MicroRNA-based engineered mesenchymal stem cell extracellular vesicles to treat visual deficits after blast-induced trauma.

ABSTRACT

Our previous studies have shown the benefit of intravitreal injection of a mesenchymal stem cell (MSC)- derived secretome to treat visual deficits in a mild traumatic brain injury (mTBI) mouse model. In this study, we have addressed whether MSC-derived extracellular vesicles (EV) overexpressing miR424, which particularly targets neuroinflammation, show similar benefits in the mTBI model. Adult C57BL/6 mice were subjected to a 50-psi air pulse on the left side, overlying the forebrain, resulting in mTBI. Sham-blast mice were controls. Within an hour of blast injury, 3 µl (∼7.5 × 108 particles) of miR424-EVs, native-EVs, or saline was delivered intravitreally. One month later, retinal morphology was observed through optical coherence tomography (OCT); visual function was assessed using optokinetic nystagmus (OKN) and electroretinogram (ERG), followed by immunohistological analysis. A separate study in adult mice tested the dose-response of EVs for safety. Blast injury mice with saline showed decreased visual acuity compared with the sham group (0.30 ± 0.03 vs. 0.39 ± 0.01 c/d, p < 0.02), improved with miR424-EVs (0.39 ± 0.02 c/d, p < 0.01) but not native-EVs (0.33 ± 0.04 c/d, p > 0.05). Contrast sensitivity thresholds of blast mice receiving saline increased compared with the sham group (85.3 ± 5.9 vs. 19.9 ± 4.8, %, p < 0.001), rescued by miR424-EVs (23.6 ± 7.3 %, p < 0.001) and native-EVs (45.6 ± 10.7 %, p < 0.01). Blast injury decreased "b" wave amplitude compared to sham mice (94.6 ± 24.0 vs. 279.2 ± 25.3 µV, p < 0.001), improved with miR424-EVs (173.0 ± 27.2 µV, p < 0.03) and native-EVs (230.2 ± 37.2 µV, p < 0.01) with a similar decrease in a-wave amplitude in blast mice improved with both miR424-EVs and native-EVs. Immunohistology showed increased GFAP and IBA1 in blast mice with saline compared with sham (GFAP 11.9 ± 1.49 vs. 9.1 ± 0.8, mean intensity/100,000 µm2 area, p < 0.03; IBA1 36.08 ± 4.3 vs. 24.0 ± 1.54, mean intensity/100,000 µm2 area, p < 0.01), with no changes with native-EVs (GFAP 12.6 ± 0.79, p > 0.05; IBA1 32.8 ± 2.9, p > 0.05), and miR424-EV (GFAP 13.14 ± 0.76, p > 0.05; IBA1 31.4 ± 2.7, p > 0.05). Both native-EVs and miR424-EVs exhibited vitreous aggregation, as evidenced by particulates in the vitreous by OCT, and increased vascular structures, as evidenced by αSMA and CD31 immunostainings. The number of capillary lumens in the ganglion cell layer increased with increased particles in the eye, with native EVs showing the worst effects. In conclusion, our study highlights the promise of EV-based therapies for treating visual dysfunction caused by mTBI, with miR424-EVs showing particularly strong neuroprotective benefits. Both miR424-EVs and native-EVs provided similar protection, but issues with EV aggregation and astrogliosis or microglial/macrophage activation at the current dosage call for improved delivery methods and dosage adjustments. Future research should investigate the mechanisms behind EVs' effects and optimize miR424 delivery strategies to enhance therapeutic outcomes and reduce complications.