Proceedings of the 2023 Division of Translational Toxicology Satellite Symposium.

The 2023 annual Division of Translational Toxicology (DTT) Satellite Symposium, entitled "Pathology Potpourri," was held in Summerlin, Nevada, at the Society of Toxicologic Pathology's 41st annual meeting. The goal of this symposium was to present and discuss challenging diagnostic pathology and/or nomenclature issues. This article presents summaries of the speakers' talks along with select images that were used by the audience for voting and discussion. Various lesions and topics covered during the symposium included induced and spontaneous neoplastic and nonneoplastic lesions in the mouse liver, infectious and proliferative lesions in nonhuman primates, interesting presentations of mononuclear cell infiltrates in various animal models and a complex oral tumor in a rat.

[Lenvatinib down-regulates IGF1R/Mek/Erk signaling pathway in the treatment of regorafenib-resistant hepatocellular carcinoma].

Objective: To investigate the therapeutic effect and mechanism of lenvatinib on regorafenib-resistant hepatocellular carcinoma cells. Methods: CCK-8 and clone formation assay were used to observe the inhibitory effect of lenvatinib on the growth of hepatocellular carcinoma cells. Flow cytometry was used to detect the apoptosis of regorafenib-resistant hepatocellular carcinoma cells treated with lenvatinib. The expression levels of related proteins were detected by western blot and immunohistochemical staining. The inhibitory effect of lenvatinib on the tumor formation ability of regorafenib-resistant hepatocellular carcinoma cells in vivo was observed by subcutaneous tumor formation experiment in mice. Results: CCK-8 and clone formation assay showed that lenvatinib could inhibit the proliferation of regorafenib-resistant hepatocellular carcinoma cells. The number of clones of HepG2, SMMC7721 and regorafenib-resistant HepG2, SMMC7721 cells in lenvatinib group (120.67±11.06, 53.00±11.14, 55.00±9.54, 78.67±14.64) were all lower than those in control group (478.00±24.52, 566.00±27.87, 333.67±7.02, 210.00±12.77, all P<0.05). Flow cytometry showed that lenvatinib could promote apoptosis of regorafenib-resistant hepatocellular carcinoma cells, the apoptosis rates of HepG2, SMMC7721 and regorafenib-resistant HepG2, SMMC7721 cells in lenvatinib group [(12.30±0.70)%, (9.83±0.38)%, (15.90±1.32)%, (10.60±0.00)%] were all higher than those in control group [(7.50±0.87)%, (5.00±1.21)%, (8.10±1.61)%, (7.05±0.78)%, all P<0.05]. The apoptosis-related protein levels suggested that apoptosis was increased in the treatment of lenvatinib. The animal study showed that lenvatinib can inhibit the growth of regorafenib-resistant cells in vivo. Immunohistochemistry and western blot results showed that lenvatinib could down-regulate the abnormally activated IGF1R/Mek/Erk signaling pathway in regorafenib-resistant cells. Conclusion: Lenvatinib can reverse regorafenib resistance in hepatocellular carcinoma, possibly by down-regulating IGF1R/Mek/Erk signaling pathway.

Tissue Inhibitor Of Metalloproteinase 3 As A Potential Staging Biomarker In Hepatocellular Carcinoma.

