抑癌素Oncostatin M 與白血抑制因子Leukemia inhibitory factor 對於類風濕性關節炎的致病機轉以及嗎啡止痛效果之探討

Abstract

細胞激素 (cytokine) 是一種由多胜肽鏈 (polypeptide) 所組成的小分子，主要是由免疫細胞所分泌，在人體內扮演著重要訊息傳遞的角色。近年來也發現細胞激素和許多疾病的發生有著密切的關係。在本研究中，擬探討細胞激素在類風濕性關節炎 (rheumatoid arthritis) 以及疼痛所扮演的角色為何。 類風濕性關節炎是一種罕見的自體免疫疾病，目前發病的原因仍然不清楚，但和基因以及環境因素有關。在臨床上常可以觀察到許多免疫細胞聚集在關節處，活化破骨細胞 (osteoclast) 並刺激滑膜纖維母細胞 (synovial fibroblast) 分泌大量的細胞激素諸如白細胞激素1 (interleukin-1) 、白細胞激素6 (interleukin-6) 以及A型腫瘤壞死因子 (TNF-α)，最終導致關節腫脹以及破壞組織。先前的報導中發現抑癌激素 (oncostatin M) 會大量表現在患有類風濕性關節炎的關節滑膜中 (synovium)。從我們的實驗可以發現，抑癌激素可以刺激類風濕性關節炎病人的滑膜纖維母細胞進而分泌大量的胎盤生長因子 (placenta growth factor)，胎盤生長因子同時也被認為是類風濕性關節炎重要的致病因子。經由給予Jak以及PI3K的抑制劑，我們發現抑癌激素的作用可以被消除。透過啟動Jak/STAT以及PI3K/Akt這兩條訊息傳遞路徑，抑癌素可以造成STAT磷酸化 (phosphorylation) 並進行細胞核內的轉移，進入到核內的STAT可以直接連接到胎盤生長因子的啟動子上進而增加胎盤生長因子的轉錄作用。這些證據顯示在類風濕性關節炎病人的關節中，大量表現的抑癌素可以作為發展藥物的標的。 對於嚴重型的類風濕性關節炎患者而言，關節持續劇烈的疼痛往往是最大的問題。當傳統的止痛藥物無法抑制疼痛時，臨床上有時會給予嗎啡 (morphine) 來舒緩疼痛。嗎啡一直以來都被視為極佳的強烈止痛劑，但長期使用嗎啡會使身體產生耐受性 (tolerance)，此時便需要給予更高的劑量才能達到相同的止痛效果。但是更高的劑量往往伴隨著更強的副作用，包括便秘、噁心、頭暈以及呼吸抑制。在我們的研究成果中發現，長期注射嗎啡的老鼠，其脊髓中會大量表現抑癌素以及白血抑制因子 (leukemia inhibitory factor)，而這兩種細胞激素可以藉由增加鉀離子通道的外流來加強嗎啡的止痛效果。除此之外，抑癌素以及白血抑制因子也可以藉由抑制神經微膠細胞 (microglia) 大大的延緩嗎啡耐受性的產生。我們更進一步比較因嚴重癌症疼痛而接受高劑量鴉片類藥物治療並產生耐受性的病患，以及健康的受試者當作對照組。結果顯示當病患產生鴉片類藥物耐受性時，其腦脊髓液 (cerebral spinal fluid) 中的白血抑制因子表現量較正常受試者來的高，此外，隨著鴉片類藥物的使用劑量增加，白血抑制因子的表現量也同樣增加。根據這些結果，我們認為藉由提高抑癌素以及白血抑制因子的表現可以增強嗎啡止痛效果，並可作為抑制鴉片類藥物產生耐受性的目標。 綜合以上的實驗結果，我們可以了解到細胞激素參與在許多不同的病理反應當中。藉由更深入的研究細胞激素在各個疾病中所扮演的角色，我們提供了一個未來藥物研發的方向。

Cytokines are small molecular polypeptides that mainly secreted by immune cells. They play a pivotal role in signaling transduction and are involved in a wide range of biological activities. In addition to their role in immune system, cytokines are also linked with numerous diseases. In this thesis, we tried to (1) examine the role of cytokines in the pathogenesis of rheumatoid arthritis (RA), and (2) to elucidate the effect of cytokines in pain management. Rheumatoid arthritis (RA) is an autoimmune disease. The cause of RA is still obscure, although a genetic component and environmental factors appear likely. In RA patients, numerous of immune cells accumulate in the joint, leading to activate osteoclast and stimulate synovial fibroblast (SF) to secret lots of cytokines such as IL-1β, IL-6 and TNF-α eventually lead to cartilage erosion and joint destruction. Previous studies have shown that the levels of oncostatin M (OSM) are upregulated in RA patients’ synovial fluid, but how OSM participates in RA is still unclear. Here we found that OSM could increase the mRNA and protein levels of placenta growth actor (PLGF) in RA synovial fibroblast (RASF) derived from RA patients. The enhancement effect of OSM was antagonized by the addition of Jak and PI3K inhibitors. Treatment of OSM in RASF could activate Jak/STAT and PI3K/Akt signaling pathways, activated-STAT will further translocate into nucleus and bind to the promoter region of PLGF. The accumulation of PLGF may contribute to inflammation and angiogenesis of RA. These results revealed that OSM could be a good target for developing drugs to treat RA. To the patients who have moderate to severe RA, persistent pain is the most annoying problem. Clinicians sometimes use opioids to reduce severe pain when traditional painkiller not able to control pain. However, long-term use of opioids is usually confined by the development of tolerance, which means a higher dose is required to achieve the same analgesic level. Higher doses of opioids are associated with more serious side effects including constipation, nausea, dizziness and respiratory depression. In our second and third studies, we found that long-term treatment of morphine could increase the levels of OSM and leukemia inhibitory factor (LIF) in rat spinal cord. The upregulated OSM and LIF could potentiate the analgesic effect of morphine by enhancing the effect of opioid on potassium outward current in dorsal horn neurons. Moreover, OSM and LIF could markedly attenuate the development of tolerance by suppressing microglia activation. We further collected sample from patients with different degrees of opioid tolerant cancer patients and compared with opioid naive surgical patients. It was found that the levels of LIF were significantly higher in opioid tolerant patients, there was a positive correlation between daily equivalent opioid dose and LIF expression. These results indicate that LIF might be a novel target to inhibit the development of opioid tolerance. In summary, our results implicate novel mechanism of OSM and LIF in the pathogenesis of RA and in the development of opioid tolerance. These chemokines may be a good drug target for developing new drugs.