Dichotomy in hypoxia-induced mitochondrial fission in placental mesenchymal cells during development and preeclampsia: consequences for trophoblast mitochondrial homeostasis.

Dynamic changes in physiologic oxygen are required for proper placenta development; yet, when low-oxygen levels persist, placental development is halted, culminating in preeclampsia (PE), a serious complication of pregnancy. Considering mitochondria's function is intimately linked to oxygen changes, we investigated the impact of oxygen on mitochondrial dynamics in placental mesenchymal stromal cells (pMSCs) that are vital for proper placental development. Transmission electron microscopy, proximity ligation assays for mitochondrial VDAC1 and endoplasmic reticulum IP3R, and immunoanalyses of p-DRP1 and OPA1, demonstrate that low-oxygen conditions in early 1st trimester and PE promote mitochondrial fission in pMSCs. Increased mitochondrial fission of mesenchymal cells was confirmed in whole PE placental tissue sections. Inhibition of DRP1 oligomerization with MDiVi-1 shows that low oxygen-induced mitochondrial fission is a direct consequence of DRP1 activation, likely via HIF1. Mitophagy, a downstream event prompted by mitochondrial fission, is a prominent outcome in PE, but not 1st trimester pMSCs. We also investigated whether mesenchymal-epithelial interactions affect mitochondrial dynamics of trophoblasts in PE placentae. Exposure of trophoblastic JEG3 cells to exosomes of preeclamptic pMSCs caused heightened mitochondrial fission in the cells via a sphingomyelin-dependent mechanism that was restored by MDiVi-1. Our data uncovered dichotomous regulation of mitochondrial fission and health in human placental mesenchymal cells under physiologic and pathologic hypoxic conditions and its impact on neighboring trophoblast cells.

JMJD6 Dysfunction Due to Iron Deficiency in Preeclampsia Disrupts Fibronectin Homeostasis Resulting in Diminished Trophoblast Migration.

The mechanisms contributing to excessive fibronectin in preeclampsia, a pregnancy-related disorder, remain unknown. Herein, we investigated the role of JMJD6, an O2- and Fe2+-dependent enzyme, in mediating placental fibronectin processing and function. MALDI-TOF identified fibronectin as a novel target of JMJD6-mediated lysyl hydroxylation, preceding fibronectin glycosylation, deposition, and degradation. In preeclamptic placentae, fibronectin accumulated primarily in lysosomes of the mesenchyme. Using primary placental mesenchymal cells (pMSCs), we found that fibronectin fibril formation and turnover were markedly impeded in preeclamptic pMSCs, partly due to impaired lysosomal degradation. JMJD6 knockdown in control pMSCs recapitulated the preeclamptic FN phenotype. Importantly, preeclamptic pMSCs had less total and labile Fe2+ and Hinokitiol treatment rescued fibronectin assembly and promoted lysosomal degradation. Time-lapse imaging demonstrated that defective ECM deposition by preeclamptic pMSCs impeded HTR-8/SVneo cell migration, which was rescued upon Hinokitiol exposure. Our findings reveal new Fe2+-dependent mechanisms controlling fibronectin homeostasis/function in the placenta that go awry in preeclampsia.

Increased placental mitochondrial fusion in gestational diabetes mellitus: an adaptive mechanism to optimize feto-placental metabolic homeostasis?

INTRODUCTION: Gestational diabetes mellitus (GDM), a common pregnancy disorder, increases the risk of fetal overgrowth and later metabolic morbidity in the offspring. The placenta likely mediates these sequelae, but the exact mechanisms remain elusive. Mitochondrial dynamics refers to the joining and division of these organelles, in attempts to maintain cellular homeostasis in stress conditions or alterations in oxygen and fuel availability. These remodeling processes are critical to optimize mitochondrial function, and their disturbances characterize diabetes and obesity. METHODS AND RESULTS: Herein we show that placental mitochondrial dynamics are tilted toward fusion in GDM, as evidenced by transmission electron microscopy and changes in the expression of key mechanochemical enzymes such as OPA1 and active phosphorylated DRP1. In vitro experiments using choriocarcinoma JEG-3 cells demonstrated that increased exposure to insulin, which typifies GDM, promotes mitochondrial fusion. As placental ceramide induces mitochondrial fission in pre-eclampsia, we also examined ceramide content in GDM and control placentae and observed a reduction in placental ceramide enrichment in GDM, likely due to an insulin-dependent increase in ceramide-degrading ASAH1 expression. CONCLUSIONS: Placental mitochondrial fusion is enhanced in GDM, possibly as a compensatory response to maternal and fetal metabolic derangements. Alterations in placental insulin exposure and sphingolipid metabolism are among potential contributing factors. Overall, our results suggest that GDM has profound impacts on placental mitochondrial dynamics and metabolism, with plausible implications for the short-term and long-term health of the offspring.

