| Objective: The present study was conducted to investigate the potential of curcumin on neovascularization in diabetic ischemia as well as its effects on regulating the proliferative,migratory,angiogenic functions and senescence of endothelial progenitor cells(EPCs).In addition,this study also investigated the capacity of curcumin on human umbilical vein endothelial cells(HUVECs)under hyperglycemic condition.Methods: Streptozocin(STZ)was applied to induce type 1 diabetic mice model.Both diabetic and non-diabetic mice both received unilateral hindlimb ischemic surgery.Curcumin was then administrated to the mice by lavage for 14 days consecutively.Laser Doppler perfusion imaging(LDPI)was conducted to demonstrate the blood flow reperfusion on day 0,day 7 and day 14.Capillary density was measured in ischemic gastrocnemius muscle.The in vitro assays include(I)EPCs were isolated from bone marrow and characterized by DiI-Ac-LDL uptake assay and flow cytometry.In addition,angiogenesis,migration,proliferation abilities and senescence were determined in EPCs isolated from diabetic and non-diabetic mice by tube formation assay,transwell assay,colony-formation assay and SA-?-gal assay,respectively.Quantitative PCR was then used to determine the mRNA expression of VEGF-? and Ang-I in EPCs.(II)HUVECs under hyperglycemic condition were treated with curcumin in a dose-escalating strategy.Tube formation and SA-?-gal assay were conducted.Results: The blood glucose levels of mice were about 20mmol/L 14-day post STZ injection,which confirmed the successful induction of type 1 diabetes.Hindlimb ischemia mice model was also constructed using diabetic and non-diabetic mice.The blood perfusion measured by laser Doppler perfusion imaging(LDPI)at day 7 and 14 after ischemic operation demonstrated that blood perfusion in curcumin-treated group was significantly enhanced compared with untreated groups,and recovery was similar to non-diabetic group.Immunohistochemistry revealed the capillary density in curcumin-treated group was elevated than control groups,indicating the proangiogenic effect of curcumin.The characterization of EPCs defined by DiI-Ac-LDL uptake assay and flow cytometry was in accordance with the previous reports.SA-?-gal assay indicated that the senescence ratio of curcumin-treated diabetic EPCs was obviously lower than control group(P<0.05,olive oil group: 43.32±1.83% vs curcumin treatment: 20.33±5.90%),indicating than curcumin could reverse the senescence of diabetic EPCs.The tube formation assay,transwell assay and colony formation assay also revealed that the angiogenesis,migration and proliferation abilities of EPCs were improved by curcumin application.Quantitative PCR confirmed the overexpression of VEGF-? and Ang-I in EPCs after curcumin treatment.Moreover,curcumin-treated HUVECs under high glucose condition showed promoted differentiation function and reversed senescence.The optimum concentration is 5?mol/L.Conclusion: The results of this study revealed that lavage application of curcumin could ameliorate the decreased blood flow reperfusion in diabetic mice.In addition,EPCs isolated from curcumin-treated diabetic mice showed enhanced ability in migration,angiogenesis and proliferation,and decreased senescence.Furthermore,VEGF-? and Ang-I expression was significantly upregulated,which may be induced by curcumin treatment.Curcumin also presented the regulatory effects on the differentiation ability and senescence of HUVECs. |
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