致死剂量γ射线照射小鼠淋巴结淋巴细胞凋亡动力学及其与相关蛋白表达的关系

用原位末端标记、原位杂交和碱磷酶免疫组化技术,观察致死剂量γ射线照射小鼠淋巴结淋巴细胞的凋亡动力学变化,及其与Bax、Bcl-2和Bcl-XL表达的关系。结果表明,照射后24h淋巴结淋巴细胞凋亡指数迅速升高,在6—12Gy范围内与照射剂量呈正比,≥15Gy照射后变化不明显。6Gy照射后24h,淋巴细胞凋亡达高峰,尔后开始降低;然而直至照射后6个月和12个月,凋亡指数仍明显高于对照组。6Gy照射后24h,淋巴细胞Bax蛋白表达即出现升高,当剂量为12Gy时达到峰值,15Gy和20Gy照射后未观察到这种剂量效应关系。而Bcl-2和Bcl-XL蛋白表达在照射后24h明显下降。bax和bcl-2mRNA的表达显示出相似趋势。以上结果显示,≤12Gy照射后细胞凋亡是淋巴细胞的重要死亡方式。照射后Bax的上调及Bcl-XL的下调在致死剂量照射引起的淋巴细胞凋亡调控中起重要作用。     

Impact of aging and life-long calorie restriction on expression of apoptosis-related genes in male F344 rat liver

Aging enhances apoptosis of hepatocytes under normal physiological conditions and increases the susceptibility to apoptosis of hepatocytes, whereas chronic calorie restriction (CR) suppresses the age-enhanced susceptibility to apoptosis. To clarify the subcellular mechanisms of age-associated dysregulation of apoptosis and the effects of CR, we analyzed the expression of genes promoting apoptosis (p53, Fas receptor, Fas ligand, TNF receptor 1, TNFalpha, Bax, TGF beta 1) and genes preventing apoptosis (Bcl-2 and Bcl-XL) in the livers of 3-, 6-, 15-, and 24-month-old male F344 rats that were either fed ad libitum or subjected to a 30% reduction in food intake (CR). After the age of 6 months, expression of p53, Fas receptor, Fas ligand, and TNFalpha mRNAs was up-regulated with aging. CR suppressed this age-enhanced p53 and Fas receptor mRNA expression, but expression of the other genes was not altered significantly by aging or CR. Expression of Fas receptor in hepatocytes, as detected immunohistochemically, increased with age, but CR suppressed age-accelerated Fas receptor expression. Our findings suggest that TNF ligand/TNF receptor family signaling, particularly Fas receptor expression, is important in age- and CR-modulated apoptosis of hepatocytes. Hepatocytes that were immunoreactive for p53 had slightly increased with aging, suggesting that p53 may mediate the age-enhanced up-regulation of Fas receptor in hepatocytes.     

Exploring the selectivity of inhibitor complexes with Bcl-2 and Bcl-XL: A molecular dynamics simulation approach

B-cell lymphoma 2 (Bcl-2) family proteins are potential drug targets in cancer and have a relatively flat and flexible binding site. ABT-199 is one of the most promising selective Bcl-2 inhibitors, and A-1155463 selectively inhibits Bcl-XL. Although the amino acid sequences of the binding sites of these two inhibitors are similar, the inhibitors selectively bind the target protein. In order to determine the origin of the selectivity of these inhibitors, we conducted molecular dynamics simulations using protein-inhibitor modeling. We confirmed that ASP103 of Bcl-2 is a key residue and that hydrogen bonding between ASP103 and ABT-199 confers the Bcl-2 selectivity of this inhibitor. For Bcl-XL selectivity, the secondary structure of α-helix 3 is a key factor. PHE105, SER106, and LEU108 in the loose α-helix 3 interact with A-1155463 to confer Bcl-XL selectivity. These findings provide important insights into the molecular mechanisms of selective inhibitors of Bcl-2 family proteins.