Figure 5.

In Vitro Effect of IL-13 and Novel Single-Stranded RNAi Agent on the Expression of POSTN of HRECs

Structure of canonical double-stranded siRNA and novel single-stranded RNAi agent (A), expression of IL-13 determined by immunohistochemistry in OIR retina (C), and the NK0144-mediated effects on POSTN expression induced by IL-13 in HRECs (B and D). Cultured HRECs transfected or not transfected with NK0000 or NK0144 were stimulated by IL-13. Total RNA was extracted at 24 hr after IL-13 stimulation and was analyzed by real-time RT-PCR (C) and ELISA (D). (A) The novel single-stranded RNAi agent was prepared as a single-stranded RNA oligomers that had the sequences of canonical double-stranded siRNA. (B) The expression of POSTN in HRECs stimulated by IL-13 was significantly increased in a dose-dependent manner (n = 4/group). *p < 0.01, **p < 0.0001. (C) Retinal sections show that IL-13 (green) is increased in CD4-positive cells (red) in OIR retina compared with control retina. (D) The expression of POSTN mRNA in HRECs induced by IL-13 was significantly reduced following transfection with 10 nM NK0144, whereas transfection with 10 nM of single-stranded scramble RNAi agent (NK0000) as a negative control RNAi agent had no significant inhibitory effect on the expression (n = 4/group). *p < 0.01. The protein level of POSTN in the supernatant from HRECs stimulated by IL-13 after the transfection with NK0144 was significantly decreased compared to that in the control group (n = 4/group). *p < 0.01, **p < 0.001. Error bars are SEM.