7,8-二羟基黄酮联合施万细胞促进大鼠坐骨神经损伤轴突再生的作用
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摘要
目的探讨7,8-二羟基黄酮(7,8-DHF)与施万细胞(SCs)对大鼠脱细胞同种异体神经支架(ANA)移植坐骨神经缺损后轴突再生和功能恢复的影响。方法 45只成年SD大鼠随机分为脱细胞神经支架(ANA)组、SCs组和7,8-DHF+SCs组。采用坐骨神经功能指数(SFI)和电生理方法检测运动功能的恢复,Western bolt检测再生神经内BDNF及其受体TrkB的蛋白表达,免疫荧光示踪PKH26标记的移植SCs,NF和S100免疫荧光法检测再生神经内轴突再生和髓鞘形成情况。结果与SCs组比较,7,8-DHF+SCs组再生神经内BDNF和TrkB蛋白相对表达明显增高,PKH-26标记的SCs数量显著增高,NF和S100表达增强(P<0. 05)。与对照组比较,SCs组和7,8-DHF+SCs组神经传导速度增快、波幅增高、延迟期缩短、SFI指数增高,其中7,8-DHF+SCs组电生理和SFI指标优于SCs组(P<0. 05)。结论 7,8-DHF联合SCs对坐骨神经损伤后轴突再生、髓鞘形成和功能恢复的作用优于SCs组,其机制可能与7,8-DHF促进再生神经BDNF及其受体TrkB表达,进而促进移植的SCs存活有关。
Objective To study the effect of 7,8-dihydroxyflavone (7,8-DHF) and Schwann cells (SCs) transplantation on axon regeneration and function recovery after acellular nerve allograft (ANA) repair of the sciatic nerve gap in mice.Methods A total of 45 healthy SD rat were randomly divided into control group,SCs group and 7,8-DHF + SCs group. SFI and electrophysiology were used to evaluate functional recovery. BDNF and TrkB protein were evaluated by western bolt,observe the fluorescence signal of SCs labeled with PKH-26 by using immunofluorescence,observe the axonal regeneration and myelination by using NF,S100 immunofluorescence. Results Compared with SCs group,the expression of BDNF,TrkB,NF,S100 and PKH-26 labeled SCs in 7,8-DHF + SCs group were increased (P< 0. 05). Compared with control group,nerve conduction velocity and wave amplitude were significantly increased in SCs group and 7,8-DHF + SCs group (P < 0. 05),however,incubation period were significantly decreased in two treatment group (P < 0. 05),the effect of 7,8-DHF + SCs group was more powerful than SCs group (P< 0. 05),moreover,the SFI score of 7,8-DHF + SCs group was higher than control group and SCs group. Conclusion The combined 7,8-DHF and SCs transplantation treatment promoted axonal regeneration,myelination and function recovery significantly more than SCs treatments,which may be related to 7,8-DHF upregulating BDNF and TrkB expression in the ANA,and promoting SCs survival.
Objective To study the effect of 7,8-dihydroxyflavone (7,8-DHF) and Schwann cells (SCs) transplantation on axon regeneration and function recovery after acellular nerve allograft (ANA) repair of the sciatic nerve gap in mice.Methods A total of 45 healthy SD rat were randomly divided into control group,SCs group and 7,8-DHF + SCs group. SFI and electrophysiology were used to evaluate functional recovery. BDNF and TrkB protein were evaluated by western bolt,observe the fluorescence signal of SCs labeled with PKH-26 by using immunofluorescence,observe the axonal regeneration and myelination by using NF,S100 immunofluorescence. Results Compared with SCs group,the expression of BDNF,TrkB,NF,S100 and PKH-26 labeled SCs in 7,8-DHF + SCs group were increased (P< 0. 05). Compared with control group,nerve conduction velocity and wave amplitude were significantly increased in SCs group and 7,8-DHF + SCs group (P < 0. 05),however,incubation period were significantly decreased in two treatment group (P < 0. 05),the effect of 7,8-DHF + SCs group was more powerful than SCs group (P< 0. 05),moreover,the SFI score of 7,8-DHF + SCs group was higher than control group and SCs group. Conclusion The combined 7,8-DHF and SCs transplantation treatment promoted axonal regeneration,myelination and function recovery significantly more than SCs treatments,which may be related to 7,8-DHF upregulating BDNF and TrkB expression in the ANA,and promoting SCs survival.
引文
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[13]ZHAO Y,WANG Y,GONG J,et al.Chitosan degradation products facilitate peripheral nerve regeneration by improving macrophage-constructed microenvironments[J].Biomaterials,2017,134(4):64-77.
[14]XIANG F,WEI D,YANG Y,et al.Tissue-engineered nerve graft with tetramethylpyrazine for repair of sciatic nerve defects in rats[J].Neurosci Lett,2017,638:114-120.
