Angiogenesis is Enhanced by Sustained Delivery of rhGM-CSF from Porous Collagen-Chitosan Scaffolds

Huafeng Sun, Xingang Wang, Weijie Yu, Xinlei Hu, Chuangang You, Hang Hu, Chunmao Han* Department of Burn, Second Affiliated Hospital of Zhejiang University, Hangzhou, 310009, China *Corresponding Author Email: [email protected] Introduction Local delivery of GM-CSF (granulocyte-macrophage colony-stimulating factor) has been has been used for treating human chronic skin ulcers and deep seconddegree burns wounds [1, 2], with good therapeutic effects. As a pleiotropic cytokine, GM-CSF regulates the proliferation, differentiation and survival of hematopoetic cells [3], by mobilizing and recruiting hematopoietic and endothelial stem cells from the bone marrow [4]. GMCSF secreted by keratinocytes, mediates epidermal cell proliferation in an autocrine manner [5]. Moreover, many other cells involved in wound healing, such as macrophages, lymphocytes, fibroblasts, endothelial cells, and dendritic cells also synthesize GM-CSF and/or are GM-CSF targets [6]. Nevertheless, continuous injection or external application is another big issue restricting better utilization in clinical practice. Thus, in the present study, we integrated recombinant human (rh) GM-CSF into a heparinized collagen-chitosan scaffold to control local and sustained release of the cytokine, as previously described [7], considering the slow angiogenesis of artificial threedimensional (3D) scaffolds during repair of full-thickness skin defects. We tested the hypothesis that a heparinized collagen-chitosan scaffold combined with rhGM-CSF accelerates angiogenesis for tissue engineering. Materials and Methods A cytokine-containing porous collagen-chitosan scaffold was fabricated by incorporating rhGM-CSF using freezedrying and further cross-linking treatment with EDC/NHS and heparin. In vitro release kinetics was investigated by ELISA. Scaffolds with/without heparin (Ø6mm) were loaded with rhGM-CSF (10μg/cm) overnight, and then incubated in PBS at 37°C . Supernatants were replaced