Sympathetic Nervous System Reactivity in Women Following Preeclamptic Pregnancies

Women who have had preeclamptic pregnancies are at risk for life-long cardiovascular disease. However, the factors contributing to this risk have yet to be established. Sympathetic nervous system dysregulation has been proposed to contribute to cardiovascular dysfunction during preeclamptic pregnancies. Therefore, we examined muscle sympathetic nerve activity (MSNA) at baseline and during a chemoreflex stimulus in women 6-24 months postpartum following a preeclamptic pregnancy (PE; n=6, age 28±2 y, BMI 27±3 kg/m2, 17±4 months postpartum). We hypothesized that MSNA responses to apnea would be greater in PE relative to control subjects, that is, women 6-24 months following a healthy pregnancy and with no history of disordered pregnancies (HP; n=6, 31±6 y, BMI 29±5 kg/m2, 17±4 months postpartum). Integrated MSNA recordings were obtained at baseline and during a voluntary end-inspiratory apnea. Baseline mean arterial pressure (MAP; 87±10 vs 95±10 mmHg, P=0.2), total peripheral resistance (TPR; 13±3 vs 14±1 mmHg/L/min, P=0.4), and heart rate (HR; 74±5 vs 74±13, P=0.9) were similar in PE vs HP. Baseline MSNA was higher in PE compared to HP (26±9 vs 14±6 bursts/100heartbeats, P\u3c0.01). The voluntary apnea was maintained for a similar duration in PE and HP (44±17 and 45±10 sec, P=0.9), without any difference in mean MAP (93±14 and 99±11, P=0.4), TPR (14±4 and 14±2, P=0.6), or HR (74±8 and 81±22, P=0.5) between groups. To discern between mild and moderate phases of chemoreflex stress, the apnea was divided into initial (i.e. first half) and latter (i.e. second half) phases for subsequent analyses. The initial phase of the apnea elicited a large increase in MSNA in the PE women which exceeded that observed in HP (37±13 vs 19±11, P=0.03, respectively). The peak sympathetic response observed in the latter half of the apnea was similar between PE and HP (56±21 vs 49±13 bursts/100hb, P=0.5). Thus, the sympathetic nervous system response to a mild chemoreflex stimulus is exaggerated in women who have had preeclampsia within the past 6-24 months relative to women without a history of preeclampsia. We have demonstrated that a recent history of preeclampsia is associated with chronic sympathetic activation as well as greater sympathetic reactivity. We propose these changes to the sympathetic nervous system contribute to the life-long risk for cardiovascular disease in formerly preeclamptic women. Funded by the Paul Titus Fellowship, Department of Obstetrics, Gynecology and Reproductive Sciences, Yale School of Medicin

PreImplantation Factor (PIF) promoting role in embryo implantation: increases endometrial Integrin-α2β3, amphiregulin and epiregulin while reducing betacellulin expression via MAPK in decidua

BACKGROUND: Viable embryos secrete preimplantation factor (PIF), a peptide that has autocrine effects where levels correlate with cultured embryos development. sPIF (PIF synthetic analog) promotes implantation by regulating decidual-cells immunity, adhesion, apoptosis and enhances trophoblastic cell invasion. Herein sPIF priming effects on non-decidualized endometrium and decidualized-stroma are investigated, assessing elements critical for effective embryo-maternal cross-talk, prior to and at implantation. METHODS: We tested sPIF effect on human non-pregnant endometrial epithelial and non-decidualized stroma α2β3 integrin expression (IHC and flow cytometry), comparing with scrambled PIF (PIFscr-control). We examined sPIF effect on decidualized non-pregnant human endometrial stromal cells (HESC) determining pro-inflammatory mediators expression and secretion (ELISA) and growth factors (GFs) expression (Affymetrix global gene array). We tested sPIF effect on HESC Phospho-kinases (BioPlex) and isolated kinases activity (FastKinase). RESULTS: sPIF up-regulates α2β3 integrin expression in epithelial cells, (P < 0.05) while PIFscr had no effect. In contrast, in stromal cell cultures sPIF had no effect on the same. In HESC, sPIF up-regulates pro-inflammatory cytokines; IL8, IL1β and IL6 expression. The major increase in GRO-α, ICAM-1 and MCP-3 expression is coupled with same ligands secretion (P < 0.05). sPIF modulates in HESC GFs expression: up-regulates amphiregulin and epiregulin- critical for implantation and enhances several fibroblast growth factors (FGF) relevant for decidual function. In contrast, sPIF down-regulates major pro-proliferative ligands, betacellulin and IGF1 expression. sPIF modulatory effect on GFs is exerted by down-regulating pro-proliferative phospho-activated MAPkinases, p-MEK1 and p-ERK (P < 0.01, P < 0.04, respectively). Stress-induced p-38-MAPK (P = 0.04) and c-Jun kinase signaling involved MAPK8IP2 (−2.1 fold) expression decreased which protects against reactive oxygen species. Although pro-inflammatory p-NFkB (P = 0.06) decrease was mild, its promoter TNFRS11 expression markedly (−25-fold) decreased. In contrast, anti-proliferative phosphatases PTPRZ1 and PPP2R2C expression increased. CONCLUSIONS: sPIF post-fertilization primes endometrial-epithelium, while during implantation creates a beneficial pro-inflammatory milieu. PIF acts by balancing decidual pro-implantation properties while controlling excessive pro-proliferative and inflammatory signals expression. Overall, PIF influences critical peri-implantation events in a sequential coordinated fashion which facilitates embryo implantation

