High throughput quantitative whole transcriptome analysis of adult mouse distal lung mesenchyme
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ABSTRACT:
PROVIDER: PRJNA430888 | ENA |
REPOSITORIES: ENA
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Project description:We used microfluidic single cell RNA-seq on 47 individual mouse lung mesenchymal cells at adulthood to measure the transcriptional state. We were able to classify these cells into distinct populations as well as map their signaling interactions with the distal lung epithelium and endothelium. In this manner, single cell RNA-seq can be used to unbiasedly determine the signals the dictate construction and maintenance of the alveolus.
2018-02-19 | GSE109444 | GEO
Project description:We used microfluidic single cell RNA-seq on 42 individual mouse lung mesenchymal and 69 endothelial cells at E18.5 to measure the transcriptional state. We were able to classify these cells into distinct populations as well as map their signaling interactions with the distal lung epithelium. In this manner, single cell RNA-seq can be used to unbiasedly determine the signals the dictate construction and maintenance of the alveolus.
2022-08-17 | GSE196874 | GEO
Project description:Distal limb mesenchyme cells were collected from embryonic chickens at Hamburger Hamilton (HH) stages HH24, HH27, and HH29, and also from samples that had been grafted from HH27 to HH20 and left to develop for 24 hours (equivalent to HH24). Samples were collected in triplicate (with 10 distal tips pooled per replicate), the RNA extracted, and sequenced on a HiSeq 2000 (Instrument: ST300). Bioinformatic and clustering analyses were performed to determine whether grafted cells are more similar to HH29 or HH24 samples, ie. which genes can be reset to expression levels observed in an earlier developmental stage.
2018-04-20 | E-MTAB-6437 | biostudies-arrayexpress
Project description:We used microfluidic single cell RNA-seq on 198 individual mouse lung epithelial cells at 4 different stages throughout development to measure the transcriptional states which define the developmental and cellular hierarchy of the distal mouse lung epithelium. We classified 80 cells comprising the distal lung epithelium at E18.5 into distinct populations using an unbiased genome-wide approach that did not require a priori knowledge of the underlying cell types or prior purification of cell types. This M-bM-^@M-^\reverse tissue engineeringM-bM-^@M-^] approach confirmed the basic outlines of the conventional model of cell type diversity in the distal lung and led to the discovery of a large number of novel transcriptional regulators and cell type markers that discriminate between the different populations. Moreover, we reconstructed the steps during maturation of bipotential progenitors into both alveolar lineages based on the presence of undifferentiated, differentiated as well as differentiation intermediate cells at the single time point E18.5. Finally, we followed Sftpc-positive cells throughout their lifecycle (E14.5, E16.5, E18.5, adult) and identified 7 gene sets that differentiate between the multipotential, bipotential, mature, as well as intermediate states of the AT2 lineage. 198 single-cell transcriptomes from mouse lung epithelium were analyzed in total, two 200-cell bulk control samples as well as one no-cell control; All single cell and control samples contain 92 external RNA spike-ins; For time point E18.5, three individual experiments were performed using 3 different pregnant mice (3 biological replicates): Replicate 1 (pooled sibling lungs) yielded 20 single cell transcriptomes, replicate 2 (one single embryonic lung) yielded 34 single cell transcriptomes and replicate 3 (pooled siibling lungs) yielded 26 single cell transcriptomes; In addition, a 200-cell bulk control sample was prepared for E18.5 replicate 1 and E18.5 replicate 3 experiments; A no-cell control sample was generated for the E18.5 replicate 1 experiment; For time point E14.5, one experiment (one pregnant mouse, pooled sibling lungs) was performed yielding 45 single cell transcriptomes; For time point E16.5, one experiment (one pregnant mouse, pooled sibling lungs) was performed yielding 27 single cell transcriptomes; For the adult time point, one 107 day old mouse was used and transcriptomes of 46 single cells were obtained; All single cell samples were processed on the microfluidic platform, 200-cell-bulk and no-cell control samples were processed in microliter volumes in PCR tubes.
