The Effects Of Co-transplantation Of Adipose-derived Mesenchymal Stem Cells On Blood-Retinal Barrier In Diabetic Rats

Purpose:To investigate the effects of co-transplantation of adipose-derived mesenchymal stem cells (AMSCs) and endothelial-like cells (ELCs) or human umbilical vascular endothelial cells (HUVECs) in rats with diabetic retinopathy (DR). To evaluate the dose-effect of AMSCs in DR rats.Methods:Diabetes was induced in 80 healthy adult Wistar rats with a single intraperitoneal injection of streptozocin (STZ). Close monitor on the blood glucose and body weight was performed during the following weeks. At week 30, retinal digesting preparation was used to determine the vascular morphology change caused by diabetes. Human AMSCs and HUVECs were isolated, cultured and then injected intravenously. ELCs were derived from AMSCs by being cultured in endothelial cell growth medium for 3 days. Experimental rats were divided into untreated (U) group, AMSCs + ELCs (AE) group, AMSCs + HUVECs (AU) group, high dose AMSCs (H) group, and low dose AMSCs (L) group. Blood-retinal barrier (BRB) changes were evaluated by retinal Evans blue (EB) content test. The expression of intercellular cell adhesion molecule-1 (ICAM-1), vascular endothelial growth factor (VEGF), and angiopoietin-2 (Ang-2) were measured via qPCR and Western-blot. The retinal morphological changes were assessed through HE staining. The distribution of transplanted cells was observed by immunohistochemical staining and co-focusing experiment. The safety of transplantation in other organs was assessed via HE staining and immunohistochemical staining.Results:No significant changes of blood glucose or body weight were found among the groups after the intervention. Two weeks after the intervention (W2), compared with the U group, the BRB function was improved in both the AE group and the L group by measuring EB leakage (P<0.05,0.001, respectively). Four weeks after the intervention (W4), the BRB function was the worst in the AE group compared with other three treatment groups. The gene transcription level and the protein translation level were not consistent for ICAM-1 and VEGF, respectively. Taken the U group as the control, the expression of ICAM-1 on protein level increased in the AE group (W2) and decreased in the AE group (W4), the AH group (W4), the H group (W4) and the L group (W2, W4); the expression of VEGF on protein level increased in the AE group (W2, W4) and decreased in the L group (W4); there was no difference of Ang-2 expression on protein level among all the groups at the two testing points. The outer nuclear layer edema and the retinal nerve fiber layer edema were lightened only in the L group (W2). The improvement of the outer nuclear layer edema were achieved in all the treatment groups (W4). The inner nuclear layer edema were also lightened in all the treatment groups except the AE group (W4). A decrease of the edema degree of the inner plexiform layer was found in both the AH group and the H group (W4). The number of transplanted cells homing to the retina was quite small. There were a lot of transplanted cells homing to the spleen and the renal tumor.Conclusions:AMSCs could repair the BRB defect by anti-inflammatory effects. Low dose of AMSCs injection improves the BRB function in DR rats the quickest. The effect of AMSCs transplantation alone is better than that of the co-transplantation of AMSCs and ELCs.