基于整合素信号通路探讨补肾填精法治疗慢性再生障碍性贫血的分子机制

我们在前期研究课题中，初步认为慢性再生障碍性贫血（CAA）的造血异常与整合素VLA亚家族黏附分子表达异常相关，我们推测此结果可能与由整合素介导的信号途径异常有关，为印证该假设并从不同角度验证前期研究的可靠性，同时研究补肾填精法对之影响，我们设计了此课题。本研究主要采用钙荧光指示剂法检测信号转导细胞内Ca2+水平变化；Western Blot法检测整合素VLA亚家族黏附分子、整合素介导信号途径的关键酶类、整合素连接激酶ILK及电压门控钾通道Kv1.3蛋白表达；采用RT-PCR法检测整合素VLA亚家族黏附分子、整合素介导信号途径的关键酶类、ILK及Kv1.3通道的基因mRNA表达水平，筛选出与本病发病相关的靶分子及信号转导通路异常的机理，进而在蛋白、酶、基因和分子水平上阐明CAA病机的生物学基础，并为补肾填精中药调节CAA患者造血黏附信号转导通路提供科学依据。

In our previous research, it preliminarily shows that the hematopoiesis abnormal of chronic aplastic anemia (CAA) is in connection with abnormal expression of integrin VLA subfamily adhesion molecule. We speculate that this result may be led by the signal pathway abnormalities of the whole integrin mediate. To prove the hypothesis and verify the reliability of previous studies from different aspects, and study the impact of methods of tonifying kidney and replenishing essence as while, hence, we design this project. This study mainly used method of calcium fluorescent indicator to detect he signal transduction changes of intracellular Ca2+ level; use Western Blot method to detect the integrin VLA subfamily adhesion molecule, key enzymes of the signal pathway for the integrin mediate, integrin connecting with the kinase ILK and expressed by the voltage gating with potassium channel Kv1.3; apply RT-PCR method to detect the integrin VLA subfamily adhesion molecule, key enzymes of the signal pathway for the integrin mediate, expression level of gene mRNA for ILK and channel Kv1.3, to screen out mechanism of target molecule and signal transduction pathway abnormities relating to pathogenesis, so as to demonstrate the biological basis of CAA pathogenesis based on the proteins, enzymes, genes and the molecular level, and provide a scientific basis for using traditional Chinese medicine which tonifies kidney and replenish essence to regulate haemopoiesis adhesion signal transduction pathway for the patients.