| 摘要: | 1.風濕性關節炎是一個長期的發炎性的病變，通常會造成滑液纖維破壞。滑液纖維母細胞是關節中重要的細胞，當發炎物質刺激時會造成細胞分泌更大量的發炎物質。而peptidoglycan (PGN) 是格蘭氏陽性菌細胞壁的主要成分，在之前的研究中指出與免疫反應及cytokines and chemokines的產生有密切的關係，然而PGN是否會造成滑液纖維母細胞的發炎反應並不明朗。
從我們的結果發現給予PGN於滑液纖維母細胞中會造成IL-6的產生，而給予PGN會造成TLR2 mRNA表現增加，另外給予TLR2 siRNA也能抑制PGN所誘導的IL-6表現。
分析PGN經由那些訊息傳遞路徑參與在IL-6的表現當中，應是一個治療風濕性關節炎的指標。由我們的結果中發現給予FAK mutant、PI3K、Akt與AP-1抑制劑可以抑制由PGN所增加的IL-6表現，另外當給予PGN後，也能增加FAK、PI3K、Akt的磷酸化，而處理PGN後也能促使滑液纖維母細胞c-jun磷酸化，並且進入細胞核中。除此之外，AP-1 promoter的冷光活性測試與c-jun結合到AP-1 element之能力皆在處理PGN後而增加。最後給予FAK mutant、PI3K與Akt抑制劑也都能抑制細胞核中c-jun之磷酸化、AP-1 promoter的冷光活性測試與c-jun結合到AP-1 element之能力。
從我們的研究結果中發現，PGN會藉由TLR2 receptor/FAK/PI3K/Akt之訊息傳遞，增加人類滑液纖維母細胞IL-6表現量，使關節炎更加嚴重。而了解這些訊息傳遞路徑將可以了解格蘭氏陽性菌感染造成關節炎病變的過程以提供臨床上治療上的可能性。
2.脂肪組織是一個人體的重要能量儲存組織，脂肪組織提供一些身體功能包含了許多器官的保護作用，這些器官有：皮膚、身體的器宮、心臟及關節。而脂肪細胞在身體之內除了產生脂肪之外還有許多的功能，包括合成及分泌發炎介質、補體、訊息傳遞分子、成長因子及黏著因子，由這些資訊發現脂肪組織或細胞與發炎的作用有相當大的關連性。
Adiponectin是脂肪組織主要分泌的一種赫爾蒙蛋白，並且調控著能量恆定的功能。在風濕性關節炎與退化性關節炎的病人中，也發現了adiponectin的大量表現。而在關節炎病人的軟骨遭受破壞時，MMP-3扮演著一個重要的角色，因此我們試圖研究adiponectin的大量表現，與MMP-3是否相關。
從我們qPCR、western blot與ELISA的研究結果中發現，將人類初代軟骨細胞處理adiponectin後能增加MMP-3的表現量。而我們利用AdipoR1的siRNA能有效的抑制adiponectin誘導的MMP-3表現，但是AdipoR2則沒有效果。當我們前處理5’-AMP-activated protein kinase (AMPK) (araA and compound C)、p38 (SB203580)與NF-κB (PDTC and TPCK)的抑制劑，也能有效的抑制adiponectin所產生的效果。從我們的結果中也發現處理adiponectin後，的確會活化AMPK、p38與NF-κB的訊息路徑。
從我們的結果發現adiponectin藉由AdipoR1/AMPK/p38/NF-κB之訊息傳遞路徑增加MMP-3的表現量，並且加速關節炎病人病症的惡化。
1.Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by robust infiltration of leukocytes into the synovium, resulting in hyperplasia of the synovial lining, progressive cartilage destruction, and erosion of the underlying bone. Synovial fibroblasts mediate joint destruction in RA by producing chemokines that facilitate the expansion and invasion of synovial fibroblasts into the adjacent tissue, and the regulation of these events has been a primary target of therapeutic intervention in RA.
Peptidoglycan (PGN), the major component of the cell wall of Gram-positive bacteria, activates the innate immune system of the host and induces the release of cytokines and chemokines.
We investigated the signaling pathway involved in IL-6 production stimulated by PGN in rheumatoid arthritis synovial fibroblasts. PGN caused concentration- and time-dependent increases in IL-6 production. PGN mediated IL-6 production was attenuated by TLR2 small interfering RNA and nucleotide-binding oligomerization domain 2 small interfering RNA.
Pretreatment with PI3K inhibitor (Ly294002 and wortmannin), Akt inhibitor, and AP-1 inhibitor (tanshinone IIA) also inhibited the potentiating action of PGN. PGN increased the focal adhesion kinase (FAK), PI3K, and Akt phosphorylation. Stimulation of rheumatoid arthritis synovial fibroblast cells with PGN increased the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to the AP-1 element on the IL-6 promoter. PGN mediated an increase in the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to AP-1 element was inhibited by Ly294002, Akt inhibitor, and FAK mutant. Our results suggest that PGN increased IL-6 production in human synovial fibroblasts via the TLR2 receptor/FAK/PI3K/Akt and AP-1 signaling pathway.
2.Adipose tissue is a ubiquitous tissue, which can be found as a structural component of many organs of the human body, including the skin, gut, heart, and joints. Adipocyte has the ability to synthesize and release proinflammatory molecules, complement factors, signaling molecules, growth factors, and adhesion molecules, suggesting an integrated function of adipocytes in tissue inflammation.
Adiponectin is a protein hormone secreted predominantly by differentiated adipocytes and is involved in energy homeostasis. The adiponectin is significantly higher in synovial fluid of patients with osteoarthritis and rheumatoid arthritis.
Matrix metalloproteinases-3 (MMP-3) may contribute to the breakdown of articular cartilage during arthritis. We investigated the signaling pathway involved in MMP-3 caused by adiponectin in human chondrocytes. Adiponectin increased the secretion of MMP-3 in cultured human chondrocytes, as shown by qPCR, western blot and ELISA analysis. Adiponectin-mediated MMP-3 expression was attenuated by AdipoR1 but not AdipoR2 siRNA. Pretreatment with 5’-AMP-activated protein kinase (AMPK) inhibitor (araA and compound C), p38 inhibitor (SB203580) and NF-κB inhibitor (PDTC and TPCK) also inhibited the potentiating action of adiponectin. Activations of p38, AMPK and NF-κB pathways after adiponectin treatment were demonstrated.
Taken together, our results provide evidence that adiponectin acts through AdipoR1 to activate p38 and AMPK, resulting in the activations of NF-kB on the MMP-3 promoter and contribute cartilage destruction during arthritis. |
