Abstract
Objective Paraquat (PQ), a widely used toxic herbicide, induces lung inflammation through mechanisms that remain incompletely understood. In a previous study, we found that the plasma MUC5B mucin level was implicated in PQ poisoning in patients. Here, we hypothesize that MUC5B is a critical mediator in PQ-induced cell inflammation. Methods A mouse model of PQ-induced lung injury was used to examine the MUC5B expression level. A549 cells (alveolar epithelial cells line) were exposed to PQ in dose-dependent and time-dependent manners. Cell viability was detected by CCK-8 assays. The expression levels of MUC5B were examined by dot blot enzyme-linked immunosorbent assay (ELISA) and RT-qPCR. Western blotting was used to detect the levels of proteins in the MAPK and NF-κB pathways. Inflammatory factors in the cell culture medium were measured by ELISA. NF-κB and MAPK pathway inhibitors and MUC5B siRNA (siMUC5B) were used to determine the function of MUC5B. Finally, N-acetyl-cysteine (NAC) was added and its regulatory effect on the MAPK-NF-κB-MUC5B pathway was examined in PQ-induced cell inflammation. Results MUC5B was significantly upregulated accompanying the increases in TNF-α and IL-6 secretion following PQ treatment in mouse and also in A549 cells after treatment with 50 μM PQ at 24 hours. Furthermore, MAPK and NF-κB pathway inhibitors could dramatically decrease the expression of MUC5B and the secretion of TNF-α and IL-6. Importantly, siMUC5B could significantly attenuate the secretion of TNF-α and IL-6 induced by PQ. As expected, the addition of NAC efficiently suppresses the TNF-α and IL-6 secretion stimulated from PQ and also downregulated ERK, JNK, and p65 phosphorylation (ERK/JNK MAPK and NF-κB pathways) as well as MUC5B expression. Conclusion Our findings suggest that MUC5B participates in the process of PQ-induced cell inflammation and is downstream of the NF-κB and MAPK pathways. NAC can attenuate PQ-induced cell inflammation at least in part by suppressing the MAPK-NF-κB-MUC5B pathway. These results nominate MUC5B as a new biomarker and therapeutic target for PQ-induced lung inflammation.
Highlights
Paraquat (PQ, 1, 1′-dimethyl-4, 4′-bipyridinium) is a highly toxic herbicide that is used worldwide, especially in developing countries [1, 2]
Compared with the saline group, significant neutrophil burst was observed from hematoxylin and eosin (HE) staining at day 3 post PQ treatment (Figure 1(i))
The results indicated that the cell viability was not significantly different compared to that of the control groups when the PQ concentration was less than 200 μM (Figure 2(a))
Summary
Paraquat (PQ, 1, 1′-dimethyl-4, 4′-bipyridinium) is a highly toxic herbicide that is used worldwide, especially in developing countries [1, 2]. In the last several years, the incidence of PQ poisoning by accident or suicide has increased in Asia, notably in China [3]. Recent studies have reported that PQ cytotoxicity involves reactive oxygen species (ROS) generation, inflammation. Mucins (MUCs) are a group of highly glycosylated proteins that are classified into membrane-associated MUC and secretory MUC according to their characteristics [7, 8]. MUC5B, a type of secretory mucin, is the most important component of the respiratory secretory system. It is secreted by the goblet cells
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