Evaluation of Combined p57KIP2 Immunohistochemistry and Fluorescent in situ Hybridization Analysis for Hydatidiform Moles Compared with Genotyping Diagnosis.

Immunostaining with p57KIP2 is a widely used diagnostic technique to differentiate complete hydatidiform moles (CHMs) from partial hydatidiform moles (PHM) and non-molar hydropic abortion. However, distinguishing between PHMs and non-molar hydropic abortions using histopathology alone is often challenging. This study aimed to evaluate the technical validity and additional benefits of using fluorescence in situ hybridization (FISH) in combination with p57KIP2 immunostaining to diagnose molar and non-molar conceptuses. The study involved 80 specimens, which underwent genetic diagnosis using short tandem repeat analysis, including 44 androgenetic CHMs, 20 diandric monogynic PHMs, 14 biparental non-molar hydropic abortions, 1 monoandric digynic triploid abortion, and 1 vaginal specimen of gestational trophoblastic neoplasia. Two pathologists independently diagnosed the cases based on morphology and p57KIP2 immunostaining while the clinical information was masked. FISH analysis was performed using 3 probes (CEP17, CEPX, and CEPY), which revealed that all androgenetic CHM and biparental diploid non-molar hydropic abortion specimens were diploid. Among the 20 diandric monogynic PHM cases examined by analyzing short tandem repeat polymorphisms, 18 were triploid, and the remaining 2 were diploid. These two specimens were possibly androgenetic/biparental mosaics based on FISH analysis, where the three-signal ratios counting 50 cells were clearly within the diploid ranges. Eight of the 20 genetic PHMs and 2 of the 14 genetically confirmed non-molar hydropic abortions that were falsely diagnosed based on morphology and immunohistochemistry by at least 1 pathologist were correctly diagnosed as PHM and non-molar hydropic abortion, respectively, by FISH analysis. However, 1 monoandric digynic villus was classified as triploid by FISH analysis, leading to a false PHM diagnosis. In conclusion, the combination of FISH analysis with p57KIP2 immunostaining helps in diagnosing molar and non-molar conceptuses in numerous cases; nevertheless, exceptional cases should be considered.

ITGB6-Knockout Suppresses Cholangiocarcinoma Cell Migration and Invasion with Declining PODXL2 Expression.

Intrahepatic cholangiocarcinoma (iCCA) is a heterogeneous bile duct cancer with a poor prognosis. Integrin αvß6 (ß6) has been shown to be upregulated in iCCA and is associated with its subclassification and clinicopathological features. In the present study, two ITGB6-knockout HuCCT1 CCA cell lines (ITGB6-ko cells) were established using the clustered regulatory interspaced short palindromic repeats (CRISPR), an associated nuclease 9 (Cas9) system, and single-cell cloning. RNA sequencing analysis, real-time polymerase chain reaction (PCR), and immunofluorescent methods were applied to explore possible downstream factors. ITGB6-ko cells showed significantly decreased expression of integrin ß6 on flow cytometric analysis. Both cell lines exhibited significant inhibition of cell migration and invasion, decreased wound-healing capability, decreased colony formation ability, and cell cycle dysregulation. RNA sequencing and real-time PCR analysis revealed a remarkable decrease in podocalyxin-like protein 2 (PODXL2) expression in ITGB6-ko cells. Colocalization of PODXL2 and integrin ß6 was also observed. S100 calcium-binding protein P and mucin 1, which are associated with CCA subclassification, were downregulated in ITGB6-ko cells. These results describe the successful generation of ITGB6-ko CCA cell clones with decreased migration and invasion and downregulation of PODXL2, suggesting the utility of integrin ß6 as a possible therapeutic target or diagnostic marker candidate.

Application of Immunohistochemistry in the Pathological Diagnosis of Liver Tumors.

