scholarly journals 2566 Personalized models of distal airway epithelial-stromal unit in COPD

2018 ◽  
Vol 2 (S1) ◽  
pp. 23-23
Author(s):  
Seyed B. Mahjour ◽  
Kazunori Gomi ◽  
Busub Lee ◽  
Olivier Elemento ◽  
Scott Randell ◽  
...  
Keyword(s):  
Cross Talk ◽  
Stromal Cells ◽  
Airway Remodeling ◽  
Squamous Metaplasia ◽  
Mucous Cell ◽  
Patient Specific ◽  
Pcr Analysis ◽  
Airway Epithelial ◽  
Chronic Lung Diseases ◽  
Distal Airway

OBJECTIVES/SPECIFIC AIMS: The objective of this study is to develop patient-derived “personalized” organotypic models of human distal airways, in which basal stem cells (BCs) isolated from the pre-/terminal conducting airway region are co-cultured with autologous stromal cells from the same region to reproduce patient-specific distal airway epithelial-stromal units and their remodeling in COPD. METHODS/STUDY POPULATION: We established a protocol to isolate and propagate epithelial BCs, fibroblasts, and endothelial cells from the distal airways of normal and COPD lung donors. Heterogeneous cellular and molecular phenotypes in the human distal airways were characterized using immunofluorescence and single-cell RNA sequencing. Patient-specific distal airway epithelial-stromal units were reconstructed by co-culturing BCs and autologous stromal cells using an air-liquid interface-based airway wall model and a bronchosphere-based 3D distal airway organoid assay. RESULTS/ANTICIPATED RESULTS: Histologic analysis of derived epithelial-stromal units revealed heterogeneous patient-specific phenotypes characterized by hypo-/hyper-/metaplastic lesions (hypo-regenerative phenotype, mucous cell hyperplasia, squamous metaplasia, distal-to-proximal repatterning) in the epithelial compartment, accompanied, in some samples, by stromal remodeling. Candidate epithelial-stromal cross-talk mechanisms were identified using quantitative real-time RT-PCR analysis of autologous epithelial and stromal compartments of established patient-specific distal airway unit models. DISCUSSION/SIGNIFICANCE OF IMPACT: Epithelial and stromal cells isolated from distal airways of subjects with and without COPD can be assembled into functional, organ-level tissue which mimics the architecture of human distal airways and, in patients with COPD, reproduces several distal airway remodeling phenotypes. Patient-specific models of distal airway epithelial-stromal cross-talk established in this study can be used to identify candidate pathways that mediate disease-relevant airway remodeling and potentially utilized as pre-clinical platforms for developing personalized therapeutic approaches to suppress the progression of distal airway remodeling in chronic lung diseases, including COPD.

scholarly journals 3055 Reconstruction of Patient-specific Distal Airway Regeneration Patterns in COPD

2019 ◽  
Vol 3 (s1) ◽  
pp. 154-154
Author(s):  
Seyed Babak Mahjour ◽  
Kazunori Gomi ◽  
Samir Rustam ◽  
Phurbu Dolma ◽  
Jamuna Krishnan ◽  
...  
Keyword(s):  
Single Cell ◽  
Stromal Cells ◽  
Airway Remodeling ◽  
Expression Patterns ◽  
Mucous Cell ◽  
Patient Specific ◽  
Mucin 1 ◽  
Cell Hyperplasia ◽  
Chronic Lung Diseases ◽  
Distal Airway

