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Rationale: Bronchopulmonary dysplasia, a chronic respiratory condition originating from preterm birth, is associated with abnormal neurodevelopment. Currently, there is an absence of effective therapies for bronchopulmonary dysplasia and its associated brain injury. In preclinical trials, mesenchymal stromal cell therapies demonstrate promise as a therapeutic alternative for bronchopulmonary dysplasia. Objectives: To investigate whether a multifactorial neonatal mouse model of lung injury perturbs neural progenitor cell function and to assess the ability of human umbilical cord-derived mesenchymal stromal cell extracellular vesicles to mitigate pulmonary and neurologic injury. Methods: Mice at Postnatal Day 7 or 8 were injected intraperitoneally with LPS and ventilated with 40% oxygen at Postnatal Day 9 or 10 for 8 hours. Treated animals received umbilical cord-mesenchymal stromal cell-derived extracellular vesicles intratracheally preceding ventilation. Lung morphology, vascularity, and inflammation were quantified. Neural progenitor cells were isolated from the subventricular zone and hippocampus and assessed for self-renewal, in vitro differentiation ability, and transcriptional profiles. Measurements and Main Results: The multifactorial lung injury model produced alveolar and vascular rarefaction mimicking bronchopulmonary dysplasia. Neural progenitor cells from lung injury mice showed reduced neurosphere and oligodendrocyte formation, as well as inflammatory transcriptional signatures. Mice treated with mesenchymal stromal cell extracellular vesicles showed significant improvement in lung architecture, vessel formation, and inflammatory modulation. In addition, we observed significantly increased in vitro neurosphere formation and altered neural progenitor cell transcriptional signatures. Conclusions: Our multifactorial lung injury model impairs neural progenitor cell function. Observed pulmonary and neurologic alterations are mitigated by intratracheal treatment with mesenchymal stromal cell-derived extracellular vesicles.
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Displasia Broncopulmonar , Vesículas Extracelulares , Lesión Pulmonar , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Nacimiento Prematuro , Animales , Displasia Broncopulmonar/terapia , Femenino , Humanos , Recién Nacido , Pulmón , Lesión Pulmonar/terapia , Ratones , EmbarazoRESUMEN
A sneak peek into the @EarlyCareerERS session at #ERSCongress 2020 and the experience of organising an @EuroRespSoc Research Seminar http://bit.ly/39yncgO.
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The prenatal and perinatal environments can have profound effects on the development of chronic inflammatory diseases. However, mechanistic insight into how the early-life microenvironment can impact upon development of the lung and immune system and consequent initiation and progression of respiratory diseases is still emerging. Recent studies investigating the developmental origins of lung diseases have started to delineate the effects of early-life changes in the lung, environmental exposures and immune maturation on the development of childhood and adult lung diseases. While the influencing factors have been described and studied in mostly animal models, it remains challenging to pinpoint exactly which factors and at which time point are detrimental in lung development leading to respiratory disease later in life. To advance our understanding of early origins of chronic lung disease and to allow for proper dissemination and application of this knowledge, we propose four major focus areas: 1) policy and education; 2) clinical assessment; 3) basic and translational research; and 4) infrastructure and tools, and discuss future directions for advancement. This review is a follow-up of the discussions at the European Respiratory Society Research Seminar "Early origins of lung disease: towards an interdisciplinary approach" (Lisbon, Portugal, November 2019).
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Enfermedades Pulmonares , Enfermedades Respiratorias , Animales , Enfermedad Crónica , Exposición a Riesgos Ambientales , Femenino , Humanos , Pulmón , Enfermedades Pulmonares/diagnóstico , Enfermedades Pulmonares/epidemiología , Enfermedades Pulmonares/etiología , EmbarazoRESUMEN
In this article, the Group Chairs and early career members of the European Respiratory Society (ERS) Paediatric Assembly highlight some of the most interesting findings in the field of paediatrics which were presented at the 2018 international ERS Congress.
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.@EarlyCareerERS looks back on #LSC2018 http://ow.ly/6hjS30jB6P9.
