基于“脾胃为气血生化之源”理论探讨逆灸对化疗后骨髓抑制骨髓基质细胞及其分泌的细胞因子的影响

The bone marrow suppression after chemotherapy on cancer patients severely limits the application of the chemotherapy ,affects the curative efficacy, and has more complications.At present,although the western medicine can use G-CSF、GM-CSF to promote the blood cells recovered,but there are many problems such as the high cost,heavy burden,side effects, especially can not be a preventive measure. Traditional chinese medicine advocates “prevention before disease onset”,and thinks that “spleen and stomach being the foundation of acquired constitution and the origin of producing qi and blood”.Deficiency of qi and blood、spleen and stomach weakness is the most basic syndrome of the bone marrow suppression after chemotherapy,and the moxibustion possesses the advantages of being easily accepted by the patients.So,the research about using moxibustion to prevent and treat the myelosuppression is a subject needed to be solved at present. Our previous study found that acupuncture can affect ECs through bFGF and VEGF；And acupuncture serum has significant effect on the proliferation, migration and adhesion of bone marrow-derived EPCs cultured in vitro. EPCs is the priority support cell for HSCs in the bone marrow microenvironment. Our research,using the method of preventive moxibustion, by intervening the spleen and stomach, and by injecting cyclophosphamide to make bone marrow suppression model on mice，aims to detect the peripheral bloods cells、the number and cell cycle of HSCs、the number of EPCs、osteoblasts 、osteoclastsand adipocytes in bone marrow microenvironment and many cytokines which secreted, then analysis the effect of preventive moxibustion on the bone marrow microenvironment for the myelosuppression systematicly.At the same time, to deepen the understanding of “spleen and stomach being origin of producing qi and blood”theory.

肿瘤患者化疗后的骨髓抑制严重限制了化疗应用，西医虽可应用G-CSF、GM-CSF等促进血细胞回升，但不宜预防性给药。中医提倡“未病先防”，认为：脾胃为后天之本、气血生化之源，气血亏虚、脾胃虚弱是其最基本证型，而艾灸具有易被患者接受的优点。因此，探索逆灸防治本病是目前亟待解决的课题。我们前期研究发现针灸能通过bFGF、VEGF影响ECs、针灸血清对体外培养的骨髓源性EPCs功能有显著影响。而EPCs是骨髓微环境中HSC的优先支持细胞。基于前期研究，本项目旨在运用逆灸的方法，通过干预脾胃，以注射环磷酰胺制成的化疗骨髓抑制小鼠为模型，综合检测骨髓外周血各种血细胞、HSC数量及周期、EPCs数量、成骨细胞、破骨细胞、脂肪细胞等基质细胞及其分泌的细胞因子等指标，系统分析逆灸对化疗后骨髓抑制骨髓微环境的影响，为充分发挥其临床效果提供科学依据；同时对深化认识“脾胃为气血生化之源”理论具有重要科学价值。

肿瘤患者化疗后骨髓抑制严重限制了化疗的应用。中医提倡“未病先防”，而艾灸具有易被患者接受的优点。针对化疗后患者气血亏虚、脾胃虚弱的基本证型，结合前期研究，本实验基于“脾胃为气血生化之源”理论探讨了逆灸对化疗后骨髓抑制骨髓基质细胞及其分泌的细胞因子的影响，同时，比较胃经下合穴及脾胃背俞穴的效应差异，为临床优化选穴提供科学依据，并分析比较逆灸及艾灸作用特点，为灸法的介入时机提供科学支持。方法：选取雄性KM小鼠60只，将其随机分为6组，包括空白组、模型组、逆灸足三里组、逆灸脾胃俞组、艾灸足三里组、艾灸脾胃俞组，进行不同干预后，运用全自动血细胞分析仪、ELISA法、流式细胞术、骨髓基质细胞培养、Western blot、Real time PCR等方法检测各组小鼠外周血象指标以及TPO、EPO、GM-CSF、骨髓基质细胞OPG、PPARγ、RUNX2、RANKL蛋白表达及其分泌的细胞因子。结果表明：（1）艾灸脾胃俞穴位可逆转化疗后HGB、红细胞的下降，在一定程度上影响TPO水平进而调节血小板数量；逆灸时选择足三里穴可提升WBC数量，而逆灸脾胃俞穴可提升化疗后骨髓中的EPO水平的表达，提高HSCs、EPCs数量。（2）艾灸脾胃俞可显著提高VEGF、b-FGF基因表达水平；艾灸组显著上调OPG、RUNX2的蛋白表达，降低RANKL蛋白表达水平；艾灸足三里可上调PPARγ蛋白表达；逆灸组可上调RUNX2蛋白表达水平，使RANKL蛋白表达水平显著降低；逆灸足三里可下调PPARγ蛋白表达水平。与中医“治未病”、“脾胃为气血生化之源”理论相一致，说明在化疗前后选择适当的时机运用灸法干预，可以减轻化疗后骨髓抑制，达到比较好的治疗效果。