血脂异常及动脉粥样硬化不同痰瘀证候血浆蛋白差异表达谱的研究

目的通过研究血脂异常及动脉粥样硬化不同痰瘀证候(包括痰证、血瘀证、痰瘀互阻证)血浆蛋白质的表达差异,寻找不同痰瘀证候的特异性标志蛋白群。方法收集146例北京地区汉族血脂异常及动脉粥样硬化中医痰证、血瘀证、痰瘀互阻证和非痰非瘀类证候患者及37名健康对照的血浆样品,采用二维凝胶电泳(two-dimensional polyacrylamide gel electrophoresis,2-DE)方法进行电泳和图像扫描及差异度分析,找出不同痰瘀证候的差异蛋白质表达谱;从凝胶上切取相应差异蛋白质斑点,酶切后于Q-TOF-MS仪上进行二级质谱分析,进一步采用Fisher判别法对差异蛋白进行典型性分析,筛选出能够显著区分不同痰瘀证候可能的标志蛋白群。结果在组间匹配变异度超过100%的11个斑点中除2个斑点未确定外,发现有7种不同的蛋白质。经对这9种不同的蛋白质进行典型性分析后发现:能区分血脂异常及动脉粥样硬化患者与正常人群间的血浆标志蛋白群可能是结合珠蛋白前体和纤维蛋白原γ链;能区分血脂异常及动脉粥样硬化患者痰瘀类证候与非痰瘀类证候的血浆标志蛋白群可能是纤维蛋白原β链和载脂蛋白AI前体;能区分痰证与痰瘀互阻证可能的标志蛋白群是纤维蛋白原γ链、白蛋白和载脂蛋白AI前体;能区分痰证与瘀证可能的标志蛋白群是结合珠蛋白前体、肾上腺髓质素结合蛋白前体、白蛋白和补体C4;能区分瘀证与痰瘀互阻证可能的标志蛋白群是白蛋白和肾上腺髓质素结合蛋白前体。结果还发现上述可能的标志蛋白的表达差异可随痰瘀证候间不同的传变方式而有不同的变化规律。结论采用蛋白质组学方法获得了可显著区分血脂异常及动脉粥样硬化不同痰瘀证候及痰瘀类证候与非痰瘀类证候间可能的标志蛋白群的特异性组合,及在功能蛋白质水平上有关痰瘀证候的传变主要是由痰致瘀进而形成痰瘀互结的初步证据。

Differential Plasma Protein Profiles in Patients with Hyperlipidemia & Atherosclerosis of Different Patterns of Phlegm-stasis Syndrome

Objective To investigate the differential plasma protein profiles in patients with hyperlipidemia atherosclerosis （HA） of different patterns of phlegm-stasis syndrome （PSS） for seeking their biomarker proteins. Methods Two-dimensional gel electrophoresis and gel screening graphical analysis were performed on plasma proteins got from 146 patients; corresponding protein spots were fetched from the gel for two-stage mass-spectrometric analysis by quadruple time-of-flight mass spectrometry; then the differential proteins for PSS were discriminated by Fisher discriminate analysis. Results Excepting two uncertain proteins, 7 differential proteins were screened out from the 11 differentially expressed plasma protein spots with variability over 100% in the inter-block matching. Classic analysis found that haptoglobin precursor and fibrinogen γ chain were possibly the plasma biomarker proteins for HA; fibrinogen β chain and apolipoproteinA-I precursor were that set apart PSS from non-PSS; fibrinogen γ chain, albumin and apolipoproteinA-I precursor were for phlegm syndrome; haptoglobin precursor, adrenomedullin binding protein precursor, albumin and complement component C4 were for stasis syndrome; albumin and adrenomedullin binding protein precursor were for the phlegm-stasis mutual blocking syndrome. Moreover, the above mentioned expressions of possible marker proteins had their own special rule of changing in the transforming progress of PSS. Conclusion This study reported, for the first time, the existence of evident variation of functional protein constitution in different patterns of PSS, and definite compatibility being detected in some functional proteins, which may be the marker proteins for making diagnosis and prognosis of PSS in HA. Besides, preliminary proof for the transformation of PSS has gained at the functional protein level.