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Bronchopulmonary dysplasia (BPD), the most common complication of extreme preterm birth, can be caused by oxygen-related lung injury and is characterized by impaired alveolar and vascular development. Mesenchymal stromal cells (MSCs) have lung protective effects. Conversely, BPD is associated with increased MSCs in tracheal aspirates. We hypothesized that endogenous lung (L-)MSCs are perturbed in a well-established oxygen-induced rat model mimicking BPD features. Rat pups were exposed to 21% or 95% oxygen from birth to postnatal day 10. On day 12, CD146+ L-MSCs were isolated and characterized according to the International Society for Cellular Therapy criteria. Epithelial and vascular repair potential were tested by scratch assay and endothelial network formation, respectively, immune function by mixed lymphocyte reaction assay. Microarray analysis was performed using the Affymetrix GeneChip and gene set enrichment analysis software. CD146+ L-MSCs isolated from rat pups exposed to hyperoxia had decreased CD73 expression and inhibited lung endothelial network formation. CD146+ L-MSCs indiscriminately promoted epithelial wound healing and limited T cell proliferation. Expression of potent antiangiogenic genes of the axonal guidance cue and CDC42 pathways was increased after in vivo hyperoxia, whereas genes of the anti-inflammatory Janus kinase (JAK)/signal transducer and activator of transcription (STAT) and lung/vascular growth-promoting fibroblast growth factor (FGF) pathways were decreased. In conclusion, in vivo hyperoxia exposure alters the proangiogenic effects and FGF expression of L-MSCs. In addition, decreased CD73 and JAK/STAT expression suggests decreased immune function. L-MSC function may be perturbed and contribute to BPD pathogenesis. These findings may lead to improvements in manufacturing exogenous MSCs with superior repair capabilities.
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Displasia Broncopulmonar/metabolismo , Lesión Pulmonar/metabolismo , Células Madre Mesenquimatosas/metabolismo , Oxígeno/efectos adversos , Animales , Animales Recién Nacidos , Displasia Broncopulmonar/etiología , Displasia Broncopulmonar/fisiopatología , Antígeno CD146/genética , Proliferación Celular/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Humanos , Pulmón/metabolismo , Pulmón/patología , Lesión Pulmonar/inducido químicamente , Lesión Pulmonar/patología , Células Madre Mesenquimatosas/patología , Oxígeno/administración & dosificación , Ratas , Linfocitos T/metabolismo , Linfocitos T/patologíaRESUMEN
BACKGROUND AIMS: Bronchopulmonary dysplasia (BPD), a chronic lung disease characterized by disrupted lung growth, is the most common complication in extreme premature infants. BPD leads to persistent pulmonary disease later in life. Alveolar epithelial type 2 cells (AEC2s), a subset of which represent distal lung progenitor cells (LPCs), promote normal lung growth and repair. AEC2 depletion may contribute to persistent lung injury in BPD. We hypothesized that induced pluripotent stem cell (iPSC)-derived AECs prevent lung damage in experimental oxygen-induced BPD. METHODS: Mouse AECs (mAECs), miPSCs/mouse embryonic stem sells, human umbilical cord mesenchymal stromal cells (hUCMSCs), human (h)iPSCs, hiPSC-derived LPCs and hiPSC-derived AECs were delivered intratracheally to hyperoxia-exposed newborn mice. Cells were pre-labeled with a red fluorescent dye for in vivo tracking. RESULTS: Airway delivery of primary mAECs and undifferentiated murine pluripotent cells prevented hyperoxia-induced impairment in lung function and alveolar growth in neonatal mice. Similar to hUCMSC therapy, undifferentiated hiPSCs also preserved lung function and alveolar growth in hyperoxia-exposed neonatal NOD/SCID mice. Long-term assessment of hiPSC administration revealed local teratoma formation and cellular infiltration in various organs. To develop a clinically relevant cell therapy, we used a highly efficient method to differentiate hiPSCs into a homogenous population of AEC2s. Airway delivery of hiPSC-derived AEC2s and hiPSC-derived LPCs, improved lung function and structure and resulted in long-term engraftment without evidence of tumor formation. CONCLUSIONS: hiPSC-derived AEC2 therapy appears effective and safe in this model and warrants further exploration as a therapeutic option for BPD and other lung diseases characterized by AEC injury.
