SAHA对骨癌痛大鼠脊髓背角内谷氨酸转运体1表达的影响

目的:通过观察辛二酰苯胺异羟肟酸(Suberoylanilide Hydroxamic Acid,SAHA)对于骨癌痛(cancerinduced bone pain,CIBP)大鼠脊髓内谷氨酸转运体1(glutamate transporter 1,GLT-1)表达的影响,探讨GLT-1参与SAHA镇痛的相关机制。方法:将体重180~220 g雌性SD大鼠随机分为4组:Sham+vehicle组(A)、Sham+SAHA组(B)、CIBP+vehicle组(C)、CIBP+SAHA组(D)。经腹膜腔注射50 mg/kg SAHA,1/d。利用von Frey纤维测量大鼠手术侧机械缩足反射阈值(paw withdrawal threshod,PWT)。上述4组大鼠在不同时间点处死后取其腰膨大段脊髓,应用免疫组织化学染色和Western Blot方法观察大鼠腰膨大节段手术侧脊髓背角内GLT-1的表达情况。结果:与A组相比,C组大鼠PWT自术后第7 d开始显著下降,且一直持续至术后第14 d(P0.01),脊髓背角内GLT-1水平逐渐下调,呈时间依赖性(P0.05)。D组与C组相比,PWT升高(P0.05),术侧脊髓背角内GLT-1表达增加(P0.05)。结论:腹膜腔注射SAHA能够有效缓解骨癌痛,其部分机制可能与促进脊髓背角内GLT-1的表达有关。     

SAHA对糖尿病大鼠肾组织Twist和HDAC1表达的影响

目的:通过观察辛二酰苯胺异羟肟酸(SAHA)对糖尿病(DM)大鼠肾组织Twist、组蛋白去乙酰化酶1(HDAC1)及相关上皮-间充质转化和纤维化指标表达的影响,初步探讨SAHA对糖尿病肾病Twist表达变化的可能干预机制。方法:以链脲佐菌素复制DM大鼠模型,实验分为正常对照(NC)组、DM组和SAHA组,每组12只。造模成功8周后,SAHA组用SAHA(25 mg·kg~(-1)·d~(-1))进行灌胃。治疗8周后处死大鼠,HE和Masson染色观察肾组织形态变化;免疫组织化学染色观察肾组织中Twist的表达变化;Western blot法检测肾组织中Twist、HDAC1、上皮钙黏蛋白(E-cadherin)、α-平滑肌肌动蛋白(α-SMA)和IV型胶原(Col-Ⅳ)蛋白的表达;real-time PCR检测肾组织中Twist的mRNA表达。结果:DM组的血糖、24 h尿蛋白量和肾脏指数均显著高于NC组;而SAHA组肾脏指数与DM组相比有明显降低(P0.05),24 h尿蛋白量虽有降低,但差异无统计学显著性,而血糖无明显改变。与NC组对比,DM组大鼠肾组织中HDAC1、α-SMA和Col-Ⅳ蛋白表达上调,E-cadherin蛋白表达下调,Twist的mRNA和蛋白表达上调(P0.05);SAHA组大鼠肾组织中的Twist、HDAC1、α-SMA和Col-Ⅳ蛋白表达量明显低于DM组(P0.05),且Twist的mRNA的表达比DM组低,E-cadherin蛋白表达较DM组有所上升(P0.05)。相关性结果显示,在糖尿病大鼠肾组织中,Twist与HDAC1蛋白表达呈正相关(P0.05)。结论:SAHA可下调DM大鼠肾组织中Twist的表达,减轻糖尿病肾病大鼠肾纤维化病变,其机制可能与抑制HDAC1和Twist形成进而促进Ecadherin转录有关。     

