药学学报, 2016, 51(3): 338-346
杨盛, 何然, 张飞燕, 薛强, 徐晓玉. 细胞共培养模型及其在中枢神经系统疾病研究中的应用[J]. 药学学报, 2016, 51(3): 338-346.
YANG Sheng, HE Ran, ZHANG Fei-yan, XUE Qiang, XU Xiao-yu. Application of cell co-culture techniques in central nervous system diseases[J]. Acta Pharmaceutica Sinica, 2016, 51(3): 338-346.

杨盛, 何然, 张飞燕, 薛强, 徐晓玉
西南大学药学院 & 中医药学院, 重庆 400715
关键词:    脑卒中      帕金森病      阿尔茨海默病      肌萎缩侧索硬化症      细胞共培养      神经血管单元     
Application of cell co-culture techniques in central nervous system diseases
YANG Sheng, HE Ran, ZHANG Fei-yan, XUE Qiang, XU Xiao-yu
College of Pharmaceutical Sciences and College of Chinese Medicine, Southwest University, Chongqing 400715, China
The study of central nervous system disease is dependent on in vitro culture of neuronal cells. However, it is hard to determine the interaction between cells in culture of single type of neuronal cells. The co-culture system is able to mimic the cell-cell interaction in the brain and to facilitate investigation into the interaction between different types of cells, as well as cell-environment interaction. The co-culture of neurocytes is more and more popular in the disease study of central nervous system in vitro, such as stroke, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, etc. Neurovascular unit (NVU), which is composed of neurons, brain microvascular endothelial cells and astrocytes, can reflect the structure and function of brain in state of the art. Establishment of NVU in vitro is important in the study of the brain diseases. In this paper, several co-culture models of the central nervous system are reviewed in techniques for two-dimensional and three-dimensional culturing. Cell contact and non-contact methods are compared. Moreover, their application in the relevant research and the future direction are explored.
Key words:    stroke    Parkinson's disease    Alzheimer's disease    amyotrophic lateral sclerosis    co-culture of cells    neurovascular unit   
收稿日期: 2015-07-01
DOI: 10.16438/j.0513-4870.2015-0583
基金项目: 国家自然科学基金面上项目(81473549);国家科技重大新药创制专项(2014ZX09304-306-04);教育部中央高校基本科研业务费(XDJK2014D023).
通讯作者: 徐晓玉
Email: xuxiaoyu@swu.edu.cn
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薛强  在本刊中的所有文章
徐晓玉  在本刊中的所有文章

[1] Miron V, Antel JP. Isolation and culture of primary human CNS neural cells[M]//Frederoff S. Protocols for Neural Cell Culture, 4th ed. New Jersey:Humana Press, 2010:87-104.
[2] Patabendige A, Skinner RA, Abbott NJ. Establishment of a simplified in vitro porcine blood-brain barrier model with high transendothelial electrical resistance[J]. Brain Res, 2013, 1521:1-15.
[3] Sun DA, Sombati S, DeLorenzo RJ. Glutamate injuryinduced epileptogenesis in hippocampal neurons:an in vitro model of stroke-induced "epilepsy"[J]. Stroke, 2001, 32:2344-2350.
[4] Guo S, Lo EH. Dysfunctional cell-cell signaling in the neurovascular unit as a paradigm for central nervous system disease[J]. Stroke, 2009, 40:S4-S7.
[5] Bath PM. Albumin for hyperacute stroke:another failed neuroprotectant[J]. Lancet Neurol, 2013, 12:1036-1037.
[6] Liu Y, Wang L, Long Z, et al. Protoplasmic astrocytes enhance the ability of neural stem cells to differentiate into neurons in vitro[J]. PLoS One, 2012, 7:e38243.
[7] Romão LF, Mendes FA, Feitosa NM, et al. Connective tissue growth factor (CTGF/CCN2) is negatively regulated during neuron-glioblastoma interaction[J]. PLoS One, 2013, 8:e55605.
[8] Xue Q, Liu Y, Qi H, et al. A novel brain neurovascular unit model with neurons, astrocytes and microvascular endothelial cells of rat[J]. Int J Biol Sci, 2013, 9:174-189.