Objectives: To assessthe potential role of tissue inhibitor of metalloproteinase 3 as a staging marker of hepatocellular carcinoma. Method: The experimental study was conducted at Faculty of Pharmacy, Kafrelsheikh University, Egypt, from December 2020 to March 2022 after approval from the Faculty of Pharmacy, Kafrelsheikh University, Egypt, and comprised male albino rats. The subjects were divided into 4 groups. The control group was administrated a single intraperitoneal injection of normal saline, while the other groups were generated post-induction of hepatocellular carcinoma. The induction was done by injecting rats intraperitoneally with a single dose of 200mg/kg diethyl nitrosamine, followed by the administration of 0.05% phenobarbital sodium in drinking water daily. Rats were euthanised at 8, 16 and 24 weeks after the injection to obtain three groups related to the 3 stages of hepatocellular carcinoma. Serum was used for measuring the alpha protein level. Liver homogenates were used for the assessment of the hepatic tissue inhibitor of metalloproteinase 3 expression, B-cell lymphoma 2-associated X protein expression, and the hepatic content of matrix metalloproteinase -9 and cyclin D1. Data was analysed using Graph Prism 8. RESULTS: Of the 24 rats with weight range 120-130g, 6(25%) were in each of the 4 groups. The relative protein and messenger ribonucleic acid tissue inhibitor of metalloproteinase 3 expressions were significantly decreased in the intervention groups compared to the control group, with more decline as the hepatocellular carcinoma stage increased. The matrix metalloproteinase -9 and cyclin D1 concentrations and the relative hepatic protein Ki67 expression were significantly raised in the intervention groups compared to the control group (p<0.05). The relative expression of hepatic B-cell lymphoma 2-associated X protein was markedly decreased in the intervention groups compared to the control group (p<0.05). CONCLUSIONS: Tissue inhibitor of metalloproteinase 3 might be a promising diagnostic and staging biomarker in hepatocellular carcinoma.

[Urine metabolomics study of hepatocellular carcinoma].

Objective: To investigate the urinary small molecular metabolites and their metabolic characteristics of patients with hepatocellular carcinoma (HCC). Methods: High throughput ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was used to detect the small molecular metabolites in urine of healthy control (n=10), patients with hepatic hemangioma (n=10) and patients with HCC (n=10). The orthogonal projections to latent structures-discriminant analysis (OPLS-DA), hierarchical cluster analysis of multivariate analysis and univariate analysis were used to analyze the differential metabolites of the three groups. Results: The metabolic profiles of the three groups showed that the total of 381 differential metabolites were identified and divided into 96 up-regulated metabolites and 285 down-regulated metabolites. There were 55 urinary metabolites specifically related to HCC. Twenty-one of them were significantly up-regulated, including Acetyl-DL-Leucine, Ala Asp, HoPhe-Gly-OH, while 34 were significantly down-regulated, including Selenocystathionine, Met Trp Met Cys, Valsartan acid and so on. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the differential metabolites were mainly enriched in glutamine/glutamate metabolism, lysine biosynthesis, tricarboxylic acid cycle and purine metabolism. Conclusions: The occurrence of HCC is accompanied by the abnormalities of multiple metabolites and metabolic pathways. The analysis of the characteristic metabolic profile of urine in patients with HCC is helpful to find metabolic markers and potential therapeutic targets for liver cancer.

[The influence of HBsAg expression in liver tissue on the postoperative recurrence of HCC patients].

Objective: To investigate the influence of HBsAg expression in peritumoral tissue of hepatocellular carcinoma (HCC) patients on their postoperative recurrence. Methods: The HCC patients treated in Shanghai Eastern Hepatobiliary Surgery Hospital from October 2009 to August 2010 were selected. The clinicopathological data and adjacent tissues of 718 patients were collected, and dextran polymer immunohistochemical staining was used to detect the expression of HBsAg in adjacent tissues. According to the expression of HBsAg in adjacent tissues, the tissues were divided into HBsAg positive group and HBsAg negative group. Kaplan-Meier method and Log rank test were used for survival analysis, and Cox regression model was used for multivariate analysis. Results: Among the 718 patients in the whole group, 153 were HBsAg negative and 565 were HBsAg positive. There was a statistically significant difference in serum HBV DNA level between HBsAg-positive and HBsAg-negative patients (P<0.001). The number of patients with serum DNA≥2 000 IU/ml and<2 000 IU/ml in HBsAg negative group were 52 and 93, while the patients in HBsAg positive group were 325 and 205. The cumulative recurrence rates of all patients at 1, 3, and 5 years after surgery were 30.2%, 54.3%, and 62.7%, respectively. The expression of HBsAg was related to the recurrence (P=0.038). Multivariate analysis showed that γ-GT, PT, multiple tumors, tumor length, and portal vein invasion were independent risk factors for recurrence of HCC (P<0.05). In HBeAg-negative patients with low viral load (HBV DNA <2 000 IU/ml) and without cirrhosis, the recurrence rates of HBsAg-positive patients were 14.3% and 31.0% at 3 and 5 years, respectively, compared with HBsAg negative patients (all 0), the difference was statistically significant (P=0.021). Conclusion: The positive expression of HBsAg in peritumoral tissue increases the postoperative recurrence risk of HCC patients.

