Park Hee Chul, Seong Jinsil, An Jung Hee, Kim Jiyoung, Kim Un Jung, Lee Bae Whan
Department of Radiation Oncology, Hallym University, Chuncheon, South Korea.
Int J Radiat Oncol Biol Phys. 2005 Apr 1;61(5):1523-34. doi: 10.1016/j.ijrobp.2004.12.070.
Although radiotherapy is highly effective in relieving bone pain due to cancer invasion, its mechanism remains unclear. The aim of this study was to explore this mechanism in an animal model system.
A hind paw model of cancer pain was developed by transplanting a murine hepatocarcinoma, HCa-1, into the periosteal membrane of the foot dorsum of C3H/HeJ mice. Bone invasion from HCa-1 was histopathologically confirmed from sequential tumor sampling. For three experimental groups, a control (N), tumor without radiation (T), and tumor with radiation (TR), the development and level of pain were objectively examined in mice with a growing tumor by assessing pain-associated behavior. The differential expression of pain-related signals in the spinal cord was analyzed by proteomic analysis using high-resolution two-dimensional gel electrophoresis (2-DE) and mass spectrometry, and those of proteins by Western blotting. The pain-mediating neurotransmitters in the spinal cord were also examined by immunohistochemical staining for calcitonin gene-related peptide (CGRP) and substance P.
In the histopathologic examinations, bone invasion from HCa-1 was seen from Day 7 and was evident at Day 14 after transplantation, and measurable pain-associated behaviors were developed from Day 7. After 25 Gy of radiation to the tumors, the objective level of pain in the TR group decreased, with higher thresholds to mechanical and thermal stimulation than in the T group. From the 2-DE of spinal cord, 107 spots were identified; 12 proteins were changed more than fivefold because of tumor formation but then reversed after radiation in the tumor-bearing mice. The proteins involved included secretagogin, syntenin, P2X purinoreceptor 6 (P2X6), and Ca(2+)/Calmodulin-dependent protein kinase 1 (CaM kinase 1), the functions of which have been known to be involved in the Ca(2+)-signaling cascade, ATP-mediated fast synaptic transmission, or control of vesicular trafficking. Validations using Western blotting were successful for the CaM kinase and P2X6. In immunohistochemical staining of the spinal cord, a significant decrease after irradiation was shown in the expression of CGRP, but not in substance P.
We developed a novel model for bone pain due to cancer invasion, which was confirmed by histopathologic examination and measurable pain-associated behaviors. Radiotherapy decreased the objective level of pain. The underlying mechanism seems to be related to the Ca(2+)-signaling cascade or control of vesicular trafficking.
尽管放射疗法在缓解癌症侵袭所致骨痛方面非常有效，但其机制仍不清楚。本研究的目的是在动物模型系统中探索这一机制。
通过将小鼠肝癌HCa-1移植到C3H/HeJ小鼠足背的骨膜中，建立癌症疼痛的后爪模型。通过连续肿瘤取样，从组织病理学上证实了HCa-1的骨侵袭。对于三个实验组，即对照组（N）、未接受放疗的肿瘤组（T）和接受放疗的肿瘤组（TR），通过评估与疼痛相关的行为，客观地检测了肿瘤生长小鼠的疼痛发展和程度。使用高分辨率二维凝胶电泳（2-DE）和质谱分析，通过蛋白质组学分析脊髓中疼痛相关信号的差异表达，并通过蛋白质印迹法分析蛋白质的差异表达。还通过降钙素基因相关肽（CGRP）和P物质的免疫组织化学染色，检测了脊髓中疼痛介导的神经递质。
在组织病理学检查中，移植后第7天可见HCa-1的骨侵袭，第14天明显，且从第7天开始出现可测量的与疼痛相关的行为。对肿瘤进行25 Gy的放疗后，TR组的客观疼痛程度降低，与T组相比，对机械和热刺激的阈值更高。从脊髓的二维凝胶电泳中，鉴定出107个斑点；12种蛋白质因肿瘤形成而变化超过五倍，但在荷瘤小鼠放疗后恢复。所涉及的蛋白质包括分泌素、syntenin、P2X嘌呤受体6（P2X6）和钙/钙调蛋白依赖性蛋白激酶１（CaM激酶1），已知其功能参与钙信号级联、ATP介导的快速突触传递或囊泡运输的控制。使用蛋白质印迹法对CaM激酶和P2X6进行的验证是成功的。在脊髓的免疫组织化学染色中，照射后CGRP的表达显著降低，但P物质的表达未降低。
我们建立了一种因癌症侵袭所致骨痛的新模型，该模型通过组织病理学检查和可测量的与疼痛相关的行为得到证实。放射疗法降低了客观疼痛程度。其潜在机制似乎与钙信号级联或囊泡运输的控制有关。
