Project description:Introduction of Hermansky-Pudlak Syndrome-associated mutations with CRISPR/Cas9 genome editing, allows for disease modeling in 3D cultures of hPSC-derived lung organoids.
Project description:The human small airway epithelium (SAE) plays a central role in the early events in the pathogenesis of most chronic inherited and acquired lung disorders. Little is known about the molecular phenotypes of the specific cell populations comprising the SAE, and the contribution of specific cell populations to the pathogenesis of human disease. There was cell type-specific expression of the genes relevant to the pathogenesis of the inherited pulmonary disorders, genes associated with risk of chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF), and (non-mutated) driver genes for lung cancers. Some unexpected observations included high expression of CFTR (cystic fibrosis) and SCNN1A and B (bronchiectasis) in ionocytes, DTNBP1 (Hermansky-Pudlak syndrome) in mast cells, FAM13A (COPD) in neuroendocrine cells, RIN3 (COPD) in mast cells, NSPA1L (IPF) in neuroendocrine cells and a variety of lung cancer-related genes expressed in different cell types that, if mutated, become driver genes. Cigarette smoking significantly altered the cell-specific transcriptome of the different cell populations. Many of the genes relevant to the hereditary and acquired disorders exhibited cell-specific modulation by cigarette smoking, including MUC5B (IPF) down-regulation specifically in intermediate, club and mucus-producing cells and SFTPB (surfactant deficiency) up-regulation in ionocytes.
Project description:The longitudinal cellular interactions that drive pulmonary fibrosis are not well understood. To investigate the disease underpinnings associated with fibrosis onset and progression, we generated a scRNAseq atlas of lungs from young and aged mouse models of multiple subtypes of Hermansky-Pudlak syndrome (HPS), a collection of rare autosomal recessive diseases associated with albinism, platelet dysfunction, and pulmonary fibrosis. We identified an age-dependent increase in SAA3+ inflammatory lung fibroblasts in HPS mice, including in double-mutant HPS1-2 mice which develop spontaneous fibrosis. HPS1 fibroblasts showed increased expression of IL-1R1, whereas alveolar type II epithelial cells from HPS2 mice induced the inflammatory gene signature in co-cultured fibroblasts. scRNAseq of lung tissue from three HPS1 patients similarly showed the presence of inflammatory fibroblasts and increased IL1R1 expression on fibroblasts. These data posit complex interactions between dysfunctional epithelial cells, inflammatory fibroblasts, and recruited immune cells, suggesting potential opportunities for mitigation of the fibrotic cascade.
Project description:Hermansky-Pudlak syndrome type 1 (HPS-1) is a rare, autosomal recessive disorder caused by defects in the biogenesis of lysosome-related organelles complex-3 (BLOC-3). Impaired kidney function is among its clinical manifestations. To investigate HPS-1 renal involvement, we employed 1D-gel-LC–MS/MS and compared the protein composition of urinary extracellular vesicles (uEVs) from HPS-1 patients to normal control individuals. We identified 1029 proteins, 149 of which were altered in HPS-1 uEVs. Ingenuity Pathway Analysis revealed disruptions in mitochondrial function and the LXR/RXR pathway that regulates lipid metabolism, which is supported by our novel Hps1 knockout mouse. Serum concentration of the LXR/RXR pathway protein ApoA1 in our patient cohort was positively correlated with kidney function (with the estimated glomerular filtration rate or eGFR). uEVs can be used to study epithelial cell protein trafficking in HPS-1 and may provide outcome measures for HPS-1 therapeutic interventions.
