Despite many clinical aspects are taken into consideration for the diagnosis of HCC, histological examination still represents the gold standard. However, well differentiated forms of HCC can be difficult to separate histomorphologically from benign lesions such as hepatocellular adenoma (HCA), particularly in small biopsies [4,5]. Because distinction between HCC and HCA is vitally important in determining appropriate therapy and assessing prognosis, many attempts have been made using immunohistochemical stains to aid in such differential diagnosis [6]. However, the clinical application and reproducibility of these immunostains still need to be further investigated [7].

In this study, the AMACR protein was detected in the cytoplasm of normal hepatocytes, and the staining were either scant faint (background staining) or weakly positive. These findings are consistent with results previously published [13]. We demonstrated that AMACR expression is significantly increased in the HCC compared with HCA, CN and NLT with respect to the intensity of immunostaining. The high expression of AMACR was found in 82% (42/51) of HCC, while only 11% (1/9) of HCA, 13% (6/48) of CN and 6% (1/16) of NLT showed high expression for AMACR. These findings suggest that AMACR may be involved in the hepatocarcinogenesis. AMACR is important in β-oxidation of branched-chain fatty acids. From pathogenesis point of view, there is a possibility that a molecular link exists between this metabolic fatty acid enzyme and HCC. Experimental studies showed that overexpression of acyl-CoA oxidase, regulated by AMACR, can transform cells in vitro [19].

Separation of HCC from HCA, in particular the well-differentiated variant, is a difficult challenge for pathologists. Although helpful morphological criteria have been worked out, there are still cases that cannot be reliably resolved in routine practice [4]. Clinical parameters such as sex, age, history of steroids, hepatitis or level of alpha-fetoprotein may give indications, but not proof, in the differential diagnosis. Since methods of treatment such as embolization and surgery, are different for HCA and HCC, an early correct diagnosis is of major importance. Therefore, it is critical to develop relevant sensitive markers to assist in making an accurate diagnosis. We assessed the potential diagnostic utility of AMACR in HCC. In our study, high expression of AMACR was found in 82% of well-differentiated HCC. In contrast, only 11% of HCA showed high expression for AMACR. These results suggest that AMACR may serve as a useful marker to facilitate accurate diagnosis in specific settings, particularly in distinguishing well-differentiated HCC from HCA given that high AMACR expression is at much lower frequency in HCA. Results may warrant further studies using large volume samples to evaluate the value of AMACR as a diagnostic marker in HCC.

Analysis of AMACR expression in relation to clinicopathological features showed that high AMACR in the HCC was significantly associated with venous invasion. This data suggests an important role of AMACR in tumor invasiveness and progression of HCC. Venous invasion is one of the most important pathological features that lead to postoperative tumor recurrence after resection of HCC [20,21]. The exact mechanism of venous invasion in HCC remains unclear, but active neovascularization of the tumor is likely to play an important role. It is not clear whether and to what extent AMACR is involved in angiogenesis during HCC progression. Further studies are needed to clarify the mechanisms.