[Relationship of operation manner and postoperative recurrence of hepatocellular carcinoma].

Hepatocellular carcinoma (HCC) owns the high morbidity and mortality rates. Surgical resection is still the major pathway for the longer survival of HCC patients. Postoperative recurrence and metastasis have become the key impairment of prognosis of HCC patients. The relationship between tumor recurrence and surgical manner underwent by HCC patients is complicated and multiple factors are included. When the liver tumor was pressured during operation, tumor cells could be squeezed into blood flow via the broken vessels, which resulted in tumor metastasis. Besides, ischemia-reperfusion injury induced by Pringle maneuver during the liver blood blockade resulted in the immune destruction of liver and induced tumor recurrence.The destruction of physical barriers consisted of interstitial cells and normal liver cells was also a key factor for tumor recurrence. This paper summarizes the possible relationship between postoperative recurrence and surgical manner in HCC patients to provide the preventive suggestions for the postoperative recurrence of HCC patients.

[Advances and controversies of statins application in prevention and treatment of hepatocellular carcinoma].

Statins, as lipid-regulating drugs, have been widely used in the treatment for hyperlipidemia and the primary and secondary prevention of cardio-cerebrovascular diseases. Hepatocellular carcinoma (HCC) is a serious burden of liver disease in China with poor prognosis, thus effective adjuvant drug used for HCC treatment has attracted much attention. Statins can suppress tumor growth, decrease the risk of tumorigenesis and postoperative recurrence of HCC, extend the survival time and improve the therapeutic effect of other treatment, therefore might increase the benefit obtained by the HCC patients. Statins also can impact the expression of MAPK/ERK signaling pathway, promote the apoptosis of malignant cells and ameliorate the HCC risk of hepatitis B virus infected patients. Statins not only prevents the HCC, but also has part therapeutic effect on the different stage of HCC. Although it can't replace the operation, radiofrequency ablation, molecular targeted treatment and immunotherapy currently, statins may be a potential adjuvant drug to provide clinical benefit for HCC patients. The advancement of statins application in the prevention and treatment of HCC has attracted more attention recently, however, discussion and controversy also existed about whether it can eventually become an adjuvant therapy for HCC. The purpose of this paper is to summarize and comment on the new development and disputes of statins application in the prevention and treatment of HCC in recent years, to provide help for the future clinical practice.

[Application of superb microvascular imaging in focal liver lesions].

