Second harmonic generation microscopy provides accurate automated staging of liver fibrosis in patients with non-alcoholic fatty liver disease

BACKGROUND: Assessment of severity of liver fibrosis is essential in the management of non-alcoholic fatty liver disease (NAFLD). Second Harmonic Generation (SHG) microscopy is a novel optical tissue imaging system that provides automated quantification of fibrosis based on unique architectural features of collagen. This study aims to develop and validate a SHG-based index for automated staging of liver fibrosis in patients with NAFLD. METHODS: SHG microscopy was performed on archived liver biopsy specimens from 83 patients with NAFLD. A unique algorithm was developed to identify specific SHG parameters that correlated with fibrosis stage. The accuracy of the algorithm was compared against clinical assessment by experienced liver histopathologists using the Brunt fibrosis staging and further validated using the leave-one-out cross-validation method. RESULTS: Mean age of the study cohort was 51.8 ± 11.7 years, with 41% males. A fibrosis index (SHG B-index) was developed comprising 14 unique SHG-based collagen parameters that correlated with severity of NAFLD fibrosis in a continuous fashion. The SHG B-index had excellent correlation with Brunt fibrosis stage (Spearman’s correlation 0.820, p<0.001). AUROCs for prediction of Brunt fibrosis stages 1, 2, 3 and 4 were 0.853, 0.967, 0.985 and 0.941 respectively. In the cross-validation analysis, the SHG B-index demonstrated high specificity for diagnosis of all grades of fibrosis. A SHG B-index score of >1.76 had an overall diagnostic accuracy of 98.5% for prediction of presence of bridging fibrosis (Brunt stage ≥3) with sensitivity of 87.5%, specificity 98.0%, positive predictive value 96.6% and negative predictive value 92.6%. CONCLUSION: The SHG B-index is a unique SHG-based index that provides accurate automated assessment of fibrosis stage in NAFLD patients.