Project description:Gene expression patterns of bronchiolar progenitors and club cells in mouse lung were examined by microarray experiments. Although it has not yet been fully characterized, a subset of epithelial cells lining bronchioles are best understood as bronchiolar progenitors that self-renew over the long term and that can differentiate into more differentiated club cells and ciliated cells. The bronchiolar progenitors are distinct from club cells and characteristically express the alveolar type 2 cell marker, prosurfactant protein C, with lower levels of club cell secretory protein/Scgb1a1. There are also functional differences between them; while club cells can be depleted by naphthalene because of the abundance of cytochrome P450 enzyme Cyp2f2, bronchiolar progenitors are resistant to naphthalene-induced depletion because of defects in the enzyme.
Project description:Peripheral blood mononuclear cells (PBMC) were collected from ficoll gradients of whole blood and immediately processed for total RNA and stored at -80oC until use. Blood was from allergic patients at different timepoints pre or post immunotherapy (RIT). Gene expression was compared for the different timepoints. Blood samples were collected from allergic patients before or after rush immunotherapy (RIT), total RNA was prepared and gene expression assessed by Illumina. Samples were collected pre-rush or at 1 week, 7 weeks, 4 months (and for one patient 5 months) after beginning rush immunotherapy. Blood was taken at each timepoint prior to the patient receiving immunotherapy at each timepoint.
Project description:The club cell, a small airway epithelial (SAE) secretory cell that uniquely expresses SCGB1A1, plays a central role in host defense in the human lung. Based on data demonstrating that ~50% of club cells express MUC5B, a secretory mucin critical for mucociliary clearance, we hypothesized that subpopulations of club cells with distinct functions may exist. To evaluate this, the SAE of normal nonsmokers and healthy cigarette smokers was sampled by bronchoscopy and brushing followed by single cell sequencing using Drop-seq technology. Subpopulations of SCGCB1A1+KRT5loMUC5AC- club cells were assessed by unsupervised clustering to evaluate club cell subpopulations. Immunostaining of SAE in lung sections, brushed SAE cells, and in vitro air-liquid interface culture was utilized to confirm the transcriptomic-based observations. Unsupervised clustering of SCGCB1A1+KRT5loMUC5AC‾ club cells in the SAE identified 3 unique club cell populations that differed by differentiation state and function, including: (1) progenitor; (2) proliferating; and (3) effector subpopulations. The progenitor club cell population was energetically active with high expression of mitochondrial and ribosomal proteins and the highest KRT5 levels vs other club cell populations. The proliferating population, defined by high expression of cyclins and proliferation markers, was the smallest, representing 2% of club cells. The effector club cell cluster expressed transcripts for host defense genes, xenobiotic metabo-lism, and barrier functions commonly associated with club cell function. Comparison of the club cell subpopulations in smokers vs nonsmokers demonstrated that the proportion of the club cell effector population was significantly decreased in smokers with a concomitant significant in-crease in the proliferating cell population. These observations provide novel insights into both the makeup of human SAE club cell subpopulations and smoking-induced changes in club cell biology.
Project description:Club (“Clara”) cells, dome shaped cells with disease cytoplasmic granules and microvilli, are the major secretory cell of the human small (>6 generation) airways. Little is known regarding the biology of club cells in the human airway, nor of the ontogeny of this cell type. Taking advantage of the SCGB1A1 as the marker for club cells and our ability to sample the normal small airway epithelium by bronchoscopy and brushing healthy volunteers, we defined the transcriptome of the normal human airway club cell using single cell transcriptome sequencing. Analysis of the human small airway epithelial single cell transcriptome together with in vitro validation provides novel insights into the molecular phenotype and biological functions of the human club cell population and identifies the basal cell as the human progenitor cells for club cells.