Proteomic profiling of hepatocellular carcinoma in Chinese cohort reveals heat-shock proteins (Hsp27, Hsp70, GRP78) up-regulation and their associated prognostic values

To facilitate the identification of candidate molecular biomarkers that are linked to the pathogenesis of hepatocellular carcinoma (HCC), we investigated protein-expression profiles of 146 tissue specimens including 67 pairs of tumors and adjacent non-tumors resected from HCC patients as well as 12 normal livers by 2-DE. Among the 1800 spots displayed in the liver proteome, a total of 90 protein species were found to be significantly different between the three groups (P < 0.05). Three of the top candidate markers up-regulated in HCC, with high receiver operating characteristic (ROC) curves, were identified by MS/MS analysis and belonged to the chaperone members: heat-shock protein (Hsp)27, Hsp70 and glucose-regulated protein (GRP)78. Over-expression of these chaperone proteins in HCC tissues was confirmed by Western blotting and immunohistochemistry. In correlation with clinico-pathological parameters, expression of Hsp27 was linked to alpha-fetoprotein level (P = 0.007) whereas up-regulation of GRP78 was associated with tumor venous infiltration (P = 0.035). No significant association of Hsp70 with any pathologic features was observed. The present HCC proteome analysis revealed that in response to the stressful cancerous microenvironment, tumor cells strived to increase the expression of chaperone proteins for cyto-protective function and to enhance tumor growth and metastasis.     

Treatment with vector-expressed small hairpin RNAs against Ki67 RNA-induced cell growth inhibition and apoptosis in human renal carcinoma cells

Short hairpin RNAs (shRNAs) transcribed by.RNA polymerase Ⅲ promoters can triggersequence-selective gene silencing in mammalian cells.By virtue of their excellent function in knocking downexpression of cancer-associated genes,shRNAs could be used as new therapeutic agents for cancer.Asoverexpression of Ki67 in renal cancer has been correlated to a more aggressive tumor phenotype,inhibitionof Ki67 protein expression by means of shRNAs seems to be a promising approach for the therapy of renalcancer.In this study,we constructed an expression plasmid encoding shRNAs against the Ki67 gene,namedpSilencerKi67,and transfected it into human renal carcinoma cells.The pSilencerKi67 was shown to signifi-cantly knock down the expression of the Ki67 gene in human renal carcinoma cells,resulting in inhibitingproliferation and inducing apoptotic cell death that can be maintained for at least 6d.These findings offer thepromise of using vector-based shRNAs against Ki67 in renal cancer gene therapy.     

Second intron of mouse nestin gene directs its expression in pluripotent embryonic carcinoma cells through POU factor binding site

Nestin,an intermediate filament protein,is expressed in the neural stem cells of the developingcentral nervous system.This tissue-specific expression is driven by the neural stem cell-specific enhancer inthe second intron of the nestin gene.In this study,we showed that the mouse nestin gene was expressed inpluripotent embryonic carcinoma (EC) P19 and F9 cells,not in the differentiated cell types.This cell type-specific expression was conferred by the enhancer in the second intron.Mutation of the conserved POUfactor-binding site in the enhancer abolished the reporter gene expression in EC cells.Oct4,a Class V POUfactor,was found to be coexpressed with nestin in EC cells.Electrophoretic mobility-shift assays and supershiftassays showed that a unique protein-DNA complex was formed specifically with nuclear extracts of ECcells,and Oct4 protein was included.Together,these results suggest the functional relevance between theconserved POU factor-binding site and the expression of the nestin gene in pluripotent EC cells.     

