A potential role of the JNK pathway in hyperoxia-induced cell death, myofibroblast transdifferentiation and TGF-β1-mediated injury in the developing murine lung

<p>Abstract</p> <p>Background</p> <p>Transforming growth factor-beta 1 (TGF-β1) has been implicated in hyperoxia-induced cell death and impaired alveolarization in the developing lung. In addition, the c-JunNH2-terminal kinase (JNK) pathway has been shown to have a role for TGF-β1-mediated effects. We hypothesized that the JNK pathway is an important regulator of hyperoxia-induced pulmonary responses in the developing murine lung.</p> <p>Results</p> <p>We used cultured human lung epithelial cells, fetal rat lung fibroblasts and a neonatal TGF-β1 transgenic mouse model. We demonstrate that hyperoxia inhibits cell proliferation, activates cell death mediators and causes cell death, and promotes myofibroblast transdifferentiation, in a dose-dependent manner. Except for fibroblast proliferation, the effects were mediated via the JNK pathway. In addition, since we observed increased expression of TGF-β1 by epithelial cells on exposure to hyperoxia, we used a TGF-β1 transgenic mouse model to determine the role of JNK activation in TGF-β1 induced effects on lung development and on exposure to hyperoxia. We noted that, in this model, inhibition of JNK signaling significantly improved the spontaneously impaired alveolarization in room air and decreased mortality on exposure to hyperoxia.</p> <p>Conclusions</p> <p>When viewed in combination, these studies demonstrate that hyperoxia-induced cell death, myofibroblast transdifferentiation, TGF-β1- and hyperoxia-mediated pulmonary responses are mediated, at least in part, via signaling through the JNK pathway.</p

A potential role of the JNK pathway in hyperoxia-induced cell death, myofibroblast transdifferentiation and TGF-β1-mediated injury in the developing murine lung.

BackgroundTransforming growth factor-beta 1 (TGF-β1) has been implicated in hyperoxia-induced cell death and impaired alveolarization in the developing lung. In addition, the c-JunNH2-terminal kinase (JNK) pathway has been shown to have a role for TGF-β1-mediated effects. We hypothesized that the JNK pathway is an important regulator of hyperoxia-induced pulmonary responses in the developing murine lung.ResultsWe used cultured human lung epithelial cells, fetal rat lung fibroblasts and a neonatal TGF-β1 transgenic mouse model. We demonstrate that hyperoxia inhibits cell proliferation, activates cell death mediators and causes cell death, and promotes myofibroblast transdifferentiation, in a dose-dependent manner. Except for fibroblast proliferation, the effects were mediated via the JNK pathway. In addition, since we observed increased expression of TGF-β1 by epithelial cells on exposure to hyperoxia, we used a TGF-β1 transgenic mouse model to determine the role of JNK activation in TGF-β1 induced effects on lung development and on exposure to hyperoxia. We noted that, in this model, inhibition of JNK signaling significantly improved the spontaneously impaired alveolarization in room air and decreased mortality on exposure to hyperoxia.ConclusionsWhen viewed in combination, these studies demonstrate that hyperoxia-induced cell death, myofibroblast transdifferentiation, TGF-β1- and hyperoxia-mediated pulmonary responses are mediated, at least in part, via signaling through the JNK pathway

A potential role of the JNK pathway in hyperoxia-induced cell death, myofibroblast transdifferentiation and TGF-beta1-mediated injury in the developing murine lung