大麻素CB_1受体活化对神经病理性疼痛大鼠脊髓背角嘌呤能P2X_2受体表达的抑制作用

目的:观察脊髓背角大麻素CB_1受体(CB_1R)在坐骨神经缩窄性损伤(CCI)所致的神经病理性疼痛中的作用及其对嘌呤能P2X_2受体表达的调节。方法:7~8周龄SD大鼠分为4组:(1)sham组;(2)CCI组;(3)CP55940+CCI组;(4)AM251+CP55940+CCI组。分别于CCI术前1 d,术后1、3、5、7、10、14 d测定热缩足反射潜伏期(TWL);免疫印迹技术检测各组大鼠损伤侧L_4~L_6段脊髓背角P2X_2受体表达。结果:CCI术后大鼠出现热痛敏,TWL明显缩短;鞘内给予非选择性大麻素受体激动剂CP55940可明显延长CCI大鼠TWL(P0.05);预先鞘内注射CB_1R拮抗剂AM251(0.05 mg/kg)可显著降低CP55940的镇痛效果(P0.05)。免疫印迹实验结果显示:CCI大鼠脊髓背角P2X_2受体在术后7、14 d表达明显增加(P0.05);鞘内给予CP55940可显著降低P2X_2受体表达(P0.05),而预先给予AM251可降低CP55940抑制P2X_2受体表达的效应(P0.05)。结论:脊髓背角CB_1受体激活对CCI所致的神经病理性疼痛具有良好的镇痛作用,其镇痛效应可能与抑制CCI大鼠嘌呤能P2X_2受体表达有关。     

老龄化对辣椒素受体1(TVRV1)在大鼠脊髓背角表达的影响(英文)

已有报道提出老龄化进程可能对伤害性行为表现有一定的影响 ,但是相关的机制仍不很明确。辣椒素受体 VR1(现命名为 TRPV1)已被证实为热伤害性感受器 ,老龄化进程中 TRPV1在痛传导路中的表达水平是否随年龄改变尚不清楚。本研究中应用免疫组织化学方法对三个年龄段 (青龄 :2～ 3月 ;中龄 :17～ 19月 ;老龄 2 4～ 2 6月 )大鼠脊髓背角的 TRPV1的表达进行了观察 ,结果显示 :(1) TRPV1在正常大鼠脊髓背角浅层的分布密度及其分布面积随年龄增长而逐渐减小 ;(2 )在外周致炎状态下 ,脊髓背角 TRPV1免疫反应产物密度在同年龄组内比较 ,青龄组减少 ,中龄和老龄组增加 ,而三年龄组内比较其分布面积时都明显增大 ,并且在非致炎组存在的组间差异消失。在青龄组大鼠 ,TRPV1免疫反应产物密度结果和分布面积结果并不一致 ,其原因未知。综上所述 ,本结果提示正常大鼠脊髓含 TRPV1的纤维终末随年龄增长而减少 ,但是在致炎情况下 ,TRPV1支配面积减少的现象消失。这可能是由于外周炎症刺激重新募集了背根节神经元 ,而老龄大鼠这种能力更强。本实验还提示 ,用免疫组织化学技术检测脊髓化学物质有无变化 ,测量光密度值和测量面积的方法都应该予以考虑     

小鼠中央杏仁核中GluR1的表达变化参与骨癌痛形成

目的:观察GluR1在骨癌痛(bone cancer pain,BCP)模型小鼠中央杏仁核中的表达变化。方法:健康C57小鼠分为假手术对照组(n=100)和骨癌痛组(n=100),每组均在术前和术后7、14、21、28 d先进行行为学检测,检测完毕后取材,进行免疫组化染色和Western Blot检测,观察GluR1在中央杏仁核中的表达变化。结果:从癌细胞接种后第7 d开始,BCP组出现自发缩足次数增多、PWT值降低,14 d后与Sham组比较开始有明显差异,表明模型建立成功;免疫组织化学染色显示GluR1在正常C57小鼠中央杏仁核中的表达水平较低,但在建模术后小鼠中央杏仁核中GluR1的表达开始逐渐升高,图像分析表明GluR1的光密度与对照组比较,第14 d时差异有统计学意义(P<0.05),BCP组21 d时GluR1的表达达高峰(P<0.01),Western Blot检测结果亦与之相符。结论:骨癌痛小鼠GluR1可能在杏仁核参与神经病理性痛的过程中具有重要作用。     

中枢神经大麻素受体分布的细胞类型研究

目的研究不同脑区大麻素CB1、CB2受体分布的细胞类型,探索大麻素受体在中枢神经系统中的可能作用。方法运用免疫荧光单标、双标的方法研究2种大麻素受体在成年大鼠不同脑区、不同类型细胞中的表达分布情况。结果成年大鼠不同脑区的神经元中有CB1、CB2受体的表达,海马、大脑皮层、脑干以及小脑的浦肯野细胞层的神经元有较高表达,且2种大麻素受体的表达差异较小,基底神经节区有中等表达,而胼胝体区未发现有神经元表达。少突胶质细胞及星型胶质细胞中发现CB1、CB2受体的表达。结论大麻素受体CB1、CB2在中枢神经系统多种类型的细胞中均有分布,可能通过多种途径参与神经系统功能调节。     

