dependence of corneal stem cells on normal ocular surface innervation

Dependence of Corneal Stem Cellson Normal Ocular Surface Innervation

H. Ueno, G. Ferrari, T. Hattori,

T. Funaki, D.R. Saban, S.K. Chauhan,

R. DanaThe authors have no financial interest in the subject matter of this poster.

Schepens Eye Research Institute, Harvard Medical School

Background

Neurotrophic keratopathy (NK) is a degenerative disease of the cornea characterized by impaired healing.

Seitz B, Kruse FE.Ophthalmologe. 2005

Corneal sensory nerve degeneration is responsible for corneal epithelial defects, stromal thinning, and perforation.

Purpose

To identify the relationship between limbal stem cells and trigeminal innervation using a mouse model of corneal denervation.

Hypothesis: Neurotrophic keratopathy results in epithelial stem cell deficiency.

Approach: Analysis of stem cells in a mouse model of neurotrophic keratopathy. 　

Methods

１ . Real Time-PCR

2. Immunohistochemistry

7days

3. Flow cytometry

Trigeminal Stereotactic Electrolysis (TSE)

no blink reflex

Ophthalmic nerve

G. Ferrari et.al.preparation

ABCG2, p63, Hes1

ABCG2, p63

Side population

Cornea

Contralateral eyeb

-tub

ulin

III

Denervated eye

Expression of beta-III tubulin (neuronal marker) is decreased in the denervated cornea.

Normal Denervated

ABCG2

p63

2DAPI ABCG2 DAPI

DAPI DAPIp63

Expression of stem cell markers (ABCG2 & p63) is decreased in the denervated cornea.

ABCG2 p63 Hes1-1.66533453693773E-16

0.2

0.4

0.6

0.8

1

1.2

Rel

ativ

e E

xpre

ssio

n of

mR

NA Normal Expression

of mRNA:1.0

p 0.01≦p 0.01≦

RNA expression levels for both stem cell markers ABCG2 and Hes1 are significantly decreased in denervated corneas.

Summary

Corneal stem cells are significantly reduced in

this murine model of neurotrophic keratopathy.

Expression of stem cell markers (ABCG2, P63, Hes1) in

denervated corneas is decreased at both the mRNA and

protein levels.

Conclusion

Our novel data clearly demonstrate that

corneal nerves play a critical role in maintaining

the corneal stem cells.

References

1. Touhami A, Grueterich M, Tseng SC. The role of NGF

signaling in human limbal epithelium expanded by amniotic

membrane Invest Ophthalmol Vis Sci. 2002 ;43:987-94.

2. Qi H, Chuang EY, Yoon KC, de Paiva CS, Shine HD, Jones DB,

Pflugfelder SC, Li DQ. Patterned expression of neurotrophic

factors and receptors in human limbal and corneal regions. Mol

Vis. 2007 ;13:1934-41.

3. Qi H, Li DQ, Shine HD, Chen Z, Yoon KC, Jones DB, Pflugfelder

SC. Nerve growth factor and its receptor TrkA serve as potential

markers for human corneal epithelial progenitor cells. Exp Eye

Res. 2008 ;86:34-40.