Abstract
Abuse of pyrrolidinophenone derivatives (PPs) is known to cause severe damage to the central nervous system due to their high lipophilicity. In this study, we compared sensitivity to toxicity elicited by 4′-iodo-α-pyrrolidinononanophenone (I-α-PNP), one of the most potent cytotoxic derivatives among PPs synthesized previously, between SH-SY5Y cells differentiated by all-trans-retinoic acid (ATRA) and the undifferentiated cells, and found that the differentiated cells are more sensitive to I-α-PNP toxicity than the undifferentiated cells. Treatment with I-α-PNP elicited some apoptotic alterations (Bax expression, loss of mitrochondrial membrane potential, and activation of caspases) in the differentiated cells, whose patterns were similar to those in the undifferentiated cells. I-α-PNP treatment resulted in no significant alteration in Bcl-2 expression in the undifferentiated cells, whereas it considerably downregulated the protein expression in the differentiated cells, suggesting that the high I-α-PNP sensitivity of the differentiated cells is mainly due to downregulation of Bcl-2 expression. I-α-PNP treatment decreased nitric oxide (NO) production and neuronal NOS (nNOS) expression in the differentiated cells, and the patterns of I-α-PNP-evoked alterations in phosphorylation of cAMP response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF) expression were almost the same as that in nNOS expression. Additionally, the addition of an NO donor restored the I-α-PNP-evoked alterations in expressions of Bcl-2, BDNF, and nNOS in the differentiated cells. These findings suggest that the downregulation of Bcl-2 expression by I-α-PNP in differentiated cells is attributed to the acceleration of two negative feedback loops (nNOS/NO/CREB loop and CREB/BDNF loop) triggered by decreased NO production.
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Abbreviations
- AMC:
-
7-Amido-4-methylcoumarin
- ATRA:
-
All-trans-retinoic acid
- BDNF:
-
Brain-derived neurotrophic factor
- ATRA ( −):
-
SH-SY5Y without ATRA treatment
- ATRA ( +):
-
Differentiated SH-SY5Y with ATRA treatment
- C-PTIO:
-
2-(4-Carboxyphenyl)-4,4,5,5-tetramethyl imidazoline-1-oxyl-3-oxide
- CREB:
-
CAMP response element-binding protein
- DAF2-DA:
-
Diaminofluorescein-2 diacetate
- DAPI:
-
4′,6-Diamidino-2-phenylindole
- DETA/NO:
-
Diethylenetriamine NONOate
- DPBS:
-
Dulbecco’s phosphate-buffered saline
- ERK:
-
Extracellular signal-regulated kinase
- FBS:
-
Fetal bovine serum
- FITC:
-
Fluorescein isothiocyanate
- iNOS:
-
Inducible NO synthase
- I-α-PNP:
-
4′-Iodo-α-pyrrolidinononanophenone
- MDPV:
-
3,4-Methylenedioxypyrovalerone
- NAC:
-
N-Acetyl-l-cysteine
- nNOS:
-
Neuronal NO synthase
- NO:
-
Nitric oxide
- PCR:
-
Polymerase chain reaction
- PPs:
-
Pyrrolidinophenones
- ROS:
-
Reactive oxygen species
- WST:
-
1,2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium monosodium salt
- α-PNP:
-
α-Pyrrolidinononanophenone
- α-PVP:
-
α-Pyrrolidinovalerophenone
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All the authors contributed to the study conception and design. Material preparation, and data collection and analysis were performed by Yuji Sakai, Yoshifumi Morikawa, Yukino Nagao, and Junta Hattori. Supervision and project administration were performed by Koichi Suenami, Emiko Yanase, Tomohiro Takayama, Akira Ikari, and Toshiyuki Matsunaga. The first draft of the manuscript was written by Yuji Sakai, and all the authors commented on previous versions of the manuscript. All the authors read and approved the final manuscript.
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Sakai, Y., Morikawa, Y., Nagao, Y. et al. 4′-Iodo-α-Pyrrolidinononanophenone Provokes Differentiated SH-SY5Y Cell Apoptosis Through Downregulating Nitric Oxide Production and Bcl-2 Expression. Neurotox Res 40, 1322–1336 (2022). https://doi.org/10.1007/s12640-022-00546-y
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DOI: https://doi.org/10.1007/s12640-022-00546-y