糖尿病机械性痛模型大鼠P2X3受体的时空表达

目的探讨糖尿病机械性痛(DMA)模型大鼠中P2X3受体(P2X3R)的时空表达变化。方法腹腔注射链脲菌素(STZ)建立大鼠DMA模型。采用von Frey细丝法测定DMA大鼠机械性痛阈,在不同时间点取腰髓4~5(L4~5)节段的脊髓背角(SDH)和背根神经节(DRG)以及足底皮肤,通过免疫荧光观察P2X3R阳性产物的表达变化。进一步Western blotting检测SDH和DRG中P2X3R蛋白的变化。结果与对照组(CON组)相比,注射STZ 7d后大鼠机械性痛阈显著降低,在14d时达到最低并持续至28d(P0.05)。免疫荧光和Western blotting结果均显示L4~5节段DRG中P2X3R表达在14d和21d时显著增多,与CON组相比差异有统计学意义(P0.05),28d时恢复至CON组水平;而在SDH及皮肤中,P2X3R表达上调出现在21d和28d(P0.05)。结论在STZ诱导的DMA大鼠机械性痛的不同时程中,DRG、SDH和皮肤中P2X3R的表达均呈现与痛敏变化几乎平行的变化趋势,但后两者比前者的变化在时间上略晚。这些结果说明P2X3R可能在DMA机械性痛敏的维持阶段起重要作用。     

脊髓辣椒素受体1(VR1)参与热敏感性的证据:老龄大鼠行为学和形态学相结合观察结果(英文)

以往的研究表明外周热伤害性感受器辣椒素受体 1(VR1,也称为 TRPV1)在热伤害性感觉中发挥着重要作用 ,但是脊髓 VR1在其中扮演的角色尚不清楚。因此本实验利用老龄大鼠蜜蜂毒 (BV)模型热痛敏 (仅持续 2 4h)和机械性痛敏 (持续 1月以上 )时程分离的行为学特点研究了在老龄大鼠中脊髓 VR1在外周组织损伤和炎性痛状态下的热敏感性中扮演的角色。在 BV注射后 4h(即热痛敏和机械性痛敏均存在时 )、2周 (即热痛敏消失而机械性痛敏存在时 )和 2月 (即两种痛敏均消失时 ) ,验证了热敏感性 (辐射热刺激 )和机械敏感性 (von-Frey纤维刺激 ) ,随后进行脊髓背角的 VR1免疫组织化学染色。结果如下 :(1)在未处理老龄大鼠组 ,VR1样免疫反应产物 (L I)在脊髓背角主要分布于 I、II层 ;(2 )在经 BV处理老龄大鼠组 ,外周损伤后 4h脊髓背角 VR1-L I有轻微增加 ,但在 2周后 VR1-LI表达水平明显低于对照水平 ,而且在 BV注射 2月后 ,即两种痛敏均消失时 ,VR1-L I表达水平下调仍非常明显。本结果表明脊髓背角的 VR1在空间分布上不随年龄改变而改变 ,但在周围化学组织损伤或炎症状态下 ,VR1-LI的表达水平可能与热敏感性的变化有动态相关性。由此我们提出除了外周位点 ,脊髓背角的 VR1也可能参与了热痛阈水平的维持和热痛敏的产生     

