ABSTRACT
@#[摘 要] P54nrb/NonO是一种能够行使多种生物学功能的核蛋白，在人体多种组织和细胞系中广泛表达并具有高度的种间保守 性，与多种疾病的发生、发展密切相关。近年来， P54nrb/NonO在多种肿瘤中的作用报道逐渐增多，然而其具体的肿瘤生物学作用 和分子机制尚不明确。P54nrb/NonO在乳腺癌、前列腺癌、结直肠癌、黑色素瘤、鼻咽癌、肺癌、血液系统肿瘤等多种肿瘤中发挥重 要作用。
ABSTRACT
Objective To explore the effects of p54nrb gene silencing on the proliferation, migration of human umbilical vein endothelial cell (HUVEC) and angiogenesis in vitro. Methods HUVECs were infected by p54nrb-specific shRNA-containing lentiviruses for 24h, then the cells were cultured in media containing puromycin until stable establishment of p54nrb-silencing HUVEC cell line. The efficiency of p54nrb gene silencing technique was determined by Western blotting, and the effects of this technique on the proliferation of HUVECs was assessed by the Cell Counting Kit-8 (CCK-8) assay. The migration of p54nrbsilencing HUVECs was measured by wound healing assay and Transwell motility assay. Vessel formation assay was used to detect the angiogenesis ability of p54nrb-silencing HUVECs. Results Western blotting showed that the expression of p54nrb protein in p54nrb-silencing HUVECs decreased significantly, implying a stable establishment of p54nrb-silencing HUVEC cell line. The CCK-8 assay revealed that knockdown of p54nrb gene promoted the proliferation of HUVECs slightly. Wound healing assay and Transwell motility assay displayed that knockdown of p54nrb significantly inhibited the motility of HUVECs. Vessel formation assay showed that knockdown of p54nrb inhibited in vitro the formation of vessel-like structures of HUVEC cells. Conclusion p54nrb significantly promote the migration and angiogenesis in vitro of HUVECs, and may be an important modulin involved in angiogenesis.
ABSTRACT
BACKGROUND: CTLA-4 (Cytotoxic T Lymphocyte associated Antigen 4, CD152) has been known as a homologue of CD28, an accessory molecule providing a key costimulatory signal for successful antigen-driven activations of T lymphocyte. Most of biochemical and cell biological characteristics of the CD152 protein remain unknown while those of CD28 have been characterized in detail. METHODS: In this study CD152 expression in both CD4+ and CD8+ PBLs was studied by using flow cytometry. And intracellular CD152 multiprotein complex was purified and used for generating antibodies recognizing proteins composing of intracellular CTLA-4 multi protein complex. RESULTS: Level of surface expression of this molecule was peaked at 2 days of PHA stimulation in flow cytometric analysis. 40~45% of PHA blast cells were CD152+ in both of two subsets at this stage and the level of expression were equivalent in both two subsets. Contrary to this surface expression, intracellular expression was peaked at day 3 and it was preferentially induced in CD8+ cells and about 60% of CD8+ cells were CD152+ at this stage. High molecular weight (> 350 kD) intacellular CD152 protein complex purified by using preparative electrophoresis were immunized into rabbits and then 3 different anti-P34PC4, anti-P34PC7 and anti-P34PC8 antibodies were obtained. Using these 3 antibodies two unknown antigens associated with intracellular CD152 multiprotein complex were found and their molecular weights were 54 kD and 75 kD, respectively. Among these, the former was present as 110 kD homodimer in non- reducing condition. CONCLUSION: It seemed that 34 kD intracellular CD152 molecule forms high molecular weight multiprotein complex at least with 2 proteins of 75 kD monomer and 110 kD homodimer.