Effects of aspirin and clopidogrel on neural stem cells

Abstract

Cerebral infarction causes severe morbidity and mortality. Most patients with cerebral infarction should take antiplatelet drugs daily, so the effects of those drugs on the regeneration of the brain need to be investigated. Aspirin and clopidogrel are the most widely used antiplatelet drugs for the prevention of ischemic stroke. We investigated the effects of aspirin and clopidogrel on neural stem cells (NSCs). NSCs were dissociated from fetal rat cortex and cultured with basic fibroblast growth factor and N2 medium. To measure the effects of aspirin and clopidogrel on NSCs, NSCs were treated with several concentrations of aspirin, clopidogrel bisulfate, and clopidogrel resinate for 24 h. After the treatment, we measured cell viability by cell counting kit-8, MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, trypan blue staining, flow cytometry, and lactate dehydrogenase assay. To evaluate their effects on NSC proliferation, we performed BrdU cell proliferation assay and colony-forming unit assay. We compared the intracellular protein level in the NSCs treated with aspirin and two types of clopidogrel, by proteomics analysis. Various viability tests showed that clopidogrel resinate and clopidogrel bisulfate did not affect the viability and proliferation of NSCs whereas aspirin decreased them even at low concentrations which are clinically relevant. Moreover, through the proteomics, it was confirmed that the toxicity of aspirin to NSCs might be associated with the alteration of several intracellular proteins. Taken together, these results suggest that clopidogrel resinate and clopidogrel bisulfate are safe but aspirin could be toxic to NSCs. Therefore, when these antiplatelet agents are prescribed over the long-term, the finding that aspirin could be toxic to NSCs should be considered.

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Abbreviations

NSCs:

Neural stem cells

MTT:

3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide

CNS:

Central nervous system

SVZ:

Subventricular zone

SGZ:

Subgranular zone

ASA:

Acetylsalicylic acid

DAPT:

Dual antiplatelet therapy

bFGF:

Basic fibroblast growth factor

DMSO:

Dimethyl sulfoxide

CCK-8:

Cell counting kit-8

LDH:

Lactate dehydrogenase

WST-8:

2-(2-Methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium

DTT:

Dithiothreitol

CFU:

Colony-forming unit

2-DE:

Two-dimensional gel electrophoresis

CHAPS:

3-[(3-Cholamidopropyl) dimethylammonio]-1-propanesulfonate

IEF:

Isoelectric focusing

CBB:

Coomassie brilliant blue R-350

PMF:

Peptide mass fingerprinting

MALDI-TOF/MS:

Matrix-assisted laser desorption/ionization-time of flight mass spectrometer

COX-2:

Cyclooxygenase-2

NDUFA10:

NADH dehydrogenase 1 subunit 10

P5C:

1-Pyrroline-5-carboxylate

ProDH:

Proline dehydrogenase

P5CDH:

P5C dehydrogenase

CKB:

Creatine kinase brain-type

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Funding information

This work was supported by the Basic Science Research Program of the National Research Foundation of Korea funded by the Ministry of Science, ICT and Future Planning (2015R1A2A2A04004865), by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number: HI17C2160), and by Korea Drug Development Fund (KDDF) funded by Ministry of Science and ICT, Ministry of Trade, Industry & Energy and Ministry of Health & Welfare (KDDF-201609-02, Republic of Korea).

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Correspondence to Seong-Ho Koh.

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Hwang, M., Park, H., Choi, H. et al. Effects of aspirin and clopidogrel on neural stem cells. Cell Biol Toxicol 34, 219–232 (2018). https://doi.org/10.1007/s10565-017-9412-y

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Keywords

  • Neural stem cells
  • Aspirin
  • Clopidogrel bisulfate
  • Clopidogrel resinate
  • Proliferation
  • Viability