Lecture: The Future of Limbal Stem Cell Transplantation

DR NARIANI: Well, good morning, good afternoon, good evening to each and every one of you.

My name is Ashiyana Nariani, and it is truly an honor to be with you today on Cybersight.

The most renowned platform that I know of, globally, that makes such an impact in education.

Today’s topic is the future of limbal stem cell transplantation.

I have no relevant financial disclosures.

Before we get started, let’s start off with a few questions as a pretest.

Question number one — and we’re gonna do this in the form of a poll — which of the following ocular surface disease or diseases is or are associated with limbal stem cell deficiency?

And that is A: Contact lens wear, B: Aniridia, C: Radiation, or D: All of the above.

We’re gonna give time for participants to chime in on the poll.

Okay. And we have as answers: 5% for contact lens wear, 3% for aniridia, 8% for radiation, and 84% for all of the above.

So I’m going to keep a stoic face, because we are going to find out the answers at the end of the lecture.

Question number two. Which of the following is true regarding staging of limbal stem cell deficiency, based on the global consensus by the international stem cell deficiency working group? Limbal stem cell deficiency working group. My apologies. That should say international limbal stem cell deficiency working group.

And the answer choices are: A, stage 1B is defined as normal central corneal epithelium with 100% limbal involvement.

B, stage IV indicates that the entire corneal surface is affected.

C, stage II A is defined as central 5 millimeter zone of cornea being affected, with less than 50% of limbal involvement.

Or D, stage II C is defined as the central 5 millimeter zone of cornea being affected, with greater than or equal to 50% but less than 100% limbal involvement.

Let’s give time to all of us to give in our best answers on the poll.

Okay. We’ve got A, 6%. B, 40%, which is saying that stage IV is the entire corneal surface being affected, C, 25%, and D, 29%. So we’re kind of all over the board.

But the majority has chimed in that B is — like I said, we’re gonna go through the answers at the end. So stay tuned.

Question number three. What does the limbal stem cell transplant technique SOMET stand for? S-O-M-E-T? Is that A, simple oral mucosal epithelial transplant?

B, serum oral mucosal epithelial transplant? C, synergistic oral

mucosal epithelial transplant?

Or D, simple oral mucosal endothelial transplant?

Give it your best shot, everyone.

All righty. We’ve got 65% of participants saying that SOMET stands for simple oral mucosal epithelial transplant, with 17% saying simple oral mucosal endothelial transplant, 11% for synergistic oral mucosal epithelial transplant, and lastly, 7% of participants saying serum oral mucosal epithelial transplant.

Question number four. What is a possible finding on anterior segment OCT imaging in limbal stem cell deficiency as compared to normal subjects?

And here are your options. A, on AS OCT, are we seeing in limbal stem cell deficiency a thinner central corneal epithelial thickness?

Or B, thicker maximum limbal epithelial thickness?

C, thinner central stromal thickness?

Or D, no change in maximum limbal epithelial thickness?

Which of the following has been found on anterior segment OCT in limbal stem cell deficiency?

Okay. So… Woo! Divided. 40% say A, thinner central corneal epithelial thickness. Epithelium thickness. With another 40% of participants saying thicker maximum limbal epithelial thickness.

6% for thinner central stromal thickness. And 13% with no change in maximum limbal epithelial thickness. I love this. Okay?

So this is going to be a great session. I am looking forward to this. Limbal stem cell deficiency is one of my favorite topics.

And we will, at the end, go through the answers that are the correct answers for those four questions.

So let us start by defining limbal stem cell deficiency.

Limbal stem cell deficiency is an ocular surface disease caused by a decrease in the population and/or function of corneal epithelial stem/progenitor cells. This decrease leads to the inability to sustain the normal homeostasis of the corneal epithelium.

Now, this is very important, that we know the source. So this was per the international limbal stem cell deficiency working group.

And this is a great paper that I have cited here for all of us to really dive in deep. Because this is the latest, in terms of defining limbal stem cell deficiency.

And from that, beyond definition, we also have the classification and the diagnosis and the staging.

The disease is characterized by conjunctivalization, which is the replacement of the normal corneal epithelium by conjunctival epithelium.

And/or signs of epithelial dysfunction, such as persistent or recurrent epithelial defects.

With or without neovascularization, ocular surface inflammation and scarring.

