The TβR II-targeted aptamer S58 prevents fibrosis after glaucoma filtration surgery

Abstract

Glaucoma filtration surgery (GFS) is an effective clinical treatment for glaucoma when intraocular pressure (IOP) control is poor. However, the occurrence of conjunctival scarring at the surgical site is the main reason for failure of the surgery. In a previous study, we isolated and developed S58, a novel nucleic acid aptamer targeting TβR II, by systematic evolution of ligands by exponential enrichment (SELEX). Here, we show how S58 sterically inhibits the TβR II interaction with TGF-β. The effects of topical S58 treatment were studied in a rabbit model of GFS. At 6 postoperative weeks, S58 reduced fibrosis and prolonged bleb survival in rabbits after GFS. Further in vitro tests showed that the levels of fibrosis in S58 treated-Human Conjunctival Fibroblasts (HConFs) were decreased and that antioxidant defense was increased. In addition, the loss of nuclear factor erythroid 2related factor 2 (Nrf2) or the inhibition of phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) reversed the anti-fibrotic effects of S58. The present work suggests that S58 could effectively improve GFS surgical outcomes by activating the intracellular antioxidant defense PI3K/Akt/Nrf2 signaling pathway.

Eye Drops of Metformin Prevents Fibrosis After Glaucoma Filtration Surgery in Rats via Activating AMPK/Nrf2 Signaling Pathway

“…The sections were dewaxed, rehydrated, and treated with 3% hydrogen peroxide at room temperature for 10 min. Then, the sections were treated with 0.4% pepsin at 37°C for 30 min to recover the antigen, followed by blocking with 1% goat serum albumin at room temperature for 40 min ( Li et al., 2020 ). After incubating with primary antibodies(α-SMA, vimentin and type 1 collagen, 1:200) at 4°C overnight, the sections were incubated with an appropriate biotinylated secondary antibody at 37°C for 1 h, and the image was captured under an optical microscope (Olympus Inc., Tokyo, Japan).…”

“…At room temperature, the blots and HRP-conjugated secondary antibody (CST, 1:1,000) were washed three times (15 min/times) with TBST at the beginning and end of the 1-hour incubation on a shaker. An enhanced chemiluminescence detection system (Bio-Rad Laboratories) and Image J software (Bethesda, Maryland, USA) were used, the intensity of bands normalized to GADPH statistics was observed and analyzed ( Li et al., 2020 ).…”

“…qRT-PCR was performed by SYBR Green qPCR master mix (Takara, Japan). The total reaction volume was 20 μl = 10 μl (SYBR solution, 2.5× Real Master Mix) + 0.5 μl (Forward primer,10 μmol/L) + 0.5 μl (Reverse primer, 10 μmol/L) + 1 μl cDNA + 8 μl (ultra-pure water) ( Li et al., 2020 ). Reaction conditions: Total mRNA was pre-denatured at 95 °C for 10 min, denatured at 95 °C for 15 s, annealed at 62 °C for 30 s, extended at 68 °C for 30 s, and 50 cycles.…”

“…According to the previous experimental method (Li et al, 2020), to assess the oxidative stress level of HConFs, cells were planted into 6-well plates (4 × 10 4 cells −1 ). After different treatment, the wells were decanted and washed two times.…”

“…Slides were washed (two times) and fixed in 4% paraformaldehyde solution at 4°C overnight. It was blocked with 1% goat serum (1 h, 37°C) after infiltration with 0.1% Triton-X100 (Li et al, 2020). Primary antibody fibronectin (1:200) was incubated at 4°C overnight.…”