Cellular therapy of corneal epithelial defect by adipose mesenchymal stem cell-derived epithelial progenitors

Abstract

BackgroundPersistent epithelial defects (PED), associated with limbal stem cell deficiency (LSCD), require ocular surface reconstruction with a stable corneal epithelium (CE). This study investigated CE reformation using human adipose mesenchymal stem cells (ADSC), which derived epithelial progenitors via mesenchymal-epithelial transition (MET).MethodsSTEMPRO human ADSC were cultured with specific inhibitors antagonizing glycogen synthase kinase-3 and transforming growth factor-β signaling, followed by culture under a defined progenitor cell targeted-epithelial differentiation condition to generate epithelial-like cells (MET-Epi), which were characterized for cell viability, mesenchymal, and epithelial phenotypes using immunofluorescence and flow cytometry. Tissue-engineered (TE) MET-Epi cells on fibrin gel were transplanted to corneal surface of the rat LSCD model caused by alkali injury. Epithelial healing, corneal edema, and haze grading, CE formation were assessed by fluorescein staining, slit lamp bio-microscopy, anterior segment optical coherence tomography, and immunohistochemistry.ResultsCD73high/CD90high/CD105high/CD166high/CD14negative/CD31negative human ADSC underwent MET, giving viable epithelial-like progenitors expressing δNp63, CDH1 (E-cadherin), epidermal growth factor receptor, integrin-β4, and cytokeratin (CK)-5, 9. Under defined epithelial differentiation culture, these progenitors generated MET-Epi cells expressing cell junction proteins ZO1 and occludin. When transplanted onto rat corneal surface with LSCD-induced PED, TE-MET-Epi achieved more efficient epithelial healing, suppressed corneal edema, and opacities, when compared to corneas without treatment or transplanted with TE-ADSC. CE markers (CK3, 12, and CDH1) were expressed on TE-MET-Epi-transplanted corneas but not in other control groups.ConclusionHuman ADSC-derived epithelial-like cells, via MET, recovered the CE from PED associated with LSCD. ADSC can be a viable adult stem cell source for potential autologous epithelial cell-based therapy for corneal surface disorders.

Epithelial differentiation of gingival mesenchymal stem cells enhances re-epithelialization for full-thickness cutaneous wound healing

“…These structures are similar to our SEM observations with GMSCs, indicating that epithelial differentiation of GMSCs can be successfully induced by epithelial cell growth medium. Consistent with previous ndings [12][13][14][15], we found that MET and stemness-related genes and proteins expression decreased while epithelial-related genes and proteins expression increased during differentiation induction. However, there were differences between the different studies regarding epithelial-related genes and proteins that were up-regulated.…”

Epithelial differentiation of gingival mesenchymal stem cells enhances re-epithelialization for full-thickness cutaneous wound healing

“…These structures are similar to our SEM observations with GMSCs, indicating that epithelial differentiation of GMSCs can be successfully induced by epithelial cell growth medium. Consistent with previous ndings [12][13][14][15], we found that MET and stemness-related genes and proteins expression decreased while epithelial-related genes and proteins expression increased during differentiation induction. However, there were differences between the different studies regarding epithelial-related genes and proteins that were up-regulated.…”

Generation of a Biomimetic Substitute of the Corneal Limbus Using Decellularized Scaffolds

“…However, hADSCs were initially negative for these three markers, as is the case of all types of human MSCs, but became positive or slightly positive for the three epithelial markers after 21 days of ex vivo differentiation induction using conditioning media. These results confirm the differentiation capability of hADSCs to the epithelial cell lineage under certain circumstances, as previously suggested [57,58]. Previous results published by our group demonstrated that these cells, which can be harvested autologously, can be differentiated ex vivo using conditioning media, although differentiation is not complete ex vivo and the in vivo environment is required for terminal differentiation [58].…”

Therapeutic Potential of Mesenchymal Stem Cells in the Treatment of Ocular Graft-Versus-Host Disease

“…By using a rabbit model of alkaline-induced corneal injury, Guo and colleagues showed that human BM-MSC differentiated in corneal epithelial cells and migrated in the damaged corneal stroma where improved survival of corneal stromal cells was observed which finally resulted in corneal regeneration and the attenuation of alkaline-induced corneal damage [ 14 ]. In line with these findings are results obtained by Bandeira and coworkers who demonstrated the therapeutic potential of AT-MSC in the treatment of LSCD [ 15 ]. Human AT-MSC, which were cultured on fibrin gel and grafted onto the damaged corneal surface of experimental mice, managed to re-populate limbal stem cells and regenerate the injured corneal epithelium [ 16 ].…”

