FBX8 Acts as an Invasion and Metastasis Suppressor and Correlates with Poor Survival in Hepatocellular Carcinoma

Background

F-box only protein 8 (FBX8), a novel component of F-box proteins, is lost in several cancers and has been associated with invasiveness of cancer cells. However, its expression pattern and role in the progression of hepatocellular carcinoma remain unclear. This study investigated the prognostic significance of FBX8 in hepatocellular carcinoma samples and analyzed FBX8 function in hepatocellular carcinoma cells by gene manipulation.

Methodology

The expression of FBX8 was detected in 120 cases of clinical paraffin-embedded hepatocellular carcinoma tissues, 20 matched pairs of fresh tissues and five hepatocellular carcinoma cell lines by immunohistochemistry with clinicopathological analyses, real-time RT-PCR or Western blot. The correlation of FBX8 expression with cell proliferation and invasion in five HCC cell lines was analyzed. Moreover, loss of function and gain of function assays were performed to evaluate the effect of FBX8 on cell proliferation, motility, invasion in vitro and metastasis in vivo.

Conclusions

We found that FBX8 was obviously down-regulated in HCC tissues and cell lines (P<0.05). The FBX8 down-regulation correlated significantly with poor prognosis, and FBX8 status was identified as an independent significant prognostic factor. Over-expression of FBX8 decreased proliferation, migration and invasion in HepG2 and 97H cells, while knock-down of FBX8 in 7721 cells showed the opposite effect. FBX8 negatively correlated with cell proliferation and invasion in 7701, M3, HepG2 and 97H cell lines. In vivo functional assays showed FBX8 suppressed tumor growth and pulmonary metastatic potential in mice. Our results indicate that down-regulation of FBX8 significantly correlates with invasion, metastasis and poor survival in hepatocellular carcinoma patients. It may be a useful biomarker for therapeutic strategy and control in hepatocellular carcinoma treatment.     

Akt2 Regulates Metastatic Potential in Neuroblastoma

Activation of PI3K/AKT pathway correlates with poor prognosis in patients with neuroblastoma. Our previous studies have demonstrated that PI3K/AKT signaling is critical for the oncogenic transformations induced by gastrin-releasing peptide (GRP) and its receptor, GRP-R, in neuroblastoma. Moreover, PI3K/AKT-dependent oncogenic transformations require N-myc, an extensively studied oncogene in neuroblastoma. Whether AKT directly regulates the expression of N-myc oncogene is yet to be determined. Here, we report a novel finding that of the three AKT isoforms, AKT2 specifically regulated N-myc expression in neuroblastoma cells. We also confirmed that GRP-R is upstream of AKT2 and in turn, regulated N-myc expression via AKT2 in neuroblastoma cells. Functional assays demonstrated that attenuation of AKT2 impaired cell proliferation and anchorage-independent cell growth, and decreased the secretion of angiogenic factor VEGF in vitro. Furthermore, silencing AKT2 inhibited migration and invasion of neuroblastoma cells in vitro. Xenografts established by injecting AKT2 silenced human neuroblastoma cells into murine spleen expressed decreased levels of AKT2 and resulted in fewer liver metastases compared to controls in vivo. Hence, our study highlights the potential molecular mechanism(s) mediating the oncogenic role of GRP/GRP-R and demonstrates a novel role for AKT2 in neuroblastoma tumorigenesis, indicating that targeting the GRP/GRP-R/AKT2 axis may be important for developing novel therapeutics in the treatment of clinically aggressive neuroblastoma.     

The Diverse Distribution of Risk Factors between Breast Cancer Subtypes of ER,PR and HER2: A 10-Year Retrospective Multi-Center Study in China

Introduction

Hormone receptors, human epidermal growth factor receptor 2 and some risk factors determine therapies and prognosis of breast cancer. The risk factors distributed differently between patients with receptors. This study aimed to investigate the distribution of risk factors between subtypes of breast cancer by the 3 receptors in Chinese native women with a large sample size.

Methods

The multi-center study analyzed 4211 patient medical records from 1999 to 2008 in 7 regions of China. Data on patients’ demographic information, risk factors (menopausal status, parity, body mass index) and receptor statuses were extracted. Breast cancer subtypes included ER (+/−), PR (+/−), HER2 (+/−), 4 ER/PR and 4 molecular subtypes. Wilcoxon and Chi-square tests were used to estimate the difference. The unconditional logistic regression model was used for analysis, and presented p-value after Bonferroni correction in the results.

