Post-translational deregulation of YAP1 is genetically controlled in rat liver cancer and determines the fate and stem-like behavior of the human disease

Previous studies showed that YAP1 is over-expressed in hepatocellular carcinoma (HCC). Here we observed higher expression of Yap1/Ctgf axis in dysplastic nodules and HCC chemically-induced in F344 rats, genetically susceptible to hepatocarcinogenesis, than in lesions induced in resistant BN rats. In BN rats, highest increase in Yap1-tyr357, p73 phosphorylation and Caspase 3 cleavage occurred. In human HCCs with poorer prognosis ( 3 years survival; HCCB). In the latter, higher levels of phosphorylated YAP1-ser127, YAP1-tyr357 and p73, YAP1 ubiquitination, and Caspase 3 cleavage occurred. Expression of stemness markers NANOG, OCT-3/4, and CD133 were highest in HCCP and correlated with YAP1 and YAP1-TEAD levels. In HepG2, Huh7, and Hep3B cells, forced YAP1 over-expression led to stem cell markers expression and increased cell viability, whereas inhibition of YAP1 expression by specific siRNA, or transfection of mutant YAP1 which does not bind to TEAD, induced opposite alterations. These changes were associated, in Huh7 cells transfected with YAP1 or YAP1 siRNA, with stimulation or inhibition of cell migration and invasivity, respectively. Furthermore, transcriptome analysis showed that YAP1 transfection in Huh7 cells induces over-expression of genes involved in tumor stemness. In conclusion, Yap1 post-translational modifications favoring its ubiquitination and apoptosis characterize HCC with better prognosis, whereas conditions favoring the formation of YAP1-TEAD complexes are associated with aggressiveness and acquisition of stemness features by HCC cells

The study of applying heat to enhance moisture transfer in knitted spacer structures

The aim of the article is to report the research of the Advanced Textiles Research Group on the application of heat to enhance the moisture transmission in knitted spacer structures. The current trend in the design and development of moisture management textiles is to use knitted spacer structures. Generally, in moisture management textiles, the moisture is transmitted through the fabric due to capillary forces, which are influenced by the hydrostatic pressure difference between the two fabric layers and the geometry and the dimensions of the capillaries of the sandwiched fibre layer of a knitted spacer structures. However, the hydrostatic pressure difference is also influenced by the outer environmental changes. The research has demonstrated that the moisture transfer rate of up to 30% per 100 cm2 of fabric area can be achieved by creating a temperature gradient between the two layers of a knitted spacer structures. This temperature gradient was achieved by application of heat at one layer of the knitted spacer structures, which influenced the hydrostatic pressure difference of the knitted spacer structures. Application of heat to the knitted spacer structures was achieved by knitting small heater elements on side of knitted spacer structures to create an active moisture management structure. Wash tests, temperature rise rates and moisture wettability experiments of the active moisture management structure were performed, and the results are discussed in the publication

Mapping a Sex Hormone–Sensitive Gene Determining Female Resistance to Liver Carcinogenesis in a Congenic F344.BN-Hcs4 Rat

Hepatocellular carcinoma (HCC) is prevalent in human and rodent males. Hepatocarcinogenesis is controlled by various genes in susceptible F344 and resistant Brown Norway (BN) rats. B alleles at Hcs4 locus, on RNO16, control neoplastic nodule volume. We constructed the F344.BN-Hcs4 recombinant congenic strain (RCS) by introgressing a 4.41-cM portion of Hcs4 from BN strain in an isogenic F344 background. Preneoplastic and neoplastic lesions were induced by the ''resistant hepatocyte'' protocol. Eight weeks after initiation, lesion volume and positivity for proliferating cell nuclear antigen (PCNA) were much higher in lesions of F344 than BN rats of both sexes. These variables were lower in females than in males. Lesion volume and PCNA values of male RCS were similar to those of F344 rats, but in females corresponded to those of BN females. Carcinomatous nodules and HCC developed at 32 and 60 weeks, respectively, in male F344 and congenics and, rarely, in F344 females. BN and congenic females developed only eosinophilic/clear cells nodules. Gonadectomy of congenic males, followed by B-estradiol administration, caused a decrease in Ar expression, an increase in Er-a expression, and development of preneoplastic lesions comparable to those from BN females. Administration of testosterone to gonadectomized females led to Ar increase and development of preneoplastic lesions as in F344 males. This indicates a role of homozygous B alleles at Hcs4 in the determination of phenotypic patterns of female RCS and presence at Hcs4 locus of a high penetrance gene(s), activated by estrogens and inhibited/unaffected by testosterone, conferring resistance to females in which the B alleles provide higher resistance. (Cancer Res 2006; 66(21): 10384-90

aberrant inos signaling is under genetic control in rodent liver cancer and potentially prognostic for the human disease

