IGF-1 Reduces BACE-1 Expression in PC12 Cells via Activation of PI3-K/Akt and MAPK/ERK1/2 Signaling Pathways

Insulin-like growth factor 1 (IGF-1) stimulates α-secretase processing of amyloid precursor protein (APP) and decreases Aβ production. Little is known about the relationship between IGF-1 and β-site amyloid precursor protein cleaving enzyme 1 (BACE-1), the protease essential for the production of β-amyloid peptides (Aβ). Here, we investigated the effect of IGF-1 on BACE-1 in PC12 cells. Quantitative polymerase chain reaction analysis and western blot showed that treatment of cells with IGF-1 significantly decreased the levels of BACE-1 mRNA and protein. Furthermore, IGF-1 increased the phosphorylation of Akt and ERK1/2. The presence of the phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002 and the mitogen-activated protein kinase kinases (MEK) inhibitor PD98059 blocked the effect of IGF-1 on BACE-1. Our data indicated that IGF-1-induced reduction of BACE-1 might involve the PI3-K/Akt and MAPK/ERK1/2 signaling pathways.     

CDH1 -160C>A gene polymorphism is an ethnicity-dependent risk factor for gastric cancer

The associations between E-cadherin (CDH1) gene polymorphisms and gastric cancer (GC) susceptibility are still controversial. Given this uncertainty, we carried out a meta-analysis of published case-control studies to derive more precise estimations of these relationships. Relevant studies were identified from PubMed and EMBASE up to March 2011. Seventeen studies with 3511 GC cases and 4826 controls were selected. Crude odds ratios (OR) and 95% confidence intervals (CI) were used to investigate the strength of the associations. No associations between CDH1 (+54T>C, -160C>A, -347G>GA, -616G>C, -2076C>T and -3159T>C) gene polymorphisms and GC risk for all genetic models were found. As for CDH1 -160C>A polymorphism, subgroup analyses by country, gender, study design, smoking status, Helicobacter pylori infection, and the Lauren classification of GC did not change the results. When stratified by ethnicity, we found the A allele carriers had a significantly increased risk of GC among Caucasians (AA vs. CA+CC: OR=1.50, 95% CI=1.03-2.19, P=0.03), but not among Asians (AA vs. CA+CC: OR=0.87, 95% CI=0.56-1.37, P=0.56). No publication bias was found in the present study. This meta-analysis suggests that CDH1 -160C>A gene polymorphism may contribute to increased risk of GC among Caucasians.     

One-step knockin for inducible expression in mouse embryonic stem cells

Transgenesis enables the elucidation of gene function; however, constant transgene expression is not always desired. The tetracycline responsive system was devised to turn on and off transgene expression at will. It has two components: a doxycycline (dox)-controlled transactivator (TA) and an inducible expression cassette. Integration of these transgenes requires two transfection steps usually accomplished by sequential random integration. Unfortunately, random integration can be problematic due to chromatin position effects, integration of variable transgene units, and mutation at the integration site. Therefore, targeted transgenesis and knockin were developed to target the TA and the inducible expression cassette to a specific location, but these approaches can be costly in time, labor, and money. Here, we describe a one-step Cre-mediated knockin system in mouse embryonic stem cells that positions the TA and inducible expression cassette to a single location. Using this system, we show dox-dependent regulation of eGFP at the DNA topoisomerase 3β promoter. Because Cre-mediated recombination is used in lieu of gene targeting, this system is fast and efficient.     

Prevalence, whole genome characterization and phylogenetic analysis of hepatitis B virus in captive orangutan and gibbon

Background Hepatitis B virus (HBV) is a public health problem worldwide and apart from infecting humans, HBV has been found in non‐human primates. Methods We subjected 93 non‐human primates comprising 12 species to ELISA screening for the serological markers HBsAg, antiHBs and antiHBc. Subsequently, we detected HBV DNA, sequenced the whole HBV genome and performed phylogenetic analysis. Results HBV infection was detected in gibbon (4/15) and orangutan (7/53). HBV DNA isolates from two gibbons and seven orangutans were chosen for complete genome amplification. We aligned the Pre‐S/S, Pre‐C/C and entire genomes with HBV sequences and performed phylogenetic analysis. The gibbon and orangutan viruses clustered within their respective groups. Conclusions Both geographic location and host species influence which HBV variants are found in gibbons and orangutans. Hence, HBV transmission between humans and non‐human primates might be a distinct possibility and additional studies will be required to further investigate this potential risk.     