大鼠硬膜外双电极脊髓电刺激模型的建立

目的:为研究脊髓电刺激(spinal cord stimulation,SCS)镇痛作用的机制提供方便、实用和有效的SD大鼠脊髓硬膜外双电极刺激模型。方法:选取250~350 g雄性SD大鼠,结扎其左侧L5脊神经制作神经病理性痛模型。在此基础上,将制作的电极置入脊髓背侧硬膜外间隙(T11~T12),电极尾端经皮下隧道从颈后部引出、并固定于皮肤。术后恢复5 d,行脊髓电刺激测试。用电子Von Frey测试仪测量建模前后大鼠后肢的机械性缩足阈值,评估硬膜外双电极刺激对其术侧后肢机械性缩足阈值的影响。SCS测试后第2 d,在大鼠腹腔内注射大麻素1型受体(CB1)的拮抗剂AM251,然后观察AM251对大鼠SCS镇痛作用的影响。结果:大鼠左侧后肢机械性缩足的基础阈值为49.37±6.99 g,L5脊神经结扎及硬膜外电极置入术后机械性缩足的阈值为19.23±5.12 g,行SCS(20 Hz,150~200 mV)30 min后术侧机械性缩足阈值为35.62 g±7.27 g,与给予SCS刺激前比较具有显著性差异(P0.01);而与手术对侧(右侧)相比,机械性缩足阈值无明显变化(P0.05)。腹腔注射AM251可翻转SCS的镇痛作用(15.00±1.01 g,P0.01)。结论:硬膜外双电极植入方法取材容易,简单易行,与当前临床普遍应用的电刺激装置极为相似,为进一步研究脊髓电刺激的镇痛机理提供了可靠的模型,并为其他领域脊髓硬膜外电刺激实验动物模型的制作提供了参考。本文结果还提示内源性大麻素CB1受体可能参与SCS的镇痛机制。     

大麻素抑制坐骨神经损伤导致的神经痛及脊髓背角HCN4通道表达

目的:观察CB1受体在坐骨神经缩窄性损伤(CCI)所致的神经痛中的作用及对CCI大鼠脊髓背角HCN4通道表达的影响。方法:7~8周龄SD大鼠分为4组:(1)sham组(假手术组);(2)CCI组;(3)CP55940+CCI组;(4)AM251+CP55940+CCI组。采用von Frey电子测痛仪测定各组大鼠损伤侧机械缩足阈值(MWT);免疫印迹法检测损伤侧L_4~L_6脊髓背角HCN4的表达。结果:CCI术后1~14 d大鼠MWT明显降低,呈现稳定的机械痛敏;鞘内给予大麻素受体激动剂CP55940(0.05 mg/kg)可显著升高CCI大鼠MWT(P0.05);预先给予CB1受体拮抗剂AM251(0.05 mg/kg)可明显阻断CP55940的镇痛效果(P0.05)。免疫印迹检测结果显示,CCI大鼠损伤侧L_4~L_6脊髓背角HCN4通道表达明显增加(P0.05);鞘内给予CP55940可显著降低CCI大鼠的HCN4表达(P0.05),CP55940抑制HCN4表达的效应可被AM251阻断(P0.05)。结论:脊髓CB1受体激活对外周神经损伤导致的神经痛具有良好的镇痛作用,其镇痛效应可能与抑制神经痛大鼠脊髓背角HCN4通道表达有关。     

Involvement of spinal monocyte chemoattractant protein-1 (MCP-1) in cancer-induced bone pain in rats

In this study, we examined the involvement of chemokine monocyte chemoattractant protein-1 (MCP-1) in the spinal cord of a rat model of cancer-induced bone pain (CIBP). In this model, CIBP was established by an intramedullary injection of Walker 256 cells into the tibia of rats. We observed a significant increase in expression levels of MCP-1 and its receptor CCR2 in the spinal cord of CIBP rats. Furthermore, the intrathecal administration of an anti-MCP-1 neutralizing antibody attenuated the mechanical allodynia established in CIBP rats. Likewise, an intrathecal injection of exogenous recombinant MCP-1 induced a striking mechanical allodynia in naïve rats. These results suggest that increases in spinal MCP-1 and CCR2 expression are involved in the development of mechanical allodynia associated with bone cancer rats.     

Immunohistochemical distribution of the cannabinoid receptor 1 and fatty acid amide hydrolase in the dog claustrum

Cannabinoid receptor 1 (CB1R) and fatty acid amide hydrolase (FAAH) are part of the endocannabinoid system (ECB) which exerts a neuromodulatory activity on different brain functions and plays a key role in neurogenesis. Although many studies have reported FAAH and CB1R expression in the brain of different animal species, to the best of our knowledge they have never been described in the canine claustrum. Claustrum samples, obtained from necropsy of four neurologically normal dogs, were formalin fixed for paraffin embedding. Sections were either stained for morpho-histological analysis or immunostained for CB1R and FAAH. Analysis of adjacent sections incubated with the two antisera showed a complementary labeling pattern in the claustrum, with CB1R antibody staining fibers while anti-FAAH antibody stained cell bodies and the proximal portion of dendrites; this particular anatomical relationship suggests a retrograde endocannabinoid action via CB1R. CB1R and FAAH complementary immunostaining and their cellular localization reported here provide the first anatomical evidence for existence of the ECB in the dog claustrum.     

