Three-dimensional (3D) brain microphysiological system for organophosphates and neurochemical agent toxicity screening

Autoři: Lumei Liu aff001;  Youngmi Koo aff001;  Chukwuma Akwitti aff001;  Teal Russell aff001;  Elaine Gay aff002;  Daniel T. Laskowitz aff003;  Yeoheung Yun aff001
Působiště autorů: FIT BEST Laboratory, Department of Chemical, Biological, and Bio Engineering, North Carolina Agricultural and Technical State University, Greensboro, North Carolina, United States of America aff001;  Center for Drug Discovery, RTI International, Research Triangle Park, Durham, North Carolina, United States of America aff002;  Departments of Neurology, Anesthesiology, and Neurobiology, Brain Injury Translational Research Center, Duke University, Durham, North Carolina, United States of America aff003
Vyšlo v časopise: PLoS ONE 14(11)
Kategorie: Research Article
doi: 10.1371/journal.pone.0224657


We investigated a potential use of a 3D tetraculture brain microphysiological system (BMPS) for neurotoxic chemical agent screening. This platform consists of neuronal tissue with extracellular matrix (ECM)-embedded neuroblastoma cells, microglia, and astrocytes, and vascular tissue with dynamic flow and membrane-free culture of the endothelial layer. We tested the broader applicability of this model, focusing on organophosphates (OPs) Malathion (MT), Parathion (PT), and Chlorpyrifos (CPF), and chemicals that interact with GABA and/or opioid receptor systems, including Muscimol (MUS), Dextromethorphan (DXM), and Ethanol (EtOH). We validated the BMPS platform by measuring the neurotoxic effects on barrier integrity, acetylcholinesterase (AChE) inhibition, viability, and residual OP concentration. The results show that OPs penetrated the model blood brain barrier (BBB) and inhibited AChE activity. DXM, MUS, and EtOH also penetrated the BBB and induced moderate toxicity. The results correlate well with available in vivo data. In addition, simulation results from an in silico physiologically-based pharmacokinetic/pharmacodynamic (PBPK/PD) model that we generated show good agreement with in vivo and in vitro data. In conclusion, this paper demonstrates the potential utility of a membrane-free tetraculture BMPS that can recapitulate brain complexity as a cost-effective alternative to animal models.

Klíčová slova:

Astrocytes – Blood – Esterases – Gamma-aminobutyric acid – Microglial cells – Neurons – Toxic agents – Toxicity


1. Eto M. Organophosphorus pesticides: CRC press; 2018.

2. Slotkin T. Does early-life exposure to organophosphate insecticides lead to prediabetes and obesity? Reprod Toxicol. 2011;31(3):297–301. doi: 10.1016/j.reprotox.2010.07.012 20850519

3. Slotkin TA, Levin ED, Seidler FJ. Comparative Developmental Neurotoxicity of Organophosphate Insecticides: Effects on Brain Development Are Separable from Systemic Toxicity. Environmental Health Perspectives. 2006;114(5):746–51. doi: 10.1289/ehp.8828 PubMed PMID: PMC1459930. 16675431

4. Koo Y, Hawkins BT, Yun Y. Three-dimensional (3D) tetra-culture brain on chip platform for organophosphate toxicity screening. Scientific reports. 2018;8(1):2841. doi: 10.1038/s41598-018-20876-2 29434277

5. Hawkins BT, Hu T, Dougherty ER, Grego S. Modeling neuroinflammatory effects after chemical exposures in a scalable, three-dimensional cell culture system. Applied in vitro toxicology. 2016;2(4):223–34.

6. Gearhart JM, Jepson GW, Clewell HJ, Andersen ME, Conolly RB. Physiologically based pharmacokinetic model for the inhibition of acetylcholinesterase by organophosphate esters. Environmental health perspectives. 1994;102(suppl 11):51–60.