Mounting evidence underlines the role of inducible nitric oxidesynthase (iNOS) in hepatocellular carcinoma (HCC) develop-ment, but its functional interactions with pathways involved inHCC progression remain uninvestigated. Here, we analyzed inpreneoplastic and neoplastic livers from Fisher 344 and BrownNorway rats, possessing different genetic predisposition to HCC,in transforming growth factor-a (TGF-a) and c-Myc–TGF-atransgenic mice, characterized by different susceptibility toHCC, and in human HCC: (i) iNOS function and interactionswith nuclear factor-kB (NF-kB) and Ha-RAS/extracellularsignal-regulated kinase (ERK) during hepatocarcinogenesis;(ii) influence of genetic predisposition to liver cancer on thesepathways and role of these cascades in determining a susceptibleor resistant phenotype and (iii) iNOS prognostic value in humanHCC. We found progressive iNos induction in rat and mouse liverlesions, always at higher levels in the most aggressive models rep-resented by HCC of rats genetically susceptible to hepatocarcino-genesis and c-Myc–TGF-a transgenic mice. iNOS, inhibitor of kBkinase/NF-kB and RAS/ERK upregulation was significantly higherin HCC with poorer prognosis (as defined by patients' survivallength) and positively correlated with tumor proliferation, genomicinstability and microvascularization and negatively with apoptosis.Suppression of iNOS signaling by aminoguanidine led to decreasedHCC growth and NF-kB and RAS/ERK expression and increasedapoptosis both in vivo and in vitro. Conversely, block of NF-kBsignaling by sulfasalazine or short interfering RNA (siRNA) orERK signaling by UO126 caused iNOS downregulation in HCCcell lines. These findings indicate that iNOS cross talk with NF-kB and Ha-RAS/ERK cascades influences HCC growth and prog-nosis, suggesting that key component of iNOS signaling could rep-resent important therapeutic targets for human HCC.IntroductionHepatocellular carcinoma (HCC) is one of the most frequent anddeadliest human cancers worldwide. Current therapies do not improvesignificantly the prognosis of patients with unresectable HCC (1,2).This emphasizes the need to investigate the molecular mechanismsresponsible for HCC development to identify new targets for earlydiagnosis, chemoprevention and treatment.Numerous genes regulating susceptibility to HCC and controllinggrowth, progression and redifferentiation of preneoplastic and neo-plastic lesions have been mapped in rodents (3). Decrease in growthability and/or marked redifferentiation of preneoplastic lesion char-acterizes rodent strains resistant to hepatocarcinogenesis (3,4). Con-sequently, studies on the mechanisms underlying the acquisition ofa phenotype susceptible/resistantto hepatocarcinogenesis in rodentstrains, carrying preneoplastic lesions differently prone to progressto HCC, may lead to the discovery of prognostic markers and ther-apeutic targets for the human disease. Dysplastic nodules and HCCinduced in susceptible Fisher 344 (F344) rats show upregulation ofc-Myc, Cyclin D1, E and A and E2f1 genes, increased cyclinD1–Cdk4, cyclin E–Cdk2 and E2f1–Dp1 complexes and retinoblas-toma protein (pRb) hyperphosphorylation (4–6). These changes areabsent or less pronounced in liver lesions from resistant Brown Norway(BN) rats, where a block of

Identification of DUSP4/6 overexpression as a potential rheostat to NRAS-induced hepatocarcinogenesis