Although radiological diagnostics have been progressing, pathological diagnosis remains the most reliable method for diagnosing liver tumors. In some cases, definite pathological diagnosis cannot be obtained by histological evaluation alone, especially when the sample is a small biopsy; in such cases, immunohistochemical staining is very useful. Immunohistochemistry is the most frequently used technique for molecular pathological diagnosis due to its broad application, ease of performance and evaluation, and reasonable cost. The results occasionally reflect specific genetic mutations. The immunohistochemical markers of hepatocellular carcinoma include those of hepatocellular differentiation-such as hepatocyte paraffin 1 and arginase-1-and those of malignant hepatocytes-such as glypican-3, heat shock protein 70, and glutamine synthetase (GS). To classify the subtypes of hepatocellular adenoma, examination of several immunohistochemical markers, such as liver fatty acid-binding protein, GS, and serum amyloid A, is indispensable. Immunohistochemical staining for GS is also important for the diagnosis of focal nodular hyperplasia. The representative immunohistochemical markers of intrahepatic cholangiocarcinoma include cytokeratin (CK) 7 and CK19. In this article, we provide an overview of the application of immunohistochemistry in the pathological diagnosis of liver tumors referring to the association with genetic alterations. Furthermore, we aimed to explain the practical points in the differential diagnosis of liver tumors by immunohistochemical staining.

Combined hepatocellular-cholangiocarcinoma with angiosarcomatoid change: A case report with immunohistochemical study.

Sarcomatoid combined hepatocellular-cholangiocarcinoma (cHCC-CCA) is a rare condition, with only 16 cases reported to date; however, there have been no reports of hepatic sarcomatoid carcinoma with angiosarcomatous features. Here, we report a rare case of cHCC-CCA with angiosarcomatoid changes in a 77-year-old man. The tumor was biphasic with malignant epithelial and mesenchymal components. Histologically, the epithelial component was concordant with classical type cHCC-CCA. The mesenchymal component included angiosarcomatoid cells growing in a vasoformative structure and undifferentiated spindle cells. Immunohistochemical analyses showed that angiosarcomatoid cells were positive for CD31, factor VIII-related antigen, and other angiosarcoma markers. Characteristically diffuse and strong expression of p53 protein was observed in the nuclei of angiosarcomatoid cells but not in carcinoma cells, suggesting that TP53 gene mutations commonly occur in these cells. Transitional zones were observed between HCC and spindle cells and between HCC and angiosarcomatoid cells. A small portion of undifferentiated spindle cells expressed pan-cytokeratin. These findings suggested that this extremely rare tumor developed from dedifferentiation or metaplastic changes of cHCC-CCA.

Clinicopathological investigation of steatohepatitic hepatocellular carcinoma: A multicenter study using immunohistochemical analysis of adenoma-related markers.

AIMS: Steatohepatitic hepatocellular carcinoma (SH-HCC) is a newly proposed concept, which shows histological features of steatohepatitis in HCC lesions, and it is strongly associated with metabolic syndrome (MS) and steatosis/steatohepatitis in non-cancerous lesions. Recently, a substantial number of HCC associated with MS were reported to have developed from pre-existing inflammatory hepatocellular adenoma (HCA). To elucidate the characteristic features of SH-HCC, we clinicopathologically investigated strictly diagnosed SH-HCC and non-SH-HCC (standard HCC). METHODS: This was a retrospective multicenter study. A clinicopathological investigation was undertaken to compare 62 cases with SH-HCC features to 31 age- and sex-matched standard HCC cases, including an immunohistochemical study using markers for classification of HCA and diagnosis of HCC. RESULTS: The characteristic features of SH-HCC compared with standard HCC include a higher rate of complications of MS, more frequent non-alcoholic fatty liver disease as an underlying liver disease, and HCC development in non-cirrhotic liver. The rate of solitary tumors showed no difference between the two groups, but the median diameter of the main tumor was greater in SH-HCCs (45 mm/20 mm, P = 0.01). The HCCs were mostly moderately differentiated, and the patterns were mainly trabecular in both groups. Positive findings for serum amyloid A and C-reactive proteins, classification markers of inflammatory HCA, were significantly higher in cancerous lesions of SH-HCC cases (50%/13%, P < 0.01 and 42%/16%, respectively; P = 0.01). CONCLUSIONS: We confirmed that SH-HCC was strongly associated with MS and NAFLD, and found that classification markers of inflammatory HCA were significantly higher in SH-HCC. Further studies are needed to elucidate the relationship between SH-CCC and HCA for understanding the carcinogenic pathways in these diseases.

ß4 and ß6 Integrin Expression Is Associated with the Subclassification and Clinicopathological Features of Intrahepatic Cholangiocarcinoma.

