A Prospective Study on Risk Prediction of Preeclampsia Using Bi-Platform Calibration and Machine Learning.

Preeclampsia is a pregnancy syndrome characterized by complex symptoms which cause maternal and fetal problems and deaths. The aim of this study is to achieve preeclampsia risk prediction and early risk prediction in Xinjiang, China, based on the placental growth factor measured using the SiMoA or Elecsys platform. A novel reliable calibration modeling method and missing data imputing method are proposed, in which different strategies are used to adapt to small samples, training data, test data, independent features, and dependent feature pairs. Multiple machine learning algorithms were applied to train models using various datasets, such as single-platform versus bi-platform data, early pregnancy versus early plus non-early pregnancy data, and real versus real plus augmented data. It was found that a combination of two types of mono-platform data could improve risk prediction performance, and non-early pregnancy data could enhance early risk prediction performance when limited early pregnancy data were available. Additionally, the inclusion of augmented data resulted in achieving a high but unstable performance. The models in this study significantly reduced the incidence of preeclampsia in the region from 7.2% to 2.0%, and the mortality rate was reduced to 0%.

Pravastatin Protects Cytotrophoblasts from Hyperglycemia-Induced Preeclampsia Phenotype.

There are no effective therapies to prevent preeclampsia (PE). Pravastatin shows promise by attenuating processes associated with PE such as decreased cytotrophoblast (CTB) migration, aberrant angiogenesis, and increased oxidative stress. This study assesses the effects of pravastatin on hyperglycemia-induced CTB dysfunction. METHODS: Human CTB cells were treated with 100, 150, 200, 300, or 400 mg/dL glucose for 48 h. Some cells were pretreated with pravastatin (1 µg/mL), while others were cotreated with pravastatin and glucose. The expression of urokinase plasminogen activator (uPA), plasminogen activator inhibitor 1 (PAI-1) mRNA, vascular endothelial growth factor (VEGF), placenta growth factor (PlGF), soluble fms-like tyrosine kinase-1 (sFlt-1), and soluble endoglin (sEng) were measured. CTB migration was assayed using a CytoSelect migration assay kit. Statistical comparisons were performed using an analysis of variance with Duncan's post hoc test. RESULTS: The hyperglycemia-induced downregulation of uPA was attenuated in CTB cells pretreated with pravastatin at glucose levels > 200 mg/dL and cotreated at glucose levels > 300 mg/dL (p < 0.05). Hyperglycemia-induced decreases in VEGF and PlGF and increases in sEng and sFlt-1 were attenuated in both the pretreatment and cotreatment samples regardless of glucose dose (p < 0.05). Pravastatin attenuated hyperglycemia-induced dysfunction of CTB migration. CONCLUSIONS: Pravastatin mitigates stress signaling responses in hyperglycemic conditions, weakening processes leading to abnormal CTB migration and invasion associated with PE in pregnancy.

Placental growth factor alleviates hyperglycemia-induced trophoblast pyroptosis by regulating mitophagy.

INTRODUCTION: Hyperglycemia is closely related to trophoblast dysfunction during pregnancy and results in suppressed invasion, migration, and pro-inflammatory cell death of trophoblasts. Hyperglycemia is a dependent risk factor for gestational hypertension accompanied by decreased placental growth factor (PLGF), which is important for maternal and fetal development. However, there is currently a lack of evidence to support whether PLGF can alleviate trophoblast cell dysfunction caused by high blood sugar. Here, we aim to clarify the effect of hyperglycemia on trophoblast dysfunction and determine how PLGF affects this process. METHODS: The changes in placental tissue histomorphology from gestational diabetes mellitus (GDM) patients were compared with those of normal placentas. HTR8/SVneo cells were cultured in different amounts of glucose to examine cellular pyroptosis, migration, and invasion as well as PLGF levels. Furthermore, the levels of pyroptosis-related proteins (NLRP3, pro-caspase1, caspase1, IL-1ß, and Gasdermin D [GSDMD]) as well as autophagy-related proteins (LC3-II, Beclin1, and p62) were examined by Western blotting. The GFP-mRFP-LC3-II system and transmission electron microscopy were used to detect mitophagy levels, and small interfering RNAs targeting BCL2 Interacting Protein 3 (siBNIP3) and PTEN-induced kinase 1 (siPINK1) were used to determine the role of mitophagy in pyroptotic death of HTR-8/SVneo cells. RESULTS: Our results show that hyperglycemia upregulates NLRP3, pro-caspase1, caspase1, IL-1ß at the protein level in GDM patients. High glucose (HG, 25 mM) inhibits viability, invasion, and migration of trophoblast cells while suppressing superoxide dismutase levels and promoting malondialdehyde production, thus leading to a senescence associated beta-gal-positive cell burst. PLGF levels in nucleus and the cytosol are also inhibited by HG, whereas PLGF treatment inhibited pyroptosis-related protein levels of NLRP3, pro-caspase1, caspase1, IL-1ß, and GSDMD, Gasdermin D N-terminal domain (GSDMD-N). HG-induced mitochondrial dysfunction and BNIP3 and PINK1/Parkin expression. Knocking down BINP3 and PINK1 abolished the protective role of PLGF by preventing mitophagy. CONCLUSION: PLGF inhibited hyperglycemia, while PLGF reversed hyperglycemic injury by promoting mitophagy via the BNIP3/PINK1/Parkin pathway. Altogether, these results suggest that PLGF may protect against trophoblast dysfunction in diabetes.