Background Upregulation of the mitogen-activated protein kinase (MAPK) cascade is common in hepatocellular carcinoma (HCC). Neuroblastoma RAS viral oncogene homolog (NRAS) is mutated in a small percentage of HCC and is hitherto considered insufficient for hepatocarcinogenesis. We aimed to characterize the process of N-Ras-dependent carcinogenesis in the liver and to identify potential therapeutic vulnerabilities. Methods NRAS V12 plasmid was delivered into the mouse liver via hydrodynamic tail vein injection (HTVI). The resulting tumours, preneoplastic lesions, and normal tissue were characterized by NanoString® gene expression analysis, Western Blot, and Immunohistochemistry (IHC). The results were further confirmed by in vitro analyses of HCC cell lines. Results HTVI with NRAS V12 plasmid resulted in the gradual formation of preneoplastic and neoplastic lesions in the liver three months post-injection. These lesions mostly showed characteristics of HCC, with some exceptions of spindle cell/ cholangiocellular differentiation. Progressive upregulation of the RAS/RAF/MEK/ERK signalling was detectable in the lesions by Western Blot and IHC. NanoString® gene expression analysis of preneoplastic and tumorous tissue revealed a gradual overexpression of the cancer stem cell marker CD133 and Dual Specificity Phosphatases 4 and 6 (DUSP4/6). In vitro, transfection of HCC cell lines with NRAS V12 plasmid resulted in a coherent upregulation of DUSP4 and DUSP6. Paradoxically, this upregulation in PLC/PRF/5 cells was accompanied by a downregulation of phosphorylated extracellular-signal-regulated kinase (pERK), suggesting an overshooting compensation. Silencing of DUSP4 and DUSP6 increased proliferation in HCC cell lines. Conclusions Contrary to prior assumptions, the G12V NRAS mutant form is sufficient to elicit hepatocarcinogenesis in the mouse. Furthermore, the upregulation of the MAPK cascade was paralleled by the overexpression of DUSP4, DUSP6, and CD133 in vivo and in vitro. Therefore, DUSP4 and DUSP6 might fine-tune the excessive MAPK activation, a mechanism that can potentially be harnessed therapeutically

Health behaviors and risk factors in those who use complementary and alternative medicine

<p>Abstract</p> <p>Background</p> <p>Surveys have generally found that individuals more likely to use complementary and alternative medicine are female, live in the western United States, are likely to have a health complaint, and have a higher socioeconomic status than do nonusers. What is not known is the extent to which those who use complementary and alternative medicine also engage in positive health behaviors, such as smoking cessation or increased physical activity and/or exhibit fewer health risk factors such as obesity. This has been identified as a key research question in a recent Institute of Medicine report. In the present study we sought to determine whether the use of complementary and alternative medicine is associated with health behaviors or risk factors known to impact on health status.</p> <p>Methods</p> <p>The current study is a cross-sectional regression analysis using data from the 2002 National Health Interview Survey. Data were collected in-person from 31,044 adults throughout the 50 states and the District of Columbia.</p> <p>Results</p> <p>After controlling for a range of other factors, we found that engaging in leisure-time physical activity, having consumed alcohol in one's life but not being a current heavy drinker, and being a former smoker are independently associated with the use of CAM. Obese individuals are slightly less likely to use CAM than individuals with a healthy body-mass index. No significant associations were observed between receipt of an influenza vaccine and CAM use.</p> <p>Conclusion</p> <p>Those engaging in positive health behaviors and exhibiting fewer health risk factors are more likely to use CAM than those who forgo positive health behaviors or exhibit more health risk factors. The fact that users of CAM tend to pursue generally healthy lifestyles suggests that they may be open to additional recommendations toward optimizing their health.</p

Post-translational deregulation of YAP1 is genetically controlled in rat liver cancer and determines the fate and stem-like behavior of the human disease

Previous studies showed that YAP1 is over-expressed in hepatocellular carcinoma (HCC). Here we observed higher expression of Yap1/Ctgf axis in dysplastic nodules and HCC chemically-induced in F344 rats, genetically susceptible to hepatocarcinogenesis, than in lesions induced in resistant BN rats. In BN rats, highest increase in Yap1- tyr357, p73 phosphorylation and Caspase 3 cleavage occurred. In human HCCs with poorer prognosis (< 3 years survival after partial liver resection, HCCP), levels of YAP1, CTGF, 14–3–3, and TEAD proteins, and YAP1-14-3-3 and YAP1-TEAD complexes were higher than in HCCs with better outcome (> 3 years survival; HCCB). In the latter, higher levels of phosphorylated YAP1-ser127, YAP1-tyr357 and p73, YAP1 ubiquitination, and Caspase 3 cleavage occurred. Expression of stemness markers NANOG, OCT-3/4, and CD133 were highest in HCCP and correlated with YAP1 and YAP1-TEAD levels. In HepG2, Huh7, and Hep3B cells, forced YAP1 over-expression led to stem cell markers expression and increased cell viability, whereas inhibition of YAP1 expression by specific siRNA, or transfection of mutant YAP1 which does not bind to TEAD, induced opposite alterations. These changes were associated, in Huh7 cells transfected with YAP1 or YAP1 siRNA, with stimulation or inhibition of cell migration and invasivity, respectively. Furthermore, transcriptome analysis showed that YAP1 transfection in Huh7 cells induces over-expression of genes involved in tumor stemness. In conclusion, Yap1 post-translational modifications favoring its ubiquitination and apoptosis characterize HCC with better prognosis, whereas conditions favoring the formation of YAP1-TEAD complexes are associated with aggressiveness and acquisition of stemness features by HCC cells