Intrahepatic cholangiocarcinoma (ICC) is a heterogeneous group of cancers of the intrahepatic biliary tract. However, few studies have evaluated integrin expression according to an ICC subgroup. We immunohistochemically investigated α6ß4 (ß4) and αvß6 (ß6) integrin expressions in 48 ICCs, and evaluated their relationship with clinical and pathological parameters and ligand expression, as well as transforming growth factor (TGF)-ß1. ß4 and ß6 expressions were detected in 46 (96%) and 35 (73%) ICC cases, respectively. We classified ICC into negative, low (ß4, 29 cases; ß6, 36 cases), or high (ß4, 19 cases; ß6, 12 cases) integrin expression groups. ß4 and ß6 integrin levels were higher in the non-peripheral central localization type ICC than in the peripheral localization type; they were also higher in the periductal-infiltrating or intraductal-growth types than in the mass-forming type ICC; lastly, they were higher in the well-differentiated type than in the poorly-differentiated type ICC. High expression was related to bile duct invasion. In addition, ß4 and ß6 expressions were associated with mucin production and the expression of cytoplasmic epithelial membrane antigen, laminin-5, and tenascin-C. TGF-ß1 was correlated with ß6 expression and poor overall survival. These results suggest that integrin expression is associated with subclassification and clinicopathological features of ICC through the coincident expression of their ligands and TGF-ß1.

Clinical and pathological significance of myeloid differentiation factor 88 expression in human hepatocellular carcinoma tissues.

The innate immune system, which includes toll-like receptor (TLR) signaling, plays an important role in inflammation and oncogenesis. Although TLR common adaptor myeloid differentiation factor 88 (MyD88) is known to have multiple effects on carcinogenesis, the role of MyD88 in hepatocarcinogenesis remains unknown. In this study, MyD88 expression was examined in 105 samples of human hepatocellular carcinoma (HCC) tissue by immunohistochemistry, Western blot, and quantitative reverse-transcriptase polymerase chain reaction methods. The relationships between MyD88 expression and clinical and pathological parameters were analyzed. The results showed that attenuated expression of MyD88 in HCC tissue tumor cells was significantly related to hepatitis B virus infection, large tumor size, positive vascular invasion, and intrahepatic metastasis (P < 0.05). Western blot analysis of MyD88 protein in nine normal livers and 28 HCCs showed gender disparity (P < 0.01, P < 0.05), and attenuated expression in cirrhotic livers (P < 0.05). Low expression of MyD88 mRNA was evident in HCCs with vascular invasion (P < 0.01). In contrast to previous reports, these results suggest that attenuated expression of MyD88 in HCC is associated with tumor progression.

Loss of the cell polarity determinant human Discs-large is a novel molecular marker of nodal involvement and poor prognosis in endometrial cancer.

BACKGROUND: Recent Drosophila studies showed that Discs-large (Dlg) is critical for regulation of cell polarity and tissue architecture. We investigated the possibility that loss of the human homologue of Drosophila Dlg (DLG1) is involved in endometrial carcinogenesis. METHODS: We analysed DLG1 expression in 160 endometrial cancers by immunohistochemical staining. Its expression was confirmed by quantitative real-time PCR (RT-PCR). We investigated the roles of DLG1 in growth and invasion by knockdown experiment in endometrial cancer cell lines. RESULTS: Human DLG1 localises at cellular membrane in normal endometrial tissues. Loss of DLG1 was observed in 37 cases (23.1%). Loss of DLG1 was observed in patients with advanced stage and high-grade histology. It was also observed in patients with nodal metastasis, deep myometrial invasion, and negative oestrogen and progesterone receptors. Patients with loss of DLG1 showed poorer overall survival (P=0.0019). Immunohistochemistry data correlated with RT-PCR data. Knockdown of Dlg1 in endometrial cancer cells resulted in accelerated tumour migration and invasion in vitro. CONCLUSIONS: Tissue polarity disturbance because of loss of DLG1 was shown to confer more aggressive characteristics to endometrial cancer cells. Our study revealed that DLG1 expression is a novel molecular biomarker of nodal metastasis, high-grade histology, and poor prognosis in endometrial cancer.

