Loïc Blanchon

A Role for Mesenchyme Dynamics in Mouse Lung Branching Morphogenesis

Mammalian airways are highly ramified tree-like structures that develop by the repetitive branching of the lung epithelium into the surrounding mesenchyme through reciprocal interactions. Based on a morphometric analysis of the epithelial tree, it has been recently proposed that the complete branching scheme is specified early in each lineage by a programme using elementary patterning routines at specific sites and times in the developing lung. However, the coupled dynamics of both the epithelium and mesenchyme have been overlooked in this process. Using a qualitative and quantitative in vivo morphometric analysis of the E11.25 to E13.5 mouse whole right cranial lobe structure, we show that beyond the first generations, the branching stereotypy relaxes and both spatial and temporal variations are common. The branching pattern and branching rate are sensitive to the dynamic changes of the mesoderm shape that is in turn mainly dependent upon the volume and shape of the surrounding intrathoracic organs. Spatial and temporal variations of the tree architecture are related to local and subtle modifications of the mesoderm growth. Remarkably, buds never meet after suffering branching variations and continue to homogenously fill the opening spaces in the mesenchyme. Moreover despite inter-specimen variations, the growth of the epithelial tree and the mesenchyme remains highly correlated over time at the whole lobe level, implying a long-range regulation of the lung lobe morphogenesis. Together, these findings indicate that the lung epithelial tree is likely to adapt in real time to fill the available space in the mesenchyme, rather than being rigidly specified and predefined by a global programme. Our results strongly support the idea that a comprehensive understanding of lung branching mechanisms cannot be inferred from the branching pattern or behavior alone. Rather it needs to be elaborated upon with the reconsideration of mesenchyme-epithelium coupled growth and lung tissues mechanics.

Esterification of Vitamin A by the Human Placenta Involves Villous Mesenchymal Fibroblasts

Vitamin A (retinol) and its active derivatives (retinoic acids) are essential for growth and development of the mammalian fetus. Maternally derived retinol must pass the placenta to reach the developing fetus. Despite its apparent importance, little is known concerning placental transfer and metabolism of retinol, and particularly of placental production and storage of retinyl esters. To elucidate this metabolic pathway, we incubated, in the presence of retinol, 1) human full-term placental explants and 2) primary cultures of major cells types contributing to placental function: trophoblasts and villous mesenchymal fibroblasts. We used HPLC to determine the types and concentrations of retinyl esters produced by these explants and cells. About 14% of total cellular retinol in placental explants was esterified. The most abundant esters were myristate and palmitate. Primary cell cultures showed that fibroblasts efficiently produced retinyl esters, but trophoblasts did not. In both types of experiments, no retinyl esters were detected in the culture medium, suggesting that retinyl esters were produced for storage purpose. These results suggest that villous mesenchymal fibroblasts are primary sites of retinol esterification and storage in the placenta.

Placental Implications of Peroxisome Proliferator-Activated Receptors in Gestation and Parturition

The placenta is a transitory structure indispensable for the proper development of the embryo and fetus during mammalian gestation. Like other members of the nuclear receptor family, the peroxisome proliferator-activated receptors (PPARs) are known to be involved in the physiological and pathological events occurring during the placentation. This placental involvement has been recently reviewed focusing on the early stages of placental development (implantation and invasion, etc.), mouse PPARs knockout phenotypes, and cytotrophoblast physiology. In this review, we describe the placental involvement of PPARs (e.g., fat transport and metabolism, etc.) during the late stages of gestation and in the amniotic membranes, highlighting their roles in the inflammation process (e.g., chorioamnionitis), metabolic disorders (e.g., diabetes), and parturition.

