The activation of ROS-NLRP3-IL-1β signaling axis induced by hyperosmotic stress (HS) has been recognized as a key priming stage of epithelial inflammation in dry eye pathogenesis. The current study aims to investigate whether calcitriol, the active metabolite of vitamin D3, could protect cells against HS-induced inflammation through modulating this critical step. Human corneal epithelial cells (iHCECs) were cultured in hyperosmotic medium (450 mOsM) with various concentrations of calcitriol. Small interfering RNA (siRNA) was used to knock down the expression of vitamin D receptor (VDR) in iHCECs. NLRP3 activation and IL-1β generation were detected by RT-qPCR or ELISA, respectively. Oxidative stress markers including ROS and 8-OHdG were examined by fluorometric analysis. The nuclear translocation of NRF2 was assessed by western blotting. Calcitriol could protect cells against HS-induced injury through inhibiting ROS-NLRP3-IL-1β signaling axis. Calcitriol remarkably suppressed the expression of NLRP3 inflammasome related genes and the production of IL-1β in cells that were exposed to HS. It could also significantly attenuate HS-induced oxidative stress, shown as the reduced intracellular ROS generation and 8-OHdG staining cells after calcitriol treatment. Calcitriol induced the translocation of NRF2 to the nucleus, and thereby triggered the expression of several antioxidant enzymes. The current study indicated that calcitriol could inhibit the priming stage of HS-induced cellular inflammation, highlighting its potential capacity to prevent and mitigate dry eye related corneal inflammation at an earlier stage.

Purpose: To investigate the applicability of a newly developed corneal topographer in assessing tear film stability. Methods: This is a prospective, case–control study. Forty-four Chinese dry eye patients and 41 normal subjects were recruited. Noninvasive tear film break-up time (NI-BUT) was measured using a new method based on a corneal topographer equipped with modified scan software. The reliability of the measurements was determined. Then, the correlations between the NI-BUT and the traditional fluorescein tear film break-up time, Schirmer I test values, and inferior tear meniscus height measurements were determined. The receiver operating characteristic curve technique was used to evaluate the NI-BUT examination in the diagnosis of dry eye. Results: In total, a significant difference between the NI-BUT and the fluorescein tear film break-up time was found (3.2 ± 2.3 seconds vs. 5.2 ± 3.4 seconds; P < 0.001). The coefficient of variation and intraclass correlation coefficient values of NI-BUT were 12.8% and 0.93, respectively, for NI-BUT for intraobserver repeatability and 15.4% and 0.88, respectively for interobserver repeatability. The NI-BUT showed a good correlation with other dry eye examinations (all P < 0.05). In addition, the NI-BUT was significantly shorter in dry eye patients (2.0 ± 0.2 seconds) than in normal subjects (4.3 ± 0.3 seconds; P < 0.001). When the cutoff value was set at <2.65 seconds, the sensitivity and specificity of the test were 84.1% and 75.6%, respectively. Conclusions: Measurements of NI-BUT obtained with the newly developed corneal topographer may provide a simple, noninvasive screening test for dry eyes with acceptable sensitivity, specificity, and repeatability.

Purpose.: To measure corneal and limbal epithelial thickness (ET) in normal subjects and to evaluate its variation with age by using anterior segment optical coherence tomography (AS-OCT). Methods.: A total of 180 normal subjects (180 healthy eyes) were enrolled and divided into four groups according to age: A (0–20 years), B (21–40 years), C (41–60 years), and D (>60 years). Cornea and limbus were imaged with OCT. Corneal ET (CET) was obtained automatically by the built-in analysis software of the OCT system. Limbal ET (LET) in four quadrants was manually measured from OCT images. Results.: Corneal ET of a central 2-mm diameter zone in groups A, B, C, and D were 53.4 ± 2.8 μm, 53.4 ± 2.7 μm, 53.2 ± 3.0 μm, and 52.9 ± 3.3 μm, respectively, N showing no significant change with aging. In the paracentral zone extending to 6-mm diameter, correlation analysis suggested that CET was inversely associated with age (P < 0.05). Limbal ET in the nasal and the temporal quadrants were similar and decreased with aging, the averages were 58.3 ± 8.1 μm, 54.1 ± 6.1 μm, 51.2 ± 6.1 μm, 51.6 ± 5.2 μm for groups A, B, C and D, respectively; while age seemed to have no effect on LET of the superior and the inferior quadrant. Conclusions.: The paracentral corneal epithelium, as well as the nasal and temporal limbal epithelium, became thinner with aging, while the central CET seemed to remain constant. Measurement with AS-OCT of the corneal and limbal ET could aid in clinical assessment and planning treatments of the cornea.

