Novel role of p66Shc in ROS-dependent VEGF signaling and angiogenesis in endothelial cells

p66Shc, a longevity adaptor protein, is demonstrated as a key regulator of reactive oxygen species (ROS) metabolism involved in aging and cardiovascular diseases. Vascular endothelial growth factor (VEGF) stimulates endothelial cell (EC) migration and proliferation primarily through the VEGF receptor-2 (VEGFR2). We have shown that ROS derived from Rac1-dependent NADPH oxidase are involved in VEGFR2 autophosphorylation and angiogenic-related responses in ECs. However, a role of p66Shc in VEGF signaling and physiological responses in ECs is unknown. Here we show that VEGF promotes p66Shc phosphorylation at Ser36 through the JNK/ERK or PKC pathway as well as Rac1 binding to a nonphosphorylated form of p66Shc in ECs. Depletion of endogenous p66Shc with short interfering RNA inhibits VEGF-induced Rac1 activity and ROS production. Fractionation of caveolin-enriched lipid raft demonstrates that p66Shc plays a critical role in VEGFR2 phosphorylation in caveolae/lipid rafts as well as downstream p38MAP kinase activation. This in turn stimulates VEGF-induced EC migration, proliferation, and capillary-like tube formation. These studies uncover a novel role of p66Shc as a positive regulator for ROS-dependent VEGFR2 signaling linked to angiogenesis in ECs and suggest p66Shc as a potential therapeutic target for various angiogenesis-dependent diseases.     

Survival trends of cancer amongst the south Asian and non-south Asian population under 30 years of age in Yorkshire, UK

Introduction: Several studies have shown differences in survival trends between ethnic groups across adults with cancer in the UK. It is unclear whether these differences exist exclusively in the older adult population or whether they begin to emerge in children and young adults. Methods: Subjects (n = 3534) diagnosed with cancer under 30 years of age in Yorkshire between 1990 and 2005 were analysed. Differences in survival rates for diagnostic subgroups were estimated by ethnic group (south Asian or not) using Kaplan–Meier estimation and Cox regression. Results: When compared to non-south Asians (all other ethnic groups excluding south Asians) a significant increased risk of death was seen for south Asians with leukaemia (hazard ratio (HR) = 1.75; 95% confidence interval (CI) = 1.11–2.76) and lymphoma (HR = 2.05; 95% CI = 1.09–3.87), whereas south Asians with solid tumours other than central nervous system tumours had a significantly reduced risk of death(HR = 0.50; 95% CI = 0.28–0.89). This was independent of socioeconomic deprivation. Conclusion: We found evidence of poorer survival outcomes for south Asians compared to non-south Asian children and young adults with leukaemia and lymphoma, but better outcomes for south Asian children and young adults with other solid tumours. This needs to be explained, and carefully addressed in the on-going development of cancer services.     

Stable isotope resolved metabolomics analysis of ribonucleotide and RNA metabolism in human lung cancer cells

We have developed a simple NMR-based method to determine the turnover of nucleotides and incorporation into RNA by stable isotope resolved metabolomics (SIRM) in A549 lung cancer cells. This method requires no chemical degradation of the nucleotides or chromatography. During cell growth, the free ribonucleotide pool is rapidly replaced by de novo synthesized nucleotides. Using [U-¹³C]-glucose and [U-¹³C,¹⁵N]-glutamine as tracers, we showed that virtually all of the carbons in the nucleotide riboses were derived from glucose, whereas glutamine was preferentially utilized over glucose for pyrimidine ring biosynthesis, via the synthesis of Asp through the Krebs cycle. Incorporation of the glutamine amido nitrogen into the N3 and N9 positions of the purine rings was also demonstrated by proton-detected ¹⁵N NMR. The incorporation of ¹³C from glucose into total RNA was measured and shown to be a major sink for the nucleotides during cell proliferation. This method was applied to determine the metabolic action of an anti-cancer selenium agent (methylseleninic acid or MSA) on A549 cells. We found that MSA inhibited nucleotide turnover and incorporation into RNA, implicating an important role of nucleotide metabolism in the toxic action of MSA on cancer cells.     

