Decreased vascular reactivity associated with increased IL-8 in 6-month-old infants of mothers with pre-eclampsia.

BACKGROUND: Offspring born to mothers with pre-eclampsia (Pre-E) suffer higher risks of adult cardiovascular diseases, suggesting that exposure to an antiangiogenic environment in-utero has a lasting impact on the development of endothelial function. The goal of this study is to test the hypothesis that in-utero exposure to Pre-E results in alterations of angiogenic factors/cytokines that negatively impact vascular development during infancy. METHODS: Infants born from mothers with and without Pre-E were recruited and followed up at 6 months. Plasma cytokines, blood pressure, microvessel density, and vascular reactivity were assessed. RESULTS: 6-month-old infants born to mothers with Pre-E had unchanged blood pressure (p = 0.86) and microvessel density (p = 0.57). Vascular reactivity was decreased in infants born to mothers with Pre-E compared to infants born to healthy mothers (p = 0.0345). Interleukin 8 (IL-8) (p = 0.03) and Angiopoeitin-2 (Ang-2) (p = 0.04) were increased in infants born to mothers with Pre-E. We observed that higher IL-8 was associated with lower vascular reactivity (rho = -0.14, p < 0.0001). CONCLUSION: At 6 months of age, infants born to mothers with Pre-E had impaired vascular reactivity and higher IL-8 and Ang-2, but similar blood pressure and microvessel density compared to infants born to non-Pre-E mothers. IMPACT STATEMENT: Changes in cord blood antiangiogenic factors are documented in infants of mothers with pre-eclampsia and may contribute to offspring risks of adult cardiovascular disease. How these factors evolve during early infancy and their correlation with offspring vascular development have not been studied. This study found that 6-month-old infants born to mothers with pre-eclampsia had decreased vascular reactivity, which was correlated with higher IL-8. These findings underscore the lasting impact of maternal pre-eclampsia on offspring vascular development and highlight the need for long-term follow-up in children born to mothers with pre-eclampsia.

Long-term childhood outcomes for babies born at term who were exposed to antenatal corticosteroids.

BACKGROUND: Antenatal corticosteroids improve neonatal outcomes when administered to infants who are at risk of preterm delivery. Many women who receive antenatal corticosteroids for threatened preterm labor proceed to deliver at term. Thus, long-term outcomes should be evaluated for term-born infants who were exposed to antenatal corticosteroids in utero. OBJECTIVE: This study aimed to compare long-term outcomes between term-born children aged ≥5 years who were born to women who received antenatal corticosteroids for threatened preterm labor and children whose mothers were also evaluated for threatened preterm labor but did not receive antenatal corticosteroids. STUDY DESIGN: We performed a retrospective cohort study of children born at ≥37 weeks' gestation, aged ≥5 years, and born to mothers diagnosed with threatened preterm labor during pregnancy. The primary exposure of interest was receiving antenatal corticosteroids. Among the collected childhood medical conditions, the primary outcome of interest was a diagnosis of asthma. RESULTS: Of the 3556 term-born children aged ≥5 years, 629 (17.6%) were exposed to antenatal corticosteroids (all betamethasone), and 2927 (82.3%) were controls whose mothers were evaluated for threatened preterm birth but did not get antenatal corticosteroid injections. Women receiving antenatal corticosteroids had higher rates of maternal comorbidities (diabetes mellitus, hypertension; P≤.01). Antenatal corticosteroid-exposed children had no difference in diagnosis of asthma (12.6% vs 11.6%), attention deficit disorder, or developmental delay (P=.47, .54, and .10, respectively). Controlling for maternal and neonatal characteristics, asthma was not different between those exposed to antenatal corticosteroids and controls (odds ratio, 1.05; 95% confidence interval, 0.79-1.39). The odds of the child's weight percentile being <10% were increased for antenatal corticosteroid-exposed children born at term (odds ratio, 2.00; 95% confidence interval, 1.22-3.25). CONCLUSION: Children born at term who were exposed to antenatal corticosteroids may have increased odds of being in a lower growth percentile than those not exposed. However, rates of diagnoses such as asthma, developmental delay, and attention deficit disorders were not different.

Inhibition of BTK and PI3Kδ impairs the development of human JMML stem and progenitor cells.