Frequent consequences are decreased vision and discomfort leading to reduced quality of life. Limbal stem cell deficiency may present alone as a single entity or associated with abnormalities of other components of the ocular surface, such as the conjunctiva, meibomian glands, lacrimal glands, tears, corneal nerves, and immune system.

So let us understand partial versus total limbal stem cell deficiency.

Partial limbal stem cell deficiency is characterized by the incomplete conjunctivalization of the corneal surface.

And the presence of residual limbal and consequent corneal epithelial cells.

And this can beautifully be depicted in the photographs on the right that stand for 1C and 1D.

You can appreciate that there is good presence still of limbal stem cell cells and epithelial cells, and that are visible both on slit lamp, white light, on the left, 1C, and on fluorescein lighting, 1D.

Total limbal stem cell deficiency is characterized in 1E and 1F. The images on your right.

By conjunctivalization of the entire corneal surface. Due to a complete loss of corneal epithelial stem progenitor cells.

And this is very important, because staging is critical to then understanding what is it that we’re gonna do about it.

So the staging based on the international classification by the international limbal stem cell deficiency working group is… And this was one of the questions that came up, regarding the pretest…

So let’s go through it. Staging actually comes into three stages.

Not four. So that was a trick question. Three stages. Stage one refers to normal corneal epithelium within the central 5 millimeter zone of the cornea.

So stages really are relating to the status of the cornea. And then the A, B, and C relate to the limbal involvement.

So when we talk about stage 1A, that would be with less than 50% of limbal involvement.

B, 1B, would be greater than or equal to 50%. But less than 100% limbal involvement. And C would be total. 100% limbal involvement.

Stage 2 is where… Remember, we’re referring to the corneal status, when we talk about the stages. So stage 2 refers to that central 5 millimeter zone of cornea being affected.

A continues to remain… A and B continue to remain the same definitions. A less than 50% limbal involvement, B, greater than or equal to 50% but less than 100% limbal involvement, and notice here that there is no C.

Stage 3 is where the entire corneal surface is affected. And there’s no need for A or B or C in that situation.

So what are the causes of limbal stem cell deficiency? Well, it’s important to note that you can categorize it between hereditary and acquired.

Hereditary causes include aniridia, multiple endocrine deficiency, epidermal dysplasia, dyskeratosis congenita, and the acquired causes include but are not limited to…

Chemical or thermal injury, Stevens-Johnson syndrome, toxic epidermal necrolysis, mucous membrane pemphigoid, MMP, ocular surgeries involving the limbus, contact lens wear, severe microbial infection extending to the limbus,

antimetabolites such as 5 fluorouracil or mitomycin C can cause limbal stem cell deficiency, and we must be attuned to that when performing glaucoma or pterygium procedures or other procedures.

Radiation, chronic limbitis, vernal or atopic conjunctivitis, bullous keratopathy, and neurotrophic keratitis.

So let’s dive in a little bit deeper into limbal stem cell deficiency. As we defined it earlier, partial versus total.

So with partial, we have to ask ourself: Question number one, is the visual axis affected? Yes or no. That will define how you move ahead into your treatment of choice.

So if the visual axis is affected, follow me here — through the diaphragm.

If the visual axis is affected, yes, then you have to ask yourself: Is the adnexa affected? Yes or no?

And if it is, you want to correct the adnexal abnormality before moving ahead into any sort of ocular surface procedure.

And that’s one of the pearls that I want to bring to you today.

Is always correct the eyelids before moving into any abnormalities of the ocular surface.

If the adnexa is not affected, then you look into: Is there fiber vascular pannus?

Yes or no?

Then consider doing a sequential sectoral conjunctival epitheliectomy, removal, plus or minus amniotic membrane, or consider stem cell therapies.

If there is no fiber vascular pannus, then at this point, consider doing sequential sectoral conjunctival epitheliectomy, mechanical brushing or scraping of… As you can see…

(audio drop)

If the visual axis is not affected, consider topical lubrication alone. If you have total limbal stem cell deficiency, then you have to consider:

Is the visual axis affected? Yes or no. If it is, then again, ask yourself: Is the adnexa affected? Yes or no. And if it is affected, correction of that abnormality. Always start there.

And if no, then ask yourself: Is it unilateral or bilateral?

If the visual axis is not affected, topical lubrication suffice.

Now, this is where we go into the unilateral or bilateral.

