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Therapeutic effect of total saponins of honeysuckle on secretory otitis media in rats after noise exposure and effect on cochlear TLR4-NLRP3 signaling pathway
- HUANG Yu
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2023, 39(22):
3060-3066.
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Objective To investigate the therapeutic effect of total saponins of honeysuckle on secretory otitis media in rats after noise exposure and the effect on toll-like receptors-4(TLR-4)-NLR family pyrin domain containing 3(NLRP3) signaling pathway. Methods Totally 50 Wistar rats were divided into normal control group,model control group,low-,medium-and high-dose groups of total saponins of honeysuckle. Except for the normal control group,the rats in the other groups were exposed to a sound pressure level(SPL) of 100 dB and 10 KHz octave noise in the organic glass box for two hours for 15 consecutive days to establish the secretory otitis media model,each drug group was given corresponding drug intervention,while the normal control group and model control group were given an equal volume of normal saline,and the treatment cycle was two weeks. After the treatment,the threshold level of audiological evaluation(ABR) of the rats was determined,the microstructure of the rat cochlear basement membrane was detected by silver nitrate staining,the pathological changes of the rat cochlea were detected by electron microscopy,and the real-time fluorescent reverse transcription method(RT-PCR) method and Western blotting were used to detect the mRNA protein levels of TLR4 and NLRP3 in cochlear tissue,and enzyme-linked immunosorbent assay(ELISA) was used to detect matrix metalloproteinase 9(MMP-9),myeloperoxidase(MPO),tumor necrosis factor-α(TNF-α) levels. Results In the normal control group,the structure of the inner and outer hair cells of the cochlea was normal. In the model control group,the structure of the inner and outer hair cells of the cochlea was severely damaged(inverted,missing),arranged in disorder,and collapsed in a large area,ciliated epithelium swelled,cilia disordered arrangement and partially shedded,and middle ear mucosa thickened,with inflammatory cell infiltration. After the intervention treatment with total saponins of honeysuckle,the degree of damage to the inner and outer hair cells of the cochlea of rats in each group was significantly improved. The structure of cochlear hair cells in the normal control group was normal,while the arrangement of inner and outer hair cells in the model control group was disordered,and there were gaps in the combination with the periprocess of spiral ganglion cells. After the intervention of total saponins of honeysuckle,the arrangement of inner and outer hair cells in the cochlea tended to be normal,there was still a gap at the junction with the periprocess of the spiral ganglion cells,but the gap was smaller. The threshold level of ABR,mRNA protein levels of cochlear TLR4[(0.73±0.12)vs(4.29±0.74),(0.24±0.06)vs(1.06±0.17)/GAPDH] and NLRP3[(0.84±0.14)vs(3.88±0.63),(0.30±0.05) vs(1.12±0.18)/GAPDH)],and the levels of MPO,TNF-α,and MMP in model control group were higher than those in the normal control group(P<0.05). The threshold level of ABR,mRNA protein levels of cochlear TLR4[(4.29±0.74)vs(3.18±0.50),(2.34±0.34),(1.52±0.24),(1.06±0.17)vs(0.90±0.15),(0.59±0.10),(0.40±0.06)/GAPDH)] and NLRP3[(3.88±0.63)vs(3.02±0.48),(2.18±0.37),(1.46±0.23),(1.12±0.18)vs(0.83±0.13),(0.54±0.09),(0.36±0.06)/GAPDH],and the levels of MPO,TNF-α,and MMP in low-,medium-and high-dose groups of total saponins of honeysuckle were lower than those in model control group(P<0.05). The threshold level of ABR,mRNA protein levels of cochlear TLR4[(3.18±0.50)vs(2.34±0.34),(1.52±0.24),(0.90±0.15)vs(0.59±0.10),(0.40±0.06)/GAPDH],and NLRP3[(3.02±0.48)vs(2.18±0.37),(1.46±0.23),(0.83±0.13)vs(0.54±0.09),(0.36±0.06)/GAPDH], and the levels of MPO,TNF-α,and MMP in medium- and high-dose groups of total saponins of honeysuckle were lower than those in low-dose group(P<0.01). The threshold level of ABR,mRNA protein levels of cochlear TLR4[(2.34±0.34)vs(1.52±0.24),(0.59±0.10)vs(0.40±0.06)/GAPDH] and NLRP3[(2.18±0.37)vs(1.46±0.23),(0.54±0.09)vs(0.36±0.06)/GAPDH] and the levels of MPO,TNF-α,and MMP in high-dose group of total saponins of honeysuckle were lower than those in medium-dose group(P<0.01),the dose-effect relationship was obvious(P<0.01). Conclusion The total saponins of honeysuckle have a significant therapeutic effect on secretory otitis media in rats after noise exposure,and the mechanism may be related to the inhibition of activation of inflammatory TLR-4-NLRP3 pathway by total saponins of honeysuckle.