[Construction of dermal skeleton by double cross-linking with glutaraldehyde and ultraviolet radiation].
OBJECTIVE
To investigate the effects of preemptive freezing with different temperature and cross-linking methods on the ultrastructure of collagen membrane and its influence on human fibroblast proliferation.


METHODS
Bovine collagen type I solution in concentration of 10 g/L was preliminarily frozen at -20 degrees C or - 80 degrees C for 12 hours, and lyophilized at -70 degrees C for 48 hours. The diameter of apertures in collagen membranes prepared with two different preliminary temperatures were observed by scanning electron microscope (SEM) and compared. The preliminary freezing temperature of - 80 degrees C was used for the following study. The apertures of collagen membrane performed with cross-linking glutaraldehyde and ultraviolet (UV) radiation cross-linking with glutaraldehyde (double cross - linking) after preliminary freezing were also compared. The proliferation of human fibroblasts inoculated in above cross-linking collagens were assessed by MTT assay, in terms of absorption value.


RESULTS
The mean diameter of apertures of collagen membrane pre-frozen at -20 degrees C was (172 +/- 374 microm, while that at -80 degrees C was (99 +/- 24) microm. The apertures of collagen membrane were reduced in size after glutaraldehyde cross-linking, while those of double cross-linking showed no change in size. There was obvious difference in absorption value of fibroblasts 8 days after seeding between above two cross-linking methods (1.534 +/- 0.013 for glutaraldehyde cross-linking, 3.778 +/- 0.010 for double cross-linking, P < 0.05).


CONCLUSIONS
The collagen membrane after preliminary freezing at - 80 degrees C and double cross-linking with UV radiation and glutaraldehyde may be used as a dermal skeleton substitute.
Gu
Hua
H
Department of Dermatology, First Affiliated Hospital of Kunming Medical College, Kunming 650031, PR China.
He
Li
L
Liu
Ling
L
Jin
Yi-Chao
YC
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D018920:Coculture Techniques; D003094:Collagen / Q000648:ultrastructure*; D003432:Cross-Linking Reagents / Q000737:chemistry; D020405:Dermis / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000528:radiation effects; D005615:Freezing; D005976:Glutaral / Q000737:chemistry*; D006801:Humans; D017768:Skin, Artificial; D054457:Tissue Scaffolds; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D003094:Collagen; D005976:Glutaral