Investigation of apoptotic and inflammatory markers in hypoxia induced human cornea epithelial cells co‐cultured with limbal mesenchymal stem cells

Aims/Purpose: Here we aim to investigate the effect of hypoxic conditions (5% O2) on human cornea epithelial cells (HCECs) and also by creating a system in which limbal mesenchymal stem cells (LMSCs) are co‐cultured with HCECs, the effect of LMSC is examined on this subject via paracrine mechanism. Methods: HCECs are either mono‐cultured or co‐cultured with LMSC under hypoxic and normoxic (%21 O2) conditions. HCECs of after 6, 24, and 48 h of incubation were harvested for comprehension of changes in RNA and protein expression levels. Bcl2, Bax, Erk1/2 protein levels are checked through western blot analysis, whereas expression levels of cytokines, which are IL‐6, IL‐8, IL‐10 and TGF‐β1, are investigated via RT‐qPCR. Results: Both anti‐inflammatory and pro‐inflammatory cytokine gene expression levels in co‐cultured model increased upon hypoxia and normoxia conditions compared to cytokines in mono cultured HCECs. In addition to the increase in cytokine levels in the co‐culture condition, this increase is less in the hypoxia exposed co‐culture in comparison to the normoxic co‐culture model. Furthermore, anti‐ and pro‐apoptotic protein expression levels have significantly changed upon co‐cultured both in normoxia and hypoxia conditions. Conclusions: In conclusion, it is observed that hypoxia with co‐cultured model decreased inflammation and acted as crucial role in cell death mechanism. These results were further confirmed with apoptotic protein ratio in hypoxic co‐cultured cells. The underlying mechanism will be examined in future experiments.