Okay? If it is unilateral, then the stem cell therapies can be thought of — and this thing — where, when, how, what do we do? This is very important. Stay tuned to this particular slide.

If it is unilateral, then the stem cell therapies can be then further classified into autologous, versus allogeneic. And we’re gonna go into each of these individually.

Autologous options being CLET, SLET, and COMET. Allogeneic including allogeneic CLET, lr-CLAL, KLAL.

Now, it’s tough when both eyes are affected. So with bilateral limbal stem cell deficiency, ask yourself: Is the ocular surface wet? Really consider keratoprosthesis. If the ocular surface is wet, you can consider keratoprosthesis. But you can also consider CLET, SLET, or COMET. Simple limbal epithelial transplantation — how can those be done if it’s bilateral? Studies have shown…

And this is done by Dr. Gita, a great friend of mine, doing SLET in bilateral cases, when you do have some area in which you can harvest tissue.

Allogenic options in stem cell therapy, in the realm of stem cell therapy options, include allogenic CLET, lr-CLAL, and KLAL. And in severe unilateral or bilateral disease — remember, this is all about definitions here — Cincinnati procedure as being an option for you.

And the Cincinnati procedure, by Dr. Edward Holland and Dr. Albert Cheung, is living related CLAL, plus KLAL. The combination.

Okay. Here we go. So let’s break it down further. The principles of autologous limbal stem cell techniques.

There are four major autologous limbal stem cell techniques.

And they are: CLAU,

where you have harvest of conjunctiva and limbal tissue from the contralateral eye,

and I’m happy to share my slides with anyone who needs it thereafter. Because these slides really help walk you through the procedures and definitely takes time beyond the time we have today to really go into it in depth, for those who are looking to start limbal stem cell transplantation.

So you have 2 to 4 strips from the superior and inferior donor limbus. And there’s direct transfer of tissue into the recipient eye.

This is a single procedure. Limitations here are… This is predominantly for unilateral cases of limbal stem cell deficiency. There is of course risk to the donor eye. The live donor — only a part of the donor limbus can be transplanted.

And the procedure cannot be repeated if it fails.

The next procedure in the autologous realm is CLET. Autologous CLET. And that is where we have harvest from the limbal stem cells from the contralateral eye, ex vivo expansion into limbal epithelial sheets, and this is a staged procedure.

Limitations include risks to the donor eye, requires presence of a small area of preserved limbus, and the requirement of ex vivo stem cell expansion.

What does that mean? When we take it into the laboratory for ex vivo stem cell expansion, there are those additional risks that this can fail.

It’s time-consuming to do so. And the transplantation has been innovated by my dear friend, Dr. Virender Sangwan, out of India. And this is the combination of CLAU plus CLET.

It is the harvest of limbal stem cells from the contralateral eye, but remember, we talked about it being in bilateral cases. So there have been case reports and studies by Dr. Gita regarding doing this bilaterally.

But generally speaking, it is harvest of limbal stem cells from the contralateral eye, direct transfer of limbal stem cells into the recipient eye, and it is simple. It’s a one stage procedure. There is no staging, I should say.

It’s a one time procedure. And this, per Dr. Sangwan, he said — up to three times. So that’s a real benefit here for patients.

What are the limitations? Well, there’s the risk to the donor eye. It’s primarily studied in unilateral cases.

The long-term efficacy has not yet been proven. Although… Because it is the newest player on the block… But at the moment, it is not necessarily… It’s not new-new. So we do have great results and reports out there, in terms of longer time period.

But still, it’s the newest kid on the block. And so… Longer term efficacy has yet to be shown.

Minimal efficacy for data, for patients undergoing keratoplasty… Though I have done it in patients with pediatric limbal stem cell deficiency, secondary to what we call lime injury, doing SLET plus a keratoplasty. And I do feel that there’s a lot more that needs to be done in this realm, for sure.

COMET is harvesting from oral mucosa with ex vivo expansion of epithelial stem cell sheets.

And here, ex vivo stem cell expansion is required. So again… It can fail… It’s time-consuming. And it’s expensive.

What are the principles for allogenic stem cell techniques? Defined as allogenic CLET, where we have ex vivo expansion of limbal stem cells into limbal epithelial cell sheets. For unilateral or bilateral disease, you can have living donor or cadaver, and it’s a single procedure.