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Células Epiteliales Alveolares/citología , Hiperoxia/complicaciones , Células Madre Pluripotentes Inducidas/citología , Lesión Pulmonar/etiología , Lesión Pulmonar/terapia , Animales , Animales Recién Nacidos , Diferenciación Celular , Modelos Animales de Enfermedad , Humanos , Células Madre Pluripotentes Inducidas/ultraestructura , Lesión Pulmonar/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Oxígeno , Teratoma/patologíaRESUMEN
Yearly more than 15 million babies are born premature (<37 weeks gestational age), accounting for more than 1 in 10 births worldwide. Lung injury caused by maternal chorioamnionitis or preeclampsia, postnatal ventilation, hyperoxia, or inflammation can lead to the development of bronchopulmonary dysplasia (BPD), one of the most common adverse outcomes in these preterm neonates. BPD patients have an arrest in alveolar and microvascular development and more frequently develop asthma and early-onset emphysema as they age. Understanding how the alveoli develop, and repair, and regenerate after injury is critical for the development of therapies, as unfortunately there is still no cure for BPD. In this review, we aim to provide an overview of emerging new concepts in the understanding of perinatal lung development and injury from a molecular and cellular point of view and how this is paving the way for new therapeutic options to prevent or treat BPD, as well as a reflection on current treatment procedures.
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Sepsis is the main cause of morbidity and mortality in neonates. Mesenchymal stromal cells (MSCs) are potent immune-modulatory cells. Their effect in neonatal sepsis has never been explored. We hypothesized that human umbilical cord-derived MSCs (hUC-MSCs) improve survival in experimental neonatal sepsis. Sepsis was induced in 3-day-old rats by intravenous injection of Escherichia coli (5 × 105/rat). One hour after infection, rats were treated intravenously with normal saline, hUC-MSCs, or with interferon-γ preconditioned hUC-MSCs (107 cells/kg). Eighteen hours after infection, survival, bacterial counts, lung neutrophil and macrophage influx, phagocytosis and apoptosis of splenocytes plasma, and LL-37 concentration were evaluated. Animals were observed for survival for 72 h after E. coli injection. Treatment with either hUC-MSCs or preconditioned hUC-MSCs significantly increased survival (hUC-MSCs, 81%; preconditioned hUC-MSCs, 89%; saline, 51%; P < 0.05). Both hUC-MSCs and preconditioned hUC-MSCs enhanced bacterial clearance. Lung neutrophil influx was decreased with preconditioned hUC-MSCs. The number of activated macrophages (CD206+) in the spleen was increased with hUC-MSCs and preconditioned hUC-MSCs; preconditioned hUC-MSCs increased the phagocytic activity of CD206+ macrophages. hUC-MSCs and preconditioned hUC-MSCs decreased splenocyte apoptosis in E. coli infected rats. Finally, LL-37 plasma levels were elevated in neonatal rats treated with hUC-MSCs or preconditioned hUC-MSCs. hUC-MSCs enhance survival and bacterial clearance in experimental neonatal sepsis. hUC-MSCs may be an effective adjunct therapy to reduce neonatal sepsis-related morbidity and mortality.
