探寻反映证候本质的微观指标是证候研究的重要科学问题。唾液淀粉酶（sAA）活性比值是公认能较好反映脾虚证的微观指标之一，但临床检出率为60-70%，作为单一指标反映脾虚证的生物学信息还不够全面，故须重新认识其发生机制。根据基因拷贝数变异及GnT-Ⅰ/Ⅱ介导N-糖基化缺陷与sAA活性相关的研究基础，本项目从影响sAA活性的基因、翻译后修饰两个水平探索脾气虚证sAA活性改变的机制。本项目研究：①慢性浅表性胃炎和重症肌无力脾气虚患者酸刺激前后的AMY1基因拷贝数，sAA的活性、含量和N-糖基化程度，及这些指标与临床症候的相关性；②脾气虚大鼠酸刺激前后sAA和GnT-Ⅰ/Ⅱ的表达与活性、sAA N-糖基化程度，并观察益气健脾方药的干预作用。冀通过对sAA活性改变机制研究，获得更全面反映脾气虚证的生物学信息，并与sAA活性比值合参以提高微观指标的临床检出率，为指导临床诊疗提供理论依据和技术参考。

It is important for syndrome research that exploring and finding micro index reflecting syndrome essence commendably. The ratio of salivary alpha-amylase (sAA) after and before acid stimulation, which received as one of micro index reflecting syndrome essence well, can’t reflect comprehensively the essence of spleen qi deficiency syndrome (SQDS) as only one index and the clinical positive rate of this index is only 60-70%.So we must restudy the mechanism of sAA activity-ratio changing. According to copy number variations(CNVs) and N-glycosylation defection mediated by GnT-Ⅰ/Ⅱ associated with sAA activity closely, we will study the mechanism of sAA activity altered in SQDS from gene and posttranslational modification level of sAA. The activity and content of sAA, extent of N-glycosylation in sAA, and CNVs of AMY1 will be detected in the chronic superficial gastritis and myasthenia gravis with SQDS before and after acid stimulation. The relationship of these index and symptom information will be discussed as well. After building animal model with SQDS, the expression and activity of sAA and GnT-Ⅰ/Ⅱ, and the extent of N-glycosylation in sAA will be tested before and after acid stimulation. The intervention effect of Chinese drugs with invigorating Qi and spleen on these indexes will be observed at the same time. In order to explore the biological information reflecting the essence of SQDS better, We uncover the mechanisms of sAA activity altered in this study. Meanwhile, synthesis of these indexes above and sAA activity-ratio will be done to improve the clinical detecting rate of micro index in SQDS and provide the scientific basis for clinical diagnosis and treatment.

唾液淀粉酶（sAA）活性比值是公认能较好反映脾虚证的微观指标之一，但临床检出率为60-70%，作为单一指标反映脾虚证的生物学信息还不够全面，故须重新认识其发生机制。本课题从影响sAA活性的生化、分子两个水平探索脾气虚证sAA活性改变的机制。在慢性非萎缩性胃炎和重症肌无力脾气虚患者上研究了：① 流率、pH值、sAA总活性、总蛋白浓度、Ca2+和Cl-浓度等生化因素对脾气虚证sAA活性比值改变的影响，以及多项指标合参后脾气虚证的临床检出率；② AMY1基因拷贝数，酸刺激前后sAA的活性、含量和N-糖基化程度；③ 优化利血平致脾气虚大鼠模型及其动物体征的半定量评价方法。 与健康人比较的结果表明：①两种疾病脾气虚证不仅表现在sAA活性比值下降，还表现在柠檬酸刺激后sAA总活性、流率及pH值降低，而总蛋白浓度、Ca2+和Cl-浓度升高， sAA活性、流率和pH值进行合参后在脾气虚证的检出率较单用sAA活性比值提高约11.00%；② 两种疾病脾气虚证均发现，AMY1拷贝数较健康人未发生明显变异，N-糖基化蛋白明显缺失，生物活性较弱的高甘露糖型糖链含量升高、生物活性较强复合型糖链含量降低；③ 0.8mg/kg.d利血平颈部皮下注射9天可成功构建脾气虚模型，其客观评价方法：体征变化评分7分以上、体重下降率达40%-45%、肛温35℃左右、摄食量及饮水量趋于0，后续实验发现模型动物在酸刺激后唾液淀粉酶活性比值明显下降。 综上，脾气虚证不仅仅表现在sAA活性比值，还表现在sAA总活性、流率、pH值、总蛋白浓度、Ca2+和Cl-浓度浓度、及N-糖基化程度改变，更全面反映了脾气虚证的生物学信息； N-糖基化程度下降可能是脾气虚证sAA活性改变的主要机制之一，进一步丰富了“脾主涎”的理论内涵。