Effects of long-term fixation on histological quality of undecalcified murine bones embedded in methylmethacrylate

While long-term fixation and storage of specimens is common and useful for many research projects, it is particularly important for space flight investigations where samples may not be returned to Earth for several months (International Space Station) or years (manned mission to Mars). We examined two critical challenges of space flight experimentation: the effect of long-term fixation on the quality of mouse bone preservation and the preservation of antigens and enzymes for both histochemical and immunohistochemical analyses, and how the animal/sample processing affects the preservation. We show that long-term fixation minimally affects standard histological staining, but that enzyme histochemistry and immunolabeling are greatly compromised. Further, we demonstrate that whole animal preservation is not as suitable as whole leg or stripped leg preservation for long-term fixation and all histological analyses. Overall, we recommend whole leg processing for long-term storage of bone specimens in fixative prior to embedding in plastic for histological examination.     

Do Exotics Homogenize or Differentiate Communities? Roles of Sampling and Exotic Species Richness

It is commonly asserted that exotic species promote the homogenization of biological communities. However, theoretical evidence implies that exotic species may often have the opposite effect, of differentiating biological communities where different exotic species become established in different areas. Furthermore, few empirical studies have directly measured the homogenizing effects of exotics. In this study, I used the plant inventories of 20 localities in the United States to measure whether exotic plant increased the similarity of those localities. I calculated Jaccard's index of similarity (JI) for exotic species and then for native species to see if the exotic JI was consistently higher when comparing two localities. I found that JI for both exotic and native species decline exponentially with increasing distance and latitude separation between localities so that localities that share many native species also tend to share many exotic species. More importantly, in nearly half of the pairwise locality comparisons (87 of 190), the (JI_exotic/JI_native) ratio is less than one, indicating a slight tendency for exotic species to differentiate rather than homogenize the localities analyzed. Also, the pattern of differentiation versus homogenization is strongly related to exotic species richness. When both localities have relatively few exotics, there is a clear tendency for exotics to produce biotic differentiation (JI_exotic/JI_native < 1). Much of this pattern seems related to the right-skewed frequency distribution of the geographic ranges of exotic species. As with native species, most exotics occur in few localities so there is a high probability that localities with low numbers of exotic species will share very few, if any, of those exotics. As exotic richness increases, the homogenizing effects become increasingly pronounced.     

三种兰科植物的保护遗传学研究初探

采用随机扩增多态 DNA(RAPD)分析研究了中国3种珍稀濒危兰科植物硬叶兜兰(Paphiopedilum micranthum Tang et Wang)、麻栗坡兜兰(P. malipoense S.C.Chen et Tsi)和独花兰(Changnienia amoena Chien)的遗传多样性与群体遗传结构.12个RAPD引物在2种兜兰中共扩增出131条带.对4个硬叶兜兰群体的检测表明其物种水平的多态条带百分率(PPB)为 71.6%,Nei 的基因多样度(h)为 0.217 1,Shannon多样性指数 (I) 为 0.330 1;4个群体的平均多样性水平为 PPB = 45.2%,h = 0.145 7,I = 0.220 4,低于远交兰花的平均水平.在总遗传变异中,群体间遗传变异占20.31%,略高于远交物种的平均水平.在物种水平上,麻栗坡兜兰的PPB为49.5%,h为0.117 4,I为0.176 4,均大大低于硬叶兜兰.对11个独花兰群体采用16个RAPD引物共扩增出119条带.物种水平PPB=76.5%,h=0.194 1,I=0.305 8;在群体水平上,上述3个指标的平均值则分别为37.2%、0.119 7和0.181 0,均低于远交兰花的平均水平.群体间的遗传变异占45.27%,遗传分化明显高于远交物种的平均水平.导致3个物种遗传多样性偏低而群体间遗传分化较高的主要原因在于人为的过度采挖和生境的片断化.研究结果为兰花保护策略和措施的制定提供了理论基础.     

The Development of Multi-epitope Vaccines: Epitope Identification, Vaccine Design and Clinical Evaluation

We have developed efficient methods for epitope identification and vaccine design. Our process for epitope selection based on the combined use of motif analyses, binding assays and immunogenicity evaluations is described. We also describe how the projected population coverage and vaccine design can be optimized. Finally, it is discussed how vaccine potency is evaluated by immunogenicity and antigenicity assays.