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Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/citología , Sepsis Neonatal/microbiología , Sepsis Neonatal/terapia , Cordón Umbilical/citología , Animales , Péptidos Catiónicos Antimicrobianos , Catelicidinas/sangre , Escherichia coli/fisiología , Humanos , Inflamación/patología , Pulmón/patología , Macrófagos/metabolismo , Sepsis Neonatal/sangre , Neutrófilos/metabolismo , Fagocitosis , Ratas , Bazo/patología , Análisis de SupervivenciaRESUMEN
Mesenchymal stromal cells (MSCs) are increasingly recognized for their therapeutic potential in a wide range of diseases, including lung diseases. Besides the use of bone marrow and umbilical cord MSCs for exogenous cell therapy, there is also increasing interest in the repair and regenerative potential of resident tissue MSCs. Moreover, they likely have a role in normal organ development, and have been attributed roles in disease, particularly those with a fibrotic nature. The main hurdle for the study of these resident tissue MSCs is the lack of a clear marker for the isolation and identification of these cells. The isolation technique described here applies multiple characteristics of lung resident MSCs (L-MSCs). Upon sacrifice of the rats, lungs are removed and rinsed multiple times to remove blood. Following mechanical dissociation by scalpel, the lungs are digested for 2-3 hr using a mix of collagenase type I, neutral protease and DNase type I. The obtained single cell suspension is subsequently washed and layered over density gradient medium (density 1.073 g/ml). After centrifugation, cells from the interphase are washed and plated in culture-treated flasks. Cells are cultured for 4-7 days in physiological 5% O2, 5% CO2 conditions. To deplete fibroblasts (CD146(-)) and to ensure a population of only L-MSCs (CD146(+)), positive selection for CD146(+) cells is performed through magnetic bead selection. In summary, this procedure reliably produces a population of primary L-MSCs for further in vitro study and manipulation. Because of the nature of the protocol, it can easily be translated to other experimental animal models.
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Pulmón , Células Madre Mesenquimatosas , Animales , Biomarcadores , Diferenciación Celular , Tratamiento Basado en Trasplante de Células y Tejidos , RatasRESUMEN
BACKGROUND: Little is known about the effects of propofol on oxidative stress and its effect on key structures of the contractile apparatus as the myosin light chain 2 (MLC2) and the p38MAPK survival pathway in the preterm heart. We hypothesized that propofol administration could attenuate the hypoxic myocardial injury after birth asphyxia. METHODS: Pregnant ewes were randomized to receive either propofol or isoflurane anesthesia. A total of 44 late-preterm lambs were subjected to in utero umbilical cord occlusion (UCO), resulting in asphyxia and cardiac arrest, or sham treatment. After emergency cesarean delivery, each fetus was resuscitated, mechanically ventilated, and supported under anesthesia for 8 h using the same anesthetic as the one received by its mother. RESULTS: At 8 h after UCO, occurrence of reactive oxygen species and activation of inducible nitric oxide synthase in the heart were lower in association with propofol anesthesia than with isoflurane. This was accompanied by less degradation of MLC2 but higher p38MAPK level and in echocardiography with a trend toward a higher median left ventricular fractional shortening. CONCLUSION: The use of propofol resulted in less oxidative stress and was associated with less cytoskeletal damage of the contractile apparatus than the use of isoflurane anesthesia.
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Asfixia/fisiopatología , Paro Cardíaco/fisiopatología , Estrés Oxidativo/efectos de los fármacos , Propofol/administración & dosificación , Animales , Animales Recién Nacidos , Asfixia Neonatal/fisiopatología , Miosinas Cardíacas/metabolismo , Ecocardiografía , Femenino , Feto/efectos de los fármacos , Corazón/fisiopatología , Isoflurano/administración & dosificación , Peroxidación de Lípido , Cadenas Ligeras de Miosina/metabolismo , Embarazo , Preñez , Distribución Aleatoria , Oveja Doméstica , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Skeletal muscle development has been the focus of intensive study for many decades. Recent advances in genetic manipulation of the mouse have increased our understanding of the cell signalling involved in the development of muscle progenitors which give rise to adult skeletal muscles and their stem cell populations. However, the influence of a vital tissue type - the peripheral nerve-has largely been ignored since its earliest descriptions. Here we carefully describe the timing in which myogenic progenitors expressing Pax3 and Pax7 (the earliest markers of myogenic cells) enter the limb buds of rat and mouse embryos, as well as the spatiotemporal relationship between these progenitors and the ingrowing peripheral nerve. We show that progenitors expressing Pax3 enter the limb bud one full day ahead of the first neurites and that Pax7-expressing progenitors (associated with secondary myogenesis in the limb) are first seen in the limb bud at the time of nerve entry and in close proximity to the nerve. The initial entry of the nerve also coincides with the first expression of myosin heavy chain showing that the first contact between nerves and myogenic cells correlates with the onset of myogenic differentiation. Furthermore, as the nerve grows into the limb, Pax3 expression is progressively replaced by Pax7 expression in myogenic progenitors. These findings indicate that the ingrowing nerve enters the limb presumptive muscle masses earlier than what was generally described and raises the possibility that nerve may influence the differentiation of muscle progenitors in rodent limbs.
