運用互補性去氧核糖核酸（cDNA）微陣列技術來評估雷公藤對僵直性脊椎炎患者周邊血液中之單核球細胞所表現出來的各種細胞激素及和細胞凋亡及發炎反應有關的各種分子的基因表現的影響

Application of cDNA Microarray to Measure the Effect of Tripterygium Wilfordii Hook F (TWH) on the Mononuclear Cytokine, Apoptotic, and other Inflammatory Gene Expressions in Patients with Ankylosing Spondylitis

雷公藤在過去的許多研究中已經被證實對許多包括類風濕性關節炎在內的發炎性關節疾患具有相當程度的療效。在本篇研究裡，我們使用了cDNA微陣列來偵測雷公藤對於僵直性脊椎炎患者周邊血液中的單核球細胞所表現出來的和發炎反應有關的各種分子的基因表現的影響。我們將周邊血液裡的單核細胞從兩位僵直性脊椎炎患者及一位正常健康人身上分離出來，加入脂多醣體以刺激其產生免疫反應後，再加入雷公藤的萃取液一同培養。在加入雷公藤的之前及之後，我們分別向這些周邊血液中的單核球細胞進行mRNA的抽取。接著，這些mRNA會被轉錄成cDNA。藉著將這些轉錄而來的cDNA和已經預先選定、作好、且按照一定的排列順序貼附在微陣列薄膜上的DNA殖株進行雜交的方式，我們將可以對由那些單核球細胞所表現出來的各種和發炎反應有關的分子的基因表現進行定量。實驗的結果顯示雷公藤對於包括干擾素、腫瘤壞死因子、介白質－－8、以及許多細胞間黏著分子等在內的和發炎反應有關的各種分子在僵直性脊椎炎患者身上的基因表現可能具有顯著的抑制作用。除此之外，我們還發現雷公藤可以對一種在發炎性關節疾病的關節組織破壞過程中伴演很重要角色的一種蛋白質分解酶－－基質金屬蛋白酶的基因表現產生明顯的抑制作用。儘管只有少數的研究對象，本篇研究提供了在免疫致病機轉上的可能證據來支持雷公藤對於僵直性脊椎炎患者周邊血液單核球細胞內的許多發炎性細胞激素及其它和發炎反應有關的分子的基因表現具有強烈的抑制作用的說法。至於雷公藤對於僵直性脊椎炎患者的臨床療效則有待進一步的研究。

Tripterygium wilfordii Hook F (TWHf) has been shown as an effective therapy to many arthritic diseases including rheumatoid arthritis (RA). In this study, we used cDNA microarray to evaluate the effect of TWHf on the gene expressions of peripheral blood mononuclear cytokines and other inflammation-related molecules. Peripheral blood mononuclear cells (PBMCs) from two patients with ankylosing spondylitis (AS) and one healthy subject were stimulated with lipopolysaccharides (LPSs), and then co-cultured with a TWHf extract. Extractions of mRNA from these mononuclear cells were carried out before and after addition of TWHf separately. After these extracted mRNA were transcribed into cDNA, a quantification of gene expressions of assorted molecules in interest were performed by way of a hybridization to those selected target DNA clones spotted on the microarray membrane. The results showed gene expressions of assorted inflammatory cytokines, including interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukine-8 (IL-8), and of many adhesion molecules, including integrin-,β5, selectin-P, selectin-P ligand, CD47, and lymphocyte-function-αssociated-αntigen-3 (LFA-3 or CD58), were declined with a probable or possible significance after treated with TWHf extract among AS patients. Besides, TWHf was also shown to exert a prominent inhibitory effect on the gene expression of an important proteolytic enzyme, matrix metalloproteinase, which is vital to tissue destruction in many chronic arthritic diseases. This preliminary case control study reconfirms that TWHf is a very potent herbal remedy to suppress mononuclear expressions of many inflammatory cytokines and pro- inflammatory molecules. Further clinical investigations are required to clarify the efficacy of TWHf on AS patients.