—  SPECIALTY CONFERENCE  —

Liver Pathology

Case 5 - Well Differentiated Hepatocellular Carcinoma with Persistent Portal Blood Supply

John Hart
University of Chicago
Chicago, IL


Click on each slide thumbnail image for an enlarged view
Case History
This 57-year-old male was diagnosed with chronic HBV hepatitis in 1991. Laboratory evaluation at that time revealed a positive serum HBsAg test but a negative HBeAg test. A liver biopsy in 1994 documented the presence of cirrhosis. An abdominal ultrasound examination and CT scan performed in 1998 revealed two nodules that were more than 4 cm in greatest dimension, as well as several smaller lesions. None of the lesions was regarded as radiographically diagnostic of hepatocellular carcinoma. The serum AFP at that time was 30 ng/ml. A percutaneous needle biopsy of the largest nodule was read as atypical macroregenerative nodule (borderline lesion). The patient was treated with Lamivudine and followed with serial MRI examinations and serum AFP levels. The two large lesions remained essentially unchanged and the serum AFP level remained stable.

On 4/3/01 the patient underwent orthotopic liver transplantation. Examination of the explanted native liver revealed cirrhosis with six distinct nodules, including 6.0 cm and 5.0 cm masses. Sections revealed that two of the smaller lesions (3.5 and 3 cm) were well-differentiated hepatocellular carcinomas. There were also two macroregenerative nodules. Sections of the two largest lesions were more difficult to precisely classify. A section from the largest mass is submitted for your review. The patient has had no evidence of recurrent hepatocellular carcinoma in the post-transplant period, with last follow-up on 3/6/05.


Case 5 - Figure 1 - Interface between largest nodule and adjacent liver.
Case 5 - Figure 2 - Reticulin stain showing interface between nodule and adjacent liver.
Case 5 - Figure 3 - Medium power view of nodule.



Case 5 - Figure 4 - Non-triadal vessels within nodule.
Case 5 - Figure 5 - Vessels within nodule
Case 5 - Figure 6 - Reticulin stain of nodule



Case 5 - Figure 7 - Cytoplasmic inclusions within hepatocytes
Case 5 - Figure 8 - Inflammatory infiltrate in adjacent liver
Case 5 - Figure 9 - Additional section of nodule


Radiographic Findings
The radiographic diagnosis of hepatocellular carcinoma takes advantage of the hypervascular arterial supply of these tumors. In the arterial phase of the examination the contrast agent injected into the arterial blood brightly delineates the tumor. During the venous phase of the examination the tumor appears dark compared to the surrounding cirrhotic parenchyma, which now is highlighted by the contrast agent. This distinctive radiographic appearance is regarded as diagnostic of hepatocellular carcinoma and obviates the need for biopsy confirmation if the lesion is greater than 2 cm in diameter [1, 2] .

Four MRI examinations were performed in the patient under discussion between 1998 and 2001. They revealed two large lesions that grew very slowly, if at all. Neither was clearly evident in the arterial phase images; instead they were best seen in the subsequent venous phase portions of the examinations. In the last MRI study two new smaller lesions were identified that were hypervascular, being seen best in the arterial phase images. Soon afterward the patient underwent liver transplantation.

Pathologic Features
Gross examination of the explanted native liver revealed established cirrhosis. Sections exposed six distinct nodules, including 6 and 5 cm masses that corresponded to the hypovascular lesions seen in the MRI studies. These two lesions were grossly distinctive in that they were quite homogenous, and were demarcated by unusually thin capsules given their large sizes. In contrast, the smaller hypervascular lesions were grossly typical hepatocellular carcinomas, with variegated cut surfaces and thick capsules.

Sections of the large lesions revealed that they were composed of cytologically bland and monotonous hepatocytes with minimal architectural atypia. The reticulin stains reveal no thickening of the trabecular cords and no loss of reticulin. There is no evidence of stromal or vascular invasion. There are, however, scattered foci of small cell change and pseudo-acinar change. Steatosis and clear cell change were not evident. Interestingly, unpaired arteries are notably sparse. In contrast, sections of the smaller tumors reveal significant cytologic and architectural distortion, with loss of reticulin, thickening of trabecular cords, and numerous unpaired arteries.