Objective: To investigate the ability of superb-microvascular imaging (SMI) to detect microvascular characteristics of focal liver lesions (FLLs) and analyze the relationship between vascular index (VI) and microvascular density (MVD) and Ki-67 levels. Methods: The imaging data of patients diagnosed as FLLs at Tianjin Medical University Cancer Hospital in 2018 were collected. A total of 166 FLLs patients were divided into non-hepatocellular liver cancer (non-HCC group, 96) and HCC group (70), respectively. The whole group of patients were subjected to color Doppler blood flow imaging (CDFI) and SMI examination. The patient's Adler's semi-quantitative grading (0 to 3 levels) and vascular morphological characteristics (a-f type) were analyzed. The receiver operating characteristic (ROC) curve was used to evaluate the detection ability of HCC with SMI and CDFI blood flow characteristics, The Pearson correlation analysis was used to evaluate the correlation between HCC patients VI and MVD and the Spearman correlation analysis was used to evaluate the correlation between VI and Ki-67. Results: In HCC group, SMI detected 50 cases of high-level blood flow (Adler's semi-quantitative grade 2 to 3) patients, higher than 22 cases of CDFI (P=0.033). In HCC group, SMI detected 52 cases of blood-rich mode (e, f type), higher than 18 cases of CDFI (P<0.001). In non-HCC group, the difference of blood flow characteristics detection between CDFI and SMI was not statistically significant. In HCC group, SMI detected 52 cases of rich blood supply patterns, which was higher than 14 cases of non-HCC group (P<0.001). The area under the ROC curve of SMI was 0.760 (sensitivity was 74.3%, specificity was 85.4%), and the SMI rich blood supply mode had the best diagnostic effect on HCC based on the blood-rich mode as the HCC diagnostic standard. In HCC group, VI was positively correlated with MVD and Ki-67 (r=0.698 and r=0.669, respectively, P<0.05). Conclusions: SMI has better detection ability than CDFI for HCC microvascular characteristics, HCC has more blood-rich mode than non-HCC. In HCC, VI is positively correlated with MVD and Ki-67 expression levels.

[Effect and mechanism of siRNA targeting α-enolase gene combined with paclitaxel on proliferation, invasion and apoptosis of hepatocellular carcinoma cell].

Objective: To investigate the effect of siRNA targeting inhibition of α-enolase (ENO1) combined with paclitaxel on the proliferation, invasion and apoptosis of hepatocellular carcinoma SK-HEP-1 cell and its mechanism. Methods: siRNA-ENO1 (siRNA-ENO1 group) and siRNA-negative control (siRNA-NC group) were transfected into SK-HEP-1 cells in vitro, the untransfected SK-HEP-1 cells were used as the control group, and the transfection effect was detected by real-time fluorescent quantitative polymerase chain reaction and western blotting. After SK-HEP-1 cells were treated with 0, 2.5, 5, 10, 20 and 40 µg/L paclitaxel for 48 hours, the cell survival rate was measured by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT) method and the semi inhibitory concentration of paclitaxel was calculated. SK-HEP-1 cells transfected with siRNA-ENO1 or siRNA-NC were treated with 10 µg/L paclitaxel as paclitaxel+ siRNA-ENO1 group and paclitaxel+ siRNA-NC group. The proliferation, clonogenesis, invasion and apoptosis of siRNA-NC group, siRNA-ENO1 group, paclitaxel+ siRNA-ENO1 group and paclitaxel+ siRNA-NC group were detected by MTT, clonogenesis, Transwell chamber and flow cytometry respectively. The expression levels of the phosphorylation of phosphatidylinositol-3-kinase (p-PI3K), p-protein kinase B (Akt) and proliferating cell nuclear antigen (PCNA), matrix metalloproteinase 9 (MMP-9) and B lymphocytoma-2 gene (Bcl-2) were detected by western blotting. Results: Compared with the control group (1.00±0.00 and 0.69±0.04, respectively), the expression levels of ENO1 mRNA and protein (0.25±0.03 and 0.23±0.02, respectively) in siRNA-ENO1 group decreased significantly (P<0.05), but there were no significant differences in the expression levels of ENO1 mRNA and protein in siRNA-NC group (P>0.05). Compared without treatment group [(100.00±0.00)%, P<0.05], the survival rates of SK-HEP-1 cells treated with 2.5, 5, 10, 20 and 40 µg/L paclitaxel [(88.65±6.46)%, (72.36±6.08)%, (60.48±4.23)%, (38.52±3.56)% and (20.75±2.32)%, respectively] decreased significantly (P<0.05), and the semi inhibitory concentration of paclitaxel was 13.26 µg/L. The cell survival rate and clone formation rate of siRNA-ENO1 group [(68.86±5.12)% and (18.12±2.25)%, respectively] were lower than those of siRNA-NC group [(100.00±0.00)% and (29.65±3.06)%, respectively, P<0.05]. The cell survival rate and clone formation rate of the paclitaxel+ siRNA-ENO1 group [(43.28±2.64)% and (8.72±0.52)%, respectively] were significantly different from those of the paclitaxel+ siRNA-NC group [(61.75±5.06)% and (13.48±2.16)%, respectively, P<0.05] and siRNA-ENO1 groups [(68.86±5.12)% and (18.12±2.25)%, respectively, P<0.05]. Cell invasion number in paclitaxel+ siRNA-ENO1 group (23.64±2.12) was lower than that in siRNA-ENO1 group and paclitaxel+ siRNA-NC group (42.16±2.75 and 37.35±2.42, respectively, P<0.05). The apoptosis rates of paclitaxel+ siRNA-NC group and siRNA-ENO1 group [(17.49±1.35)% and (15.29±1.50)%, respectively] were higher than that of siRNA-NC group [(7.21±0.70)%, P<0.05]. The apoptosis rate in the paclitaxel+ siRNA-ENO1 group [(24.59±2.40)%] was higher than those in the paclitaxel+ siRNA-NC group and siRNA-ENO1 group [(17.49±1.35)% and (15.29±1.50)%, respectively, P<0.05]. The expression levels of ENO1, PI3K/Akt signaling pathway related proteins including p-PI3K and p-Akt and the expression levels of PCNA, MMP-9 and Bcl-2 in siRNA-ENO1 group and paclitaxel+ siRNA-NC group were lower than those in siRNA-NC group (P<0.05). The expression levels of ENO1, p-PI3K, p-Akt, PCNA, MMP-9 and Bcl-2 in paclitaxel+ siRNA-ENO1 group were lower than those in siRNA-ENO1 group or paclitaxel+ siRNA-NC group (P<0.05). Conclusion: siRNA targeting inhibition of ENO1 expression can enhance the inhibitory effect of paclitaxel on proliferation, invasion and apoptosis of SK-HEP-1 cells, and its mechanism may be related to the inhibition of PI3K/AKT signaling pathway.

