Presenilin-1 processing of ErbB4 in fetal type II cells is necessary for control of fetal lung maturation

AbstractMaturation of pulmonary fetal type II cells to initiate adequate surfactant production is crucial for postnatal respiratory function. Little is known about specific mechanisms of signal transduction controlling type II cell maturation. The ErbB4 receptor and its ligand neuregulin (NRG) are critical for lung development. ErbB4 is cleaved at the cell membrane by the γ-secretase enzyme complex whose active component is either presenilin-1 (PSEN-1) or presenilin-2. ErbB4 cleavage releases the 80kDa intracellular domain (4ICD), which associates with chaperone proteins such as YAP (Yes-associated protein) and translocates to the nucleus to regulate gene expression. We hypothesized that PSEN-1 and YAP have a development-specific expression in fetal type II cells and are important for ErbB4 signaling in surfactant production. In primary fetal mouse E16, E17, and E18 type II cells, PSEN-1 and YAP expression increased at E17 and E18 over E16. Subcellular fractionation showed a strong cytosolic and a weaker membrane location of both PSEN-1 and YAP. This was enhanced by NRG stimulation. Co-immunoprecipitations showed ErbB4 associated separately with PSEN-1 and with YAP. Their association, phosphorylation, and co-localization were induced by NRG. Confocal immunofluorescence and nuclear fractionation confirmed these associations in a time-dependent manner after NRG stimulation. Primary ErbB4-deleted E17 type II cells were transfected with a mutant ErbB4 lacking the γ-secretase binding site. When compared to transfection with wild-type ErbB4, the stimulatory effect of NRG on surfactant protein mRNA expression was lost. We conclude that PSEN-1 and YAP have crucial roles in ErbB4 signal transduction during type II cell maturation

ErbB4 regulates the timely progression of late fetal lung development

AbstractThe ErbB4 receptor has an important function in fetal lung maturation. Deletion of ErbB4 leads to alveolar hypoplasia and hyperreactive airways similar to the changes in bronchopulmonary dysplasia (BPD). BPD is a chronic pulmonary disorder affecting premature infants as a consequence of lung immaturity, lung damage, and abnormal repair. We hypothesized that proper ErbB4 function is needed for the timely progression of fetal lung development. An ErbB4 transgenic cardiac rescue mouse model was used to study the effect of ErbB4 deletion on fetal lung structure, surfactant protein (SP) expression, and synthesis, and inflammation. Morphometric analyses revealed a delayed structural development with a significant decrease in saccular size at E18 and more pronounced changes at E17, keeping these lungs in the canalicular stage. SP-B mRNA expression was significantly down regulated at E17 with a subsequent decrease in SP-B protein expression at E18. SP-D protein expression was significantly decreased at E18. Surfactant phospholipid synthesis was significantly decreased on both days, and secretion was down regulated at E18. We conclude that pulmonary ErbB4 deletion results in a structural and functional delay in fetal lung development, indicating a crucial regulatory role of ErbB4 in the timely progression of fetal lung development

A novel European H5N8 influenza A virus has increased virulence in ducks but low zoonotic potential

We investigated in a unique setup of animal models and a human lung explant culture biological properties, including zoonotic potential, of a representative 2016 highly pathogenic avian influenza virus (HPAIV) H5N8, clade 2.3.4.4 group B (H5N8B), that spread rapidly in a huge and ongoing outbreak series in Europe and caused high mortality in waterfowl and domestic birds. HPAIV H5N8B showed increased virulence with rapid onset of severe disease and mortality in Pekin ducks due to pronounced neuro- and hepatotropism. Cross-species infection was evaluated in mice, ferrets, and in a human lung explant culture model. While the H5N8B isolate was highly virulent for Balb/c mice, virulence and transmissibility were grossly reduced in ferrets, which was mirrored by marginal replication in human lung cultures infected ex vivo. Our data indicate that the 2016 HPAIV H5N8B is avian-adapted with augmented virulence for waterfowl, but has low zoonotic potential. The here tested combination of animal studies with the inoculation of human explants provides a promising future workflow to evaluate zoonotic potential, mammalian replication competence and avian virulence of HPAIV.Peer Reviewe

Pneumolysin induced mitochondrial dysfunction leads to release of mitochondrial DNA

Abstract Streptococcus pneumoniae (S.pn.) is the most common bacterial pathogen causing community acquired pneumonia. The pore-forming toxin pneumolysin (PLY) is the major virulence factor of S.pn. and supposed to affect alveolar epithelial cells thereby activating the immune system by liberation of danger-associated molecular patterns (DAMP). To test this hypothesis, we established a novel live-cell imaging based assay to analyse mitochondrial function and associated release of mitochondrial DNA (mtDNA) as DAMP in real-time. We first revealed that bacterially released PLY caused significant changes of the cellular ATP homeostasis and led to morphologic alterations of mitochondria in human alveolar epithelial cells in vitro and, by use of spectral live-tissue imaging, in human alveoli. This was accompanied by strong mitochondrial calcium influx and loss of mitochondrial membrane potential resulting in opening of the mitochondrial permeability transition pore and mtDNA release without activation of intrinsic apoptosis. Moreover, our data indicate cellular mtDNA liberation via microvesicles, which may contribute to S.pn. related pro-inflammatory immune activation in the human alveolar compartment

Estrogen-induced upregulation of Sftpb requires transcriptional control of neuregulin receptor ErbB4 in mouse lung type II epithelial cells

AbstractEstrogen is known for its positive stimulatory effects on surfactant proteins. ErbB4 receptor and its ligand neuregulin (NRG) positively stimulate lung development. ErbB receptors interact with nuclear receptors and ErbB4 co-regulates estrogen receptor (ER)α expression in breast cells. ERβ is highly expressed in pneumocytes and its deletion leads to fewer alveoli and reduced elastic recoil. A similar picture was seen in ErbB4-deleted lungs. We hypothesized that estrogen signals its effect on surfactant protein B (Sftpb) expression through interactions of ERβ and ErbB4. Estrogen and NRG treatment decreased cell numbers and stimulated Sftpb expression in type II cells. Estrogen and NRG both stimulated phosphorylation of ERβ and co-localization of both receptors. Overexpression of ERβ increased the cell number and Sftpb expression, which was further augmented by estrogen and NRG. Finally, estrogen and NRG stimulated ERβ and ErbB4 binding to the Sftpb promoter. Overexpression of these receptors stimulated Sftpb promoter activation, which was further enhanced by estrogen and NRG. The stimulatory effect of estrogen and NRG was abolished in ErbB4 deletion and reconstituted by re-expression of full-length ErbB4 in fetal ErbB4-deleted type II cells. Estrogen-induced nuclear translocation of ErbB4 required the intact γ-secretase cleavage site but not the nuclear localization sequence of the ErbB4 receptor, suggesting that ERβ might function as a nuclear chaperone for ErbB4. These studies demonstrate that estrogen effects on Sftpb expression require an interaction of ERβ and ErbB4. We speculate that the stimulatory effects of estrogen on Sftpb are under transcriptional control of ErbB4