Involvement of EphB1 receptor/ephrinB1 ligand in bone cancer pain

In prior studies, Eph/ephrin system was demonstrated to be involved in inflammatory and neuropathic pain modulation. The present study was to investigate whether the spinal Eph/ephrin signaling was involved in modulation of spinal inflammatory cytokines in bone cancer pain (BCP) of rats. BCP was induced by intra-tibial inoculation of Walker 256 mammary gland carcinoma cells. The expressions of EphB1/ephrinB1 in spinal cord (SC) and dorsal root ganglia (DRG) were determined. At 16 days post inoculation, the pain relieving effect and the mRNA levels of inflammatory cytokines were detected after intrathecal administration of EphB1-Fc (blocker of EphB1 receptor, 10μg). The results showed that the EphB1/ephrinB1 expression was significantly increased in SC, but ephrinB1 was decreased in DRG after Walker 256 inoculation. The mechanical allodynia induced by bone cancer was significantly alleviated by intrathecal administration of EphB1-Fc. Furthermore, the RT-PCR analysis showed that the mRNA levels of IL-1β, IL-6 and TNF-α were significantly increased at 16 days post Walker 256 inoculation and were significantly suppressed by intrathecal administration of EphB1-Fc in SC. We concluded that Eph/ephrin might be involved in the maintenance of mechanical allodynia, via modulating the expression of spinal inflammatory cytokines, in the present rat model of BCP. This study suggested that Eph/ephrin signaling would be a potential target for the treatment of BCP.     

Regional changes in type 1 cannabinoid receptor availability in Parkinson's disease in vivo

The type 1 cannabinoid receptor (CB1) is a crucial modulator of synaptic transmission in brain and has been proposed as a potential therapeutic target in Parkinson's disease (PD), especially for treatment of levodopa-induced dyskinesias (LID). Our aim was to measure CB1 levels in brains of PD patients in vivo and to investigate the relation between CB1 availability and LID. We studied 12 healthy controls and 29 PD patients (9 drug-naïve patients with early PD, 10 patients with advanced PD and LID, and 10 patients with advanced PD without LID). PD patients were examined using the Unified Parkinson's Disease Rating Scale (UPDRS) and the modified Abnormal Involuntary Movement Scale (mAIMS). All subjects underwent positron emission tomography (PET) with the CB1-selective radioligand [¹⁸F] MK-9470 and magnetic resonance imaging (MRI). PD patients showed an absolute decrease in CB1 availability in the substantia nigra. By contrast, CB1 availability was relatively increased in nigrostriatal, mesolimbic, and mesocortical dopaminergic projection areas. CB1 availability did not differ significantly between advanced PD patients with and without LID. Within the group of PD patients with LID, there was no significant correlation between CB1 availability and LID severity. These data demonstrate regional changes in CB1 availability in PD in vivo, but do not support a role for dysregulation of CB1 levels in the pathogenesis of LID.     

Interleukin 1beta facilitates bone cancer pain in rats by enhancing NMDA receptor NR-1 subunit phosphorylation

It has been shown that interleukin-1beta (IL-1beta) facilitates nociception during neuropathic and inflammatory pain, but its involvement in bone cancer pain and its mechanisms have not previously been established. This study is an investigation of IL-1beta spinal expression and the N-methyl-D-aspartate (NMDA) receptor (NMDAR) NR1 subunit phosphorylation during cancer pain, co-localization of IL-1 receptor type I (IL-1RI) and NMDAR in the spinal cord, and the effects of IL-1 receptor antagonist (IL-1ra) on NMDAR1 (NR1) phosphorylation and hyperalgesia in a rat model of bone cancer pain. Cancer was induced by injecting AT-3.1 prostate cancer cells into the tibia of the male Copenhagen rat. Phosphorylation of NR1, an essential subunit of the NMDAR, is known to modulate NMDAR activity and facilitate pain. Mechanical hyperalgesia, established by a decrease in paw withdrawal pressure threshold (PWPT), was measured at baseline and 2 h after IL-1ra treatment. IL-1ra was given (i.t.) daily for 7 days between days 13 and 19 after the cancer cell inoculation. Spinal cords were removed for Western blot to measure IL-1beta and NR1 phosphorylation and for double immunostaining of IL-1RI and NR1. The data showed that 1) spinal IL-1beta was up-regulated and NR1 phosphorylation was increased, 2) IL-1ra at 0.1 mg/rat significantly (P<0.05) inhibited mechanical hyperalgesia, increasing PWPT on day 14 from 71.1+/-3.1-85.3+/-4.6 g and on day 19 from 73.5.0+/-3.5-87.1+/-3.7 g, and inhibited NR1 phosphorylation compared with saline control, and 3) IL-1RI is localized in NR1-immunoreactive neurons within the spinal cord. The results suggest that spinal IL-1beta enhances NR1 phosphorylation to facilitate bone cancer pain.     

