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中文摘要
長久以來，人們認為中樞神經系統(central nervus system)是一個免疫特權區(immunologically privileged site) ，主要是因為中樞神經系統缺乏淋巴組織，且其血管外層還包圍了一層由星狀細胞(astrocyte)所組成的血腦障壁(blood-brain barrier, BBB)，以及FasL(CD95L) 及TGFβ (transformation factor-β) 的表現所致。但在過去的研究中發現，週邊給予LPS (lipopolysaccharide)刺激後所產生的細胞激素(cytokine) 如TNFα (tumor necrosis factor-α)、IL-1β(interleukin-1β)、IL-6 (interleukin-6)等，可活化下視丘-腦垂體-腎上腺軸(hypothalamic-pituitary-adrenocortical axis, HPA axis)，進而引起生理現象的改變如發燒。這些結果顯示出，中樞神經系統與週邊免疫系統間的交流是存在的。一般認為腦部因為有血腦障壁的存在，所以週邊發炎所產生的細胞激素是不會進入到腦部的，於是細胞激素是如何影響中樞神經系統的疑問就產生了。在我們的研究中，我們利用完全佛蘭氏佐劑(complete Freund’s adjuvant，CFA)或完全佛蘭氏佐劑混poly(Glu60 Ala30 Tyr10 , GAT )給予老鼠週邊刺激，引起鼠腦表現細胞激素。以免疫化學染色的方法我們發現原本在腦中不表現的TNFα及COX-2 (cyclooxygenase type 2)，於完全佛蘭氏佐劑或完全佛蘭氏佐劑混和GAT刺激後可表現在微血管上，也發現IL-1β只表現在免疫完全佛蘭氏佐劑混和GAT的鼠腦中。相反的原本表現在腦部神經束細胞的IL-1α及ependymal cell的IL-6經此刺激後卻會下降。利用半定量的反轉錄-聚合連鎖反應證明了腦部有自行產生細胞激素的能力。我們亦利用會持續表現β-galactosidase 的M4 E. coli 當作偵測血腦障壁通透性的工具，結果發現在免疫完全佛蘭氏佐劑混和GAT的老鼠中，其血腦障壁通透性最大的時間點是48小時；而免疫完全佛蘭氏佐劑的老鼠，其血腦障壁通透性最大的時間點是96小時。且血腦障壁通透性增加的現象可被NS398 (COX-2抑制劑)及TNFα的抗血清所抑制。另外也由免疫化學染色發現在處理過NS398後，鼠腦中的TNFα表現量會降低；而在處理過TNFα抗血清後鼠腦的COX-2表現量會降低；我們也發現在免疫完全佛蘭氏佐劑混和GAT的老鼠在48小時時會產生發燒現象，此發燒的現象可被NS398或是indomethacin (COX-1及COX-2抑制劑)所抑制，但卻不能被TNFα的抗血清所抑制，顯示出cyclooxygenase 可參與發燒反應。綜合上述結果，我們認為週邊的發炎反應除了可引起腦部微血管細胞表現TNFα、IL-1β之外，在微血管的TNFα可促使COX-2的產生，進而導致血腦障壁通透性的增加，而IL-1β也透過COX-2的作用引起發燒反應。
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英文摘要
The CNS (central nervous system) of mammal is considered to be an immunologically privileged site because it lacks lymphatic drainage, the brain is separated from the blood compartment by the blood-brain barrier (BBB). The brain also expresses FasL and TGFβ that can down-modulate the immune response. However, peripheral administration of lipopolysaccharide has been found to stimulate the hypothalamic- pituitary- adrenocortical axis to induce fever and influence behavior. This result showed that communication between peripheral immune system and CNS exists. It is general believed that cytokines produced in peripheral tissue can not cross the BBB because of the unique features of brain microvascular endothelial cells that wrapped by astrocyte. In this study, we reported that the peripheral stimulation with complete Freund’s adjuvant (CFA) induced the cytokine expression in brain. With immunohistochemical staining, TNFα and cyclooxygenase type 2 (COX-2) were detected on brain microvascular vessels post CFA or poly (Glu60 Ala30 Tyr10 , GAT ) in CFA injection. IL-1βwas detected in microvascular vessels only in mice injected with GAT/CFA. On the contrary, the constitutive expression of IL-6 on brain microvascular vessels and IL-1α on brain neuron bundle were found to decrease. The cytokine induction was also confirmed by semiquantitative reverse-transcription polymerase chain reaction, indicating its de novo synthesis. Using the M4 E. coli that constitutive expresses β-galactosidaseas as a tracer to quantitate the increased permeability of BBB, we found that the vasopermeability of BBB increased maximum at 96 h post CFA injection and 48 h post GAT/CFA injection. The increase of BBB permeability was inhibited by either NS398 (COX-2 inhibitor) or anti-TNFα antiserum. Using immunohistochemistry staining, the TNFαexpression was inhibited by NS398 treatment while COX-2 induction was blocked by anti-TNFαadministration. Furthermore, peripheral administration of GAT/CFA were found to induce fever at 48 h. The fever was not inhibited by anti-TNFαantiserum, but could be inhibited by NS398 or indomethacin (COX-1 and COX-2 inhibitor). Based on the above data, we conclude that peripheral inflammation stimulates TNFα or IL-1βexpression on brain microvascular vessels. The TNFαwould further cause COX-2 induction that involved in the increase of BBB permeability. The IL-1βinduction might be responsible for the fever that can be inhibited by COX inhibitor.
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