Gadolinium-based Contrast Agent Accumulates in the Brain Even in Subjects without Severe Renal Dysfunction: Evaluation of Autopsy Brain Specimens with Inductively Coupled Plasma Mass Spectroscopy.

PURPOSE: To use inductively coupled plasma mass spectroscopy (ICP-MS) to evaluate gadolinium accumulation in brain tissues, including the dentate nucleus (DN) and globus pallidus (GP), in subjects who received a gadolinium-based contrast agent (GBCA). MATERIALS AND METHODS: Institutional review board approval was obtained for this study. Written informed consent for postmortem investigation was obtained either from the subject prior to his or her death or afterward from the subject's relatives. Brain tissues obtained at autopsy in five subjects who received a linear GBCA (GBCA group) and five subjects with no history of GBCA administration (non-GBCA group) were examined with ICP-MS. Formalin-fixed DN tissue, the inner segment of the GP, cerebellar white matter, the frontal lobe cortex, and frontal lobe white matter were obtained, and their gadolinium concentrations were measured. None of the subjects had received a diagnosis of severely compromised renal function (estimated glomerular filtration rate <45 mL/min/1.73 m(2)) or acute renal failure. Fisher permutation test was used to compare gadolinium concentrations between the two groups and among brain regions. RESULTS: Gadolinium was detected in all specimens in the GBCA agent group (mean, 0.25 µg per gram of brain tissue ± 0.44 [standard deviation]), with significantly higher concentrations in each region (P = .004 vs the non-GBCA group for all regions). In the GBCA group, the DN and GP showed significantly higher gadolinium concentrations (mean, 0.44 µg/g ± 0.63) than other regions (0.12 µg/g ± 0.16) (P = .029). CONCLUSION: Even in subjects without severe renal dysfunction, GBCA administration causes gadolinium accumulation in the brain, especially in the DN and GP.

Clinicopathological characteristics of serum amyloid A-positive hepatocellular neoplasms/nodules arising in alcoholic cirrhosis.

AIMS: To characterize serum amyloid A (SAA)-positive hepatocellular neoplasms/nodules arising in alcoholic cirrhosis, which are detected as hypervascular hepatocellular nodules resembling hepatocellular carcinoma on imaging. METHODS AND RESULTS: Fifty-three hepatocellular nodules were examined with immunostaining for SAA, glutamine synthetase and glypican-3 in 23 patients (four women and 19 men) with alcoholic cirrhosis. Sixteen nodules were examined with magnetic resonance imaging with gadolinium ethoxybenzyl diethylenetriaminepentaacetic acid enhancement (EOB-MRI). Somatic mutations in IL6ST, GNAS and STAT3 were examined in 19 nodules. Thirty-six nodules in 18 patients were diagnosed as SAA-positive hepatocellular neoplasms/nodules, and the remaining 17 nodules in eight patients were SAA-negative focal nodular hyperplasia (FNH)-like nodules. SAA-positive hepatocellular neoplasms/nodules showed significantly more extensive sinusoidal dilatation, inflammatory reaction, abnormally thick arteries and cellular atypia than FNH-like nodules (P < 0.05). Eight SAA-positive hepatocellular neoplasms/nodules (67%) showed slight hypointensity in the hepatobiliary phase on EOB-MRI, whereas all four FNH-like nodules showed iso-intensity (P < 0.05). STAT3 mutations were detected in two of 17 SAA-positive hepatocellular neoplasms/nodules. CONCLUSIONS: This study showed that approximately two-thirds of hypervascular hepatocellular nodules arising in alcoholic cirrhosis were SAA-positive hepatocellular neoplasms/nodules, which show different findings on the EOB-MRI. STAT3 mutations were detected in 11.8% of SAA-positive hepatocellular neoplasms/nodules, supporting a neoplastic nature.

Preserved or enhanced OATP1B3 expression in hepatocellular adenoma subtypes with nuclear accumulation of ß-catenin.

