ABSTRACT
There is a growing amount of data uncovering the cellular diversity of the pulmonary circulation and mechanisms governing vascular repair after injury. However, the molecular and cellular mechanisms contributing to the morphogenesis and growth of the pulmonary vasculature during embryonic development are less clear. Importantly, deficits in vascular development lead to significant pediatric lung diseases, indicating a need to uncover fetal programs promoting vascular growth. To address this, we used a transgenic mouse reporter for expression of Cxcl12, an arterial endothelial hallmark gene, and performed single-cell RNA sequencing on isolated Cxcl12-DsRed+ endothelium to assess cellular heterogeneity within pulmonary endothelium. Combining cell annotation with gene ontology and histological analysis allowed us to segregate the developing artery endothelium into functionally and spatially distinct subpopulations. Expression of Cxcl12 is highest in the distal arterial endothelial subpopulation, a compartment enriched in genes for vascular development. Accordingly, disruption of CXCL12 signaling led to, not only abnormal branching, but also distal vascular hypoplasia. These data provide evidence for arterial endothelial functional heterogeneity and reveal conserved signaling mechanisms essential for pulmonary vascular development.
Subject(s)
Endothelium, Vascular , Lung , Mice , Pregnancy , Animals , Female , Endothelium, Vascular/metabolism , Morphogenesis , Mice, Transgenic , Embryonic DevelopmentABSTRACT
During the stepwise specification and differentiation of tissue-specific multipotent progenitors, lineage-specific transcriptional networks are activated or repressed to orchestrate cell specification. The gas-exchange niche in the lung contains two major epithelial cell types, alveolar type 1 (AT1) and AT2 cells, and the timing of lineage specification of these cells is critical for the correct formation of this niche and postnatal survival. Integrating cell-specific lineage tracing studies, spatially specific mRNA transcript and protein expression, and single-cell RNA-sequencing analysis, we demonstrate that specification of alveolar epithelial cell fate begins concomitantly with the proximal-distal specification of epithelial progenitors and branching morphogenesis earlier than previously appreciated. By using a newly developed dual-lineage tracing system, we show that bipotent alveolar cells that give rise to AT1 and AT2 cells are a minor contributor to the alveolar epithelial population. Furthermore, single-cell assessment of the transcriptome identifies specified AT1 and AT2 progenitors rather than bipotent cells during sacculation. These data reveal a paradigm of organ formation whereby lineage specification occurs during the nascent stages of development coincident with broad tissue-patterning processes, including axial patterning of the endoderm and branching morphogenesis.
Subject(s)
Cell Lineage , Lung/cytology , Pulmonary Alveoli/cytology , Animals , Cell Differentiation , Female , In Situ Hybridization, Fluorescence , Mice , Pregnancy , TranscriptomeABSTRACT
BACKGROUND: In today's world, antibiotic-resistant microorganisms are a major concern. There is solid evidence that metal nanoparticles (NPs) tend to have antimicrobial properties. The most effective substitute for antibiotic resistance is the incorporation of metal NPs. The antibacterial properties of NPs are currently being explored and shown to be successful. Zinc (Zn) NPs that are biosynthesized from marine Actinobacterium proved to be more biocompatible, bioactive, and affordable. Aim: This study aims to investigate the synthesis of ZnNPs from Actinobacterium Streptomyces species and their antimicrobial effects against gram-positive and gram-negative bacteria. MATERIALS AND METHODS: The current study uses natural, considerably safer processes to synthesize ZnNPs from marine Actinobacteria with little to no negative side effects. It involves sample collection, identification, and isolation of Actinobacterium Streptomyces species. The isolated sample was air-dried, and extracts of ZnNPs were taken. Among the isolates from marine sediment, two Actinobacteria that generate bioactive secondary metabolites-Streptomyces species (MOSEL-ME28) and Rhodococcus rhodochrous (MOSEL-ME29)-were selected for extracellular synthesis of ZnNPs. The antimicrobial activity of the biosynthesized ZnNPs from marine Actinobacteria was analyzed against Staphylococcus (MRSA), Klebsiella pneumoniae, and Streptococcus mutans. The results were statistically analyzed and graphs were created. RESULTS: ZnNPs obtained from Actinobacterium Streptomyces species exhibited antimicrobial effects against Staphylococcus (MRSA), Klebsiella, and Streptococcus mutans. At 280 nm wavelength, analysis of the UV spectrum showed a notable absorbance value of 1.8. The antibacterial efficacy against Staphylococcus MRSA, Klebsiella species, and Streptococcus mutans was assessed by measuring the zone of inhibition in diameter. The zones of inhibition were 8, 8, and 7 mm on the evaluation for Streptococcus mutans, S. aureus, and Klebsiella species, respectively, at a dose of 75 µg/mL. When the dosage was increased to 100 µg/mL, the inhibition zones were found to be 9.5, 9, and 7.5 mm for the respective bacterial strains. CONCLUSION: ZnNPs are biosynthesized from marine Actinobacterium Streptomyces species in this research study. They have a significant antimicrobial activity against both gram-positive and negative bacteria. This indicates that ZnNPs have enormous antimicrobial potential and have an extensive spectrum of applications. However, clinical trials must be completed before it can be used safely on patients.