Abstract Background Transforming growth factor-beta 1 (TGF-β1) has been implicated in hyperoxia-induced cell death and impaired alveolarization in the developing lung. In addition, the c-JunNH2-terminal kinase (JNK) pathway has been shown to have a role for TGF-β1-mediated effects. We hypothesized that the JNK pathway is an important regulator of hyperoxia-induced pulmonary responses in the developing murine lung. Results We used cultured human lung epithelial cells, fetal rat lung fibroblasts and a neonatal TGF-β1 transgenic mouse model. We demonstrate that hyperoxia inhibits cell proliferation, activates cell death mediators and causes cell death, and promotes myofibroblast transdifferentiation, in a dose-dependent manner. Except for fibroblast proliferation, the effects were mediated via the JNK pathway. In addition, since we observed increased expression of TGF-β1 by epithelial cells on exposure to hyperoxia, we used a TGF-β1 transgenic mouse model to determine the role of JNK activation in TGF-β1 induced effects on lung development and on exposure to hyperoxia. We noted that, in this model, inhibition of JNK signaling significantly improved the spontaneously impaired alveolarization in room air and decreased mortality on exposure to hyperoxia. Conclusions When viewed in combination, these studies demonstrate that hyperoxia-induced cell death, myofibroblast transdifferentiation, TGF-β1- and hyperoxia-mediated pulmonary responses are mediated, at least in part, via signaling through the JNK pathway

Modeling the Pathogenesis of Charcot-Marie-Tooth Disease Type 1A Using Patient-Specific iPSCs

Summary: Charcot-Marie-Tooth disease type 1A (CMT1A), one of the most frequent inherited peripheral neuropathies, is associated with PMP22 gene duplication. Previous studies of CMT1A mainly relied on rodent models, and it is not yet clear how PMP22 overexpression leads to the phenotype in patients. Here, we generated the human induced pluripotent stem cell (hiPSC) lines from two CMT1A patients as an in vitro cell model. We found that, unlike the normal control cells, CMT1A hiPSCs rarely generated Schwann cells through neural crest stem cells (NCSCs). Instead, CMT1A NCSCs produced numerous endoneurial fibroblast-like cells in the Schwann cell differentiation system, and similar results were obtained in a PMP22-overexpressing iPSC model. Therefore, despite the demyelination-remyelination and/or dysmyelination theory for CMT1A pathogenesis, developmental disabilities of Schwann cells may be considered as an underlying cause of CMT1A. Our results may have important implications for the uncovering of the underlying mechanism and the development of a promising therapeutic strategy for CMT1A neuropathy

A Critical Regulatory Role for Macrophage Migration Inhibitory Factor in Hyperoxia-Induced Injury in the Developing Murine Lung

<div><p>Background</p><p>The role and mechanism of action of MIF in hyperoxia-induced acute lung injury (HALI) in the newborn lung are not known. We hypothesized that MIF is a critical regulatory molecule in HALI in the developing lung.</p><p>Methodology</p><p>We studied newborn wild type (WT), MIF knockout (MIFKO), and MIF lung transgenic (MIFTG) mice in room air and hyperoxia exposure for 7 postnatal (PN) days. Lung morphometry was performed and mRNA and protein expression of vascular mediators were analyzed.</p><p>Results</p><p>MIF mRNA and protein expression were significantly increased in WT lungs at PN7 of hyperoxia exposure. The pattern of expression of Angiopoietin 2 protein (in MIFKO>WT>MIFTG) was similar to the mortality pattern (MIFKO>WT>MIFTG) in hyperoxia at PN7. In room air, MIFKO and MIFTG had modest but significant increases in chord length, compared to WT. This was associated with decreased expression of Angiopoietin 1 and Tie 2 proteins in the MIFKO and MIFTG, as compared to the WT control lungs in room air. However, on hyperoxia exposure, while the chord length was increased from their respective room air controls, there were no differences between the 3 genotypes.</p><p>Conclusion</p><p>These data point to the potential roles of Angiopoietins 1, 2 and their receptor Tie2 in the MIF-regulated response in room air and upon hyperoxia exposure in the neonatal lung.</p></div

Effect of hyperoxia on lung architecture, BAL total and differential cell counts, BAL total protein concentrations and IL-6 levels in NB MIF KO and MIF TG mice at PN7.