Juvenile myelomonocytic leukemia (JMML) is an aggressive myeloproliferative neoplasia that lacks effective targeted chemotherapies. Clinically, JMML manifests as monocytic leukocytosis, splenomegaly with consequential thrombocytopenia. Most commonly, patients have gain-of-function (GOF) oncogenic mutations in PTPN11 (SHP2), leading to Erk and Akt hyperactivation. Mechanism(s) involved in co-regulation of Erk and Akt in the context of GOF SHP2 are poorly understood. Here, we show that Bruton's tyrosine kinase (BTK) is hyperphosphorylated in GOF Shp2-bearing cells and utilizes B cell adaptor for PI3K to cooperate with p110δ, the catalytic subunit of PI3K. Dual inhibition of BTK and p110δ reduces the activation of both Erk and Akt. In vivo, individual targeting of BTK or p110δ in a mouse model of human JMML equally reduces monocytosis and splenomegaly; however, the combined treatment results in a more robust inhibition and uniquely rescues anemia and thrombocytopenia. RNA-seq analysis of drug-treated mice showed a profound reduction in the expression of genes associated with leukemic cell migration and inflammation, leading to correction in the infiltration of leukemic cells in the lung, liver, and spleen. Remarkably, in a patient derived xenograft model of JMML, leukemia-initiating stem and progenitor cells were potently inhibited in response to the dual drug treatment.

Are newborn outcomes different for term babies who were exposed to antenatal corticosteroids?

BACKGROUND: Antenatal corticosteroids improve newborn outcomes for preterm infants. However, predicting which women presenting for threatened preterm labor will have preterm infants is inaccurate, and many women receive antenatal corticosteroids but then go on to deliver at term. OBJECTIVE: This study aimed to compare the short-term outcomes of infants born at term to women who received betamethasone for threatened preterm labor with infants who were not exposed to betamethasone in utero. STUDY DESIGN: We performed a retrospective cohort study of infants born at or after 37 weeks' gestational age to mothers diagnosed as having threatened preterm labor during pregnancy. The primary neonatal outcomes of interest included transient tachypnea of the newborn, neonatal intensive care unit admission, and small for gestational age and were evaluated for their association with betamethasone exposure while adjusting for covariates using multiple logistic regression. RESULTS: Of 5330 women, 1459 women (27.5%) received betamethasone at a mean gestational age of 32.2±3.3 weeks. The mean age of women was 27±5.9 years and the mean gestational age at delivery was 38.9±1.1 weeks. Women receiving betamethasone had higher rates of maternal comorbidities (P<.001 for diabetes mellitus, asthma, and hypertensive disorder) and were more likely to self-identify as White (P=.022). Betamethasone-exposed neonates had increased rates of transient tachypnea of the newborn, neonatal intensive care unit admission, small for gestational age, hyperbilirubinemia, and hypoglycemia (all, P<.05). Controlling for maternal characteristics and gestational age at delivery, betamethasone exposure was not associated with a diagnosis of transient tachypnea of the newborn (adjusted odds ratio, 1.10; 95% confidence interval, 0.80-1.51), although it was associated with more neonatal intensive care unit admissions (adjusted odds ratio, 1.49; 95% confidence interval, 1.19-1.86) and higher odds of the baby being small for gestational age (adjusted odds ratio, 1.78; 95% confidence interval, 1.48-2.14). CONCLUSION: Compared with women evaluated for preterm labor who did not receive betamethasone, women receiving betamethasone had infants with higher rates of neonatal intensive care unit admission and small for gestational age. Although the benefits of betamethasone to infants born preterm are clear, there may be negative impacts for infants delivered at term.

Stereoselective Analysis of Methadone and EDDP in Laboring Women and Neonates in Plasma and Dried Blood Spots and Association with Neonatal Abstinence Syndrome.

