title: Un-cross-linked fibrin substrates inhibit keratinocyte spreading and replication: correction with fibronectin and factor XIII cross-linking.
abstract: Wound repair is characterized by the presence of a fibrin-rich matrix, but the effect of fibrin on re-epithelialization remains unclear. In this study, we determined the effects of different fibrin matrices on cultured human neonatal keratinocytes. Using purified fibrinogen and fibrin gels generated by the enzymatic action of thrombin, batroxobin (it leads to retention of fibrinopeptide B), or Agkistrodon contortrix thrombin-like enzyme (ACTE; it leads to retention of fibrinopeptide A), we determined the effect of each of these matrices on keratinocyte morphology, attachment, spreading, and replication as compared to tissue culture plastic. Morphologically, keratinocytes seeded on fibrin surfaces were more rounded and formed three-dimensional structures. Specific cell attachment, as measured at either 37 degrees C or 4 degrees C, was not altered on the different fibrin substrates (P > .05) but was increased on fibrinogen and factor XIII cross-linked fibrin (P < .01). However, keratinocytes seeded on fibrin, regardless of the presence or absence of fibrinopeptides A or B, showed a marked decrease (up to 71%) in cell numbers by days 5 (P = .0357) and 10 (P = .0114). Keratinocyte spreading was decreased by 78.8% (P = .0006), 80.3% (P = .0001), and 89.2% (P = .0001) on thrombin-, batroxobin-, and ACTE-generated fibrin, respectively, but not on fibrinogen-coated dishes. However, either the addition of fibronectin or cross-linking of fibrin with factor XIII allowed full keratinocyte spreading to occur (P = .0002 and P = .0013, respectively). We conclude that fibrin inhibits keratinocyte spreading in the absence of other matrix or plasma proteins or cross-linking by factor XIII.
authors.affiliation: Department of Dermatology and Cutaneous Surgery, University of Miami School of Medicine, Florida, 33136, USA.
authors.forename: E
authors.initials: E
authors.lastname: Weiss
authors.forename: Y
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: A
authors.initials: A
authors.lastname: Falabella
authors.forename: S
authors.initials: S
authors.lastname: Crane
authors.forename: Y
authors.initials: Y
authors.lastname: Tokuda
authors.forename: V
authors.initials: V
authors.lastname: Falanga
mesh_terms: D002448:Cell Adhesion; D002455:Cell Division; D002465:Cell Movement; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D005176:Factor XIII; D005337:Fibrin; D005353:Fibronectins; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism; D008297:Male
chemical_list: D003432:Cross-Linking Reagents; D005353:Fibronectins; D005337:Fibrin; D005176:Factor XIII