Background: The ways in which peripheral sensory nerves interact with osteosarcomas are important to understand because it could lead to development of new approaches to treat bone cancer pain. This study aimed to determine how cancer affects sensory nerve density and distribution in a murine model of osteosarcoma-induced bone pain. Methods: The femoral marrow cavities of male C3H/HeNHsd mice were injected with either NCTC 2472 primary osteosarcoma (cancer) cells or phosphate buffered saline (control). Pain behavior was assessed using limb use score and static weight bearing assays. At the experimental endpoint, femurs were collected, decalcified, immunolabeled, cleared and imaged using light sheet microscopy (Ultramicroscope Blaze, Miltenyi Biotec). The distribution of sensory nerves was traced through the marrow cavity of the proximal femur and the periosteum overlying the third trochanter (Imaris, Bitplane). Results: Weight bearing on the injected limb was decreased in osteosarcoma-injected but not saline-injected mice. Filament tracing revealed a reduced density of neurofilament 200 kDa-positive (NF200+; myelinated nerve marker) but not calcitonin gene-related peptide-positive (CGRP+; peptidergic nerve marker) sensory nerves in the marrow cavity of osteosarcoma-injected relative to saline-injected mice. There was increased density of CGRP+ but not NF200+ nerves in the periosteum of osteosarcoma-injected relative to saline-injected mice. Conclusions: Osteosarcoma differentially affects the density and distribution of different subtypes of peripheral sensory nerves in bone. Understanding how osteosarcomas affect different populations of sensory nerves could lead to more targeted mechanism-based treatments for bone cancer-induced pain.
背景：明确外周感觉神经与骨肉瘤的相互作用机制具有重要意义，这有助于开发治疗骨癌疼痛的新方法。本研究旨在通过小鼠骨肉瘤疼痛模型，探究肿瘤对感觉神经密度与分布的影响。方法：向雄性C3H/HeNHsd小鼠股骨骨髓腔内注射NCTC 2472原代骨肉瘤细胞（实验组）或磷酸盐缓冲液（对照组）。通过肢体使用评分和静态负重实验评估疼痛行为。实验终点采集股骨样本，经脱钙、免疫标记、透明化处理后，采用光片显微镜（Ultramicroscope Blaze，Miltenyi Biotec）成像。使用Imaris软件（Bitplane）对股骨近端骨髓腔及第三转子骨膜区域的感觉神经分布进行三维重建追踪。结果：骨肉瘤组小鼠注射侧肢体负重能力显著降低，而对照组未出现此现象。神经纤维示踪显示：与对照组相比，骨肉瘤组骨髓腔内神经丝蛋白200kDa阳性（NF200+；有髓神经标记）感觉神经密度降低，而降钙素基因相关肽阳性（CGRP+；肽能神经标记）感觉神经密度无显著变化；骨膜区域CGRP+神经密度增加，而NF200+神经密度无显著变化。结论：骨肉瘤对骨组织中不同亚型外周感觉神经的密度与分布存在差异化影响。深入解析骨肉瘤对感觉神经群体的特异性调控机制，可为开发基于精准靶向机制的骨癌疼痛治疗方案提供理论依据。
肿瘤(癌症)患者之家