PneumaCult™-Ex Medium

Serum- and BPE-free medium for expansion of primary human airway epithelial cells
PneumaCult™-Ex Medium

Serum- and BPE-free medium for expansion of primary human airway epithelial cells

500 mL
Catalog # 05008
168 USD
Required Products
  1. Trypsin-EDTA (0.05%)
    Trypsin-EDTA (0.05%)

    Enzymatic cell dissociation reagent

  2. HBSS without Ca++ and Mg++
    HBSS, Modified (Without Ca++ and Mg++)

    Hanks' Balanced Salt Solution (HBSS) without calcium and magnesium

  3. Hydrocortisone Stock Solution|07925
    Hydrocortisone Stock Solution

    Cell culture supplement

  4. D-PBS Without Ca++ and Mg++
    D-PBS (Without Ca++ and Mg++)

    Dulbecco’s phosphate-buffered saline without calcium and magnesium

Overview

PneumaCult™-Ex is a defined, serum- and BPE-free cell culture medium that supports rapid expansion of human airway epithelial cells.

Primary airway epithelial cells cultured in PneumaCult™-Ex expand rapidly over at least 3 passages while maintaining a cobblestone morphology and uniform expression of the basal cell markers p63 and p75NTR. Additionally, cells cultured in PneumaCult™-Ex can be differentiated to form a pseudostratified mucociliary epithelium when cultured at the air-liquid interface in PneumaCult™-ALI.

Together, PneumaCult™-Ex and PneumaCult™-ALI constitute a fully integrated BPE-free culture system for in vitro human airway modeling. This robust and defined system is a valuable tool for basic respiratory research, toxicity studies, and drug development.

Learn how to culture human airway epithelial cells at the ALI in our On-Demand Pulmonary Course or browse our Frequently Asked Questions (FAQs) about the ALI culture workflow using PneumaCult™.
Advantages
• PneumaCult™-Ex is a defined (BPE-free) medium that delivers consistent performance for expansion of HBECs
• When used together, PneumaCult™-Ex and PneumaCult™-ALI constitute a complete cell culture media system for expansion of primary human airway cells and their subsequent differentiation to a pseudostratified mucociliary epithelium
Components
  • PneumaCult™-Ex Basal Medium, 490 mL
  • PneumaCult™-Ex 50X Supplement, 10 mL
Subtype
Specialized Media
Cell Type
Airway Cells
Species
Human
Application
Cell Culture, Expansion, Maintenance
Brand
PneumaCult
Area of Interest
Epithelial Cell Biology
Formulation
Serum-Free

Scientific Resources

Product Documentation

Document Type Product Name Catalog # Lot # Language
Document Type
Product Information Sheet
Product Name
PneumaCult™-Ex Medium
Catalog #
05008
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
PneumaCult™-Ex Medium
Catalog #
05008
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
PneumaCult™-Ex Medium
Catalog #
05008
Lot #
All
Language
English

Educational Materials (10)

Brochure
PneumaCult™ Culture Media for Human Airway Epithelial Cells
Technical Bulletin
Air-Liquid Interface Culture for Respiratory Research
Video
How to Model the Human Airway at the Air-Liquid Interface: Differentiation of HBECs
5:03
How to Model the Human Airway at the Air-Liquid Interface: Differentiation of HBECs
Video
Why Use Air-Liquid Interface Cultures for Respiratory Research: Q&A with Dr. Wadsworth
3:55
Why Use Air-Liquid Interface Cultures for Respiratory Research: Q&A with Dr. Wadsworth
Video
How to Model the Human Airway at the Air-Liquid Interface: Introduction
2:13
How to Model the Human Airway at the Air-Liquid Interface: Introduction
Video
How to Model the Human Airway at the Air-Liquid Interface: Expansion of HBECs
7:42
How to Model the Human Airway at the Air-Liquid Interface: Expansion of HBECs
Video
Air-Liquid Interface (ALI) Culture of Bronchial Epithelial Cells with PneumaCult™-ALI Medium
2:01
Air-Liquid Interface (ALI) Culture of Bronchial Epithelial Cells with PneumaCult™-ALI Medium
Webinar
PneumaCult™-ALI: An Improved Medium Formulation for the Differentiation of Human Bronchial Epithelial Cells
47:25
PneumaCult™-ALI: An Improved Medium Formulation for the Differentiation of Human Bronchial Epithelial Cells
Mini Review
Airway Epithelial Cells of the Human Respiratory System
Scientific Poster
PneumaCult™: an Integrated Culture Medium System for in Vitro Human Airway Modeling

