Cell Model Resources

Cell models have been critical to enhancing our understanding of the physiological, biochemical, and genetic mechanisms underlying cystic fibrosis and to developing therapeutic strategies. The Cystic Fibrosis Foundation provides funding and support to make several cell model systems available to researchers.

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Summary
  • Although primary human bronchial epithelial (HBE) cells are the “gold-standard” for drug discovery in cystic fibrosis research, scientists are limited in the genotypes available because of how they are collected. For this reason, a number of other cell models have been developed to fill the gap.
  • Several cell models are available upon request.
  • These cell models include: Fischer Rat Thyroid cells, immortalized HBE cells, gene-edited HBE cells, CF and non-CF respiratory cell lines, human nasal epithelial cells, and induced pluripotent stem cells.

Although primary human bronchial epithelial (HBE) cells are the “gold-standard” for drug discovery in cystic fibrosis research, scientists are limited in the genotypes available because of how they are collected. For this reason, a number of other cell models have been developed to fill the gap.

Below is a listing of some of cell models available through the Cystic Fibrosis Foundation or other sources to advance CF research.

Stable Cell Lines

Fischer Rat Thyroid Cells

Fischer Rat Thyroid (FRT) cells have been used extensively for studies of cystic fibrosis transmembrane conductance regulator (CFTR) protein for more than 20 years and have been a ”workhorse” for drug discovery in the disease.1 The cells have been used by the U.S. Food and Drug Administration (FDA) as relevant to expanding drug label and therapeutic access for individuals with very rare forms of cystic fibrosis.2,3

FRT cells expressing CFTR are established using Flp-In™ technology.4 All cells are based on the background of a clonal FRT Flp baseline (cells bearing a single FRT insertion site), FRT Flp4, to generate CFTR variant lines. FRT Flp4 cells are clonal and selected based on zeocin and β-galactosidase activities expressed from the base construct (pFRT/lacZeo).4-6 FRT Flp4 base cells contain a single copy of lacZ-Zeocin fusion gene as shown by Southern blot and copy number variance analysis (Applied Biosystems).5 FRT wild-type and mutant CFTR cells express CFTR on an isogenic background generated using FLP-mediated recombination between pairs of FRT sites.5 All of the listed FRT Flp-CFTR cells are clonal and hygromycin resistant and selected based on very similar mRNA expression levels.5 FRT cells are commonly used in the following types of studies:

  • Electrophysiological
    • Ussing chamber analysis/short-circuit current (Isc)
    • Patch clamp/single channel measurements
    • Medium-throughput conductance measurement
  • Western blot
  • Compound library screening in a high-throughput manner
  • Immunoprecipitation
  • Immunofluorescence

RELEVANT LINKS:

To request FRT cells, please contact:

Jeong S. Hong, PhD, Andras Rab, PhD, or Eric J. Sorscher, MD
Emory University
jshong5@emory.edu
arab@emory.edu
esorscher@emory.edu
404-727-3519

REFERENCES

  1. Sheppard DN et al. Expression of cystic fibrosis transmembrane conductance regulator in a model epithelium. Am J Physiol. 1994 266(Lung Cell Mol Physiol. 10):L405-13.
  2. Durmowicz T and Pacanowski M. Novel approach allows expansion of indication for cystic fibrosis drug. FDA News & Events. 2017. https://www.fda.gov/Drugs/NewsEvents/ucm559051.htm
  3. Kingwell K. FDA OKs first in vitro route to expanded approval. Nature Reviews Drug Discovery. 2017 Sep 1;16(9):591-592
  4. Flp-In System: For Generating Constitutive Expression Cell Lines. Thermo Fisher Scientific. https://www.thermofisher.com/us/en/home/references/protocols/proteins-expression-isolation-and-analysis/protein-expression-protocol/flp-in-system-for-generating-constitutive-expression-cell-lines.html
  5. Rab A et al. Progress and challenges toward Cystic Fibrosis personalized therapy: The role of in vitro model systems. Manuscript in preparation.
  6. Han ST et al. Residual function of cystic fibrosis mutants predicts response to small molecule CFTR modulators. JCI Insights 2018, 3(14). PMID: 30046002

CF Bronchial Epithelial Cells

Immortalized human CF bronchial epithelial cells (CFBE41o- generated by D. Gruenert, UCSF) were modified using a single Flp recombination target to express CFTR cDNA to generate CF8Flp cells (as reported in Gottschalk et al. 2016). CF8 cells are commonly used in the following types of studies:

  • Electrophysiological/short-circuit current (Isc)
  • Western blot

RELEVANT LINKS:

To request CF8Flp cells, please contact:

CFTR2 Cell Center
PI: Garry Cutting, MD
Johns Hopkins School of Medicine
cftr2cellcenter@jhmi.edu

REFERENCES

Gene-Edited Human Bronchial Epithelial Cells

Immortalized, CFTR wild-type expressing 16HBE14o- cells (generated by D. Gruenert, UCSF) were gene edited at the endogenous CFTR locus using CRISPR/Cas9 to create isogenic cell lines (16HBEge). During the immortalization of the parental cells, SV40 sequence was inserted into one of the two CFTR alleles in these cells, rendering these cells monoallelic with respect to functional CFTR expression. (Full SV40 sequence information and whole genome sequencing data for parental cell line available upon request.) These cells have the native CFTR gene construct and are commonly used in the following types of studies:

  • Electrophysiological/short-circuit current (Isc)
  • Studying CFTR mutations in the native gene context
  • Western blot
  • Analysis of mRNA abundance (due to the native gene context, premature termination codon (PTC) mutations are sensitive to nonsense-mediated mRNA decay)

RELEVANT LINKS:

Cells can be requested from the Cystic Fibrosis Foundation Therapeutics Lab (CFFT) by filling out a Material Transfer Request form.

