骨髓间充质干细胞移植治疗大鼠复杂性区域疼痛综合征Ⅰ型的疗效及机制研究
|
摘要
目的:研究骨髓间充质干细胞(BMSCs)移植治疗复杂性区域疼痛综合征Ⅰ(CRPS-I)型的疗效及机制。方法:骨髓贴壁法分离培养SD大鼠BMSCs。建立CRPS-I型模型,随机分为空白、模型、治疗和对照组(n=15)。机械性缩足反射阈值(thermal withdrawal latency, TWL)和热缩足反射潜伏期(mechanical withdrawal threshold, MWL)检测痛阈。PCR和Western-blot检测背根神经节脑源性神经营养因子(brain derived neurotrophic factor, BDNF)、降钙素基因相关肽(calcitonin gene related peptide,CGRP)、P物质(substance P,SP)等的表达。结果:CRPS-I型大鼠TWL值和MWL值均显著降低(P<0.05)。BMSCs移植后7、10及14 d TWL值和MWL值均有明显上升(P<0.05)。CRPS-I型大鼠背根神经节中BDNF、CGRP、SP、TNF-α和IL-6 mRNA和蛋白水平较假手术组均显著升高(P<0.05),而BMSCs移植后,上述因子的表达上调被逆转(P <0.05)。结论:BMSCs移植显著改善CRPS-I型模型大鼠的疼痛,其机制与减少受损神经组织中神经肽和炎症因子的释放有关。
Objective: The main aim of the study is to investigate the effects and mechanisms of bone marrow mesenchymal stem cell transplantation in treatment of complex regional pain syndrome type I. Methods: Bone marrow isolated from SD rats was purified and cultured to obtain the BMSCs using the whole bone marrow adherence method. CRPS-I rat model was established. The SD rats were randomly divided into 4 groups, namely blank group(rats without any intervention), experimental group(CRPS-I rat model without any intervention),treatment group(CRPS-I rat model with intrathecal BMSCs transplantation) and control group(rats with intrathecal cell culture fluid injection). Thermal withdrawal latency(TWL) and mechanical withdrawal threshold(MWL) were used to determine the pain threshold. The mRNA and protein expression of BDNF、CGRP、SP、TNF-α and IL-6 in rat dorsal root ganglia were measured by reverse transcription polymerase chain reaction(RTPCR) and Western blot analysis. Results: BMSCs were confirmed by microscopic observation, stem cell phenotype analysis and lipogenic/osteogenic induction and differentiation. The TWL and MWL of CRPS-I rats were significantly decreased(P < 0.05). After transplantation of BMSCs, the TWL and MWL of treatment group increased at 7,10 and 14 days after the operation(P < 0.05). The mRNA and protein expression of BDNF, CGRP, SP, TNF-α, IL-6 in dorsal root ganglion of CRPS-I model rats was significantly higher than that of normal rats(P < 0.05). After BMSCs transplantation, the mRNA and protein expression of these factors significantly decreased(P < 0.05). Conclusion:BMSCs transplantation can relieve pain and hyperalgesia of CRPS-I rats. The mechanism may be related with reducing the release of neuropeptides and inflammatory cytokines in damaged nerve tissue.
Objective: The main aim of the study is to investigate the effects and mechanisms of bone marrow mesenchymal stem cell transplantation in treatment of complex regional pain syndrome type I. Methods: Bone marrow isolated from SD rats was purified and cultured to obtain the BMSCs using the whole bone marrow adherence method. CRPS-I rat model was established. The SD rats were randomly divided into 4 groups, namely blank group(rats without any intervention), experimental group(CRPS-I rat model without any intervention),treatment group(CRPS-I rat model with intrathecal BMSCs transplantation) and control group(rats with intrathecal cell culture fluid injection). Thermal withdrawal latency(TWL) and mechanical withdrawal threshold(MWL) were used to determine the pain threshold. The mRNA and protein expression of BDNF、CGRP、SP、TNF-α and IL-6 in rat dorsal root ganglia were measured by reverse transcription polymerase chain reaction(RTPCR) and Western blot analysis. Results: BMSCs were confirmed by microscopic observation, stem cell phenotype analysis and lipogenic/osteogenic induction and differentiation. The TWL and MWL of CRPS-I rats were significantly decreased(P < 0.05). After transplantation of BMSCs, the TWL and MWL of treatment group increased at 7,10 and 14 days after the operation(P < 0.05). The mRNA and protein expression of BDNF, CGRP, SP, TNF-α, IL-6 in dorsal root ganglion of CRPS-I model rats was significantly higher than that of normal rats(P < 0.05). After BMSCs transplantation, the mRNA and protein expression of these factors significantly decreased(P < 0.05). Conclusion:BMSCs transplantation can relieve pain and hyperalgesia of CRPS-I rats. The mechanism may be related with reducing the release of neuropeptides and inflammatory cytokines in damaged nerve tissue.