[Effect of ultrasound contrast agent targeting gelatin on uptake of mouse ascites hepatocellular carcinoma cell lines with high lymphatic metastasis].

Objective: To analyze the effect of ultrasound contrast agent targeting gelatin on uptake of high lymphatic metastasis cell lines of mouse hepatocellular carcinoma with peritoneal effusion. Methods: The modified double emulsifying solvent evaporation method was used to construct the macromolecule contrast agent PLGA-Cooh. The carbodiimide was used to connect the monoclonal antibody of gelatin with the contrast agent PLGA-Cooh, and the targeted ultrasound contrast agent Gsn-PLGA was established. The particle size and Zeta potential of the targeted ultrasound contrast agent were measured by laser particle size analyzer. The surface binding of the contrast agent to the gelatin monoclonal antibody was evaluated by immunofluorescence. Hca-F cells with high lymphatic metastasis were cultured in mice with peritoneal effusion hepatocellular carcinoma. Target-seeking ability in vitro was evaluated by in vitro uptake test, and the imaging effect of the contrast agent in vitro was evaluated by in vitro developing test. Results: The contrast agent is white powder with good water solubility. The average particle size and surface potential were (569.68±6.96) nm and (-10.95±2.43) mV, respectively. The fluorescent antibody binding rate of non-targeted and targeted ultrasound contrast agent labeled with DiI were 0.84% and 95.89%, respectively. The results showed that the targeted ultrasound contrast agent Gsn-PLGA had a better of developing effect in vitro. Hca-F cells with high expression of gelsolin protein had stronger uptake ability of targeted ultrasound contrast agent and stronger green fluorescence in vitro than those with low expression of gelsolin protein (P<0.05). Moreover, targeted ultrasound contrast agent Gsn-PLGA had stronger targeting to the gelsolin protein. The echo of the targeted ultrasound contrast agent Gsn-PLGA was uniform and fine, without attenuating echo of the back. Simultaneously, the development effect was more obvious with the increase of contrast agent concentration (P<0.05). Conclusion: Ultrasound contrast agent Gsn-PLGA targeting gelatin can bind Hca-F cells with high expression of gelatin and display a good imaging effect in vitro.

