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Preeclampsia is a leading cause of maternal morbidity and mortality worldwide, complicating 2–8% of pregnancies, and is recognized as an independent risk factor for CVD by the ACC/AHA. The etiology of preeclampsia is poorly understood. However, the role of placental extracellular matrix (pECM) and extravillous trophoblast in placental blood vessel formation and preeclampsia is understood. The objective is to identify phenotypical changes in extravillous trophoblast (HTR8/SVneo) in a preeclampsia microenvironment using healthy and preeclamptic patient placental samples vs Dahl S rats (SS) as an animal model of superimposed preeclampsia and explore a novel therapeutic modality restoring trophoblastic phenotype by phototherapy using a 670nm light (NIR). Placentas from rats and patients were decellularized and characterized using pECM markers galectin‐3 and heparan sulfate proteoglycan 2 (HSP2) by western blot and immunofluorescence. pECM was used as substrate for HTR8/SVneo. Proliferation, migration, and apoptosis assays were used to determine cell behavior in the placental microenvironment. Immunofluorescence for placental growth factor (PLGF) and transforming growth factor (TGFβ) was performed on HTR8/SVneo cultured on both pECM. NIR was applied to HTR8/SVneo cultured on patient and rat pECM with 4J/cm2 intensity. In patient samples galectin‐3 expression was reduced by 34.21%±13.16 in preeclamptic PE compared to control patient pECM. HSP2 was reduced by 14.75%±6.11 in PE compared to control patient pECM. In rats galectin‐3 expression was reduced to 51.58%±3.78 in SS compared to control rat pECM. HSP2 was reduced to 45.15%± 0.63 in SS compared to control rat pECM. Migration of HTR8/SVneo on pECM (patient: control 11.08±0.82, PE 6.45±0.88; rats: control 3.87±0.96, SS 4.42±1.18) and proliferation of HTR8/SVneo on pECM were decreased in patients (# of cells X 1000) control: 59.125±6262.9, PE:67.125±7706.5 and rat control 5.16±0.79, SS 4.25±0.6 samples. Proliferation and migration were restored by NIR treatment: proliferation patient: control 101.375±2075.4; PE 106.875±4870.7, migration rat: control 12.58±7.06, SS 5.11±2.55. An increased number of apoptotic HTR8/SVneo was observed in preeclamptic pECM compared to healthy pECM: patient control 2.4±0.24, PE 3±0.4 and rat apoptotic cells control 5.2±0.89; SS 7.4±1.23). NIR decreased the number of apoptotic cells, patient:1.2 ±0.58, PE 1.4±0.5; rat control 3.9±0.43, SS 5.4±0.73. PLGF and TGFβ were downregulated in HTR8/SVneo when cultured on preeclamptic pECM from patients compared to healthy pECM and unchanged in rats. Fluorescence Intensity of PLGF in patient pECM; control 102.12±14.91, PE 68.77±9.73, TGFβ control 88.86±2.51, PE 63.59±4.83 and rats; PLGF control 65.18±0.79, SS 61.09±0.68; TGFβ control 60.78±0.48, SS 61.47 ± 0.45. The decrease was reversed by NIR treatment in patients; PLGF control 165.05±12.32, PE 175.14±12.48, TGFβ control 74.99±7.17, PE 91.03±2.27; rat PLGF control 66.58±0.99, SS 65.57±1.52, TGFβ control 66.67±1.58, SS 63.82±1.53). Extracellular matrix...

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Preeclampsia is a serious pregnancy disorder which in extreme cases may lead to maternal and fetal injury or death. Preexisting conditions which increase oxidative stress, e.g., hypertension and diabetes, increase the mother’s risk to develop preeclampsia. Previously, we established that when the extracellular matrix is exposed to oxidative stress, trophoblast function is impaired, and this may lead to improper placentation. We investigated how the oxidative ECM present in preeclampsia alters the behavior of first trimester extravillous trophoblasts. We demonstrate elevated levels of advanced glycation end products (AGE) and lipid oxidation end product 4-hydroxynonenal in preeclamptic ECM (28%, and 32% increase vs control, respectively) accompanied with 35% and 82% more 3-chlorotyrosine and 3-nitrotyrosine vs control, respectively. Furthermore, we hypothesized that 670 nm phototherapy, which has antioxidant properties, reverses the observed trophoblast dysfunction as depicted in the improved migration and reduction in apoptosis. Since NO is critical for placentation, we examined eNOS activity in preeclamptic placentas compared to healthy ones and found no differences; however, 670 nm light treatment triggered enhanced NO availability presumably by using alternative NO sources. Light exposure decreased apoptosis and restored trophoblast migration to levels in trophoblasts cultured on preeclamptic ECM. Moreover, 670 nm irradiation restored expression of Transforming Growth Factor (TGFβ) and Placental Growth Factor (PLGF) to levels observed in trophoblasts cultured on healthy placental ECM. We conclude the application of 670 nm light can successfully mitigate the damaged placental microenvironment of late onset preeclampsia as depicted by the restored trophoblast behavior.

Preeclampsia is a leading cause of maternal morbidity and mortality worldwide, complicating 2-8% of pregnancies. The etiology of PE is poorly understood, the role of placental extracellular matrix (pECM) in PE is understudied. Extravillous trophoblasts (HTR8/svneo) and pECM are crucial in placental blood vessel formation, including spiral arteries. The objective is to identify changes in signaling mechanisms in late onset PE and how HTR8/svneo respond. Additionally, we explored a novel therapeutic to restore HTR8/svneo phenotype by phototherapy using a 670nm LED light. Western blot analysis shows eNOS, phospho-eNOS and Hsp90 expressions are unchanged in late onset PE placental samples despite previous reports in early onset of PE. The association of Hsp90 to eNOS and eNOS to phospho-eNOS is reduced in the PE sample. Soluble Flt, a diagnostic marker for PE, levels are increased in media from HTR8/svneo trophoblasts cultured on PE pECM. Endothelial function is significantly reduced in an isolated vessel as determined by acetylcholine stimulation. 670nm light reverses this impairment. Migration of HTR8/SVneo on PE pECM is reduced (control 86.29±18.58, PE 30.16±15.49) but proliferation of HTR8/SVneo on PE pECM is unaffected in patients (control: 59.125±6262.9, PE:67.125±7706.5). Proliferation and migration are improved by 670nm light (proliferation control 101.375±2075.4; PE 106.875±4870.7, migration control 125±11.97, PE 144.84±16.01). An increase of apoptotic HTR8/SVneo cultured on PE pECM is observed compared to healthy pECM in patients control 2.4±0.24, PE 3±0.4 samples. 670nm light treatment decreases the number of apoptotic cells, control 1.2 ±0.58, PE 1.4±0.5. Conclusion: In late onset preeclampsia, endothelial function is decreased, sFlt release is increased, the eNOS/Hsp90/p-eNOS axis is slightly reduced pointing to O 2 - synthesis. Migration is decreased likely due to impaired trophoblast invasion, but proliferation is unaffected. 670nm light treatment can reverse some of the effects of the preeclamptic microenvironment. Extracellular matrix is altered in late onset PE leading to an abnormal HTR8/SVneo phenotype. 670nm light restores the phenotype of HTR8/SVneo on PE pECM suggesting 670nm light as a therapeutic treatment.