Differential production of apoptosis-modulating proteins in patients with hypertrophic burn scar.
BACKGROUND
The biochemical and cellular pathways resulting in the production of proliferative scar in the thermally injured patient remain incompletely elucidated. A promising area of investigation is the phenomenon of programmed cell death and its modulation. The following study was designed to quantify differential levels of the bcl-2 protooncogene and the Fas cell surface receptor, two apoptosis-modulating proteins, in the peripheral blood mononuclear cell (PBMC) fractions of burn patients with hypertrophic scar versus those considered to have healed normally. The study also encompassed an immunohistochemical examination of fibroblasts in vitro, to identify differential levels of Fas, bcl-2, and interleukin converting enzyme (ICE).


METHODS
PBMC fractions were isolated from two matched burn patient cohorts of 10 patients each, the experimental group carrying the clinical and histopathologic diagnosis of hypertrophic burn scar. The supernatant from each mitogenically stimulated specimen was halved and subjected to the Fas/APO-1 enzyme-linked immunosorbent assay (ELISA) and the bcl-2 ELISA. Results for each assay were compared between groups by unpaired t tests. Further biopsy specimens of isolated proliferative scar were used in vitro to analyze the role of these apoptosis-modulating proteins and ICE. This immunoperoxidase technique was analyzed qualitatively.


RESULTS
The expression of the bcl-2 protein in the PBMC fractions of the burn patients with hypertrophic scar is significantly elevated in comparison to the control cohort (307.72 +/- 72.29 u/ml vs 31.55 +/- 6.73 u/ml; P = 0.0042). The quantitative levels of the Fas receptor did not differ significantly between the groups, respectively (0.3988 +/- 0.179 u/ml vs 0.2899 +/- 0.066 u/ml; P = 0.5787). Immunoperoxidase staining of proliferative scar fibroblasts and those from surrounding skin revealed relatively decreased levels of membrane-bound Fas and ICE. bcl-2 was not detectable in these specimens.


CONCLUSIONS
Differential expression of the bcl-2 protooncogene and the Fas cell surface receptor in the PBMC fraction of patients with burn injuries may suggest a disequilibrium in a complex biochemical signaling mechanism mediating programmed cell death. The increased levels of bcl-2 could be responsible for delayed fibroblast apoptosis, resulting in the disruption of normal healing and subsequent hypertrophic scarring. This is confirmed by an in vitro examination of wound fibroblasts versus those from surrounding uninjured skin. This immunoperoxidase technique reveals a localized relative decrease in Fas and ICE, two apoptosis-inducing proteins, at the level of the fibroblast in the proliferative scar specimen.
Wassermann
R J
RJ
Division of Plastic & Reconstructive Surgery, University of South Carolina School of Medicine, Columbia 29203, USA.
Polo
M
M
Smith
P
P
Wang
X
X
Ko
F
F
Robson
M C
MC
D000328:Adult; D017209:Apoptosis; D002056:Burns / Q000150:complications*; D020170:Caspase 1; D017439:Cicatrix, Hypertrophic / Q000209:etiology / Q000378:metabolism*; D015331:Cohort Studies; D003546:Cysteine Endopeptidases / Q000096:biosynthesis; D053222:Fas Ligand Protein; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007124:Immunoenzyme Techniques; D008297:Male; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D014176:Protein Biosynthesis; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis
C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D019253:Proto-Oncogene Proteins c-bcl-2; D003546:Cysteine Endopeptidases; D020170:Caspase 1