Results

Compared to patients with negative progesterone receptor, the positive patients were younger at diagnosis, and reported less likely in postmenopausal status and lower parity (p<0.05). Comparing with the subtype of ER+/PR+, ER+/PR− subtype were 4-year older at diagnosis (OR = 1.02), more likely to be postmenopausal (OR = 1.91) and more likely to have >1 parity (OR = 1.36) (p<0.05); ER−/PR− subtype were more likely to be postmenopausal (OR = 1.33) and have >1 parity (OR = 1.19) (p<0.05). In contrast to the luminal A subtype, triple negative subtype had a lower BMI (OR = 0.96) and ORs of overweight and obesity reduced by >20% (p<0.05).

Conclusion

In this study, it was found that Chinese female patients did have statistically significant differences of age, menopausal status, parity and body mass index between breast cancer subtypes. Studies are warranted to further investigate the risk factors between subtypes, which was meaningful for prevention and treatment among Chinese females.     

Induction of Porcine Host Defense Peptide Gene Expression by Short-Chain Fatty Acids and Their Analogs

Dietary modulation of the synthesis of endogenous host defense peptides (HDPs) represents a novel antimicrobial approach for disease control and prevention, particularly against antibiotic-resistant infections. However, HDP regulation by dietary compounds such as butyrate is species-dependent. To examine whether butyrate could induce HDP expression in pigs, we evaluated the expressions of a panel of porcine HDPs in IPEC-J2 intestinal epithelial cells, 3D4/31 macrophages, and primary monocytes in response to sodium butyrate treatment by real-time PCR. We revealed that butyrate is a potent inducer of multiple, but not all, HDP genes. Porcine β-defensin 2 (pBD2), pBD3, epididymis protein 2 splicing variant C (pEP2C), and protegrins were induced markedly in response to butyrate, whereas pBD1 expression remained largely unaltered in any cell type. Additionally, a comparison of the HDP-inducing efficacy among saturated free fatty acids of different aliphatic chain lengths revealed that fatty acids containing 3–8 carbons showed an obvious induction of HDP expression in IPEC-J2 cells, with butyrate being the most potent and long-chain fatty acids having only a marginal effect. We further investigated a panel of butyrate analogs for their efficacy in HDP induction, and found glyceryl tributyrate, benzyl butyrate, and 4-phenylbutyrate to be comparable with butyrate. Identification of butyrate and several analogs with a strong capacity to induce HDP gene expression in pigs provides attractive candidates for further evaluation of their potential as novel alternatives to antibiotics in augmenting innate immunity and disease resistance of pigs.     

Predictors of Intention to Smoke among Junior High School Students in Shanghai,China: An Empirical Test of the Information-Motivation-Behavioral Skills (IMB) Model

Background

Adolescent smoking is a worldwide problem that is particularly severe in low- and middle-income countries. Many endogenous and environmental factors affect the intention to smoke, so a comprehensive model is needed to understand the significance and relationship of predictors. The study aimed to test the associations among information-motivation-behavioral skills (IMB) model constructs as predictors of intention to smoke in junior high school students in Shanghai, China.

Methods

We conducted a cross-sectional study of 16,500 junior high school students in Shanghai, China. Data on tobacco-related information, motivation, behavioral skills, and behaviors were collected from students. Structural equation model (SEM) was used to assess the IMB model.

Results

The mean age of participants was 13.8 years old (standard deviation = 1.02; range 11–17). The experimental smoking rate among junior high school students was 6.6% and 8.7% of the participants expected that they would be smokers in 5 years. The IMB model provided acceptable fit to the data (comparative fit index = 0.984, root mean square error of approximation = 0.04). Intention to smoke was predicted by behavioral skills (β= 0.670, P < 0.001) and motivation (β= 0.095, P<0.001) among junior high school students.

Conclusion

The IMB model provides a good understanding of the predictors of intention to smoke and it suggests future interventions among junior high school students should focus on improving motivation and behavioral skills.     

Dietary N-Carbamylglutamate Supplementation Boosts Intestinal Mucosal Immunity in Escherichia coli Challenged Piglets

