Using Stem Cells to
Treat Ocular Surface
Diseases and
Disorders
C.S. Woodson, E. Meyer-Blazejewska, F.E. Kruse, M.K. Call, W.K.
Edith J. Crawley Vision Research Center, University of Cincinnati
Background
Limbal epithelial stem cells are located in the limbus, a thin band
along the edge of the iris. These cells are responsible for
replenishing the cells of the transparent, avascular cornea and
separating them from the opaque,
conjunctival tissue of the eye that
contains blood vessels. Corneal injury
or genetic defects can prevent proper
cell barrier and replenishment function
or destroy them all together, a disorder
called Limbal Stem Cell Deficiency
(LSCD). As a result, the conjunctiva
grows uncontrollably over the clear
cornea and causes blindness.
Currently, patients with unilateral LSCD are able to transplant
healthy limbal stem cells from the healthy eye to the diseased one
to treat LSCD. However, patients with bilateral LSCD cannot do this
process and if they accept a transplant from a donor, rejection of
the donated cells is possible.
Purpose
To explore the therapeutic potential of bulge-derived hair follicle
stem cells to treat LSCD.
Hypothesis
In this study, we addressed the hypothesis that when transplanted,
Hair Follicle Stem Cells (HFSCs) will develop into the corneal
epithelial cells needed to treat LSCD.
Method
Tri-transgenic mice (K12rtTA /tetO-Cre/RosamTmG) were utilized to
harvest HFSCs. Wild-type mice were then given LSCD by the
complete removal of all epithelium with a corneal rust-ring remover.
The isolated HFSCs were transplanted on a fibrin carrier onto wildtype LSCD mice. Factors in the corneal environment signaled HFSCs
to differentiate into corneal cells, exhibited by expression of K12 (in
green).
I. Hair Follicle Stem Cell (HFSC) Isolation
Murine HFSCs were isolated, in order to access the bulge region
harboring the epithelial stem cells, which express keratin 15 (Krt15) in
green.
Results
I. HFSC transplant
The HFCS from K12rtTA /tetO-Cre/RosamTmG mouse were placed on a
wild-type C57/Black6 mouse with induced LSCD (A) and sutured on
(B). The eye was then examined post-transplantation under red light
(C) and green light (D).
II. Keratin12 (K12) production
post-transplant
Keratin 12, considered a molecular
marker of corneal epithelial cells,
is detected with EGFP within 3
days (A) and 2 weeks (B) posttransplantation.
III. HFSC in action
HFSCs replaced nonfunctional limbal stem cells and began barrier
function and replenishment of avascular cornea once more.
A-H are 4 weeks after epithelial tissue was removed (induced LSCD). A-D are images from an LSCD mouse that received
a transplants. Images E-H are a control LSCD mouse.
Conclusion
HFSCs can differentiate into corneal epithelial cells.
HFSCs have therapeutic potential for LSCD patients with an 80%
success rate.
Next Steps
Now, we have begun examining environmental factors that cue
maintenance and differentiation of limbal stem cells. PPARɣ, a
transcription factor, is thought to be one of these factors. This is
evident in the human limbus and cornea where PPARɣ density is
high in limbus and decreases as you move towards central cornea
(A). In mice, PPARɣ is expressed in limbus (B) and only weakly in the
central cornea.
(A)
(B)
Human limbus
Human corneal epithelium
Mouse limbus
Mouse corneal epithelium