Faulty oxygen sensing disrupts angiomotin function in trophoblast cell migration and predisposes to preeclampsia.

Human placenta development and a successful pregnancy is incumbent upon precise oxygen-dependent control of trophoblast migration/invasion. Persistent low oxygen leading to failed trophoblast invasion promotes inadequate spiral artery remodeling, a characteristic of preeclampsia. Angiomotin (AMOT) is a multifaceted scaffolding protein involved in cell polarity and migration, yet its upstream regulation and significance in the human placenta remain unknown. Herein, we show that AMOT is primarily expressed in migratory extravillous trophoblast cells (EVTs) of the intermediate and distal anchoring column. Its expression increases after 10 weeks of gestation when oxygen tension rises and EVT migration/invasion peaks. Time-lapse imaging confirmed that the AMOT 80-kDa isoform promotes migration of trophoblastic JEG3 and HTR-8/SVneo cells. In preeclampsia, however, AMOT expression is decreased and its localization to migratory fetomaternal interface EVTs is disrupted. We demonstrate that Jumonji C domain-containing protein 6 (JMJD6), an oxygen sensor, positively regulates AMOT via oxygen-dependent lysyl hydroxylation. Furthermore, in vitro and ex vivo studies show that transforming growth factor-ß (TGF-ß) regulates AMOT expression, its interaction with polarity protein PAR6, and its subcellular redistribution from tight junctions to cytoskeleton. Our data reveal an oxygen- and TGF-ß-driven migratory function for AMOT in the human placenta, and implicate its deficiency in impaired trophoblast migration that plagues preeclampsia.

Ceramide-induced BOK promotes mitochondrial fission in preeclampsia.

Mitochondria are in a constant balance of fusing and dividing in response to cellular cues. Fusion creates healthy mitochondria, whereas fission results in removal of non-functional organelles. Changes in mitochondrial dynamics typify several human diseases. However, the contribution of mitochondrial dynamics to preeclampsia, a hypertensive disorder of pregnancy characterized by placental cell autophagy and death, remains unknown. Herein, we show that the mitochondrial dynamic balance in preeclamptic placentae is tilted toward fission (increased DRP1 expression/activation and decreased OPA1 expression). Increased phosphorylation of DRP1 (p-DRP1) in mitochondrial isolates from preeclamptic placentae and transmission electron microscopy corroborated augmented mitochondrial fragmentation in cytotrophoblast cells of PE placentae. Increased fission was accompanied by build-up of ceramides (CERs) in mitochondria from preeclamptic placentae relative to controls. Treatment of human choriocarcinoma JEG3 cells and primary isolated cytrophoblast cells with CER 16:0 enhanced mitochondrial fission. Loss- and gain-of-function experiments showed that Bcl-2 member BOK, whose expression is increased by CER, positively regulated p-DRP1/DRP1 and MFN2 expression, and localized mitochondrial fission events to the ER/MAM compartments. We also identified that the BH3 and transmembrane domains of BOK were vital for BOK regulation of fission. Moreover, we found that full-length PTEN-induced putative kinase 1 (PINK1) and Parkin, were elevated in mitochondria from PE placentae, implicating mitophagy as the process that degrades excess mitochondria fragments produced from CER/BOK-induced fission in preeclampsia. In summary, our study uncovered a novel CER/BOK-induced regulation of mitochondrial fission and its functional consequence for heightened trophoblast cell autophagy in preeclampsia.