Aqueous Humor Cytokine Analysis in Age-Related Macular Degeneration After Switching From Aflibercept to Faricimab.

Purpose: To examine the changes in aqueous humor cytokine levels and clinical outcomes of switching from aflibercept to faricimab in eyes with neovascular age-related macular degeneration (nAMD). Methods: Fifty-four eyes of 54 patients with AMD undergoing treatment with aflibercept under a treat-and-extend (TAE) regimen were switched to faricimab and studied prospectively. Best-corrected visual acuity (BCVA; in logarithm of the minimum angle of resolution), central retinal thickness (CRT), central choroidal thickness (CCT), and exudative status were analyzed using optical coherence tomography. Aqueous humor was collected before and after the switch, and angiopoietin-2 (Ang-2), placental growth factor (PlGF), and vascular endothelial growth factor (VEGF) A levels were measured. Results: After switching from aflibercept to faricimab, exudative changes improved in 28 eyes (52%), remained stable in eight eyes (15%), and worsened in 18 eyes (33%). BCVA changed from 0.27 ± 0.31 to 0.26 ± 0.29 (P = 0.46), CRT decreased from 306.2 ± 147.5 µm to 278.6 ± 100.4 µm (P = 0.11), and CCT changed from 189.5 ± 92.8 µm to 186.8 ± 93.9 µm (P = 0.21). VEGF-A levels were below the detection sensitivity in many cases throughout the pre- and post-switching periods. Ang-2 significantly decreased from 23.8 ± 23.5 pg/mL to 16.4 ± 21.9 pg/mL (P < 0.001), and PlGF significantly increased from 0.86 ± 0.85 pg/mL to 1.72 ± 1.39 pg/mL (P < 0.001). Conclusions: Switching from aflibercept to faricimab in patients with nAMD may not only suppress VEGF-A but also Ang-2 and reduce exudative changes.

Recombinant human placental growth factor-2 in post-infarction left ventricular dysfunction: a randomized, placebo-controlled, preclinical study.

Placental growth factor (PlGF)-2 induces angio- and arteriogenesis in rodents but its therapeutic potential in a clinically representative post-infarction left ventricular (LV) dysfunction model remains unclear. We, therefore, investigated the safety and efficacy of recombinant human (rh)PlGF-2 in the infarcted porcine heart in a randomized, placebo-controlled blinded study. We induced myocardial infarction (MI) in pigs using 75 min mid-LAD balloon occlusion followed by reperfusion. After 4 w, we randomized pigs with marked LV dysfunction (LVEF < 40%) to receive continuous intravenous infusion of 5, 15, 45 µg/kg/day rhPlGF-2 or PBS (CON) for 2 w using osmotic pumps. We evaluated the treatment effect at 8 w using comprehensive MRI and immunohistochemistry and measured myocardial PlGF-2 receptor transcript levels. At 4 w after MI, infarct size was 16-18 ± 4% of LV mass, resulting in significantly impaired systolic function (LVEF 34 ± 4%). In the pilot study (3 pigs/dose), PIGF administration showed sustained dose-dependent increases in plasma concentrations for 14 days without systemic toxicity and was associated with favorable post-infarct remodeling. In the second phase (n = 42), we detected no significant differences at 8 w between CON and PlGF-treated pigs in infarct size, capillary or arteriolar density, global LV function and regional myocardial blood flow at rest or during stress. Molecular analysis showed significant downregulation of the main PlGF-2 receptor, pVEGFR-1, in dysfunctional myocardium. Chronic rhPIGF-2 infusion was safe but failed to induce therapeutic neovascularization and improve global cardiac function after myocardial infarction in pigs. Our data emphasize the critical need for properly designed trials in representative large animal models before translating presumed promising therapies to patients.

