Human platelet concentrates: a source of solvent/detergent-treated highly enriched brain-derived neurotrophic factor.
BACKGROUND
Human blood platelets (PLTs) contain brain-derived neurotrophic factor (BDNF), a neurotrophin that binds to neurotrophic tropomyosin-related kinase B (TrkB) receptor on central nervous system cells. This binding promotes neural synaptic plasticity and memory and prevents neuronal degeneration. Alterations in BDNF homeostasis are associated with aging and are found in several neurodegenerative conditions such as Alzheimer's, Huntington's, and Parkinson's diseases and multiple sclerosis. We have developed PLT viral inactivation and chromatographic fractionation processes and decided here to identify fractions enriched in BDNF.


METHODS
PLT concentrates (PCs) were treated by solvent/detergent (S/D), extracted by oil, and subjected to fractionation (C18, sulfopropyl [SP]-Sepharose, diethylaminoethyl [DEAE]-Sepharose, or activated charcoal). BDNF and pro-BDNF were evaluated by enzyme-linked immunosorbent assay, and Western blot. TrkB was studied by Western blot. Tri-n-butyl phosphate (TnBP) was quantified by high-performance liquid chromatography, and Triton X-45 by gas chromatography.


RESULTS
The mean BDNF content of 2.9 ± 0.7 ng/mL in PC was noted to increase to 56.2 ± 2.4 ng/mL after S/D treatment and remained stable during oil extraction. Approximately 70% of the BDNF content was recovered after C18 chromatography. BDNF did not bind to DEAE-Sepharose and was almost completely adsorbed by charcoal. Chromatography on SP-Sepharose yielded a highly enriched 13-kDa mature BDNF fraction that was more than 170-fold purified, with a mean of 137 ± 29.4 ng/mL and 82% chromatographic recovery, devoid of detectable TnBP and Triton X-45. Pro-BDNF and TrkB proteins were not detected in the PLT extracts.


CONCLUSIONS
We obtained a S/D-treated, highly enriched mature PLT-derived BDNF fraction that could help unveil the pharmacokinetics, pharmacodynamic, and potential therapeutic applications of the BDNF neurotrophin.
College of Oral Medicine, Taipei Medical University, Taipei, Taiwan.
Thierry
T
Burnouf
Ya-Po
YP
Kuo
David
D
Blum
Sylvie
S
Burnouf
Chen-Yao
CY
Su
D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D015153:Blotting, Western / Q000379:methods; D019208:Brain-Derived Neurotrophic Factor / Q000302:isolation & purification* / Q000378:metabolism*; D002458:Cell Fractionation / Q000379:methods*; D002849:Chromatography, Gas / Q000379:methods; D002853:Chromatography, Liquid / Q000379:methods; D003902:Detergents / Q000494:pharmacology*; D006624:Hippocampus / Q000378:metabolism; D006801:Humans; D051379:Mice; D010976:Platelet Count; D010983:Plateletpheresis / Q000379:methods; D020813:Receptor, trkB / Q000378:metabolism; D012997:Solvents / Q000494:pharmacology*
D019208:Brain-Derived Neurotrophic Factor; D003902:Detergents; D012997:Solvents; D020813:Receptor, trkB