基于"脾胃相关"理论研究萎缩性胃炎黏膜微环境中CAMKⅡ信号通路的响应及健脾理气和胃法的干预作用

Gansu Province is the region for high incidence of chronic atrophic gastritis (CAG) and gastric cancer. Based on clinical practice of treating CAG and blocking gastric cancer with Traditional Chinese Medicine effectively and pathogenesis of CAG, combined with'Spleen and stomach associated'theory and our preliminary research, the study hypothesis was established, which is CaMKⅡ signal transduction pathways play an important role in the gastric mucosa microenvironment of CAG and the therapy with Spleen-strengthening, Qi-combing and Stomach-reconciling therapy on it has some possible intervention functions. So CAG model rats are study object, then detect the activity of Ca2+/ CaM-PK II and the dynamic expression law of CaM mRNA in the gastric mucosa at different time points in CAMK II signal pathway by using immunohistochemistry, Western blotting and RT-PCR technique. On this basis, the relationship between the expression change of CaM-PK II and PCNA protein, Caspase-3 protein, NF-κB, SS, PGE2 and MOT will be explored further. At the same time, the model rats are treated with Xiangsha Liujunzi decoction. Then through correlation analysis, further reveal the mechanism of CAG on the basis of CAMK II signal pathway key molecular and the scientific connotations of Spleen-strengthening, Qi-combing and Stomach-reconciling therapy on CAG, and provide theoretical support for clinical application of this therapy and for developing drugs against CAG.

甘肃省为慢性萎缩性胃炎（CAG）及胃癌高发区。本研究基于中医药干预CAG可有效阻断胃癌发生的临床实践和CAG脏腑病机规律，以"脾胃相关"理论为基础并结合前期工作，建立"钙调素蛋白激酶Ⅱ（CAMKⅡ）信号通路在CAG胃黏膜微环境中的响应及健脾理气和胃法干预的可能作用"工作假说，以CAG大鼠为研究对象，采用免疫组化、免疫印迹和PCR技术检测CAMKⅡ信号通路中Ca2+/CaM-PKII活性与CaM mRNA在胃黏膜组织中的动态表达规律，在此基础上进一步观察CaM-PK II变化与PCNA 蛋白、Caspase-3蛋白、NF-κB、SS、PGE2、MOT等之间的关系，并应用以健脾理气和胃法为指导的香砂六君子汤进行干预，通过相关性分析，拟从CAMKⅡ信号通路关键分子角度揭示CAG的发生机制以及健脾理气和胃法对其干预作用，并为该治法的临床应用和开发防治CAG优势中成药的关键技术提供理论支撑。

项目按照计划实施并取得预期成果。我们结合临床应用香砂六君子汤或健脾理气和胃相关方药防治CAG的体会，探讨了基于“脾胃相关”CAG病机规律及中医药防治思路；同时结合名老中医应用健脾理气和胃方药防治CAG临床验案，验证“健脾理气和胃”治法的临床指导意义。.研究结果：（1）模型组大鼠一般生存状况较差，平均每日摄食量和体重增加量、胃排空率和胃蛋白酶活性降低，胃液总酸和游离酸含量降低；胃组织SDH活性降低，LDH活性升高；与模型组比较，香砂六君子汤能改善大鼠生存状况，提高平均每日摄食量和体重增加量以及胃排空率、胃蛋白酶活性、SDH活性而降低LDH活性，并能提高胃液总酸和游离酸含量。（2）模型组大鼠胃组织NF-κB p65基因和蛋白表达、IL-1β、IL-6、TNF-αmRNA和含量升高，而IL-10 mRNA和含量降低；与模型组比较，香砂六君子汤组大鼠NF-κB p65基因和蛋白表达水平、IL-1β、IL-6 、TNF-αmRNA和含量降低，而IL-10 mRNA和含量升高，说明该复方可通过下调NF-κB p65基因和蛋白表达，调节促炎/抑炎因子平衡分泌。（3）模型组大鼠胃组织PGE2、GAS、SS、MOT水平下降，说明CAG病程中存在胃肠激素分泌紊乱；与模型组比较，香砂六君子汤可提高PGE2、GAS、SS、MOT水平，提示该复方具有调节胃肠激素分泌和胃动力，增加胃黏膜血流量而修复胃黏膜的作用。（4）模型组大鼠胃组织游离Ca2+浓度降低，CaM、CaMKⅡ、HSP70基因和蛋白表达降低，HIF-1α、PCNA基因和蛋白表达升高，说明胃组织因炎性、萎缩等病变因HIF-1α、PCNA基因和蛋白高表达，Ca2+/CaM-CaMKⅡ信号通路关键分子低表达诱发；经治疗后与模型组比较，香砂六君子汤组大鼠胃组织游离Ca2+浓度、CaM、CaMKⅡ、HSP70基因和蛋白表达升高，而HIF-1α、PCNA基因和蛋白表达下调，且均以高剂量作用显著，提示该复方具有激活Ca2+/CaM-CaMKⅡ信号通路关键分子而促进胃平滑肌蠕动，调节胃肠激素正常分泌，同时可下调HIF-1α、PCNA基因和蛋白表达，抑制细胞炎性反应和异常增殖的途径防治CAG。.培养青年教师3名，博士和硕士生各1名（均在读），指导2名本科生参加课外科技活动，发表学术论文6篇。