Hypoxia regulates placental angiogenesis via alternatively activated macrophages

Uterine and placental macrophages play critical roles in maintaining a normal pregnancy. The majority of these macrophages are believed to be alternatively activated macrophages (M2).

Mouse bone marrow cells were differentiated into macrophages and polarized to M2 in vitro by treatment with IL-4 [M2a] or IL-10 [M2c] and M1 with LPS/IFN-γ as controls. Macrophage subtypes were confirmed by surface markers and characterized by gene expression profiles.

Compared to M1, M2 showed strong pro-angiogenic activity by expressing higher mRNA for angiogenic-associated factors (Cxcl12, Vegfa, PlGF, Mmp2). M2c produced the highest levels of PlGF, Mmp2, and Cxcr4. To mimic the normal placental microenvironment, M2 were exposed to hypoxia and sex hormones (progesterone, estrogen). In both M2c and M2a, severe hypoxic (1%-3% O₂ ) exposure significantly suppressed PlGF, Cxcl12, and Mmp2 mRNA, but not Vegfa, compared to normoxia (21% O₂ ) or physiological hypoxia (5% O₂ ). mRNA expression returned to normal when hypoxic cells were returned to normoxia. Hypoxia (1%) reduced antioxidant levels in M2 and re-exposure to normoxia significantly increased superoxide dismutase (Sod1, Sod2) and heme oxygenase-1 (HO-1) levels in M2a, and only glutathione peroxidase (Gpx1, Gpx3, Gpx4) in M2c. However, progesterone and estrogen treatment had minimal effects on angiogenic factor expression in M2.

M2, particularly M2c, displayed strong pro-angiogenic potential, which decreased under severe hypoxia such as in early pregnancy. Thus, conditions that alter the placental oxygen microenvironment or macrophage response in early pregnancy might cause aberrant angiogenesis and vascular remodeling, and lead to abnormal placental vascular development.