7. Timchalk C, Nolan R, Mendrala A, Dittenber D, Brzak K, Mattsson J. A physiologically based pharmacokinetic and pharmacodynamic (PBPK/PD) model for the organophosphate insecticide chlorpyrifos in rats and humans. Toxicological Sciences. 2002;66(1):34–53. doi: 10.1093/toxsci/66.1.34 11861971

8. Yi Y, Park J, Lim J, Lee CJ, Lee S-H. Central nervous system and its disease models on a chip. Trends in biotechnology. 2015;33(12):762–76. doi: 10.1016/j.tibtech.2015.09.007 26497426

9. Rahman NA, Rasil ANaHM, Meyding-Lamade U, Craemer EM, Diah S, Tuah AA, et al. Immortalized endothelial cell lines for in vitro blood–brain barrier models: A systematic review. Brain research. 2016;1642:532–45. doi: 10.1016/j.brainres.2016.04.024 27086967

10. Lenz DE, Clarkson ED, Schulz SM, Cerasoli DM. Butyrylcholinesterase as a therapeutic drug for protection against percutaneous VX. Chemico-biological interactions. 2010;187(1–3):249–52. doi: 10.1016/j.cbi.2010.05.014 20513442

11. EVARREST® Fibrin Sealant Patch, authors FDA Approval Letter. 2012.

12. Jo SH, Mathiasen RA, Gurushanthaiah D. Prospective, randomized, controlled trial of a hemostatic sealant in children undergoing adenotonsillectomy. Otolaryngology—Head and Neck Surgery. 2007;137(3):454–8. doi: 10.1016/j.otohns.2006.09.020 17765775

13. Mozet C, Prettin C, Dietze M, Fickweiler U, Dietz A. Use of Floseal and effects on wound healing and pain in adults undergoing tonsillectomy: randomised comparison versus electrocautery. European Archives of Oto-Rhino-Laryngology. 2012;269(10):2247–54. doi: 10.1007/s00405-011-1904-4 22207530

14. Nesheim O, Criswell J. Pesticide Applicator Certification Series: toxicity of pesticides [LD50 values, includes lists of common and trade names]. OSU extension facts-Cooperative Extension Service, Oklahoma State University (USA). 1982.

15. Koziol FS, Witkowski JF. Synergism studies with binary mixtures of permethrin plus methyl parathion, chlorpyrifos, and malathion on European corn borer larvae. Journal of Economic Entomology. 1982;75(1):28–30.

16. Budavari S. The Merck Index, an encyclopedia of chemical drug, and biologicals. Merck, 1989.

17. Echave M, Oyagüez I, Casado MA. Use of Floseal®, a human gelatine-thrombin matrix sealant, in surgery: a systematic review. BMC surgery. 2014;14(1):111.

18. Itzhak BRaB. The Risks of Haemostatic Materials in Tonsillectomy. Archives of Otolaryngology and Rhinology. 2015;1(2):046–7.

19. Gearhart JM, Jepson GW, Clewell III HJ, Andersen ME, Conolly RB. Physiologically based pharmacokinetic and pharmacodynamic model for the inhibition of acetylcholinesterase by diisopropyfluorophosphate. Toxicology and applied pharmacology. 1990;106(2):295–310. doi: 10.1016/0041-008x(90)90249-t 2256118

20. Poet TS, Timchalk C, Hotchkiss JA, Bartels MJ. Chlorpyrifos PBPK/PD model for multiple routes of exposure. Xenobiotica. 2014;44(10):868–81. doi: 10.3109/00498254.2014.918295 24839995

21. Wevers NR, Van Vught R, Wilschut KJ, Nicolas A, Chiang C, Lanz HL, et al. High-throughput compound evaluation on 3D networks of neurons and glia in a microfluidic platform. Scientific reports. 2016;6:38856. doi: 10.1038/srep38856 27934939

22. Hole G. Eight things you need to know about interpreting correlations. 2014.

23. Rush T, Liu X, Hjelmhaug J, Lobner D. Mechanisms of chlorpyrifos and diazinon induced neurotoxicity in cortical culture. Neuroscience. 2010;166(3):899–906. doi: 10.1016/j.neuroscience.2010.01.025 20096330

