Identification of a novel HLA-A2-restricted cytotoxic T lymphocyte epitope from cancer-testis antigen PLAC1 in breast cancer

Identification of cytotoxic T lymphocyte (CTL) epitopes from tumor antigens is essential for the development of peptide vaccines against tumor immunotherapy. Among all the tumor antigens, the caner-testis (CT) antigens are the most widely studied and promising targets. PLAC1 (placenta-specific 1, CT92) was considered as a novel member of caner-testis antigen, which expressed in a wide range of human malignancies, most frequently in breast cancer. In this study, three native peptides and their analogues derived from PLAC1 were predicted by T cell epitope prediction programs including SYFPEITHI, BIMAS and NetCTL 1.2. Binding affinity and stability assays in T2 cells showed that two native peptides, p28 and p31, and their analogues (p28-1Y9?V, p31-1Y2L) had more potent binding activity towards HLA-A*0201 molecule. In ELISPOT assay, the CTLs induced by these four peptides could release IFN-γ. The CTLs induced by these four peptides from the peripheral blood mononuclear cells (PBMCs) of HLA-A*02⁺ healthy donor could lyse MCF-7 breast cancer cells (HLA-A*0201⁺, PLAC1⁺) in vitro. When immunized in HLA-A2.1/K^b transgenic mice, the peptide p28 could induce the most potent peptide-specific CTLs among these peptides. Therefore, our results indicated that the peptide p28 (VLCSIDWFM) could serve as a novel candidate epitope for the development of peptide vaccines against PLAC1-positive breast cancer.     

Promoting the formation of tetramolecular G-quadruplexes under freezing condition

In the study, we demonstrated that freezing was able to facilitate the short DNA strand T(4)G(4) to self-assemble into a tetramolecular G-quadruplex with a low DNA concentration in the absence of any ligand, providing a fast approach to configure intermolecular G-quadruplexes. The fast formation of intermolecular G-quadruplexes may offer a convenient tool.     

Free radical-operated proteotoxic stress in macrophages primed with lipopolysaccharide

The free-radical-operated mechanism of death of activated macrophages at sites of inflammation is unclear, but it is important to define it in order to find targets to prevent further tissue dysfunction. A well-defined model of macrophage activation at sites of inflammation is the treatment of RAW 264.7 cells with lipopolysaccharide (LPS), with the resulting production of reactive oxygen species (ROS). ROS and other free radicals can be trapped with the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO), a cell-permeable probe with antioxidant properties, which thus interferes with free-radical-operated oxidation processes. Here we have used immuno-spin trapping to investigate the role of free-radical-operated protein oxidation in LPS-induced cytotoxicity in macrophages. Treatment of RAW 264.7 cells with LPS resulted in increased ROS production, oxidation of proteins, cell morphological changes and cytotoxicity. DMPO was found to trap protein radicals to form protein-DMPO nitrone adducts, to reduce protein carbonyls, and to block LPS-induced cell death. N-Acetylcysteine (a source of reduced glutathione), diphenyleneiodonium (an inhibitor of NADPH oxidase), and 2,2'-dipyridyl (a chelator of Fe(2+)) prevented LPS-induced oxidative stress and cell death and reduced DMPO-nitrone adduct formation, suggesting a critical role of ROS, metals, and protein-radical formation in LPS-induced cell cytotoxicity. We also determined the subcellular localization of protein-DMPO nitrone adducts and identified some candidate proteins for DMPO attachment by LC-MS/MS. The LC-MS/MS data are consistent with glyceraldehyde-3-phosphate dehydrogenase, one of the most abundant, sensitive, and ubiquitous proteins in the cell, becoming labeled with DMPO when the cell is primed with LPS. This information will help find strategies to treat inflammation-associated tissue dysfunction by focusing on preventing free radical-operated proteotoxic stress and death of macrophages.     

IL6 gene polymorphisms and susceptibility to colorectal cancer: a meta-analysis and review