ABSTRACT
Hyaluronan (HA) accumulates in the extracellular matrix to regulate organ morphogenesis. The spatiotemporal dynamics of its production and function are poorly understood due to its instability. Here, we present a protocol using the embryonic chicken intestine as a binary in vivo system for HA visualization and manipulation. We describe steps for pharmacological manipulation and in ovo electroporation to target HA production and accumulation. We then detail HA-binding protein assay to detect HA accumulation and quantification of tissue morphology changes. For complete details on the use and execution of this protocol, please refer to Sivakumar et al. (2018)1 and Sanketi et al. (2022).2.
ABSTRACT
BACKGROUND: Foot eczema in children is a commonly encountered condition but is associated with diverse etiology, thereby posing a diagnostic challenge. These include atopic eczema, juvenile plantar dermatoses, irritant contact dermatitis, and allergic contact dermatitis. Because of the diverse etiology, it is often difficult to distinguish between the conditions clinically, thereby warranting patch testing for the correct diagnosis. OBJECTIVES: The study was conducted to determine the frequency of positive patch tests among children with foot eczema and to compare them with children with and without atopy. METHODS: This was a prospective observational study done on 86 children aged 4 to 17 years presenting with foot eczema in whom patch testing was done using Indian standard series and footwear series and patients' own footwear. RESULTS: The clinical diagnosis of foot eczema in our study was allergic contact dermatitis (37%), followed by atopic eczema (30%), juvenile plantar dermatoses (17%), and lichen simplex chronicus (15%). Patch test positivity was present in 36% of the cases with clinical relevance in 65% of the cases. The common allergens were rubber allergens and disperse dyes. Of the children who were patch tested, highest patch test positivity rates were seen in allergic contact dermatitis (50%) and juvenile plantar dermatoses (53%). No significant differences were observed with regard to patch test positivity or relevance between children with and without atopy. CONCLUSIONS: The diagnosis of foot eczema in children cannot be made on clinical grounds alone, and patch testing should be recommended in children with any morphological pattern of foot eczema irrespective of presence or absence of atopy, especially in those with long-standing and recalcitrant disease with frequent exacerbations.
Subject(s)
Dermatitis, Allergic Contact , Dermatitis, Atopic , Eczema , Foot Diseases , Allergens/adverse effects , Child , Coloring Agents , Dermatitis, Allergic Contact/diagnosis , Dermatitis, Allergic Contact/etiology , Dermatitis, Atopic/complications , Dermatitis, Atopic/diagnosis , Eczema/diagnosis , Eczema/etiology , Humans , Irritants , Patch Tests/adverse effects , RubberABSTRACT
Regeneration of the architecturally complex alveolar niche of the lung requires precise temporal and spatial control of epithelial cell behavior. Injury can lead to a permanent reduction in gas exchange surface area and respiratory function. Using mouse models, we show that alveolar type 1 (AT1) cell plasticity is a major and unappreciated mechanism that drives regeneration, beginning in the early postnatal period during alveolar maturation. Upon acute neonatal lung injury, AT1 cells reprogram into alveolar type 2 (AT2) cells, promoting alveolar regeneration. In contrast, the ability of AT2 cells to regenerate AT1 cells is restricted to the mature lung. Unbiased genomic assessment reveals that this previously unappreciated level of plasticity is governed by the preferential activity of Hippo signaling in the AT1 cell lineage. Thus, cellular plasticity is a temporally acquired trait of the alveolar epithelium and presents an alternative mode of tissue regeneration in the postnatal lung.
