Mechanisms of cancer pain at the level of the peripheral nerve system

背景：越来越多的证据表明内皮素-1（ET-1）在动物和人类外周伤害性信号传导中起作用。然而，ET-1的伤害性作用的机制尚未完全了解。本研究采用细胞内钙离子测定和组织化学方法研究了ET-1对培养的成年小鼠背根神经节（DRG）神经元对辣椒素反应的影响。用数字图像分析系统检测培养的DRG细胞经fura-2 AM负载后的[Ca^<2+>] i图像。结果ET-1（10 nM）可使辣椒素（10 nM）引起的细胞内[Ca^2+] i从87.6 +-11.6 nM增加到414.8 +-62.3 nM;抑制内皮素A受体可明显抑制ET-1的增强作用。此外，非选择性PKC抑制剂显著抑制ET-1的增强作用。肉豆蔻酰标记的膜渗透性PKCD V1.2抑制肽也显著抑制ET-1的增强作用。在免疫细胞化学研究中，大多数TRPV 1阳性神经元中发现PKCD免疫反应。在TRPV 1阳性神经元中，ET-1可使PKCD免疫反应性从胞浆转移到细胞膜上。结论ET-1可增强DRG神经元对辣椒素的反应，其机制可能与PKC的依赖性有关。我们的发现可能会导致一种新的策略来治疗与ET-1相关的疼痛。

Background: Increasing evidence indicates that endothelin-1 (ET-1) has a role for peripheral nociceptive signaling in animals and humans. However, the mechanisms of the nociceptive effects of ET-1 have not been fully understood. The current study investigated the effects of ET-1 on the response of cultured adult mice dorsal root ganglion (DRG) neurons to capsaicin using intracellular calcium measurement and histochemical analyses.Methods: DRG were harvested from adult male C57B6N mice and were cultured. With a digital image analysis system, we detected the [Ca^<2+>]_i image of cultured DRG cells after loading with fura-2 AM. In addition, co-localization of protein kinase C (PKCL) with transient receptor potential Vi (TRPV1) and the translocation of PKCD were investigated using immunohistochemical methods.ResultsET-1 (10 nM) enhanced an increase in [Ca^<2+>]_i by capsaicin (10 nM) from 87.6 +- 11.6 nM to 414.8 +- 62.3 nM. The inhibition of endothelin A receptor significantly suppressed the enhancing effect of ET-1. In addition, a nonselective PKC inhibitor significantly suppressed the enhancing effect of ET-1. A myristoyl-tagged membrane-permeant-PKCD V1.2 inhibitory peptide also significantly suppressed the enhancing effect of ET-1. In the immunocytochemical study, PKCD immunoreactivity was found in most of TRPV1-positive neurons. After ET -1 application, PKCD immunoreactivity was observed to be translocated from the cytosol to the cell membrane in TRPV1-positive neurons,ConclusionOur results indicate that ET-1 enhances the response of DRG neurons to capsaicin in a PKC dependent manner. Our findings may lead to a new strategy to treat pain associated with ET-1.