[9] Wang CN, Lin L, Duan ZF, et al. Primary co-culture of cortical neurons and astrocytes of new-born SD rats[J]. Acta Pharm Sin (药学学报), 2013, 48:1729-1732.
[10] Kempuraj D, Khan MM, Thangavel R, et al. Glia maturation factor induces interleukin-33 release from astrocytes:implications for neurodegenerative diseases[J]. J Neuroimmune Pharmacol, 2013, 8:643-650.
[11] Shi Y, Zhou M, Jiang M. A protocol for primary dissociated astrocyte and neuron co-culture[J]. Acta Physiol Sin (生理学报), 2013, 65:72-76.
[12] Silva SL, Osório C, Vaz AR, et al. Dynamics of neuron-glia interplay upon exposure to unconjugated bilirubin[J]. J Neurochem, 2011, 117:412-424.
[13] Luo ZZ, Wang J, Kong M, et al. Effect of neuronal injury severity on microglial phenotype[J]. J Huazhong Univ Sci Technol (Health Sci) (华中科技大学学报医学版), 2015, 44:47-51.
[14] Yan R, Luo XG, Zhang Y, et al. Mechanism of neuronal differentiaton of bone marrow stromal cells induced by astrocyte conditional medium[J]. J China Med Univ (中国医科大学学报), 2013, 42:714-721.
[15] Ye JM, Liu ZH, Xie HF, et al. Protection role of silent information regultaor 1 in toxicity of activated BV-2 cells to PC12 cells[J]. Chin J Neuromed (中华神经医学杂志), 2013, 12:1112-1117.
[16] Park LC, Zhang H, Gibson GE. Co-culture with astrocytes or microglia protects metabolically impaired neurons[J]. Mech Ageing Dev, 2001, 123:21-27.
[17] Rathinam ML, Watts LT, Narasimhan M, et al. Astrocyte mediated protection of fetal cerebral cortical neurons from rotenone and paraquat[J]. Environ Toxicol Pharmacol, 2012, 33:353-360.
[18] Guo H, Ma J, Tong Y, et al. A comparative study on three models of co-culture of neurons and astrocytes[J]. Chin J Contemp Pediatr (中国当代儿科杂志), 2010, 12:984-987.
[19] Zhang ZH, Li D, Sun K, et al. Adipose-derivedstem cells differentiation into neuron-like cells induced by co-culture with schwann cells[J]. Chin J Repar Reconstr Surg (中国修复重建外科杂志), 2015, 29:97-102.
[20] Hu KH, Yao Y. Advances in application of three-dimensional cell culture[J]. J Med Mol Biol (医学分子生物学杂志), 2008, 5:185-188.
[21] Hopkins AM, DeSimone E, Chwalek K, et al. 3D in vitro modeling of the central nervous system[J]. Prog Neurobiol, 2015, 125:1-25.
[22] Cucullo L, Marchi N, Hossain M, et al. A dynamic in vitro BBB model for the study of immune cell trafficking into the central nervous system[J]. J Cereb Blood Flow Metab, 2011, 31:767-777.
[23] Wang SS, Good TA. Effect of culture in a rotating wall bioreactor on the physiology of differentiated neuron-like PC12 and SH-SY5Y cells[J]. J Cell Biochem, 2001, 83:574-584.
[24] Nishitsuji K, Hosono T, Nakamura T, et al. Apolipoprotein E regulates the integrity of tight junctions in an isoformdependent manner in an in vitro blood-brain barrier model[J]. J Biol Chem, 2011, 286:17536-17542.
[25] Matsusaki M, Case CP, Akashi M. Three-dimensional cell culture technique and pathophysiology[J]. Adv Drug Deliv Rev, 2014, 74:95-103.
[26] Johnstone B, Yoo J, Stewart M. Cell sources for cartilage tissue engineering[M]//Vunjak-Novakovic G, Freshney RI. Culture of Cells for Tissue Engineering. Hoboken New Jersey:John Wiley & Sons, 2006.
[27] Mosahebi A, Wiberg M, Terenghi G. Addition of fibronectin to alginate matrix improves peripheral nerve regeneration in tissue-engineered conduits[J]. Tissue Eng, 2003, 9:209-218.
