Gax inhibits perivascular preadipocyte biofunction mediated by IGF-1 induced FAK/Pyk2 and ERK2 cooperative pathways

Perivascular adipocyte (PVAC) biofunctions were closely related to cardiovascular diseases; its specific biological mechanisms remained unclear. How to adjust PVAC functions of vascular cells is an important topic. The present study was designed to investigate whether FAK/Pyk2 and ERK1/2 MAPK signaling pathways participate in PVAC functions, which is activated by insulin-like growth factor 1(IGF-1) and inhibited by Gax. PVACs isolated from perivascular adipocyte were cultured, dedifferentiated, and stimulated with 10 nM IGF-I. Cellular function experiments showed that IGF-1 promoted PVAC proliferation, adhesion, and migration. However Gax weakened IGF-1-mediated these function. Flow cytometry demonstrated that IGF-1 increased PVACs percent of S phase and decreased the percent of G0/G1 phase and apoptotic cells. While, Gax decreased the percent of S phase cells and increased those of G0–G1 phase and apoptotic cells. Western blotting and RT-PCR revealed that IGF-1 activated FAK/Pyk2 and ERK1/2 signaling pathways, upregulated the mRNA and protein expression of FAK, Pyk2, and ERK1/2, and suppressed p53 expression. Reversely, Gax lowered the expression of these signaling proteins and increased p53 expression. Therefore, IGF-1 mediated FAK/Pyk2 and ERK1/2 pathways to augment PVAC functions; Gax effectively counteracted these effects of IGF-1, repressed PVAC activities, and increased the cell apoptosis. Our findings suggested that FAK/Pyk2 and ERK1/2 cooperative activation mediated by IGF-1 is essential for PVAC functions, and Gax is a promising candidate gene to interfere with these signaling pathways and inhibit PVAC functions.     

The tumor suppressor p53: Cancer and aging

Aging, like many other biological processes, is subject to regulation by genes that reside in pathways that have been conserved during evolution. The insulin/ IGF-1 pathway, mTOR pathway and p53 pathway are among those conserved pathways that impact upon longevity and aging-related diseases such as cancer. Most cancers arise in the last quarter of life span with the frequency increasing exponentially with time, and mutation accumulation in critical genes (e.g. p53) in individual cells over a lifetime is thought to be the reason. Recently, we found that the efficiency of the p53 response to stress decline significantly with age in mice, and the time of onset of this decreased p53 response correlates with the life span of mice. Given the crucial role of the p53 in tumor prevention, this decline in p53 activity at older ages in animals could contribute to the observed dramatic increases in cancer frequency, and provides a plausible explanation for the correlation between tumorigenesis and aging in addition to the accumulation of DNA mutations over lifetime. We discuss here the coordination and communication between the p53 pathway and the IGF-1-mTOR pathways, and their possible impact on cancer and longevity.     

p53 Regulation of the IGF-1/AKT/mTOR Pathways and the Endosomal Compartment

In response to various stress signals, which introduce infidelity into the processes of cell growth and division, p53 initiates cell-cycle arrest, apoptosis, or senescence to maintain fidelity throughout the cell cycle. Although these functions are traditionally thought of as the major functions of the p53 protein for tumor suppression, recent studies have revealed some additional novel functions of the p53 pathway. These include the down-regulation of two central cell-growth pathways, the IGF/AKT-1 and mTOR pathways, and the up-regulation of the activities of the endosomal compartment. The IGF-1/AKT and mTOR pathways are two evolutionarily conserved pathways that play critical roles in regulation of cell proliferation, survival, and energy metabolism. In response to stress, p53 transcribes a group of critical negative regulators in these two pathways, including IGF-BP3, PTEN, TSC2, AMPK β1, and Sestrin1/2, which leads to the reduction in the activities of these two pathways. Furthermore, p53 transcribes several critical genes regulating the endosomal compartment, including TSAP6, Chmp4C, Caveolin-1, and DRAM, and increases exosome secretion, the rate of endosomal removal of growth factor receptors (e.g., EGFR) from cell surface, and enhances autophagy. These activities all function to slow down cell growth and division, conserve and recycle cellular resources, communicate with adjacent cells and dendritic cells of the immune system, and inform other tissues of the stress signals. This coordinated regulation of IGF-1/AKT/mTOR pathways and the endosomal compartment by the p53 pathway integrates the molecular, cellular, and systemic levels of activities and prevents the accumulations of errors in response to stress and restores cellular homeostasis after stress.     

转录因子HBP1抑制骨肉瘤细胞增殖

转录因子HBP1(HMG-box containing protein 1, HBP1)属HMG家族,是1个含有513个氨基酸残基的多肽,可与RB蛋白结合,抑制许多癌基因的表达,从而抑制细胞的增殖. 本工作构建了HBP1慢病毒表达载体,病毒包装后感染骨肉瘤细胞U2OS细胞.Western印迹结果显示,感染细胞cyclin D1、c-Myc蛋白质水平降低 ,而p53蛋白质水平增加;Real-time PCR检测cyclin D1、c-Myc及p53 mRNA水平与蛋白质检测结果一致. HBP1表达载体（pEFBOS-HBP1）和报告基因载体（含 cyclin D1、c-Myc或p53 promoter）共转染U2OS细胞后的荧光素酶分析发现,HBP1可抑制cyclin D1、c-Myc启动子转录激活,促进p53启动子转录激活,并且这种抑制或激活作用具有HBP1剂量依赖性. 以细胞代龄群体倍增值PD （population doubling）为指标测试细胞生长结合软琼脂集落形成实验证明,HBP1慢病毒感染的U2OS细胞生长速度减缓、集落形成能力下降. 上述结果提示,HBP1可能通过调控细胞增殖相关基因的表达抑制骨肉瘤细胞的增殖.     

Autophagy of vascular smooth muscle cells in atherosclerotic lesions

Autophagy genes were first identified in the yeast system and some of their mammalian orthologues have also been characterized. Increasing lines of evidence indicate that various intracellular proteins, including G proteins, mammalian target of rapamycin (mTor) and Pl3K/Akt/PKB, of transmembrane signaling pathways are involved in the regulation of autophagy genes. We have recently discovered autophagy as a mechanism of cell death in atherosclerotic vascular smooth muscle cells (VSMCs). Tumor necrosis factor-alpha (TNF-alpha), insulin-like growth factor-1 (IGF-1), and 7-ketocholesterol can regulate the expression of autophagic genes, including microtubule-associated protein 1 light chain-3 (MAP1LC3) and Beclin 1, through Akt/PKB and c-jun N-terminal signal pathways in VSMCs. However, the balance between cell death and survival of VSMCs in the fibrous cap of atherosclerotic plaques appears to best correlate with plaque instability. Understanding the underlying cellular and molecular mechanisms of autophagy can provide key insights into the cell death machinery of atherosclerotic diseases.