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Esbozos de los Miembros/embriología , Esbozos de los Miembros/inervación , Desarrollo de Músculos/fisiología , Músculo Esquelético/embriología , Músculo Esquelético/inervación , Unión Neuromuscular/embriología , Animales , Diferenciación Celular/genética , Embrión de Mamíferos , Femenino , Regulación del Desarrollo de la Expresión Génica , Esbozos de los Miembros/metabolismo , Ratones , Ratones Endogámicos C57BL , Desarrollo de Músculos/genética , Músculo Esquelético/metabolismo , Unión Neuromuscular/genética , Unión Neuromuscular/metabolismo , Factores de Transcripción Paired Box/genética , Factores de Transcripción Paired Box/metabolismo , Embarazo , Ratas , Ratas WistarRESUMEN
Bronchopulmonary dysplasia (BPD) is the most common adverse outcome in extreme preterm neonates (born before 28 weeks gestation). BPD is characterized by interrupted lung growth and may predispose to early-onset emphysema and poor lung function in later life. At present, there is no treatment for BPD. Recent advances in stem/progenitor cell biology have enabled the exploration of endogenous lung progenitor populations in health and disease. In parallel, exogenous stem/progenitor cell administration has shown promise in protecting the lung from injury in the experimental setting. This review will provide an outline of the progenitor populations that have currently been identified in all tissue compartments of the distal lung and how they may be affected in BPD. A thorough understanding of the lung's endogenous progenitor populations during normal development, injury and repair may one day allow us to harness their regenerative capacity.
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Displasia Broncopulmonar , Recien Nacido Extremadamente Prematuro , Pulmón , Células Madre , Displasia Broncopulmonar/metabolismo , Displasia Broncopulmonar/patología , Displasia Broncopulmonar/fisiopatología , Humanos , Recién Nacido , Pulmón/metabolismo , Pulmón/patología , Pulmón/fisiopatologíaRESUMEN
BACKGROUND: Intra-amniotic lipopolysaccharide (LPS) exposure may affect neonatal outcome by altering fetal lung and immune system development. We hypothesized that intra-amniotic LPS exposure would cause persistent fetal pulmonary responses as the lungs develop in utero. METHODS: Fetal lambs were exposed to intra-amniotic LPS at 118 or at 118 and 123 d of gestational age (GA) with delivery at 125, 133, or 140 d (term = 147 d). Immune responses, PU.1 expression, Toll-like receptor (TLR)-1,2,4,6 mRNA levels, mast cell levels, and pulmonary elastin deposition were evaluated. RESULTS: After a single dose of LPS, pulmonary inflammatory responses were observed with increases of (i) PU.1 and TLR1 at 125 d GA and (ii) monocytes, lymphocytes, TLR2, and TLR6 at 133 d GA. Repetitive LPS exposure resulted in (i) increases of neutrophils, monocytes, PU.1, and TLR1 at 125 d GA; (ii) increases of neutrophils, PU.1, and TLR2 at 133 d GA; and (iii) decreases of mast cells, elastin foci, TLR4, and TLR6 at early gestation. At 140 d GA, only PU.1 was increased after repetitive LPS exposure. CONCLUSION: The preterm fetal lung can respond to a single exposure or repeated exposures from intra-amniotic LPS in multiple ways, but the absence of inflammatory and structural changes in LPS-exposed fetuses delivered near term suggest that the fetus can resolve an inflammatory stimulus in utero with time.