Alterations of Methionine Metabolism as Potential Targets for the Prevention and Therapy of Hepatocellular Carcinoma

Several researchers have analyzed the alterations of the methionine cycle associated with liver disease to clarify the pathogenesis of human hepatocellular carcinoma (HCC) and improve the preventive and the therapeutic approaches to this tumor. Different alterations of the methionine cycle leading to a decrease of S-adenosylmethionine (SAM) occur in hepatitis, liver steatosis, liver cirrhosis, and HCC. The reproduction of these changes in MAT1A-KO mice, prone to develop hepatitis and HCC, demonstrates the pathogenetic role of MAT1A gene under-regulation associated with up-regulation of the MAT2A gene (MAT1A:MAT2A switch), encoding the SAM synthesizing enzymes, methyladenosyltransferase I/III (MATI/III) and methyladenosyltransferase II (MATII), respectively. This leads to a rise of MATII, inhibited by the reaction product, with a consequent decrease of SAM synthesis. Attempts to increase the SAM pool by injecting exogenous SAM have beneficial effects in experimental alcoholic and non-alcoholic steatohepatitis and hepatocarcinogenesis. Mechanisms involved in hepatocarcinogenesis inhibition by SAM include: (1) antioxidative effects due to inhibition of nitric oxide (NO&bull;) production, a rise in reduced glutathione (GSH) synthesis, stabilization of the DNA repair protein Apurinic/Apyrimidinic Endonuclease 1 (APEX1); (2) inhibition of c-myc, H-ras, and K-ras expression, prevention of NF-kB activation, and induction of overexpression of the oncosuppressor PP2A gene; (3) an increase in expression of the ERK inhibitor DUSP1; (4) inhibition of PI3K/AKT expression and down-regulation of C/EBP&alpha; and UCA1 gene transcripts; (5) blocking LKB1/AMPK activation; (6) DNA and protein methylation. Different clinical trials have documented curative effects of SAM in alcoholic liver disease. Furthermore, SAM enhances the IFN-&alpha; antiviral activity and protects against hepatic ischemia-reperfusion injury during hepatectomy in HCC patients with chronic hepatitis B virus (HBV) infection. However, although SAM prevents experimental tumors, it is not curative against already established experimental and human HCCs. The recent observation that the inhibition of MAT2A and MAT2B expression by miRNAs leads to a rise of endogenous SAM and strong inhibition of cancer cell growth could open new perspectives to the treatment of HCC

Genetic Predisposition to Hepatocellular Carcinoma

Liver preneoplastic and neoplastic lesions of the genetically susceptible F344 and resistant BN rats cluster, respectively, with human HCC with better (HCCB) and poorer prognosis (HCCP); therefore, they represent a valid model to study the molecular alterations determining the genetic predisposition to HCC and the response to therapy. The ubiquitin-mediated proteolysis of ERK-inhibitor DUSP1, which characterizes HCC progression, favors the unrestrained ERK activity. DUSP1 represents a valuable prognostic marker, and ERK, CKS1, or SKP2 are potential therapeutic targets for human HCC. In DN (dysplastic nodule) and HCC of F344 rats and human HCCP, DUSP1 downregulation and ERK1/2 overexpression sustain SKP2-CKS1 activity through FOXM1, the expression of which is associated with a susceptible phenotype. SAM-methyl-transferase reactions and SAM/SAH ratio are regulated by GNMT. In addition, GNMT binds to CYP1A, PARP1, and NFKB and PREX2 gene promoters. MYBL2 upregulation deregulates cell cycle and induces the progression of premalignant and malignant liver. During HCC progression, the MYBL2 transcription factor positively correlates with cells proliferation and microvessel density, while it is negatively correlated to apoptosis. Hierarchical supervised analysis, regarding 6132 genes common to human and rat liver, showed a gene expression pattern common to normal liver of both strains and BN nodules, and a second pattern is observed in F344 nodules and HCC of both strains. Comparative genetics studies showed that DNs of BN rats cluster with human HCCB, while F344 DNs and HCCs cluster with HCCP