Abstract：Objective: To investigate the protective effect of human umbilical cord mesenchymal stem cell (hUCMSC)-derived exosomes carrying thrombospondin-1 (TSP-1) in a rat model of retinal vein occlusion (RVO). Methods: The hUCMSCs were randomly divided into hUCMSC group, NC-hUCMSC group and TSP-1-hUCMSC group, the NC-hUCMSC group and TSP-1-hUCMSC group were infected with the lentivirus containing TSP-1 gene sequence and the negative control lentivirus, respectively, and the cell transfection effect was determined by fluorescent inverted microscope, real-time fluorescent quantitative PCR and Western blot. The hUCMSC-derived exosomes carrying the TSP-1 gene were isolated, the morphology was observed by transmission electron microscopy, the diameter was determined by the nanoparticle tracking analysis method, and the expression of the exosomal marker proteins CD9, CD63 and Alix was detected by Western blot. The RVO model of SD rats was established by laser photodynamic method and divided into model group, hUCMSC-Exo group, TSP-1-hUCMSC-Exo group, each with 10 rats, and another 10 healthy ratswerewolf made the control group. The hUCMSC-Exo group was subconjunctivally injected with 100 μl of hUCMSC-derived exosomes, and the TSP-1-hUCMSC-Exo group was injected with 100 μL of hUCMSC-derived exosomes containing TSP-1 lentivirus infection. 1 day, 7 days, 14 days, 21 days after the establishment of the RVO model, the fundus images were observed. 21 d later, the vascular lesions were examined by fluorescein fundus angiography (FFA), and the histopathological changes of the retina were observed by HE staining, Real-time fluorescent quantitative PCR and Western blot were used to detect the mRNA and protein levels of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and vascular cell adhesion molecule-1 (VCAM-1) in retinal tissues. Results: The inverted fluorescence microscope showed that there was obvious green fluorescent protein expression in the cells transfected by the lentivirus, and the relative expression of TSP-1 mRNA and protein in hUCMSC in the TSP-1-hUCMSC group were significantly increased (P<0.05), indicates that the cell transfection was successful. The separated particles are spherical with a diameter of approximately 100 nm. CD9, CD63 and Alix proteins are all highly expressed, indicating that exosomes have been successfully isolated. Compared with the control group, different degrees of white occluded blood vessels and plaques appeared in the eyes of the model group on the 1st, 7th, 14th and 21th day of the model, and the model was judged to be successful; in addition, the retinal blood vessels of the model group showed tortuous and dilated veins. Uneven, fluorescence staining and exudation of blood vessel wall, retinal tissue structure was obviously damaged, and the relative expression of HIF-1α, VEGF and VCAM-1 mRNA and protein in the tissue were significantly increased (P<0.05). Compared with the model group, the hUCMSC-Exo group and TSP-1-hUCMSC-Exo group reduced the fluorescence staining of the blood vessel wall, restored the retinal structure, and the relative expression of HIF-1α, VEGF and VCAM-1 mRNA and protein decreased significantly (P<0.05). Compared with the hUCMSC-Exo group, the TSP-1-hUCMSC-Exo group did not see obvious fluorescence leakage in the blood vessels, the retina structure was clearer, and the relative expressions of HIF-1α, VEGF and VCAM-1 mRNA and protein in the tissue were significantly decreased ( P<0.05). Conclusion: HUCMSC-derived exosomes carrying TSP-1 can alleviate retinal damage in RVO rats and protect the retina. The mechanism may be related to the inhibition of the expression of HIF-1α, VEGF and VCAM-1.
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2022, Vol. 28 
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