Multi-organ histopathological changes in a mouse hepatitis virus model of COVID-19

Infection with SARS-CoV-2, the virus responsible for the global COVID-19 pandemic, causes a respiratory illness that can severely impact other organ systems and is possibly precipitated by cytokine storm, septic shock, thrombosis, and oxidative stress. SARS-CoV-2 infected individuals may be asymptomatic or may experience mild, moderate, or severe symptoms with or without pneumonia. The mechanisms by which SARS-CoV-2 infects humans are largely unknown. Mouse hepatitis virus 1 (MHV-1)-induced infection was used as a highly relevant surrogate animal model for this study. We further characterized this animal model and compared it with SARS-CoV-2 infection in humans. MHV-1 inoculated mice displayed death as well as weight loss, as reported ear-lier. We showed that MHV-1-infected mice at days 7–8 exhibit severe lung inflammation, peribron-chiolar interstitial infiltration, bronchiolar epithelial cell necrosis and intra-alveolar necrotic debris, alveolar exudation (surrounding alveolar walls have capillaries that are dilated and filled with red blood cells), mononuclear cell infiltration, hyaline membrane formation, the presence of hemo-siderin-laden macrophages, and interstitial edema. When compared to uninfected mice, the infected mice showed severe liver vascular congestion, luminal thrombosis of portal and sinusoidal vessels, hepatocyte degeneration, cell necrosis, and hemorrhagic changes. Proximal and distal tubular ne-crosis, hemorrhage in interstitial tissue, and the vacuolation of renal tubules were observed. The heart showed severe interstitial edema, vascular congestion, and dilation, as well as red blood cell extravasation into the interstitium. Upon examination of the MHV-1 infected mice brain, we observed congested blood vessels, perivascular cavitation, cortical pericellular halos, vacuolation of neuropils, darkly stained nuclei, pyknotic nuclei, and associated vacuolation of the neuropil in the cortex, as well as acute eosinophilic necrosis and necrotic neurons with fragmented nuclei and vac-uolation in the hippocampus. Our findings suggest that the widespread thrombotic events observed in the surrogate animal model for SARS-CoV-2 mimic the reported findings in SARS-CoV-2 infected humans, representing a highly relevant and safe animal model for the study of the pathophysiologic mechanisms of SARS-CoV-2 for potential therapeutic interventions

PIF* promotes brain re-myelination locally while regulating systemic inflammation- clinically relevant multiple sclerosis M.smegmatis model.

Neurologic disease diagnosis and treatment is challenging. Multiple Sclerosis (MS) is a demyelinating autoimmune disease with few clinical forms and uncertain etiology. Current studies suggest that it is likely caused by infection(s) triggering a systemic immune response resulting in antigen/non-antigen-related autoimmune response in central nervous system (CNS). New therapeutic approaches are needed. Secreted by viable embryos, PreImplantation Factor (PIF) possesses a local and systemic immunity regulatory role. Synthetic PIF (PIF) duplicates endogenous peptide's protective effect in pre-clinical autoimmune and transplantation models. PIF protects against brain hypoxia-ischemia by directly targeting microglia and neurons. In chronic experimental autoimmune encephalitis (EAE) model PIF reverses paralysis while promoting neural repair. Herein we report that PIF directly promotes brain re-myelination and reverses paralysis in relapsing remitting EAE MS model. PIF crosses the blood-brain barrier targeting microglia. Systemically, PIF decreases pro-inflammatory IL23/IL17 cytokines, while preserving CNS-specific T-cell repertoire. Global brain gene analysis revealed that PIF regulates critical Na+/K+/Ca++ ions, amino acid and glucose transport genes expression. Further, PIF modulates oxidative stress, DNA methylation, cell cycle regulation, and protein ubiquitination while regulating multiple genes. In cultured astrocytes, PIF promotes BDNF-myelin synthesis promoter and SLC2A1 (glucose transport) while reducing deleterious E2F5, and HSP90ab1 (oxidative stress) genes expression. In cultured microglia, PIF increases anti-inflammatory IL10 while reducing pro-inflammatory IFNγ expression. Collectively, PIF promotes brain re-myelination and neuroprotection in relapsing remitting EAE MS model. Coupled with ongoing, Fast-Track FDA approved clinical trial, NCT#02239562 (immune disorder), current data supports PIF's translation for neurodegenerative disorders therapy

Preimplantation factor modulates oligodendrocytes by H19-induced demethylation of NCOR2.