[28] Go AS, Mozaffarian D, Roger VL, et al. Heart disease and stroke statistics-2013 update:a report from the American Heart Association[J]. Circulation, 2013, 127:e6-e245.
[29] del Zoppo GJ, Hallenbeck JM. Advances in the vascular pathophysiology of ischemic stroke[J]. Thromb Res, 2000, 98:73-81.
[30] Wang Y, Reis C, Applegate R 2nd, et al. Ischemic conditioninginduced endogenous brain protection:applications pre-, per-or post-stroke[J]. Exp Neurol, 2015, 272:26-40.
[31] Han HS, Suk K. The function and integrity of the neurovascular unit rests upon the integration of the vascular and inflammatory cell systems[J]. Curr Neurovasc Res, 2005, 2:409-423.
[32] Woodruff TM, Thundyil J, Tang SC, et al. Pathophysiology, treatment, and animal and cellular models of human ischemic stroke[J]. Mol Neurodegener, 2011, 6:11.
[33] del Zoppo GJ, Sharp FR, Heiss WD, et al. Heterogeneity in the penumbra[J]. J Cereb Blood Flow Metab, 2011, 31:1836-1851.
[34] Mo SJ, Tong XZ, Zhong Q, et al. Bone marrow-derived mesenchymal stem cells protect against hypoxia-induced injury and apoptosis of PC12 cells via up-regulation of erythropoietin expression[J]. Chin J Pathophysiol (中国病理生理杂志), 2013, 29:62-69.
[35] Puyal J, Ginet V, Clarke PG. Multiple interacting cell death mechanisms in the mediation of excitotoxicity and ischemic brain damage:a challenge for neuroprotection[J]. Prog Neurobiol, 2013, 105:24-48.
[36] Brown DR. Neurons depend on astrocytes in a coculture system for protection from glutamate toxicity[J]. Mol Cell Neurosci, 1999, 13:379-389.
[37] Haseloff RF, Dithmer S, Winkler L, et al. Transmembrane proteins of the tight junctions at the blood-brain barrier:structural and functional aspects[J]. Semin Cell Dev Biol, 2015, 38:16-25.
[38] Dirnagl U, Iadecola C, Moskowitz MA. Pathobiology of ischaemic stroke:an integrated view[J]. Trends Neurosci, 1999, 22:391-397.
[39] Kuhlmann CR, Tamaki R, Gamerdinger M, et al. Inhibition of the myosin light chain kinase prevents hypoxia-induced blood-brain barrier disruption[J]. J Neurochem, 2007, 102:501-507.
[40] Kuhlmann CR, Zehendner CM, Gerigk M, et al. MK801 blocks hypoxic blood-brain-barrier disruption and leukocyte adhesion[J]. Neurosci Lett, 2009, 449:168-172.
[41] Muoio V, Persson PB, Sendeski M. The neurovascular unit-concept review[J]. Acta Physiol, 2014, 210:790-798.
[42] Bekris LM, Mata IF, Zabetian CP. The genetics of Parkinson disease[J]. J Geriatr Psychiatry Neurol, 2010, 23:228-242.
[43] Jenner P, Olanow CW. The pathogenesis of cell death in Parkinson's disease[J]. Neurology, 2006, 66:S24-S36.
[44] Rudow G, O'Brien R, Savonenko AV, et al. Morphometry of the human substantia nigra in ageing and Parkinson's disease[J]. Acta Neuropathol, 2008, 115:461-470.
[45] Ribeiro RP, Moreira EL, Santos DB, et al. Probucol affords neuroprotection in a 6-OHDA mouse model of Parkinson's disease[J]. Neurochem Res, 2013, 38:660-668.
[46] Lu C, Zhang J, Shi X, et al. Neuroprotective effects of tetramethylpyrazine against dopaminergic neuron injury in a rat model of Parkinson's disease induced by MPTP[J]. Int J Biol Sci, 2014, 10:350-357.