“…In line with these findings are results obtained in several experimental and clinical studies which demonstrated the capacity of MSC to repair and regenerate the injured corneal epithelium, meibomian and lacrimal glands, indicating their therapeutic potential in the treatment of oGVHD [ 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 ].…”

Epithelial differentiation of gingival mesenchymal stem cells enhances re-epithelialization for full-thickness cutaneous wound healing

“…These structures are similar to our SEM observations with GMSCs, indicating that epithelial differentiation of GMSCs can be successfully induced by epithelial cell growth medium. Consistent with previous ndings [12][13][14][15], we found that MET and stemness-related genes and proteins expression decreased while epithelial-related genes and proteins expression increased during differentiation induction. However, there were differences between the different studies regarding epithelial-related genes and proteins that were up-regulated.…”

Generation of a Biomimetic Substitute of the Corneal Limbus Using Decellularized Scaffolds

“…However, hADSCs were initially negative for these three markers, as is the case of all types of human MSCs, but became positive or slightly positive for the three epithelial markers after 21 days of ex vivo differentiation induction using conditioning media. These results confirm the differentiation capability of hADSCs to the epithelial cell lineage under certain circumstances, as previously suggested [57,58]. Previous results published by our group demonstrated that these cells, which can be harvested autologously, can be differentiated ex vivo using conditioning media, although differentiation is not complete ex vivo and the in vivo environment is required for terminal differentiation [58].…”

Therapeutic Potential of Mesenchymal Stem Cells in the Treatment of Ocular Graft-Versus-Host Disease

“…By using a rabbit model of alkaline-induced corneal injury, Guo and colleagues showed that human BM-MSC differentiated in corneal epithelial cells and migrated in the damaged corneal stroma where improved survival of corneal stromal cells was observed which finally resulted in corneal regeneration and the attenuation of alkaline-induced corneal damage [ 14 ]. In line with these findings are results obtained by Bandeira and coworkers who demonstrated the therapeutic potential of AT-MSC in the treatment of LSCD [ 15 ]. Human AT-MSC, which were cultured on fibrin gel and grafted onto the damaged corneal surface of experimental mice, managed to re-populate limbal stem cells and regenerate the injured corneal epithelium [ 16 ].…”

“…In line with these findings are results obtained in several experimental and clinical studies which demonstrated the capacity of MSC to repair and regenerate the injured corneal epithelium, meibomian and lacrimal glands, indicating their therapeutic potential in the treatment of oGVHD [ 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 ].…”

Epithelial differentiation of gingival mesenchymal stem cells enhances re-epithelialization for full-thickness cutaneous wound healing

“…These structures are similar to our SEM observations with GMSCs, indicating that epithelial differentiation of GMSCs can be successfully induced by epithelial cell growth medium. Consistent with previous ndings [12][13][14][15], we found that MET and stemness-related genes and proteins expression decreased while epithelial-related genes and proteins expression increased during differentiation induction. However, there were differences between the different studies regarding epithelial-related genes and proteins that were up-regulated.…”

Generation of a Biomimetic Substitute of the Corneal Limbus Using Decellularized Scaffolds

“…However, hADSCs were initially negative for these three markers, as is the case of all types of human MSCs, but became positive or slightly positive for the three epithelial markers after 21 days of ex vivo differentiation induction using conditioning media. These results confirm the differentiation capability of hADSCs to the epithelial cell lineage under certain circumstances, as previously suggested [57,58]. Previous results published by our group demonstrated that these cells, which can be harvested autologously, can be differentiated ex vivo using conditioning media, although differentiation is not complete ex vivo and the in vivo environment is required for terminal differentiation [58].…”

Therapeutic Potential of Mesenchymal Stem Cells in the Treatment of Ocular Graft-Versus-Host Disease

“…By using a rabbit model of alkaline-induced corneal injury, Guo and colleagues showed that human BM-MSC differentiated in corneal epithelial cells and migrated in the damaged corneal stroma where improved survival of corneal stromal cells was observed which finally resulted in corneal regeneration and the attenuation of alkaline-induced corneal damage [ 14 ]. In line with these findings are results obtained by Bandeira and coworkers who demonstrated the therapeutic potential of AT-MSC in the treatment of LSCD [ 15 ]. Human AT-MSC, which were cultured on fibrin gel and grafted onto the damaged corneal surface of experimental mice, managed to re-populate limbal stem cells and regenerate the injured corneal epithelium [ 16 ].…”

“…In line with these findings are results obtained in several experimental and clinical studies which demonstrated the capacity of MSC to repair and regenerate the injured corneal epithelium, meibomian and lacrimal glands, indicating their therapeutic potential in the treatment of oGVHD [ 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 ].…”