引文
[1]Butler S.Qualitative research in complex regional pain syndrome(CRPS)[J].Scand J Pain,2015,9(1):62-63.
[2]Coderre TJ,Bennett GJ.A hypothesis for the cause of complex regional pain syndrome-type I(reflex sympathetic dystrophy):pain due to deep-tissue microvascular pathology[J].Pain Med,2010,11(8):1224-1238.
[3]Wang F,Zhang KH,Hu HM,et al.Alternatively activated microglia co-cultured with BMSCS offers a new strategy in the treatment of CNS-associated disease[J].Cell Biol Int,2015,39(3):341-349.
[4]Coderre TJ,Xanthos DN,Francis L,et al.Chronic post-ischemia pain(CPIP):a novel animal model of complex regional pain syndrome-type I(CRPS-I;reflex sympathetic dystrophy)produced by prolonged hindpaw ischemia and reperfusion in the rat[J].Pain,2004,112(1-2):94-105
[5]Bytner B,Huang YH,Yu LC,et al.Nociceptin/orphanin FQ into the rat periaqueductal gray decreases the withdrawal latency to heat and loading,an effect reversed by(Nphe(1)nociceptin(1-13)NH(2)[J].Brain Res,2001,922(1):118-124.
[6]Chen JY,Yeh GC,Tao PL,et al.Prenatal exposure to methamphetamine alters the mechanical withdrawal threshold and tonic hyperalgesia in the offspring[J]Neurotoxicology,2010,31(5):432-438.
[7]徐宵寒,许力,黄宇光.炎症反应和自身免疫在复杂性区域疼痛综合征中作用[J].中国疼痛医学杂志2016,22(11):847-850.
[8]Freund W,Wunderlich AP,Stuber G,et al.Different activation of opercular and posterior cingulate cortex(PCC)in patients with complex regional pain syndrome(CRPS I)compared with healthy controls during perception of electrically induced pain:a functional MRIstudy[J].Clin J Pain,2010,26(4):339-347.
[9]Zhang ZH,Pan YY,Jing RS,et al.Protective effects of BMSCs in combination with erythropoietin in bronchopulmonary dysplasia-induced lung injury[J].Mol Med Rep,2016,14(2):1302-1308.
[11]Wang Q,Sun G,Gao C,et al.Bone marrow mesenchymal stem cells attenuate 2,5-hexanedione-induced neuronal apoptosis through a NGF/AKT-dependent pathway[J].Sci Rep,2016,6:34715.
[12]Zong X,Wu S,Li F,et al.Transplantation of VEGF-mediated bone marrow mesenchymal stem cells promotes functional improvement in a rat acute cerebral infarction model[J].Brain Res,2017,1676:9-18.
[13]He X,Jiang L,Dan QQ,et al.Bone marrow stromal cells promote neuroplasticity of cerebral ischemic rats via a phosphorylated CRMP2-mediated mechanism[J].Behav Brain Res,2017,320:494-503.
[14]Yin F,Battiwalla M,Ito S,et al.Bone marrow mesenchymal stromal cells to treat tissue damage in allogeneic stem cell transplant recipients:correlation of biological markers with clinical responses[J].Stem Cells2014,32(5):1278-1288.
[15]Lopes C,Goncalves NP,Gomes CP,et al.BDNF gene delivery mediated by neuron-targeted nanoparticles is neuroprotective in peripheral nerve injury[J].Biomaterials,2017,121:83-96.
[16]Butler S.Qualitative research in complex regional pain syndrome(CRPS)[J].Scand J Pain,2015,9(1):62-63.
[17]Karsan N,Goadsby PJ.CGRP mechanism antagonists and migraine management[J].Curr Neurol Neurosci Rep,2015,15(5):25.
