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Probenecid relieves LPS-induced neuroinflammation by modulating the NLRP1 inflammasome pathway in BV2 cells
Xiaoxuan Wang1, Haozhe Tian2,3, Shifei Chen1
1Clinical Laboratory, Second People's Hospital of Bengbu City, Bengbu, 233000, China.
Abstract:
Inflammation is a key factor in neurodegenerative diseases (NDs) development, such as Alzheimer's disease (AD) and Parkinson's disease (PD). Microglial cells maintain immune microenvironment homeostasis in the central nervous system (CNS) and are actively involved in neuroinflammation. Therefore, inhibiting microglia-mediated neuroinflammation may provide therapeutic benefits in treating NDs. Probenecid (Prob), a sulfonamide derivative, has demonstrated neuroprotective and anti-inflammatory properties. This study investigated the anti-inflammatory and antioxidative properties of Prob in a neuroinflammatory model which was established by stimulating BV2 cells with lipopolysaccharide (LPS). The oxidative stress was assessed by measuring the nitric oxide (NO), reactive oxygen species (ROS), superoxide dismutase (SOD), and malondialdehyde (MDA) levels. Tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were quantified using enzyme-linked immunosorbent assay (ELISA) and RT-qPCR. The wound-healing assay and phagocytosis assay were employed to examine the migration and phagocytosis capacity of BV2 cells, respectively. The mRNA expression of CD16, CD206, iNOS, and Arg-1 was measured using RT-qPCR, while Western blot was used to analyze inflammasome-associated protein levels. The results indicated that Prob suppressed LPS-induced microglial activation and reduced TNF-α and IL-1β production. Moreover, Prob attenuated the migration and phagocytosis capacities of BV2 cells and inhibited oxidative stress. Prob regulated the immune microenvironment, as shown by reduced M1 markers (iNOS and CD16) and increased M2 markers (Arg-1 and CD206). However, the NLRP1 inflammasome activator (muramyl dipeptide, MDP) reversed the effects of Prob. In conclusion, Prob could alleviate LPS-induced inflammation by modulating the NLRP1 inflammasome pathway.
Insights
Probenecid (Prob) reduces neuroinflammation by inhibiting microglial activation and modulating the NLRP1 inflammasome pathway. This drug shows potential for treating neurodegenerative diseases by decreasing inflammatory markers and oxidative stress.
Area of Science:
- Neuroscience
- Immunology
- Pharmacology
Background:
- Neuroinflammation, driven by microglial cells, is central to neurodegenerative diseases (NDs) like Alzheimer's and Parkinson's.
- Inhibiting microglial-mediated neuroinflammation offers a therapeutic strategy for NDs.
- Probenecid (Prob), a known anti-inflammatory agent, exhibits neuroprotective properties.
Purpose of the Study:
- To investigate the anti-inflammatory and antioxidative effects of Probenecid (Prob) in a lipopolysaccharide (LPS)-induced neuroinflammation model using BV2 microglial cells.
- To elucidate the underlying mechanisms, including the role of the NLRP1 inflammasome pathway.
Main Methods:
- BV2 cells were stimulated with LPS to induce neuroinflammation.
- Probenecid's effects were assessed by measuring oxidative stress markers (NO, ROS, SOD, MDA), inflammatory cytokines (TNF-α, IL-1β), cell migration, and phagocytosis.
- Gene and protein expression of microglial markers (CD16, CD206, iNOS, Arg-1) and inflammasome components were analyzed via RT-qPCR and Western blot.
- The role of the NLRP1 inflammasome was investigated using muramyl dipeptide (MDP).
Main Results:
- Probenecid significantly suppressed LPS-induced microglial activation, reducing TNF-α and IL-1β levels.
- Prob attenuated BV2 cell migration and phagocytosis, while inhibiting oxidative stress markers.
- Probenecid modulated the microglial immune microenvironment, decreasing M1 markers (iNOS, CD16) and increasing M2 markers (Arg-1, CD206).
- The effects of Prob were reversed by MDP, indicating the involvement of the NLRP1 inflammasome.
Conclusions:
- Probenecid exhibits significant anti-inflammatory and antioxidative properties in a microglial cell model of neuroinflammation.
- Prob alleviates neuroinflammation by modulating the NLRP1 inflammasome pathway and shifting microglial polarization from M1 to M2 phenotype.
- These findings suggest Probenecid as a potential therapeutic agent for neurodegenerative diseases characterized by neuroinflammation.