Engineered nanovesicles mediated cardiomyocyte survival and neovascularization for the therapy of myocardial infarction.

Myocardial infarction (MI) leads to substantial cellular necrosis as a consequence of reduced blood flow and oxygen deprivation. Stimulating cardiomyocyte proliferation and angiogenesis can promote functional recovery after cardiac events. In this study, we explored a novel therapeutic strategy for MI by synthesizing a biomimetic nanovesicle (NV). This biomimetic NVs are composed of exosomes sourced from umbilical cord mesenchymal stem cells, which have been loaded with placental growth factors (PLGF) and surface-engineered with a cardiac-targeting peptide (CHP) through covalent bonding, termed Exo-P-C NVs. With the help of the myocardial targeting effect of homing peptides, NVs can be enriched in the MI site, thus improve cardiac regeneration, reduce fibrosis, stimulate cardiomyocyte proliferation, and promote angiogenesis, ultimately resulted in improved cardiac functional recovery. It was demonstrated that Exo-P-C NVs have the potential to offer novel therapeutic strategies for the improvement of cardiac function and management of myocardial infarction.

Placental growth factor mediates pathological uterine angiogenesis by activating the NFAT5-SGK1 signaling axis in the endometrium: implications for preeclampsia development.

After menstruation the uterine spiral arteries are repaired through angiogenesis. This process is tightly regulated by the paracrine communication between endometrial stromal cells (EnSCs) and endothelial cells. Any molecular aberration in these processes can lead to complications in pregnancy including miscarriage or preeclampsia (PE). Placental growth factor (PlGF) is a known contributing factor for pathological angiogenesis but the mechanisms remain poorly understood. In this study, we investigated whether PlGF contributes to pathological uterine angiogenesis by disrupting EnSCs and endothelial paracrine communication. We observed that PlGF mediates a tonicity-independent activation of nuclear factor of activated T cells 5 (NFAT5) in EnSCs. NFAT5 activated downstream targets including SGK1, HIF-1α and VEGF-A. In depth characterization of PlGF - conditioned medium (CM) from EnSCs using mass spectrometry and ELISA methods revealed low VEGF-A and an abundance of extracellular matrix organization associated proteins. Secreted factors in PlGF-CM impeded normal angiogenic cues in endothelial cells (HUVECs) by downregulating Notch-VEGF signaling. Interestingly, PlGF-CM failed to support human placental (BeWo) cell invasion through HUVEC monolayer. Inhibition of SGK1 in EnSCs improved angiogenic effects in HUVECs and promoted BeWo invasion, revealing SGK1 as a key intermediate player modulating PlGF mediated anti-angiogenic signaling. Taken together, perturbed PlGF-NFAT5-SGK1 signaling in the endometrium can contribute to pathological uterine angiogenesis by negatively regulating EnSCs-endothelial crosstalk resulting in poor quality vessels in the uterine microenvironment. Taken together the signaling may impact on normal trophoblast invasion and thus placentation and, may be associated with an increased risk of complications such as PE.

Protective effect of leukotriene receptor antagonist, montelukast, against cyclophosphamide-induced placental toxicity via modulation of NLRP3/IL-1ß signaling pathway in rats.