24. Park JH, Ko J, Hwang J, Koh HC. Dynamin-related protein 1 mediates mitochondria-dependent apoptosis in chlorpyrifos-treated SH-SY5Y cells. Neurotoxicology. 2015;51:145–57. doi: 10.1016/j.neuro.2015.10.008 26598294

25. Carlson K, Jortner BS, Ehrich M. Organophosphorus compound-induced apoptosis in SH-SY5Y human neuroblastoma cells. Toxicology and applied pharmacology. 2000;168(2):102–13. doi: 10.1006/taap.2000.8997 11032765

26. Kashyap M, Singh A, Siddiqui M, Kumar V, Tripathi V, Khanna V, et al. Caspase cascade regulated mitochondria mediated apoptosis in monocrotophos exposed PC12 cells. Chemical research in toxicology. 2010;23(11):1663–72. doi: 10.1021/tx100234m 20957986

27. Kashyap MP, Singh AK, Kumar V, Yadav DK, Khan F, Jahan S, et al. Pkb/Akt1 mediates Wnt/GSK3β/β-catenin signaling-induced apoptosis in human cord blood stem cells exposed to organophosphate pesticide monocrotophos. Stem cells and development. 2012;22(2):224–38. doi: 10.1089/scd.2012.0220 22897592

28. Tiffany-Castiglioni E, Ehrich M, Dees L, Costa L, Kodavanti P, Lasley S, et al. Bridging the gap between in vitro and in vivo models for neurotoxicology. Toxicological sciences: an official journal of the Society of Toxicology. 1999;51(2):178–83.

29. Ariwodola OJ, Weiner JL. Ethanol potentiation of GABAergic synaptic transmission may be self-limiting: role of presynaptic GABAB receptors. Journal of Neuroscience. 2004;24(47):10679–86. doi: 10.1523/JNEUROSCI.1768-04.2004 15564584

30. Chen C-L, Cheng M-H, Kuo C-F, Cheng Y-L, Li M-H, Chang C-P, et al. Dextromethorphan Attenuates NADPH Oxidase-regulated GSK-3β and NF-κB Activation and Reduces Nitric Oxide Production in Group A Streptococcal Infection. Antimicrobial agents and chemotherapy. 2018:AAC. 02045–17.

31. Van den Pol AN, Obrietan K, Chen G. Excitatory actions of GABA after neuronal trauma. Journal of Neuroscience. 1996;16(13):4283–92. doi: 10.1523/JNEUROSCI.16-13-04283.1996 8753889

32. Werling LL, Lauterbach EC, Calef U. Dextromethorphan as a potential neuroprotective agent with unique mechanisms of action. The neurologist. 2007;13(5):272–93. doi: 10.1097/NRL.0b013e3180f60bd8 17848867

33. Liu Y, Qin L, Li G, Zhang W, An L, Liu B, et al. Dextromethorphan protects dopaminergic neurons against inflammation-mediated degeneration through inhibition of microglial activation. Journal of Pharmacology and Experimental Therapeutics. 2003;305(1):212–8. doi: 10.1124/jpet.102.043166 12649371

34. Larson TA, Wang T-W, Gale SD, Miller KE, Thatra NM, Caras ML, et al. Postsynaptic neural activity regulates neuronal addition in the adult avian song control system. Proceedings of the National Academy of Sciences. 2013:201310237.

35. Tateno M, Saito T. Biological studies on alcohol-induced neuronal damage. Psychiatry investigation. 2008;5(1):21–7. doi: 10.4306/pi.2008.5.1.21 20046404

36. Ziemann U, Chen R, Cohen LG, Hallett M. Dextromethorphan decreases the excitability of the human motor cortex. Neurology. 1998;51(5):1320–4. doi: 10.1212/wnl.51.5.1320 9818853

Článek vyšel v časopise


2019 Číslo 11