Surfactant protein A (SP-A) gene targeted mice

AbstractMice lacking surfactant protein A (SP-A) mRNA and protein in vivo were generated using gene targeting techniques. SP-A (−/−) mice have normal levels of SP-B, SP-C and SP-D mRNA and protein and survive and breed normally in vivarium conditions. Phospholipid composition, secretion and clearance, and incorporation of phospholipid precursors are normal in the SP-A (−/−) mice. Lungs of SP-A (−/−) mice have markedly decreased tubular myelin figures and clear Group B streptococci and Pseudomonas aeruginosa less efficiently than SP-A wild type mice. These studies of SP-A (−/−) mice demonstrate that SP-A has an important role in the innate immune system of the lung in vivo

Gene expression and biological processes influenced by deletion of Stat3 in pulmonary type II epithelial cells

<p>Abstract</p> <p>Background</p> <p>The signal transducer and activator of transcription 3 (STAT3) mediates gene expression in response to numerous growth factors and cytokines, playing an important role in many cellular processes. To better understand the molecular mechanisms by which <it>Stat3 </it>influences gene expression in the lung, the effect of pulmonary epithelial cell specific deletion of <it>Stat3 </it>on genome wide mRNA expression profiling was assessed. Differentially expressed genes were identified from Affymetrix Murine GeneChips analysis and subjected to gene ontology classification, promoter analysis, pathway mapping and literature mining.</p> <p>Results</p> <p>Total of 791 mRNAs were significantly increased and 314 mRNAs were decreased in response to the deletion of <it>Stat3</it>^Δ/Δin the lung. STAT is the most enriched cis-elements in the promoter regions of those differentially expressed genes. Deletion of <it>Stat3 </it>induced genes influencing protein metabolism, transport, chemotaxis and apoptosis and decreased the expression of genes mediating lipid synthesis and metabolism. Expression of <it>Srebf1 </it>and <it>2</it>, genes encoding key regulators of fatty acid and steroid biosynthesis, was decreased in type II cells from the <it>Stat3</it>^Δ/Δmice, consistent with the observation that lung surfactant phospholipids content was decreased. <it>Stat3 </it>influenced both pro- and anti-apoptotic pathways that determine cell death or survival. <it>Akt</it>, a potential transcriptional target of <it>Stat3</it>, was identified as an important participant in <it>Stat3 </it>mediated pathways including Jak-Stat signaling, apoptosis, Mapk signaling, cholesterol and fatty acid biosynthesis.</p> <p>Conclusion</p> <p>Deletion of <it>Stat3 </it>from type II epithelial cells altered the expression of genes regulating diverse cellular processes, including cell growth, apoptosis and lipid metabolism. Pathway analysis indicates that STAT3 regulates cellular homeostasis through a complex regulatory network that likely enhances alveolar epithelial cell survival and surfactant/lipid synthesis, necessary for the protection of the lung during injury.</p

The pulmonary surfactant system matures upon pipping in the freshwater turtle Chelydra serpentina

© The Company of BiologistsPulmonary surfactant (PS), a mixture of phospholipids (PL), neutral lipids and surfactant proteins (SP), lowers surface tension within the lung, which increases lung compliance and improves the removal of fluid at birth. Here, we have examined the expression of thyroid transcription factor-1 (TTF-1) and the surfactant protein SP-B, and also the composition of pulmonary surfactant lipids in the developing lung of the turtle Chelydra serpentina. Lavage and lung tissue were collected from late embryonic, pipped and hatchling turtles. TTF-1, a regulator of gene expression of surfactant proteins and cell differentiation in mammals, was detected using immunohistochemistry in epithelia of the gas-exchange area and conducting airways during late development. Expression declined in hatchlings. SP-B was detected in subsets of cells within the respiratory epithelium at all stages sampled. The same cell types also stained for TTF-1. Turtle surfactant lipids matured toward the end of incubation. Maximal secretion of both total phospholipids and disaturated phospholipid (DSP) occurred at the time of pipping, coincident with the onset of breathing. The DSP/PL ratio increased after pipping, whereas cholesterol levels (Chol) increased prior to pipping. This resulted in a decrease in the Chol/PL and Chol/DSP ratios after pipping. Thus, TTF-1 and SP-B appear to be highly conserved within the vertebrates. Maturation of surfactant phospholipid content occurred with the commencement of pulmonary ventilation.Sonya D. Johnston, Christopher B. Daniels, David Cenzato, Jeffrey A. Whitsett and Sandra Orgei

Temporal effects of Sprouty on lung morphogenesis

AbstractParacrine signaling mediated by FGF-10 and the FGF-R2IIIb receptor is required for formation of the lung. To determine the temporal requirements for FGF signaling during pulmonary morphogenesis, Sprouty-4 (Spry-4), an intracellular FGF receptor antagonist, was expressed in epithelial cells of the fetal lung under control of a doxycycline-inducible system. Severe defects in lobulation and severe lung hypoplasia were observed when Spry-4 was expressed throughout fetal lung development (E6.5–E18.5) or from E6.5 until E13.5. Effects of Spry-4 on branching were substantially reversed by removal of doxycycline from the dam at E12.5, but not at E13.5. In contrast, when initiated late in development (E12.5 to birth), Spry-4 caused less severe pulmonary hypoplasia. Expression of Spry-4 from E16.5 to E18.5 reduced lung growth and resulted in perinatal death due to respiratory failure. Expression of Spry-4 during the saccular and alveolar stages, from E18.5 to postnatal day 21, caused mild emphysema. These findings demonstrate that the embryonic-pseudoglandular stage is a critical time period during which Spry-sensitive pathways are required for branching morphogenesis, lobulation, and formation of the peripheral lung parenchyma

