Cellular senescence in chronic pain and aging

项目摘要 慢性疼痛是一种令人衰弱的疾病，三分之一的美国人患有这种疾病，给社会带来了高昂的代价。老龄化 是慢性疼痛的主要危险因素，65岁以上的成年人中有50%患有AT 至少有一种慢性疼痛症状。不幸的是，人们还不能很好地理解为什么年龄是高血压的危险因素。 疼痛状况的发展。因此，迫切需要利用老年动物模型进行基础研究 研究年龄和疼痛之间的潜在交互作用，并最终为定制的 针对这一特定人群的治疗。随着人类和动物年龄的增长，衰老细胞在许多组织中积累。 并通过分泌引起炎症的因子来破坏组织的动态平衡，称为 衰老相关分泌表型(SASP)。有趣的是，这些SASP因素中有几个是已知的 痛诱导细胞因子在背根神经节(DRG)释放，在那里初级感觉神经元细胞 身体驻留，并驱使痛苦。令人惊讶的是，衰老细胞还没有在痛觉环路中被研究 老龄小鼠，甚至幼年小鼠周围神经损伤后。我们假设衰老的细胞；1)是 存在于老年小鼠的痛觉回路中，2)在神经损伤后进一步积累，3)导致慢性 通过分泌SASP因子的反射性和情感性疼痛反应。为了支持我们的假设，我们有 强劲的初步数据显示，与未受损伤的老年背根节相比，衰老神经元的数量增加了4倍 致未受伤的年轻DRG。此外，我们还证明了早期衰老标志物p21在 损伤(ATF3+)和未损伤(ATF3-)的TRPV1+伤害性神经元群体，提示旁分泌诱导 衰老的迹象。我们将SASP因子和疼痛介质IL6定位于这些p21+细胞，提供了证据 衰老细胞是疼痛回路中这类因素的细胞来源。最后，我们有初步数据表明 提示使用感觉性药物治疗可改善幸免神经损伤(SNI)所致的机械性痛觉异常，同时 维持幼年和老年小鼠的整体感觉功能。因此，潜在的机制是 老年小鼠损伤后疼痛超敏反应增强可能是年龄相关和损伤共同作用的结果 诱导衰老细胞。在这项建议中，我们的目标是进一步表征SNI后衰老细胞的诱导 通过分析衰老标志物在单个细胞内的共表达，确定其特定的细胞 识别和量化SASP因子在基线(未受伤)以及急性和慢性损伤后的表达 幼年和老年小鼠的时间点。进一步，我们将研究衰老细胞对神经元的贡献。 在体外使用双光子钙成像和电生理学的超兴奋性，以及在体内使用疼痛 老年小鼠与年轻成年小鼠在用特定的感觉剂治疗后的行为范式比较， 这可能选择性地诱导DRG中各亚型衰老细胞的凋亡。这项研究将是第一次 在神经病理性疼痛的临床前小鼠模型中研究细胞衰老，并具有 有可能开辟一条新的治疗途径，使用感觉剂来缓解疼痛。

Project Summary Chronic pain is a debilitating condition from which one in three Americans suffer, at a high cost to society. Aging is a major risk factor for the development of chronic pain with 50% of adults over the age of 65 suffering from at least one chronic pain condition. Unfortunately, it is not well understood why age is a risk factor for the development of pain conditions. Thus, there is an urgent need for basic research using aged animal models to examine the underlying interaction between age and pain and ultimately inform the development of tailored treatments for this specific population. As humans and animals age, senescent cells accumulate in many tissues throughout the body and disrupt tissue homeostasis by secreting factors that induce inflammation, known as senescence-associated secretory phenotype (SASP). Interestingly, several of these SASP factors are known pain-inducing cytokines that are released in the dorsal root ganglion (DRG), where primary sensory neuron cell bodies reside, and drive pain. Surprisingly, senescent cells have yet to be investigated within the pain circuit in aged mice, or even in young mice after peripheral nerve injury. We hypothesize that senescent cells; 1) are present in the pain circuit of aged mice, 2) further accumulate following nerve injury, and 3) contribute to chronic reflexive and affective pain responses through secretion of SASP factors. In support of our hypothesis, we have robust preliminary data demonstrating a 4-fold increase in senescent neurons in uninjured aged DRG compared to uninjured young DRG. Additionally, we demonstrate expression of the early senescence marker, p21, in injured (ATF3+) and uninjured (ATF3-) populations of Trpv1+ nociceptive neurons, suggesting paracrine induction of senescence. We localize the SASP factor and pain mediator, IL6, to these p21+ cells providing evidence that senescent cells are a cellular source of such factors in the pain circuit. Finally, we have preliminary data that indicate treatment with a senolytic drug improves spared nerve injury (SNI)-induced mechanical allodynia while maintaining overall sensory function in young adult and aged mice. Therefore, a potential mechanism underlying enhanced pain hypersensitivity following injury in aged mice may be the combination of age-related and injury- induced senescent cells. In this proposal, we aim to further characterize senescent cell induction following SNI by analyzing the co-expression of senescent markers within individual cells, determining their specific cellular identities, and quantifying SASP factor expression, at baseline (uninjured) and at acute and chronic post-injury time points in young and aged mice. Further, we will investigate the senescent cell contribution to neuronal hyperexcitability in vitro using 2-photon calcium imaging and electrophysiology, as well as in vivo using pain behavioral paradigms in aged mice compared to young adult mice after treatment with specific senolytic agents, which may selectively induce apoptosis of subtypes of senescent cells in the DRG. This research will be the first of its kind to investigate cellular senescence in a pre-clinical mouse model of neuropathic pain and has the potential to open a new therapeutic avenue, using senolytic agents, to alleviate pain.