OBJECTIVE: This pilot study evaluated the relationship between maternal and neonatal R- and S-methadone and R- and S-2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) exposure and the severity of neonatal abstinence syndrome (NAS). The use of dried blood spots (DBS) as an alternative for plasma in assessing methadone and EDDP was also assessed. STUDY DESIGN: Women receiving methadone for medication assisted treatment of opioid use disorder during pregnancy were eligible for recruitment. Plasma and DBS samples were collected from mothers during labor, from cord blood, and from newborns during genetic screen. R-/S-methadone and EDDP were measured by high-performance liquid chromatography tandem mass spectrometry (HPLC/MS/MS). Associations between methadone exposure, neonatal morphine requirements, and severity of NAS were examined. RESULTS: Twenty women and infants completed the study. Maternal methadone dose at delivery was 112 mg/day (range = 60-180 mg/day). Sixteen neonates experienced NAS requiring morphine; three also required phenobarbital. Higher cord blood concentrations of R-methadone, R- and S-EDDP were associated with higher maximum doses of morphine (p < 0.05). CONCLUSION: Maternal methadone and cord blood concentration at delivery are variable and may be potential markers of neonatal abstinence syndrome.

Effects of Pregnancy on the Pharmacokinetics of Metformin.

This study's primary objective was to fully characterize the pharmacokinetics of metformin in pregnant women with gestational diabetes mellitus (GDM) versus nonpregnant controls. Steady-state oral metformin pharmacokinetics in pregnant women with GDM receiving either metformin monotherapy (n = 24) or a combination with glyburide (n = 30) as well as in nonpregnant women with type 2 diabetes mellitus (T2DM) (n = 24) were determined utilizing noncompartmental techniques. Maternal and umbilical cord blood samples were collected at delivery from 38 women. With both 500- and 1000-mg doses, metformin bioavailability, volume of distribution beta (V ß ), clearance, and renal clearance were significantly increased during pregnancy. In addition, in the women receiving metformin 500 mg, significantly higher metformin apparent oral clearance (CL/F) (27%), weight-adjusted renal secretion clearance (64%), and apparent oral volume of distribution beta (V ß /F) (33%) were seen during pregnancy. Creatinine clearance was significantly higher during pregnancy. Increasing metformin dose from 500 to 1000 mg orally twice daily significantly increased V ß /F by 28%, weight-adjusted V ß /F by 32% and CL/F by 25%, and weight-adjusted CL/F by 28% during pregnancy. Mean metformin umbilical cord arterial-to-venous plasma concentration ratio was 1.0 ± 0.1, venous umbilical cord-to-maternal concentration ratio was 1.4 ± 0.5, and arterial umbilical cord-to-maternal concentration ratio was 1.5 ± 0.5. Systemic exposure after a 500-mg dose of metformin was lower during pregnancy compared with the nonpregnant women with T2DM. However, in patients receiving metformin 1000 mg, changes in estimated bioavailability during pregnancy offset the changes in clearance leading to no significant change in CL/F with the higher dose. SIGNIFICANCE STATEMENT: Gestational diabetes mellitus complicates 5%-13% of pregnancies and is often treated with metformin. Pregnant women undergo physiological changes that alter drug disposition. Preliminary data suggest that pregnancy lowers metformin concentrations, potentially affecting efficacy and safety. This study definitively describes pregnancy's effects on metformin pharmacokinetics and expands the mechanistic understanding of pharmacokinetic changes across the dosage range. Here we report the nonlinearity of metformin pharmacokinetics and the increase in bioavailability, clearance, renal clearance, and volume of distribution during pregnancy.

Endothelial Colony-Forming Cell Function Is Reduced During HIV Infection.

BACKGROUND: Human immunodeficiency virus (HIV) may be related to cardiovascular disease through monocyte activation-associated endothelial dysfunction. METHODS: Blood samples from 15 HIV-negative participants (the uninfected group), 8 HIV-positive participants who were not receiving antiretroviral therapy (ART) (the infected, untreated group), and 15 HIV-positive participants who were receiving ART (the infected, treated group) underwent flow cytometry of endothelial colony-forming cells (ECFCs) and monocyte proportions. IncuCyte live cell imaging of 8 capillary proliferative capacity parameters were obtained from cord blood ECFCs treated with participant plasma. RESULTS: The ECFC percentage determined by flow cytometry was not different between the study groups; however, values of the majority of capillary proliferative capacity parameters (ie, cell area, network length, network branch points, number of networks, and average tube width uniformity) were significantly lower in infected, untreated participants as compared to values for uninfected participants or infected, treated participants (P < .00625 for all comparisons). CD14+CD16+ intermediate monocytes and soluble CD163 were significantly and negatively correlated with several plasma-treated, cord blood ECFC proliferative capacity parameters in the combined HIV-positive groups but not in the uninfected group. CONCLUSIONS: Cord blood ECFC proliferative capacity was significantly impaired by plasma from infected, untreated patients, compared with plasma from uninfected participants and from infected, treated participants. Several ECFC functional parameters were adversely associated with monocyte activation in the HIV-positive groups, thereby suggesting a mechanism by which HIV-related inflammation may impair vascular reparative potential and consequently increase the risk of cardiovascular disease during HIV infection.

