Human Macrophage ATP7A is Localized in the trans-Golgi Apparatus, Controls Intracellular Copper Levels, and Mediates Macrophage Responses to Dermal Wounds

Abstract

The copper transporter ATP7A has attracted significant attention since the discovery of its gene mutation leading to human Menkes disease. We previously reported that ATP7A is highly expressed in the human vasculature and identified a novel vascular function of ATP7A in modulation of the expression and activity of extracellular superoxide dismutase. We recently identified that ATP7A expression in THP-1 cells (a monocyte/macrophage model cell line) plays a role in the oxidation of low density lipoproteins, indicating that it is necessary to further investigate its expression and function in monocytes/macrophages. In the current study, we demonstrated the protein and mRNA expression of ATP7A in human peripheral blood mononuclear cell (PBMC)-derived macrophages and alveolar macrophages. ATP7A was strongly co-localized with the trans-Golgi apparatus in PBMC-derived macrophages. Intracellular copper, detected by synchrotron X-ray fluorescence microscopy, was found to be distributed to the nucleus and cytoplasm in human THP-1 cells. To confirm the role of endogenous ATP7A in macrophage copper homeostasis, we performed inductively coupled plasma mass spectrometry in murine peritoneal macrophages, which showed markedly increased intracellular copper levels in macrophages isolated from ATP7A-deficient mice versus control mice. Moreover, the role of ATP7A in regulating macrophage responses to dermal wounds was studied by introduction of control and ATP7A-downregulated THP-1 cells into dermal wounds of nude mice. Infiltration of THP-1 cells into the wounded area (detected by expression of human macrophage markers MAC2 and CD68) was reduced in response to downregulation of ATP7A, hinting decreased macrophage accumulation subsequent to dermal wounds. In summary, alongside our previous studies, these findings indicate that human macrophage ATP7A is localized in the trans-Golgi apparatus, regulates intracellular copper levels, and mediates macrophage responses to a dermal wound.

Behind the Link between Copper and Angiogenesis: Established Mechanisms and an Overview on the Role of Vascular Copper Transport Systems

“…ATP7A is highly expressed in the vasculature (endothelial cells and VSMCs), macrophages and fibroblasts [118,134,135,136]. The actual relevance of ATP7A to angiogenesis is underscored by the etiological role of mutant transporters, responsible for the human Menkes syndrome, in the onset of congenital cardiovascular abnormalities [137].…”

“…Kim et al [134] showed the role of endogenous ATP7A in regulating the copper content in macrophages. When the pump was down-regulated, cells showed a reduced migratory potential towards dermal wounds experimentally inflicted on nude mice [134]. In particular, they observed a slight, but not significant, increase in copper content within the injured area at day 11 after wounding.…”

“…It has been shown that TM-induced copper starvation in a co-culture of malignant breast cancer cells and macrophages does not interfere with the local release of angiogenic cytokines, but compromises downstream angiogenic-dependent cell signalling [253]. An in vitro study on RAW264.7 macrophage cell cultures exposed to hypoxia, a condition of a real tumour context, showed relevant changes in the expression and activity of cell copper transport proteins, reflecting a higher delivery of the metal to ceruloplasmin [134], and an obvious elevation of blood-circulating copper in a bioavailable form. …”

Bone Marrow from Blotchy Mice Is Dispensable to Regulate Blood Copper and Aortic Pathologies but Required for Inflammatory Mediator Production in LDLR-Deficient Mice during Chronic Angiotensin II Infusion

“…In a previous study using THP-1 cells as a MΦ model cell, 65 ATP7A downregulation in THP-1 cells is associated with reduced cell accumulation in dermal wounds. 5 In addition, ATP7A downregulation inhibits the migration of vascular smooth muscle cells in response to platelet-derived growth factor and wound scratch. 20 Thus, it is highly likely that ATP7A downregulation inhibits the migration of hematopoietic cells via decreasing the expression of MCPs.…”

“…ATP7A, also known as Menkes disease protein or copper-transporting P-type ATPase, regulates the copper transport at 2 different levels. At a cellular level, this transporter regulates copper egress, as indicated by an increase of copper levels in cultured fibroblasts, 1 lymphoblasts, 2 muscle cells, 3 and macrophages (MΦs) 4,5 isolated from Menkes disease patients and mouse models. At the organ level, ATP7A primarily controls dietary copper absorption in the intestine, as indicated by the increased copper levels in intestine and decreased copper levels in most other tissues.…”