Department of Drug Sciences, Pharmacology Section, University of Pavia, 27100 Pavia, Italy.
Department of Biotechnology and Biosciences, University of Milan-Bicocca, 20126 Milan, Italy.
Int J Mol Sci. 2023 Dec 22;25(1):206. doi: 10.3390/ijms25010206.
N-acetylcysteine (NAC), a mucolytic agent and an antidote to acetaminophen intoxication, has been studied in experimental conditions and trials exploring its analgesic activity based on its antioxidant and anti-inflammatory properties. The purpose of this study is to investigate additional mechanisms, namely, the inhibition of nerve growth factor (NGF) and the activation of the Tropomyosin receptor kinase A (TrkA) receptor, which is responsible for nociception. In silico studies were conducted to evaluate dithiothreitol and NAC's interaction with TrkA. We also measured the autophosphorylation of TrkA in SH-SY5Y cells via ELISA to assess NAC's in vitro activity against NGF-induced TrkA activation. The in silico and in vitro tests show that NAC interferes with NGF-induced TrkA activation. In particular, NAC breaks the disulfide-bound Cys 300-345 of TrkA, perturbing the NGF-TrkA interaction and producing a rearrangement of the binding site, inducing a consequent loss of their molecular recognition and spatial reorganization, which are necessary for the induction of the autophosphorylation process. The latter was inhibited by 40% using 20 mM NAC. These findings suggest that NAC could have a role as a TrkA antagonist, an action that may contribute to the activity and use of NAC in various pain states (acute, chronic, nociplastic) sustained by NGF hyperactivity and/or accompanied by spinal cord sensitization.
N-乙酰半胱氨酸（NAC）是一种黏液溶解剂和对乙酰氨基酚中毒的解毒剂，已在实验条件下和探索其镇痛活性的试验中进行了研究，其依据是其抗氧化和抗炎特性。本研究的目的是研究其他机制，即神经生长因子（NGF）的抑制和 Tropomyosin receptor kinase A（TrkA）受体的激活，该受体负责伤害感受。进行了计算机模拟研究以评估二硫苏糖醇和 NAC 与 TrkA 的相互作用。我们还通过 ELISA 测量了 SH-SY5Y 细胞中 TrkA 的自动磷酸化，以评估 NAC 对 NGF 诱导的 TrkA 激活的体外活性。计算机模拟和体外试验表明，NAC 干扰 NGF 诱导的 TrkA 激活。具体而言，NAC 破坏了 TrkA 上 Cys 300-345 之间的二硫键，扰乱了 NGF-TrkA 相互作用，并产生了结合位点的重排，从而导致它们的分子识别和空间重排丧失，这对于诱导自动磷酸化过程是必需的。使用 20mM NAC 可抑制 40％的磷酸化。这些发现表明，NAC 可以作为 TrkA 拮抗剂发挥作用，这种作用可能有助于 NAC 在由 NGF 过度活跃和/或伴随脊髓敏化引起的各种疼痛状态（急性，慢性，伤害感受性）中的活性和使用。