BACKGROUNDS & AIM: Placental insufficiency is a serious complication that affects pregnancy and fetal growth. Cyclophosphamide (CYC) is considered one of the chemotherapeutic agents. Unfortunately, CYC not only affects tumor cells but also affects healthy cells causing multiple injuries including the placenta. The present study aimed to evaluate the effect of cysteinyl leukotriene receptor antagonist; montelukast (MK), on CYC-induced placental injury in rats. MATERIALS AND METHODS: Forty-eight female Wister rats were randomly divided into 8 experimental groups. Group 1: control pregnant group; Group 2: MK 5 mg-treated pregnant rats; Group 3: MK 10 mg-treated pregnant rats; Group 4: MK 20 mg-treated pregnant rats; Group 5: pregnant rats received CYC (20 mg/kg, i.p); Group 6: pregnant rats received MK 5 mg and CYC; Group 7: pregnant rats received MK 10 mg and CYC; Group 8: pregnant rats received MK 20 mg and CYC. Placental malondialdehyde (MDA), reduced glutathione (GSH), total antioxidant capacity (TAC), placental growth factor (PlGF), and Nod-like receptor p3 (NLRP3) inflammasome were measured. Histological changes, interleukin-1ß (IL-1ß), and cleaved caspase-3 immuno-expressions were also evaluated. RESULTS: CYC showed a significant decrease in placental GSH, TAC, and PlGF with a significant increase in placental MDA, NLRP3, and immuno-expression of IL-1ß and caspase-3. MK showed significant improvement in all oxidative stress (MDA, GSH and TAC), inflammatory (NLRP3 and IL-1ß), and apoptotic (caspase-3) parameters. CONCLUSION: According to the findings, MK was proved to have a possible protective role in CYC-induced placental injury via modulation of NLRP3/IL-1ß signaling pathway with anti-oxidant, anti-inflammatory, and anti-apoptotic effects.

Placental biophysical model for prediction of early onset fetal growth restriction in first and second trimester of pregnancy: A prospective cohort study.

INTRODUCTION: To assess the placental biometry, placental biomarkers and uterine artery Doppler in each trimester of pregnancy for prediction of early-onset fetal growth restriction (EO FGR). METHODS: In this prospective cohort study placental biometry; biomarkers PAPP-A, sFLT-1, and PlGF along with the uterine artery blood flow evaluation was done serially at 11-14, 20-24 and 28-32 weeks of gestation. The above parameters were compared between women with early onset FGR and controls. RESULTS: Out of 1008 fully followed cases, the small for gestational age fetuses were 227/1008 (22.5 %), and EO FGR were 84/1008(8.3 %).The placental length, volume, and PlGF levels were significantly lower, whereas the uterine artery PI(Ut PI) was significantly higher at all time points among cases. The sFLT-1 level showed a significant increase among cases, whereas it decreased among controls from the first to the second trimester. The detection rate using PV/UtA PI was 60 % in the first trimester and 66.7 % in the second trimester at 30 % FPR. CONCLUSION: The PV/Ut PI in first and the second trimester was a good marker for the prediction of pregnancies at increased risk of developing EO FGR.

An examination of the mechanisms driving the therapeutic effects of an AAV expressing a soluble variant of VEGF receptor-1.

In previous animal model studies, we demonstrated the potential of rAAV2-sVEGFRv-1, which encodes a truncated variant of the alternatively spliced soluble version of VEGF receptor-1 (VEGFR1), as a human gene therapy for age-related macular degeneration (AMD) and diabetic retinopathy (DR). Here, we elucidate in vitro some of the mechanisms by which rAAV2-sVEGFRv-1 exerts its therapeutic effects. Human umbilical vein endothelial cells (HUVECs) were infected with rAAV2-sVEGFRv-1 or a control virus vector in the presence of members of the VEGF family to identify potential binding partners via ELISA, which showed that VEGF-A, VEGF-B, and placental growth factor (PlGF) are all ligands of its transgene product. In order to determine the effects of rAAV2-sVEGFRv-1 on cell proliferation and permeability, processes that are important to the progression AMD and DR, HUVECs were infected with the therapeutic virus vector under the stimulation of VEGF-A, the major driver of the neovascularization that characterizes the forms of these conditions most associated with vision loss. rAAV2-sVEGFRv-1 treatment, as a result, markedly reduced the extent to which these processes occurred, with the latter determined by measuring zonula occludens 1 expression. Finally, the human microglial HMC3 cell line was used to show the effects of the therapeutic virus vector upon inflammatory processes, another major contributor to angiogenic eye disease pathophysiology, with rAAV2-sVEGFRv-1 reducing therein the secretion of pro-inflammatory cytokines interleukin (IL)-1ß and IL-6. Combined with our previously published in vivo data, the in vitro activity of the expressed transgene here further demonstrates the great promise of rAAV2-sVEGFRv-1 as a potential human gene therapeutic for addressing angiogenic ocular conditions.

PlGF and VEGF-A/PlGF Heterodimer are Crucial for Recruitment and Activation of Immune Cells During Choroid Neovascularization.