A number of case-control studies were conducted to investigate the association of IL6 gene polymorphisms with colorectal cancer (CRC). However, the results were not always consistent. We performed a systematic review and meta-analysis to examine the association between the IL6 gene polymorphisms and CRC. Data were collected from the following electronic databases: PubMed, EMBASE, Web of Science, BIOSIS Previews, HuGENet, and Chinese Biomedical Literature Database, with the last report up to July 2011. A total of 17 studies involving 4 SNPs were included (16 for rs1800795, 2 for rs1800796, 2 for rs1800797, and 1 for rs13306435). Overall, no significant association of these polymorphisms with CRC was found in heterozygote comparisons as well as homozygote comparison, dominant genetic model and recessive model. In subgroup analysis, among studies using population-based controls, fulfilling Hardy-Weinberg equilibrium, or using Taqman genotyping method, we did not find any significant association. However, the rs1800795 C allele was significantly associated with reduced risk for CRC among persons who regularly or currently took NSAIDs (four studies, OR = 0.750; 95 % CI, 0.64-0.88; P = 0.474 for heterogeneity test), and with increased risk for CRC among persons who drank (one study, OR = 1.97; 95 % CI, 1.32-2.94). Individuals with the rs1800795 C allele in the IL6 gene have a significantly lower risk of CRC, but in the setting of NSAIDs use. Further studies are merited to assess the association between the IL6 gene polymorphisms and CRC risk among persons who take NSAIDs, drink or smoke, etc.     

Water-soluble o-hydroxycinnamate as an efficient photoremovable protecting group of alcohols with fluorescence reporting

Two new o-hydroxycinnamates have been prepared for photoremovable protecting groups, and their photochemistry has been investigated. The photolysis of two caged compounds can efficiently release the corresponding alcohol in aqueous solutions, and the uncaging reaction proceeds with large one-photon excitation cross sections (1919 and 1535 M(-1) cm(-1)). The uncaging process has been observed by NMR spectroscopy. The caged compounds exhibit good aqueous solubility and excellent resistance to hydrolysis in a buffer solution.     

GWAS identifies novel susceptibility loci on 6p21.32 and 21q21.3 for hepatocellular carcinoma in chronic hepatitis B virus carriers

Genome-wide association studies (GWAS) have recently identified KIF1B as susceptibility locus for hepatitis B virus (HBV)–related hepatocellular carcinoma (HCC). To further identify novel susceptibility loci associated with HBV–related HCC and replicate the previously reported association, we performed a large three-stage GWAS in the Han Chinese population. 523,663 autosomal SNPs in 1,538 HBV–positive HCC patients and 1,465 chronic HBV carriers were genotyped for the discovery stage. Top candidate SNPs were genotyped in the initial validation samples of 2,112 HBV–positive HCC cases and 2,208 HBV carriers and then in the second validation samples of 1,021 cases and 1,491 HBV carriers. We discovered two novel associations at rs9272105 (HLA-DQA1/DRB1) on 6p21.32 (OR = 1.30, P = 1.13×10⁻¹⁹) and rs455804 (GRIK1) on 21q21.3 (OR = 0.84, P = 1.86×10⁻⁸), which were further replicated in the fourth independent sample of 1,298 cases and 1,026 controls (rs9272105: OR = 1.25, P = 1.71×10⁻⁴; rs455804: OR = 0.84, P = 6.92×10⁻³). We also revealed the associations of HLA-DRB1*0405 and 0901*0602, which could partially account for the association at rs9272105. The association at rs455804 implicates GRIK1 as a novel susceptibility gene for HBV–related HCC, suggesting the involvement of glutamate signaling in the development of HBV–related HCC.     

Epigenome-wide scans identify differentially methylated regions for age and age-related phenotypes in a healthy ageing population

Age-related changes in DNA methylation have been implicated in cellular senescence and longevity, yet the causes and functional consequences of these variants remain unclear. To elucidate the role of age-related epigenetic changes in healthy ageing and potential longevity, we tested for association between whole-blood DNA methylation patterns in 172 female twins aged 32 to 80 with age and age-related phenotypes. Twin-based DNA methylation levels at 26,690 CpG-sites showed evidence for mean genome-wide heritability of 18%, which was supported by the identification of 1,537 CpG-sites with methylation QTLs in cis at FDR 5%. We performed genome-wide analyses to discover differentially methylated regions (DMRs) for sixteen age-related phenotypes (ap-DMRs) and chronological age (a-DMRs). Epigenome-wide association scans (EWAS) identified age-related phenotype DMRs (ap-DMRs) associated with LDL (STAT5A), lung function (WT1), and maternal longevity (ARL4A, TBX20). In contrast, EWAS for chronological age identified hundreds of predominantly hyper-methylated age DMRs (490 a-DMRs at FDR 5%), of which only one (TBX20) was also associated with an age-related phenotype. Therefore, the majority of age-related changes in DNA methylation are not associated with phenotypic measures of healthy ageing in later life. We replicated a large proportion of a-DMRs in a sample of 44 younger adult MZ twins aged 20 to 61, suggesting that a-DMRs may initiate at an earlier age. We next explored potential genetic and environmental mechanisms underlying a-DMRs and ap-DMRs. Genome-wide overlap across cis-meQTLs, genotype-phenotype associations, and EWAS ap-DMRs identified CpG-sites that had cis-meQTLs with evidence for genotype-phenotype association, where the CpG-site was also an ap-DMR for the same phenotype. Monozygotic twin methylation difference analyses identified one potential environmentally-mediated ap-DMR associated with total cholesterol and LDL (CSMD1). Our results suggest that in a small set of genes DNA methylation may be a candidate mechanism of mediating not only environmental, but also genetic effects on age-related phenotypes.     

