Objective

2

To construct traditional Chinese medicine （TCM） disease and syndrome combination model of laryngopharyngeal reflux disease （LPRD） in rats with spleen deficiency and evaluate the model.

Methods

2

Rats were randomly divided into the control group and the model group， with 8 rats in each group. The model group was subjected to sleep deprivation and a high-fat diet， while the control group received normal sleep and a high-fat diet. The general condition of the rats was observed， and hematoxylin and eosin （HE） staining was used to observe the pathological changes of the laryngeal mucosa. Electron microscopy was employed to observe the ultrastructure of the laryngeal mucosa.

Results

2

At the 8th week of modeling， pH measurements of the oropharynx in the model group showed varying degrees of laryngopharyngeal reflux， indicating the successful establishment of the LPRD model. During the modeling process， one rat in the model group died due to repeated drowning leading to a pulmonary infection. Compared with the control group， the model group showed significant "weight loss"， with body weight

lower than that of the control group （

P

<

0.05）. Fur was sparse， hair color was withered， and tail defecation rate was high. The grasping strength was low （

P

<

0.05）， and the rats exhibited slow responses with a reduced reaction to environmental stimuli. The R-value of tongue color analysis was lower in the model group （

P

<

0.05）， while the G and B values were higher， but the difference was not statistically significant （

P

>

0.05）. The laryngeal mucosa of model group showed ciliary breakage and loss in the epithelial layer， with irregular columnar epithelial morphology and areas showing squamous epithelium and fibrous scaffolds. The glandular lumens in the submucosa were dilated， with secretions accumulating inside， and inflammatory cell infiltration was observed throughout the mucosa， with tumor-like changes in some areas. The mucosa was significantly thickened， and the laryngeal cartilage was not visible in the field of view. Under electron microscopy， the laryngeal mucosal epithelium of rats in the model group showed mitochondrial swelling， most of which exhibited vacuolar changes， with mitochondrial membrane defects， broken cristae， and ribosome reduction. The mucosal tight junctions were shorter than those of the control group， with larger gaps and looser structure.

Conclusion

2

Long-term sleep deprivation and a high-fat diet can lead to spleen deficiency-type LPRD in rats， with pathological changes in the subglottic region and "spleen deficiency" mitochondrial changes in the laryngeal tissues. It is speculated that reflux events influence the regulation of key processes such as mitochondrial dynamics and mitochondrial autophagy in laryngeal cells， resulting in impaired laryngeal cell function and cell barrier functions， including intercellular space， gap junctions， bridge corpuscles， and adhesive bands. These changes manifest as shorter mucosal tight junctions， larger junction gaps， and a looser structure， which contribute to the susceptibility to LPRD. This method is simple to operate and successfully establishes a TCM disease-syndrome combination model of LPRD.