2014-04-08 | E-GEOD-52583 | biostudies-arrayexpress
Project description:We used microfluidic single cell RNA-seq on 198 individual mouse lung epithelial cells at 4 different stages throughout development to measure the transcriptional states which define the developmental and cellular hierarchy of the distal mouse lung epithelium. We classified 80 cells comprising the distal lung epithelium at E18.5 into distinct populations using an unbiased genome-wide approach that did not require a priori knowledge of the underlying cell types or prior purification of cell types. This “reverse tissue engineering” approach confirmed the basic outlines of the conventional model of cell type diversity in the distal lung and led to the discovery of a large number of novel transcriptional regulators and cell type markers that discriminate between the different populations. Moreover, we reconstructed the steps during maturation of bipotential progenitors into both alveolar lineages based on the presence of undifferentiated, differentiated as well as differentiation intermediate cells at the single time point E18.5. Finally, we followed Sftpc-positive cells throughout their lifecycle (E14.5, E16.5, E18.5, adult) and identified 7 gene sets that differentiate between the multipotential, bipotential, mature, as well as intermediate states of the AT2 lineage.
2014-04-08 | GSE52583 | GEO
Project description:Distal limb mesenchyme cells were collected from embryonic chickens at Hamburger Hamilton (HH) stages HH24 and HH27, and from samples that had been grafted from HH20 to HH24 and left to develop for 24 hours. Bioinformatics and clustering analyses were then performed to determine whether grafted cells are more similar to HH24 (graft stage) or HH27 (host stage).
2023-08-24 | GSE232959 | GEO
Project description:Co-cultures of lung epithelium and mesenchyme are useful tools to study epithelial-mesenchymal crosstalk in lung development and disease. However, many previous attempts to generate such co-cultures have yielded poor juxtaposition between the epithelial and the mesenchymal lineage. In addition, induced pluripotent stem cell (iPSC)-derived co-cultures often contain generic mesenchyme that is not necessarily lung-specific. We sought to establish co-cultures of purified mouse iPSC-derived lung-specific mesenchyme and iPSC-derived lung epithelial progenitors. We used a mouse iPSC line carrying a lung mesenchyme-specific reporter/tracer (Tbx4-LERGFP) to generate lung mesenchymal progenitors by directed differentiation via a lateral plate mesodermal progenitor state (induced lung mesenchyme, iLM). In parallel we differentiated a mouse embryonic stem (ES) cell line carrying a Nkx2-1mCherry reporter into lung epithelial progenitor cells using our established directed differentiation protocol. We then combined the purified lung epithelial and mesenchymal progenitor cells and co-cultured them in distal or proximal differentiation media for 1 week on Matrigel. We find that cells self-organize into complex 3-dimensional organoids with closely juxtaposed epithelial and mesenchymal cells. Furthermore, co-culture affects the molecular phenotype of both lineages. Our iPSC-derived co-culture model can provide an inexhaustible source of cells for studying lung development, modeling diseases, and developing therapeutics.
2023-05-08 | GSE228313 | GEO
Project description:Previous studies have suggested that Bmp4 is a key Msx1-dependent mesenchymal odontogenic signal for driving tooth morphogenesis through the bud-to-cap transition. Whereas the bud stage tooth developmental arrest in Msx1-/- mutant mice was accompanied by reduction in mesenchymal Bmp4 mRNA expression, we show that depleting functional Bmp4 mRNAs in the tooth mesenchyme, through neural crest-specific gene inactivation in Bmp4f/f;Wnt1Cre mice, caused mandibular molar developmental arrest at the bud stage but allowed maxillary molars and incisors to develop to mineralized teeth. We show that the Wnt inhibitors Dkk2 and Wif1 were much more abundantly expressed in the mandibular than maxillary molar mesenchyme in wildtype embryos and that Dkk2 expression was significantly unregulated in the tooth mesenchyme in Bmp4f/f;Wnt1Cre embryos. In addition, expression of Osr2, which encodes a zinc finger protein that antagonizes Msx1-mediated activation of odontogenic mesenchyme, is significantly upregulated in the molar mesenchyme in Bmp4f/f;Wnt1Cre embryos. Msx1 heterozygosity enhanced maxillary molar developmental defects whereas Osr2 heterozygosity rescued mandibular first molar morphogenesis in Bmp4f/f;Wnt1Cre mice. Moreover, in contrast to complete lack of supernumerary tooth initiation in Msx1-/-Osr2-/- mutant mice, Osr2-/-Bmp4f/f;Wnt1Cre compound mutant mice exhibit formation and subsequent arrest of supernumerary tooth germs that correlated with down regulation of Msx1 expression in the tooth mesenchyme. Taken together, our data indicate that, while reduction in mesenchymal Bmp4 expression alone could not account for the tooth bud arrest phenotype in Msx1-/- mutant mice, Bmp4 signaling synergizes with Msx1 and antagonizes Osr2 to activate mesenchymal odontogenic activity to drive tooth morphogenesis and sequential tooth formation. E13.5 mouse embryos tooth germs were microdissected by laser capture microdissection (LCM), and the mandibular molar and maxillary molar were separated. 3 pairs of control and mutant samples were pooled for the RNA extraction.