[The prognostic analysis of hepatocellular carcinoma based on the tumor neo-vessels, macrophages and α-SMA in tumor microenvironment].

Objective: To analyze the quantitative expression and prognostic significance of tumor neo-vessels, macrophages and fibroblasts in tumor microenvironment of hepatocellular carcinoma (HCC). Methods: The clinic-pathological features and tissue samples for 101 HCC cases were collected. Immunohistochemistry was used to stain the tumor neo-vessels, macrophages and fibroblasts on tumor tissue. The distribution results and quantitative data of above key components were acquired by inverted microscopy equipped with CRi Nuance multispectral analysis system. The number of tumor neo-vessels and macrophages on HCC tissue were counted and the thickness of cancer stroma based on the expression of fibroblasts was measured. The clinic-pathological characteristics and overall survival were analyzed. Results: The median disease free survival (DFS) of 101 HCC cases was 5 month. The quantitative analysis of tumor neo-vessels, macrophages and fibroblasts showed that the expression range was 51-429 with median 218, 110-555 with median 259, 35.61-555.35 with median 246.98, respectively. To take the median as cutoff, all the cases could be classified into high and low expression group. The survival analysis demonstrated that the high density group of macrophages (P=0.022) and fibroblasts (P<0.001) has shorter DFS than low density group, with statistical significance. The high tumor neo-vessels group has shorter DFS with median 5 month than low density group with median 7 month. However, there was no statistical significance between these two group (P=0.197). Combined with above the three stromal components, all the cases could be classified into low, middle and high group. The survival analysis demonstrated that the high density group of stromal components has shorter DFS than the other two groups with median 3 month (P=0.001). Multivariate analysis by Cox regression indicated that cirrhosis, metastasis status, macrophages and fibroblasts density were the independent prognostic factors. Conclusion: The key elements in tumor microenvironment including tumor neo-vessels, macrophages and fibroblasts were heterogenic in HCC tissues and played significant roles in HCC invasion and metastasis.

Hepatocellular neoplasms arising in genetic metabolic disorders: steatosis is common in both the tumor and background liver.

Hepatocellular neoplasms can develop in multiple genetic metabolic disorders. While there have been rare case reports, clinical and pathological characterizations have not been systematically performed. We conducted a retrospective study in 9 patients with these rare genetic metabolic disorders, including glycogen storage disease type 1, ornithine carbamyl transferase deficiency, hereditary tyrosinemia type 1, and Navajo neurohepatopathy, who developed hepatocellular neoplasms. Our results show that steatosis is a common finding in both tumor (6/9 cases, 67%) and background liver parenchyma (8/9 cases, 89%), underlying a possible role for steatosis in tumorigenesis in these genetic metabolic disorders. Our findings also raise a consideration of underlying genetic metabolic disorder when young patients with hepatocellular neoplasm show steatosis in both the tumor and background liver.

The heterogenic tumor microenvironment of hepatocellular carcinoma and prognostic analysis based on tumor neo-vessels, macrophages and α-SMA.

