IC-55: Limbal stem cell deficiency and its management
Limbal stem cell deficiency is a painful and blinding disease characterised by
recurrent epithelial defects, corneal vascularisation and corneal conjunctivalisation.
There are many causes including hereditary (such as aniridia) and acquired causes
(such as chemical eye burns and inflammatory eye diseases). There are medical and
surgical measures for this debilitating disease, including cultured limbal stem cell
therapy. This course will describe the conventional management, including
conservative management and whole tissue limbal grafting, as well as more
contemporary management such as cultured limbal stem cell transplantation. The
different methods and regulatory issues involved in limbal stem cell culture will also
be highlighted.
Objectives:
1. Limbal stem cell deficiency and its causes
2. Diagnosis of limbal stem cell deficiency
3. Medical management of limbal stem cell deficiency
4. Surgical management of limbal stem cell deficiency: whole tissue limbal
grafting and cultured limbal stem cell transplantation
5. Different methods for culturing limbal stem cells
6. Regulatory requirements for cultured limbal stem cell transplantation
Schedule:
0-20
minutes
20-30 minutes
30-50 minutes
50-60 minutes
60-75 minutes
75-90 minutes
90-105 minutes
105-120 minutes
Limbal stem cell deficiency, its causes and diagnostic tools
Questions
Management options for limbal stem cell deficiency
Questions
Culture methods for limbal stem cells
Transplantation of cultured limbal stem cells
The regulation of cultured limbal stem cell transplantation
Questions
1) Limbal stem cell deficiency, its causes and diagnostic tools
Limbal stem cell deficiency
The limbus has two important functions with regards to the corneal epithelium.
Firstly, the corneal epithelium is renewed by stem cells located at the limbus, the socalled limbal stem cells. Secondly, the limbus also acts as a barrier preventing the
conjunctival epithelium and its blood vessels from encroaching on to the corneal
surface. When the limbal stem cells become deficient or dysfunctional, the disease of
limbal stem cell deficiency results. The corneal epithelium cannot be maintained and
conjunctivalisation ensues. The result is a disease which causes chronic ocular surface
inflammation and epithelial breakdown, and visual impairment.
Causes
Limbal stem cells become deficient or dysfunctional from a variety of causes. These
can be broken down into primary or hereditary causes and secondary or acquired
causes. Primary causes include aniridia and ectodermal dysplasia. Secondary causes
include chemical and thermal burns, inflammatory ocular surface disease (Stevens-
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Johnson syndrome and ocular cicatricial pemphigoid), iatrogenic causes (extensive
limbal surgery and cryotherapy, and radiotherapy) and contact lens related keratitis.
Diagnosis
The diagnosis of limbal stem cell deficiency is very much made on natural history of
the disease and clinical signs of conjunctivalisation of the corneal surface. Delayed
fluorescein staining of the corneal surface is a characteristic feature. Corneal surface
impression cytology is a useful diagnostic tool to confirm conjunctivalisation of the
corneal surface. Corneal impression cytology staining for goblet cells (which are
present in conjunctival but not corneal epithelium) and for cytokeratin 19 (a marker
for conjunctival epithelium) are commonly used. Imaging of the cornea and limbus,
such as confocal microscopy, are also being investigated but their role in diagnosis is
not yet clear.
2) Management options for limbal stem cell deficiency
Medical or conservative: Lubricants (preservative free) and bandage contact lens
wear are the mainstay. Topical steroids and antibiotics (preservative free) for periods
of significant ocular surface inflammation and epithelial breakdown. Autologous
serum drops have a role in epithelial wound healing but the exact nature of their
action is unclear. Punctal plugs for improvement of dry eye also have a role
Surgical: Prior to any surgical management of the limbal stem cell deficiency, any
eyelid disease and dry eye need to be optimised. Lid margin malposition (cicatricial
entropion) and trichiasis need to be treated. Punctal occlusion and lateral tarsorrhaphy
may be necessary to reduce dry eye. Only once eyelid disease and dry eye have been
treated effectively should one consider surgical management of limbal stem cell
deficiency. The surgical options for limbal stem cell deficiency are based on the
transplantation of healthy limbal epithelium. This can be in the form of whole tissue
grafts or cultured cells. The source of the tissue can be autologous (in unilateral
disease) or allogeneic from matched living donors or cadaveric donors.
3) Culture methods for limbal epithelium
The culture of human limbal epithelium has many variations:
Suspension versus explant
3T3 fibroblast co-culture versus amniotic membrane versus other methods
Foetal calf serum versus autologous serum
The use of airlifting
The duration of culture expansion
Each of these different culture methods will be described in detail (Osei-Bempong et
al Regenerative Medicine 2009).
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4) Transplantation of cultured limbal epithelium
The use of cultured limbal epithelial cell therapy for severe limbal stem cell
deficiency essentially involves three steps:
Procurement of limbal epithelium from donor eye: The removal of a small amount
of limbal tissue (1-2mm X 1-2mm)
Culture: Ex vivo expansion of the limbal epithelium.
Transplantation of ex vivo expanded limbal epithelium: After performing a
superficial keratectomy to remove the conjunctival tissue from the corneo-limbal
surface, the cultured limbal epithelial sheet is sutured to the eye.
There are some significant variations in the procedure which will be discussed:
removal of the explant from the culture versus limbal positioning of explant; the use
of a bandage amniotic membrane (as part of a sandwich technique) versus bandage
contact lens; and post-operative lid closure using a botulinum toxin induced ptosis or
central tarsorrhaphy.
The results from published cases and case series will be discussed (Baylis et al 2011
Journal of Cell Biochem). The overall success rate of cultured limbal epithelial cell
transplantation is 76% (77% for autografts and 73% for allografts).
5) The regulation of cultured limbal epithelial cell therapy
The culture expansion of human limbal epithelium and its clinical application as part
of a clinical trial is highly regulated (for example EMA and MHRA). The actual
culture process has to be performed in accordance with good manufacturing practise
in order to produce transplant grade cultures. The culture process therefore has to be
performed in specialised “clean” laboratories and the culture has to be monitored for
sterility and quality.
The success of cultured limbal epithelial cell transplants is difficult to determine as no
formal clinical trial data is available to date. It is envisaged that EMA or MHRA
approved clinical trials will be conducted to determine the true efficacy of this
treatment modality for limbal stem cell deficiency.
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