基于胶质细胞及相关免疫炎症机制研究α7-nAChRs在骨关节炎疼痛中的作用

Pain is the defining clinical presentation of osteoarthritis (OA). Current management of OA pain falls short of patient needs both in terms of providing adequate and sustained pain relief and in terms of undesirable adverse events and health risks. Relief from OA pain is what drives people to seek out care, but available treatments provide modest relief at best. OA pain pathways are subject to complex levels of control and modulation, both in the periphery and in the central nervous system. Ongoing clinical and basic research suggest inflammation and sensitization play a prominent role in the mechanisms of OA pain. It has been long known that nicotinic natural products have analgesic effects, but the discovery of the remarkable high analgesic potency of the frog alkaloid epibatidine prompted extensive research on nicotinic compounds as potential novel pain treatments. However, the identity of the nicotinic acetylcholine receptors (nAChRs) that are responsible for the spinal control of nociception is currently unknown. Recent studies suggest that the nicotinic anti-inflammatory pathway may have important clinical implications, as treatment with nicotinic agonists can potentially modulate the production of pro-inflammatory cytokines from immune cells via interactions at α7-nAChRs. Thus, α7-nAChRs may serve as a crucial link between inflammation and OA pain and could represent a pharmacological target for potential pain management in OA. Our previous data have demonstrated that ① In primary cultured mouse astrocytes, pretreatment with nicotine suppressed LPS-induced astrocyte activation, as evidenced by both decreased production of TNF-α and inhibition of extracellular regulated kinase1/2 (Erk1/2) and p38 activation in astrocytes, and these effects were also reversed by the α7-nAChR-selective antagonist methyllycaconitine (MLA). ②Nicotine exerts a protective effect on H2O2-induced astrocyte apoptosis and glial cell-derived neurotrophic factor downregulation, and this effect was abolished by MLA. The underlying mechanisms might involve alleviation of mitochondrial membrane potential loss, stabilization of the Bax/Bcl-2 balance, and inhibition of cleaved caspase-9 activity through α7-nAChR activation. Those data imply that α7-nAChR-mediated inhibition of astrocyte activation and alleviation of astrocyte dysfunction might be important mechanisms underlying OA pain. We also found that systemic administration of nicotine alleviated monosodium iodoacetate (MIA)- induced joint degradation. The protective effects of nicotine were abolished by administration of the α7-nAChR-selective antagonist. In primary cultured rat chondrocytes, pretreatment with nicotine suppressed both p38, Erk1/2 and c-Jun-N-terminal kinase (JNK) mitogen-activated protein kinases (MAPK) phosphorylation and phosphorylated nuclear factor-kappa B (NF-κB) p65 activation induced by MIA- or IL-1β, and these effects were also reversed by MLA. Activation of α7-nAChRs is closely associated with the pathophysiological mechanism of OA pain. Therefore, the objective of the present study was to determine whether nicotine, acting at α7-nAChRs, could regulate glia-mediated inflammation and sensitization mechanisms, and may provide some insight into changes in cholinergic signaling associated with OA pathogenesis and present valuable information for new drug therapies. We test hypothesis about α7-nAChRs providing a new therapeutic strategy for treatment of OA pain. Nicotinic agonists that specifically target the α7-nAChR might be developed and serve as potential therapeutic agents for OA pain and other neuropathic pain.

骨性关节炎（Osteoarthritis, OA）疼痛病理生理机制复杂，免疫炎症反应学说及痛觉敏化学说一直是OA疼痛主要病理机制研究和关注的热点。流行病学统计显示：吸烟或许是OA的相关因素，烟碱乙酰胆碱受体（Nicotinic acetylcholine receptors, nAChRs）在其中发挥重要作用。α7-nAChRs是近年来多种疾病的研究热点，胆碱能抗炎通路中的关键受体。申请人前期研究结果发现：激活α7-nAChRs可抑制LPS诱导的星形胶质细胞活化；激活α7-nAChRs可抑制氧化应激诱导的星形胶质细胞的凋亡；激活α7-nAChRs可以显著改善OA大鼠模型膝关节软骨的退变。因此，我们提出假说：“α7-nAChRs可通过胶质细胞及相关免疫炎症机制在OA疼痛中发挥重要作用。”阐明α7-nAChRs在OA疼痛发生、发展中的机制，为研发理想的多潜能关节保护剂和镇痛药物提供新的靶标。

骨性关节炎（Osteoarthritis, OA）疼痛病理生理机制复杂，烟碱乙酰胆碱受体（Nicotinic acetylcholine receptors, nAChRs）或许在其中发挥重要作用。我们检索了PubMed，Embase和Cochrane图书馆数据库从建库到2020年8月31日的相关研究。共纳入了32项研究，涉及近30万名参与者，发现吸烟与膝骨关节炎的疼痛呈负相关，随后，我们应用野生型（WT）小鼠建立MIA OA模型，连续28天腹腔给予烟碱（Nic）可以显著改善OA小鼠机械性超敏反应，软骨退化和MMP-9的上调，选择性α7-nAChRs阻断剂甲基牛扁亭碱（MLA）可拮抗上述Nic的作用。在RAW264.7细胞和小鼠原代骨髓来源的巨噬细胞中，Nic显著抑制脂多糖（LPS）诱导的MMP-9的产生。机制有可能为Nic通过α7-nAChRs调节了磷脂酰肌醇3-激酶/蛋白激酶B（PI3K/ Akt）信号通路，及B细胞核因子κ轻链增强子（NF-κB）核转位有关。第三，我们发现OA患者膝关节软骨组织中α7-nAChRs表达减少，软骨细胞凋亡与自噬失衡。在体研究中，敲除（KO）α7-nAChRs加重软骨退化和自噬向凋亡的转化，离体研究发现，敲除（KO）α7-nAChRs同样加重自噬向凋亡的转化，Nic能改善自噬与凋亡间的平衡。机制与Beclin-1，p62，LC3 (I and II)，Bcl-2，Bax，cleaved caspase-3，caspase-3等信号通路有关。近年来研究表明，神经胶质细胞尤其是小胶质细胞和星形胶质细胞在慢性疼痛的发生发展中起重要作用。OA等引起的慢性疼痛状态下，脊髓星形胶质细胞比小胶质细胞呈现出更持久的活化，该活化与慢性疼痛行为密切相关。我们发现α7-nAChR选择性激动剂PNU-282987在神经毒素MPP+处理的原代星形胶质细胞中有抗凋亡作用，在神经毒素MPP+处理的SH-SY5Y细胞中，Nic和PNU-282987预处理通过激活α7-nAChRs/MAPK/p53轴也显示出神经保护性抗凋亡作用。在微小RNA干扰α7‐nAChRs的表达后，其抗凋亡作用减弱。因此，我们阐明了α7-nAChRs在OA疼痛发生、发展中的机制，为研发理想的多潜能关节保护剂和镇痛药物提供新的靶标。