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Lipopolisacáridos/farmacología , Pulmón/embriología , Preñez , Ovinos/embriología , Animales , Peso Corporal , Femenino , Pulmón/efectos de los fármacos , Tamaño de los Órganos , EmbarazoRESUMEN
SIGNIFICANCE: Bronchopulmonary dysplasia (BPD) is a disease of the developing lung that afflicts extreme preterm infants in the neonatal intensive care unit. Follow-up studies into adulthood show that BPD is not merely a problem of the neonatal period, as it also may predispose to early-onset emphysema and poor lung function in later life. RECENT ADVANCES: The increasing promise of bone marrow- or umbilical cord-derived mesenchymal stromal cells (MSCs) to repair neonatal and adult lung diseases may for the first time offer the chance to make substantial strides in improving the outcome of extreme premature infants at risk of developing BPD. As more knowledge has been obtained on MSCs over the past decades, it has become clear that each organ has its own reservoir of endogenous MSCs, including the lung. CRITICAL ISSUES: We have only barely scratched the surface on what resident lung MSCs exactly are and what their role and function in lung development may be. Moreover, what happens to these putative repair cells in BPD when alveolar development goes awry and why do their counterparts from the bone marrow and umbilical cord succeed in restoring normal alveolar development when they themselves do not? FUTURE DIRECTIONS: Much work remains to be carried out to validate lung MSCs, but with the high potential of MSC-based treatment for BPD and other lung diseases, a thorough understanding of the endogenous lung MSC will be pivotal to get to the bottom of these diseases.
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Displasia Broncopulmonar/patología , Pulmón/patología , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/patología , Animales , Animales Recién Nacidos , Humanos , Recién Nacido , Recien Nacido PrematuroRESUMEN
BACKGROUND: Antenatal inflammation and maternal corticosteroids induce fetal lung maturation but interfere with late lung development. Canonical Wingless-Int (Wnt) signaling directs lung development and repair. We showed that intra-amniotic (IA) lipopolysaccharide (LPS) exposure disrupted developmental signaling pathways in the preterm lamb lungs. Therefore, we hypothesized that pulmonary Wnt signaling was altered by exposure to IA LPS and/or antenatal corticosteroids. METHODS: Ovine fetuses were exposed to IA LPS, maternal intramuscular betamethasone, a control saline injection, or a combination thereof at 107 and/or 114 d gestational age (term = 150 d gestational age) before delivery at 121 d gestational age. RESULTS: IA LPS exposure decreased the lung expression of lymphoid enhancer-binding factor 1 (LEF1), a major Wnt pathway effector. WNT1, WNT4, and downstream messenger ß-catenin decreased after LPS exposure. WNT7b mRNA increased fourfold 14 d post-LPS exposure. Betamethasone treatment 7 d before LPS exposure prevented the reduction in LEF1 expression, whereas betamethasone administration after LPS normalized the LPS-induced increase in Wnt7b mRNA. CONCLUSION: IA LPS exposure decreased canonical Wnt signaling in the developing lung. Antenatal corticosteroids before or after IA inflammation had different effects on pulmonary Wnt signaling. This study provides new insights into possible mechanisms by which prenatal inflammation affects lung development and how corticosteroid can be beneficial in this setting.
