Nastassia Pham

Introduction. Pre-eclampsia is a pregnancy induced hypertensive disorder characterized by systolic blood pressure greater than 140 mmHg and diastolic blood pressure greater than 90 mmHg that usually presents after 20 weeks of gestation^1,2. The pathology begins with malfunction during trophoblast invasion, resulting in reduced uteroplacental perfusion¹. This stresses the placenta, leading to endothelial dysregulation. Vascular dysfunction of the placenta promotes release of humoral factors into maternal circulation, triggering multiple organ injuries¹. Nitric oxide derived from endothelial nitric oxide synthase (eNOS) prevents vascular inflammation². The transcription factor NF-kB has been shown to play a role in inflammation of endothelial cells³. Studies have shown that NF-kB activating stimuli, such as tumor necrosis factor (TNF- α), suppress eNOS mRNA and protein levels indirectly by upregulating miR-155 levels under inflammatory conditions³. These findings suggest miR-155 suppression as a potential therapeutic target for preventing vascular inflammation in pre-eclampsia³.  Methods. Human umbilical vein endothelial cells (HUVECs) were isolated from human umbilical cord veins by collagenase treatment and grown in M199 media³. The cells were then transfected with antigomiR-155, miR-155 mimics, a negative control of antagomiR-101 and miR-101, Dicer siRNA, or scrambled control³. The levels of miR-155 were determined using polymerase chain reaction³. Western blot was used to determine eNOS protein levels³. Chromatin immunoprecipitation assay was used to determine where NF-kB binds on the pGL3-MIR155 host gene promoter Luc construct³. Results. TNF- α increased MIR155 host gene (MIR155HG) expression by increasing transcriptional activity of the MIR155HG promoter construct at the 1150 nucleotide region upstream from the promoter, thus increasing miR-155 generation in HUVECs^3,5. The NF-kB dependent activation of miR-155 biogenesis suppressed eNOS expression⁴. Among the different miRNAs induced in HUVECs, miR155 was found to target the 3’ UTR of the human eNOS mRNA³. NF-kB inhibitors reversed TNF- α mediated transcriptional expression of MIR155HG and suppression of eNOS expression³. Conclusions. Studies found that NF-kB binding to the promoter is responsible for MIR155HG dependent miR155 expression³. The -1150 NF-kB binding site is involved in negatively regulating eNOS expression in endothelial cells via miR155 synthesis^3,5. miR-155 binds to the 3’ UTR region of the eNOS mRNA and destabilizes the mRNA strand, preventing translation³. As a result, activation of NF-kB leads to inhibition of eNOS expression and nitric oxide production, triggering endothelial dysfunction in pre-eclamptic women³.

1.  Burton GJ, Redman CW, Roberts JM, Moffett A. Pre-eclampsia: Pathophysiology and Clinical Implications. Bmj. 2019:l2381. doi:10.1136/bmj.l2381
2. Matsubara K, Higaki T, Matsubara Y, Nawa A. Nitric Oxide and Reactive Oxygen Species in the Pathogenesis of Preeclampsia. International Journal of Molecular Sciences. 2015;16(3):4600-4614. doi:10.3390/ijms16034600
3. Lee K-S, Kim J, Kwak S-N, et al. Functional Role of NF-κB in Expression of Human Endothelial Nitric Oxide Synthase. Biochemical and Biophysical Research Communications. 2014;448(1):101-107. doi:10.1016/j.bbrc.2014.04.079
4. Kim S, Lee K-S, Choi S, et al. NF-κB–responsive miRNA-31-5p Elicits Endothelial Dysfunction Associated with Preeclampsia via Down Regulation of Endothelial Nitric Oxide Synthase. Journal of Biological Chemistry. 2018;293(49):18989-19000. doi:10.1074/jbc.ra118.005197
5. Choi S, Park M, Kim J, et al. TNF- α Elicits Phenotypic and Functional Alterations of Vascular Smooth Muscle Cells by miR-155-5p–dependent Down Regulation of cGMP-dependent Kinase 1. Journal of Biological Chemistry. 2018;293(38):14812-14822. doi:10.1074/jbc.ra118.004220