Establishment of rabbit dry eye cells model and evaluation of its biological characteristics / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To further explore effective drugs for dry eye treatment by isolating and culturing lacrimal gland epithelial cells<i> in vitro</i>, establishing a dry eye cell model and analyzing relevant inflammatory factors. <p>METHODS: Rabbit lacrimal gland epithelial cells were <i>in vitro</i> isolated and cultured, and the activity and purity of primary cells were identified by cell proliferation experiment and immunofluorescence experiment. In addition, 0.5 times IC₅₀ of lipopolysaccharide LPS and TNF-α were used respectively to stimulate rabbit lacrimal gland epithelial cells and then establish two dry eye cell models. Finally, through cell proliferation experiment, ELISA and flow cytometry, the biological characteristics of these two dry eye cell models were compared. <p>RESULTS:After 12h of culture, the primary cells of lacrimal gland epithelial cells basically adhered to the wall of culture bottles; and 48h later, the cells stretched and almost each of them presented a shape of a long triangle. The activity of primary cells of lacrimal gland epithelium was 92%, and the positive rate of marker Pan-rkeratin was more than 90%, which accorded with the experimental requirements. The IC₅₀ of LPS and TNF-α are 20μg/mL and 4.996ng/mL respectively. After 12h of intervention with LPS(10μg/mL)and TNF-α(2.5ng/mL), the cell activity of the two groups was significantly lower than that of control group(<i>P</i><0.01); compared between these two groups, the apoptosis rate of TNF-α group is higher than that of LPS group(<i>P</i><0.01). The levers of IL-1β and IL-6 in the cell supernatants of the two groups were significantly higher than those of the control group(<i>P</i><0.01); compared between the two groups, IL-1β and IL-6 in TNF-α group were significantly higher than those in LPS group(<i>P</i><0.01). It was suggested that TNF-α was superior to LPS in simulating inflammatory response of dry eye. <p>CONCLUSION: This study successfully established a relatively simple and rapid rabbit dry eye cell model with high cell purity and stability, which provided a more stable <i>in vitro</i> experimental model for the basic research on the function of rabbit lacrimal gland epithelial cells and dry eye.

Effect of Buddleia flavonoids drug-containing plasma on the expression of STAT1 phosphoprotein in lacrimal gland epithelial cells in vitro / 国际眼科杂志(Guoji Yanke Zazhi)

AIM: To explore the effect of Buddleia flavonoids drug-containing plasma and androgen receptor(AR) blocker on the expression of STAT1 phosphoprotein.METHODS: In vitro lacrimal gland epithelial cells were cultivated with H2O2 to establish the dry eye apoptosis state. Blank plasma group, Buddleia officinalis plasma total flavonoids interfere with drug-containing group, and the intervention group of testosterone propionate were set. The expressions of STAT1 phosphoprotein of each group were observed by Western blot and AR blocker flutamide was used to explore the intended androgen effect of Buddleia flavonoids.RESULTS: After the intervention of drug-containing plasma, the expression of STAT1 Phosphoprotein in Buddleja officinalis drug-containing plasma intervention group(0.353±0.494) and testosterone propionate intervention group(0.502±0.036) were enhanced and the differences between the two groups were significant (P<0.01). After using the AR blocker in all groups, the expression of STAT1 phosphoprotein in each group (0.268±0.061,0.283±0.106,0.213±0.071) had no difference.CONCLUSION: Buddleja officinalis drug-containing plasma total flavonoids can promote the expression of STAT1 phosphorylation.