Inducible HGF-secreting Human Umbilical Cord Blood-derived MSCs Produced via TALEN-mediated Genome Editing Promoted Angiogenesis

Abstract

Mesenchymal stem cells (MSCs) promote therapeutic angiogenesis to cure serious vascular disorders. However, their survival period and cytokine-secretory capacity are limited. Although hepatocyte growth factor (HGF) can accelerate the rate of angiogenesis, recombinant HGF is limited because of its very short half-life (<3–5 minutes). Thus, continuous treatment with HGF is required to obtain an effective therapeutic response. To overcome these limitations, we produced genome-edited MSCs that secreted HGF upon drug-specific induction. The inducible HGF expression cassette was integrated into a safe harbor site in an MSC chromosome using the TALEN system, resulting in the production of TetOn-HGF/human umbilical cord blood-derived (hUCB)-MSCs. Functional assessment of the TetOn-HGF/hUCB-MSCs showed that they had enhanced mobility upon the induction of HGF expression. Moreover, long-term exposure by doxycycline (Dox)-treated TetOn-HGF/hUCB-MSCs enhanced the anti-apoptotic responses of genome-edited MSCs subjected to oxidative stress and improved the tube-formation ability. Furthermore, TetOn-HGF/hUCB-MSCs encapsulated by arginine-glycine-aspartic acid (RGD)-alginate microgel induced to express HGF improved in vivo angiogenesis in a mouse hindlimb ischemia model. This study showed that the inducible HGF-expressing hUCB-MSCs are competent to continuously express and secrete HGF in a controlled manner. Thus, the MSCs that express HGF in an inducible manner are a useful therapeutic modality for the treatment of vascular diseases requiring angiogenesis.

Optimization of lentiviral transduction parameters and its application for CRISPR-based secretome modification of human endometrial mesenchymal stem cells

“…In order to overcome these limitations, guided nucleases can be used. In the context of MSCs secretome modification, we were able to find the only article where HGF-secreting MSCs were obtained by TALEN-mediated genome editing [64]. In the present article, we applied another type of guided nucleases, CRISPR/Cas9, that is characterized by target design simplicity and possibility of multiplex gene expression regulation compared to TALENs and ZFNs [65].…”

“…Additionally, Flt-1-engineered MSCs caused regression of endometriotic lesions [50]. A plenty of other fascinating results are obtained by applying MSCs with genetically modified paracrine activity [57,58,63,64]. However, in the vast majority of studies, the gain-of-function genetic perturbations are obtained by the use of cDNA overexpression systems.…”

Therapeutic Potential of Human Mesenchymal Stem Cells for Treating Ischemic Limb Diseases

“…These aggregates were also shown to reduce the fibrotic areas in the ischemic limb tissues of mice because of their bulkiness and effective entrapment at the transplantation sites (6). Combination therapies such as microgels with gene modifications or spheroids with low-level light have also been developed to enhance angiogenesis in the context of animal models of limb ischemia (12, 32). …”

IFN‐γ enhances the wound healing effect of late EPCs (LEPCs) via BST2‐mediated adhesion to endothelial cells

“…Clinical interest has been focused on therapeutic angiogenesis by injecting ex vivo expanded LEPCs into patients. Various clinical studies have shown improved vascular outcomes in patients with ischemic and chronic wounds . However, one of the major challenges for this approach is that a low number of intravenously injected LEPCs reach the site of diseased target area due to their circulatory nature and low adhesion rate .…”

“…Clinical interest has been focused on therapeutic angiogenesis by injecting ex vivo expanded LEPCs into patients. Various clinical studies have shown improved vascular outcomes in patients with ischemic and chronic wounds [2,6]. However, one of the major challenges for this approach is that a low number of intravenously injected LEPCs reach the site of diseased target area due to their circulatory nature and low adhesion rate Abbreviations BST2, bone marrow stromal cell antigen 2; ECD, extracellular domain; ECs, Endothelial cells; HUVECs, human umbilical vein endothelial cells; IFN-c, Interferon gamma; LEPCs, Late endothelial progenitor cells.…”