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Betametasona/administración & dosificación , Lipopolisacáridos/administración & dosificación , Pulmón/patología , Vía de Señalización Wnt , Animales , Betametasona/química , Femenino , Glucógeno Sintasa Quinasa 3/metabolismo , Glucógeno Sintasa Quinasa 3 beta , Inflamación , Lipopolisacáridos/química , Pulmón/metabolismo , Factor de Unión 1 al Potenciador Linfoide/metabolismo , Exposición Materna , Fosforilación , Embarazo , Preñez , Ovinos , Oveja Doméstica , Factores de Tiempo , Proteínas Wnt/metabolismo , beta Catenina/metabolismoRESUMEN
Understanding how alveoli and the underlying capillary network develop and how these mechanisms are disrupted in disease states is critical for developing effective therapies for lung regeneration. Recent evidence suggests that lung angiogenesis promotes lung development and repair. Vascular endothelial growth factor (VEGF) preserves lung angiogenesis and alveolarization in experimental O2-induced arrested alveolar growth in newborn rats, but combined VEGF+angiopoietin 1 treatment is necessary to correct VEGF-induced vessel leakiness. Hypoxia-inducible factors (HIFs) are transcription factors that activate multiple O2-sensitive genes, including those encoding for angiogenic growth factors, but their role during postnatal lung growth is incompletely understood. By inducing the expression of a range of angiogenic factors in a coordinated fashion, HIF may orchestrate efficient and safe angiogenesis superior to VEGF. We hypothesized that HIF inhibition impairs alveolarization and that HIF activation regenerates irreversible O2-induced arrested alveolar growth. HIF inhibition by intratracheal dominant-negative adenovirus (dnHIF-1α)-mediated gene transfer or chetomin decreased lung HIF-1α, HIF-2α, and VEGF expression and led to air space enlargement and arrested lung vascular growth. In experimental O2-induced arrested alveolar growth in newborn rats, the characteristic features of air space enlargement and loss of lung capillaries were associated with decreased lung HIF-1α and HIF-2α expression. Intratracheal administration of Ad.HIF-1α restored HIF-1α, endothelial nitric oxide synthase, VEGF, VEGFR2, and Tie2 expression and preserved and rescued alveolar growth and lung capillary formation in this model. HIFs promote normal alveolar development and may be useful targets for alveolar regeneration.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Hipoxia/metabolismo , Alveolos Pulmonares/fisiopatología , Regeneración/fisiología , Animales , Animales Recién Nacidos , Neovascularización Patológica/metabolismo , Neovascularización Patológica/fisiopatología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Oxígeno/metabolismo , Alveolos Pulmonares/metabolismo , Ratas , Receptor TIE-2/metabolismo , Factores de Transcripción/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Hypoxic-ischemic encephalopathy (HIE) is common in preterm infants, but currently no curative therapy is available. Cell-based therapy has a great potential in the treatment of hypoxic-ischemic preterm brain injury. Granulocyte-colony stimulating factor (G-CSF) is known to mobilize endogenous hematopoietic stem cells (HSC) and promotes proliferation of endogenous neural stem cells. On these grounds, we hypothesized that systemic G-CSF would be neuroprotective in a large translational animal model of hypoxic-ischemic injury in the preterm brain. Global hypoxia-ischemia (HI) was induced by transient umbilical cord occlusion in instrumented preterm sheep. G-CSF treatment (100µg/kg intravenously, during five consecutive days) was started one day before the global HI insult to ascertain mobilization of endogenous stem cells within the acute phase after global HI. Mobilization of HSC and neutrophils was studied by flow cytometry. Brain sections were stained for microglia (IBA-1), myelin basic protein (MBP) and myeloperoxidase (MPO) to study microglial proliferation, white matter injury and neutrophil invasion respectively. Electrographic seizure activity was analyzed using amplitude-integrated electroencephalogram (aEEG). G-CSF effectively mobilized CD34-positive HSC in the preterm sheep. In addition, G-CSF caused marked mobilization of neutrophils, but did not influence enhanced invasion of neutrophils into the preterm brain after global HI. Microglial proliferation and hypomyelination following global HI were reduced as a result of G-CSF treatment. G-CSF did not cause a reduction of the electrographic seizure activity after global HI. In conclusion, G-CSF induced mobilization of endogenous stem cells which was associated with modulation of the cerebral inflammatory response and reduced white matter injury in an ovine model of preterm brain injury after global HI. G-CSF treatment did not improve neuronal function as shown by seizure analysis. Our study shows that G-CSF treatment has neuroprotective potential following hypoxic-ischemic injury in the preterm brain.