Failed or altered gliogenesis is a major characteristic of diffuse white matter injury in survivors of premature birth. The developmentally regulated long noncoding RNA (lncRNA) H19 inhibits S-adenosylhomocysteine hydrolase (SAHH) and contributes to methylation of diverse cellular components, such as DNA, RNA, proteins, lipids, and neurotransmitters. We showed that the pregnancy-derived synthetic PreImplantation Factor (sPIF) induces expression of the nuclear receptor corepressor 2 (NCOR2) via H19/SAHH-mediated DNA demethylation. In turn, NCOR2 affects oligodendrocyte differentiation markers. Accordingly, after hypoxic-ischemic brain injury in rodents, myelin protection and oligodendrocytes' fate are in part modulated by sPIF and H19. Our results revealed an unexpected mechanism of the H19/SAHH axis underlying myelin preservation during brain recovery and its use in treating neurodegenerative diseases can be envisioned

Effects of Uterine Cells-Conditioned Media on Expression of DNMT3B and DNMT3C in Mouse Embryos Cultured in a Microfluidic Device.

Abstract presented at the 66th Annual Meeting of the Society for Reproductive Investigation, 12-16 Mar 2019, Paris, France

MicroRNA Analysis in Maternal Blood of Pregnancies with Preterm Premature Rupture of Membranes Reveals a Distinct Expression Profile

OBJECTIVE: To determine the expression profile of microRNAs in the peripheral blood of pregnant women with preterm premature rupture of membranes (PPROM) compared to that of healthy pregnant women. STUDY DESIGN: This was a pilot study with case-control design in pregnant patients enrolled between January 2017 and June 2019. Patients with healthy pregnancies and those affected by PPROM between 20- and 33+6 weeks of gestation were matched by gestational age and selected for inclusion to the study. Patients were excluded for multiple gestation and presence of a major obstetrical complication such as preeclampsia, diabetes, fetal growth restriction and stillbirth. A total of ten (n = 10) controls and ten (n = 10) patients with PPROM were enrolled in the study. Specimens were obtained before administration of betamethasone or intravenous antibiotics. MicroRNA expression was analyzed for 800 microRNAs in each sample using the NanoString nCounter Expression Assay. Differential expression was calculated after normalization and log2- transformation using the false discovery rate (FDR) method at an alpha level of 5%. RESULTS: Demographic characteristics were similar between the two groups. Of the 800 miRNAs analyzed, 116 were differentially expressed after normalization. However, only four reached FDR-adjusted statistical significance. Pregnancies affected by PPROM were characterized by upregulation of miR-199a-5p, miR-130a-3p and miR-26a-5p and downregulation of miR-513b-5p (FDR adjusted p-values CONCLUSION: Patients with PPROM have a distinct peripheral blood microRNA profile compared to healthy pregnancies as measured by the NanoString Expression Assay

Wharton's Jelly Mesenchymal Stem Cells Protect the Immature Brain in Rats and Modulate Cell Fate.

The development of a mammalian brain is a complex and long-lasting process. Not surprisingly, preterm birth is the leading cause of death in newborns and children. Advances in perinatal care reduced mortality, but morbidity still represents a major burden. New therapeutic approaches are thus desperately needed. Given that mesenchymal stem/stromal cells (MSCs) emerged as a promising candidate for cell therapy, we transplanted MSCs derived from the Wharton's Jelly (WJ-MSCs) to reduce the burden of immature brain injury in a murine animal model. WJ-MSCs transplantation resulted in protective activity characterized by reduced myelin loss and astroglial activation. WJ-MSCs improved locomotor behavior as well. To address the underlying mechanisms, we tested the key regulators of responses to DNA-damaging agents, such as cyclic AMP-dependent protein kinase/calcium-dependent protein kinase (PKA/PKC), cyclin-dependent kinase (CDK), ataxia-telangiectasia-mutated/ATM- and Rad3-related (ATM/ATR) substrates, protein kinase B (Akt), and 14-3-3 binding protein partners. We characterized WJ-MSCs using a specific profiler polymerase chain reaction array. We provide evidence that WJ-MSCs target pivotal regulators of the cell fate such as CDK/14-3-3/Akt signaling. We identified leukemia inhibitory factor as a potential candidate of WJ-MSCs' induced modifications as well. We hypothesize that WJ-MSCs may exert adaptive responses depending on the type of injury they are facing, making them prominent candidates for cell therapy in perinatal injuries

Evidence for/Against Administration of Fibrinogen Concentrate and Coagulation Factor Concentrate During an Obstetrical Hemorrhage

Correction of hypofibrinogenemia is increasingly being recognized as an important component of treatment of obstetric hemorrhage. Fibrinogen levels below 2 g/L are predictive of severe hemorrhage. Fibrinogen concentrate is an attractive means of fibrinogen replacement and has advantages over both fresh frozen plasma and cryoprecipitate. Data are beginning to emerge supporting the use of fibrinogen concentrate in the obstetric population, especially when guided by thromboelastometry/thromboelastography-driven protocols. Prevention of clot breakdown via use of antifibrinolytic therapy is an important component as well