Protective effect of lipopolysaccharide-preconditioned astrocyte extracellular vesicles on neurons

doi:10.12307/2022.322

Abstract

Abstract: BACKGROUND: Central nervous system diseases are often closely related to neuronal apoptosis. Astrocytes are closely related to neurons. It remains poorly understood whether extracellular vesicles, which are derived from astrocytes, especially activated astrocytes, may inhibit glutamate-induced neuronal apoptosis, thereby exerting neuroprotective effect.
OBJECTIVE: To investigate the role and possible mechanism of astrocyte extracellular vesicles (AS-EVs) and lipopolysaccharide-preconditioned astrocyte extracellular vesicles (LPAS-EVs) in glutamate-induced neuronal injury.
METHODS: Astrocyte was cultured in vitro and induced activation model was established by lipopolysaccharide. The supernatant of cells of two groups was collected and extracellular vesicles were extracted by hypervelocity centrifuges, and then identified. PC12 cells were used for preliminary experiments. First, we investigated the effects of AS-EVs and LPAS-EVs on PC12 cell proliferation, and then established the PC12 cells and primary neurons injury model induced by glutamate. Experiment was divided into four groups: PBS group (PBS of the same volume as EVs groups, 24 hours), glutamate group (10 mmol/L glutamate, 24 hours), AS-EVs group (200 mg/L AS-EVs, 24 hours + 10 mmol/L glutamate, 24 hours), and LPAS-EVs group (200 mg/L LPAS-EVs, 24 hours + 10 mmol/L glutamate, 24 hours). The survival rate of PC12 cells was determined by CCK-8 assay. Western blot assay was used to detect the expression of apoptosis related proteins Bax and Bcl-2.
RESULTS AND CONCLUSION: (1) Astrocyte was extracted and lipopolysaccharide-induced astrocyte activation model was established successfully. (2) AS-EVs and LPAS-EVs were successfully extracted, which met the morphological standards of extracellular vesicles and showed positive expression of related markers. (3) AS-EVs and LPAS-EVs had no significant effect on PC12 cells proliferation. (4) Compared with glutamate group, AS-EVs and LPAS-EVs could improve the survival rate of PC12 cells, and the effect of LPAS-EVs was stronger than that of AS-EVs (all P < 0.05, n=3). (5) AS-EVs and LPAS-EVs could inhibit the pro-apoptotic protein Bax expression and promote the expression of anti-apoptotic protein Bcl-2 (all P < 0.01, n=3), and then inhibited apoptosis of PC12 cells and primary neurons, thus playing a neuroprotective role, and the effect of LPAS-EVs was stronger than that of AS-EVs (all P < 0.05). (6) The results conclude that extracellular vesicles derived from astrocytes can inhibit the apoptosis of neurons injured by glutamate, which has a neuroprotective effect, and this effect is enhanced after being stimulated by lipopolysaccharide.

Key words: astrocytes, extracellular vesicles, lipopolysaccharide, PC12 cells, neurons, apoptosis

CLC Number:

Sun Haitao, Huang Yonghui, Gong Aihua, Cao Xingbing, Li Zhen. Protective effect of lipopolysaccharide-preconditioned astrocyte extracellular vesicles on neurons[J]. Chinese Journal of Tissue Engineering Research, 2022, 26(13): 1985-1992.

Figures/Tables 6

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