[47] Wang F, Awan UF, Wang Y, et al. Damage of neuroblastoma cell SH-SY5Y mediated by MPP+ inhibits proliferation of T-cell leukemia Jurkat by co-culture system[J]. Int J Mol Sci, 2014, 15:10738-10750.
[48] Rappold PM, Tieu K. Astrocytes and therapeutics for Parkinson's disease[J]. Neurotherapeutics, 2010, 7:413-423.
[49] Terashvili M, Sarkar P, Nostrand MV, et al. The protective effect of astrocyte-derived 14, 15-epoxyeicosatrienoic acid on hydrogen peroxide-induced cell injury in astrocyte-dopaminergic neuronal cell line co-culture[J]. Neuroscience, 2012, 223:68-76.
[50] Ikemoto S. Dopamine reward circuitry:two projection systems from the ventral midbrain to the nucleus accumbensolfactory tubercle complex[J]. Brain Res Rev, 2007, 56:27-78.
[51] Liu S, Tian Z, Yin F, et al. Generation of dopaminergic neurons from human fetal mesencephalic progenitors after co-culture with striatal-conditioned media and exposure to lowered oxygen[J]. Brain Res Bull, 2009, 80:62-68.
[52] Ittner LM, Götz J. Amyloid-β and tau-a toxic pas de deux in Alzheimer's disease[J]. Nat Rev Neurosci, 2011, 12:65-72.
[53] Qin B, Cartier L, Dubois-Dauphin M, et al. A key role for the microglial NADPH oxidase in APP-dependent killing of neurons[J]. Neurobiol Aging, 2006, 27:1577-1587
[54] Hu J, Van Eldik LJ. Glial-derived proteins activate cultured astrocytes and enhance amyloid-induced glial activation[J]. Brain Res, 1999, 842:46-54.
[55] Culbert AA, Skaper SD, Howlett DR, et al. MAPK-activated protein kinase 2 deficiency in microglia inhibits pro-inflammatory mediator release and resultant neurotoxicity. Relevance to neuroinflammation a transgenic mouse model of Alzheimer disease[J]. J Biol Chem, 2006, 281:23658-23667.
[56] Liu R, Zhang TT, Wu CX, et al. Targeting the neurovascular unit:development of a new model and consideration for novel strategy for Alzheimer's disease[J]. Brain Res Bull, 2011, 86:13-21.
[57] Turner MR, Bowser R, Bruijn L, et al. Mechanisms, models and biomarkers in amyotrophic lateral sclerosis[J]. Amyotroph Lateral Scler Frontotemporal Degener, 2013, 14:19-32.
[58] Wen W, Sanelli T, Ge W, et al. Activated microglial supernatant induced motor neuron cytotoxicity is associated with upregulation of the TNFR1 receptor[J]. Neurosci Res, 2006, 55:87-95.
[59] Naruse H, Iwata A, Takahashi Y, et al. Familial amyotrophic lateral sclerosis with novel A4D SOD1 mutation with late age at onset and rapid progressive course[J]. Neurol Clin Neurosci, 2013, 1:45-47.
[60] Kunze A, Lengacher S, Dirren E, et al. Astrocyte-neuron co-culture on microchips based on the model of SOD mutation to mimic ALS[J]. Integr Biol, 2013, 5:964-975.
[61] Haastert K, Grosskreutz J, Jaeckel M, et al. Rat embryonic motoneurons in long-term co-culture with Schwann cells-a system to investigate motoneuron diseases on a cellular level in vitro[J]. J Neurosci Methods, 2005, 142:275-284.
[62] Tohda C, Kuboyama T. Current and future therapeutic strategies for functional repair of spinal cord injury[J]. Pharmacol Ther, 2011, 132:57-71.
[63] Pinkernelle J, Fansa H, Ebmeyer U, et al. Prolonged minocycline treatment impairs motor neuronal survival and glial function in organotypic rat spinal cord cultures[J]. PLoS One, 2013, 8:e73422.
[64] Alberts B, Bray D, Hopkin K, et al. Essential Cell Biology[M]. New York:Garland Science, 2013.
[65] Hawkins BT, Davis TP. The blood-brain barrier/neurovascular unit in health and disease[J]. Pharmacol Rev, 2005, 57:173-185.
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