Cellular senescence in chronic pain and aging

项目摘要 慢性疼痛是一种使人衰弱的疾病，三分之一的美国人患有这种疾病，社会为此付出了高昂的代价。老化 是慢性疼痛发展的主要危险因素，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，人们还不清楚为什么年龄是一个危险因素， 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相互作用，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 并通过分泌诱导炎症的因子破坏组织稳态， 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， 在背根神经节（DRG）中释放的疼痛诱导细胞因子，其中初级感觉神经元细胞 身体存在，并驱动疼痛。令人惊讶的是，衰老细胞尚未在疼痛回路中进行研究。 老年小鼠，甚至周围神经损伤后的年轻小鼠。我们假设衰老细胞：1） 存在于老年小鼠的疼痛回路中，2）在神经损伤后进一步积累，和3）有助于慢性疼痛。 反射性和情感性疼痛反应通过分泌SASP因子。为了支持我们的假设，我们 稳健的初步数据表明，与对照组相比， 未受伤的年轻DRG此外，我们还证实了早期衰老标志物p21在人乳腺癌中的表达。 Trpv 1+伤害感受神经元的损伤（ATF 3+）和未损伤（ATF 3-）群体，表明旁分泌诱导 衰老我们将SASP因子和疼痛介质IL 6定位于这些p21+细胞，提供了证据， 衰老细胞是疼痛回路中这些因子的细胞来源。最后，我们有初步的数据， 用衰老清除药物治疗可改善保留神经损伤（SNI）诱导的机械性异常性疼痛， 维持年轻成年和老年小鼠的整体感觉功能。因此，潜在的机制 老年小鼠损伤后疼痛超敏性增强可能是年龄相关和损伤的组合， 诱导衰老细胞。在这个提议中，我们的目标是进一步表征SNI后的衰老细胞诱导 通过分析单个细胞内衰老标记物的共表达，确定它们的特异性细胞内标记物， 在基线（未受伤）和急性和慢性损伤后， 在年轻和老年小鼠中的时间点。此外，我们将研究衰老细胞对神经元的贡献。 使用双光子钙成像和电生理学进行体外兴奋过度，以及使用疼痛进行体内兴奋过度 用特定衰老清除剂处理后老年小鼠与年轻成年小鼠相比的行为模式， 其可以选择性地诱导DRG中衰老细胞亚型的凋亡。这项研究将是第一个 研究神经病理性疼痛的临床前小鼠模型中的细胞衰老， 有可能开辟一条新的治疗途径，使用衰老清除剂，以减轻疼痛。

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.