title: Photochemical tissue bonding: a potential strategy for treating limbal stem cell deficiency.
abstract: OBJECTIVE
  To determine the feasibility of attaching human amniotic membrane (HAM), pre-cultured with limbal stem cells (LSCs), to cornea using a novel, light-activated tissue bonding method.
  
  
  METHODS
  LSCs were isolated from rabbit eyes, and then cultured on de-epithelialized HAM to create grafts (HAM/LSC). These were then transplanted onto rabbit eyes with surgically created limbal stem cell deficiency (LSCD). The grafts were secured either by sutures or by a light-activated method called photochemical tissue bonding (PTB). Outcomes included corneal opacity, inflammation, neovascularization, and collagen alignment.
  
  
  RESULTS
  The isolated and cultured cells were verified to be LSCs based on their K19+/intergrin β1+/P63+/K3 profile. Securing the HAM/LSC graft with PTB provided better outcomes. At 28 days post-surgery, the corneal opacity scores were significantly lower after securing the graft with PTB compared with suture attachment (0.8 ± 0.5 vs. 1.8 ± 0.5, P < 0.01). Similarly, neovascularization scores were lower after PTB (0.8 ± 0.5 vs. 1.5 ± 0.6, P < 0.01). Quantification of MPO and CD31 levels from immunofluorecent staining indicated that PTB stimulated less neutrophil infiltration (5.3 ± 2.2 vs. 13.3 ± 3.1, P < 0.01) and less new blood vessels formation (2.0 ± 0.8 vs. 6.3 ± 1.3, P < 0.01) at the wound site. The collagen alignment in PTB-treated corneas, as shown by immunofluorescence and second harmonic generation image, was better organized in the PTB-treated group than in the suture group.
  
  
  CONCLUSIONS
  Bonding LSC grafts with PTB produced improved outcomes compared to suture attachment. This light-activated method is a promising modality for treating patients with LSCD.
authors.affiliation: Department of Burns and Plastic Surgery, No. 3 People's Hospital, and Institute of Traumatic Medicine; School of Medicine, Shanghai Jiao Tong University, Shanghai 201900, China.
authors.forename: Chuan
authors.initials: C
authors.lastname: Gu
authors.forename: Tao
authors.initials: T
authors.lastname: Ni
authors.forename: Erol E
authors.initials: EE
authors.lastname: Verter
authors.forename: Robert W
authors.initials: RW
authors.lastname: Redmond
authors.forename: Irene E
authors.initials: IE
authors.lastname: Kochevar
authors.forename: Min
authors.initials: M
authors.lastname: Yao
mesh_terms: D000650:Amnion / Q000637:transplantation*; D000818:Animals; D003316:Corneal Diseases / Q000601:surgery*; D048988:Corneal Surgery, Laser / Q000379:methods*; D005240:Feasibility Studies; D005456:Fluorescent Dyes / Q000627:therapeutic use; D006801:Humans; D053844:Lasers, Solid-State / Q000627:therapeutic use; D016850:Limbus Corneae / Q000166:cytology* / Q000503:physiopathology; D055668:Photochemical Processes; D011817:Rabbits; D011897:Random Allocation; D012395:Rose Bengal / Q000627:therapeutic use; D033581:Stem Cell Transplantation / Q000379:methods*; D013536:Suture Techniques; D014014:Tissue Adhesives / Q000627:therapeutic use; D014183:Transplantation, Heterologous; D016896:Treatment Outcome; D014945:Wound Healing
chemical_list: D005456:Fluorescent Dyes; D014014:Tissue Adhesives; D012395:Rose Bengal