Ex Vivo Porcine Corneas Express Lower Angiogenesis And Inflammatory Markers After Oxidative Stress Induction

Oxidative stress plays a crucial role in several ocular injuries or diseases. Hence, the effect of oxidative stress on the cornea was investigated using ex vivo porcine tissue.

Laboratory investigation, Experimental Eye Research Institute at University Eye Hospital, Ruhr-University Bochum, Bochum, Germany.

Ex vivo porcine corneas were placed in 5 groups (n=13/group). Four groups were incubated H₂O₂ to induce oxidative stress: (1) 100 mM for 30 min, (2) 200 mM for 30 min, (3) 100 mM for 3 h, (4) 200 mM for 3 h. Untreated corneas were used as a control group (5). All five study groups were cultivated for a total of 24 h. Then, samples were processed for RT-qPCR (n=7/group) or (immuno)-histology (n=6/group). RNA expression of inflammatory (IL1B, IL6, IL8), angiogenesis (VEGF, VEGFB), tight junction (TJP1), and aldehyde dehydrogenase markers (ALDH3A1) was investigated. Also, histological sections were stained for H&E and ZO-1. Afterwards staining area was evaluated.

IL1B expression was significantly decreased in both 3 h H₂O₂ groups (both p<0.050). No changes were noted regarding IL6 and IL10 expression. 30 min incubation with 100 mM (p=0.031) or 200 mM H₂O₂ (p=0.005) lead to a decreased IL8 mRNA expression. This was also seen with 100 mM H₂O₂ for 3 h (p=0.003). VEGF expression decreased trough H₂O₂, significant effects were seen for 30 min 200 mM H₂O₂ (p=0.014) and 3 h 100 mM H₂O₂ (p=0.010). VEGFB and TJP1 expression was comparable in all groups. Regarding ALDH3A1, a lower expression was noted with 3 h 100 mM or 200 mM H₂O₂ (both p<0.050). With increasing dose and duration of H₂O₂, more tissue damage was noted in H&E stained sections. ZO-1 area was significantly decreased in the H₂O₂ groups (p<0.050).

Acute oxidative stress decreased the expression of inflammatory, angiogenesis, tight junction, and aldehyde dehydrogenase markers in ex vivo corneas. A damage of corneal tissue was also noted via (immuno-)histology. Hence, this ex vivo model is suitable to mimic oxidative stress. In addition, it can be used to gain further insights into disease pathomechanisms occurring in corneal injuries, like chemical damage, as well as in corneal diseases, including Fuchs' endothelial dystrophy.