Limitations are immunosuppression. And that you can see is a running theme for our cases. Ex vivo stem cell expansion is required. It can fail. As a result, when we do move into a staged procedure here…

With ex vivo stem cell expansion, time-consuming, and it’s expensive.

The second of the five procedures I want to highlight to you is living related CLAL. It’s similar to the CLAU technique.

It can be used for unilateral or bilateral disease. Here you need a living related donor. Hence the term lr for “living related”.

The donor and recipient immunological matching base is critical, based on blood type and antigens. And tissue is fresher here than KLAL.

Limitations here: Immunosuppression, risk to the donor eye, it’s a live donor, so only part of the donor limbus can be transplanted. If it fails, another donor must be found.

Now, with KLAL, 360-degree ring that consists of the entire donor eye’s limbus, with cadaver donor, is used. This can be done for unilateral or bilateral disease.

Here, limitations include immunosuppression, graft up to 24 hours old before retrieval, tissue not immunocompatible, and the risk of rejection and failure rate is high.

Now, the final two procedures, limbal stem cell types, transplant types, in the allogenic category, are the Cincinnati procedure and the modified Cincinnati procedure.

Remember what we talked about a few slides before. The definition of the Cincinnati procedure is that you have a living related CLAL, plus KLAL both. This involves living donor tissue, of someone who is related, and this is predominantly used for severe unilateral or bilateral disease.

In these conditions, living donor tissue is used from the 6:00 and 12:00 meridians, and cadaver tissue, 3:00 and 9:00 meridians.

Limitations: Again, immunosuppression. With the living related CLAL part, there is the risk to the donor eye, and the live donor, only part of the donor limbus can be transplanted.

If it fails, another donor must be found. When it comes to the KLAL part, what are the limitations? Well, the graft up to 24 hours old, before retrieval. Tissue not being compatible. Risk of rejection.

And failure rate being high. The modified Cincinnati procedure is CLAU, instead of living related CLAL. Plus KLAL.

Here you have living donor tissue, which is the self, and this is used for severe unilateral disease, as compared to the Cincinnati procedure, which can be used for severe unilateral or bilateral disease.

Here, living donor tissue can be used from the 6:00 and 12:00 meridians, the cadaver tissue from the 3:00 and 9:00 meridians.

Here, when you break it down to what the limitations are, for CLAU, they include the risk to the donor eye, and the procedure cannot be repeated if it fails. Risks regarding the limitations of KLAL — immunosuppression and of course the other related KLAL limitations that are mentioned above. With the Cincinnati procedure.

Graft up to 24 hours old before retrieval, tissue not compatible, and the risk of rejection and failure rate being high.

So there was a question — I really appreciate a few of you have already submitted questions prior to today’s lecture.

And one of them was: How do you go about preparing amniotic membrane? Because amniotic membrane is a critical part of a number of the procedures that are performed in limbal stem cell deficiency.

Not to mention that amniotic membrane alone, amniotic membrane grafting alone, with either glue or vicryl suture, without doing a limbal stem cell transplant, can be so therapeutic. And why?

It is because factors that are present in epithelialization, like epidermal growth factor, keratinocyte growth factor, transforming growth factor α, and so forth, as you can see here — also amniotic membrane suppresses angiogenesis.

Which is so critical. It suppresses inflammation, scarring, as well, and it has an antimicrobial function.

So this is a video that I saw. Forgive me for it not being the clearest. But it is a video. That was… (audio drop) society. So there is in amniotic membrane — remember, there’s the epithelial side and basement membrane.

So when we are preparing amniotic membrane tissue, and this is really important for my colleagues in Ghana, Ethiopia, South Sudan, that I work with, who are so interested in going about bringing amniotic membrane for their ocular surface disease patients…

So you have to (inaudible) from the chorion. And this is what you’re seeing here in the video. This is your shiny amniotic membrane tissue. And once you dissect this amniotic membrane away from the chorion, you have to make sure to — all those pieces of chorion tissue that remain —

to go ahead and remove that. That is critical. Then you use it within nitroglycerol paper. You’re gonna implant this on nitroglycerol paper.

And you are going to have to ensure that your epithelial side is up. Now, how do you know which side is epithelial side? Well, classic question.

Weck-Cel sponge always upon the epithelial side, and you will see that it will lift up so nicely. So that’s a key tip using the Weck-Cel sponge. Once you place it on the nitroglycerol paper, you’re ready to use it in the surgery. The paper you use has to be presterilized. So that’s very important.

