Mast Cells Localize in Hypoxic Zones of Tumors and Secrete CCL-2 under Hypoxia through Activation of L-Type Calcium Channels

Abstract

Hypoxia is a condition that together with low pH, high amounts of reactive oxygen species (ROS), and increased adenosine levels characterize tumor microenvironment. Mast cells (MCs) are part of tumor microenvironment, but the effect of hypoxia on the production of MC-derived cytokines has not been fully described. Using the hypoxia marker pimonidazole in vivo, we found that MCs were largely located in the low-oxygen areas within B16-F1 mice melanoma tumors. In vitro, hypoxia promoted ROS production, a ROS-dependent increase of intracellular calcium, and the production of MCP 1 (CCL-2) in murine bone marrow–derived MCs. Hypoxia-induced CCL-2 production was sensitive to the antioxidant trolox and to nifedipine, a blocker of L-type voltage-dependent Ca2+ channels (LVDCCs). Simultaneously with CCL-2 production, hypoxia caused the ROS-dependent glutathionylation and membrane translocation of the α1c subunit of Cav1.2 LVDCCs. Relationship between ROS production, calcium rise, and CCL-2 synthesis was also observed when cells were treated with H2O2. In vivo, high CCL-2 production was detected on hypoxic zones of melanoma tumors (where tryptase-positive MCs were also found). Pimonidazole and CCL-2 positive staining diminished when B16-F1 cell–inoculated animals were treated with trolox, nifedipine, or the adenosine receptor 2A antagonist KW6002. Our results show that MCs are located preferentially in hypoxic zones of melanoma tumors, hypoxia-induced CCL-2 production in MCs requires calcium rise mediated by glutathionylation and membrane translocation of LVDCCs, and this mechanism of CCL-2 synthesis seems to operate in other cells inside melanoma tumors, with the participation of the adenosine receptor 2A.

Phenotypic and Functional Heterogeneity of Low-Density and High-Density Human Lung Macrophages

“…We cannot exclude the possibility that a percentage of LDMs or HDMs are tumor-associated macrophages [ 19 , 84 ]. There is also the possibility that in vivo exposure to altered tumor microenvironment, such as low pH [ 85 ], hypoxia [ 86 , 87 ], lactate [ 88 ] or adenosine [ 89 ] may have affected the phenotypic and the functional activity of lung macrophages. In addition, it has been recently demonstrated that macrophages isolated from lung adenocarcinoma differ phenotypically from those obtained from distant and macroscopically normal lung parenchyma [ 39 ].…”

Mast Cells and Skin and Breast Cancers: A Complicated and Microenvironment-Dependent Role

“…Hypoxic environments are highly immunosuppressive due in part to low pH and elevated concentrations of ROS, adenosine, and nitric oxide [ 143 ]. Interestingly, mast cells are frequently localized in hypoxic regions of tissues such as melanoma and associated with the active angiogenic processes initiated in these microenvironments [ 5 , 144 ].…”

“…A moderate level of hypoxia does not induce significant cell death of bone marrow-derived mast cells, nor does it inhibit their degranulation [ 144 ]. Instead, hypoxic conditions drive HIF1α and subsequent ROS production in exposed mast cells as seen in vitro studies [ 144 ].…”

“…A moderate level of hypoxia does not induce significant cell death of bone marrow-derived mast cells, nor does it inhibit their degranulation [ 144 ]. Instead, hypoxic conditions drive HIF1α and subsequent ROS production in exposed mast cells as seen in vitro studies [ 144 ]. Significantly, hypoxia transiently elevates mRNA levels of VEGF-A and TNF above baseline and stably promotes CCL2 protein production, the latter being dependent on ROS and LVDCC [ 144 ].…”

“…Instead, hypoxic conditions drive HIF1α and subsequent ROS production in exposed mast cells as seen in vitro studies [ 144 ]. Significantly, hypoxia transiently elevates mRNA levels of VEGF-A and TNF above baseline and stably promotes CCL2 protein production, the latter being dependent on ROS and LVDCC [ 144 ]. As determined by confocal microscopy, there is high overlap between tryptase + mast cells and hypoxic regions (pimonidazole + cells) in murine melanomas including B16-F1 and -F10, along with overlap with CCL2 expression in the former [ 144 ].…”