"Stem cell and drug delivery systems to promote angiogenesis and myogenesis in ischemic limbs" 

Peripheral artery disease (PAD) is a widespread atherosclerotic disorder affecting more than 200 million people globally, with critical limb ischemia (CLI) being its most severe manifestation. Stem cell therapy has been a promising approach for CLI treatment, but harsh conditions in the ischemic limb, such as hypoxia, high reactive oxygen species (ROS) pressure, lack of support matrix, trophic factors, and nutrients, compromise the therapeutic efficacy by reducing the survival rate or function of transplanted cells. This thesis presents various biomaterial systems designed for stem cell and drug delivery to improve the cell microenvironment and promote angiogenesis and myogenesis in ischemic limbs.

In four chapters, therapeutic systems consisting of polymer-derived biomaterial, drugs, and/or stem cells were developed and tested both in vitro and in vivo. Firstly, a therapeutic system has been developed to treat critical limb ischemia (CLI), which consists of a biodegradable and thermosensitive hydrogel (pNHA), oxygen release microspheres (ORMs), and mesenchymal stromal cells (MSCs). The ORMs was conjugated by catalase to produce and release oxygen from loaded H₂O₂ for up to 14 days without increasing reactive oxygen species (ROS) pressure. This system can increase oxygen content to promote MSC paracrine effect, which enhances cell survival. In vivo studies have demonstrated that this system promotes cell survival, proliferation, and paracrine effect, regulates inflammation, and accelerates angiogenesis and myogenesis.

Secondly, a N-cadherin mimic peptide HAV was conjugated to the thermosensitive and biodegradable hydrogel poly(NIPAAm-co-MAPEG-co-AOLA-co-NAS) (pNMAN) (Gel). This HAV-functionalized hydrogel (Gel-HAV) can increase the expression of N-cadherin in engrafted induced pluripotent stem cell-derived mesenchymal stem cells (iMSCs), resulting in promoted cell-matrix interaction, cell survival and paracrine effect under hypoxic condition. The delivery of Gel-HAV/MSC also lead to a significant improvement in ischemic limb tissue regeneration.

Thirdly, the cell membrane repair-related protein TRIM72 was modified with an ischemia-targeted peptide IMTP and an MMP2 substrate sequence to create the engineered TRIM72 protein (ETRIM72). Platelet membrane coated ETRIM72 nanoparticles (ETRIM72 NP) and oxygen release nanoparticles (O₂ NP) were then delivered to diabetic mice with CLI. This system can increase oxygen tension and improve cell membrane repair to reduce cell death and improve cell proliferation, resulting in a significantly accelerated tissue regeneration without the use of stem cells.

Finally, an ischemia-targeted PLGA nanoparticle was utilized to encapsulate a GDH1 inhibitor, R162, and an antioxidant peptide, SS-31, to increase glutamine concentration, decrease ROS content and regulate M1 macrophage secretory function in cells. This system shows great potential in clinical use. The co-treatment of R162 NP (RNP) and SS-31 NP (SNP) in diabetic CLI mice promoted ischemic tissue angiogenesis or myogenesis.

"Stem cell and drug delivery systems to promote angiogenesis and myogenesis in ischemic limbs" 

Peripheral artery disease (PAD) is a widespread atherosclerotic disorder affecting more than 200 million people globally, with critical limb ischemia (CLI) being its most severe manifestation. Stem cell therapy has been a promising approach for CLI treatment, but harsh conditions in the ischemic limb, such as hypoxia, high reactive oxygen species (ROS) pressure, lack of support matrix, trophic factors, and nutrients, compromise the therapeutic efficacy by reducing the survival rate or function of transplanted cells. This thesis presents various biomaterial systems designed for stem cell and drug delivery to improve the cell microenvironment and promote angiogenesis and myogenesis in ischemic limbs.

In four chapters, therapeutic systems consisting of polymer-derived biomaterial, drugs, and/or stem cells were developed and tested both in vitro and in vivo. Firstly, a therapeutic system has been developed to treat critical limb ischemia (CLI), which consists of a biodegradable and thermosensitive hydrogel (pNHA), oxygen release microspheres (ORMs), and mesenchymal stromal cells (MSCs). The ORMs was conjugated by catalase to produce and release oxygen from loaded H₂O₂ for up to 14 days without increasing reactive oxygen species (ROS) pressure. This system can increase oxygen content to promote MSC paracrine effect, which enhances cell survival. In vivo studies have demonstrated that this system promotes cell survival, proliferation, and paracrine effect, regulates inflammation, and accelerates angiogenesis and myogenesis.

Secondly, a N-cadherin mimic peptide HAV was conjugated to the thermosensitive and biodegradable hydrogel poly(NIPAAm-co-MAPEG-co-AOLA-co-NAS) (pNMAN) (Gel). This HAV-functionalized hydrogel (Gel-HAV) can increase the expression of N-cadherin in engrafted induced pluripotent stem cell-derived mesenchymal stem cells (iMSCs), resulting in promoted cell-matrix interaction, cell survival and paracrine effect under hypoxic condition. The delivery of Gel-HAV/MSC also lead to a significant improvement in ischemic limb tissue regeneration.

Thirdly, the cell membrane repair-related protein TRIM72 was modified with an ischemia-targeted peptide IMTP and an MMP2 substrate sequence to create the engineered TRIM72 protein (ETRIM72). Platelet membrane coated ETRIM72 nanoparticles (ETRIM72 NP) and oxygen release nanoparticles (O₂ NP) were then delivered to diabetic mice with CLI. This system can increase oxygen tension and improve cell membrane repair to reduce cell death and improve cell proliferation, resulting in a significantly accelerated tissue regeneration without the use of stem cells.

Finally, an ischemia-targeted PLGA nanoparticle was utilized to encapsulate a GDH1 inhibitor, R162, and an antioxidant peptide, SS-31, to increase glutamine concentration, decrease ROS content and regulate M1 macrophage secretory function in cells. This system shows great potential in clinical use. The co-treatment of R162 NP (RNP) and SS-31 NP (SNP) in diabetic CLI mice promoted ischemic tissue angiogenesis or myogenesis.