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Encefalitis/patología , Hipoxia Fetal/complicaciones , Factor Estimulante de Colonias de Granulocitos/farmacología , Hipoxia-Isquemia Encefálica/complicaciones , Fármacos Neuroprotectores/farmacología , Animales , Modelos Animales de Enfermedad , Electrocardiografía , Electroencefalografía , Encefalitis/etiología , Hipoxia Fetal/patología , Feto , Citometría de Flujo , Movilización de Célula Madre Hematopoyética , Hipoxia-Isquemia Encefálica/patología , Inmunohistoquímica , Fibras Nerviosas Mielínicas/efectos de los fármacos , Convulsiones/etiología , OvinosRESUMEN
Lung diseases characterized by alveolar damage such as bronchopulmonary dysplasia (BPD) in premature infants and emphysema lack efficient treatments. Understanding the mechanisms contributing to normal and impaired alveolar growth and repair may identify new therapeutic targets for these lung diseases. Axonal guidance cues are molecules that guide the outgrowth of axons. Amongst these axonal guidance cues, members of the Semaphorin family, in particular Semaphorin 3C (Sema3C), contribute to early lung branching morphogenesis. The role of Sema3C during alveolar growth and repair is unknown. We hypothesized that Sema3C promotes alveolar development and repair. In vivo Sema3C knock down using intranasal siRNA during the postnatal stage of alveolar development in rats caused significant air space enlargement reminiscent of BPD. Sema3C knock down was associated with increased TLR3 expression and lung inflammatory cells influx. In a model of O2-induced arrested alveolar growth in newborn rats mimicking BPD, air space enlargement was associated with decreased lung Sema3C mRNA expression. In vitro, Sema3C treatment preserved alveolar epithelial cell viability in hyperoxia and accelerated alveolar epithelial cell wound healing. Sema3C preserved lung microvascular endothelial cell vascular network formation in vitro under hyperoxic conditions. In vivo, Sema3C treatment of hyperoxic rats decreased lung neutrophil influx and preserved alveolar and lung vascular growth. Sema3C also preserved lung plexinA2 and Sema3C expression, alveolar epithelial cell proliferation and decreased lung apoptosis. In conclusion, the axonal guidance cue Sema3C promotes normal alveolar growth and may be worthwhile further investigating as a potential therapeutic target for lung repair.
Asunto(s)
Células Epiteliales Alveolares/fisiología , Proliferación Celular , Péptidos y Proteínas de Señalización Intracelular/fisiología , Proteínas del Tejido Nervioso/fisiología , Animales , Animales Recién Nacidos , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/metabolismo , Apoptosis , Displasia Broncopulmonar/metabolismo , Displasia Broncopulmonar/patología , Línea Celular , Supervivencia Celular , Células Cultivadas , Células Endoteliales/fisiología , Técnicas de Silenciamiento del Gen , Hiperoxia/metabolismo , Hiperoxia/patología , Pulmón/irrigación sanguínea , Pulmón/inervación , Pulmón/fisiopatología , Microvasos/patología , Microvasos/fisiopatología , Neovascularización Fisiológica , Fibras Nerviosas Mielínicas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Peroxidasa/metabolismo , ARN Interferente Pequeño/genética , Ratas , Receptores de Superficie Celular/metabolismo , Transducción de Señal , Cicatrización de HeridasRESUMEN
BACKGROUND: Bronchopulmonary dysplasia (BPD) is one of the most common complications after preterm birth and is associated with intrauterine exposure to bacteria. Transforming growth factor-ß (TGFß) is implicated in the development of BPD. OBJECTIVES: We hypothesized that different and/or multiple bacterial signals could elicit divergent TGFß signaling responses in the developing lung. METHODS: Time-mated pregnant Merino ewes received an intra-amniotic injection of lipopolysaccharide (LPS) and/or Ureaplasma parvum serovar 3 (UP) at 117 days' and/or 121/122 days' gestational age (GA). Controls received an equivalent injection of saline and or media. Lambs were euthanized at 124 days' GA (term = 150 days' GA). TGFß1, TGFß2, TGFß3, TGFß receptor (R)1 and TGFßR2 protein levels, Smad2 phosphorylation and elastin deposition were evaluated in lung tissue. RESULTS: Total TGFß1 and TGFß2 decreased by 24 and 51% after combined UP+LPS exposure, whereas total TGFß1 increased by 31% after 7 days' LPS exposure but not after double exposures. Alveolar expression of TGFßR2 decreased 75% after UP, but remained unaltered after double exposures. Decreased focal elastin deposition after single LPS exposure was prevented by double exposures. CONCLUSIONS: TGFß signaling components and elastin responded differently to intrauterine LPS and UP exposure. Multiple bacterial exposures attenuated TGFß signaling and normalized elastin deposition.