Soluble Forms and Ligands of the Receptor for Advanced Glycation End-Products in Patients with Acute Respiratory Distress Syndrome: An Observational Prospective Study

Background The main soluble form of the receptor for advanced glycation end-products (sRAGE) is elevated during acute respiratory distress syndrome (ARDS). However other RAGE isoforms and multiple ligands have been poorly reported in the clinical setting, and their respective contribution to RAGE activation during ARDS remains unclear. Our goal was therefore to describe main RAGE isoforms and ligands levels during ARDS. Methods 30 ARDS patients and 30 mechanically ventilated controls were prospectively included in this monocenter observational study. Arterial, superior vena cava and alveolar fluid levels of sRAGE, endogenous-secretory RAGE (esRAGE), high mobility group box-1 protein (HMGB1), S100A12 and advanced glycation end-products (AGEs) were measured in duplicate ELISA on day 0, day 3 and day 6. In patients with ARDS, baseline lung morphology was assessed with computed tomography. Results ARDS patients had higher arterial, central venous and alveolar levels of sRAGE, HMGB1 and S100A12, but lower levels of esRAGE and AGEs, than controls. Baseline arterial sRAGE, HMGB1 and S100A12 were correlated with nonfocal ARDS (AUC 0.79, 0.65 and 0.63, respectively). Baseline arterial sRAGE, esRAGE, S100A12 and AGEs were associated with severity as assessed by PaO2/FiO2. Conclusions This is the first kinetics study of levels of RAGE main isoforms and ligands during ARDS. Elevated sRAGE, HMGB1 and S100A12, with decreased esRAGE and AGEs, were found to distinguish patients with ARDS from those without. Our findings should prompt future studies aimed at elucidating RAGE/HMGB1/S100A12 axis involvement in ARDS. Trial Registration clinicaltrials.gov Identifier: NCT01270295.

Nuclear retinoid receptors and pregnancy: placental transfer, functions, and pharmacological aspects.

Animal models of vitamin A (retinol) deficiency have highlighted its crucial role in reproduction and placentation, whereas an excess of retinoids (structurally or functionally related entities) can cause toxic and teratogenic effects in the embryo and foetus, especially in the first trimester of human pregnancy. Knock-out experimental strategies-targeting retinoid nuclear receptors RARs and RXRs have confirmed that the effects of vitamin A are mediated by retinoic acid (especially all-trans retinoic acid) and that this vitamin is essential for the developmental process. All these data show that the vitamin A pathway and metabolism are as important for the well-being of the foetus, as they are for that of the adult. Accordingly, during this last decade, extensive research on retinoid metabolism has yielded detailed knowledge on all the actors in this pathway, spurring the development of antagonists and agonists for therapeutic and research applications. Natural and synthetic retinoids are currently used in clinical practice, most often on the skin for the treatment of acne, and as anti-oncogenic agents in acute promyelocytic leukaemia. However, because of the toxicity and teratogenicity of retinoids during pregnancy, their pharmacological use needs a sound knowledge of their metabolism, molecular aspects, placental transfer, and action.

Net alveolar fluid clearance is associated with lung morphology phenotypes in acute respiratory distress syndrome

BACKGROUND The acute respiratory distress syndrome (ARDS) is a heterogeneous syndrome that encompasses multiple phenotypes, e.g. with regards to lung morphology as assessed by computed tomography (CT). Focal or non-focal lung morphology may influence the response to positive end-expiratory pressure (PEEP), recruitment manoeuvres and prone position. Lung morphology has been hypothesized to be associated with alveolar fluid clearance (AFC), thus explaining various responses to such therapeutic interventions; however, this hypothesis has not been specifically studied in humans. METHODS We measured net AFC rates in 30 patients with ARDS as a secondary data analysis of a prospective single-centre study. Net AFC rates were compared between patients with focal ARDS and those with non-focal ARDS, as assessed by lung CT-scans. RESULTS Net AFC rates were significantly lower in patients with non-focal ARDS (n=23; median [interquartile range], 1.5 [0-5.5] %/h) as compared to those with focal ARDS (n=7; 10.3 [4.5-15] %/h) (P=0.01). The area under the receiver-operating characteristic curve when net AFC rates were used to differentiate the presence from absence of non-focal ARDS was 0.93 (95% confidence interval, 0.81-1). Tidal volumes and PEEP levels differed between focal and non-focal ARDS patients, but there was no difference in arterial oxygenation or in alveolar-capillary permeability. CONCLUSIONS Non-focal lung morphology may be characterized by a functional endotype consistent with marked AFC impairment. Despite study limitations and the need for validating studies in larger cohorts, such novel findings may reinforce our understanding of the association between ARDS phenotypes and therapeutic responses.

