Assessment of collagen and elastic fibres in various stages of oral submucous fibrosis using Masson's trichrome, Verhoeff vangieson and picrosirius staining under light and polarizing microscopy

Abstract

Background: Oral Submucous Fibrosis (OSMF) is a collagen related disorder seen in habitual betel-quid chewers. This is a high risk precancerous condition which clinically presents vertical palpable fibrous bands in buccal mucosa, generalized fibrosis of oral soft tissues with restricted mouth opening. Present study was undertaken to ascertain the changes occurring in collagen type I & III and elastic fibres in OSMF in relation to orientation, density and thickness. Study Methods: The study was performed on 15 cases in each group such as Stage I OSMF, Stage II OSMF, Stage III OSMF and normal oral mucosa (NOM). The biopsied samples were routinely processed for paraffin embedding; stained with Hematoxylin and Eosin as well as special stains like Masson's Trichrome (MT), Verhoeff Van Gieson (VVG) and Picrosirius red (PSR) and examined under light and polarized microscope respectively. MT stain demonstrated all types of collagen fibers collectively while PSR stain under polarized microscopy demonstrated collagen type I and type III separately with enhanced birefringence. VVG stain demonstrated very fine black colored elastic fibers, thus changes taken place could be accurately ascertained with progression of the disease. Results: Change in the orientation of collagen type I has been observed in stage II and stage III of OSMF from haphazard to parallel in relation to surface epithelium, while no change was noted in type III collagen fibres as well as elastic fibres in any stage of OSMF, and remained haphazardly arranged. Increased density of type I collagen was observed with increasing stage of OSMF from moderately dense to dense, while decrease in density of type III collagen was noticed in stage III than in stage II OSMF from moderately dense to sparse. The density of elastic fibres was decreasing from dense in stage I to sparse in stage III. Thickness of collagen type I was increasing with increasing grades while type III collagen and elastic fibres remained unchanged. Conclusion: The alterations in orientation, density and thickness of collagen fibres and density of elastic fibres in various grades of OSMF contribute to the clinical presentation of trismus with progression of the disease. Key Words: Collagen type I, Collagen type III, Elastic fibres, Masson's trichrome stain, Verhoeff Van Gieson stain, Picrosirius red stain