|
中文摘要
鏈球菌致熱性外毒素B (SPE B) 是一種cysteine protease，所有A群鏈球菌 (GAS) 都攜帶致熱性外毒素B之基因，在A群鏈球菌侵入性感染的病程中，致熱性外毒素B扮演一個重要的致病角色。根據實驗室先前的研究結果顯示，致熱性外毒素B能夠引發U937細胞的凋亡結果，同時降低U937細胞的吞噬能力。冬蟲夏草是中國傳統的中草藥，被認為具有調節免疫能力的功效。本論文係利用人類單核細胞株U937細胞及A群鏈球菌感染模式來探討冬蟲夏草之免疫調節機轉，我們證實冬蟲夏草水提液確實可以活化U937細胞之吞噬能力，若是冬蟲夏草水提液經加熱或trypsin酵素處理後，並不會對冬蟲夏草的功能造成影響。由於致熱性外毒素B能夠降低U937細胞的吞噬能力，我們進一步利用A群鏈球菌感染小鼠的模式以及外給U937細胞SPE B的條件下，探討冬蟲夏草是否能夠活化吞噬能力。實驗結果顯示，冬蟲夏草能夠回復致熱性外毒素B對U937細胞所引起之吞噬能力下降。另一方面，冬蟲夏草與U937細胞作用十個小時能觀察到其吞噬能力上升，但作用六小時則不具活化效果。根據這樣的結果，我們取冬蟲夏草與U937細胞共同作用之培養液進一步與U937細胞共同培養六小時後測試其吞噬能力，實驗結果發現U937細胞受冬蟲夏草刺激會釋放某些因子，而這些因子具有增強吞噬能力、可以緩解致熱性外毒素B的抑制效果。當細胞培養液經加熱處理，其活化吞噬能力便明顯降低，因此推測U937細胞釋放增強吞噬能力的因子，可能是如細胞激素之類的蛋白質。我們發現冬蟲夏草確實能夠活化U937細胞在TNF-��, IL-12 p35, IL-12 p40以及IFN-�袕o些細胞激素mRNA的表現，在抗體中和試驗中，培養液活化吞噬能力的效果可被IFN-�蛂BIL-12及TNF-�悝凗撽狶磻謘C利用小鼠動物模式進一步探討胃管餵食冬蟲夏草之小鼠，在背部氣囊感染A群鏈球菌後，其存活率有回復增加的結果，根據菌血量以及氣囊回抽液菌數，餵食冬蟲夏草的小鼠較控制組之菌數為低，而且在控制組的小鼠能夠觀察到A群鏈球菌菌血症及細菌散佈到脾臟、肝藏及腎臟，而餵食冬蟲夏草的小鼠則沒有這些的情形。在血清中AST和ALT的表現，控制組小鼠感染A群鏈球菌後較蟲草組為高。這些實驗結果顯示冬蟲夏草對A群鏈球菌的感染具有修復的效果，冬蟲夏草的成份中，具有能夠調節免疫反應的特殊因子，特別是可以活化U937細胞的吞噬能力及刺激細胞激素的產生、提高感染A群鏈球菌小鼠的存活率，對於冬蟲夏草的免疫調節能力值得更深入研究。
|
|
Abstract:
Streptococcal pyrogenic exotoxin B (SPE B) is a cysteine protease and carried by every strain of Streptococcus pyogenes. SPE B has been shown to play an important role in invasive group A streptococcal (GAS) infection. We previously showed that SPE B could induce apoptosis and reduce the phagocytic activity in U937 cells. Cordyceps sinensis (CS) is a Chinese herbal medicine, which is known to possess immunomodulatory activity. In this study, we use human monocytic U937 cells and GAS infection mouse model to study the mechanisms of immunomodulatory effects of CS. We had demonstrated the elevation of phagocytic activity of U937 cells by CS extract. Heat or trypsin treatment did not affect the enhancing effect of CS. Whether the phagocytic enhancing properties of CS may exert modulatory effect on the pathogenicity of GAS, especially the role played by SPE B, was further investigated. Results showed that SPE B-mediated suppression of U937 phagocytic activity could be abrogated by CS extract. The enhancing effect was detectable at 10 h after coincubation with CS, but not at 6 h. The conditioned medium containing factors released from CS extract-treated U937 cells augmented the phagocytic activity and also abrogated SPE B-mediated suppression of phagocytosis. Moreover, heat treatment eliminated the enhancing effect of conditioned medium, suggesting that the protein products, such as cytokines, released by U937 cells caused augmentation of phagocytosis. CS can induce TNF-��, IL-12 p35, IL-12 p40 and IFN-�� mRNA expression in U937 cells. In antibody neutralization test, the enhancing effect of conditioned medium on phagocytosis was suppressed by anti-IFN-��, anti-IL-12 and anti-TNF-�� antibodies. Further studies in the mouse model showed that force-feeding with CS was able to increase the survival rate after air pouch inoculation with GAS. Bacterial numbers in the air pouch exudates from CS-treated mice were lower than those from control mice. Bacteremia and bacterial dissemination to spleen, liver and kidney could be observed in mice infected with GAS, but not in those pretreated with CS. The levels of AST and ALT in control mice sera after GAS infection were higher than CS-pretreated mice. These results suggest that CS confers protective effect in the animal model of GAS infection. The CS contains factors that can modulate the immune responses, especially phagocytic activity of U937 cells and cytokine expression. The CS extract can increase survival rate and decrease bacteremia in GAS infection. The immunomodulatory activity of CS is worthy for further investigation.
|