GM-CSF Regulates Alveolar Macrophage Differentiation and Innate Immunity in the Lung through PU.1

AbstractGM-CSF gene targeted (GM−/−) mice are susceptible to respiratory infections and develop alveolar proteinosis due to defects in innate immune function and surfactant catabolism in alveolar macrophages (AMs), respectively. Reduced cell adhesion, phagocytosis, pathogen killing, mannose- and Toll-like receptor expression, and LPS- or peptidoglycan-stimulated TNFα release were observed in AMs from GM−/− mice. The transcription factor PU.1 was markedly reduced in AMs of GM−/− mice in vivo and was restored by selective expression of GM-CSF in the lungs of SPC-GM/GM−/− transgenic mice. Retrovirus-mediated expression of PU.1 in AMs from GM−/− mice rescued host defense functions and surfactant catabolism by AMs. We conclude that PU.1 mediates GM-CSF-dependent effects on terminal differentiation of AMs regulating innate immune functions and surfactant catabolism by AMs

A systems approach to mapping transcriptional networks controlling surfactant homeostasis

<p>Abstract</p> <p>Background</p> <p>Pulmonary surfactant is required for lung function at birth and throughout life. Lung lipid and surfactant homeostasis requires regulation among multi-tiered processes, coordinating the synthesis of surfactant proteins and lipids, their assembly, trafficking, and storage in type II cells of the lung. The mechanisms regulating these interrelated processes are largely unknown.</p> <p>Results</p> <p>We integrated mRNA microarray data with array independent knowledge using Gene Ontology (GO) similarity analysis, promoter motif searching, protein interaction and literature mining to elucidate genetic networks regulating lipid related biological processes in lung. A Transcription factor (TF) - target gene (TG) similarity matrix was generated by integrating data from different analytic methods. A scoring function was built to rank the likely TF-TG pairs. Using this strategy, we identified and verified critical components of a transcriptional network directing lipogenesis, lipid trafficking and surfactant homeostasis in the mouse lung.</p> <p>Conclusions</p> <p>Within the transcriptional network, SREBP, CEBPA, FOXA2, ETSF, GATA6 and IRF1 were identified as regulatory hubs displaying high connectivity. SREBP, FOXA2 and CEBPA together form a common core regulatory module that controls surfactant lipid homeostasis. The core module cooperates with other factors to regulate lipid metabolism and transport, cell growth and development, cell death and cell mediated immune response. Coordinated interactions of the TFs influence surfactant homeostasis and regulate lung function at birth.</p

Pluripotent Transcription Factors Possess Distinct Roles in Normal versus Transformed Human Stem Cells

Cancer and normal stem cells (SCs) share proliferative properties of self-renewal and expression of key transcription factors (TFs). Despite similar TF identities, the functional role of specific TFs responsible for retaining SC state has yet to be examined in cancer.Here, we compare the role of Oct4 and Nanog, two-core pluripotent TFs, in transformed (t-hPSCs), and normal human pluripotent stem cells (hPSCs). Unlike normal SCs, self-renewal and survival of t-hPSCs were found to be independent of Oct4. In contrast, t-hPSCs exhibit hypersensitivity to reduction in Nanog and demonstrate complete loss of self-renewal coupled with apoptosis. Dual and sequential knockdown of Oct4 and Nanog revealed that sensitivity of t-hPSCs to Nanog was Oct4 dependent.Our study indicates a bifurcation for the role of two-core SC and cancer related TFs in self-renewal and survival processes. We suggest that the divergent roles of these TFs establish a paradigm to develop novel therapeutics towards selective destruction of aggressive tumors harboring cancer stem cells (CSCs) with similar molecular signatures

Transcript Profiling of Elf5+/− Mammary Glands during Pregnancy Identifies Novel Targets of Elf5

Background: Elf5, an epithelial specific Ets transcription factor, plays a crucial role in the pregnancy-associated development of the mouse mammary gland. Elf5 2/2 embryos do not survive, however the Elf5 +/2 mammary gland displays a severe pregnancy-associated developmental defect. While it is known that Elf5 is crucial for correct mammary development and lactation, the molecular mechanisms employed by Elf5 to exert its effects on the mammary gland are largely unknown. Principal Findings: Transcript profiling was used to investigate the transcriptional changes that occur as a result of Elf5 haploinsufficiency in the Elf5 +/2 mouse model. We show that the development of the mouse Elf5 +/2 mammary gland is delayed at a transcriptional and morphological level, due to the delayed increase in Elf5 protein in these glands. We also identify a number of potential Elf5 target genes, including Mucin 4, whose expression, is directly regulated by the binding of Elf5 to an Ets binding site within its promoter. Conclusion: We identify novel transcriptional targets of Elf5 and show that Muc4 is a direct target of Elf5, further elucidatin