And do not do this unless you have direct guidance first by someone who has experience in this.

Is my big caution. Because there are the sterility matters that you really want to take into account before you move ahead on the journey.

But I’m very excited for those colleagues that have asked for knowing more about the preparation.

So coming to the topic today, which is the future of limbal stem cell transplantation, we have… If you go into PubMed, and you type in limbal stem cells, 100 publications as of 2020 and 115 publications as of 2022, if you were to go into PubMed and type out limbal stem cells…

You can break down… I’ve gone through the

literature, and you can find them classified into five major categories. And that is: Surgical techniques, pathophysiology, immunosuppression, limbal stem cell diagnostics, and stromal stem cells.

So we’re gonna go through these, and I’m gonna give you the highlights. I’m not gonna give you 142 here today.

But I want to give you and share with you the highlights from each of these five categories.

Another place and resource for you is… We had put together a limbal stem cell advancements course that is available for all.

On YouTube. Limbal stem cell advancements 2020, with Virender Sangwan, Albert Cheung, Shigeru Kinoshita, Melissa Daluvoy, and Bhaskar Srinivasan. So I highly recommend going back to that as well for some further information. It’s about a two and a half hour course.

So focusing in on the highlights of the future, as it relates — future and current highlights of limbal stem cell transplantation is allo-SLET, which (inaudible) has case reports on. Allo-SLET, better for earlier phase of chemical injury in grade four and above.

There’s also as we talked about earlier… How do we combine PKP, penetrating keratoplasty, along with a limbal stem cell transplant?

And this paper had come out, regarding PKP, following CLAU.

And the thought was that waiting for at least one year post-CLAU transplantation to perform a PKP will increase corneal clarity.

So again, this idea of staging, even in my hands, I have done it, more and more of the importance of doing a staged procedure, getting that ocular surface to be healed, and then moving into doing a penetrating keratoplasty.

But the future is yet to show us more. Living related CLAL versus KLAL for cases of bilateral total limbal stem cell deficiency — a paper showed there that limbal related CLAL demonstrates lower rejection rates, improved graph survival, and better best corrected visual acuity, compared with KLAL.

Allogenic CLET was highlighted for cases of bilateral total limbal stem cell deficiency.

And they found in that paper that 70%, 7 out of 10 cases, had failed. The conclusion being that it may be considered in patients with limbal stem cell deficiency due to burns or congenital etiology such as aniridia, but its benefits are limited for patients with immunological diseases such as OCP or idiopathic panuveitis.

There was a case report regarding the use of solid activated platelet-rich plasma. With allogenic limbal epithelial stem cell transplantation from a cadaveric donor.

Allogenic CLET versus cultivated autologous oral mucosal epithelial cell sheet in the literature… There was a comparison study — a large evaluation done on cases of total bilateral limbal stem cell deficiency.

And it concluded that comparison between studies was difficult, due to the lack of universal and objective grading tools for assessing postoperative results.

The definition of clinical improvement was also found to be problematic. And with autologous COMET, there is no risk of immune activation and no immunosuppression, but increased risk of persistent epithelial defects and graft failure, compared with allogenic CLET.

And finally, in the surgical techniques category, PROSE scleral lenses were highlighted as potentially improving epithelial defect resolution in limbal stem cell deficiency cases.

A few more discussion articles in this realm were corneal neurotization, the use of SMILE lenticules for limbal stem cell deficiency, SLET — of course, we know that it has been there since 2012, and 31 articles were reviewed between 2012 to 2020.

But further, additional studies like the allogenic, the allo-SLET procedure is further in the works. The option of xenofree alternatives for culturing limbal epithelial stem cells for evaluated. The autologous transplantation of conjunctiva by modifying simple limbal epithelial transplantation for limbal stem cell deficiency and femtosecond laser assisted KLAL.

This is one of the questions that was asked. What does SOMET stand for? Difficult question. But…

It stands for simple oral mucosal epithelial transplantation.

It is a bounce off of the COMET procedure that was innovated by Shigeru Kinoshita out of Japan. And here in this rabbit model, you can appreciate in the SOMET group:

Epithelium expanding over the corneal surface around the grafts at one week after surgery and epithelialization being complete by two weeks, with no fluorescein staining of the cornea being observed at two weeks.

