Skin Wound Repair Is Not Altered in the Absence of Endogenous AnxA1 or AnxA5, but Pharmacological Concentrations of AnxA4 and AnxA5 Inhibit Wound Hemostasis

Abstract

Skin injury induces the cell surface exposure of phosphatidylserine (PS) on damaged and dying cells to activate coagulation and repair processes. Annexins can bind to PS and may modulate the healing response. Here, we determine the relevance of annexins for skin wound healing using AnxA1- and AnxA5-deficient mice and recombinant annexins with distinct PS binding properties. Wound inflammation, closure and the formation of granulation tissue were not altered in AnxA1- or AnxA5-deficient mice or after increasing AnxA5 serum concentrations (100 n<smlcap>M</smlcap>) in wild-type mice. Increased serum concentrations (1 µ<smlcap>M</smlcap>) of AnxA5 induced massive bleeding, but wound hemostasis was not delayed by AnxA1. Both annexins interact with PS, but only AnxA5 can form 2-dimensional (2D) arrays on the cell surface. The injection of an AnxA5 mutant that binds to PS but lacks the ability of 2D array formation failed to induce bleeding. 2D lattice-forming AnxA4, with high affinity to PS also caused bleeding, while hemostasis was not affected by AnxA8 with low affinity or the AnxA8 mutant with medium affinity for PS and the lack of 2D formation. Increased concentrations of AnxA4 and AnxA5 also delayed coagulation pathway activation in vitro. This effect was attenuated for the AnxA5 mutant as well as for AnxA1 and AnxA8. In conclusion, endogenous AnxA1 and AnxA5 are dispensable for wound hemostasis and repair, but pharmacologically excessive concentrations of AnxA4 and AnxA5 inhibit hemostasis in skin wounds.

Gene Expression Profiling of the Extracellular Matrix Signature in Macrophages of Different Activation Status: Relevance for Skin Wound Healing

“…At early time points of the healing process, the wound is mainly repopulated by hematopoietic cells [32]. At day three and five post wounding the majority of the cells are macrophages whereas fibroblasts, vascular cells, neutrophils or platelets are hardly found within the wound bed [14,32,33]. We confirmed by immunofluorescence studies that at day three post wounding F4/80 + macrophages are found within the wound bed (Figure 5a).…”

“…To demonstrate that ECM components could be important for macrophage-modulated wound healing, immunofluorescence and immunoblot analysis of mouse skin wounds at the peak of macrophage infiltration were performed. At early time points of the healing process, the wound is mainly repopulated by hematopoietic cells [32]. At day three and five post wounding the majority of the cells are macrophages whereas fibroblasts, vascular cells, neutrophils or platelets are hardly found within the wound bed [14,32,33].…”

Annexin A12–26 Treatment Improves Skin Heterologous Transplantation by Modulating Inflammation and Angiogenesis Processes

“…It is noteworthy to mention that angiogenesis induced by VEGF-A treatment was similar in wild type (WT) and AnxA1 null mice ( Supplementary Figure 2 ), showing that endogenous AnxA1 is not relevant to the VEGF-A induced angiogenesis in the skin. As previously mentioned, it was recently showed the normal skin repair in AnxA1 null mice ( Kreft et al, 2016 ).…”

“…However, the role of AnxA1 on skin repair is not well established. Secreted AnxA1 is not essential to skin wound healing, as wound inflammation, closure and the formation of granulation tissue were not altered in AnxA1 null mice ( Kreft et al, 2016 ). Conversely, we recently showed, for the first time, that the systemic pharmacological treatment using AnxA1 2–26 in mice increased skin allograft survival by inducing the resolution of inflammation, as it caused impaired migration of neutrophils into tissue and enhanced apoptosis of these cells in the site of allograft transplantation ( Teixeira et al, 2016 ).…”

Annexin A4 inhibits sulfatide‐induced activation of coagulation factor XII

“…One well-known physiological role of several ANXs is the regulation of blood coagulability during coagulation (including thrombosis) and fibrinolysis. [24][25][26] ANXA5 is the most studied ANX in terms of its anticoagulant activity and inhibits coagulation by binding to negatively charged phospholipid membranes, such as those containing phosphatidylserine, and preventing vitamin K-dependent proteins from forming protease complexes on the membrane. [27][28][29] Moreover, an ANXA5 knock-out mouse study showed an increase in placental thrombosis and subsequent fetal loss.…”