Surgeon: Ex vivo expansion of limbal cells viable for treating limbal cell deficiencies

BARCELONA — The ex vivo expansion of limbal stem cells may be an option for treating severe corneal conditions related to total limbal cell deficiencies, according to a surgeon speaking here at the European Society of Cataract and Refractive Surgeons Winter Refractive Surgery Meeting.

Frederick E. Kruse, MD, of Erlangen-Nürnberg University discussed his method for culturing such cells, which involves a small piece of limbal tissue about 1 mm × 2 mm in size placed on a suitable carrier of amniotic membrane or fibrin glue matrix. Over 2 to 3 weeks, the tissue is left to grow into a transplantable cell sheet 2 cm to 3 cm in diameter.

The cultured cell sheet is then trimmed to the desired size and transferred onto the ocular surface, Dr. Kruse said.

"We compared all carrier systems in vitro and found that the appearance of the cells on fibrin and amniotic membrane is similar," Dr. Kruse said. "Also, electron microscopy does not show any significant difference; the current stem cell markers are similarly expressed on fibrin and amniotic membrane."

The most successful study on the use of autologous tissue-engineered limbal cell sheets was published by Paolo Rama and colleagues in Italy, which involved a fibrin glue matrix as a carrier, he said.

"Using this method, they treated corneas that were severely damaged and, in some cases, had undergone one or more transplantation procedures, obtaining a high rate of success. It was after reading this paper that we chose to use fibrin glue instead of amniotic membrane, which has considerable biological variability," Dr. Kruse said.

Clonal analysis was another improvement suggested by the Italian group, he noted.

"Limbal stem cells have the unique capacity to form colonies, [and] clonal analysis helps differentiate between stem cells with higher or lower capabilities to form colonies," Dr. Kruse said. "It is also important to localize the small pockets (niches) in the limbus where these stem cells reside."

For bilateral transplantations, the use of stem cells obtained from other bodily regions has recently started being investigated, Dr. Kruse added. Kohji Nishida, MD, advocated the use of cell sheets comprised of autologous oral mucosal epithelium. These cells are cultured on temperature-responding sheets, amniotic membrane or fibrin gel.

"Initial results are favorable," Dr. Kruse said.

Two other potential sources of stem cells - the conjunctiva and skin - are also currently under investigation, he said.