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|Disease Keywords||Lung, cancer, fibrosis|
|Relevance||Identification of putative human lung stem cells. HSLC are capable of expanding indefinitely and harbour both self-renewal capacity and the potency to differentiate in vitro and in vivo. Unlike previously identified putative human lung stem cells this HLSC cell line has a distinctive epithelial origin and as shown in kidney grafts these cells do not differentiate into mesenchymal or endothelial cells.|
|Production Details||Human lung tissue was obtained from patients undergoing lung resection. After pleura was separated bluntly, lung specimens were finely minced and resuspended in collagenase (0.5–3 mg/ml, Whorthington)/dispase (1 mg/ml, Invitrogen) containing DMEM (Invitrogen) and incubated for 30–45 min at 37°C in a shaking incubator. The suspension was spun for 5 min at 1200, r.p.m. and the supernatant removed. The pellet was resuspended in fresh DMEM containing 0.1 mg/ml DNase (optional) and incubated for further 5–10 min. The suspension was washed with PBS, filtered through cell strainers (100, 70 μm, BD) and treated with red blood cell lysis buffer (Roche Applied Science). Following further filtration (40 μm mesh) and centrifugation (5 min at 1200, r.p.m.), the isolated cells were cultured in RH‐B (Stem Cell Science) medium containing 2% FCS, with additional insulin (5 μg/ml, Pepro Tech), EGF (10 ng/ml, Pepro Tech) and FGF2 (20 ng/ml, Pepro Tech) for 2 days. This was then was replaced with fresh, serum‐free medium containing growth factors (37°C in a 7% humidified CO2 incubator).|
|Research Area||Stem Cell Biology|
|Growth/Phenotype Keywords||E-Cadlow/Lgr6+ HSLC cells grow clonally forming aggregates that have been successfully expanded in vitro for over 50 passages expressing lung specific (SP-C, CC-10, AQ4 5), epithelial (E-Cad), and stem cell markers (Sox9, Lgr5/6, Integrin-beta6).|
|Recommended Growing Conditions||Serum‐free basal stem cell culture medium containing growth factors FGF (10 ng ml-1) and EGF (20 ng ml-1) (37°C in a 7% humidified CO2 incubator)|
HSLC are capable of regenerating damaged lung tissue in vivo.
They are characterised by low expression of E-Cadlow and positivity to Lgr6 marker.
The exact area where the biopsy was taken is unknown, the cells were extracted from a piece of tissue containing mostly alveoli but also some small bronchioles and of course all the surrounding tissue.
They are not derived from tracheal/bronchial tissue.
The pool of lung cells were negative selected for CD45/CD31/aSMA and then positive selection Lgr6/E-Cad. Later on we used other markers to determine their profile and characterise.
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