AIM: Recent studies have indicated that hepatocellular adenoma (HCA) is a heterogenous group of benign tumors with various genetic and clinicopathological characteristics. We delineated the clinicopathological characteristics of HCA subtypes and evaluated the expression of transporter protein OATP1B3 in HCA. METHODS: HCA in 34 Japanese patients were investigated immunohistochemically and classified into four subtypes (HNF1α-inactivated type, H-HCA; ß-catenin-activated type, b-HCA; inflammatory type, I-HCA; unclassified type, u-HCA). Immunostaining of OATP1B3 protein in HCA tissue sections was performed to determine the association between OATP1B3 expression and HCA subtypes. RESULTS: HCA was categorized into the following four subtypes and two combined subtypes: 10 H-HCA (29%), 10 I-HCA (29%), seven b-HCA (21%), two b-HCA/H-HCA (6%), two b-HCA/I-HCA (6%) and three u-HCA (9%). The male-to-female ratio was 18:16. Oral contraceptive use was rare but seven HCA were found in patients with glycogen storage disease, congenital absence of the portal vein and idiopathic portal hypertension. OATP1B3 expression was decreased in 24 HCA but was preserved or increased in 10 HCA. All nine HCA with nuclear staining for ß-catenin showed preserved or enhanced OATP1B3 expression, indicating a significant association between nuclear ß-catenin accumulation and OATP1B3 expression in HCA. CONCLUSION: HCA subtype classification was validated in 91% of our Japanese subjects although their clinical backgrounds including rare contraceptive use were different from European subjects. A close association between preserved or enhanced OATP1B3 expression and b-HCA subtype indicated important modalities for clinical decisions in the treatment and follow up of patients with HCA.

A case of ß-catenin-positive hepatocellular adenoma with MR imaging sign of diffuse intratumoral fat deposition.

Hepatocellular adenoma (HCA) is a rare primary benign tumor of the liver, which occurs predominantly in young and middle-aged women. Recently, the subclassification of HCA was proposed by the Bordeaux group. Subsequently, characteristic radiological and clinical features have been revealed in each HCA subtype. According to the previous literature, diffuse intratumoral fat deposition is a very common finding in hepatocyte nuclear factor 1α-negative HCA, but this finding has been reported in ß-catenin-positive HCA in the literature for only one case. In this case report, we report the second case of ß-catenin-positive HCA with MR imaging sign of diffuse intratumoral fat deposition, confirmed immunohistologically on the basis of a surgical specimen. In addition, our case showed hypovascularity and isointensity on the hepatobiliary phase which have been reported as characteristic findings in ß-catenin-positive HCA. Diffuse intratumoral fat deposition can be observed in ß-catenin-positive HCA, which has a greater probability of malignant transformation than other types of HCA.

Stromal miR-21 is more important than miR-21 of tumour cells for the progression of gastric cancer.

BACKGROUND: Gastric cancer (GC) is a common cancer globally. miRNA-21 (miR-21) appears to be important in the tumourigenesis of almost all types of human cancer. However its precise localization in GC has yet to be clarified. We thus examined miR-21 localization in GC and revealed its clinicopathological importance. METHODS: Tissue arrays of 469 GCs from 454 patients were examined for miR-21 using in situ hybridization (ISH). The positivity was evaluated separately in tumour cells and stromal cells. Conventional sections of 10 GCs were also stained. Eight cases were examined by quantitative RT-PCR (qRT-PCR). RESULTS: miR-21 was highly expressed in tumour cells of 44% of cases and in cancer stroma of 51% of cases. miR-21 of tumour cells was not related to clinicopathological factors, whereas stromal miR-21 was related to many factors including tumour stage, size, and nodal metastasis. Stromal miR-21 gradually increased during tumour progression. ISH of whole sections showed stronger stromal positivity in invasive areas with desmoplastic reaction. Cancer stroma also showed higher miR-21 expression than tumour and non-tumourous tissue in the qRT-PCR study. CONCLUSION: Stromal miR-21 is closely related to tumour progression in GC. Stromal miR-21 of tumours might be a target of treatment.

Pathological diagnosis of benign hepatocellular nodular lesions based on the new World Health Organization classification.

There are various types of benign hepatocellular nodular lesions, and their diagnostic criteria were formulated in detail. However, in 2010, the new World Health Organization (WHO) classification introduced immunohistochemical diagnostic criteria for hepatocellular adenoma (HCA) reflecting molecular pathological properties, and HCA was classified into 4 subtypes. These criteria were useful for its differential diagnosis from focal nodular hyperplasia (FNH). They were also useful for the diagnosis of HCA, its subtyping, and differentiation from FNH in Japan. However, the new WHO classification is based on principles that differ from those of conventional definitions of disease concepts and methods for the differential diagnosis. Therefore, it has caused disagreements in the diagnosis in some cases. Based on this background, we present a new perspective on the diagnosis of benign hepatocellular nodular lesions.

