HMGB1 siRNA can reduce damage to retinal cells induced by high glucose in vitro and in vivo

Abstract

BackgroundDiabetic retinopathy (DR), one of the most common complications of late-phase diabetes, is associated with many risk factors, among which continuous low-grade inflammation is one of the principal ones. As such, lowering inflammation levels and maintain the viability of human retinal endothelial cells (HRECs) are critical for DR therapy. HMGB1 is a well-known proinflammatory cytokine. However, whether HMGB1 small interfering RNA (siRNA) can protect retina cells under a high-glucose environment from morphological changes and functional abnormalities remain undetermined. We aimed to investigate the effect of HMGB1 siRNA on retinal cells in DR.Materials and methodsA total of 80 adult Wistar rats were randomly divided into four groups (n=20 each): normal control, diabetes mellitus (DM), scrambled (Scr) siRNA, and HMGB1 siRNA. Rats in the DM, Scr siRNA, and siRNA groups were established by intraperitoneal injection of streptozotocin. At 16 weeks after injection, rats in the siRNA and Scr-siRNA groups were intravitreally injected with 2 μL HMGB1 siRNA and 2 μL Scr-siRNA, while rats in the control and DM groups were intravitreally injected with the same dose of sterile saline. At 1 week after injections, we performed the following experiments. Immunohistochemical staining and real-time quantitative polymerase chain reaction were performed to test HMGB1 protein and messenger RNA expression in retinas. We performed TUNEL assays to detect retinal cell apoptosis and electroretinography to detect retinal function. In HRECs treated with high glucose, proliferation, morphology, apoptosis, super-oxide dismutase (SOD), and reactive oxygen species production were detected. Western blot was applied to determine the expressions of HMGB1 and its related protein and apoptosis protein.ResultsIntravitreal injection of HMGB1 siRNA reduced protein and messenger RNA expression of HMGB1 (both P<0.05). Intravitreal injection of HMGB1 siRNA reduced apoptosis of retinal cells (P<0.05), protected morphological changes in the retina, and improved the function of the retina (P<0.05). In HRECs treated with high glucose, HMGB1 siRNA pretreatment increased cell viability, reduced cell apoptosis, and reduced oxidative damage to cells (all P<0.05). Western blot detection found that HMGB1 siRNA pretreatment can inhibit the expression of cleaved caspase 3 and improve the expression of BCL2 (P<0.05). HMGB1 and NFκB expression increased in a time-dependent manner in the high-glucose environment and IKKβ and NFκB protein expression decreased significantly after HMGB1 silencing.ConclusionAs a therapeutic target, HMGB1 siRNA can reduce retinal cell damage induced by high glucose in vitro and in vivo and delay DR progress through the HMGB1–IKKβ–NFκB signaling pathway.

The Complex Relationship between Diabetic Retinopathy and High-Mobility Group Box: A Review of Molecular Pathways and Therapeutic Strategies

“…HMGB1 suppression, mediated by intravitreal injections of siRNA, is capable, in diabetic rats, of reducing retinal apoptosis rates as well as improving retinal function. In HRECs exposed to high glucose concentrations, siRNA HMGB1 improved cell viability and reduced the oxidative damage lowering ROS production [105]. The same study group demonstrated the protective role of HMGB1 inhibition in murine DR models.…”

Glycyrrhizin Protects the Diabetic Retina against Permeability, Neuronal, and Vascular Damage through Anti-Inflammatory Mechanisms

“…Inhibition of HMGB1 could be developed as a therapy for diabetic disease. HMGB1 siRNA improved electroretinogram (ERG) measurements, retinal morphology, and reduced retinal cell apoptosis when given intravitreally to diabetic rats or REC grown in high glucose (Jiang and Chen 2017 [33]). Studies in ARPE-19 cells demonstrated that HMGB1 upregulated angiogenic and fibrogenic factors in response to hypoxia (Chang et al 2017 [34]).…”

High Mobility Group Box-1 and Diabetes Mellitus Complications: State of the Art and Future Perspectives

“…They also showed that HMGB1 induced cleaved caspase-3, IL-1β and PARP-1, and that this effect is inhibited by administration of GA. Interestingly, in a work by Sohn and coworkers, the administration in diabetic rats of extract of Polygonum cuspidatum (PCE), a dried root with anti-inflammatory action, reduced HMGB1, RAGE and NF-kB expression and it ameliorated vascular retinal permeability, inhibiting tight junction leakage [133]. To support evidences of HMGB1 activity on progression of DR, Jiang and coworkers demonstrated that intravitreal injection of HMGB1 siRNA in rats reduced retinal damage and cellular death and it improved retinal function [135]. Furthermore, in human retinal endothelial cells treated with high glucose, HMGB1siRNA reduced oxidative stress and cellular apoptosis.…”