Transgelin induces dysfunction of fetal endothelial colony-forming cells from gestational diabetic pregnancies.

Fetal exposure to gestational diabetes mellitus (GDM) predisposes children to future health complications including hypertension and cardiovascular disease. A key mechanism by which these complications occur is through the functional impairment of vascular progenitor cells, including endothelial colony-forming cells (ECFCs). Previously, we showed that fetal ECFCs exposed to GDM have decreased vasculogenic potential and altered gene expression. In this study, we evaluate whether transgelin (TAGLN), which is increased in GDM-exposed ECFCs, contributes to vasculogenic dysfunction. TAGLN is an actin-binding protein involved in the regulation of cytoskeletal rearrangement. We hypothesized that increased TAGLN expression in GDM-exposed fetal ECFCs decreases network formation by impairing cytoskeletal rearrangement resulting in reduced cell migration. To determine if TAGLN is required and/or sufficient to impair ECFC network formation, TAGLN was reduced and overexpressed in ECFCs from GDM and uncomplicated pregnancies, respectively. Decreasing TAGLN expression in GDM-exposed ECFCs improved network formation and stability as well as increased migration. In contrast, overexpressing TAGLN in ECFCs from uncomplicated pregnancies decreased network formation, network stability, migration, and alignment to laminar flow. Overall, these data suggest that increased TAGLN likely contributes to the vasculogenic dysfunction observed in GDM-exposed ECFCs, as it impairs ECFC migration, cell alignment, and network formation. Identifying the molecular mechanisms underlying fetal ECFC dysfunction following GDM exposure is key to ascertain mechanistically the basis for cardiovascular disease predisposition later in life.

Impact of Pregnancy History and 17-Hydroxyprogesterone Caproate on Cervical Cytokines and Matrix Metalloproteinases.

OBJECTIVE: The objective of this study was to evaluate the impact of pregnancy history and 17-hydroxyprogesterone caproate (17-OHPC) treatment on cervical fluid cytokines and matrix metalloproteinases (MMPs). STUDY DESIGN: Cervical fluid was obtained between 160/7 and 246/7 weeks from women with only prior term births (controls, n = 26), women with one or more prior spontaneous preterm births (SPTBs) choosing to receive 17-OHPC (17-OHPC, n = 24), or to not receive 17-OHPC (refusers, n = 12). Cervical fluid collections were repeated 2, 4, and 8 weeks after the first sample and concentrations of MMPs and cytokines were measured by multiplex immune assay. RESULTS: Among women whose earliest prior delivery occurred between 16 and 23 weeks, cervical fluid concentration of interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α at baseline were significantly elevated when compared with cervical cytokines of women whose earliest delivery occurred between 32 and 36 weeks (relative risk ratio was 3.37 for IL-6 [95% confidence interval, CI, 1.08-10.53, p < 0.05], 2.81 for IL-10 [95% CI, 1.39-5.70, p < 0.05], and 6.34 for TNF-α [95% CI, 2.19-18.68, p < 0.001]). Treatment with 17-OHPC had no significant impact on these cytokines. CONCLUSION: The cervical fluid of women with a history of an early prior SPTB is characterized by inflammation that is unaffected by 17-OHPC.

Kinetic analyses of vasculogenesis inform mechanistic studies.