So what is this about? Basically, when we talk about limbal — simple limbal epithelial transplantation, SLET, right, you have taken limbal stem cells from unilateral disease, and transplanting it into the affected eye.

With SOMET, you’re taking a little bit of oral mucosa and transplanting it in a single procedure, no staged procedure, no ex vivo stem cell expansion in the laboratory, direct transplantation into the eye.

When we come to the second category of the different aspects of what’s hot in the literature is diagnostics. And as far as diagnostics, there was the discussion of custom made web-based grading system, based on vascularization, haze, and integrity.

And the use of OCT angiography has also been explored to provide novel quantitative and non-invasive parameters to assess limbal stem cell deficiency.

And they have found markers such as maximum corneal vascular extension, COVE, from the limbus to the furthest vessel over the cornea, as well as corneal vascular thickness, as potential markers.

They’ve also used 3D micro/nanostructures of the stem cell niche, and evaluation and imaging of that. Anterior segment OCT — this was a question that was brought up in the pretest. And we’re gonna go over the answers.

But listen up here: Central corneal epithelium thickness, CET, and maximum limbal epithelium thickness, mLET, were thinner, guys, thinner, in patients with limbal stem cell deficiency, compared to normal subjects.

It has also been discussed in the literature of the use of keratograph 5M non-invasive tear break-up time as a useful tool for evaluating the ocular surface in limbal stem cell deficiency.

Coming to pathophysiology…

Coming to pathophysiology here…

What’s hot in the literature has been anti-VEGF therapy.

Using porcine animal models of limbal epithelial stem cells. Human adipose mesenchymal stem cells as a possible viable source for adult stem cells.

Ocular surface in Stevens-Johnson, they have identified a higher proportion of pathogenic microorganisms, including pseudomonas, staphylococcus, streptococcus, shown in the Stevens-Johnson group, as compared to normal.

Transcriptase phenotype… (audio drop) keratin 12 mRNA expression possibly serving as an early corneal marker for limbal explant cultures, and possible limbal stem cell deficiency after glaucoma surgery.

Very interesting paper. When it comes to explant site of the limbal stem cells, where do you go? It has been found when you look at these three areas, LCOR, LM, or LCONJ, LM standing for middle, which one has been the best?

In terms of the success for transplantation, they have found expression of these two factors that were checked in LCONJ and LM that have been previously shown in having successful transplantation.

Hot off the press in diagnostics also — we brought up these five points.

And finally, immunosuppression. What’s hot off the press in the realm of immunosuppression include comparison of immunosuppression for allogenic limbal grafts and allogenic limbal epithelial stem cell transplantation, oral cyclosporine A was evaluated at different doses. However, studies using oral mycophenolate mofetil and tacrolimus were used as well. Cyclosporine A was evaluated in Stevens-Johnson and TEN and found no difference.

They looked at epidermal growth factor ointment, as possibly reducing symptoms and promoting corneal epithelialization of refractory persistent epithelial defects.

And the use of autologous serum tears can medically reverse severe contact lens induced severe limbal stem cell deficiency and prevent the need for further surgical intervention.

So going back to the beginning of today’s lecture, we’re going to go through the answers for those four questions.

Question number one was: Which of the following ocular surface diseases is or are associated with limbal stem cell deficiency?

And you all got it right. Those of you that — I should say — the majority of you got it right. With D being all of the above.

Contact lens wear, aniridia, radiation. And today we have gone through the list of acquired and hereditary causes of limbal stem cell deficiency.

Very important to note: The causes, because once you understand the cause — anything is potentially reversible. Such as with contact lens wear. Really getting to that problem before moving ahead into therapies.

Question number two: Which of the following is true regarding staging of limbal stem cell deficiency based on the global consensus by the international stem cell deficiency working group?

And the answer here was C, stage II A. The central 5 millimeter zone of the cornea is affected with less than 50% of the limbal involvement.

That was the only one of these… (audio drop) B, for those of you, the majority, if I remember correctly, had chosen… That the entire corneal surface was affected.

Remember that there was no stage IV. There were only three stages in the classification.

So to come up with what was right — it was that stage III is (audio drop) entire corneal surface was affected.

Question number three: What does the limbal stem cell technique SOMET, S-O-M-E-T, stand for? And the answer was: Simple oral mucosal epithelial transplant.