Subject(s)
Alveolar Epithelial Cells , Lung , Animals , Homeostasis , Mice , Respiratory Mucosa , Signal TransductionABSTRACT
Lymphangioleiomyomatosis (LAM) is a rare fatal cystic lung disease due to bi-allelic inactivating mutations in tuberous sclerosis complex (TSC1/TSC2) genes coding for suppressors of the mechanistic target of rapamycin complex 1 (mTORC1). The origin of LAM cells is still unknown. Here, we profile a LAM lung compared to an age- and sex-matched healthy control lung as a hypothesis-generating approach to identify cell subtypes that are specific to LAM. Our single-cell RNA sequencing (scRNA-seq) analysis reveals novel mesenchymal and transitional alveolar epithelial states unique to LAM lung. This analysis identifies a mesenchymal cell hub coordinating the LAM disease phenotype. Mesenchymal-restricted deletion of Tsc2 in the mouse lung produces a mTORC1-driven pulmonary phenotype, with a progressive disruption of alveolar structure, a decline in pulmonary function, increase of rapamycin-sensitive expression of WNT ligands, and profound female-specific changes in mesenchymal and epithelial lung cell gene expression. Genetic inactivation of WNT signaling reverses age-dependent changes of mTORC1-driven lung phenotype, but WNT activation alone in lung mesenchyme is not sufficient for the development of mouse LAM-like phenotype. The alterations in gene expression are driven by distinctive crosstalk between mesenchymal and epithelial subsets of cells observed in mesenchymal Tsc2-deficient lungs. This study identifies sex- and age-specific gene changes in the mTORC1-activated lung mesenchyme and establishes the importance of the WNT signaling pathway in the mTORC1-driven lung phenotype.
Subject(s)
Lung/metabolism , Lymphangioleiomyomatosis/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , Mesoderm/metabolism , Age Factors , Aged , Animals , Female , Humans , Lung/drug effects , Lung/physiopathology , Lymphangioleiomyomatosis/drug therapy , Lymphangioleiomyomatosis/genetics , Lymphangioleiomyomatosis/physiopathology , Male , Mechanistic Target of Rapamycin Complex 1/genetics , Mesoderm/drug effects , Mice , Sex Factors , Sirolimus/administration & dosage , Tuberous Sclerosis Complex 2 Protein/genetics , Tuberous Sclerosis Complex 2 Protein/metabolism , Wnt Signaling PathwayABSTRACT
Alveolar epithelial regeneration is essential for recovery from devastating lung diseases. This process occurs when type II alveolar pneumocytes (AT2 cells) proliferate and transdifferentiate into type I alveolar pneumocytes (AT1 cells). We used genome-wide analysis of chromatin accessibility and gene expression following acute lung injury to elucidate repair mechanisms. AT2 chromatin accessibility changed substantially following injury to reveal STAT3 binding motifs adjacent to genes that regulate essential regenerative pathways. Single-cell transcriptome analysis identified brain-derived neurotrophic factor (Bdnf) as a STAT3 target gene with newly accessible chromatin in a unique population of regenerating AT2 cells. Furthermore, the BDNF receptor tropomyosin receptor kinase B (TrkB) was enriched on mesenchymal alveolar niche cells (MANCs). Loss or blockade of AT2-specific Stat3, Bdnf or mesenchyme-specific TrkB compromised repair and reduced Fgf7 expression by niche cells. A TrkB agonist improved outcomes in vivo following lung injury. These data highlight the biological and therapeutic importance of the STAT3-BDNF-TrkB axis in orchestrating alveolar epithelial regeneration.
