The Beneficial Effect of Melatonin in Brain Endothelial Cells against Oxygen-Glucose Deprivation Followed by Reperfusion-Induced Injury
Figure 3
The measurement of the tight junction protein in bEND.3 cells after OGD/R-induced injury. (a) The level of Claudin 5, a tight junction protein, was evaluated by immunocytochemistry. This image shows that expression of Claudin 5 in the experimental control (EC) group decreased compared to the normal control (NC) group. Melatonin increased the expression of Claudin 5 under OGD/R injury (green). In the Mel (10 nM) and Mel (100 nM) groups, the expression of Claudin 5 was higher than in the EC group. Claudin 5 was preserved in the melatonin treatment group, following OGD/R-induced injury. Scale bar: 200 µm, Claudin 5: red, and 4′,6-diamidino-2-phenylindole (DAPI): blue. (b) Western blotting showed that the relative protein level of Claudin 5 was reduced in EC compared to the NC group. The relative level of Claudin 5 was increased in Mel (10 nM) and Mel (100 nM) groups, compared to the EC group. The bar graph shows the quantification of Claudin 5 protein in all groups. β-Actin was used as an internal control. Data are expressed as mean ± S.E.M. (*). (i) Normal control (NC): bEnd.3 cells cultured with normal media without OGD injury, (ii) experimental control (EC): bEnd.3 cells cultured in nontreated medium for 18 h after 6 h of OGD injury, and (iii) 10 nM melatonin (Mel 10 nM): bEnd.3 cells treated with 10 nM melatonin for 24 h before 6 h of OGD injury. These cells were then cultured in nontreated medium for 18 hr. (iv) 100 nM melatonin (Mel 100 nM): bEnd.3 cells were also treated with 100 nM melatonin (100 nM melatonin group) for 24 h before 6 h of OGD injury. These cells were then cultured in nontreated medium for 18 h.
