This study aimed to investigate the effect of mesenchymal stem cell (MSC)-derived small extracellular vesicles (sEVs) on diabetic retinopathy (DR) and its underlying mechanism. In vivo, MSC-sEVs were injected intravitreally into diabetic rats to determine the therapeutic efficacy. In vitro, MSC-sEVs with/without miR-22-3p inhibition were cocultured with advanced glycation end-products (AGEs)-induced microglia with/without NLRP3 overexpression to explore the molecular mechanism. In vivo, MSC-sEVs inhibited NLRP3 inflammasome activation, suppressed microglial activation, decreased inflammatory cytokines levels in the retina, and alleviated DR as evidenced by improved histological morphology and blood-retinal barrier function. Based on miRNA sequencing of MSC-sEVs, bioinformatic software, and dual-luciferase reporter assay, miR-22-3p stood out as the critical molecule for the role of MSC-sEVs in regulating NLRP3 inflammasome activation. Diabetic rats had lower level of miR-22-3p in their retina than those of control and sEV-treated rats. Confocal microscopy revealed that sEV could be internalized by microglia both in vivo and in vitro. In vitro, compared with sEV, the anti-inflammation effect of sEV^{miR-22-3p (-)} on AGEs-induced microglia was compromised as they gave a lower suppression of NLRP3 inflammasome activation and inflammatory cytokines. In addition, NLRP3 overexpression in microglia damped the anti-inflammatory effect of sEV. These results indicated that MSC-sEVs alleviated DR via delivering miR-22-3p to inhibit NLRP3 inflammasome activation. Our findings indicate that MSC-sEVs might be a potential therapeutic method for DR.

The effects of epigallocatechingallate (EGCG) on the migration and expression of MMP-2 of uveal melanoma cells have not been reported. We studied this effect and relevant signaling pathways in a human uveal melanoma cell line (M17). MTT study found that EGCG did not affect the cell viability of M17 cells up to 100 µM. Wound-healing assay showed that EGCG significantly reduced the migration of melanoma cells in a dose-dependent manner from 20 to 100 µM. Gelatin zymography showed that secreted MMP-2 activity was dose-dependently inhibited by EGCG, whereas the MMP-2 expression at protein and mRNA levels was not affected as determined by western blot and RT-PCR analysis. EGCG significantly increased the expressions of MMP-2 endogenous inhibitors (TIMP-2 and RECK) in M17 cells. Western blot analysis of MAPK signal pathways showed that EGCG significantly decreased phosphorylated ERK1/2 levels, but not p38 and JNK levels, in melanoma cells. ERK1/2 inhibitors also reduced the migration and activity of MMP-2 in M17 cells. The present study suggested EGCG at nontoxic levels could inhibit migration of melanoma cells via downregulation of activities of secreted MMP-2 through the inhibition of the ERK1/2 phosphorylation. Therefore, EGCG may be a promising agent to be explored for the prevention of metastasis of uveal melanoma.

Hypoxia is the most important stimulus leading to upregulation of VEGF in the retina and this is caused by accumulation of hypoxia-inducible factors-1α(HIF-1α) protein. The effects of zeaxanthin, a natural phytochemical, on the VEGF and HIF-1αexpression in the primary culture of human retinal pigment epithelial (RPE) cells were studied. An in vitro RPE cell hypoxia model was established by placing cells under 1% oxygen pressure or by adding cobalt chloride (CoCl₂) to the culture medium. RPE cells and conditioned media were collected from cultures treated with and without zeaxanthin under normoxic and hypoxic conditions. VEGF and HIF-1αprotein and RNA levels were measured by ELISA kits and RT-PCR, respectively. Hypoxia caused a significant increase of VEGF expression and accumulation of HIF-1αin RPE cells. Zeaxanthin at 50–150 μM significantly inhibited the expression of VEGF and accumulation of HIF-1αprotein caused by hypoxia but did not affect expression of VEGF and HIF-1αunder normoxic conditions. This is the first report on the effect of zeaxanthin on VEGF and HIF-1αlevels in cultured RPE cells and suggests that zeaxanthin may have potential value in the prevention and treatment of various retinal diseases associated with vascular leakage and neovascularization.