[15]PLANT C D,PLANT G W.Viral Transduction of Schwann Cells for Peripheral Nerve Repair[J].Methods Mol Biol,2018,1 739:455-466.
[16]PAINTER M W.Aging Schwann cells:mechanisms,implications,future directions[J].Curr Opin Neurobiol,2017,47:203-208.
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[20]LIU S,SANDNER B,SCHACKEL T,et al.Regulated viral BDNF delivery in combination with Schwann cells promotes axonal regeneration through capillary alginate hydrogels after spinal cord injury[J].Acta Biomater,2017,60:167-180.
[21]SCHAAKXS D,KALBERMATTEN D F,PRALONG E,et al.Poly-3-hydroxybutyrate strips seeded with regenerative cells are effective promoters of peripheral nerve repair[J].J Tissue Eng Regen Med,2017,11(3):812-821.
[22]MARUYAMA M,WAKABAYASHI S,OTA H,et al.Reconstruction of the Shallow Acetabulum With a Combination of Autologous Bulk and Impaction Bone Grafting Fixed by Cement[J].Clin Orthop Relat Res,2017,475(2):387-395.
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[24]GERSEY Z C,BURKS S S,ANDERSON K D,et al.First human experience with autologous Schwann cells to supplement sciatic nerve repair:report of 2 cases with longterm follow-up[J].Neurosurg Focus,2017,42(3):E2.
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[26]JIANG M,PENG Q,LIU X,et al.Small-molecule Trk B receptor agonists improve motor function and extend survival in a mouse model of Huntington's disease[J].Hum Mol Genet,2013,22(12):2 462-2 470.
[27]HELAN M,ARAVAMUDAN B,HARTMAN W R,et al.BDNF secretion by human pulmonary artery endothelial cells in response to hypoxia[J].J Mol Cell Cardiol,2014,68:89-97.
[28]BOLLEN E,VANMIERLO T,AKKERMAN S,et al.7,8-Dihydroxyflavone improves memory consolidation processes in rats and mice[J].Behav Brain Res,2013,257:8-12.
[29]CHITRANSHI N,GUPTA V,KUMAR S,et al.Exploring the Molecular Interactions of 7,8-Dihydroxyflavone and Its Derivatives with Trk B and VEGFR2 Proteins[J].Int J Mol Sci,2015,16(9):21 087-21 108.
[30]ENGLISH A W,LIU K,NICOLINI J M,et al.Small-molecule trk B agonists promote axon regeneration in cut peripheral nerves[J].Proc Natl Acad Sci U S A,2013,110(40):16 217-16 222.
[2]WANG Y,JIA H,LI W Y,et al.Molecular examination of bone marrow stromal cells and chondroitinase ABC-assisted acellular nerve allograft for peripheral nerve regeneration[J].Exp Ther Med,2016,12(4):1 980-1 992.
[3]王莹,李文媛,尹国华,等.二氢睾酮联合ADSC对大鼠脊髓损伤BDNF及其受体Trk B的影响[J].中国实用神经疾病杂志,2017,20(5):17-20.
[4]ZHANG X L,MCGLOTHAN J L,MIRY O,et al.From the Cover:7,8-Dihydroxyflavone Rescues Lead-Induced Impairment of Vesicular Release:A Novel Therapeutic Approach for Lead Intoxicated Children[J].Toxicol Sci,2018,161(1):186-195.
[5]PRINCE C S,MALOYAN A,MYATT L.Tropomyosin Receptor Kinase B Agonist,7,8-Dihydroxyflavone,Improves Mitochondrial Respiration in Placentas From Obese Women[J].Reprod Sci,2018,25(3):452-462.
[6]KRISHNA G,AGRAWAL R,ZHUANG Y,et al.7,8-Dihydroxyflavone facilitates the action exercise to restore plasticity and functionality:Implications for early brain trauma recovery[J].Biochim Biophys Acta,2017,1863(6):1 204-1 213.
[7]CHEN C,WANG Z,ZHANG Z,et al.The prodrug of 7,8-dihydroxyflavone development and therapeutic efficacy for treating Alzheimer's disease[J].Proc Natl Acad Sci U SA,2018,115(3):578-583.
[8]AYTAN N,CHOI J K,CARRERAS I,et al.Protective effects of 7,8-dihydroxyflavone on neuropathological and neurochemical changes in a mouse model of Alzheimer's disease[J].Eur J Pharmacol,2018,828(1):9-17.
[9]SUN T,CHEN S,HUANG H,et al.Metabolic profile study of 7,8-dihydroxyflavone in monkey plasma using high performance liquid chromatography-tandem mass spectrometry[J].J Chromatogr B Analyt Technol Biomed Life Sci,2017,1 061-1 062:97-102.