Integrins αvß6, α6ß4 and α3ß1 are down-regulated in cholangiolocellular carcinoma but not cholangiocarcinoma.

AIM: The aim of this study was to evaluate integrin expression and immunolocalization of the extracellular matrix proteins in cholangiolocellular carcinoma (CoCC). METHODS: Tissue specimens of 23 CoCC, 28 cholangiocarcinomas (CCC), 42 hepatocellular carcinomas (HCC) and 11 classical type combined hepatocellular cholangiocarcinomas (CHC) were immunostained for ß6, ß4 and α3 integrins, fibronectin and laminin. ITGB6, B4 and A3 mRNA levels in six HCC cell lines, five CCC cell lines and two CHC cell lines were quantified by quantitative reverse transcription polymerase chain reaction. RESULTS: Little or no positivity for ß6, ß4 and α3 integrins was shown in 91%, 91% and 52% of CoCC and 100%, 98% and 81% of HCC, respectively, according to immunostaining, whereas intense positive staining for these integrins was demonstrated in 64%, 96% and 75% of CCC, respectively. There was a close correlation between ß4 and α3 integrin expression and intracytoplasmic laminin in CoCC, CCC and HCC, but not between ß6 expression and its ligand, fibronectin. Integrin mRNA levels were increased in four of five CCC cell lines, but nearly undetectable in five of six HCC cell lines and one CHC cell line. Tubular differentiation of a CHC cell line cultured in collagen gel matrix induced upregulation of these integrins. CONCLUSION: Our results first indicated downregulation of αvß6, α6ß4 and α3ß1 integrins in CoCC, in contrast to its high expression in CCC, suggesting a diagnostic value of integrins in the differential diagnosis of CoCC and CCC, as well as a useful inducible marker of the intermediate features of CoCC.

Discrimination of fibrotic staging of chronic hepatitis C using multiple fibrotic markers.

AIM: In order to evaluate and judge a fibrotic stage of patients with chronic hepatitis C, multivariate regression analysis was performed using multiple fibrotic markers. METHODS: A total of 581 patients from eight hepatology units and institutes were diagnosed by needle biopsy as having chronic liver disease caused by hepatitis C virus. Twenty-three variables and their natural logarithmic transformation were employed in the multivariate analysis. RESULTS: Multivariate regression analysis finally obtained the following function: z = 2.89 × ln (type IV collagen 7S) (ng/mL) - 0.011 × (platelet count) (×10(3) /mm(3) ) + 0.79 × ln (total bilirubin) (mg/dL) + 0.39 × ln (hyaluronic acid) (µg/L) - 1.87. Median values of the fibrotic score of F1 (n = 172), F2 (n = 80), F3 (n = 37) and F4 (n = 16) were calculated as 1.00, 1.45, 2.82 and 3.83, respectively. Multiple regression coefficient and coefficient of determination were 0.56 and 0.320, respectively. Validation with patient data from other institutions demonstrated good reproducibility of the fibrotic score for hepatitis C (FSC), showing 1.10 in F1 (n = 156), 2.35 in F2 (n = 73), 3.16 in F3 (n = 36) and 3.58 in F4 (n = 11). CONCLUSION: A concise multiple regression function using four laboratory parameters successfully predicted pathological fibrotic stage of patients with hepatitis C virus infection.

Focal nodular hyperplasia-like lesion of the liver with focal adenoma features associated with idiopathic portal hypertension.

Great progress has been made in the diagnosis of focal nodular hyperplasia (FNH) and hepatocellular adenoma (HCA) in the last few years due to the use of molecular criteria. This has allowed us to identify a new type of hepatic nodule. In this case report, we present a male patient with a hepatic nodule associated with idiopathic portal hypertension (IPH) pathologically exhibiting not only the morphological features of FNH, such as ductular reactions, dilated sinusoids, major vascular abnormalities and an immunohistochemical "map-like" pattern of glutamine synthetase (GS), but also the immunohistological features of focal HCA, such as strong expression of serum amyloid A and C-reactive protein and weak expression of GS. As the final diagnosis, the nodule was identified as an FNH-like lesion with focal inflammatory hepatocellular adenoma.

