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Surface Tethering of Inflammation-Modulatory Nanostimulators to Stem Cells for Ischemic Muscle Repair.

ABSTRACT: Stem cell transplantation has been a promising treatment for peripheral arterial diseases in the past decade. Stem cells act as living bioreactors of paracrine factors that orchestrate tissue regeneration. Prestimulated adipose-derived stem cells (ADSCs) have been proposed as potential candidates but have been met with challenges in activating their secretory activities for clinical use. Here, we propose that tethering the ADSC surface with nanoparticles releasing tumor necrosis factor α (TNFα), named nanostimulator, would stimulate cellular secretory activity in situ. We examined this hypothesis by complexing octadecylamine-grafted hyaluronic acid onto a liposomal carrier of TNFα. Hyaluronic acid increased the liposomal stability and association to CD44 on ADSC surface. ADSCs tethered with these TNFα carriers exhibited up-regulated secretion of proangiogenic vascular endothelial growth factor and immunomodulatory prosteoglandin E2 (PGE₂) while decreasing secretion of antiangiogenic pigment epithelium-derived factors. Accordingly, ADSCs tethered with nanostimulators promoted vascularization in a 3D microvascular chip and enhanced recovery of perfusion, walking, and muscle mass in a murine ischemic hindlimb compared to untreated ADSCs. We propose that this surface tethering strategy for in situ stimulation of stem cells would replace the costly and cumbersome preconditioning process and expedite clinical use of stem cells for improved treatments of various injuries and diseases.

SUBMITTER: Leong J

PROVIDER: S-EPMC8274413 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Mesenchymal stem cells (MSCs) hold great promise for vascular smooth muscle regeneration. However, most studies have mainly relied on extended supplementation of sophisticated biochemical regimen to drive MSC differentiation towards vascular smooth muscle cells (vSMCs). Herein we demonstrate a concomitant method that exploits the advantages of biomimetic matrix stiffness and tethered transforming growth factor ?1 (TGF-?1) to guide vSMC commitment from human MSCs. Our designed poly(ethylene glycol) hydrogels, presenting a biomimetic stiffness and tethered TGF-?1, provide an instructive environment to potently upregulate smooth muscle marker expression in vitro and in vivo. Importantly, it significantly enhances the functional contractility of vSMCs derived from MSCs within 3 days. Interestingly, compared to non-tethered one, tethered TGF-?1 enhanced the potency of vSMC commitment on hydrogels. We provide compelling evidence that combining stiffness and tethered TGF-?1 on poly(ethylene glycol) hydrogels can be a promising approach to drastically enhance maturation and function of vSMCs from stem cell differentiation in vitro and in vivo. STATEMENT OF SIGNIFICANCE: A fast, reliable and safe regeneration of vascular smooth muscle cells (vSMCs) from stem cell differentiation is promising for vascular tissue engineering and regenerative medicine applications, but remains challenging. Herein, a photo-click hydrogel platform is devised to recapitulate the stiffness of vascular tissue and appropriate presentation of transforming growth factor ?1 (TGF-?1) to guide vSMC commitment from mesenchymal stem cells (MSCs). We demonstrate that such concomitant method drastically enhanced regeneration of mature, functional vSMCs from MSCs in vitro and in vivo within only a 3-days span. This work is not only of fundamental scientific importance, revealing how physiochemical factors and the manner of their presentation direct stem cell differentiation, but also attacks the long-standing difficulty in regenerating highly functional vSMCs within a short period.

Project description:Rational: Ischemic injury of the skeletal muscle remains a serious clinical problem and currently there is no effective therapy. The aim of the present study is to determine whether human umbilical cord mesenchymal stem cells- derived exosomes (UMSC-Exo) could repair ischemic injury by releasing circular RNA. Methods and Results: To create hindlimb ischemia, we surgically ligated the left femoral artery in C57BL/6 mice. Using circRNA-seq analyses of total RNA from ischemic and control muscles, we found reduced expression of circHIPK3 in the ischemic muscle. To explore the role of circHIPK3 in ischemic injury, the mice were randomly assigned into three groups after surgery: 1) vehicle; 2) UMSC-Exo; 3) UMSC-Exo and siRNA targeting circHIPK3 (UMSC-Exo /si-circHIPK3). UMSC-Exo treatment significantly increased expression of circHIPK3 and improved blood perfusion, running distance and muscle force, which were reversed by injection of UMSC-Exo /si-circHIPK3, suggesting that UMSC-Exo improve muscle function by releasing circHIPK3. UMSC-Exo treatment also inhibited ischemia induced pyroptosis - cell death caused by inflammasome as evidenced by activation of NLRP3, cleaved caspase-1, and subsequent increase of IL-1β and IL-18, and the effects were reversed by injection UMSC-Exo /si-circHIPK3. Bioinformatic analysis identified miR-421/FOXO3a as a potential target for circHIPK3, which was confirmed by luciferase reporter assay. Knockdown of circHIPK3 in C2C12 cells resulted in increased expression of miR-421. We established an in vitro model of pyroptosis by stimulating C2C12 cells with LPS and ATP. LPS and ATP treatment resulted in reduced expression of circHIPK3 and increased expression of miR-421, which was prevented by UMSC-Exo. Western blot analysis showed reduced levels of NLRP3 and cleaved caspase-1 when cells were treated by UMSC-Exo. The expression of FOXO3a in C2C12 cells was increased in the presence of miR-421 inhibitor, and the expression was reduced when cells were treated by LPS and ATP. Importantly, the expression of FOXO3a was upregulated by UMSC-Exo but was reduced when si-circHIPK3 was present. Conclusions: Using loss/gain-of function method, we demonstrated that miR-421/FOXO3a is the direct target of circHIPK3, and UMSC-Exo prevent ischemic injury by releasing circHIPK3, which in turn down regulate miR-421, resulting in increased expression of FOXO3a, leading to inhibition of pyroptosis and release of IL-1β and IL-18.

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Surface Tethering of Inflammation-Modulatory Nanostimulators to Stem Cells for Ischemic Muscle Repair.

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