<p>Representative photomicrographs of lung histology (H&E stain) of NB MIF KO, MIF TG and WT control mice exposed to RA or survived 100% O₂ at PN7 (3A). The figures are illustrative of a minimum of 8 animals in each group. Alveolar size, as measured by chord length and septal thickness, confirmed features noted on lung histology (3B and 3C). Each bar represents the mean ± SEM of a minimum of 5 animals. BAL total and differential cell recovery of NB MIF KO, MIF TG and WT control mice exposed to RA or survived 100% O₂ at PN7 (3D and 3E). Each bar represents the mean ± SEM of a minimum of 12 animals. BAL total protein levels of NB MIF KO, MIF TG and WT control mice exposed to RA or survived 100% O₂ at PN7 (3F). Each bar represents the mean ± SEM of a minimum of 4 animals. BAL IL-6 levels of NB MIF KO, MIF TG and WT control mice exposed to RA or survived 100% O₂ at PN7 (3G). Quantitative measurements of RV hypertrophy (3H).^§Each bar represents the mean ± SEM of a minimum of 3 animals. MIF +/+: wild type; MIF −/−: macrophage migration inhibitory factor knock out; MIF TG +: macrophage migration inhibitory factor (over-expressing) transgenic; BAL: bronchoalveolar lavage; IL-6: interleukin-6; RV: right ventricle; LV: left ventricle; IVS: interventricular septum. ^§Note: MIF KO assessments were done at PN6, due to their increased mortality in hyperoxia at PN7. ##<i>P</i><0.0001, #<i>P</i>≤0.01, *<i>P</i><0.05.</p

Effect of hyperoxia on RNA expression of vascular mediators and their receptors in NB MIF KO and MIF TG mice at PN7.

<p>The ratios of mRNA of VEGF -A, and the receptors -R1 and -R3 (4A) as well as Ang1 and Ang2 and their receptor Tie2 (4B) with β-actin were quantified by densitometery. Ang1, Ang2, Tie2, and VEGF-A proteins, with β-actin as controls, were detected by western blotting (4C). The densitometric values of Ang2/Ang1 ratio and Tie2 protein expression are noted in 4D and 4E, respectively. The noted values represent assessments in a minimum of 4 animals in each group. MIF +/+: wild type; MIF −/−: macrophage migration inhibitory factor knock out; MIF TG +: macrophage migration inhibitory factor (over-expressing) transgenic; VEGF: vascular endothelial growth factor; R: receptor; Ang: angiopoietin; Tie2: Tyrosine kinase with Ig and EGF homology domain 2 (receptor for Ang1 and 2); β-actin: beta actin. ##<i>P</i><0.0001, #<i>P</i>≤0.01, *<i>P</i><0.05.</p

Effect of lung-targeted VEGF overexpression in NB MIF KO mouse lungs.

<p>Representative photomicrographs of lung histology (H&E stain) of NB MIF KO, MIF KO VEGF TG and WT litter-mate controls, on regular or dox (to activate VEGF165 in the VEGF TG mice) water mice in RA at PN7 (<b>5A</b>). The figures are illustrative of a minimum of 4 animals in each group. Alveolar size, as measured by chord length, confirmed features noted on lung histology (<b>5B</b>). Each bar represents the mean ± SEM of a minimum of three animals. BAL total cell count of NB MIF KO, MIF KO VEGF TG and WT litter-mate controls, on regular or dox (to activate VEGF165 in the VEGF TG mice) water mice in RA at PN7 (<b>5C</b>). Each bar represents the mean ± SEM of a minimum of three animals. VEGF: vascular endothelial growth factor; MIF +/+: wild type; MIF −/−: macrophage migration inhibitory factor knock out; VEGF TG+: vascular endothelial growth factor (over-expressing) transgenic; dox: doxycycline; BAL: bronchoalveolar lavage. ##<i>P</i><0.0001, #<i>P</i>≤0.01.</p