Vasculogenesis is a complex process by which endothelial stem and progenitor cells undergo de novo vessel formation. Quantitative assessment of vasculogenesis is a central readout of endothelial progenitor cell functionality. However, current assays lack kinetic measurements. To address this issue, new approaches were developed to quantitatively assess in vitro endothelial colony-forming cell (ECFC) network formation in real time. Eight parameters of network structure were quantified using novel Kinetic Analysis of Vasculogenesis (KAV) software. KAV assessment of structure complexity identified two phases of network formation. This observation guided the development of additional vasculogenic readouts. A tissue cytometry approach was established to quantify the frequency and localization of dividing ECFCs. Additionally, Fiji TrackMate was used to quantify ECFC displacement and speed at the single-cell level during network formation. These novel approaches were then implemented to identify how intrauterine exposure to maternal diabetes mellitus (DM) impairs fetal ECFC vasculogenesis. Fetal ECFCs exposed to maternal DM form fewer initial network structures, which are not stable over time. Correlation analyses demonstrated that ECFC samples with greater division in branches form fewer closed network structures. Additionally, reductions in average ECFC movement over time decrease structural connectivity. Identification of these novel phenotypes utilizing the newly established methodologies provides evidence for the cellular mechanisms contributing to aberrant ECFC vasculogenesis.

Mesenchyme Homeobox 2 Enhances Migration of Endothelial Colony Forming Cells Exposed to Intrauterine Diabetes Mellitus.

Diabetes mellitus (DM) during pregnancy has long-lasting implications for the fetus, including cardiovascular morbidity. Previously, we showed that endothelial colony forming cells (ECFCs) from DM human pregnancies have decreased vasculogenic potential. Here, we evaluate whether the molecular mechanism responsible for this phenotype involves the transcription factor, Mesenchyme Homeobox 2 (MEOX2). In human umbilical vein endothelial cells, MEOX2 upregulates cyclin-dependent kinase inhibitor expression, resulting in increased senescence and decreased proliferation. We hypothesized that dysregulated MEOX2 expression in neonatal ECFCs from DM pregnancies decreases network formation through increased senescence and altered cell cycle progression. Our studies show that nuclear MEOX2 is increased in ECFCs from DM pregnancies. To determine if MEOX2 is sufficient and/or required to induce impaired network formation, MEOX2 was overexpressed and depleted in ECFCs from control and DM pregnancies, respectively. Surprisingly, MEOX2 overexpression in control ECFCs resulted in increased network formation, altered cell cycle progression, and increased senescence. In contrast, MEOX2 knockdown in ECFCs from DM pregnancies led to decreased network formation, while cell cycle progression and senescence were unaffected. Importantly, migration studies demonstrated that MEOX2 overexpression increased migration, while MEOX2 knockdown decreased migration. Taken together, these data suggest that altered migration may be mediating the impaired vasculogenesis of ECFCs from DM pregnancies. While initially believed to be maladaptive, these data suggest that MEOX2 may serve a protective role, enabling increased vessel formation despite exposure to a DM intrauterine environment. J. Cell. Physiol. 232: 1885-1892, 2017. © 2016 Wiley Periodicals, Inc.

Safety and pharmacokinetics of pravastatin used for the prevention of preeclampsia in high-risk pregnant women: a pilot randomized controlled trial.

BACKGROUND: Preeclampsia complicates approximately 3-5% of pregnancies and remains a major cause of maternal and neonatal morbidity and mortality. It shares pathogenic similarities with adult cardiovascular disease as well as many risk factors. Pravastatin, a hydrophilic, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitor, has been shown in preclinical studies to reverse various pathophysiological pathways associated with preeclampsia, providing biological plausibility for its use for preeclampsia prevention. However, human trials are lacking. OBJECTIVE: As an initial step in evaluating the utility of pravastatin in preventing preeclampsia and after consultation with the US Food and Drug Administration, we undertook a pilot randomized controlled trial with the objective to determine pravastatin safety and pharmacokinetic parameters when used in pregnant women at high risk of preeclampsia. STUDY DESIGN: We conducted a pilot, multicenter, double-blind, placebo-controlled, randomized trial of women with singleton, nonanomalous pregnancies at high risk for preeclampsia. Women between 12(0/7) and 16(6/7) weeks' gestation were assigned to daily pravastatin 10 mg or placebo orally until delivery. Primary outcomes were maternal-fetal safety and pharmacokinetic parameters of pravastatin during pregnancy. Secondary outcomes included rates of preeclampsia and preterm delivery, gestational age at delivery, birthweight, and maternal and cord blood lipid profile (clinicaltrials.gov identifier NCT01717586). RESULTS: Ten women assigned to pravastatin and 10 to placebo completed the trial. There were no differences between the 2 groups in rates of study drug side effects, congenital anomalies, or other adverse or serious adverse events. There was no maternal, fetal, or neonatal death. Pravastatin renal clearance was significantly higher in pregnancy compared with postpartum. Four subjects in the placebo group developed preeclampsia compared with none in the pravastatin group. Although pravastatin reduced maternal cholesterol concentrations, umbilical cord cholesterol concentrations and infant birthweight were not different between the groups. The majority of umbilical cord and maternal pravastatin plasma concentrations at the time of delivery were below the lower limit of quantification of the assay. Pravastatin use was associated with a more favorable pregnancy angiogenic profile. CONCLUSION: This study provides preliminary safety and pharmacokinetic data regarding the use of pravastatin for preventing preeclampsia in high-risk pregnant women. Although the data are preliminary, no identifiable safety risks were associated with pravastatin use in this cohort. This favorable risk-benefit analysis justifies using pravastatin in a larger clinical trial with dose escalation.