For questions related to these cells:

Hillary Valley, PhD
CFFT Lab
hvalley@cff.org

REFERENCES

CF and Non-CF Respiratory Cell Lines

A large number of human respiratory cell lines, originally collected and distributed by Dr. Dieter Gruenert, have been used to facilitate research for academic and non-commercial research purposes. This collection includes the 16HBE14o- and CFBE41o- parental cell lines as well as a number of other CF lines from different respiratory tissue locations, such as the nasal passages, submucosal glands, and tracheal tissues. In addition to the respiratory cells, the collection also includes a number of other cell lines, such as the CF pancreatic ductal cells (CFPAC), and induced pluripotent stem cells.

The USCF Cell and Genome Editing Core recently began housing and distributing these cells for researchers. Cells are still being processed into the system and will be added to inventory list as that process is completed. However, if you would like to request cells that do not appear on the list, please contact cgecore@ucsf.edu to inquire about availability.

To request cells, please follow the process outlined here. If you run into any issues, please contact:

Jennifer Page
Program Director
USCF Cell and Genome Editing Core
jennifer.page@ucsf.edu

Primary Cells

Human Bronchial Epithelial Cells

HBE cells are collected from explanted lungs after transplant. These cells are considered the “gold standard” for evaluating CFTR function in a cell type representative of the CF lung. When cultured at air-liquid interface (ALI) conditions on permeable filters, the cells resemble a pseudostratified airway epithelium as seen in the airways.

Ideal for the following types of studies:

  • Short-circuit current (Isc)
  • Studying CFTR mutations in the native gene context
  • Western blot
  • Analysis of mRNA abundance (due to the native gene context, PTC mutations are sensitive to nonsense-mediated mRNA decay)

This list of mutations available in HBE cells changes regularly based on availability of cells and is not inclusive of all mutations available through the CF Foundation. If you are interested in a mutation that is not on the list, you can inquire about its availability when filling out the Material Transfer Request form by clicking on “other” and indicating the mutation of interest in the text box.

As HBE cells are a limited resource, please note that all requests will undergo a review process to determine scientific merit of the study before requests will be granted.

RELEVANT LINKS:

Human Nasal Epithelial Cells

Human nasal epithelial (HNE) cells can be collected by nasal brush or curettage and expanded for evaluation in 2D planar or 3D spheroid culture. When cultured at air-liquid interface (ALI) conditions on permeable filters, the cells form a pseudostratified respiratory epithelium.

Ideal for the following types of studies:

  • Short-circuit current (Isc)
  • Studying CFTR mutations in the native gene context
  • Western blot
  • Analysis of mRNA abundance (due to the native gene context, PTC mutations are sensitive to nonsense-mediated mRNA decay)

G551D Observational Study (GOAL) and PROSPECT Prospective Biomarkers Study HNE Cells can be requested through the CF Foundation Biorepository.

HNE cells collected as part of the RARE Study can be requested from the Cystic Fibrosis Foundation Therapeutics Lab by filling out a Material Transfer Request form. The current collection of HNE cells available through the CFFT Lab all have genotypes with two nonsense mutations. This list of mutations available in HNE cells changes regularly based on availability of cells and is not inclusive of all mutations available through the CF Foundation. If you are interested in requesting cells, you can inquire about their availability when filling out the Material Transfer Request form by clicking on “other” and indicating the mutation of interest in the text box.

To request HNE cells collected through the CF Canada-SickKids Program for Individual CF Therapy (CFIT), please contact:

Paul Eckford, PhD
Program Manager
The Hospital for Sick Children (SickKids)
peckford@sickkids.ca

Induced Pluripotent Stem Cells

Induced pluripotent stem (iPS) cells are stem cells generated from adult cells (non-embryonic) by introducing specific transcription factors that reprogram the cells to become pluripotent. iPS cells do not express CFTR in the undifferentiated state but can be differentiated into different cell types (including those that express CFTR) by using various differentiation protocols.

Ideal for the following types of studies:

  • Studying CFTR mutations in the native gene context
  • Western blot
  • Analysis of mRNA abundance (due to the native gene context, PTC mutations are sensitive to nonsense-mediated mRNA decay)

RELEVANT LINKS:

iPS cells can be requested from the CFFT Lab by filling out a Material Transfer Request form.

To request iPS cells collected through the CF Canada-SickKids Program for Individual CF Therapy (CFIT), please contact:

Paul Eckford, PhD
Program Manager
The Hospital for Sick Children (SickKids)
peckford@sickkids.ca

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