Abstract
Angiotensin II receptor blockers (ARBs) have been shown to exert neuroprotective effects by suppressing inflammatory and apoptotic responses. In the present study, the effects of the ARB telmisartan on the NLRP3 inflammasome induced by oxygen-glucose deprivation (OGD) in neural stem cells (NSCs) were investigated, as well as their possible association with the activation of the PI3K pathway. Cultured NSCs were treated with different concentrations of telmisartan and subjected to various durations of OGD. Cell counting, lactate dehydrogenase, bromodeoxyuridine, and colony-forming unit assays were performed to measure cell viability and proliferation. In addition, the activity of intracellular signaling pathways associated with the PI3K pathway and NLRP3 inflammasome was evaluated. Telmisartan alone did not affect NSCs up to a concentration of 10 μM under normal conditions but showed toxicity at a concentration of 100 μM. Moreover, OGD reduced the viability of NSCs in a time-dependent manner. Nevertheless, treatment with telmisartan increased the viability and proliferation of OGD-injured NSCs. Furthermore, telmisartan promoted the expression of survival-related proteins and mRNA while inhibiting the expression of death-related proteins induced by OGD. In particular, telmisartan attenuated OGD-dependent expression of the NLRP3 inflammasome and its related signaling proteins. These beneficial effects of telmisartan were blocked by a PI3K inhibitor. Together, these results indicate that telmisartan attenuated the activation of the NLRP3 inflammasome by triggering the PI3K pathway, thereby contributing to neuroprotection.
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We thank all members of Prof. Koh’s Lab for scientific advice and helps.
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This research was supported by the Basic Science Research Program of the National Research Foundation of Korea, which is funded by the Ministry of Science, ICT, and Future Planning (2018R1A2A2A15023219); by a grant from 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 HI18C1254); and by the Medical Research Center (2017R1A5A2015395).
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The study conception and design: K.H.S, H.J, and K.S-H. Investigation: H.J., P.H.H, and L.E-H. Data analysis and interpretation of data: K.H.S and HJ. Supervision: K.J.Y., C.H., L.K-Y., and L.Y.J. Original draft: K.H.S and H.J. Revised manuscript: K.J.Y. and K.S-H. K.H.S and H.J. contributed equally to this work.
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All animal procedures were conducted in accordance with Hanyang University’s guidelines for the care and use of laboratory animals, and approved by the Institutional Animal Care and Use Committee (IACUC) of Hanyang University (2019-0162A). All efforts were made to minimize the number of animals used and animal suffering.
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Suppl. Fig. 1
Identification of NSCs via immunocytochemistry (a) and RT-PCR (b). Scale bars for immunocytochemistry = 50 μm. RT-PCR analysis: Gapdh, 112 bp; nestin, 234 bp; Sox2, 414 bp. (PNG 629 kb)
Suppl. Fig. 2
Effects of telmisartan and OGD on the viability and cytotoxicity of NSCs. Cell viability (a, trypan blue staining) was significantly decreased and cytotoxicity (b, LDH assay) significantly increased in a time-dependent manner after exposure to OGD. Telmisartan treatment alone up to a concentration of 10 μM did not affect the viability (c) and cytotoxicity (d) of NSCs. Data are presented as mean (% of control) ± SD. Treatment groups were compared with the control group using one-way ANOVA followed by Tukey’s test (n = 4). *P < 0.05, **P < 0.01 (vs. the control group); #P < 0.05, ##P < 0.01 (vs. the group only subjected to an 8-h OGD). (PNG 384 kb)
Suppl. Fig. 3
Photographs of NSCs showing the effects of telmisartan on the viability of NSCs injured by OGD. NSCs were photographed after exposure to OGD at different time points (a), after telmisartan treatment at various concentrations (b), and after both an 8-h exposure to OGD and telmisartan treatment at various concentrations (c). (PNG 2454 kb)
Suppl. Fig. 4
Photographs of NSCs showing the effects of telmisartan on ROS generation via detection of DCF by fluorescent microscopy. (PNG 1271 kb)
Suppl. Fig. 5
Effects of GW9662 on the viability and cytotoxicity of NSCs. GW9662 treatment up to a concentration of 10 μM for 24 h did not affect the viability (a, trypan blue staining) or cytotoxicity (b, LDH assay) of NSCs (a: n = 4 and b: n = 3). (PNG 115 kb)
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Kwon, H.S., Ha, J., Kim, J.Y. et al. Telmisartan Inhibits the NLRP3 Inflammasome by Activating the PI3K Pathway in Neural Stem Cells Injured by Oxygen-Glucose Deprivation. Mol Neurobiol 58, 1806–1818 (2021). https://doi.org/10.1007/s12035-020-02253-1
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DOI: https://doi.org/10.1007/s12035-020-02253-1