Remember what that is? That we are transplanting oral mucosa directly into the eye. No staged procedure. Direct transplantation. Just like a shoot off of COMET and off of SLET, simple limbal epithelial transplantation. And finally: Question number four.

What is a possible finding on anterior segment OCT imaging in limbal stem cell deficiency as compared to normal subjects? And that was thinner central corneal epithelium thickness.

I remember 40% of participants had chosen A, and 40% of participants had chosen B.

So B would have been right, if instead of thicker, we had said thinner. So the answers… What are the anterior segment OCT imaging findings that you find in limbal stem cell deficiency patients?

It is thinner central corneal epithelial thickness and thinner maximum limbal epithelial thickness. So with that, I thank you very much for your time.

And I am going to open it up for any questions that anyone may have.

I see that we do have a question on the chat box.

By Alejandro. That says: In case of bilateral limbal stem cell deficiency, would you prefer autologous COMET from oral mucosa, or any of the allogenic techniques?

I guess the answer to that is… It depends. Right?

We definitely want to give you an answer there, but it’s case dependent. And so you really have to understand what is the cause, what is the proximity, to presentation and all of that.

Happy to answer any questions specifically on a particular case, if you have.

Let me go into this a little bit more, regarding KLAL. I think the best way to show it…

None of this is easy topics, my friend. So I am definitely going to try and explain to you visually. I’m gonna pull up my slides regarding KLAL, keratolimbal allograft here. And go through that in a little bit more detail. Okay?

So… KLAL represents an allogenic limbal stem cell technique. Allogenic being that we are using it from another donor. That could be…

A living related donor or it could be a cadaver.

In KLAL, keratolimbal allograft, we are using the tissue from a cadaveric donor.

And I want you to take a look at the bottom row here.

And look at the diagram.

PROFESSOR: Sorry, Dr. Nariani. You have to reshare your slides. We’re not seeing them.

DR NARIANI: Thank you so much for bringing that up. Okay.

So here, for KLAL… Thank you, Lawrence. Thank you.

So here I want to bring up — to the question of KLAL, it’s very important to see it visually. Okay?

So let me just repeat what I was just saying.

KLAL stands for keratolimbal allograft.

It is in the category of allogenic limbal stem cell techniques. Allogenic limbal stem cell techniques being:

That the tissue for transplantation is coming from another. Who is that other? It could either be a living related donor or other relative donor,

or it could be a cadaver. With keratolimbal allograft, it is a cadaveric donor.

And as you can see in the diagram at the bottom row…

I want you to pay attention to the diagram there. A 360-degree ring that consists of the entire donor’s eye.

Limbus. Is used. Okay? This is very important. And is very different from the one above that. That is where you’re doing transplantation. Taking a piece of tissue, and you’re either transplanting it at 12:00 and 6:00,

leaving some of the patient’s original tissue in place, or 3:00 and 9:00.

Okay? Here it’s a full 360 degrees. I have a patient with severe unilateral disease, and I want to do a KLAL. What are the steps there? I would call the eye bank. I would ask them for penetrating PKP tissue.

And I would then basically cut out the rims of the corneal scleral rim. 360. Not using the actual tissue, corneal tissue, I should say. I would use the limbal stem cells along with a little bit of the scleral rim.

And I would then do a 360 on my patient’s eye, come back, I would do a 360-degree conjunctival peritomy, at which time I would then do a suturing with 10-0 nylon suture.

So that they are not going to be absorbed. We want a non-absorbable suture.

So 10-0 nylon would be my suture of choice. And then I would go ahead and transplant that full 360 degrees. But as you can see in the imaging here, it’s not necessarily that you would take it verbatim uncut.

Here, as you can see, it can be cut into three parts. Why? Because naturally you may not get the same limbus to limbus dimensions as the person — as the donor, cadaveric donor, limbus to limbus dimensions.

And so that’s why cutting it would be just fine. I’m sorry about that, that you could not see my screen.

I would be happy to answer further questions.

But I think our time is running fast.

So I will give you my email address. Let me go ahead and share that with everyone. And that way you can all be in touch with me, should you have any questions at all in the future or specific cases.

That I can be able to answer for you. Hold on. I’m going to just… [email protected].

This is my India number. And for those of you on WhatsApp, you can contact me at any time. Thank you. It’s truly been an honor and a privilege to be with you.

And I look forward to getting everyone’s questions and learning from all of you as well. Thank you very much.