Angiogenic Properties of Concentrated Growth Factors (CGFs): The Role of Soluble Factors and Cellular Components

Abstract

Blood-derived concentrated growth factors (CGFs) represent a novel autologous biomaterial with promising applications in regenerative medicine. Angiogenesis is a key factor in tissue regeneration, but the role played by CGFs in vessel formation is not clear. The purpose of this study was to characterize the angiogenic properties of CGFs by evaluating the effects of its soluble factors and cellular components on the neovascularization in an in vitro model of angiogenesis. CGF clots were cultured for 14 days in cell culture medium; after that, CGF-conditioned medium (CGF-CM) was collected, and soluble factors and cellular components were separated and characterized. CGF-soluble factors, such as growth factors (VEGF and TGF-β1) and matrix metalloproteinases (MMP-2 and -9), were assessed by ELISA. Angiogenic properties of CGF-soluble factors were analyzed by stimulating human cultured endothelial cells with increasing concentrations (1%, 5%, 10%, or 20%) of CGF-CM, and their effect on cell migration and tubule-like formation was assessed by wound healing and Matrigel assay, respectively. The expression of endothelial angiogenic mediators was determined using qRT-PCR and ELISA assays. CGF-derived cells were characterized by immunostaining, qRT-PCR and Matrigel assay. We found that CGF-CM, consisting of essential pro-angiogenic factors, such as VEGF, TGF-β1, MMP-9, and MMP-2, promoted endothelial cell migration; tubule structure formation; and endothelial expression of multiple angiogenic mediators, including growth factors, chemokines, and metalloproteinases. Moreover, we discovered that CGF-derived cells exhibited features such as endothelial progenitor cells, since they expressed the CD34 stem cell marker and endothelial markers and participated in the neo-angiogenic process. In conclusion, our results suggest that CGFs are able to promote endothelial angiogenesis through their soluble and cellular components and that CGFs can be used as a biomaterial for therapeutic vasculogenesis in the field of tissue regeneration.

Analysis of CGF Biomolecules, Structure and Cell Population: Characterization of the Stemness Features of CGF Cells and Osteogenic Potential

“…In particular, the amount of VEGF in the CGF-conditioned medium after a 14-day incubation was even greater than the amount of VEGF extracted from the CGF. These results agree with our previous study showing that CGF-derived cells expressed and released angiogenic factors, including VEGF [22].…”

“…Recently, we demonstrated the ability of CGF to promote the osteogenic differentiation of stem cells [11]. Furthermore, we showed that CGF releases endothelial progenitor cells, which contribute to neo-angiogenesis and to the formation of endothelial tubular structures [22].…”

“…In recent years CGF was widely studied as an autologous blood derivative able to promote tissue repair, vascularization, cell migration, and differentiation [11,[19][20][21][22]. Tissue repair is a complex mechanism that takes place over four phases: inflammatory process, cell proliferation, differentiation, and ECM remodeling.…”

“…Indeed, the presence of different cell populations is known: hematopoietic stem cells, lymphocytes, monocytes, and fibroblast-like cells [1]. Our recent findings showed that when CGF, without manipulation, is released into the culture medium, cells are able to adhere to the plate and proliferate [22]. Here we show that the release of cells from CGF seemed to be rather slow, and most of the cells were found in the plate only after cutting CGF on the 14th day.…”

Pro-angiogenic approach for skeletal muscle regeneration

“…Also, matrix metalloproteinases, especially matrix metalloproteinases 1 and 2, mediate degradation of the capillary wall and the extracellular matrix components urokinase and integrins favouring ECs migration towards the extracellular matrix [37]. Angiopoietin-1 (Ang1) promotes the reorganization of the ECs in stable tubules with a central lumen, and platelet-derived growth factor (PDGF) action results in formation of a vascular network with the newly formed tubules [38,39].…”