Assessment of Changes in Birefringence and Orientation of Collagen Fibres in Different Grades of Oral Submucous Fibrosis and Oral Squamous Cell Carcinoma using Picrosirius Red and Polarising Microscopy

Abstract

Introduction: Oral submucous fibrosis (OSMF) is a potentially malignant disease with a prevalence of 7-13%. It is characterised by the fibroelastic changes due to excessive deposition of collagen which results in dense fibrous bands and epithelial changes. About 90-95% of oral cancers are Oral Squamous Cell Carcinoma (OSCC). Stroma produced by the invading neoplastic cells are rich in collagen fibres. These collagen fibers have been the main focus of study to understand the pathogenesis of these lesions. Hence, these fibres have been evaluated under polarised microscopy following staining with Picrosirius red stain. Aim: To evaluate changes in birefringence, thickness and orientation of collagen fibres in different histopathological stages of OSMF and OSCC. Materials and Methods: This observational study involved sections of clinically and histopathologically confirmed cases of OSCC and OSMF from Department of Oral Pathology and Microbiology, Rajarajeswari Dental College and Hospital, Bangalore. The study was conducted for a duration of one year from April 2017 to March 2018. The study included total 60 cases among which 20 for each-normal mucosa, OSCC and OSMF. Tissue sections were stained with Picrosirius red stain and collagen fibres were analysed for colour, thickness and orientation under polarised microscope. Chi-square test was used for statistical analysis. Significance was set at p-value <0.05. Results: Comparison of birefringence of collagen fibres between OSMF, OSCC and normal mucosa was not statistically significant (p-value=0.37). Orientation of collagen fibres between OSMF, OSCC and normal mucosa was statistically significant with (p-value=0.02). Conclusion: This study showed a change in colour from yellow orange to orange red in advanced OSMF cases which indicated progression of disease and tightly packing of collagen fibres, suggestive of presence of thick fibres in the extracellular matrix. In OSCC, the colour change from yellow orange to orange red and haphazardly arranged collagen fibres was indicative of transformation of preneoplastic to carcinoma stage.