[10]ZHENG J,SUN J,LU X,et al.BDNF promotes the axonal regrowth after sciatic nerve crush through intrinsic neuronal capabi lity upregulation and distal portion protection[J].Neurosci Lett,2016,621(2):1-8.
[11]JIA H,WANG Y,WANG T,et al.Synergistic Effects of G-CSF and Bone Marrow Stromal Cells on Nerve Regeneration with Acellular Nerve Xenografts[J].Synapse,2017,71(7):1-11.
[12]LI W Y,ZHANG W T,CHENG Y X,et al.Inhibition of KLF7-Targeting MicroRNA 146b Promotes Sciatic Nerve Regeneration[J].Neuroscience Bulletin,2018,34(3):419-437.
[13]ZHAO Y,WANG Y,GONG J,et al.Chitosan degradation products facilitate peripheral nerve regeneration by improving macrophage-constructed microenvironments[J].Biomaterials,2017,134(4):64-77.
[14]XIANG F,WEI D,YANG Y,et al.Tissue-engineered nerve graft with tetramethylpyrazine for repair of sciatic nerve defects in rats[J].Neurosci Lett,2017,638:114-120.
[15]PLANT C D,PLANT G W.Viral Transduction of Schwann Cells for Peripheral Nerve Repair[J].Methods Mol Biol,2018,1 739:455-466.
[16]PAINTER M W.Aging Schwann cells:mechanisms,implications,future directions[J].Curr Opin Neurobiol,2017,47:203-208.
[17]LIU H,LV P,WU H,et al.The Proliferation Enhancing Effects of Salidroside on Schwann Cells In Vitro[J].Evid Based Complement Alternat Med,2017,2017:4 673 289.
[18]ZHENG Y,HUANG C,LIU F,et al.Reactivation of denervated Schwann cells by neurons induced from bone marrow-derived mesenchymal stem cells[J].Brain Res Bull,2018,139:211-223.
[19]WU L M N,DENG Y,WANG J,et al.Programming of Schwann Cells by Lats1/2-TAZ/YAP Signaling Drives Malignant Peripheral Nerve Sheath Tumorigenesis[J].Cancer Cell,2018,33(2):292 e7-308 e7.
[20]LIU S,SANDNER B,SCHACKEL T,et al.Regulated viral BDNF delivery in combination with Schwann cells promotes axonal regeneration through capillary alginate hydrogels after spinal cord injury[J].Acta Biomater,2017,60:167-180.
[21]SCHAAKXS D,KALBERMATTEN D F,PRALONG E,et al.Poly-3-hydroxybutyrate strips seeded with regenerative cells are effective promoters of peripheral nerve repair[J].J Tissue Eng Regen Med,2017,11(3):812-821.
[22]MARUYAMA M,WAKABAYASHI S,OTA H,et al.Reconstruction of the Shallow Acetabulum With a Combination of Autologous Bulk and Impaction Bone Grafting Fixed by Cement[J].Clin Orthop Relat Res,2017,475(2):387-395.
[23]YAN Y,HUNTER D A,SCHELLHARDT L,et al.Nerve stepping stone has minimal impact in aiding regeneration across long acellular nerve allografts[J].Muscle Nerve,2018,57(2):260-267.doi:10.1002/mus.25659.
[24]GERSEY Z C,BURKS S S,ANDERSON K D,et al.First human experience with autologous Schwann cells to supplement sciatic nerve repair:report of 2 cases with longterm follow-up[J].Neurosurg Focus,2017,42(3):E2.
[25]TSAI T,KLAUSMEYER A,CONRAD R,et al.7,8-Dihydroxyflavone leads to survival of cultured embryonic motoneurons by activating intracellular signaling pathways[J].Mol Cell Neurosci,2013,5(10):18-28.
[26]JIANG M,PENG Q,LIU X,et al.Small-molecule Trk B receptor agonists improve motor function and extend survival in a mouse model of Huntington's disease[J].Hum Mol Genet,2013,22(12):2 462-2 470.
[27]HELAN M,ARAVAMUDAN B,HARTMAN W R,et al.BDNF secretion by human pulmonary artery endothelial cells in response to hypoxia[J].J Mol Cell Cardiol,2014,68:89-97.
[28]BOLLEN E,VANMIERLO T,AKKERMAN S,et al.7,8-Dihydroxyflavone improves memory consolidation processes in rats and mice[J].Behav Brain Res,2013,257:8-12.
[29]CHITRANSHI N,GUPTA V,KUMAR S,et al.Exploring the Molecular Interactions of 7,8-Dihydroxyflavone and Its Derivatives with Trk B and VEGFR2 Proteins[J].Int J Mol Sci,2015,16(9):21 087-21 108.
[30]ENGLISH A W,LIU K,NICOLINI J M,et al.Small-molecule trk B agonists promote axon regeneration in cut peripheral nerves[J].Proc Natl Acad Sci U S A,2013,110(40):16 217-16 222.