炭疽芽胞杆菌(Bacillus anthracis)检测质粒的构建及其应用

根据炭疽芽胞杆菌(Bacillus anthracis)毒性质粒pX01和pX02上的2个毒力相关基因cya和capA的序列特点,以pIJ2925为出发载体,采用一步重叠延伸PCR技术(One-step Overlap Extension PCR,简称OOE-PCR)构建了包含cya基因和capA基因保守区DNA片段的炭疽检测质粒pBIB2006。采用复合PCR对模拟炭疽危险品进行分析,结果表明pBIB2006可以为炭疽芽胞杆菌的检测提供准确、安全和方便的阳性参照品,从而为检测炭疽芽胞杆菌和炭疽芽胞杆菌灭活疫苗提供了便利。     

Use of moving aeration membrane bioreactor for the efficient production of tissue type plasminogen activator in serum free medium

A moving aeration-membrane (MAM) bioreactor was employed for the production of 2 μg/mL of tissue type Plasminogen Activator (tPA) in serum free medium from normal human fibroblast cells. This system could maintain high cell density for long periods of steady state conditions in perfusion cultivation. Under normal operating conditions, shear stress was as low as 0.65 dynes/cm² at the agitation speed of 80 rpm. Even though cell density gradually decreased with increasing agitation speed, tPA production increased linearly with increasing shear stress within a moderate range. This culture system allowed production of 2 μg tPA/mL while maintaining a high cell density of 1.0×10⁷ viable cells/mL.     

Induction of Apoptotic Cell Death on Human Cervix Cancer HeLa cells by Extract from <Emphasis Type="Italic">Loranthus yadoriki</Emphasis>

Loranthus yadoriki, one of the Korean mistletoe species, has been already known for anti-viral effects, but the molecular basis that it caused apoptosis in cancer cells was not definitely revealed yet. The aim of this study was to estimate the mechanisms of apoptotic cell death of the extract from Loranthus yadoriki (named as ELY) in human cervix HeLa cells. We identified that ELY prevented the proliferation of HeLa cells between 50 and 300 μg/mL which did not affect non-cancerous HaCaT cells. In addition, ELY induced a morphological change and nucleus disruption as well as an accumulation of sub-G1 phase in HeLa cells. The mechanism study, by using western blot analysis, showed that the phosphorylation of Fas-associated death domain (FADD), Bim and Bak was up-regulated by ELY treatment. Furthermore, the expression of cytochrome c and Apaf-1 was increased by ELY treatment. In immunofluorescence staining, the increased intensity of cleaved caspase-3 and cleaved PARP was also observed under ELY treatment. Sequentially, the caspase cascade was activated by ELY from caspase-8 to caspase-3 and from caspase-9 to caspase-3, in both extrinsic and intrinsic pathways. The results of this study demonstrate that ELY has anti-cancer effects on human cervix cancer HeLa cells via caspase cascade in apoptotic signaling pathways.     

Ferrous Iron Induces Nrf2 Expression in Mouse Brain Astrocytes to Prevent Neurotoxicity

Free radical damage caused by ferrous iron is involved in the pathogenesis of secondary brain injury after intracerebral hemorrhage (ICH). NF‐E2‐related factor 2 (Nrf2), a major phase II gene regulator that binds to antioxidant response element, represents an important cellular cytoprotective mechanism against oxidative damage. We hypothesized that Nrf2 might protect astrocytes from damage by Fe²⁺. Therefore, we examined cytotoxicity in primary astrocytes induced by iron overload and evaluated the effects of Fe²⁺ on Nrf2 expression. The results demonstrated that 24‐h Fe²⁺ exposure exerted time‐ and concentration‐dependent cytotoxicity in astrocytes. Furthermore, Fe²⁺ exposure in astrocytes resulted in time‐ and concentration‐dependent increases in Nrf2 expression, which preceded Fe²⁺ toxicity. Nrf2‐specific siRNA further knocked down Nrf2 levels, resulting in greater Fe²⁺‐induced astrocyte cytotoxicity. These data indicate that induction of Nrf2 expression could serve as an adaptive self‐defense mechanism, although it is insufficient to completely protect primary astrocytes from Fe²⁺‐induced neurotoxicity.     

