| Objective: To observe the protective effect of human umbilical cord mesenchymal stem cells on retinal tight junction protein and inflammatory factors in early diabetic retinopathy rats,and to explore the protective effect of human umbilical cord mesenchymal stem cells on BRB injury.Methods: Isolated and culturedhuman umbilical cord mesenchymal stem cells(HUCMSC)by the tissue adherent method and examined them by flow cytometric analysis.Seventy healthy male SD rats were randomly divided into normal control group(n = 15)and diabetic group(n = 55).Diabetic rats were induced by intraperitoneal injection of streptozotocin STZ(60mg / kg).Rats in each group were sacrificedand made retinal stretched preparation with the perfusion of Evans blue at the 1st,3rd and 5th month after the models were successfully established.The DR group was divided into DR group,DR + PBS group and DR + HUCMSC group when the diabetic retina(DR)abnormalities were observed in the retina.DR + HUCMSC group was injected with 1mlfourth generation HUCMSC,DR + PBS group were injected 1ml PBS by tail vein,DR group received no treatment,all rats were executed after observed 1 month.Evans blue retina was used to observe the changes of vascular morphology and permeability.The expression of Occludin protein was observed and detected by immunohistochemistry and Western-blot.ELISA was used to detect the the levels of inflammatory factors TNF-?,ICAM-1 and VEGF who were correlated with vascular permeability.Results: 1.Mesenchymal stem cells(MSC)could be isolated from human umbilical cord.Stem cell markers CD73,CD90 were positive while CD34,CD45 and HLA-DR were negative.2.Evans blue perfusion in retinal blood vessels: Blood vessels in the normal group had a good shape while in DM5 months,the blood vessels walking tortuous,stiff,and uneven lumen thickness,visible fluorescence leakage,microangioma and blood vessel infusion area were formed.One month after the intervention,vascular leakage was reduced.The results of retinal vascular permeability showed an increased level in DR was when comparedwith the normal group(P <0.05).HUCMSC could decrease the vascular permeability,but it could not be restored to normal.The difference was statistically significant(P <0.05).3.Occludin protein was detected by immunohistochemical staining and Western-blot.The retinal microvascular wall of the normal group was brown and yellow,but it was lighter in DR group.The color of retinal microvascular wall in HUCMSC group was deeper than that in DR group.The expression of Occludin protein in DR and DR + PBS group was significantly lower than that in normal group(P <0.05).The expression of Occludin protein in DR+HUCMSC group was significantly higher than that in DR and DR+HUCMSC group,and the difference was statistically significant(P <0.05).4.ELISA results: In the DR and DR + PBS group,the levels of VEGF,ICAM-1 and TNF-? in the retina were up-regulated.The levels of VEGF,ICAM-1 and TNF-? in the retina of DR + HUCMSC group were down-regulated,and the difference was statistically significant compared with DR group(P <0.05).The results also showed that ICAM-1 was positively correlated with VEGF expressionConclusion: 1.The HUCMSC are isolated from the human umbilical cord,and the cell purity is higher,the morphology is uniform,the spindle shape is increased,and the specific antigen of MSC is expressed.2.Diabetic retinopathy model can be induced in 5 months of diabetes rats,which presented as retinal microvascular morphology abnormalities and increased blood retinal barrier permeability.3.Intravenous injection of human umbilical cord mesenchymal stem cells reduces microvascular leakage and reduces the damage of the blood retinal barrier by increasing the level of Occludin.4.Intravenous injection of human umbilical cord mesenchymal stem cells can further protect the blood retinal barrier from damage by reducing the levels of inflammatory factors TNF-?,ICAM-1 and VEGF in the retina. |
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