Purpose: Recruitment and activation of inflammatory cells, such as retinal microglia/macrophages, in the subretinal space contribute significantly to the pathogenesis of age-related macular degeneration (AMD). This study aims to explore the functional role of vascular endothelial growth factor (VEGF-A), placental growth factor (PlGF) and VEGF-A/PlGF heterodimer in immune homeostasis and activation during pathological laser-induced choroidal neovascularization (CNV). Methods: To investigate these roles, we utilized the PlGF-DE knockin (KI) mouse model, which is the full functional knockout (KO) of PlGF. In this model, mice express a variant of PlGF, named PlGF-DE, that is unable to bind and activate VEGFR-1 but can still form heterodimer with VEGF-A. Results: Our findings demonstrate that, although there is no difference in healthy conditions, PlGF-DE-KI mice exhibit decreased microglia reactivity and reduced recruitment of both microglia and monocyte-macrophages, compared to wild-type mice during laser-induced CNV. This impairment is associated with a reduction in VEGF receptor 1 (VEGFR-1) phosphorylation in the retinae of PlGF-DE-KI mice compared to C57Bl6/J mice. Corroborating these data, intravitreal delivery of PlGF or VEGF-A/PlGF heterodimer in PlGF-DE-KI mice rescued the immune cell response at the early phase of CNV compared to VEGF-A delivery. Conclusions: In summary, our study suggests that targeting PlGF and the VEGF-A/PlGF heterodimer, thereby preventing VEGFR-1 activation, could represent a potential therapeutic approach for the management of inflammatory processes in diseases such as AMD.

Fibroblasts activation by embryonic signal switching: A novel mechanism of placental growth factor-induced cardiac remodeling.

INTRODUCTION: Cardiac remodeling is defined as cellular interstitial changes that lead dysfunction of the heart after injury. Placental growth factor (PlGF), a member of the VEGF family, has been reported to regulate cardiac hypertrophy in hemodynamic state. We therefore analyze the function of PlGF during cardiac remodeling using cardiac cells and fibroblasts, under Angiotensin II (AngII) stimulation. METHODS: PlGF overexpressed mouse embryonic fibroblasts derived from C57BL/6 mice, were made by deficient retrovirus vector, designated as C57/PlGF. Only retrovirus vector introduced C57 cells (C57/EV) were used as control. After AngII stimulation, wound scratching assay and MTT proliferation assay with or without p38 MAPK inhibitor, SB205580 were performed in retrovirally-introduced C57 cells. Reactive oxygen species (ROS) production, NF-kB activation, IL-6 and TNF-α production were also measured. Then we assessed AngII-induced cell proliferation of mouse cardiac fibroblasts (CFs) and rat primary cardiomyocytes incubating with C57/PlGF conditioned-medium. RESULTS: The PlGF production in C57/PlGF were confirmed by ELISA (1093.48 ± 3.5 pg/ml, ±SE). AngII-induced cell migration, proliferation and H2O2 production were increased in C57/PlGF compared with C57/EV. SB205580 inhibited the AngII-induced cell proliferation in C57/PlGF. In C57/PlGF cells, NF-kB activation was higher, followed by up-regulation of IL-6 and TNF-α production. CFs and cardiomyocytes proliferation increased when stimulated with C57/PlGF conditioned-medium. DISCUSSION: The activation of fibroblast is stimulated by PlGF signaling via p38 MAPK/NF-kB pathway accompanied by elevation of ROS and inflammatory response. Furthermore, these signals stimulate the activation of CFs and cardiomyocytes, indicating that high circulating level of PlGF have a potential to regulate cardiac remodeling.

Alleviation of preeclampsia-like symptoms through PlGF and eNOS regulation by hypoxia- and NF-κB-responsive miR-214-3p deletion.

Preeclampsia is caused by placental hypoxia and systemic inflammation and is associated with reduced placental growth factor (PlGF) and endothelial nitric oxide synthase (eNOS) levels. The molecular signaling axes involved in this process may play a role in the pathogenesis of preeclampsia. Here, we found that hypoxic exposure increased hypoxia-inducible factor-1α (HIF-1α)/Twist1-mediated miR-214-3p biogenesis in trophoblasts, suppressing PlGF production and trophoblast invasion. TNF-α stimulation increased NF-κB-dependent miR-214-3p expression in endothelial cells, impairing eNOS expression and causing endothelial dysfunction. Synthetic miR-214-3p administration to pregnant mice decreased PlGF and eNOS expression, resulting in preeclampsia-like symptoms, including hypertension, proteinuria, and fetal growth restriction. Conversely, miR-214-3p deletion maintained the PlGF and eNOS levels in hypoxic pregnant mice, alleviating preeclampsia-like symptoms and signs. These findings provide new insights into the role of HIF-1/Twist1- and NF-κB-responsive miR-214-3p-dependent PlGF and eNOS downregulation in the pathogenesis of preeclampsia and establish miR-214-3p as a therapeutic or preventive target for preeclampsia and its complications.