OBJECTIVES/SPECIFIC AIMS: The objective of this study was to reconstruct patient-specific distal airway patterns at the tissue- and single-cell resolution and develop personalized distal airway models based on utilization of patient-derived DABCs and autologous region-specific stromal cells. METHODS/STUDY POPULATION: Patient-specific distal airway units, containing parental small bronchiole (<2 mm in diameter, >12th generation) and daughter airway branches, including pre-terminal/terminal bronchioles, leading to alveoli (3-7 units/lung), were dissected. Epithelial and stromal cells were isolated from these units and processed for ddSeq single-cell RNA-sequencing (n=6 samples). Autologous DABCs and stromal cells were isolated, propagated, biobanked, and used for establishment of patient-specific distal airway models (3D-organoids and air-liquid interface-based airway wall model; n=10 samples). Region-specific tissue patterns were evaluated using immunofluorescence and laser-capture microdissection (LCM; n=6 samples). RESULTS/ANTICIPATED RESULTS: Single-cell-based human distal airway transcriptome map (constructed based on the analysis of >6,500 distal airway cells obtained from 6 subjects) identified physiological and COPD-relevant distal airway differentiation patterns, including distal airway-specific secretory phenotype (DASP) characterized with high expression of secretoglobins 3A2 and 3A1, surfactant proteins SFTPB and SFTPA2, and mucin 1, unique signatures of DABCs, and stromal (fibroblasts, smooth muscle, endothelial cell subpopulations) and immune (macrophage, T cells, B cell, mast cells). Immunofluorescence analysis and LCM confirmed distribution of cell type-specific markers with differential expression patterns of DABC and DASP signatures. Patient-derived DABC-stromal co-culture models reproduced 3 regenerative patterns: 1) physiological (high DABC-clonogenic potency, establishment of polarized differentiated organoids and DASP-expressing epithelia); 2) hypo-regenerative (failure of DABCs to form clones, spheres and mechanically stable differentiated epithelial barrier); and 3) hyperplastic (generation of DABC hyperplasia accompanied in some COPD samples by mucous-cell hyperplasia mimicking in vivo remodeling patterns). DISCUSSION/SIGNIFICANCE OF IMPACT: Patient-specific maps and models of distal airway regeneration patterns have been established in this study, which can be used to identify candidate pathways that mediate disease-relevant airway remodeling and potentially utilized as pre-clinical platforms for developing personalized therapeutic approaches to suppress the progression of distal airway remodeling in chronic lung diseases, including COPD.

scholarly journals Pulmonary fibrosis distal airway epithelia are dynamically and structurally dysfunctional

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ian T. Stancil ◽  
Jacob E. Michalski ◽  
Duncan Davis-Hall ◽  
Hong Wei Chu ◽  
Jin-Ah Park ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Airway Epithelium ◽  
Cell Types ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Airway Epithelia ◽  
Chronic Lung Diseases ◽  
Distal Airway ◽  
Signaling Axis

AbstractThe airway epithelium serves as the interface between the host and external environment. In many chronic lung diseases, the airway is the site of substantial remodeling after injury. While, idiopathic pulmonary fibrosis (IPF) has traditionally been considered a disease of the alveolus and lung matrix, the dominant environmental (cigarette smoking) and genetic (gain of function MUC5B promoter variant) risk factor primarily affect the distal airway epithelium. Moreover, airway-specific pathogenic features of IPF include bronchiolization of the distal airspace with abnormal airway cell-types and honeycomb cystic terminal airway-like structures with concurrent loss of terminal bronchioles in regions of minimal fibrosis. However, the pathogenic role of the airway epithelium in IPF is unknown. Combining biophysical, genetic, and signaling analyses of primary airway epithelial cells, we demonstrate that healthy and IPF airway epithelia are biophysically distinct, identifying pathologic activation of the ERBB-YAP axis as a specific and modifiable driver of prolongation of the unjammed-to-jammed transition in IPF epithelia. Furthermore, we demonstrate that this biophysical state and signaling axis correlates with epithelial-driven activation of the underlying mesenchyme. Our data illustrate the active mechanisms regulating airway epithelial-driven fibrosis and identify targets to modulate disease progression.

scholarly journals Membrane-bound mucins of the airway mucosal surfaces are densely decorated with keratan sulfate: revisiting their role in the Lung’s innate defense

Glycobiology ◽  
2020 ◽  
Author(s):  
Jerome Carpenter ◽  
Mehmet Kesimer
Keyword(s):  
Light Scattering ◽  
Keratan Sulfate ◽  
Mucosal Barrier ◽  
Airway Epithelial ◽  
Surface Binding ◽  
Force Microscopy ◽  
Innate Defense ◽  
Chronic Lung Diseases ◽  
Mucosal Surfaces ◽  
Membrane Bound

Abstract Understanding the basic elements of the airway mucosal surfaces and how they form a functional barrier is essential in understanding disease initiation, progression, pathogenesis and ultimately treating chronic lung diseases. Using primary airway epithelial cell cultures, atomic force microscopy (AFM), multiangle light scattering and quartz crystal micro balance with dissipation monitoring techniques, here we report that the membrane bound mucins (MBMs) found in the periciliary layer (PCL) of the airway surface are densely decorated with keratan sulfate (KS). AFM and immunoblotting show that the KS sidechains can be removed enzymatically with keratanase II (KII) treatment, and the antibody accessibility for B2729 (MUC1), MUCH4 (MUC4) and OC125 (MUC16) was substantially enhanced. Light scattering analysis confirmed that KII treatment removed ~40% of the mass from the mucin fractions. Surface binding experiments indicated that MBMs were able to pack into a tighter conformation following KS removal, suggesting that negatively charged KS sidechains play a role in mucin–mucin repulsion and contribute to “space filling” in the PCL. We also observed that soluble filtrate from the common airway pathogen Pseudomonas aeruginosa is capable of stripping KS from MBMs. Altogether, our findings indicate that KS glycosylation of MBMs may play an important role in the integrity of the airway mucosal barrier and its compromise in disease.

