Loss of 5hmC immunohistochemical expression is a useful diagnostic aid for distinguishing hepatocellular carcinoma in cytology FNA specimens

OBJECTIVE: Disruption of the DNA methylation pathway involving 5-hydroxymethylcytosine (5hmC) has been implicated in hepatic tumour development. The aim of this study was to investigate the expression of 5hmC in malignant and benign hepatic mass lesions and evaluate the diagnostic utility of deficient 5hmC expression in hepatocellular carcinoma (HCC). METHODS: Our study consisted of 48 fine needle aspiration (FNA) cytological cases (30 cases positive for HCC, 18 cases negative for malignancy) and 39 liver resection specimens (30 HCCs and nine hepatic adenomas [HAs]). A 5hmC immunohistochemistry score (range 0-9) was calculated by multiplying the percentage (0 = no staining; 1 = 1%-10%; 2 = 11%-50%; 3 = 51%-100%) and intensity scores (0-3+). A score of ≤2 was considered deficient for 5hmC. RESULTS: In resection specimens, 5hmC expression was deficient in 90% of HCC cases. Surrounding cirrhotic nodules and 78% of HA cases exhibited intact 5hmC expression. In FNA cytological specimens, expression of 5hmC was deficient in nearly all cases of HCC (96.7%, 29/30 cases). Admixed benign hepatocytes exhibited predominantly intact expression of 5hmC with moderate to strong nuclear staining in the majority of the benign hepatocytes. 5hmC was strongly expressed by endothelial cells, lymphocytes and neutrophils in the background. Many 5hmC-negative HCC clusters were highlighted by characteristic endothelial wrapping with strong 5hmC expression in the endothelial cells. CONCLUSIONS: 5hmC can serve as an important diagnostic tool for the diagnosis of HCC by aiding in the distinction of HCC from HA or cirrhotic nodules in both liver resection and FNA specimens.