Placental growth factor promotes neural invasion and predicts disease prognosis in resectable pancreatic cancer.

BACKGROUND: Surgery represents the only curative treatment option for pancreatic ductal adenocarcinoma (PDAC), but recurrence in more than 85% of patients limits the success of curative-intent tumor resection. Neural invasion (NI), particularly the spread of tumor cells along nerves into extratumoral regions of the pancreas, constitutes a well-recognized risk factor for recurrence. Hence, monitoring and therapeutic targeting of NI offer the potential to stratify recurrence risk and improve recurrence-free survival. Based on the evolutionary conserved dual function of axon and vessel guidance molecules, we hypothesize that the proangiogenic vessel guidance factor placental growth factor (PlGF) fosters NI. To test this hypothesis, we correlated PlGF with NI in PDAC patient samples and functionally assessed its role for the interaction of tumor cells with nerves. METHODS: Serum levels of PlGF and its soluble receptor sFlt1, and expression of PlGF mRNA transcripts in tumor tissues were determined by ELISA or qPCR in a retrospective discovery and a prospective validation cohort. Free circulating PlGF was calculated from the ratio PlGF/sFlt1. Incidence and extent of NI were quantified based on histomorphometric measurements and separately assessed for intratumoral and extratumoral nerves. PlGF function on reciprocal chemoattraction and directed neurite outgrowth was evaluated in co-cultures of PDAC cells with primary dorsal-root-ganglia neurons or Schwann cells using blocking anti-PlGF antibodies. RESULTS: Elevated circulating levels of free PlGF correlated with NI and shorter overall survival in patients with PDAC qualifying for curative-intent surgery. Furthermore, high tissue PlGF mRNA transcript levels in patients undergoing curative-intent surgery correlated with a higher incidence and greater extent of NI spreading to tumor-distant extratumoral nerves. In turn, more abundant extratumoral NI predicted shorter disease-free and overall survival. Experimentally, PlGF facilitated directional and dynamic changes in neurite outgrowth of primary dorsal-root-ganglia neurons upon exposure to PDAC derived guidance and growth factors and supported mutual chemoattraction of tumor cells with neurons and Schwann cells. CONCLUSION: Our translational results highlight PlGF as an axon guidance factor, which fosters neurite outgrowth and attracts tumor cells towards nerves. Hence, PlGF represents a promising circulating biomarker of NI and potential therapeutic target to improve the clinical outcome for patients with resectable PDAC.

Excessive endometrial PlGF- Rac1 signalling underlies endometrial cell stiffness linked to pre-eclampsia.

Cell stiffness is regulated by dynamic interaction between ras-related C3 botulinum toxin substrate 1 (Rac1) and p21 protein-activated kinase 1 (PAK1) proteins, besides other biochemical and molecular regulators. In this study, we investigated how the Placental Growth Factor (PlGF) changes endometrial mechanics by modifying the actin cytoskeleton at the maternal interface. We explored the global effects of PlGF in endometrial stromal cells (EnSCs) using the concerted approach of proteomics, atomic force microscopy (AFM), and electrical impedance spectroscopy (EIS). Proteomic analysis shows PlGF upregulated RhoGTPases activating proteins and extracellular matrix organization-associated proteins in EnSCs. Rac1 and PAK1 transcript levels, activity, and actin polymerization were significantly increased with PlGF treatment. AFM further revealed an increase in cell stiffness with PlGF treatment. The additive effect of PlGF on actin polymerization was suppressed with siRNA-mediated inhibition of Rac1, PAK1, and WAVE2. Interestingly, the increase in cell stiffness by PlGF treatment was pharmacologically reversed with pravastatin, resulting in improved trophoblast cell invasion. Taken together, aberrant PlGF levels in the endometrium can contribute to an altered pre-pregnancy maternal microenvironment and offer a unifying explanation for the pathological changes observed in conditions such as pre-eclampsia (PE).

Evaluation of salivary placental growth factor in Health and Periodontitis.

