臺灣博碩士論文加值系統

在菌血症病人的體內，內毒素由革蘭氏陰性菌的細胞壁上釋出，大量存在於血液中，活化免疫細胞。其中，巨噬細胞受內毒素活化後，釋出種種發炎性物質，諸如 cytokines、TNF-α、arachidonate metabolites、 nitric oxide 等。而 TNF-α 被認為是發炎反應的誘導物，nitric oxide 則是引發菌血性休克的決定性物質。本實驗以這二項介質為指標，探討腎上腺素作用劑對小鼠腹腔巨噬細胞受 LPS 活化的影響。
我們以體外試驗的方式，將引發至腹腔的巨噬細胞抽出培養，處理各種腎上腺素作用劑，觀察 TNF-α 及 nitric oxide 釋出量之變化情形。在腎上腺素作用劑中，β-選擇性的作用劑在抑制 LPS 活化巨噬細胞的效果較α-選擇性者佳，而其中β2-選擇性的作用劑又比β1-選擇性者有較好的效果。在β-腎上腺素作用劑的訊息傳遞中，PKA 的活化為抑制 TNF-α及 nitric oxide 釋出的必須步驟。在 LPS 活化巨噬細胞的機轉中，已知tyrosine kinase 之參與，而在 38KDa 及 42KDa 的蛋白質可見到 tyro-sine phosphorylation 的變化。因此，我們探討 isoprenaline 經由β-adrenoceptor 將訊息傳入細胞之後，與 LPS 活化巨噬細胞的此一 tyro-sine kinase 路徑之間的相互作用情形。至於 PKA 透過干擾 LPS 活化tyrosine kinase ，使 protein tyrosine phosphorylation 的作用將下游訊息阻礙的過程中，蛋白質 38KDa 及 42KDa 可能參與其中，但仍須進一步的研究加以證明。另外，α-adrenoceptor 作用劑：phenylephrine只抑制 LPS 釋放 TNF-α 而不抑制 nitric oxide 之產量，由此推論 LPS釋放 TNF-α 及 nitric oxide 的訊息傳遞機轉，可能不盡相同。
另外，我們以小鼠腹腔巨噬細胞研究氯化銅與甲基汞對內毒素作用的影響，發現：(1) 氯化銅前處理具抑制 LPS-induced NO 的效果，若同時給予則失去此作用。另以 genistein 探討氯化銅與 tyrosine kinase 路徑的關係，則認為氯化銅有可能經由與 genistein 不同的方式來干擾tyrosine kinase 的活化，尚須更多研究證實；(2)甲基汞在小鼠腹腔巨噬細胞同樣具有抑制 LPS-induced NO 效果，且是經由 PKA 的活化而作用。

LPS is one component of G(-) bacteria's outer membrane. In
sepsis patient, LPS was released to the circulatory system and
then markedly activated immune cells. The activated macrophages
would produce inflammatory substances, such as cytokines, arachi-donate metabolites, nitric oxide, etc. In the paper, we attempted to investigate the effects of catecholamines on the production of TNFα and nitric oxide (NO) from the isolated mouse peritoneal macrophages activated by LPS.
The results showed that α1-agonist, phenylephrine had no
effect on NO production but markedly inhibited TNF-α production. β-agonist, isoprenaline was the most potent in antagonizing the production of both NO & TNFα and β1 -agonist, dobutamine appeared to be less potent than β2 -agonist, metaproterenol in this aspect. By means of SDS gel electrophoresis and the specific antibodies against phosphotyrosine and ERK2, we were able to demonstrate that isoprenaline inhibited LPS in the phosphory- lation of 38KDa & 42KDa proteins. H89, an inhibitior of protein kinase A attenuated the inhibitory action of isopreterenol in the
production of NO & TNFα as well as in the protein tyrosine
phosphorylation.
In this study, we have also demonstrated that methyl mercury
apparently mimicked isoprenaline in the inhibition of LPS
mediated by the activation of PKA but cuppric chloride might
exhibit this effect through the tyrosine kinase pathway.