To compare outcomes of femtosecond laser-assisted deep anterior lamellar keratoplasty (FSL-DALK) and penetrating keratoplasty (FSL-PK) for the treatment of keratoconus. Twenty eight eyes underwent FSL-DALK (consisted of 12 eyes in the FSL-DALKa subgroup without baring the Descemet’s membrane and 16 eyes in the FSL-DALKb subgroup baring the Descemet’s membrane using big-bubble technique) were compared with 12 eyes that underwent FSL-PK for keratoconus. These patients underwent an ophthalmic examination preoperatively and 3, 6, 9, and 12 months postoperatively. The postoperative BCVA in the FSL-PK group, and the FSL-DALKb subgroup were significantly better than that in the FSL-DALKa subgroup (P < 0.05), whereas no differences were found between the FSL-DALKb subgroup and the FSL-PK group (P > 0.05). There were no significant differences in the mean spherical equivalent (SE) and astigmatism between the FSL-DALK and the FSL-PK groups, nor between the subgroups of FSL-DALK during the follow-up period (P > 0.05). At the last follow-up, the mean endothelial cell loss in the FSL-DALK group (9.12 %) was significantly less than that in the FSL-PK group (20.79 %) (P < 0.001), while there was no difference between the FSL-DALKa (9.15 %) and the FSL-DALKb (9.10 %) subgroups (P = 0.15). The FSL-DALK group seemed to have fewer graft rejections (1/28 cases) than the FSL-PK group (2/12 cases), although Kaplan-Meier curve showed no significant difference between the two groups (P = 0.144). In this retrospective study, the results suggested that FSL-DALKb gives better visual outcome, and FSL-DALKb is a better option for keratoconus whose endothelium is not compromised. However, larger and prospective studies are further required.

Diabetic retinopathy (DR) ranks among the primary causes of adult blindness globally. Oxidative stress and mitochondrial dysfunction play a critical role in the progression of DR. Mounting data indicated that small extracellular vesicles (sEVs) of mesenchymal stem cell (MSC) have the ability to transport bioactive chemicals to target cells, leading to changes in their phenotype. Nevertheless, it remains elusive how MSC-derived sEVs regulate oxidative stress and mitochondrial function in DR. MSC-sEVs was intravitreally injected to streptozotocin (STZ)-treated Sprague-Dawley rats to assess the therapeutic effects on DR. The underlying regulatory mechanism was investigated by coculturing advanced glycation end-products (AGEs)-induced rat Müller cells with/without PTP1B overexpression with MSC-sEVs in vitro, with or without miR-125a-5p suppression. Intravitreal injection of MSC-sEVs improved histological morphology and blood-retinal barrier function, alleviated Müller gliosis, decreased PTP1B expression, redox stress and apoptosis in retina of diabetic rat. MSC-sEVs decreased the accumulation of ROS and improved the structure and function of mitochondria of Müller cells with AGEs treatment. Mechanically, MSC-sEVs activated the mitophagy of AGEs-treated Müller cells, represented by an increased expression of the LC3II/LC3I ratio, TOM20, PINK1 and Parkin along with a decreased expression of P62. Importantly, miR-125a-5p inhibitor abolished the protective effects of MSC-sEVs. Furthermore, the overexpression of PTP1B in Müller cells reduced the effects of MSC-sEVs. These findings suggested that miR-125a-5p of MSC-sEVs alleviates Müller cells injury in DR by modulating PINK1/Parkin-mediated mitophagy via PTP1B pathway.

Progressive endothelial cell injury of retinal vascular is a vital factor in diabetic retinopathy (DR) pathogenesis. Mesenchymal stem cells-derived small extracellular vesicles (MSC-sEVs) showed beneficial effects on DR. However, the effects of MSC-sEVs on endothelial dysfunction of DR and the mechanism is still unclear. In this study, MSC-sEVs mitigated retinal blood-retina barrier (BRB) impairment in rats with streptozotocin (STZ)-induced DR by reducing ferroptosis in vivo and in vitro. MSC-sEVs miRNA sequencing analysis revealed that miR-125b-5p may mediate human retina microvascular endothelial cells (HRMECs) ferroptosis and P53 as a downstream target based on dual-luciferase reporter assays. Silencing miR-125b-5p in MSC-sEVs reversed the therapeutic effects of MSC-sEVs on rats with DR and advanced glycation end products (AGEs)-treated HRMECs. Additionally, overexpression of miR-125b-5p could diminish ferroptosis in HRMECs, and this effect could be effectively reversed by overexpressing P53. This study indicated the potential therapeutic effect of MSC-sEVs on vascular endothelial function maintenance and that the delivery of sEVs carrying miR-125b-5p could prevent endothelial cell ferroptosis by inhibiting P53, thereby protecting the BRB.