以差異性蛋白質體學分析PMA 和A23187 對人類嗜鹼性白血球細胞之影響   全文下載

Comparative Proteomics on Resting and Activated Human Basophilic Leukemic KU812 upon Stimulation with PMA plus A23187

人類嗜鹼性白血球細胞對過敏和免疫調節作用在近年來逐漸受到重視。本研究利用免疫促進劑（PMA 和A23187）去刺激人類嗜鹼性白血球細胞，觀察活化前後其細胞內所有蛋白質之變化，並採用二維電泳，配合介質輔助雷射脫附離子化四極棒式飛行時間分析儀（MALDI-Q-TOF）和蛋白質比對軟體，來進行差異性蛋白質體分析。實驗結果發現經PMA 和A23187 處理後，嗜鹼性白血球細胞會大量產生第二型（Th2）細胞激素如介白素4 、5 、13（IL-4, IL-5, IL-13），使反應趨向於過敏方向，並同時鑑定到2 個正向調節和15 個負向調節蛋白質。進一步以蛋白質轉漬法分析異質核醣核蛋白K（HNRPK）和烯醇?（ENO1）的產量，發現與二維電泳上蛋白質的表現量趨勢相吻合。這些蛋白質主要存在於細胞質、細胞核和細胞質膜。經生物資訊軟體分析比對後，這群蛋白質大部分和免疫、過敏失調和癌症形成有關。"

Basophil activation has been implicated in allergy reaction and immune modulation. Signaling transduction, mediator release and morphology change of basophils have been largely deciphered. This study examined the feasibility of using proteomic strategy to investigate the whole cell proteomic changes in KU812 basophils following activation with phorbol myristate acetate (PMA) plus ionophore (A23187). The two-dimensional electrophoresis (2D) coupled with matrix-assisted laser desorption/ionization-quadrupole/ time-of-flight (MALDI-Q-TOF) was selected for studying comparative proteome differences; and cytokine mRNA expressions were monitored. The differentially displayed proteins were further validated by 1D- and 2D-Western blotting for protein expressions. IL-4, IL-5, and IL-13 mRNAs in KU812 were significantly induced; a phenomenon similar to the allergy reaction. The protein expressions of heterogeneous nuclear ribonucleoprotein K (HNRPK) and a-enolase (ENO1) were shown a good correlation according to the results from the 2D, and 2D-Western blots. Since the majority of the identified proteins were associated with immunological, inflammatory disorders and tumor genesis, this study in defining the proteomic changes in basophilic cells that occur in response to treatment with PMA plus A23187 can serve as biomarkers for characterization of in vitro immuno-stimulation.