Product Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Research Area
Workflow Stages

Data and Publications

Data

HBECs Cultured in PneumaCult™-Ex Exhibit Cobblestone Morphology

Figure 1. HBECs Cultured in PneumaCult™-Ex Exhibit Cobblestone Morphology

Commercially available, cryopreserved, passage 1 (P1) HBECs were seeded into PneumaCult™-Ex or a Control medium (BEGM™, Lonza). Cells exhibit cobblestone morphology in both culture media, as seen in representative images of confluent cultures 5 days post-seeding (A,B). HBECs cultured for an additional 3 passages in both PneumaCult™-Ex and Control medium continue to expand and retain their normal cobblestone morphology, as shown by representative images of confluent P4 cultures at 7 days post-seeding (C,D). All images were taken through 10X objective.

HBECs Cultured in PneumaCult™-Ex Exhibit Uniform Expression of Basal Cell Markers

Figure 2. HBECs Cultured in PneumaCult™-Ex Exhibit Uniform Expression of Basal Cell Markers

Passage 3 HBECs cultured in PneumaCult™-Ex demonstrate extensive co-labeling of the basal cell markers p63 (red) and p75NTR (green, A). A representative merged image indicates widespread co-labeling of p63, p75NTR and the nuclear stain DAPI (blue, B).

HBECs Cultured in PneumaCult™-Ex Exhibit Comparable Expansion Rates to Cells Cultured in Control Medium

Figure 3. HBECs Cultured in PneumaCult™-Ex Exhibit Comparable Expansion Rates to Cells Cultured in Control Medium

Commercially available, cryopreserved, P1 HBECs were seeded into PneumaCult™-Ex or a Control medium (BEGM™, Lonza). In seven independent donor samples, the average fold expansion over four passages was not significantly different between cells cultured in PneumaCult™-Ex and cells cultured in the Control medium (7.1 ± 1.4 vs. 7.2 ± 1.9, mean ± SD, n = 7, p = 0.9 in paired t-test).

Publications (14)

International journal of molecular sciences 2020 sep Impact of KLF4 on Cell Proliferation and Epithelial Differentiation in the Context of Cystic Fibrosis. L. Sousa et al.
Abstract

Abstract

Cystic fibrosis (CF) cells display a more cancer-like phenotype vs. non-CF cells. KLF4 overexpression has been described in CF and this transcriptional factor acts as a negative regulator of wt-CFTR. KLF4 is described as exerting its effects in a cell-context-dependent fashion, but it is generally considered a major regulator of proliferation, differentiation, and wound healing, all the processes that are also altered in CF. Therefore, it is relevant to characterize the differential role of KLF4 in these processes in CF vs. non-CF cells. To this end, we used wt- and F508del-CFTR CFBE cells and their respective KLF4 knockout (KO) counterparts to evaluate processes like cell proliferation, polarization, and wound healing, as well as to compare the expression of several epithelial differentiation markers. Our data indicate no major impact of KLF4 KO in proliferation and a differential impact of KLF4 KO in transepithelial electrical resistance (TEER) acquisition and wound healing in wt- vs. F508del-CFTR cells. In parallel, we also observed a differential impact on the levels of some differentiation markers and epithelial-mesencymal transition (EMT)-associated transcription factors. In conclusion, KLF4 impacts TEER acquisition, wound healing, and the expression of differentiation markers in a way that is partially dependent on the CFTR-status of the cell.
Cells 2020 oct Aprotinin Inhibits SARS-CoV-2 Replication. D. Bojkova et al.