N-carbamylglutamate (NCG) has been shown to enhance performance in neonatal piglets. However, few studies have demonstrated the effect of NCG on the intestinal mucosal barrier. This study was conducted to determine the effects of dietary NCG supplementation on intestinal mucosal immunity in neonatal piglets after an Escherichia coli (E. coli) challenge. New-born piglets (4 d old) were assigned randomly to one of four treatments (n = 7), including (I) sham challenge, (II) sham challenge +50 mg/kg NCG, (III) E. coli challenge, and (IV) E. coli challenge +50 mg/kg NCG. On d 8, pigs in the E. coli challenge groups (III and IV) were orally challenged with 5 mL of E. coli K88 (10⁸ CFU/mL), whereas pigs in the sham challenge groups (I and II) were orally dosed with an equal volume of water. On d 13, all piglets were sacrificed, and samples were collected and examined. The results show that average daily gain in the E. coli challenged piglets (III and IV) was decreased (P_E.coli<0.05). However, it tended to be higher in the NCG treated piglets (II and IV). Ileum secretory IgA, as well as IFN-γ, IL-2, IL-4 and IL-10 in ileal homogenates, were increased in E. coli challenged piglets (III and IV). Similarly, ileum SIgA and IL-10 levels, and CD4⁺ percentage in NCG treated piglets (II and IV) were higher than no-NCG treated piglets (P_NCG<0.05). However, the IL-2 level was only decreased in the piglets of E. coli challenge + NCG group (IV) compared with E. coli challenge group (III) (P<0.05). No change in the IL-2 level of the sham challenged piglets (III) was observed. In conclusion, dietary NCG supplementation has some beneficial effects on intestinal mucosal immunity in E. coli challenged piglets, which might be associated with stimulated lymphocyte proliferation and cytokine synthesis. Our findings have an important implication that NCG may be used to reduce diarrhea in neonatal piglets.     

In Vivo Regulation of Brain-Derived Neurotrophic Factor in Dorsal Root Ganglia Is Mediated by Nerve Growth Factor-Triggered Akt Activation during Cystitis

The role of brain-derived neurotrophic factor (BDNF) in sensory hypersensitivity has been suggested; however the molecular mechanisms and signal transduction that regulate BDNF expression in primary afferent neurons during visceral inflammation are not clear. Here we used a rat model of cystitis and found that the mRNA and protein levels of BDNF were increased in the L6 dorsal root ganglia (DRG) in response to bladder inflammation. BDNF up-regulation in the L6 DRG was triggered by endogenous nerve growth factor (NGF) because neutralization of NGF with a specific NGF antibody reduced BDNF levels during cystitis. The neutralizing NGF antibody also subsequently reduced cystitis-induced up-regulation of the serine/threonine kinase Akt activity in L6 DRG. To examine whether the NGF-induced Akt activation led to BDNF up-regulation in DRG in cystitis, we found that in cystitis the phospho-Akt immunoreactivity was co-localized with BDNF in L6 DRG, and prevention of the endogenous Akt activity in the L6 DRG by inhibition of phosphoinositide 3-kinase (PI3K) with a potent inhibitor LY294002 reversed cystitis-induced BDNF up-regulation. Further study showed that application of NGF to the nerve terminals of the ganglion-nerve two-compartmented preparation enhanced BDNF expression in the DRG neuronal soma; which was reduced by pre-treatment of the ganglia with the PI3K inhibitor LY294002 and wortmannin. These in vivo and in vitro experiments indicated that NGF played an important role in the activation of Akt and subsequent up-regulation of BDNF in the sensory neurons in visceral inflammation such as cystitis.     

Genetic Association Study betweenSTK39 and CCDC62/HIP1R and Parkinson’s Disease

Background

The first large-scale meta-analysis of published genome-wide association studies (GWAS) in Parkinson’s disease (PD) identified 5 new genetic loci (ACMSD, STK39, MCCC1/LAMP3, SYT11, and CCDC62/HIP1R). Very recently, a large-scale replication and heterogeneity study also reported that STK39 and CCDC62/HIP1R increased risk of PD in Asian and Caucasian populations. However, their roles still remain unclear in a Han Chinese population from mainland China.

Methods

We examined genetic associations of STK39 rs2102808 and CCDC62/HIP1R rs12817488 with PD susceptibility in a Han Chinese population of 783 PD patients and 725 controls. We also performed further stratified analyses by the age of onset and accomplished in-depth clinical characteristics analyses between the different genotypes for each locus.

Results

No significant differences were observed in the minor allele frequency (MAF) among cases and controls at the two loci (STK39 rs2102808: OR = 1.06, 95% CI = 0.91, 1.23, P = 0.467; CCDC62/HIP1R rs12817488: OR = 0.88, 95% CI = 0.76, 1.01, P = 0.072). Subgroup analyses by the age of onset also showed no significant differences among different subgroups of the two loci. In addition, minor allele carriers cannot be distinguished from non-carriers based on their clinical features at the two loci.

Conclusions

We are unable to demonstrate the association between STK39 and CCDC62/HIP1R and PD susceptibility in a Han Chinese population from mainland China. Additional replication studies in other populations and functional studies are warranted to better validate the role of the two new loci in PD risk.