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

Would transcranial direct current stimulation (tDCS) enhance the effects of working memory training in older adults with mild neurocognitive disorder due to Alzheimer’s disease: study protocol for a randomized controlled trial

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Genome-wide association study in two cohorts from a multi-generational mouse advanced intercross line highlights the difficulty of replication due to study-specific heterogeneity

There has been extensive discussion of the Replication Crisis in many fields, including genome-wide association studies

A phase 1 dose-escalating study of pegylated recombinant human arginase 1 (Peg-rhArg1) in patients with advanced hepatocellular carcinoma

Background Hepatocellular carcinoma (HCC) cells are auxotrophic for arginine, depletion of which leads to tumour regression. The current study evaluated safety, pharmacokinetics (PK)/ pharmacodynamics (PD) parameters, and potential anti-tumor activity of pegylated recombinant human arginase 1 (peg-rhArg1) in advanced HCC patients. Methods Eligibility criteria included advanced HCC with measurable lesions, Child-Pugh A or B, and adequate organ function. Initial single IV bolus was followed by weekly doses of peg-rhArgI escalated from 500 U/kg to 2500 U/kg in a 3 + 3 design. Results Fifteen patients were enrolled at weekly doses of 500 U/kg (n = 3), 1000 U/kg (n = 3), 1600 U/kg (n = 3) and 2500 U/kg (n = 6). The median age was 57 years (33-74); 87% were hepatitis B carriers and 47% had prior systemic treatment. The most commonly reported drug-related non-haematological adverse events (AEs) were diarrhea (13.3%), abdominal discomfort (6.7%) and nausea (6.7%). No drug-related haematological AEs were seen. Only 1 of the six patients that received 2500U/kg peg-rhArg1 experienced DLT (grade 4 bilirubin elevation) and thus the maximum tolerated dose was 2500 U/kg. PK and PD analysis indicated that peg-rhArg1 was efficacious in inducing arginine depletion in a dose-dependent manner. Adequate arginine depletion dose was achieved in the 1,600-2,500 U/kg range and therefore the optimal biological dose was at 1600 U/kg, which was chosen as the recommended dose. The best response was stable disease for >8 weeks in 26.7% of the enrolled patients. Conclusion Peg-rhArg1 has manageable safety profile and preliminary evidence of activity in advanced HCC patients.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

Self-rated health in middle-aged and elderly Chinese : distribution, determinants and associations with cardio-metabolic risk factors

Background: Self-rated health (SRH) has been demonstrated to be an accurate reflection of a person's health and a valid predictor of incident mortality and chronic morbidity. We aimed to evaluate the distribution and factors associated with SRH and its association with biomarkers of cardio-metabolic diseases among middle-aged and elderly Chinese. Methods: Survey of 1,458 men and 1,831 women aged 50 to 70 years, conducted in one urban and two rural areas of Beijing and Shanghai in 2005. SRH status was measured and categorized as good (very good and good) vs. not good (fair, poor and very poor). Determinants of SRH and associations with biomarkers of cardio-metabolic diseases were evaluated using logistic regression. Results: Thirty two percent of participants reported good SRH. Males and rural residents tended to report good SRH. After adjusting for potential confounders, residence, physical activity, employment status, sleep quality and presence of diabetes, cardiovascular disease, and depression were the main determinants of SRH. Those free from cardiovascular disease (OR 3.68; 95%CI 2.39; 5.66), rural residents (OR 1.89; 95% CI 1.47; 2.43), non-depressed participants (OR 2.50; 95% CI 1.67; 3.73) and those with good sleep quality (OR 2.95; 95% CI 2.22; 3.91) had almost twice or over the chance of reporting good SRH compared to their counterparts. There were significant associations -and trend- between SRH and levels of inflammatory markers, insulin levels and insulin resistance. Conclusion: Only one third of middle-aged and elderly Chinese assessed their health status as good or very good. Although further longitudinal studies are required to confirm our findings, interventions targeting social inequalities, lifestyle patterns might not only contribute to prevent chronic morbidity but as well to improve populations' perceived health

An individual based computational model of intestinal crypt fission and its application to predicting unrestrictive growth of the intestinal epithelium.

