从p38MAPK信号通路研究复方参鹿颗粒调控较低危MDS骨髓CD34+细胞凋亡的机制

Ineffective hematopoiesis caused by excessive apoptosis of bone marrow CD34+ cells is the primary characteristic of lower risk MDS. According to recent studies, excessive apoptosis of bone marrow CD34+ cells induced by myelosuppressive cytokines via p38 MAPK signaling pathway is important in the pathogenesis of ineffective hematopoiesis in lower risk MDS. Early studies have shown that Compound Shenlu Granules(CSG) can reduce the production of myelosuppressive cytokines, promote the normal hematopoiesis, and is an effective compound to ameliorate the ineffective hematopoiesis in MDS. Based on these studies, we introduce and intend to prove the hypothesis that the CSG reduces the apoptosis of bone marrow CD34+ cells in lower risk MDS via regulating the p38 MAPK signaling pathway. We use the p38 MAPK signaling pathway as an entry point, apply randomized controlled trials,cell culture, FCM, RT-PCR, western blot, FISH and other technologies to clarify whether CSG affects the apoptosis of bone marrow CD34+ cells in lower risk MDS via regulating the p38 MAPK signaling pathway, thus restoring cellular quiesence and the preferential expansion of normal cell clones; the study also enters into the molecular mechanism of this compound regulating the apoptosis of CD34+ cells via p38 MAPK, and provides scientific evidence for the clinical treatment of lower risk MDS with "boosting marrow by warming kidney".

CD34+细胞过度凋亡导致骨髓的无效造血，是较低危MDS的主要特征。多种造血抑制因子共同通过p38 MAPK信号通路参与CD34+细胞凋亡调控，并且是引起骨髓无效造血的重要病理机制。 结合前期研究发现复方参鹿颗粒可以减少造血抑制因子的产生，促进骨髓正常造血，是改善MDS无效造血的有效复方，我们提出并论证“复方参鹿颗粒通过调控p38 MAPK信号通路抑制较低危MDS骨髓CD34+细胞凋亡”的假说。本研究以p38 MAPK信号通路为切入点，采用随机对照临床研究，结合细胞培养，通过FCM、RT-PCR、Western blot、FISH等技术，明确复方参鹿颗粒是否通过调控p38MAPK信号通路影响较低危MDS骨髓CD34+细胞凋亡，以恢复骨髓CD34+细胞静止和正常细胞克隆优势生长，并探讨该复方调控p38MAPK信号通路的分子机制，为临床“温肾益髓”治疗较低危MDS提供科学依据。

骨髓增生异常综合征(MDS)是一组恶性克隆性干细胞疾病，是老年人最常见的血液系统恶性肿瘤之一。较低危MDS是指IPSS≤1.0（低危/中危-1）MDS患者。临床约70%的MDS是以骨髓无效造血为特点的较低危MDS。现代医学却缺乏有效的治疗手段。在本研究中，我们发现，与非MDS引起的血细胞减少患者相比，较低危MDS存在骨髓CD34+的高凋亡，p38MAPK的过度磷酸化，骨髓CD34+细胞凋亡率和p-p38蛋白表达成正相关；p38MAPK信号通路中p-p53蛋白表达增加、Bcl-xl蛋白表达降低，Bcl-2的表达则无显著差异，提示过度激活的p38MAPK可能主要促进Bcl-xl降解，导致促凋亡蛋白从与Bcl-xl的复合体中释放出来，激活并寡聚化，启动凋亡。复方参鹿颗粒为改善较低危骨髓无造血的有效中药复方，尤其是对红系无效造血。经复方参鹿颗粒干预后，骨髓CD34+细胞凋亡减少，并且凋亡率下降与p-p38蛋白表达水平下降成正相关（P＜0.01）。提示复方参鹿颗粒能通过抑制p38MAPK的过度磷酸化来减少骨髓CD34+细胞凋亡。体外实验发现，复方参鹿颗粒含药血清可有效抑制TNF-α诱导的正常人骨髓CD34+细胞凋亡，下调p-p38和p-p53蛋白表达。结合FISH检测，我们发现FSG含药血清干预后形成的造血干/祖细胞集落中，来源于异常的MDS克隆比例显著低于对照组（P＜0.05）。相关性分析结果提示，p-p38蛋白表达水平和异常克隆细胞群体比例成正相关（P＜0.01）。提示复方参鹿颗粒含药血清能通过抑制较低危MDS骨髓细胞p38MAPK的过度磷酸化，恢复骨髓正常克隆细胞来源的造血干/祖细胞的优势生长。综上，本研究初步明确了温肾益髓中药复方治疗较低危MDS的作用机理，为临床治疗提供了一定的理论依据。但是复方参鹿颗粒对较低危MDS异常炎症信号调控作用的深层次机制仍需要进一步研究。