Induction of autophagy contributes to the neuroprotection of nicotinamide phosphoribosyltransferase in cerebral ischemia

Recent reports indicate that autophagy serves as a stress response and may participate in pathophysiology of cerebral ischemia. Nicotinamide phosphoribosyltransferase (Nampt, also known as visfatin), the rate-limiting enzyme in mammalian NAD (+) biosynthesis, protects against ischemic stroke through inhibiting neuronal apoptosis and necrosis. This study was taken to determine the involvement of autophagy in neuroprotection of Nampt in cerebral ischemia. Middle cerebral artery occlusion (MCAO) in rats and oxygen-glucose deprivation (OGD) in cultured cortical neurons were performed. Nampt was overexpressed or knocked-down using lentivirus-mediated gene transfer in vivo and in vitro. Immunochemistry (LC3-II), electron microscope and immunoblotting assays (LC3-II, beclin-1, mammalian target of rapamycin [mTOR], S6K1 and tuberous sclerosis complex-2 [TSC2]) were performed to assess autophagy. We found that overexpression of Nampt increased autophagy (LC3 puncta immunochemistry staining, LC3-II/beclin-1 expression and autophagosomes number) both in vivo and in vitro at 2 hours after MCAO. At the early stage of OGD, autophagy inducer rapamycin protected against neuronal injury induced by Nampt knockdown, whereas autophagy inhibitor 3-methyladenine abolished the neuroprotective effect of Nampt partly. Overexpression or knockdown of Nampt regulated the phosphorylation of mTOR and S6K1 signaling pathway upon OGD stress through enhancing phosphorylation of TSC2 at Ser1387 but not Thr1462 site. Furthermore, in cultured SIRT1-knockout neurons, the regulation of Nampt on autophagic proteins LC3-II and beclin-1 was abolished. Our results demonstrate that Nampt promotes neuronal survival through inducing autophagy via regulating TSC2-mTOR-S6K1 signaling pathway in a SIRT1-dependent manner during cerebral ischemia.     

纤毛虫念珠异列虫射出胞器的超微结构观察

应用扫描电镜术和透射电镜术显示,纤毛虫念珠异列虫(Anteholosticha monilata)的射出胞器早期发生在细胞质深处,附近有不同类型的囊泡结构。成熟后射出胞器向表膜迁移,结构由不同电子密度片层的体部、结晶状的中心轴杆部和多层膜的帽部组成。受外界刺激时胞器冲破皮层射出,形态呈"蘑菇"状。据上述观察结果推测:该射出胞器具有防御作用,它可能起源于高尔基体活动产生的小泡;在亲缘关系较近的纤毛虫中,其射出胞器可能具有相似的分化特征。     

SCF(Fbw7) modulates the NFkB signaling pathway by targeting NFkB2 for ubiquitination and destruction

The NFkB/Rel family of proteins play critical roles in a variety of cellular processes. Thus, their physiological activation is tightly controlled. Recently, the NFkB2/p100 precursor has been characterized as the fourth IkB type of suppressor for NFkB. However, the molecular mechanism(s) underlying regulated destruction of NFkB2 remains largely unknown. Here, we report that, unlike other IkBs, ubiquitination and destruction of NFkB2 are governed by SCF(Fbw7) in a GSK3-dependent manner. In Fbw(7-/-) cells, elevated expression of NFkB2/p100 leads to a subsequent reduction in NFkB signaling pathways and elevated sensitivity to TNFa-induced cell death. Reintroducing wild-type Fbw7, but not disease-derived mutant forms of Fbw7, rescues NFkB activity. Furthermore, T cell-specific depletion of Fbw7 also leads to reduced NFkB activity and perturbed T cell differentiation. Therefore, our work identifies Fbw7 as a physiological E3 ligase controlling NFkB20s stability. It further implicates that Fbw7 might exert its tumor-suppressor function by regulating NFkB activity.