Abstract

Severe acute respiratory syndrome virus 2 (SARS-CoV-2) is the cause of the current coronavirus disease 19 (COVID-19) pandemic. Protease inhibitors are under consideration as virus entry inhibitors that prevent the cleavage of the coronavirus spike (S) protein by cellular proteases. Herein, we showed that the protease inhibitor aprotinin (but not the protease inhibitor SERPINA1/alpha-1 antitrypsin) inhibited SARS-CoV-2 replication in therapeutically achievable concentrations. An analysis of proteomics and translatome data indicated that SARS-CoV-2 replication is associated with a downregulation of host cell protease inhibitors. Hence, aprotinin may compensate for downregulated host cell proteases during later virus replication cycles. Aprotinin displayed anti-SARS-CoV-2 activity in different cell types (Caco2, Calu-3, and primary bronchial epithelial cell air-liquid interface cultures) and against four virus isolates. In conclusion, therapeutic aprotinin concentrations exert anti-SARS-CoV-2 activity. An approved aprotinin aerosol may have potential for the early local control of SARS-CoV-2 replication and the prevention of COVID-19 progression to a severe, systemic disease.
The Journal of biological chemistry 2020 nov Genomic determinants implicated in the glucocorticoid-mediated induction of KLF9 in pulmonary epithelial cells. M. M. Mostafa et al.
Abstract

Abstract

Ligand-activated glucocorticoid receptor (GR) elicits variable glucocorticoid-modulated transcriptomes in different cell types. However, some genes, including Kr{\{u}}ppel-like factor 9 (KLF9) a putative transcriptional repressor demonstrate conserved responses. We show that glucocorticoids induce KLF9 expression in the human airways in vivo and in differentiated human bronchial epithelial (HBE) cells grown at air-liquid interface (ALI). In A549 and BEAS-2B pulmonary epithelial cells glucocorticoids induce KLF9 expression with similar kinetics to primary HBE cells in submersion culture. A549 and BEAS-2B ChIP-seq data reveal four common glucocorticoid-induced GR binding sites (GBSs). Two GBSs mapped to the 5'-proximal region relative to KLF9 transcription start site (TSS) and two occurred at distal sites. These were all confirmed in primary HBE cells. Global run-on (GRO)-sequencing indicated robust enhancer RNA (eRNA) production from three of these GBSs in BEAS-2B cells. This was confirmed in A549 cells plus submersion and ALI culture of HBE cells. Cloning each GBS into luciferase reporters revealed glucocorticoid-induced activity requiring a glucocorticoid response element (GRE) within each distal GBS. While the proximal GBSs drove modest reporter induction by glucocorticoids this region exhibited basal eRNA production RNA polymerase II enrichment and looping to the TSS plausibly underlying constitutive KLF9 expression. Post-glucocorticoid treatment interactions between distal and proximal GBSs and the TSS correlated with KLF9 induction. CBP/P300 silencing reduced proximal GBS activity but negligibly effected KLF9 expression. Overall a model for glucocorticoid-mediated regulation of KLF9 involving multiple GBSs is depicted. This work unequivocally demonstrates that mechanistic insights gained from cell-lines can translate to physiologically relevant systems."
The Journal of biological chemistry 2020 nov Loss of versican and production of hyaluronan in lung epithelial cells are associated with airway inflammation during RSV infection. G. G. Kellar et al.
Abstract