Japanese herbal medicines shosaikoto, inchinkoto, and juzentaihoto inhibit high-fat diet-induced nonalcoholic steatohepatitis in db/db mice.

Few studies have investigated the effects of Japanese herbal medicines on nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH). To the best of our knowledge, only one study has examined whether high-fat (HF) diet-fed db/db mice are appropriate animal models of NASH. We investigated the effects of four types of Japanese herbal medicines (shosaikoto (TJ-9), inchinkoto (TJ-135), juzentaihoto (TJ-48), and keishibukuryogan (TJ-25)) on hepatic lesions of HF diet-fed db/db mice. Db/db mice were divided into six groups: control diet (control); HF diet (HF); and HF diet supplemented with TJ-9, TJ-135, TJ-48, or TJ-25 (TJ-9, TJ-135, TJ-48, and TJ-25, respectively). Mice were killed after 6 weeks of treatment, and biochemical and pathological analyses were performed. Mice in the HF group consistently developed histopathological features consistent with definite NASH, and marked necroinflammation occurred. Serum alanine aminotransferase levels in the TJ-9, TJ-135, and TJ-48 groups were significantly improved compared with those in the HF group. With regard to liver histology, TJ-9 and TJ-48 significantly improved lobular inflammation, and TJ-135 significantly improved ballooning degeneration. We have shown that HF diet-fed db/db mice are animal models that correctly recapitulate the histopathology of human NASH and that TJ-9, TJ-135, and TJ-48 inhibit necroinflammatory activity in this model.

Histone acetyltransferase Hbo1 destabilizes estrogen receptor α by ubiquitination and modulates proliferation of breast cancers.

The estrogen receptor (ER) is a key molecule for growth of breast cancers. It has been a successful target for treatment of breast cancers. Elucidation of the ER expression mechanism is of importance for designing therapeutics for ER-positive breast cancers. However, the detailed mechanism of ER stability is still unclear. Here, we report that histone acetyltransferase Hbo1 promotes destabilization of estrogen receptor α (ERα) in breast cancers through lysine 48-linked ubiquitination. The acetyltransferase activity of Hbo1 is linked to its activity for ERα ubiquitination. Depletion of Hbo1 and anti-estrogen treatment displayed a potent growth suppression of breast cancer cell line. Hbo1 modulated transcription by ERα. Mutually exclusive expression of Hbo1 and ERα was observed in roughly half of the human breast tumors examined in the present study. Modulation of ER stability by Hbo1 in breast cancers may provide a novel therapeutic possibility.

Enhanced expression of farnesoid X receptor in human hepatocellular carcinoma.

AIM: The aim of this study was to investigate the expression of farnesoid X receptor (FXR) in human hepatocellular carcinoma (HCC) tissues and cell lines and evaluate its clinicopathological significance. METHODS: Expression levels of FXR protein and mRNA in hepatocytes, hepatic stellate cells (SC), and HCC cells in human liver, HCC tissues and cultured cell lines were analyzed using immunohistochemical methods, western blotting (WB), and quantitative reverse transcription polymerase chain reaction (qRT-PCR). The relationship between FXR expression and clinicopathological parameters was also investigated. RESULTS: Immunoreactivity for FXR was observed in nuclei of hepatocytes, SC and HCC cells. The intensity of nuclear FXR positive staining was comparable or increased in tumor cells of all HCC tissues when compared with hepatocytes of non-tumorous liver tissues of the same patients as well as in comparison with metastatic colon cancers. A significant level of FXR expression in four of six human HCC cell lines was also confirmed, while it was undetectable in three cholangiocarcinoma cell lines. However, FXR protein and mRNA levels in HCC tissues determined by WB and qRT-PCR were lower than those in non-tumorous liver tissues because of the high level of FXR expression in SC nuclei as detected by immunohistochemical double stain. Statistically significant relationships between FXR immunostaining intensity and high Ki-67 labeling indices or a history of transcatheter arterial chemoembolization in HCC patients were also disclosed. CONCLUSION: Contrary to previous reports, preserved or enhanced expression levels of nuclear FXR were detected in HCC, indicating that FXR may play significant roles in the biological behavior of HCC.