Subject(s)
Alveolar Epithelial Cells/cytology , Brain-Derived Neurotrophic Factor/metabolism , Lung Injury/prevention & control , Membrane Glycoproteins/metabolism , Protein-Tyrosine Kinases/metabolism , Receptor, trkB/metabolism , Regeneration , STAT3 Transcription Factor/metabolism , Alveolar Epithelial Cells/metabolism , Animals , Brain-Derived Neurotrophic Factor/genetics , Female , Humans , Lung Injury/etiology , Lung Injury/pathology , Male , Membrane Glycoproteins/genetics , Protein-Tyrosine Kinases/genetics , Receptor, trkB/genetics , STAT3 Transcription Factor/geneticsABSTRACT
PURPOSE OF REVIEW: Throughout the lifespan, lung injury impedes the primary critical function essential for life-respiration. To repair quickly and efficiently is critical and is orchestrated by a diverse repertoire of progenitor cells and their niche. This review incorporates knowledge gained from early studies in lung epithelial morphogenesis and cell fate and explores its relevance to more recent findings of lung progenitor and stem cells in development and regeneration. RECENT FINDINGS: Cell fate in the lung is organized into an early specification phase and progressive differentiation phase in lung development. The advent of single cell analysis combined with lineage analysis and projections is uncovering new functional cell types in the lung providing a topographical atlas for progenitor cell lineage commitment during development, homeostasis, and regeneration. SUMMARY: Lineage commitment of lung progenitor cells is spatiotemporally regulated during development. Single cell sequencing technologies have significantly advanced our understanding of the similarities and differences between developmental and regenerative cell fate trajectories. Subsequent unraveling of the molecular mechanisms underlying these cell fate decisions will be essential to manipulating progenitor cells for regeneration.
ABSTRACT
This case report describes a woman in her 20s with painful, red skin lesions present for 6 months that had gradually progressed from the groin to other sites.
Subject(s)
Acrodermatitis , Zinc , Humans , Acrodermatitis/diagnosisABSTRACT
The emerging field of transcriptional regulation of cell shape changes aims to address the critical question of how gene expression programs produce a change in cell shape. Together with cell growth, division, and death, changes in cell shape are essential for organ morphogenesis. Whereas most studies of cell shape focus on posttranslational events involved in protein organization and distribution, cell shape changes can be genetically programmed. This review highlights the essential role of transcriptional regulation of cell shape during morphogenesis of the heart, lungs, gastrointestinal tract, and kidneys. We emphasize the evolutionary conservation of these processes across different model organisms and discuss perspectives on open questions and research avenues that may provide mechanistic insights toward understanding birth defects.
Subject(s)
Cell Shape/physiology , Gastrointestinal Tract/embryology , Heart/embryology , Kidney/embryology , Lung/embryology , Organogenesis/physiology , Animals , Congenital Abnormalities/embryology , Gene Expression Regulation, Developmental , Humans , Transcription, Genetic/geneticsABSTRACT
For many years, biologists have focused on the role of Pitx2, expressed on the left side of developing embryos, in governing organ laterality. Here, we identify a different pathway during left-right asymmetry initiated by the right side of the embryo. Surprisingly, this conserved mechanism is orchestrated by the extracellular glycosaminoglycan, hyaluronan (HA) and is independent of Pitx2 on the left. Whereas HA is normally synthesized bilaterally as a simple polysaccharide, we show that covalent modification of HA by the enzyme Tsg6 on the right triggers distinct cell behavior necessary to drive the conserved midgut rotation and to pattern gut vasculature. HA disruption in chicken and Tsg6-/- mice results in failure to initiate midgut rotation and perturbs vascular development predisposing to midgut volvulus. Our study leads us to revise the current symmetry-breaking paradigm in vertebrates and demonstrates how enzymatic modification of HA matrices can execute the blueprint of organ laterality.