Rescue of perfluorooctanesulfonate (PFOS)-mediated Sertoli cell injury by overexpression of gap junction protein connexin 43

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Granulin-epithelin precursor is an oncofetal protein defining hepatic cancer stem cells

Background and Aims: Increasing evidence has suggested that hepatocellular carcinoma (HCC) might originate from a distinct subpopulation called cancer stem cells (CSCs), which are responsible for the limited efficacy of conventional therapies. We have previously demonstrated that granulin-epithelin precursor (GEP), a pluripotent growth factor, is upregulated in HCC but not in the adjacent non-tumor, and that GEP is a potential therapeutic target for HCC. Here, we characterized its expression pattern and stem cell properties in fetal and cancerous livers. Methods: Protein expression of GEP in fetal and adult livers was examined in human and mouse models by immunohistochemical staining and flow cytometry. Liver cancer cell lines, isolated based on their GEP and/or ATP-dependent binding cassette (ABC) drug transporter ABCB5 expression, were evaluated for hepatic CSC properties in terms of colony formation, chemoresistance and tumorigenicity. Results: We demonstrated that GEP was a hepatic oncofetal protein that expressed in the fetal livers, but not in the normal adult livers. Importantly, GEP+ fetal liver cells co-expressed the embryonic stem (ES) cell-related signaling molecules including β-catenin, Oct4, Nanog, Sox2 and DLK1, and also hepatic CSC-markers CD133, EpCAM and ABCB5. Phenotypic characterization in HCC clinical specimens and cell lines revealed that GEP+ cancer cells co-expressed these stem cell markers similarly as the GEP+ fetal liver cells. Furthermore, GEP was shown to regulate the expression of ES cell-related signaling molecules β-catenin, Oct4, Nanog, and Sox2. Isolated GEP high cancer cells showed enhanced colony formation ability and chemoresistance when compared with the GEP low counterparts. Co-expression of GEP and ABCB5 better defined the CSC populations with enhanced tumorigenic ability in immunocompromised mice. Conclusions: Our findings demonstrate that GEP is a hepatic oncofetal protein regulating ES cell-related signaling molecules. Co-expression of GEP and ABCB5 further enriches a subpopulation with enhanced CSC properties. The current data provide new insight into the therapeutic strategy. © 2011 Cheung et al.published_or_final_versio

Identification and characterization of tropomyosin 3 associated with granulin-epithelin precursor in human hepatocellular carcinoma

Background and Aim: Granulin-epithelin precursor (GEP) has previously been reported to control cancer growth, invasion, chemo-resistance, and served as novel therapeutic target for cancer treatment. However, the nature and characteristics of GEP interacting partner remain unclear. The present study aims to identify and characterize the novel predominant interacting partner of GEP using co-immunoprecipitation and mass spectrometry. Methods and Results: Specific anti-GEP monoclonal antibody was used to capture GEP and its interacting partner from the protein extract of the liver cancer cells Hep3B. The precipitated proteins were analyzed by SDS-PAGE, followed by mass spectrometry and the protein identity was demonstrated to be tropomyosin 3 (TPM3). The interaction has been validated in additional cell models using anti-TPM3 antibody and immunoblot to confirm GEP as the interacting partner. GEP and TPM3 expressions were then examined by real-time quantitative RT-PCR in clinical samples, and their transcript levels were significantly correlated. Elevated TPM3 levels were observed in liver cancer compared with the adjacent non-tumorous liver, and patients with elevated TPM3 levels were shown to have poor recurrence-free survival. Protein expression of GEP and TPM3 was observed only in the cytoplasm of liver cancer cells by immunohistochemical staining. Conclusions: TPM3 is an interacting partner of GEP and may play an important role in hepatocarcinogenesis. © 2012 Lam et al.published_or_final_versio

N-wasp Is Required For Structural Integrity Of The Blood-testis Barrier.

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Changes in endogenous Zn and Cu distribution in different cytosolic protein fractions in mouse liver after administration of a single sublethal dose of CdCl2

The time course of change in tissue Cd, Cu and Zn contents, their distribution in cellular protein fractions as well as the profile of MT gene expression in mouse liver was described over a 7 days period following a single intraperitoneal injection of 2 mg/kg of CdCl2. The result showed that Cd accumulated rapidly in mouse liver. Between 1 h and 7 days after administration, over 18% of the total Cd administered were found in the liver. Cd administration was also associated with the overexpression of the MT-mRNA. However, the time course of induction was not parallel to the change in tissue Cd content. When separated on a Sephadex G-75 column, majority of Cd was found to bind to the fractions known to contain the metal-binding protein, metallothionein (MT). From day 2 after Cd administration, a small amount of the metal was also found associated with the high molecular weight (HMW) proteins. In addition to Cd, tissue Zn content was affected most during the entire study. There was a significant decrease in tissue Zn content during the initial 8 h but tissue Zn content increased significantly throughout the following 6 days. At 1-7 days, majority of Zn was associated with the HMW protein fraction. Although there was no significant change in total tissue Cu content, distribution of Cu in different protein fractions was detected. While in control aniamls, Cu was mainly associated with the HMW proteins, some was found in the MT fraction on the second day. On the 7th day, Cu distribution had deteriorated. Together with changes seen in Cd, the results might suggest that injury had occurred in the tissue at this time. The results of the present study showed that Cd caused a change in subcellular distribution of tissue endogenous metals, which might reflect alteration of cellular functional activities. (C) 2000 Elsevier Science Ireland Ltd.link_to_subscribed_fulltex