scholarly journals Cigarette Smoke Condensate Exposure Changes RNA Content of Extracellular Vesicles Released from Small Airway Epithelial Cells

Cells ◽  
2019 ◽  
Vol 8 (12) ◽  
pp. 1652
Author(s):  
Tiziana Corsello ◽  
Andrzej S. Kudlicki ◽  
Roberto P. Garofalo ◽  
Antonella Casola
Keyword(s):  
Epithelial Cells ◽  
Cigarette Smoke ◽  
Extracellular Vesicles ◽  
Viral Infections ◽  
Cell Communication ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Cigarette Smoke Condensate ◽  
Diverse Range ◽  
Chronic Lung Diseases

Exposure to environmental tobacco smoke (ETS) is a known risk factor for the development of chronic lung diseases, cancer, and the exacerbation of viral infections. Extracellular vesicles (EVs) have been identified as novel mediators of cell–cell communication through the release of biological content. Few studies have investigated the composition/function of EVs derived from human airway epithelial cells (AECs) exposed to cigarette smoke condensate (CSC), as surrogates for ETS. Using novel high-throughput technologies, we identified a diverse range of small noncoding RNAs (sncRNAs), including microRNA (miRNAs), Piwi-interacting RNA (piRNAs), and transfer RNA (tRNAs) in EVs from control and CSC-treated SAE cells. CSC treatment resulted in significant changes in the EV content of miRNAs. A total of 289 miRNAs were identified, with five being significantly upregulated and three downregulated in CSC EVs. A total of 62 piRNAs were also detected in our EV preparations, with five significantly downregulated and two upregulated in CSC EVs. We used TargetScan and Gene Ontology (GO) analysis to predict the biological targets of hsa-miR-3913-5p, the most represented miRNA in CSC EVs. Understanding fingerprint molecules in EVs will increase our knowledge of the relationship between ETS exposure and lung disease, and might identify potential molecular targets for future treatments.

Reconstruction of Patient-Specific Distal Airway Regeneration Patterns in COPD

2019 ◽  
Author(s):  
S.B. Mahjour ◽  
K. Gomi ◽  
S. Rustam ◽  
P. Dolma ◽  
J. Krishnan ◽  
...  
Keyword(s):  
Patient Specific ◽  
Distal Airway

scholarly journals Heparin Anticoagulant for Human Bone Marrow Does Not Influence In Vitro Performance of Human Mesenchymal Stromal Cells

Cells ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 1580
Author(s):  
Yvonne Roger ◽  
Laura Burmeister ◽  
Anika Hamm ◽  
Kirsten Elger ◽  
Oliver Dittrich-Breiholz ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Mononuclear Cells ◽  
Population Doubling ◽  
Pcr Analysis ◽  
Inhibitory Influence ◽  
In Vitro Differentiation ◽  
Cell Surface Antigens

Mesenchymal stromal cells (MSCs) are a promising cell source for tissue engineering and regenerative medicine. In our lab, we found that MSC preparations from bone marrow of many different donors had a limited capacity of in vitro differentiation into osteogenic and chondrogenic lineages—a capacity claimed to be inherent to MSCs. The current study was designed to test the hypothesis that the amount of heparin used as anticoagulant during bone marrow harvest had an inhibitory influence on the in vitro differentiation capacity of isolated MSCs. Bone marrow was obtained from the femoral cavity of twelve donors during total hip arthroplasty in the absence or presence of heparin. No coagulation was observed in the absence of heparin. The number of mononuclear cells was independent of heparin addition. Isolated MSCs were characterized by morphology, population doubling times, expression of cell surface antigens and in vitro differentiation. Results of these analyses were independent of the amount of heparin. Transcriptome analyses of cells from three randomly chosen donors and quantitative realtime PCR (qRT-PCR) analysis from cells of all donors demonstrated no clear effect of heparin on the transcriptome of the cells. This excludes heparin as a potential source of disparate results.

scholarly journals Prognostic Roles of Cross-Talk between Peritumoral Hepatocytes and Stromal Cells in Hepatocellular Carcinoma Involving Peritumoral VEGF-C, VEGFR-1 and VEGFR-3

