Preclinical validation and imaging of Wnt-induced repair in human 3D lung tissue cultures.

The European respiratory journal (2015-05-02)
Franziska E Uhl, Sarah Vierkotten, Darcy E Wagner, Gerald Burgstaller, Rita Costa, Ina Koch, Michael Lindner, Silke Meiners, Oliver Eickelberg, Melanie Königshoff
RESUMEN

Chronic obstructive pulmonary disease (COPD) is characterised by a progressive loss of lung tissue. Inducing repair processes within the adult diseased lung is of major interest and Wnt/β-catenin signalling represents a promising target for lung repair. However, the translation of novel therapeutic targets from model systems into clinical use remains a major challenge.We generated murine and patient-derived three-dimensional (3D) ex vivo lung tissue cultures (LTCs), which closely mimic the 3D lung microenvironment in vivo. Using two well-known glycogen synthase kinase-3β inhibitors, lithium chloride (LiCl) and CHIR 99021 (CT), we determined Wnt/β-catenin-driven lung repair processes in high spatiotemporal resolution using quantitative PCR, Western blotting, ELISA, (immuno)histological assessment, and four-dimensional confocal live tissue imaging.Viable 3D-LTCs exhibited preserved lung structure and function for up to 5 days. We demonstrate successful Wnt/β-catenin signal activation in murine and patient-derived 3D-LTCs from COPD patients. Wnt/β-catenin signalling led to increased alveolar epithelial cell marker expression, decreased matrix metalloproteinase-12 expression, as well as altered macrophage activity and elastin remodelling. Importantly, induction of surfactant protein C significantly correlated with disease stage (per cent predicted forced expiratory volume in 1 s) in patient-derived 3D-LTCs.Patient-derived 3D-LTCs represent a valuable tool to analyse potential targets and drugs for lung repair. Enhanced Wnt/β-catenin signalling attenuated pathological features of patient-derived COPD 3D-LTCs.

MATERIALES
Referencia del producto
Marca
Descripción del producto

Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Methanol, HPLC Plus, ≥99.9%, poly-coated bottles
Sigma-Aldrich
HEPES, ≥99.5% (titration)
Sigma-Aldrich
Methanol, ACS reagent, ≥99.8%
Sigma-Aldrich
HEPES, BioPerformance Certified, ≥99.5% (titration), suitable for cell culture
Sigma-Aldrich
Sodium phosphate monobasic, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Sodium phosphate dibasic, ACS reagent, ≥99.0%
Sigma-Aldrich
Magnesium chloride hexahydrate, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium phosphate dibasic, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Magnesium chloride hexahydrate, ACS reagent, 99.0-102.0%
Sigma-Aldrich
Methanol, puriss. p.a., ACS reagent, reag. ISO, reag. Ph. Eur., ≥99.8% (GC)
Sigma-Aldrich
HEPES buffer solution, 1 M in H2O
Sigma-Aldrich
Agarose, low gelling temperature, BioReagent, for molecular biology
Sigma-Aldrich
Sodium phosphate dibasic, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium phosphate dibasic, BioReagent, suitable for cell culture, suitable for insect cell culture, ≥99.0%
Sigma-Aldrich
Sodium phosphate monobasic, BioReagent, for molecular biology, anhydrous, ≥98%
SAFC
HEPES
Sigma-Aldrich
Sodium phosphate dibasic, puriss. p.a., ACS reagent, anhydrous, ≥99.0% (T)
Sigma-Aldrich
Sodium phosphate dibasic, for molecular biology, ≥98.5% (titration)
Sigma-Aldrich
Magnesium chloride hexahydrate, BioReagent, suitable for cell culture, suitable for insect cell culture
Sigma-Aldrich
Sodium phosphate monobasic, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium phosphate dibasic solution, BioUltra, 0.5 M in H2O
Sigma-Aldrich
Sodium phosphate dibasic, puriss., meets analytical specification of Ph. Eur., BP, USP, FCC, E 339, anhydrous, 98-100.5% (calc. to the dried substance)
Sigma-Aldrich
Methanol, Laboratory Reagent, ≥99.6%
Sigma-Aldrich
HEPES, BioUltra, for molecular biology, ≥99.5% (T)
Sigma-Aldrich
Sodium phosphate monobasic, BioPerformance Certified, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99.0% (titration)
Sigma-Aldrich
Calcein-AM, BioReagent, suitable for fluorescence, ≥95.0% (HPLC)
Sigma-Aldrich
Magnesium chloride hexahydrate, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Methanol, BioReagent, ≥99.93%
Sigma-Aldrich
Methanol, ACS spectrophotometric grade, ≥99.9%