title: Comparison of collagen content in skin wounds evaluated by biochemical assay and by computer-aided histomorphometric analysis.
abstract: BACKGROUND
  The quantification of total collagen is of major importance in a wide range of research areas, including the study of cutaneous wound healing and new drugs trials.
  
  
  OBJECTIVE
  The total collagen content in skin biopsies was compared by biochemical hydroxyproline assay and by two computer-aided histomorphometric analyses of histological sections.
  
  
  METHODS
  Two methods were used to evaluate collagen formation: the hydroxyproline assay, as the gold standard and histomorphometric image analysis of the filled areas by corresponding stained collagen fibres, using picrosirius and Gomori's trichrome staining. The image analyses were determined by digital densitometry recognition using computer-aided ImageJ software. One-way ANOVA, simple linear regression and ANCOVA were applied for the statistical analysis and correlation.
  
  
  RESULTS
  In a simple linear regression analysis carried out on the 14th day period after the induction of skin injury, three techniques, picrosirius red (F = 33.57, p = 0.00), Gomori's trichrome (F = 81.61, p = 0.00) and hydroxyproline content (F = 16.85, p = 0.00) were able to detect collagen production. After scale adjustment, there were no significant differences among either the slopes (F = 1.17, p = 0.32) or the intercepts (F = 0.69, p = 0.51) of the estimated regression lines. It seems that a highly significant correlation exists between the histomorphometrical analysis and hydroxyproline assay.
  
  
  CONCLUSIONS
  The morphometric analysis proved to be adequate and can be used as a simple, rapid, low-cost technology for evaluating total collagen in cutaneous wound specimens, compared with the gold standard hydroxyproline assay.
authors.affiliation: a Division of Dermatology, Department of Internal Medicine, Ribeirão Preto School of Medicine , University of São Paulo (USP) , Ribeirão Preto , SP , Brazil.
authors.forename: Guilherme F
authors.initials: GF
authors.lastname: Caetano
authors.affiliation: b Department of Pharmacy , University of Vila Velha , Vila Velha , Espirito Santo , Brazil.
authors.forename: Marcio
authors.initials: M
authors.lastname: Fronza
authors.affiliation: a Division of Dermatology, Department of Internal Medicine, Ribeirão Preto School of Medicine , University of São Paulo (USP) , Ribeirão Preto , SP , Brazil.
authors.forename: Marcel N
authors.initials: MN
authors.lastname: Leite
authors.affiliation: c Department of Biology , University of Vila Velha , Vila Velha , Espirito Santo , Brazil.
authors.forename: Ary
authors.initials: A
authors.lastname: Gomes
authors.affiliation: a Division of Dermatology, Department of Internal Medicine, Ribeirão Preto School of Medicine , University of São Paulo (USP) , Ribeirão Preto , SP , Brazil.
authors.forename: Marco Andrey Cipriani
authors.initials: MA
authors.lastname: Frade
mesh_terms: D000818:Animals; D003094:Collagen / Q000032:analysis*; D006909:Hydroxyproline / Q000032:analysis; D007091:Image Processing, Computer-Assisted; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D012867:Skin / Q000737:chemistry / Q000293:injuries*; D014945:Wound Healing / Q000502:physiology*
chemical_list: D003094:Collagen; D006909:Hydroxyproline
keywords: Cutaneous wound healing; Gomori’s trichrome; hydroxyproline; picrosirius red