Discussion
PFKFB3 controls the LPS-induced inflammation via the NF-κB pathway and impacts trophoblasts function such as adhesion, oxidative stress, apoptosis, migration, and invasion, thereby potentially participating in the preeclamptic etiopathogenesis.
Methods
Immunohistochemistry, qRT-PCR, and Western blot analysis (WB) were used to detect the expression of PFKFB3 in preeclamptic and normal placentas. Lipopolysaccharide (LPS)-induced HTR8/SVneo cells were established as the in vitro model to simulate the overactive inflammation at the uteroplacental interface of PE, which were subsequently transfected with PFKFB3 siRNA. The expression of PFKFB3, NF-κB-p-p65, phosphorylation states of NF-κB-p65, ICAM-1, Bcl-2, BAX, and MMP2 were detected by WB. qRT-PCR was used to detect the expression of TNF-α and IL-1β. The ICAM-1 expression was also reflected by monocyte adhesion assay. Reactive Oxygen Species (ROS) levels were detected by DCFH-DA (2,7-Dichlorodi-hydrofluorescein diacetate). Apoptosis was detected using Annexin V-FITC staining. Migration and invasion were measured by wound-healing and transwell assays.
Results
PFKFB3 was up-regulated in the preeclamptic placenta. In LPS-treated HTR-8/Svneo cells, the inhibition of PFKFB3 blocked the NF-κB signal pathway, thereby downregulating the expression of proinflammatory cytokines and adhesion molecules, meanwhile, PFKFB3 knockdown significantly alleviated monocyte adhesion, oxidative stress, apoptosis, and reinstated migration and invasive capacity.