Receptor for advanced glycation end-products and ARDS prediction: a multicentre observational study

Acute respiratory distress syndrome (ARDS) prediction remains challenging despite available clinical scores. To assess soluble receptor for advanced glycation end-products (sRAGE), a marker of lung epithelial injury, as a predictor of ARDS in a high-risk population, adult patients with at least one ARDS risk factor upon admission to participating intensive care units (ICUs) were enrolled in a multicentre, prospective study between June 2014 and January 2015. Plasma sRAGE and endogenous secretory RAGE (esRAGE) were measured at baseline (ICU admission) and 24 hours later (day one). Four AGER candidate single nucleotide polymorphisms (SNPs) were also assayed because of previous reports of functionality (rs1800625, rs1800624, rs3134940, and rs2070600). The primary outcome was ARDS development within seven days. Of 500 patients enrolled, 464 patients were analysed, and 59 developed ARDS by day seven. Higher baseline and day one plasma sRAGE, but not esRAGE, were independently associated with increased ARDS risk. AGER SNP rs2070600 (Ser/Ser) was associated with increased ARDS risk and higher plasma sRAGE in this cohort, although confirmatory studies are needed to assess the role of AGER SNPs in ARDS prediction. These findings suggest that among at-risk ICU patients, higher plasma sRAGE may identify those who are more likely to develop ARDS.

Aquaporins and Fetal Membranes From Diabetic Parturient Women: Expression Abnormalities and Regulation by Insulin.

CONTEXT During pregnancy, aquaporins (AQPs) expressed in fetal membranes are essential for controlling the homeostasis of the amniotic volume, but their regulation by insulin was never explored in diabetic women. OBJECTIVE The aim of our study was to investigate the involvement of AQPs 1, 3, 8, and 9 expressed in fetal membranes in diabetic parturient women and the control of their expression by insulin. DESIGN AND PARTICIPANTS From 129 fetal membranes in four populations (controls, type 1, type 2 [T2D], and gestational diabetes [GD]), we established an expression AQP profile. In a second step, the amnion was used to study the control of the expression and functions of AQPs 3 and 9 by insulin. MAIN OUTCOMES AND MEASURES The expression of transcripts and proteins of AQPs was studied by quantitative RT-PCR and ELISA. We analyzed the regulation by insulin of the expression of AQPs 3 and 9 in the amnion. A tritiated glycerol test enabled us to measure the impact of insulin on the functional characteristics. Using an inhibitor of phosphatidylinositol 3-kinase, we analyzed the insulin intracellular signaling pathway. RESULTS The expression of AQP3 protein was significantly weaker in groups T2D and GD. In nondiabetic fetal membranes, we showed for the amnion (but not for the chorion) a significant repression by insulin of the transcriptional expression of AQPs 3 and 9, which was blocked by a phosphatidylinositol 3-kinase inhibitor. CONCLUSION In fetal membranes, the repression of AQP3 protein expression and functions observed in vivo is allowed by the hyperinsulinism described in pregnant women with T2D or GD.

DNA methyl transferases are differentially expressed in the human anterior eye segment

DNA methylation is an epigenetic mark involved in the control of genes expression. Abnormal epigenetic events have been reported in human pathologies but weakly documented in eye diseases. The purpose of this study was to establish DNMT mRNA and protein expression levels in the anterior eye segment tissues and their related (primary or immortalized) cell cultures as a first step towards future in vivo and in vitro methylomic studies.

Effects of tracheal occlusion with retinoic acid administration on normal lung development

Tracheal occlusion (TO) is an investigational therapy for severe congenital diaphragmatic hernia that decreases pulmonary hypoplasia, but sustained TO also induces deficient surfactant synthesis. Intramuscular maternal administration of retinoic acid (RA) in a surgical rabbit model of congenital diaphragmatic hernia showed a beneficial effect on lung maturation. We evaluated the potential of RA delivery into the trachea and studied the combined effects of TO and RA on normal lung development.