Role of Inhibiting Inflammation of LC3-Associated Phagocytosis in Dry Eye Disease

AbstractPurpose To confirm the expression and investigate the role of LC3-associated phagocytosis (LAP) in dry eye disease (DED).Methods The DED model of mice was established by scopolamine subcutaneous injection in a low-humidity environment chamber. Tear secretion test and corneal fluorescein sodium staining were used to evaluate the severity of DED. Expression levels of Rubicon, microtubule-associated protein light chain 3-II (LC3-II), Beclin-1 and autophagy-related gene-7 (Atg-7) in corneas of mice with DED were tested by western blot. Cell Counting Kit-8 (CCK-8) assay was used to detect the effects of different concentrations of hypertonic solutions on the proliferation activity of human corneal epithelial cells (HCECs). The expression levels of Dectin-1, IL-6 and IL-1β in HCECs after stimulation with different concentrations of hypertonic solutions were tested. The expressions of Rubicon, LC3-II, Beclin-1 and ATG-7 in HCECs were detected by reverse transcription polymerase chain reaction (RT-PCR). After being pretreated with 10 μM si-Rubicon, the severity of the disease was documented by corneal fluorescein sodium staining. And the expression levels of IL-6 and IL-1β were also tested by RT-PCR.Results Compared with the normal control group, the corneal fluorescein sodium staining scores and tear secretion were significantly reduced. Rubicon, LC3-II, Beclin-1 and ATG-7 were significantly elevated. CCK-8 showed that the 400 and 450 mOsM hypertonic solutions did not affect the proliferation activity of HCECs. The expression of Dectin-1, IL-1β and IL-6 were elevated after stimulation with 450 mOsM solution. LC3-II, Rubicon, ATG-7 and Beclin-1 increased after stimulation with 450 mOsM hyperosmolar solution in HCECs. Corneal fluorescein staining showed that si-Rubicon increased the severity of DED in mice. Moreover, the mRNA expressions of inflammatory factors IL-1β and IL-6 in the cornea of mice were significantly increased.Conclusion DED increased the expression of proteins associated with LAP. LAP could play an anti-inflammatory effect in DED.Keywords: LC3-associated phagocytosisdry eye diseaseinflammationhypertoniccornea Disclosure statementNo conflict of interest exists in the submission of this manuscript, and the manuscript is approved by all authors for publication. The work described was original research that has not been published previously, and is not under consideration for publication elsewhere, in whole or in part. All the authors listed have approved the manuscript that is enclosed. We have no conflicts of interest to disclose.Data availability statementData will be made available on request.Additional informationFundingThis work was supported by Shandong Province Key Research and Development Plan [grant number 2019GSF107022], Taishan Scholar Project of Shandong Province [grant number tsqn201812151].