected X* Y

X

X


34

genetics67


Ratios (genotype)

Phenotypes

Ratios (phenotypes)

0 1 0 1

Normal

*carrier


1 0 0 1

Parents

Father affected X* Y

X X*X XY

X X*X XY


genetics68


Ratios (genotype)

Phenotypes

Ratios (phenotypes)Normal

*carrier


Parents

Mother-carrier X Y

X*

X


35

genetics69


Ratios (genotype)

Phenotypes

Ratios (phenotypes)

1 1 1 1

Normal

*carrier


1 1 1 1

Parents

Mother-carrier X Y

X* X*X X*Y

X XX XY


Mitochondrial diseases affecting the eye

8-70

Ophthalmic manifestations of mitochondrial diseases:

cataract, retinopathy, optic atrophy, cortical visual loss, ptosis and

ophthalmoplegia

• Leber’s Hereditary Optic Neuropathy (LHON)

• Kearns-Sayre Syndrome (KSS)

• Mitochondrial Encephalopathy, Lactic Acidosis Stroke (MELAS)

• Myoclonic Epilepsy and Ragged Red Fiber myopathy

(MERRF)

36

genetics71

Patterns of Inheritance: Mitochondrial

Leber Optic Atrophy

• Presents in mid-life as acute or sub

acute central vision loss leading to central

scotoma and blindness

• Associated with many missense

mutations in the mtDNA

• Mutations can act autonomously or in

association with other mt mutations

• Final visual acuity can range from 20/50

to no light perception, depending on

severity of the mutations

Leber Optic Neuropathy with

temporal optic nerve pallor in both

eyes.

http://www.revoptom.com/continuing_education/ta

bviewtest/lessonid/108143/

Healthy optic disc

http://www.intechopen.com/books/the-mystery-of-glaucoma/

8-72

http://www.usc.edu/dept/mda/180evolution/IMAGES/wmho.html

http://www.intechopen.com/source/html/18955/media/image1.png




37

8-73

Patterns of Inheritance: Mitochondrial

Mitochondrial mutations:

•inherited ONLY from the mother

•NEVER from the father

ROM1/ROM1; ROM1/ROM1; +/ROM1 ; +/ROM1;

RDS/RDS +/RDS RDS/RDS +/RDS

ROM1/ROM1; ROM1/ROM1; +/ROM1; +/ROM1;

RDS/+ +/+ RDS+ +/+

+/ROM1; ROM1/+; +/+; +/+;

RDS/RDS +/RDS RDS/RDS +/RDS

ROM1/+; ROM1/+; +/+; +/+;

+/RDS +/+ RDS/+ +/+

Mode of inheritance: multigenic

ROM1/+; RDS/+ x ROM1/+; RDS/+

ROM1; RDS ROM1; + +; RDS +; +

ROM1; RDS

ROM1; +

+; RDS

+;+

Genotypes 1:2:1:2:4:2:1:2:1

Phenotypes: 1:3:2:1:9

Documents

Gross Anatomy of the Eye...6 Graw (2010) in Seminars in Developmental Biology 90: 343.11 Primary Eye Field in Anterior Neural Plate Eyes develop from cells in anterior neural plate