The present study was performed to quantify tumor neo-vessels, macrophages and fibroblasts in the tumor microenvironment of hepatocellular carcinoma (HCC) and explore the prognostic factors of HCC. The distribution of tumor neo-vessels, macrophages and fibroblasts was quantified by immunohistochemistry and inverted microscopy with the CRi Nuance multispectral imaging system, and the correlation of these parameters with the clinico-pathological characteristics and overall survival of the patients was analyzed. The number of tumor neo-vessels and macrophages, and density of the fibroblasts, as calculated by the thickness of the tumor stroma in the tumor microenvironment, ranged from 51-429 (median, 218), 110-555 (median, 259) and 35.6-555.5 µm (median, 247.0), respectively. Using the median values as a cutoff, the cases were stratified into high- and low-density groups. Survival analysis demonstrated that the high-density groups regarding macrophages (χ2=5.249, P=0.022) and fibroblasts (χ2=18.073, P<0.001) had a significantly shorter disease-free survival (DFS) than the low-density groups. The high-density tumor neo-vessel group had a shorter DFS with a median of 5 months than the low-density group with a median of 7 months; however, there was no statistical significance between these two groups (χ2=1.663, P=0.197). Regarding the above three stromal components combined, all of the cases were classified into low-, middle- and high-density groups. Survival analysis demonstrated that the high-density group of stromal components had a shorter DFS than the other two groups with a median of 3 months (χ2=14.439, P=0.001). Multivariate analysis by Cox regression indicated that cirrhosis, metastasis stage, as well as macrophage and fibroblast density were independent prognostic factors. In conclusion, the key elements in the tumor microenvironment, including tumor neo-vessels, macrophages and fibroblasts, were heterogenic in HCC tissues and have significant roles in HCC invasion and metastasis. Stromal components are associated with the prognosis of patients with HCC; the higher the density of stromal components, the poorer the prognosis of patients with HCC.

Contrast-enhanced ultrasonographic findings of serum amyloid A-positive hepatocellular neoplasm: Does hepatocellular adenoma arise in cirrhotic liver?

Hepatocellular adenoma (HCA) was recently classified into four pathological subtypes. There have been few studies describing the findings of contrast-enhanced ultrasonography (CEUS) of each type. Our case concerns a 78-year-old man who had undergone routine medical check-ups for hepatitis C for 11 years. Abdominal ultrasonography showed a 28 mm, hypo-echoic mass in the segment 4 of the liver. His integrating amount of drinking was 670 kg convert into ethanol. CEUS with Sonazoid demonstrated mild uniform hypo-enhancement with inflow of microbubbles from the periphery of the tumor in the arterial phase, and heterogeneously hypo-enhancement in the post vascular phase. Because the mass increased in size within 3 mo, a well differentiated hepatocellular carcinoma was suspected, and hepatic resection was performed. Microscopic findings showed homogeneous cell proliferation with low grade atypia, infiltration of inflammatory cells, ductular reactions, fatty deposit in part, and sinusoidal dilation. Immunohistochemistry revealed geographic positive for serum amyloid A (SAA), focal positive for glutamine synthetase, diffuse and strong positive for C-reactive protein, and positive for liver-type fatty acid binding protein. These pathological features corresponded to that of an inflammatory HCA. However, we could not make a clear diagnosis, because HCAs were defined as not to arise in cirrhotic liver. Finally, this tumor was diagnosed as a SAA positive hepatocellular neoplasm.

Role of biopsy sampling for diagnosis of early and progressed hepatocellular carcinoma.

The current guidelines for the diagnosis of hepatocellular carcinoma (HCC) recommend liver biopsy for hepatic nodules which do not demonstrate the typical features of HCC on imaging. Thus, while not all HCCs are biopsied for histological confirmation, the nodules that pathologists now encounter on biopsy specimens are frequently well-differentiated early HCCs. This paper reviews the pathological features of HCC and its precursor lesions on liver biopsy specimens, with special emphasis on the differential diagnosis between well-differentiated HCCs and high-grade dysplastic nodules, and discusses the different roles of liver biopsy in diagnosis and management of early and progressed HCC. The potential role of liver biopsy for the development of molecular markers to predict prognosis and response to targeted therapy is also discussed.