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Objective To investigate the mechanism by which Senegenin (SEN) alleviates microglial inflammatory response through the nuclear factor erythroid 2-related factor 2 (Nrf2)/NOD-like receptor protein 3 (NLRP3) pathway. Methods BV2 mouse microglia cells were randomly divided into control group, model group, SEN group and MCC950 group. Cells in control group were not treated, and cells in model group were added with 1 μg/ml lipopolysaccharide (LPS); Cells in SEN group were added with 1 μg/ml LPS+4 μmol/L SEN, and cells in MCC950 group were added with 1 μg/ml LPS+10 μmol/L MCC950 for 24 hours. CCK-8 method was used to detect the effect of different concentrations of SEN on the viability of BV2 cells. Griess method was used to determine the release amount of nitric oxide (NO) in the supernatant. Real-time fluorescent quantitative PCR was used to determine the mRNA expression levels of NLRP3, lymphocyte apoptosis-associated spect-like protein containing a CARD (ASC), caspase-1, interleukin (IL)-1β and IL-18 mRNA. Immunofluorescence staining was used to detect the expression levels of ASC, IL-1β, Nrf2 and heme oxygenase-1 (HO-1). Western blotting was used to detect the expression levels of NLRP3, caspase-1, ASC, IL-1β, IL-18, Nrf2, HO-1, nuclear factor kappa B (NF-κB) and inducible nitric oxide synthase (iNOS). Results The results of CCK-8 method showed that there was no significant difference in the viability of BV2 cells treated with 2~20 μmol/L SEN compared with control group (P>0.05). Compared with control group, the viability of BV2 cells in model group decreased significantly (P<0.05). Compared with model group, the viability of BV2 cells in 4 μmol/L SEN group was significantly restored (P<0.05). Compared with control group, the results of Griess method showed that the release amount of NO in cells of model group increased significantly (P<0.05); the results of real-time PCR showed that the expression levels of NLRP3, ASC, caspase-1, IL-1β and IL-18 mRNA in cells of model group increased significantly (P<0.05); the results of Western blotting showed that the protein expression levels of NLRP3, ASC, caspase-1, IL-1β and IL-18 proteins in cells of model group increased significantly (P<0.05), and the immunofluorescence staining results showed that the expression levels of iNOS and NF-κB protein in cells of model group increased, and the expression levels of Nrf2 and HO-1 decreased, with statistically significant differences (P<0.05). Compared with model group, the release amount of NO in cells of SEN group and MCC950 group decreased, and the expression levels of NLRP3, ASC, caspase-1, IL-1β and IL-18 mRNA and proteins decreased, with statistically significant differences (P<0.05); in the SEN group, the expression levels of iNOS and NF‑κB decreased, and immunofluorescence staining showed that Nrf2 was translocated into the nucleus, and the expression levels of Nrf2 and HO-1 proteins increased significantly, with statistically significant differences (P<0.05). Conclusions SEN could alleviate the inflammatory response of mouse microglia cells induced by LPS and inhibit the activation and expression of NLRP3 inflammasome, with an effect comparable to that of the inflammasome inhibitor MCC950. The mechanism may be related to the regulation of the expression of upstream factors Nrf2 and HO-1.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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