Body image and quality of life in post massive weight loss body contouring patients

Objective: Because post‐bariatric surgery patients undergo massive weight loss, the resulting skin excess can lead to both functional problems and profound dissatisfaction with appearance. Correcting skin excess could improve all these corollaries, including body image. Presently, few data are available documenting body image and weight‐related quality of life in this population. Research Methods and Procedures: Eighteen patients who underwent both bariatric surgery and body contouring completed our study. Both established surveys and new surveys designed specifically for the study were used to assess body perception and ideals, quality of life, and mood. Patients were surveyed at the following time‐points: pre‐body contouring (after massive weight loss) and both 3 and 6 month post‐body contouring. Statistical testing was performed using Student's t test and ANOVA. Results: The mean age of the patients was 46 ± 10 years (standard deviation). Quality of life improved after obesity surgery and was significantly enhanced after body contouring. Three months after body contouring, subjects ascribed thinner silhouettes to both current appearance and ideal body image. Body image also improved with body contouring surgery. Mood remained stable over 6 months. Discussion: Body contouring after surgical weight loss improved both quality‐of‐life measurements and body image. Initial body dissatisfaction did not correlate with mood. Body contouring improved body image but produced dissatisfaction with other parts of the body, suggesting that as patients become closer to their ideal, these ideals may shift. We further developed several new assessment methods that may prove useful in understanding these post‐surgical weight loss patients.     

Mechano-chemical signaling in F9 cells

We investigated the molecular mechanism by which cells recognize and respond to physical forces in their local environment. Using a model system, to study wild type mouse F9 embryonic carcinoma cells, we examined how these cells sense mechanical stresses and translate them into biochemical responses through their cell surface receptor integrin and via the focal adhesion complex (FAC). Based on studies that show that many signal transducing molecules are immobilized on the cytoskeleton at the site of integrin binding within the focal adhesion complex, we found a time-dependent increase of focal adhesion kinase (pp125(FAK)) phosphorylation possibly due to protein kinase C (PKC) activation as well as protein kinase A (PKA) activity increase upon cell adhesion/spreading. These studies provide some insight into intracellular mechano-chemical signaling.     

DBC2 is essential for transporting vesicular stomatitis virus glycoprotein

DBC2 is a tumor suppressor gene linked to breast and lung cancers. Although DBC2 belongs to the RHO GTPase family, it has a unique structure that contains a Broad-Complex/Tramtrack/Bric a Brac (BTB) domain at the C terminus instead of a typical CAAX motif. A limited number of functional studies on DBC2 have indicated its participation in diverse cellular activities, such as ubiquitination, cell-cycle control, cytoskeleton organization and protein transport. In this study, the role of DBC2 in protein transport was analyzed using vesicular stomatitis virus glycoprotein (VSVG) fused with green fluorescent protein. We discovered that DBC2 knockdown hinders the VSVG transport system in 293 cells. Previous studies have demonstrated that VSVG is transported via the microtubule motor complex. We demonstrate that DBC2 mobility depends also on an intact microtubule network. We conclude that DBC2 plays an essential role in microtubule-mediated VSVG transport from the endoplasmic reticulum to the Golgi apparatus.     

Apoptotic and anti-proliferative effects of 17β-estradiol and 17β-estradiol-like compounds in the Hep3B cell line

Since there is evidence for estrogen and estrogen-like compounds to have beneficial effect on the pathogenesis of hepatocellular carcinoma (HCC), this study was designed to investigative the apoptotic and anti-proliferative effects of these compounds on the human hepatoma Hep3B cell line. The Hep3B cells were treated with 17β-estradiol (E2), diethylstilbestrol (DES), tamoxifen, and genistein. After treatments of these compounds at the concentration of 10^-6 or 10^-8 M, the Hep3B cells were demonstrated to have significant DNA fragmentation, nucleus condensation, cytochrome-c leaking from the mitochondria and caspase-3 activation by DAPI and Western blotting. The cells were also observed to have declined proliferative potential by MTT assay, arrested cell cycle by flow-cytometry measurements. However, the cytochrome-c leaking from the mitochondria induced by E2 and E2-like compounds was blocked totally by ICI 182,780 treatment. These finding suggest that estrogen and the estrogen-like compounds may induce anti-proliferative and apoptotic effects in Hep3B cells, and the E2 and the E2-like compounds mediated apoptotic effect was estrogen receptor dependent. Among the drugs tested, E2, E2 agonists (DES and genistein) and partial antagonist (tamoxifen), all showed the stronger anti-tumor potential. The last two authors, Wei-Wen Kuo and Chih-Yang Huang, share equal contribution.