O1092: Dysregulation of Angiogenesis in a Mouse Model of Intrauterine Growth Restriction Is Associated with Abnormal Expression of the Regulatory microrna-15b

Background: Approximately 10% of pregnancies are complicated by intrauterine growth restriction (IUGR) which is responsible for up to 50% of stillbirths. Placental insufficiency, aberrant angiogenesis, and a relative hypoxic state are thought to contribute to the pathogenesis of IUGR. MicroRNAs (miRs) are short non-coding RNAs that regulate gene expression at the post-transcriptional level by targeting mRNA for degradation or suppressing mRNA translation. We hypothesize that miR-15b, an anti-angiogenic miR, is dysregulated in IUGR. We utilized a maternal caloric restricted IUGR mouse model to analyze placental gene expression of miR-15b, its downstream target - vascular endothelial growth factor a (VEGFa), as well as gene expression of proteins elevated in states of hypoxia.

Methods: Pregnant mice were provided ad libitum access to food between E1-E8. From E9-E18, dams received either a 50% caloric restricted diet (IUGR) or continued ad libitum access (controls). Placentas were harvested at E18.5. RNA was extracted using a column-based extraction technique. Primers for miR15b as well as mRNAs of interest were used to compare levels of expression between IUGR and control placentas.

Results: Ten placentas in the IUGR group were compared to ten placentas in the control group. Expression of the anti-angiogenic miR-15b was upregulated in the IUGR group compared to the control group (Figure 1). VEGFa, which is downregulated in response to increased levels of miR-15b, was suppressed. B-cell lymphoma-2 (Bcl-2), a protein regulated by VEGFa, also demonstrated lower levels of gene expression, but this did not reach statistical significance (Figure 1). We also found that Hypoxia Inducible Factor 1 Subunit alpha (HIF-1a), superoxide dismutase 2 (SOD2), and NADPH oxidase 2 (Nox2), proteins elevated in the setting of oxidative stress, were upregulated in our IUGR mouse model. Despite this hypoxic state, miR-21, a regulator of HIF1a, was not dysregulated in IUGR placentas compared to controls.

Conclusion: We demonstrate that our murine model of IUGR exhibits increased miR-15b expression which is associated with a significantly reduced VEGFa gene expression. As VEGFa contributes to the modulation of angiogenesis and apoptosis through multiple downstream effector proteins, we suspect that miR-15b may be a key driver of IUGR. In addition, HIF1a gene expression is upregulated despite no alteration in miR-21, leading us to speculate that the hypoxic state associated with IUGR stimulates HIF-1a, SOD2, and NOX2 production, but this response fails to offset the effects of miR-15b, a hypothesis supported by diminished circulation of VEGFa (Figure 2). This data provides insight into specific epigenetic drivers of angiogenesis. Furthermore, it is possible that trans-placental introduction of antisense oligonucleotides targeted to inhibit miR-15b may be a therapeutic option to mitigate the effects of IUGR.