The migratory capacity of HTR8/SVneo cells under hypoxiais regulated by KLF6

RACCA A.; RODRIGUEZ-LOMBARDI G.; KOURDOVA L.; GENTI-RAIMONDI S; MIRANDA A.L.; PANZETTA-DUTARIA GM

Buenos Aires

Congreso; 8th Latin American Symposium on Maternal-Fetal Interaction and Placenta (VIII SLIMP); 2019

IFPA

Objective: Pregnancy complications, such as preeclampsia are, at least in part, due to alterations in the transcriptional response to physiological stimuli such as hypoxia, where HIF-1α plays an essential role. KLF6 is a transcription factor early activated in response to hypoxia in a partially HIF-1α-dependent manner. In this context, we evaluated KLF6 impact on the migration capacity of trophoblast cells.Methods: The HTR8/SVneo cell line was used as a migratory trophoblast cell model. KLF6 expression was either silenced with a specific siRNA or stably overexpressed through pLenti-KLF6 transduction. Cells were cultured in a hypoxia chamber (1% O2). Migration was evaluated in wound healing assays. The expression of migration-related molecules such as MMP9, MMP2, and uPAR, as well as that of HIF-1α was analyzed by qRT-PCR and western blot, respectively. MMP activity was detected by zymography assays. Reactive oxygen species (ROS) were measured by flow cytometry using the H2DCFDA probe.Results: KLF6 silencing in hypoxia conditions leads to an increase in cell migration accompanied by an increase in MMP9 protein expression and activity, whereas MMP2 and uPAr remain unmodified. Surprisingly, HIF-1α transcript and protein levels increase when KLF6 is downregulated and HIF-1α protein level decrease in HTR8/SVneo stably overexpressing KLF6 as compared to the control cell line. HIF-1α expression is partially restored by N-acetylcysteine treatment of KLF6-silenced cells. Thus, modulation of HIF-1α expression could be a direct transcriptional consequence of KLF6 absence or an indirect response to ROS induced in KLF6-silenced cells. Conclusions: Herein, we demonstrate that KLF6 modulates the migration of trophoblasts in hypoxia since its downregulation releases the migration machinery. These results are in line with a higher KLF6-expression reported in the placental bed from preeclamptic pregnancies. In addition, present results suggest a regulation loop between two transcription factors, KLF6 and HIF-1α, relevant for trophoblast physiology and pathophysiology.