Liyan Kaiyin Formula relieves reflux pharyngitis by regulating M1 macrophage polarization via the NF-κB/NLRP3 pathway

We aimed to investigate whether Liyan Kaiyin Formula (LYKYF) can relieve reflux pharyngitis in rats by regulating M1 macrophage polarization via the nuclear factor-κB (NF-κB)/Nod-like receptor protein 3 (NLRP3) pathway.

Forty rats were randomized into a sham group, a laryngopharyngeal reflux disease (LPRD) group, a LYKYF group and a LYKYF+CHPG group (n = 10). Enzyme-linked immunosorbent assay was conducted to measure the serum levels of inflammatory factors interleukin-6 (IL-6), IL-1β and tumor necrosis factor-α (TNF-α). Western blotting and reverse transcription-polymerase chain reaction (RT-PCR) were performed to measure the expressions of proteins implicated in NF-κB/NLRP3 pathway. Western blotting was also used for the detection of M1 macrophage markers (CD86 and iNOS).

Compared to the sham group, TNF-α, IL-6 and IL-1β levels in the serum, proportion of M1 macrophages in pharyngeal tissues, p-NF-κB p65/p65 ratio, protein expressions of NLRP3, Caspase-1, Apoptosis-associated speck-like protein (ASC), cluster of differentiation 86 (CD86) and inducible nitric oxide synthase, and mRNA expressions of NF-κB p65, NLRP3, Caspase-1 and ASC in the LPRD group exhibited significant elevations (P < 0.05). Compared with the LYKYF group, the LYKYF+CHPG group had significant elevations in serum TNF-α, IL-6 and IL-1β levels, proportion of M1 macrophages in pharyngeal tissues, p-NF-κB p65/p65 ratio, protein expressions of NLRP3, Caspase-1, ASC, CD86 and iNOS, as well as NF-κB p65, NLRP3, Caspase-1 and ASC mRNA expressions (P < 0.05). The identified key target genes were significantly enriched in GO terms associated with signal transduction, protein phosphorylation regulation, and adaptive responses to external stimuli.

LYKYF may suppress M1 macrophage polarization through suppressing the NF-κB/NLRP3 pathway activation, thereby alleviating reflux pharyngitis in rats.