Abstract

Airway inflammation is a critical feature of lower respiratory tract infections caused by viruses such as respiratory syncytial virus (RSV). A growing body of literature has demonstrated the importance of extracellular matrix (ECM) changes such as the accumulation of hyaluronan (HA) and versican in the subepithelial space in promoting airway inflammation; however, whether these factors contribute to airway inflammation during RSV infection remains unknown. To test the hypothesis that RSV infection promotes inflammation via altered HA and versican production, we studied an ex vivo human bronchial epithelial cell (BEC)/human lung fibroblast (HLF) co-culture model. RSV infection of BEC/HLF co-cultures led to decreased hyaluronidase expression by HLFs, increased accumulation of HA, and enhanced adhesion of U937 cells as would be expected with increased HA. HLF production of versican was not altered following RSV infection; however, BEC production of versican was significantly downregulated following RSV infection. In vivo studies with epithelial-specific versican-deficient mice [SPC-Cre(+) Vcan-/-] demonstrated that RSV infection led to increased HA accumulation compared to control mice which also coincided with decreased hyaluronidase expression in the lung. SPC-Cre(+) Vcan-/- mice demonstrated enhanced recruitment of monocytes and neutrophils in bronchoalveolar lavage fluid and increased neutrophils in the lung compared to SPC-Cre(-) RSV-infected littermates. Taken together, these data demonstrate that altered ECM accumulation of HA occurs following RSV infection and may contribute to airway inflammation. Additionally, loss of epithelial expression of versican promotes airway inflammation during RSV infection further demonstrating that versican's role in inflammatory regulation is complex and dependent on the microenvironment.
Nature communications 2020 dec MCL-1 gains occur with high frequency in lung adenocarcinoma and can be targeted therapeutically. E. Munkhbaatar et al.
Abstract

Abstract

Evasion of programmed cell death represents a critical form of oncogene addiction in cancer cells. Understanding the molecular mechanisms underpinning cancer cell survival despite the oncogenic stress could provide a molecular basis for potential therapeutic interventions. Here we explore the role of pro-survival genes in cancer cell integrity during clonal evolution in non-small cell lung cancer (NSCLC). We identify gains of MCL-1 at high frequency in multiple independent NSCLC cohorts, occurring both clonally and subclonally. Clonal loss of functional TP53 is significantly associated with subclonal gains of MCL-1. In mice, tumour progression is delayed upon pharmacologic or genetic inhibition of MCL-1. These findings reveal that MCL-1 gains occur with high frequency in lung adenocarcinoma and can be targeted therapeutically.
International journal of cancer 2018 JUL Conditionally reprogrammed cells (CRC) methodology does not allow the in vitro expansion of patient-derived primary and metastatic lung cancer cells. G. Sette et al.

Abstract

Availability of tumor and non-tumor patient-derived models would promote the development of more effective therapeutics for non-small cell lung cancer (NSCLC). Recently, conditionally reprogrammed cells (CRC) methodology demonstrated exceptional potential for the expansion of epithelial cells from patient tissues. However, the possibility to expand patient-derived lung cancer cells using CRC protocols is controversial. Here, we used CRC approach to expand cells from non-tumoral and tumor biopsies of patients with primary or metastatic NSCLC as well as pulmonary metastases of colorectal or breast cancers. CRC cultures were obtained from both tumor and non-malignant tissues with extraordinary high efficiency. Tumor cells were tracked in vitro through tumorigenicity assay, monitoring of tumor-specific genetic alterations and marker expression. Cultures were composed of EpCAM+ lung epithelial cells lacking tumorigenic potential. NSCLC biopsies-derived cultures rapidly lost patient-specific genetic mutations or tumor antigens. Similarly, pulmonary metastases of colon or breast cancer generated CRC cultures of lung epithelial cells. All CRC cultures examined displayed epithelial lung stem cell phenotype and function. In contrast, brain metastatic lung cancer biopsies failed to generate CRC cultures. In conclusion, patient-derived primary and metastatic lung cancer cells were negatively selected under CRC conditions, limiting the expansion to non-malignant lung epithelial stem cells from either tumor or non-tumor tissue sources. Thus, CRC approach cannot be applied for direct therapeutic testing of patient lung tumor cells, as the tumor-derived CRC cultures are composed of (non-tumoral) airway basal cells.
View All Publications

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