2013-01-22 | E-GEOD-39918 | biostudies-arrayexpress
Project description:Mutations in MSX1 cause craniofacial developmental defects, including tooth agenesis, in humans and mice. Previous studies suggest that Msx1 activates Bmp4 expression in the developing tooth mesenchyme to drive early tooth organogenesis. Whereas Msx1−/− mice exhibit developmental arrest of all tooth germs at the bud stage, however, mice with neural crest-specific inactivation of Bmp4 (Bmp4ncko/ncko), which lack Bmp4 expression in the developing tooth mesenchyme, showed developmental arrest of only mandibular molars. We recently demonstrated that deletion of Osr2, which encodes a zinc finger transcription factor expressed in a lingual-to-buccal gradient in the developing tooth bud mesenchyme, rescued molar tooth morphogenesis in both Msx1−/− and Bmp4ncko/ncko mice. In this study, through RNA-seq analyses of the developing tooth mesenchyme in mutant and wildtype embryos, we found that Msx1 and Osr2 have opposite effects on expression of several secreted Wnt antagonists in the tooth bud mesenchyme. Remarkably, both Dkk2 and Sfrp2 exhibit Osr2-dependent preferential expression on the lingual side of the tooth bud mesenchyme and expression of both genes was up-regulated and expanded into the tooth bud mesenchyme in Msx1−/− and Bmp4ncko/ncko mutant embryos. We show that pharmacological activation of canonical Wnt signaling by either lithium chloride (LiCl) treatment or by inhibition of Dkk in utero was sufficient to rescue mandibular molar tooth morphogenesis in Bmp4ncko/ncko mice. Furthermore, whereas inhibition of Dkk alone was insufficient to rescue tooth morphogenesis in Msx1−/− mice, pharmacological inhibition of Dkk in combination with genetic inactivation of Sfrp2 and Sfrp3 rescued maxillary molar morphogenesis in Msx1−/− mice. Together, these data reveal a novel mechanism that the Bmp4-Msx1 pathway drives tooth organogenesis by activating Wnt signaling via regulation of the secreted Wnt antagonists.
2017-05-11 | GSE80288 | GEO
Project description:Mutations in MSX1 cause craniofacial developmental defects, including tooth agenesis, in humans and mice. Previous studies suggest that Msx1 activates Bmp4 expression in the developing tooth mesenchyme to drive early tooth organogenesis. Whereas Msx1−/− mice exhibit developmental arrest of all tooth germs at the bud stage, however, mice with neural crest-specific inactivation of Bmp4 (Bmp4ncko/ncko), which lack Bmp4 expression in the developing tooth mesenchyme, showed developmental arrest of only mandibular molars. We recently demonstrated that deletion of Osr2, which encodes a zinc finger transcription factor expressed in a lingual-to-buccal gradient in the developing tooth bud mesenchyme, rescued molar tooth morphogenesis in both Msx1−/− and Bmp4ncko/ncko mice. In this study, through RNA-seq analyses of the developing tooth mesenchyme in mutant and wildtype embryos, we found that Msx1 and Osr2 have opposite effects on expression of several secreted Wnt antagonists in the tooth bud mesenchyme. Remarkably, both Dkk2 and Sfrp2 exhibit Osr2-dependent preferential expression on the lingual side of the tooth bud mesenchyme and expression of both genes was up-regulated and expanded into the tooth bud mesenchyme in Msx1−/− and Bmp4ncko/ncko mutant embryos. We show that pharmacological activation of canonical Wnt signaling by either lithium chloride (LiCl) treatment or by inhibition of Dkk in utero was sufficient to rescue mandibular molar tooth morphogenesis in Bmp4ncko/ncko mice. Furthermore, whereas inhibition of Dkk alone was insufficient to rescue tooth morphogenesis in Msx1−/− mice, pharmacological inhibition of Dkk in combination with genetic inactivation of Sfrp2 and Sfrp3 rescued maxillary molar morphogenesis in Msx1−/− mice. Together, these data reveal a novel mechanism that the Bmp4-Msx1 pathway drives tooth organogenesis by activating Wnt signaling via regulation of the secreted Wnt antagonists.
2017-05-12 | GSE81282 | GEO