Enhanced angiogenesis by mesenchymal stem cells based on Hyaluronic Acid hydrogel combined with GM-CSF and IL-2 in a rat model of hindlimb ischemia

Background The clinical application of stem cells in restoring ischemic lower limb perfusion has been hindered by challenges. To promote the directed differentiation of mesenchymal stem cells(MSCs) into endothelial cells(ECs) and enhance the paracrine effect, this paper aim to find a combined therapy for angiogenesis. Methods MSCs based on Hyaluronic Acid(HA) hydrogel combined with GM-CSF and IL-2(HGI-MSCs) were locally injected into the femoral artery ligated ischemia model rat. Recovery of perfusion was assessed by limb temperature and exhaustive distance. Hematoxylin and eosin(H&E) staining, immunohistochemistry and immunofluorescence staining were used to evaluate the quantity of blood vessels, microvessel density(MVD) and the proliferation and maturation of neovascularization in the limb muscle. Migration and tube formation of Human umbilical vein endothelial cells(HUVECs) were evaluated by Transwell and tube formation assays. The expression levels of angiogenesis-related genes, cytokine and proteins were measured by qRT-PCR, ELISA and Western blotting, respectively. Results HGI-MSCs promoted ischemic limb angiogenesis and restored perfusion. Our study showed that HGI-MSCs could promote the proliferation and maturation of neovascularization. Moreover, HGI-MSCs promoted HUVECs migration and tube formation in vitro, up-regulated the expression of VEGF and activated PI3K/Akt signaling pathway. Conclusions HGI-MSCs showed a more robust pro-angiogenic effect than that of pure MSCs in the ischemia limb model rat. It offers novel ideas for the treatment of patients with refractory limb ischemia.