Genomics approach to identify growth factor and drug transporter associated with liver cancer recurrence and chemo-resistance

Oral PresentationBackground: Liver cancer is the third leading cause of cancer death globally. Surgery is the mainstay for early stage patients but recurrence is still common. Chemotherapy has been widely used to treat advanced liver cancer but with marginal efficacy. Recent evidences have shown the role of tumor initiating cells on chemo-resistance and recurrence, however, the key genes remind inconclusive. Recently, we have demonstrated that the novel growth factor granulin-epithelin precursor (GEP) and drug transporter ABCB5 were associated with chemo-resistance and recurrence-free survival (Gastroenterology 2011). GEP has been identified as a potential therapeutic target for liver cancer in our earlier genomic studies. The aim of the study is to identify additional drug transporters with clinical relevance. Methods: The liver cancer gene expression profiles reported previously were re-examined for all the ATP-binding transporters. The gene of interest were further validated in an independent cohort using real-time quantitative RT-PCR. Results: ABCF1 was upregulated in tumor compared with non-tumor (P/0.001), and increased ABCF1 was associated with poor recurrence- free survival (log-rank test, P = 0.001). Functionally, suppression of ABCF1 rendered the liver cancer cells sensitive to chemotherapeutic agents. Liver cancer cells that were positive for GEP and ABCB5/ABCF1 showed enhanced expression of the stem cell related signaling molecules-catenin, Oct4 and Nanog. Conclusions: In summary, chemo-resistance and cancer recurrence are dictated by a subset of cells expressing GEP and ABC transporters. Targeting the novel growth factor GEP and drug transporters, in combination with chemotherapy, could provide novel treatment modalities to eradicate the aggressive hepatic tumor initiating cells

Genomics approach to identify drug transporter ABCF1 associated with liver cancer recurrence and chemo-resistance

Poster Session 33 - New Models, Screens, and Diagnostics for Target Identification: abstract no. 882BACKGROUND: Liver cancer is the third leading cause of cancer death globally. Surgery is the mainstay for early stage patients with the best survival outcome, but cancer recurrence is still common after curative surgery. Chemotherapy has been widely used to treat advanced liver cancer but with marginal efficacy. There is an urgent need to elucidate the key genes that regulate recurrence and chemo-resistance in the clinical situation. Recently, we have demonstrated that the novel growth factor granulin-epithelin precursor (GEP) is associated with chemo-resistance and recurrence-free survival. GEP has been identified as a potential therapeutic target for liver cancer in our ...link_to_OA_fulltextThe Annual Meeting of the American Association for Cancer Research (AACR 2012), Chicago, IL., 31 March-4 April 2012

Granulin-epithelin precursor and ATP-dependent binding cassette (ABC)B5 regulate liver cancer cell chemoresistance

Background & Aims: Chemotherapy is used to treat unresectable liver cancer with marginal efficacy; this might result from hepatic cancer cells with stem cell and chemoresistant features. Gene expression profiling studies have shown that hepatic cancer cells express granulin-epithelin precursor (GEP); we investigated its role in hepatic cancer stem cell functions and chemoresistance. Methods: The effects of GEP and drug transporter signaling on chemoresistance were investigated in hepatic cancer stem cells. We analyzed the expression patterns of 142 clinical samples from liver tumors, adjacent nontumorous liver tissue, and liver tissue from patients who did not have cancer. Results: GEP regulated the expression of the adenosine triphosphatedependent binding cassette (ABC)B5 drug transporter in liver cancer cells. Chemoresistant cells that expressed GEP had increased levels of ABCB5; suppression of ABCB5 sensitized the cells to doxorubicin uptake and apoptosis. Most cells that expressed GEP and ABCB5 also expressed the hepatic cancer stem cell markers CD133 and EpCAM; blocking ABCB5 reduced their expression. Expression levels of GEP and ABCB5 were correlated in human liver tumor samples. ABCB5 levels were increased in liver cancer cells compared with nontumor liver tissue from patients with cirrhosis or hepatitis, or normal liver tissue. ABCB5 expression was associated with the recurrence of hepatocellular carcinoma after partial hepatectomy. Conclusions:: Expression of GEP and ABCB5 in liver cancer stem cells is associated with chemoresistance and reduced survival times of patients with hepatocellular carcinoma. Reagents designed to target these proteins might be developed as therapeutics and given in combination with chemotherapy to patients with liver cancer. © 2011 AGA Institute.link_to_subscribed_fulltex