Biomarkers for Diagnosis and Prognosis of Sinusoidal Obstruction Syndrome after Hematopoietic Cell Transplantation.

Reliable, noninvasive methods for diagnosing and prognosing sinusoidal obstruction syndrome (SOS) early after hematopoietic cell transplantation (HCT) are needed. We used a quantitative mass spectrometry-based proteomics approach to identify candidate biomarkers of SOS by comparing plasma pooled from 20 patients with and 20 patients without SOS. Of 494 proteins quantified, we selected 6 proteins (L-Ficolin, vascular cell adhesion molecule-1 [VCAM1], tissue inhibitor of metalloproteinase-1, von Willebrand factor, intercellular adhesion molecule-1, and CD97) based on a differential heavy/light isotope ratio of at least 2 fold, information from the literature, and immunoassay availability. Next, we evaluated the diagnostic potential of these 6 proteins and 5 selected from the literature (suppression of tumorigenicity-2 [ST2], angiopoietin-2 (ANG2), hyaluronic acid [HA], thrombomodulin, and plasminogen activator inhibitor-1) in samples from 80 patients. The results demonstrate that together ST2, ANG2, L-Ficolin, HA, and VCAM1 compose a biomarker panel for diagnosis of SOS. L-Ficolin, HA, and VCAM1 also stratified patients at risk for SOS as early as the day of HCT. Prognostic Bayesian modeling for SOS onset based on L-Ficolin, HA, and VCAM1 levels on the day of HCT and clinical characteristics showed >80% correct prognosis of SOS onset. These biomarkers may provide opportunities for preemptive intervention to minimize SOS incidence and/or severity.

Fetal hyperglycemia and a high-fat diet contribute to aberrant glucose tolerance and hematopoiesis in adult rats.

BACKGROUND: Children exposed to gestational diabetes mellitus (GDM) during pregnancy are at increased risk of obesity, diabetes, and hypertension. Our goal was to identify metabolic and hematopoietic alterations after intrauterine exposure to maternal hyperglycemia that may contribute to the pathogenesis of chronic morbidities. METHODS: Streptozotocin treatment induced maternal hyperglycemia during the last third of gestation in rat dams. Offspring of control mothers (OCM) and diabetic mothers (ODM) were evaluated for weight, glucose tolerance, insulin tolerance, and hematopoiesis defects. The effects of aging were examined in normal and high-fat diet (HFD)-fed young (8-wk-old) and aged (11-mo-old) OCM and ODM rats. RESULTS: Young adult ODM males on a normal diet, but not females, displayed improved glucose tolerance due to increased insulin levels. Aged ODM males and females gained more weight than OCM on a HFD and had worse glucose tolerance. Aged ODM males fed a HFD were also neutrophilic. Increases in bone marrow cellularity and myeloid progenitors preceded neutrophilia in ODM males fed a HFD. CONCLUSION: When combined with other risk factors like HFD and aging, changes in glucose metabolism and hematopoiesis may contribute to the increased risk of obesity, type 2 diabetes, and hypertension observed in children of GDM mothers.