[18]Tanaka T,Narazaki M,Masuda K,et al.Regulation of IL-6 in Immunity and Diseases[J].Adv Exp Med Biol,2016,941:79-88.
[19]Shi X,Guo TZ,Wei T,et al.Facilitated spinal neuropeptide signaling and upregulated inflammatory mediator expression contribute to postfracture nociceptive sensitization[J].Pain,2015,156(10):1852-1863.
[2]Coderre TJ,Bennett GJ.A hypothesis for the cause of complex regional pain syndrome-type I(reflex sympathetic dystrophy):pain due to deep-tissue microvascular pathology[J].Pain Med,2010,11(8):1224-1238.
[3]Wang F,Zhang KH,Hu HM,et al.Alternatively activated microglia co-cultured with BMSCS offers a new strategy in the treatment of CNS-associated disease[J].Cell Biol Int,2015,39(3):341-349.
[4]Coderre TJ,Xanthos DN,Francis L,et al.Chronic post-ischemia pain(CPIP):a novel animal model of complex regional pain syndrome-type I(CRPS-I;reflex sympathetic dystrophy)produced by prolonged hindpaw ischemia and reperfusion in the rat[J].Pain,2004,112(1-2):94-105
[5]Bytner B,Huang YH,Yu LC,et al.Nociceptin/orphanin FQ into the rat periaqueductal gray decreases the withdrawal latency to heat and loading,an effect reversed by(Nphe(1)nociceptin(1-13)NH(2)[J].Brain Res,2001,922(1):118-124.
[6]Chen JY,Yeh GC,Tao PL,et al.Prenatal exposure to methamphetamine alters the mechanical withdrawal threshold and tonic hyperalgesia in the offspring[J]Neurotoxicology,2010,31(5):432-438.
[7]徐宵寒,许力,黄宇光.炎症反应和自身免疫在复杂性区域疼痛综合征中作用[J].中国疼痛医学杂志2016,22(11):847-850.
[8]Freund W,Wunderlich AP,Stuber G,et al.Different activation of opercular and posterior cingulate cortex(PCC)in patients with complex regional pain syndrome(CRPS I)compared with healthy controls during perception of electrically induced pain:a functional MRIstudy[J].Clin J Pain,2010,26(4):339-347.
[9]Zhang ZH,Pan YY,Jing RS,et al.Protective effects of BMSCs in combination with erythropoietin in bronchopulmonary dysplasia-induced lung injury[J].Mol Med Rep,2016,14(2):1302-1308.
[11]Wang Q,Sun G,Gao C,et al.Bone marrow mesenchymal stem cells attenuate 2,5-hexanedione-induced neuronal apoptosis through a NGF/AKT-dependent pathway[J].Sci Rep,2016,6:34715.
[12]Zong X,Wu S,Li F,et al.Transplantation of VEGF-mediated bone marrow mesenchymal stem cells promotes functional improvement in a rat acute cerebral infarction model[J].Brain Res,2017,1676:9-18.
[13]He X,Jiang L,Dan QQ,et al.Bone marrow stromal cells promote neuroplasticity of cerebral ischemic rats via a phosphorylated CRMP2-mediated mechanism[J].Behav Brain Res,2017,320:494-503.
[14]Yin F,Battiwalla M,Ito S,et al.Bone marrow mesenchymal stromal cells to treat tissue damage in allogeneic stem cell transplant recipients:correlation of biological markers with clinical responses[J].Stem Cells2014,32(5):1278-1288.
[15]Lopes C,Goncalves NP,Gomes CP,et al.BDNF gene delivery mediated by neuron-targeted nanoparticles is neuroprotective in peripheral nerve injury[J].Biomaterials,2017,121:83-96.
[16]Butler S.Qualitative research in complex regional pain syndrome(CRPS)[J].Scand J Pain,2015,9(1):62-63.
[17]Karsan N,Goadsby PJ.CGRP mechanism antagonists and migraine management[J].Curr Neurol Neurosci Rep,2015,15(5):25.
[18]Tanaka T,Narazaki M,Masuda K,et al.Regulation of IL-6 in Immunity and Diseases[J].Adv Exp Med Biol,2016,941:79-88.
[19]Shi X,Guo TZ,Wei T,et al.Facilitated spinal neuropeptide signaling and upregulated inflammatory mediator expression contribute to postfracture nociceptive sensitization[J].Pain,2015,156(10):1852-1863.