Genetic polymorphism of glutathione S-transferase T1 and the risk of colorectal cancer: A meta-analysis

Background: Studies investigating the association between genetic polymorphism of glutathione S-transferase T1 (GSTT1) and risk of colorectal cancer have reported conflicting results. In order to clarify the effect of GSTT1 polymorphism on the risk of developing colorectal cancer, we carried out a meta-analysis using published data to obtain more precise estimates of risk. Methods: Electronic searches of PubMed and EMBASE were conducted to select studies for this meta-analysis. Papers were included if they were observational studies investigating the association between GSTT1 polymorphism and colorectal cancer risk. The principal outcome measure was the odds ratio (OR) with 95% confidence interval (CI) for the risk of colorectal cancer associated with GSTT1 null genotype. Results: We identified 30 eligible studies, which included 7635 cases and 12,911 controls. The combined results based on all studies showed that there was a statistically significant link between GSTT1 null genotype and colorectal cancer risk (OR = 1.20, 95% CI = 1.03–1.40). In the analysis of ethnic groups, we observed distinct differences associated with GSTT1 null genotype, the pooled odds ratios for the GSTT1 polymorphism were 1.32 in Caucasians (95% CI = 1.09–1.58) and 1.03 in Asians (95% CI = 0.81–1.32). As far as concerned the interaction between GSTT1 genotype and colorectal cancer risk in relation to smoking history, there was no increase in risk for smokers or nonsmokers with the GSTT1 null genotype (smokers: OR = 1.13, 95% CI = 0.80–1.60, nonsmokers: OR = 0.99, 95% CI = 0.71–1.38). When stratifying by the location of colorectal cancer, we found that there was a statistically significant link in rectal cancer (OR = 1.50, 95% CI = 1.09–2.07), but not in colon cancer (OR = 1.33, 95% CI = 0.94–1.88). No associations could be detected between null GSTT1 polymorphism and age, sex, tumor stage and differentiation. Conclusion: Our current study demonstrates that GSTT1 null genotype is associated with an increased risk of colorectal cancer, specifically, among Caucasians.     

AMPK regulates homeostasis of invasion and viability in trophoblasts by redirecting glucose metabolism: Implications for pre‐eclampsia

Pre‐eclampsia (PE) is deemed an ischemia‐induced metabolic disorder of the placenta due to defective invasion of trophoblasts during placentation; thus, the driving role of metabolism in PE pathogenesis is largely ignored. Since trophoblasts undergo substantial glycolysis, this study aimed to investigate its function and regulatory mechanism by AMPK in PE development. Metabolomics analysis of PE placentas was performed by gas chromatography–mass spectrometry (GC–MS). Trophoblast‐specific AMPKα1‐deficient mouse placentas were generated to assess morphology. A mouse PE model was established by Reduced Uterine Perfusion Pressure, and placental AMPK was modulated by nanoparticle‐delivered A769662. Trophoblast glucose uptake was measured by 2‐NBDG and 2‐deoxy‐d‐[³H] glucose uptake assays. Cellular metabolism was investigated by the Seahorse assay and GC–MS.PE complicated trophoblasts are associated with AMPK hyperactivation due not to energy deficiency. Thereafter, AMPK activation during placentation exacerbated PE manifestations but alleviated cell death in the placenta. AMPK activation in trophoblasts contributed to GLUT3 translocation and subsequent glucose metabolism, which were redirected into gluconeogenesis, resulting in deposition of glycogen and accumulation of phosphoenolpyruvate; the latter enhanced viability but compromised trophoblast invasion. However, ablation of AMPK in the mouse placenta resulted in decreased glycogen deposition and structural malformation. These data reveal a novel homeostasis between invasiveness and viability in trophoblasts, which is mechanistically relevant for switching between the ‘go’ and ‘grow’ cellular programs.

Pre‐eclampsia (PE) is associated with trophoblast AMPK hyperactivation, presumably due to LKB1 phosphorylation, and glucose uptake is consequently increased via trafficking of GLUT3 from the cytosol to the plasma membrane. Such translocation enhances glycolytic flux and redirects glucose metabolic intermediates into gluconeogenesis, resulting in PEP accumulation, which not only benefits cell survival but also suppresses invasion by repressing MMPs, and thus in turn modulates switching between the ‘go’ and ‘grow’ cellular programs.