Lung endothelial cells strengthen, but brain endothelial cells weaken barrier properties of a human alveolar epithelium cell culture model

Neuhaus, Winfried; Samwer, Fabian; Kunzmann, Steffen; Muellenbach, Ralf Michael; Wirth, Michael; Speer, Christian P.; Roewer, Norbert; Förster, Carola

doi:10.1016/j.diff.2012.08.006

Cited by 26 publications

(34 citation statements)

References 43 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These results are consistent with previous studies that have found that A549 cells are incapable of forming functional tight junctions [18, 40, 50, 55]. The contrasting barrier functions between the two cell lines may be due to the differential expression of junctional proteins.…”

Section: Discussionsupporting

confidence: 93%

“…Reported culture conditions for air-liquid culture of NCI-H441 cells vary widely in seeding density (20,000–5,000,000 cells/cm 2 ) [23, 40–43], dexamethasone concentration (10–2000 nM) [22, 27, 40–42], and the use of supplements such as ITS and T3 [27, 28]. We examined the effect of these variables on the phenotypic and functional characteristics of the model.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, none of these studies have examined sodium and chloride transport in conjunction and there is no information about the paracellular and transcellular components of net transport. The properties of these cell line models have also not been compared to recent data from primary human cell cultures [11] and there is little information on how transport protein expression and function is affected by culture conditions that vary widely in cell seeding density [23, 40–43] and the nature and concentration of supplements [22, 27, 28, 40–42]. …”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

An Optimised Human Cell Culture Model for Alveolar Epithelial Transport

2016

View full text Add to dashboard Cite

Robust and reproducible in vitro models are required for investigating the pathways involved in fluid homeostasis in the human alveolar epithelium. We performed functional and phenotypic characterisation of ion transport in the human pulmonary epithelial cell lines NCI-H441 and A549 to determine their similarity to primary human alveolar type II cells. NCI-H441 cells exhibited high expression of junctional proteins ZO-1, and E-cadherin, seal-forming claudin-3, -4, -5 and Na+-K+-ATPase while A549 cells exhibited high expression of pore-forming claudin-2. Consistent with this phenotype NCI-H441, but not A549, cells formed a functional barrier with active ion transport characterised by higher electrical resistance (529 ± 178 Ω cm2 vs 28 ± 4 Ω cm2), lower paracellular permeability ((176 ± 42) ×10−8 cm/s vs (738 ± 190) ×10−8 cm/s) and higher transepithelial potential difference (11.9 ± 4 mV vs 0 mV). Phenotypic and functional properties of NCI-H441 cells were tuned by varying cell seeding density and supplement concentrations. The cells formed a polarised monolayer typical of in vivo epithelium at seeding densities of 100,000 cells per 12-well insert while higher densities resulted in multiple cell layers. Dexamethasone and insulin-transferrin-selenium supplements were required for the development of high levels of electrical resistance, potential difference and expression of claudin-3 and Na+-K+-ATPase. Treatment of NCI-H441 cells with inhibitors and agonists of sodium and chloride channels indicated sodium absorption through ENaC under baseline and forskolin-stimulated conditions. Chloride transport was not sensitive to inhibitors of the cystic fibrosis transmembrane conductance regulator (CFTR) under either condition. Channels inhibited by 5-nitro-1-(3-phenylpropylamino) benzoic acid (NPPB) contributed to chloride secretion following forskolin stimulation, but not at baseline. These data precisely define experimental conditions for the application of NCI-H441 cells as a model for investigating ion and water transport in the human alveolar epithelium and also identify the pathways of sodium and chloride transport.

show abstract

Section: Discussionsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

An Optimised Human Cell Culture Model for Alveolar Epithelial Transport

2016

View full text Add to dashboard Cite

show abstract

“…102 The alveolar epithelium is made up of two epithelial cell types, namely, the terminally differentiated squamous alveolar epithelial type-1 (AT-I) and the surfactant-producing cuboidal alveolar epithelial type-II (AT-II) cells. 102 …”

Section: Pulmonary Modelsmentioning

confidence: 99%

TEER Measurement Techniques for In Vitro Barrier Model Systems

et al. 2015

View full text Add to dashboard Cite

Transepithelial/transendothelial electrical resistance (TEER) is a widely accepted quantitative technique to measure the integrity of tight junction dynamics in cell culture models of endothelial and epithelial monolayers. TEER values are strong indicators of the integrity of the cellular barriers before they are evaluated for transport of drugs or chemicals. TEER measurements can be performed in real-time without cell damage and generally are based on measuring ohmic resistance or measuring impedance across a wide spectrum of frequencies. TEER measurements for various cell types have been reported with commercially available measurement systems and also with custom built microfluidic implementations. Some of the barrier models that have been widely characterized utilizing TEER include the blood-brain barrier (BBB), gastrointestinal (GI) tract, and pulmonary models. Variations in TEER value can arise due to factors such as temperature, medium formulation and passage number of cells. The aim of this paper is to review the different TEER measurement techniques and analyze their strengths and weaknesses, the significance of TEER in drug toxicity studies, examine the various in vitro models and microfluidic organs-on-chips implementations utilizing TEER measurements in some widely studied barrier models (BBB, GI tract and pulmonary), and discuss the various factors that can affect TEER measurements.

show abstract

“…Consequently, microvascular endothelial cells form tighter cell–cell junctions [105], [106]. Human lung microvascular endothelial cells may also enhance the barrier function of epithelial cells [107], which actually provide a barrier with a smaller transport threshold than that of the endothelium. Compared to the 5-nm cutoff for endothelial cell–cell barriers, the epithelium provides a barrier to molecules greater than 1 nm.…”

Section: Strategies For Cellular Repopulation: Cell Types and Usmentioning

confidence: 99%

Strategies for Whole Lung Tissue Engineering

Calle

Ghaedi

Sundaram

et al. 2014

IEEE Trans. Biomed. Eng.

View full text Add to dashboard Cite

Recent work has demonstrated the feasibility of using decellularized lung extracellular matrix scaffolds to support the engineering of functional lung tissue in vitro. Rendered acellular through the use of detergents and other reagents, the scaffolds are mounted in organ-specific bioreactors where cells in the scaffold are provided with nutrients and appropriate mechanical stimuli such as ventilation and perfusion. Though initial studies are encouraging, a great deal remains to be done to advance the field and transition from rodent lungs to whole human tissue engineered lungs. To do so, a variety of hurdles must be overcome. In particular, a reliable source of human-sized scaffolds, as well as a method of terminal sterilization of scaffolds, must be identified. Continued research in lung cell and developmental biology will hopefully help identify the number and types of cells that will be required to regenerate functional lung tissue. Finally, bioreactor designs must be improved in order to provide more precise ventilation stimuli and vascular perfusion in order to avoid injury to or death of the cells cultivated within the scaffold. Ultimately, the success of efforts to engineer a functional lung in vitro will critically depend on the ability to create a fully endothelialized vascular network that provides sufficient barrier function and alveolar-capillary surface area to exchange gas at rates compatible with healthy lung function.

show abstract

Lung endothelial cells strengthen, but brain endothelial cells weaken barrier properties of a human alveolar epithelium cell culture model

Cited by 26 publications

References 43 publications

An Optimised Human Cell Culture Model for Alveolar Epithelial Transport

An Optimised Human Cell Culture Model for Alveolar Epithelial Transport

TEER Measurement Techniques for In Vitro Barrier Model Systems

Strategies for Whole Lung Tissue Engineering

Contact Info

Product

Resources

About