BACKGROUND: Various immune mediators have a role in the progression of periodontitis. Placental Growth Factor (PLGF) is important during pregnancy and also is involved in the pathology of several diseases. Hence, this study aimed to evaluate salivary PLGF in health and periodontitis that seemingly has not been reported earlier. METHODS: Fifty participants were grouped as healthy and periodontitis patients. Clinical history, periodontal parameters [Plaque Index (PI), Gingival Index (GI), probing pocket depth (PPD), clinical attachment loss (CAL), bleeding on probing (BoP)] were recorded; saliva was collected and PLGF was estimated using a commercially available ELISA kit. The data were statistically analyzed using Shapiro-Wilk's test, Kruskal-Wallis test, Dunn's post hoc test with Bonferroni correction, and Spearman's rank-order correlation coefficient. The significance level was set at p ≤ 0.05 for all tests. RESULTS: Salivary PLGF levels comparison between the two groups showed no significant difference between both groups. Quantitatively, females had higher salivary PLGF levels than males. No significant association was observed between salivary PLGF levels and the severity of periodontitis. The periodontitis group showed statistically significant correlations between salivary PLGF levels, BoP(p = 0.005) and PPD(p = 0.005), and significant correlations of PLGF with PPD (p = 0.035) for both groups. CONCLUSIONS: PLGF can be detected and measured in the saliva of healthy individuals and periodontitis patients. However, the role of PLGF in periodontal pathology needs to be further confirmed based on their salivary levels.

Ghrelin alleviates placental dysfunction by down-regulating NF-κB phosphorylation in LPS-induced rat model of preeclampsia.

In our previous study, we uncovered that ghrelin promotes angiogenesis in human umbilical vein endothelial cells (HUVECs) in vitro by activating the Jagged1/Notch2/VEGF pathway in preeclampsia (PE). However, the regulatory effects of ghrelin on placental dysfunction in PE are unclear. Therefore, we applied Normal pregnant Sprague-Dawley (SD) rats, treated with lipopolysaccharide (LPS), to establish a PE-like rat model. The hematoxylin-eosin (HE) staining method and immunohistochemistry (IHC) technology were used to detect morphological features of the placenta. IHC and Western blot were applied to examine Bax and Bcl-2 expression levels. The concentrations of serum soluble fms-like tyrosine kinase-1 (sFlt1) and placental growth factor (PIGF) were assessed by enzyme-linked immunosorbent assay (ELISA) kit. In addition, the apoptosis rates of JEG-3 and HTR-8/SVneo trophoblast cells were determined by Annexin V-FITC/PI apoptosis detection kit. Cell migratory capacities were assessed by scratch-wound assay, and RNA-sequencing assay was used to determine the mechanism of ghrelin in regulating trophoblast apoptosis. It has been found that ghrelin significantly reduced blood pressure, urinary protein, and urine creatinine in rats with PE, at the meanwhile, ameliorated placental and fetal injuries. Second, ghrelin clearly inhibited placental Bax expression and circulating sFlt-1 as well as elevated placental Bcl-2 expression and circulating PIGF, restored apoptosis and invasion deficiency of trophoblast cells caused by LPS in vitro. Finally, transcriptomics indicated that nuclear factor kappa B (NF-κB) was the potential downstream pathway of ghrelin. Our findings illustrated that ghrelin supplementation significantly improved LPS-induced PE-like symptoms and adverse pregnancy outcomes in rats by alleviating placental apoptosis and promoting trophoblast migration.

Status of alternative angiogenic pathways in glioblastoma resected under and after bevacizumab treatment.

Glioblastoma multiforme (GBM) acquires resistance to bevacizumab (Bev) treatment. Bev affects angiogenic factors other than vascular endothelial growth factor (VEGF), which are poorly understood. We investigated changes in angiogenic factors under and after Bev therapy, including angiopoietin-1 (ANGPT1), angiopoietin-2 (ANGPT2), placental growth factor (PLGF), fibroblast growth factor 2, and ephrin A2 (EphA2). Fifty-four GBM tissues, including 28 specimens from 14 cases as paired specimens from the same patient obtained in three settings: initial tumor resection (naïve Bev), tumors resected following Bev therapy (effective Bev), and recurrent tumors after Bev therapy (refractory Bev). Immunohistochemistry assessed their expressions in tumor vessels and its correlation with recurrent MRI patterns. PLGF expression was higher in the effective Bev group than in the naïve Bev group (p = 0.024) and remained high in the refractory Bev group. ANGPT2 and EphA2 expressions were higher in the refractory Bev group than in the naïve Bev group (p = 0.047 and 0.028, respectively). PLGF expression was higher in the refractory Bev group compared with the naïve Bev group for paired specimens (p = 0.036). PLGF was more abundant in T2 diffuse/circumscribe patterns (p = 0.046). This is the first study to evaluate angiogenic factors other than VEGF during effective and refractory Bev therapy in patient-derived specimens.