Dishevelled-associated activator of morphogenesis 1 (Daam1) is required for heart morphogenesis.

Dishevelled-associated activator of morphogenesis 1 (Daam1), a member of the formin protein family, plays an important role in regulating the actin cytoskeleton via mediation of linear actin assembly. Previous functional studies of Daam1 in lower species suggest its essential role in Drosophila trachea formation and Xenopus gastrulation. However, its in vivo physiological function in mammalian systems is largely unknown. We have generated Daam1-deficient mice via gene-trap technology and found that Daam1 is highly expressed in developing murine organs, including the heart. Daam1-deficient mice exhibit embryonic and neonatal lethality and suffer multiple cardiac defects, including ventricular noncompaction, double outlet right ventricles and ventricular septal defects. In vivo genetic rescue experiments further confirm that the lethality of Daam1-deficient mice results from the inherent cardiac abnormalities. In-depth analyses have revealed that Daam1 is important for regulating filamentous actin assembly and organization, and consequently for cytoskeletal function in cardiomyocytes, which contributes to proper heart morphogenesis. Daam1 is also found to be important for proper cytoskeletal architecture and functionalities in embryonic fibroblasts. Biochemical analyses indicate that Daam1 does not regulate cytoskeletal organization through RhoA, Rac1 or Cdc42. Our study highlights a crucial role for Daam1 in regulating the actin cytoskeleton and tissue morphogenesis.

Membrane and capillary components of lung diffusion and pro-angiogenic cells in infants.

Angiogenesis is a critical determinant of alveolarisation, which increases alveolar surface area and pulmonary capillary blood volume in infants; however, our understanding of this process is very limited. The purpose of our study was to measure the pulmonary membrane diffusion capacity (DM) and pulmonary capillary blood volume (VC) components of the diffusing capacity of the lung for carbon monoxide (DLCO) in healthy infants and toddlers, and evaluate whether these components were associated with pro-angiogenic circulating haematopoietic stem/progenitor cells (pCHSPCs) early in life. 21 healthy subjects (11 males), 3-25 months of age, were evaluated. DLCO was measured under normoxic and hyperoxic conditions, and DM and VC were calculated. From 1 mL venous blood, pCHSPCs were quantified by multiparametric flow cytometry. DM and VC increased with increasing body length; however, membrane resistance as a fraction of total resistance to pulmonary diffusion remained constant with somatic size. In addition, DLCO and VC, but not DM, increased with an increasing percentage of pCHSPCs. The parallel increase in the membrane and vascular components of pulmonary diffusion is consistent with alveolarisation during this period of rapid lung growth. In addition, the relationship between pCHSPCs and VC suggest that pro-angiogenic cells may contribute to this vascular process.

Gestational diabetes induces alterations in the function of neonatal endothelial colony-forming cells.

BACKGROUND: Children born to mothers with gestational diabetes mellitus (GDM) experience increased risk of developing hypertension, type 2 diabetes mellitus, and obesity. Disrupted function of endothelial colony-forming cells (ECFCs) may contribute to this enhanced risk. The goal of this study was to determine whether cord blood ECFCs from GDM pregnancies exhibit altered functionality. METHODS: ECFCs isolated from the cord blood of control and GDM pregnancies were assessed for proliferation, senescence, and Matrigel network formation. The requirement for p38MAPK in hyperglycemia-induced senescence was determined using inhibition and overexpression studies. RESULTS: GDM-exposed ECFCs were more proliferative than control ECFCs. However, GDM-exposed ECFCs exhibited decreased network-forming ability in Matrigel. Aging of ECFCs by serial passaging led to increased senescence and reduced proliferation of GDM-exposed ECFCs. ECFCs from GDM pregnancies were resistant to hyperglycemia-induced senescence compared with those from controls. In response to hyperglycemia, control ECFCs activated p38MAPK, which was required for hyperglycemia-induced senescence. In contrast, GDM-exposed ECFCs showed no change in p38MAPK activation under equivalent conditions. CONCLUSION: Intrauterine exposure of ECFCs to GDM induces unique phenotypic alterations. The resistance of GDM-exposed ECFCs to hyperglycemia-induced senescence and decreased p38MAPK activation suggest that these progenitor cells have undergone changes that induce tolerance to a hyperglycemic environment.