PLoS ONE ◽  
2013 ◽  
Vol 8 (5) ◽  
pp. e64598 ◽  
Author(s):  
Peng-Yuan Zhuang ◽  
Jun Shen ◽  
Xiao-Dong Zhu ◽  
Lu Lu ◽  
Lu Wang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cross Talk ◽  
Stromal Cells

266. Supressor of cytokine signalling 3 regulates IL-11 induced human endometrial stromal cell decidualization

2005 ◽  
Vol 17 (9) ◽  
pp. 109
Author(s):  
E. Dimitriadis ◽  
C. Stoikos ◽  
L. A. Salamonsen
Keyword(s):  
Stromal Cells ◽  
Embryo Implantation ◽  
Cellular Protein ◽  
Pcr Analysis ◽  
Cytokine Signaling ◽  
Pituitary Cells ◽  
Human Pituitary ◽  
Endometrial Stromal Cells ◽  
Socs3 Protein

Decidualization of endometrial stromal cells is critical for embryo implantation and establishment of pregnancy. Locally produced cytokines such as interleukin (IL)-11 enhance decidualization of human endometrial stromal cells (HESC). IL-11 signaling is negatively regulated by suppressor of cytokine signaling (SOCS) proteins. IL-11 stimulates SOCS3 in human pituitary cells. The aim of this study was to examine the role of SOCS3 on IL-11 induced HESC decidualization. Decidualization of HESC was assessed using an in vitro model in which estrogen (E)+progesterone (P) or cAMP was administered for 8 days to cells. Medium was collected for prolactin (PRL) assay (a decidual marker). Cellular protein was extracted for Western analysis and cellular RNA for real-time RT-PCR analysis. SOCS3 was overexpressed in HESC cells and the effect on decidualization assessed. HESC treated with E+P or cAMP secreted PRL from day 6. Treatment of HESC with E+P or cAMP increased the abundance of SOCS3 protein, coinciding with an increase in PRL secretion. cAMP maximally stimulated SOCS3 protein and mRNA during decidualization. Antiprogestin (onapristone) added to E+P or cAMP treated cells at day 6 reduced PRL secretion but had no influence on SOCS3 abundance suggesting that SOCS3 protein was not regulated via the P-receptor pathway. Addition of IL-11 to HESC increased SOCS3 abundance from 1 h. SOCS3 abundance returned to control levels following treatment of cells with IL-11 and IL-11 neutralising antibody. SOCS3 overexpression in HESC treated with cAMP reduced PRL secretion compared to mock- or non-transfected HESC. Furthermore, IL-11 mediated decidualization was diminished by SOCS3 overexpression. We have demonstrated for the first time that SOCS3 regulates IL-11 induced decidualization and that SOCS3 overexpression in HESC disrupts decidualization. This knowledge is important in understanding the mechanisms by which IL-11 promotes decidualization of HESC and thus the formation of decidua, an essential component of a functional placenta.

scholarly journals Derivation of induced pluripotent stem cells from ferret somatic cells

2020 ◽  
Vol 318 (4) ◽  
pp. L671-L683
Author(s):  
Jinghui Gao ◽  
Sophia Petraki ◽  
Xingshen Sun ◽  
Leonard A. Brooks ◽  
Thomas J. Lynch ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Patient Specific ◽  
Preclinical Model ◽  
Specific Cell ◽  
Chronic Lung Diseases ◽  
Human Ipscs ◽  
Fetal Fibroblasts ◽  
Induced Pluripotent

Ferrets are an attractive mammalian model for several diseases, especially those affecting the lungs, liver, brain, and kidneys. Many chronic human diseases have been difficult to model in rodents due to differences in size and cellular anatomy. This is particularly the case for the lung, where ferrets provide an attractive mammalian model of both acute and chronic lung diseases, such as influenza, cystic fibrosis, A1A emphysema, and obliterative bronchiolitis, closely recapitulating disease pathogenesis, as it occurs in humans. As such, ferrets have the potential to be a valuable preclinical model for the evaluation of cell-based therapies for lung regeneration and, likely, for other tissues. Induced pluripotent stem cells (iPSCs) provide a great option for provision of enough autologous cells to make patient-specific cell therapies a reality. Unfortunately, they have not been successfully created from ferrets. In this study, we demonstrate the generation of ferret iPSCs that reflect the primed pluripotent state of human iPSCs. Ferret fetal fibroblasts were reprogrammed and acquired core features of pluripotency, having the capacity for self-renewal, multilineage differentiation, and a high-level expression of the core pluripotency genes and pathways at both the transcriptional and protein level. In conclusion, we have generated ferret pluripotent stem cells that provide an opportunity for advancing our capacity to evaluate autologous cell engraftment in ferrets.

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