In Vitro Comparison of Lymphangiogenic Potential of Hypoxia Preconditioned Serum (HPS) and Platelet-Rich Plasma (PRP)

Abstract

Strategies for therapeutic lymphangiogenesis are gradually directed toward the use of growth factor preparations. In particular, blood-derived growth factor products, including Hypoxia Preconditioned Serum (HPS) and Platelet-rich Plasma (PRP), are both clinically employed for accelerating tissue repair and have received considerable attention in the field of regenerative medicine research. In this study, a comparative analysis of HPS and PRP was conducted to explore their lymphangiogenic potential. We found higher pro-lymphangiogenic growth factor concentrations of VEGF-C, PDGF-BB, and bFGF in HPS in comparison to normal serum (NS) and PRP. The proliferation and migration of lymphatic endothelial cells (LECs) were promoted considerably with both HPS and PRP, but the strongest effect was achieved with HPS-40% dilution. Tube formation of LECs showed the highest number of tubes, branching points, greater tube length, and cell-covered area with HPS-10%. Finally, the effects were double-validated using an ex vivo lymphatic ring assay, in which the highest number of sprouts and the greatest sprout length were achieved with HPS-10%. Our findings demonstrate the superior lymphangiogenic potential of a new generation blood-derived secretome obtained by hypoxic preconditioning of peripheral blood cells—a method that offers a novel alternative to PRP.

Comparison of the Effect of Different Conditioning Media on the Angiogenic Potential of Hypoxia Preconditioned Blood-Derived Secretomes: Towards Engineering Next-Generation Autologous Growth Factor Cocktails

“…HPP and HPS organically differ with respect to their protein factor composition, since they correlate with distinct wound healing phases, the former having a direct correlation with the hypoxia-induced, angiogenesis-driven proliferative phase, while the latter also incorporates the platelet-derived hemostatic phase [ 6 , 10 , 12 ]. Despite their differences, the clinical utility of these secretomes harnesses their angiogenic activity, since they can both provide a useful tool for stimulating microvessel sprouting and lymphatic vessel formation on demand [ 5 , 6 , 7 , 8 , 12 ]. As such, they could play an important role in a modern therapeutic strategy that aims to improve local tissue perfusion and accelerate tissue healing where this is a delay or stagnation [ 9 , 10 , 12 , 13 , 14 ].…”

“…The main thesis of this work is based on the notion that the net bioactivity of a growth factor-based regenerative therapy is effected through the balance of stimulatory and inhibitory signals. More specifically, it is known that a range of angiogenic inhibitors are of platelet origin and are thus released into coagulated blood-derived secretomes, e.g., HPS, as a result of platelet activation and degranulation [ 6 , 7 , 8 , 12 , 15 ]. In order to examine these effects, plasma and serum were substituted with different conditioning media, namely, hypoxia preconditioned normal saline (HPP/HPS-NaCl), phosphate buffered saline (HPP/HPS- PBS), Glucose 5% (HPP/HPS-G5%), and AIM V medium (HPP/HPS-AIM).…”

“…In our previous work, we showed that angiogenesis and lymphangiogenesis have symbiotic roles in wound healing, since they both appear to rely on overlapping growth factor mechanisms [ 5 , 6 , 7 , 8 ]. There is now strong evidence indicating that hypoxia preconditioned blood-derived secretomes could constitute a new generation of autologous, bioactive compositions that can supply the necessary biochemical signals for stimulating angiogenesis/lymphangiogenesis, thus driving wound healing to completion [ 5 , 6 , 8 , 9 , 10 , 11 , 12 , 13 , 14 ]. These growth factor compositions can be obtained through the method of hypoxia-adjusted in vitro preconditioning of peripheral blood cells (PBCs), first proposed by Hadjipanayi and Schilling [ 5 , 10 , 11 , 12 ].…”

“…Conditioning PBCs under the very same conditions encountered within a wound microenvironment, i.e., physiological temperature and hypoxia, offers a means for optimizing the angiogenic potential of Hypoxia Preconditioned Plasma (HPP) and Hypoxia Preconditioned Serum (HPS), which can be differentially prepared by adjusting blood coagulation prior to hypoxic conditioning [ 5 , 6 , 9 , 10 , 11 , 12 , 15 ]. More specifically, we have shown that the angiogenic potential of blood-derived secretomes is defined by the complex stoichiometry of their component pro- and anti-angiogenic factor proteins, rather than the concentration of one or more individual growth factors [ 6 , 8 , 12 , 15 ]. The angiogenic potency of hypoxia preconditioned secretomes is further highlighted by the fact that they maintain their pro-angiogenic activity in vitro, even when they are prepared from peripheral blood that has been obtained from patients who receive oral anticoagulation due to underlying vascular pathology or who suffer from diabetes mellitus [ 16 ].…”

Lymphangiogenesis in gastric cancer: function and mechanism

“…Inhibition of PDGF-BB can significantly reduce the ability of LEC proliferation, migration and tube formation [ 55 ]. In addition, the concentrations of VEGF-C, PDGF-BB and bFGF in hypoxic preconditioning serum (HPS) and platelet-rich plasma (PRP) were higher than those in normal serum (NS), and could significantly promote the proliferation and migration of LECs and improve the ability of lymphangiogenesis [ 56 ]. Angiopoietin-2 (Ang-2): Angiopoietin-2 (Ang-2) is the ligand of receptor tyrosine kinase Tie2, involved in lymphangiogenesis [ 57 ].…”