Discussion
It is now well established that hepatocellular carcinomas in the setting of cirrhosis arise from pre-existing dysplastic nodules with characteristic histologic features. These features well elucidated in careful morphologic studies performed by Japanese investigators of small nodules detected by radiographic studies performed as part of surveillance programs of high risk patient populations. In the early studies the histologic findings thought to indicate future malignant potential included: increased cell density, microacinar formation, cytoplasmic basophilia, clear cell change, and steatosis [3, 4, 5, 6, 7, 8] . The malignant potential of these lesions were confirmed in studies of explanted cirrhotic livers [9, 10, 11] . Standardized criteria and nomenclature to describe these borderline nodules were proposed in a 1993 article that brought the subject to the attention of Western surgical pathologists [12].

Subsequent improvements in the radiographic identification of hepatocellular carcinomas centered upon distinction from benign and premalignant lesions based on the vascular supply to the nodule [13]. This in turn led to the histologic evaluation of the number and type of vessels within these nodules. The arterial supply to dysplastic nodules was indeed increased when compared to ordinary cirrhotic nodules and macroregenerative nodules, in the form of unpaired ("isolated"} arterioles [14, 15, 16, 17, 18, 19] . Isolated arterioles were incorporated as a diagnostic feature in an international consensus classification scheme of hepatocellular nodules in cirrhotic livers [20]. This system remains in wide use, but the authors are currently at work on a revised system that is more useful in the diagnosis of needle biopsies [21]. Alteration in the phenotype of sinusoidal endothelial cells was also identified during hepatic carcinogenesis, a process termed "capillarization". The endothelial cell fenestrae are lost and basement membrane is deposited, producing an appearance similar to the endothelial cells lining systemic arteries [22, 23, 24, 25, 26] . The altered phenotype can be easily documented by use of the CD34 immunostain. Normal sinusoidal endothelial cells are negative with this antibody, while an increasing number of cells become reactive in the sequence from cirrhosis to macroregenerative nodule, to low and high grade dysplastic nodule, and finally hepatocellular carcinoma [17, 18] .

Vascular alterations that occur during the process of hepatic carcinogenesis are among the diagnostic markers helpful in the distinction of dysplastic nodules from hepatocellular carcinoma [27, 28, 29] . However, accumulating evidence suggests that neoangiogenesis may also actually be an important driver of the neoplastic process. A number of studies using molecular profiling methods have demonstrated that as a group, angiogenic molecules are among the most commonly differentially expressed during the process of hepatic carcinogenesis [30, 31, 32] . A number of endothelial cell antigens expressed in hepatocellular carcinomas are known to influence members of the cell cycle cascade, formation of tumor stroma, and growth factors [33, 34, 35] .

The mechanisms responsible for neoangiogenesis during hepatic carcinogenesis have yet to be fully elucidated. A recent study has demonstrated macrophages are increased in number in dysplastic nodules and early hepatocellular carcinomas when compared to the surrounding cirrhotic parenchyma [36]. A follow-up study showed that proteins secreted by malignant hepatocytes stimulate the migration of these macrophages. Moreover, macrophages activated by co-culture with hepatocellular carcinoma cells stimulated angiogenesis [37]. There are several reports in the literature that suggest that macrophage migration inhibitory factor may be responsible in part both for the migration of macrophages into hepatocellular carcinoma and for the stimulation of angiogenesis [38, 39, 40] .

A recent study reported the rare occurrence of neoplastic hepatic nodules in which, for unknown reasons, the conversion to an arterial supply doesn't develop [41]. These large nodules grow very slowly and do not develop thick capsules. Morphologically they are remarkably bland and homogeneous, without well-developed features of overt hepatocellular carcinoma. The incidence of such nodules in a surveillance program of patients with chronic HCV and HBV related cirrhosis was 2.6%. The large nodules present in the patient under discussion appear to be examples of this new entity, which was termed "atypical large well-differentiated hepatocellular carcinoma with benign nature" by the author of the study [41]. In the study no nodule developed into a traditional arterially supplied hepatocellular carcinoma. The existence of nodules of this type, however they are classified, provides additional evidence for the central role of the switch to an arterial blood supply in the process of hepatic carcinogenesis. Further study of the mechanisms of angiogenesis in the spectrum of hepatocellular lesions will contribute to the understanding of tumorigenesis.

Diagnosis
Well differentiated Hepatocellular Carcinoma with Persistent Portal Blood Supply

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