- 學位論文
九孔血液細胞吞噬作用中辨識受體與蛋白質激脢C 訊息傳遞路徑之研究
Study of the Recognition Receptor and Protein Kinase C Signaling Pathway in Phagocytosis of Abalone (Haliotis diversicolor) Hemocytes
摘要
無脊椎動物的免疫機制多屬先天性免疫反應，由免疫細胞進行的吞噬作用為其中最主要的一種。。本研究以九孔 (Haliotis diversicolor) 的血液細胞為材料，研究其吞噬作用的機制。在25 °C時，附著型細胞會在血液抽離個體後30分鐘之內完全附著，並具有吞噬作用的功能。九孔的血液細胞可以吞噬酵母菌，由於吞噬作用可被1％的甘露聚醣(mannan)、褐藻澱粉(laminarin)和精氨酸-安氨酸-天門冬氨酸肽鍊(RGD peptide)所抑制，因此親醣蛋白(lectin)和整合素(integrin)可能參與九孔血液細胞吞噬的酵母菌辨識。再者，由於由血清處理過的酵母菌被吞噬率較未經處理的酵母菌高，因此血清中可能含有類似調理素(opsonin)的物質可促進吞噬。九孔血液細胞也會累積由乙醯修飾的低密度脂肪球(acLDL)，由乙醯修飾或氧化(oxidized)的低密度脂肪球為清道夫受體(scavenger receptor)的配體，清道夫受體已知會廣泛的參與免疫細胞辨認外來物。實驗中發現在九孔血液細胞上，酵母菌、甘露聚醣和乙醯修飾的低密度脂肪球間沒有競爭作用。因此，九孔細胞上可能具有清道夫受體，並可能以不同的受體接合酵母菌。在血液細胞吞噬酵母菌的細胞內訊息部分，蛋白激脢C (protein kinase C)的活化劑PMA和細胞內鈣離子載體(ionophore) A23187會促進細胞吞噬酵母菌，而蛋白激脢C、磷脂醯肌醇-3激脢(phosphatidylinositol 3-kinase)和磷脂脢C (phospholipase C)的抑制劑 Calphostin C、wortmannin和U73122則會抑制細胞吞噬酵母菌，顯示與磷脂醯肌醇-3激脢、磷脂脢C和蛋白激脢C相關的訊息途徑參與九孔血液細胞的吞噬作用。整體而言，九孔血液細胞吞噬作用與脊椎動物的巨噬細胞相似，顯示此免疫反應可能有其演化上的關連性。
並列摘要
Most invertebrates do not have acquired immunity. However, innate immunity, which is faster-reacting and functions more extensively than acquired immunity, is the major type of immunity found in invertebrates. In this study, abalone (Haliotis diversicolor) hemocytes were used to elucidate the mechanism of phagocytosis, which is the most-significant function of innate immunity. Abalone hemocytes are classified into two categories: adherent and non-adherent cells. Adherent cells can complete adhesion and phagocytose within 30 min at 25 °C. Abalone hemocytes phagocytosed yeast cells, and this process was inhibited by 1% laminarin, 1% mannan, and 0.5 mM RGD peptide. These results suggest that lectin and integrin might participate in yeast recognition. Abalone hemocytes also accumulated acetylated low-density-lipoprotein (acLDL). Modified LDL, including acLDL and oxLDL (oxidized LDL), are known ligands for scavenger receptors. Due to a competition effect, the accumulation of FITC-acLDL by hemocytes was inhibited by acLDL, but not by yeast or laminarin. These results suggest that scavenger receptors or scavenger receptor-like molecules expressed on abalone hemocytes, and the hemocytes themselves may use different receptors for yeast binding. In terms of the signaling process involved, the protein kinase C activator, PMA (phorbol 12-myristate 13-acetate), and the calcium ionophore, A23187, promoted yeast phagocytosis, whereas Calphostin C, wortmannin, and U73122, respective inhibitors of protein kinase C, phosphatidylinositol 3-kinase, and phospholipase C, inhibited phagocytosis. These results suggest that the intracellular level of calcium ions can affect phagocytosis, and signaling molecules including protein kinase C, phosphatidylinositol 3-kinase, and phospholipase C may be involved in the signaling pathway. In conclusion, the mechanism of abalone hemocyte phagocytosis is similar to the mechanism used by human macrophages. Thus, these immune functions may be evolutionarily related.
參考文獻
延伸閱讀
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