Human monocytes subjected to ischaemia/reperfusion inhibit angiogenesis and wound healing in vitro

Abstract

Objectives The sequence of initial tissue ischaemia and consecutive blood flow restoration leads to ischaemia/reperfusion (I/R) injury, which is typically characterized by a specific inflammatory response. Migrating monocytes seem to mediate the immune response in ischaemic tissues and influence detrimental as well as regenerative effects during I/R injury. Materials and Methods To clarify the role of classical monocytes in I/R injury, isolated human monocytes were subjected to I/R in vitro (3 hours ischaemia followed by 24 hours of reperfusion). Cellular resilience, monocyte differentiation, cytokine secretion, as well as influence on endothelial tube formation, migration and cell recovery were investigated. Results We show that I/R supported an enhanced resilience of monocytes and induced intracellular phosphorylation of the prosurvival molecules Erk1/2 and Akt. FACS analysis showed no major alteration in monocyte subtype differentiation and surface marker expression under I/R. Further, our experiments revealed that I/R changes the cytokine secretion pattern, release of angiogenesis associated proteins and MMP‐9 activity in supernatants of monocytes exposed to I/R. Supernatants from monocytes subjected to I/R attenuated endothelial tube formation as indicator for angiogenesis as well as endothelial cell migration and recovery. Conclusion In summary, monocytes showed no significant change in cellular integrity and monocyte subtype after I/R. Functionally, monocytes might have a rather detrimental influence during the initial phase of I/R, suppressing endothelial cell migration and neoangiogenesis.

Effects of remote ischemic preconditioning (RIPC) and chronic remote ischemic preconditioning (cRIPC) on levels of plasma cytokines, cell surface characteristics of monocytes and in-vitro angiogenesis: a pilot study

“…In contrary to RIPC/cRIPC plasma, cell culture supernatants from RIPC/cRIPC monocytes did not significantly influence any of the parameters of angiogenesis investigated. In one of our previous studies cell culture supernatants from human monocytes that were subjected to 3 h of in-vitro hypoxia even negatively affected tube formation in-vitro as a surrogate parameter for angiogenesis [22]. These findings suggest that prolonged hypoxia/ischemia may have detrimental effects on human monocytes and may attenuate potentially positive effects of monocytes on angiogenesis.…”

“…Human umbilical vein endothelial cells (HUVEC) were isolated from umbilical cords as described previously [3,22] [approval by the local ethics committee of the Christian-Albrechts University Kiel, Germany (D519/18 and D518/13)] and cultured in endothelial cell growth medium ECGM (PromoCell, Heidelberg, Germany) supplemented with 4 μL/mL of endothelial cell growth supplement, 0.1 ng/ mL epidermal growth factor, 1 ng/mL basic fibroblast growth factor, 90 μg/mL heparin, 1 μg/mL hydrocortisone (all from PromoCell) and 10% fetal bovine serum (Thermo Fisher, Dreieich, Germany). The cells were maintained in a humidified atmosphere (5% carbon dioxide / 95% air) at 37 °C.…”

“…In the present work, tube formation assays utilizing human umbilical vein endothelial cells (HUVEC) were performed to estimate the pro-angiogenic capacity of RIPC/cRIPC plasma and cell culture supernatants derived from RIPC/ cRIPC treated human monocytes. Although in-vitro tube formation assays do not fully resemble all aspects of in-vivo angiogenesis, they have been used by several groups and represent a reproducible and stable system for the in-vitro analysis of early processes of angiogenesis [3,22,23,42]. Employing in-vitro tube formation assays in combination with computer-assisted analysis [4], several parameters of angiogenesis were significantly increased by RIPC/cRIPC plasma.…”

Ablation of GSDMD Improves Outcome of Ischemic Stroke Through Blocking Canonical and Non-canonical Inflammasomes Dependent Pyroptosis in Microglia

“…Neuroinflammation is considered as the main cause of secondary brain injury during I/R. Immune cells recognize certain danger signals which boost production and the release of cytokines like TNF-α, IL-1β, IL-6, MCP-1, and MIP-1α, leading to activation of inflammatory pathways and neural damage (36,37). As a pro-inflammatory process, pyroptosis causes not only rapid loss of membrane integrity but also the release of mature IL-1β and IL-18, which induces the strong pro-inflammatory activity of immune cells.…”

Strategies to Overcome the Barrier of Ischemic Microenvironment in Cell Therapy of Cardiovascular Disease

“…The transplantation of MSCs and subsequently treatment with anti-CCR2 showed promising results in the treatment of acute MI [ 77 ]. This could be explained by the initial detrimental effect of migrating monocytes/macrophages, and subsequently cytokine release in the early phase of ischemia related tissue damage, which was potentially abolished in this study [ 77 , 78 ]. Accordingly, accurate control of oxidants is required to stimulate vascularization, while targeting and modulation of i/nBZ might be further promising ways of improving the effect and survival of transplanted cells in ischemic disease.…”