Eucommia granules activate Wnt/ß-catenin pathway, and improve oxidative stress, inflammation, and endothelial injury in preeclampsia rats.

PURPOSE: Pre-eclampsia (PE) is a pregnancy-related complication. Eucommia is effective in the treatment of hypertensive disorders in pregnancy, but the specific effects and possible mechanisms of Eucommia granules (EG) in PE remain unknown. The aim of this study was to investigate the effects and possible mechanisms of EG in PE rats. METHODS: Pregnant Sprague Dawley rats were divided into five groups (n = 6): the control group, the model group, the low-dose group, the medium-dose group, and the high-dose group of EG. The PE model was established by subcutaneous injection of levonitroarginine methyl ester. Saline was given to the blank and model groups, and the Eucommia granules were given by gavage to the remaining groups. Blood pressure and urinary protein were detected. The body length and weight of the pups and the weight of the placenta were recorded. Superoxide dismutase (SOD) activity and levels of malondialdehyde (MDA), placental growth factor (PIGF), and soluble vascular endothelial growth factor receptor-1 (sFIt-1) were measured in the placenta. Pathological changes were observed by hematoxylin-eosin staining. Wnt/ß-catenin pathway-related protein expression was detected using Western blot. RESULTS: Compared with the model group, the PE rats treated with EG had lower blood pressure and urinary protein. The length and weight of the pups and placental weight were increased. Inflammation and necrosis in the placental tissue was improved. SOD level increased, MDA content and sFIt-1/PIGF ratio decreased, and Wnt/ß-catenin pathway-related protein expression level increased. Moreover, the results of EG on PE rats increased with higher doses of EG. CONCLUSIONS: EG may activate the Wnt/ß-catenin pathway and inhibit oxidative stress, inflammation, and vascular endothelial injury in PE rats, thereby improving the perinatal prognosis of preeclamptic rats. EG may inhibit oxidative stress, inflammation, and vascular endothelial injury through activation of the Wnt/ß-catenin pathway in preeclampsia rats, thereby improving perinatal outcomes in PE rats.

Placenta-anchored tadalafil liposomes rescues intrauterine growth restriction through continuous placental blood perfusion improvement.

Physiological or pathological hypoperfusion of the placenta is one of the main causes of intrauterine growth restriction (IUGR) which poses a significant risk to the health of the fetus and newborn. Tadalafil, a 5-type phosphodiesterase inhibitor, has previously been found to improve the symptoms of IUGR in various clinical studies. Unfortunately, its clinical utility is hindered by its limited water solubility, rapid metabolism, and lack of specific distribution in target tissues rendering tadalafil unable to maintain long-term placental perfusion. In this study, iRGD-modified tadalafil-loaded liposomes (iRGD-lipo@Tad) featuring a size of approximately 480 nm were designed to rectify the shortcomings of tadalafil. The prepared iRGD-lipo@Tad exhibited superior stability, sustained drug release capacity, and low cytotoxicity. The fluorescence study, tissue slice study, and drug biodistribution study together demonstrated the placenta-anchored ability of iRGD-modified liposomes. This was achieved by a dual approach consisting of the iRGD-mediated placenta-targeting effect and special particle size-mediated placenta resident effect. The pharmacokinetic study revealed a significant improvement in the in vivo process of tadalafil encapsulated by the iRGD-modified liposomes. In comparison to the tadalafil solution, the peak plasma concentration of iRGD-lipo@Tad was significantly increased, and the area under the curve was increased by about 7.88 times. In the pharmacodynamic study, iRGD-lipo@Tad achieved a continuous and efficient improvement of placental blood perfusion. This was achieved by decreasing the ratio of plasma soluble fms-like tyrosine kinase to placental growth factor and increasing the levels of cyclic guanosine monophosphate and nitric oxide. Consequently, iRGD-lipo@Tad resulted in a significant increase in embryo weight and a reduction in the miscarriage rate of N-Nitro-L-arginine methyl ester-induced IUGR pregnant mice without detectable toxicity. In summary, the nanotechnology-assisted therapy strategy presented here not only overcomes the limitations of tadalafil in the clinical treatment of IUGR but also offers new avenues to address the treatment of other placenta-originated diseases.