Evoked pain behavior and spinal glia activation is dependent on tumor necrosis factor receptor 1 and 2 in a mouse model of bone cancer pain

Bone–cancer-related pain is one of the most disabling factors in patients suffering from primary bone cancer or bone metastases. Recent studies point toward an important role of proinflammatory cytokines, example tumor necrosis factor-α (TNF), for tumor growth and bone–cancer-associated pain. Mechanisms by which TNF, through its receptor subtypes, TNF receptor 1 (TNFR1) and −2 (TNFR2), elicits altered sensation and pain behavior, are still incompletely understood. To look for a potential role of TNF in bone cancer pain, cancer-related pain was analyzed in fibrosarcoma-bearing C57Bl/6J wild type mice after systemic antagonism of TNF. To further clarify the role of TNF receptor (TNFR) in bone-cancer pain, naive and fibrosarcoma-bearing C57Bl/ 6J wild type and transgenic mice with a deficiency of TNFR1 (TNFR1ko), TNFR2 (TNFR2ko), and TNFR1+2 (TNFR1+2ko) were compared regarding cancer-related pain and hyperalgesia, tumor growth, osteoclast activation, and spinal astrogliosis. Systemic antagonism of TNF significantly alleviated tactile hypersensitivity and spontaneous bone–cancer-related pain behavior. Most interestingly, combined deletion of the TNFR1 and TNFR2, but not of either gene alone, almost completely inhibited the development of tactile hypersensitivity, whereas spontaneous pain behavior was transiently increased. Accordingly, spinal astrogliosis was markedly reduced, whereas tumor growth was significantly increased in TNFR1+2ko mice. In contrast, deletion of the TNFR1 or TNFR2 gene alone did not change tumor growth or spinal astrogliosis. Our findings suggest that the combined absence of TNFR1 and TNFR2 is necessary for the attenuation of cancer-related tactile hypersensitivity and concomitant spinal astrogliosis, whereas tumor growth seems to be inhibited by combined TNFR activation. These findings support the hypothesis of cytokine-dependent pain development in cancer pain. Differential targeting of TNFR activation could be an interesting strategy in bone–cancer-related pain conditions.     

Oxytocin ameliorates bone cancer pain by suppressing toll-like receptor 4 and proinflammatory cytokines in rat spinal cord

Abstract

Bone cancer pain is considered to be mechanistically unique compared with inflammatory or neuropathic pain states. Toll-like receptor 4 (TLR4) is a transmembrane receptor protein which has been reported to be involved in neuropathic pain. However, the role of TLR4 in bone cancer pain is still unclear. Therefore, the aim of this study is to investigate the hypothesis that oxytocin may ameliorate bone cancer pain by suppressing TLR4 in spinal cord. Behavioral analysis and molecular biological experiments were carried out. Our data demonstrated that intrathecally delivery of oxytocin significantly ameliorated the mechanical allodynia and thermal hyperalgesia in bone cancer pain rats. Moreover, oxytocin suppressed the up-regulation of TLR4 and proinflammatory cytokines TNFα and IL-1β in spinal cord of bone cancer pain rats. Therefore, we concluded that intrathecal administration of oxytocin relieves bone cancer pain by suppressing the up-regulation of TLR4, TNFα and IL-1β in spinal cord. Oxytocin possesses analgesic efficacy against bone cancer pain and deserves further to confirm its effectiveness in clinically relevant of cancer pain.