Intestinal crypt fission is a homeostatic phenomenon, observable in healthy adult mucosa, but which also plays a pathological role as the main mode of growth of some intestinal polyps. Building on our previous individual based model for the small intestinal crypt and on in vitro cultured intestinal organoids, we here model crypt fission as a budding process based on fluid mechanics at the individual cell level and extrapolated predictions for growth of the intestinal epithelium. Budding was always observed in regions of organoids with abundant Paneth cells. Our data support a model in which buds are biomechanically initiated by single stem cells surrounded by Paneth cells which exhibit greater resistance to viscoelastic deformation, a hypothesis supported by atomic force measurements of single cells. Time intervals between consecutive budding events, as simulated by the model and observed in vitro, were 2.84 and 2.62 days, respectively. Predicted cell dynamics was unaffected within the original crypt which retained its full capability of providing cells to the epithelium throughout fission. Mitotic pressure in simulated primary crypts forced upward migration of buds, which simultaneously grew into new protruding crypts at a rate equal to 1.03 days-1 in simulations and 0.99 days-1 in cultured organoids. Simulated crypts reached their final size in 4.6 days, and required 40 6.2 days to migrate to the top of the primary crypt. The growth of the secondary crypt is independent of its migration along the original crypt. Assuming unrestricted crypt fission and multiple budding events, a maximal growth rate of the intestinal epithelium of 0.10 days-1 43 is predicted and thus approximately 22 days are required for a 10-fold increase of polyp size. These predictions are in agreement with the time reported to develop macroscopic adenomas in mice after loss of Apc in intestinal stem cells

Suppression of tumorigenesis and metastasis of hepatocellular carcinoma by shRNA interference targeting on homeoprotein Six1

We previously demonstrated that the overexpression of homeoprotein Six1 in hepatocellular carcinoma (HCC) patients is associated with venous infiltration, advanced pathologic tumor metastasis (pTNM) stage and poor overall survival rate (Ng et al. Br J Cancer 2006;95:1050-5). In this study, short hairpin RNA (shRNA) interference approach was used to suppress the expression of Six1 in a metastatic HCC cell line MHCC97L. Stable transfectant MHCC97L-shSix1 carrying Six1-specific shRNA plasmid was established to downregulate Six1 expression to about 40% when compared with MHCC97L-Control. In vitro functional assays demonstrated that the growth rate and proliferation ability of MHCC97L-shSix1 cells were markedly decreased. Moreover, significant decrease of cell motility and invasiveness were observed in MHCC97L-shSix1 cells. Data from in vivo xenograft tumorigenesis model demonstrated that the size of tumor in MHCC97L-shSix1 group was dramatically reduced. Experimental and spontaneous metastasis models indicated that targeting Six1 suppression noticeably reduced the pulmonary metastasis in MHCC97L-shSix1 group. To identify Six1-regulated targets, cDNA microarray was employed to compare the expression profiles of MHCC97L-Control and MHCC97L-shSix1 cells. Twenty-eight downregulated and 24 upregulated genes with known functions were identified in MHCC97L-shSix1. The functions of these target genes are involved in diverse biological activities. Our data suggest that Six1 may be involved in regulation of proliferation and invasiveness of HCC; thus targeting suppression of Six1 is a viable option for treating HCC patients. © 2009 UICC.postprin

Polynomial Growth Harmonic Functions on Finitely Generated Abelian Groups

In the present paper, we develop geometric analytic techniques on Cayley graphs of finitely generated abelian groups to study the polynomial growth harmonic functions. We develop a geometric analytic proof of the classical Heilbronn theorem and the recent Nayar theorem on polynomial growth harmonic functions on lattices \mathds{Z}^n that does not use a representation formula for harmonic functions. We also calculate the precise dimension of the space of polynomial growth harmonic functions on finitely generated abelian groups. While the Cayley graph not only depends on the abelian group, but also on the choice of a generating set, we find that this dimension depends only on the group itself.Comment: 15 pages, to appear in Ann. Global Anal. Geo