Glucose intolerance as a consequence of hematopoietic stem cell dysfunction in offspring of obese mice.

OBJECTIVE: Maternal obesity is increasingly common and negatively impacts offspring health. Children of mothers with obesity are at higher risk of developing diseases linked to hematopoietic system abnormalities and metabolism such as type 2 diabetes. Interestingly, disease risks are often dependent on the offspring's sex, suggesting sex-specific reprogramming effect of maternal obesity on offspring hematopoietic stem and progenitor cell (HSPC) function. However, the impact of maternal obesity exposure on offspring HSPC function, and the capability of HSPC to regulate offspring metabolic health is largely understudied. This study aims to test the hypothesis that offspring of obese mice exhibit sex-differences in HSPC function that affect offspring's metabolic health. METHODS: We first assessed bone marrow hematopoietic stem and progenitor cell phenotype using postnatal day 21 (P21) and 8-week-old C57BL/6J mice born to control and diet-induced obese dams. We also sorted HSPC (Lineage-, Sca1+, cKit + cells) from P21 mice for competitive primary and secondary transplant, as well as transcriptomic analysis. Body weight, adiposity, insulin tolerance test and glucose tolerance tests were performed in primary and secondary transplant recipient animals. RESULTS: We discovered sex-differences in offspring HSPC function in response to maternal obesity exposure, where male offspring of obese dams (MatOb) showed decreased HSPC numbers and engraftment, while female MatOb offspring remained largely unaffected. RNA-seq revealed immune stimulatory pathways in female MatOb offspring. Finally, only recipients of male MatOb offspring HSPC exhibited glucose intolerance. CONCLUSIONS: This study demonstrated the lasting effect of maternal obesity exposure on offspring HSPC function and implicates HSPC in metabolic regulation.

Alkynyl nicotinamides show antileukemic activity in drug-resistant acute myeloid leukemia.

Activating mutations of FLT3 contribute to deregulated hematopoietic stem and progenitor cell (HSC/Ps) growth and survival in patients with acute myeloid leukemia (AML), leading to poor overall survival. AML patients treated with investigational drugs targeting mutant FLT3, including Quizartinib and Crenolanib, develop resistance to these drugs. Development of resistance is largely due to acquisition of cooccurring mutations and activation of additional survival pathways, as well as emergence of additional FLT3 mutations. Despite the high prevalence of FLT3 mutations and their clinical significance in AML, there are few targeted therapeutic options available. We have identified 2 novel nicotinamide-based FLT3 inhibitors (HSN608 and HSN748) that target FLT3 mutations at subnanomolar concentrations and are potently effective against drug-resistant secondary mutations of FLT3. These compounds show antileukemic activity against FLT3ITD in drug-resistant AML, relapsed/refractory AML, and in AML bearing a combination of epigenetic mutations of TET2 along with FLT3ITD. We demonstrate that HSN748 outperformed the FDA-approved FLT3 inhibitor Gilteritinib in terms of inhibitory activity against FLT3ITD in vivo.

Lack of plakoglobin in epidermis leads to keratoderma.

Loss-of-function mutation of Jup has been associated with Naxos disease, which is characterized by arrhythmogenic cardiomyopathy and the cutaneous disorder palmoplantar keratoderma. Previously, we have shown that genetic ablation of Jup in cardiomyocytes in mice leads to arrhythmogenic cardiomyopathy similar to Naxos disease in humans. Currently, to determine the pathogenesis of Naxos disease-associated keratoderma, we generated Jup mutant mice by inactivating Jup restrictively in keratinocytes. Jup mutant mice largely recapitulated the clinical features of human palmoplantar keratoderma: overcornification and thickening of the epidermis. Jup mutant mice also suffered skin ulceration and inflammation. Cell apoptosis and proliferation were significantly elevated in Jup mutant epidermis. Ultrastructural analyses revealed the disruption of the assembly of desmosomes and adherens junctions in Jup mutant epidermis. We also demonstrated the compensational increase in ß-catenin at Jup mutant cell-cell junctions without altering its signaling activities. Our findings provide important insights for understanding the pathogenesis of human palmoplantar keratoderma.