Allergic conjunctivitis is a common problem that significantly impairs patients’ quality of life. Whether air pollution serves as a risk factor for the development of allergic conjunctivitis remains elusive. In this paper, we assess the relationship between air pollutants and weather conditions with outpatient visits for allergic conjunctivitis. By using a time-series analysis based on the largest dataset ever assembled to date, we found that the number of outpatient visits for allergic conjunctivitis was significantly correlated with the levels of NO₂, O₃ and temperature, while its association with humidity was statistically marginal. No associations between PM₁₀, PM_2.5, SO₂, or wind velocity and outpatient visits were seen. Subgroup analyses showed that sex seemed to modify the effects of humidity on outpatient visits for allergic conjunctivitis, but not for NO₂, O₃, or temperature. People younger than 40 were found to be susceptible to changes of all four parameters, while those older than 40 were only consistently affected by NO₂ levels. Our findings revealed that higher levels of ambient NO₂, O₃ and temperature increase the chances of outpatient visits for allergic conjunctivitis. Ambient air pollution and weather changes may contribute to the worsening of allergic conjunctivitis.

Purpose: Inflammasome activation in response to elevated tear osmolarity behaves as an initial signal in dry eye-related corneal inflammation. Pyroptosis is another prominent consequence of inflammasome activation, which is featured by gasdermin D (GSDMD)-driven cell lysis. This study aims to explore the role of pyroptosis in dry eye, and also to verify if calcitriol, a potential therapeutic agent for dry eye, has certain effects against hyperosmotic stress (HS)-induced pyroptosis in human corneal epithelial cells (iHCECs) and the underlying mechanism. Methods: The expression of pyroptosis executor GSDMD in tears from dry eye patients was examined using western blotting. iHCECs were grown in hyperosmotic medium (450 mOsM) to mimic the feature of elevated tear osmolality of dry eye in vitro. Exogenous calcitriol or pyroptosis inhibitor disulfiram was used. The extent of pyroptosis of iHCECs under various treatments was examined by scanning electron microscopy, caspase-1 and propidium iodide (PI) double staining by flow cytometry, immunofluorescent staining for ASC speck formation, and western blotting. Cell viability was measured by a CCK-8 assay and an LDH release assay. Results: We found that pyroptosis was presented in dry eye patients, shown as the elevation of its effector GSDMD N-terminal domain (N-GSDMD) in patients’ tears. Further in vitro results showed that HS promoted pyroptosis in human corneal epithelial cells, while exogeneous supplementation of disulfiram could reduce the number of iHCECs with pyroptotic markers. More importantly, we demonstrated that, in line with the effect of disulfiram, calcitriol could also alleviate HS-induced pyroptosis, through inhibiting the NLRP3–ASC–caspase-1–GSDMD pyroptosis pathway. Conclusion: The current study provided direct evidence showing increased pyroptosis in dry eye patients. We demonstrated that calcitriol was able to effectively alleviate HS-induced corneal epithelial cell damage through inhibiting the NLRP3–ASC–caspase-1–GSDMD pyroptosis pathway. This study underlined calcitriol as a promising therapeutic agent for dry eye given its multiple therapeutic targets.