HSAEC1-KT (ATCC® CRL-4050)

Organism: Homo sapiens, human  /  Cell Type: epithelial  /  Tissue: lung, small airway  /  Disease: normal

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Organism Homo sapiens, human
Tissue lung, small airway
Cell Type epithelial
Product Format frozen
Morphology epithelial, packed cuboidal
Culture Properties adherent
Biosafety Level 2  [Cells immortalized by CDK4 and hTERT and contain SV40 promoter sequences]
Disease normal
Age 22
Gender male
Applications

The immortalized HSAEC1-KT cells show a stable epithelial morphology and differentiated cytokeratin isoforms after over 100 population doublings, express the stem cell marker p63 and high levels of p16INK4a, and have an intact p53 checkpoint pathway (RefRamirez RD, Sheridan S, Girard L, et al. Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res. 64(24):9027-9034, 2004. PubMed: 15604268).  

The HSAEC1-KT cells also exhibit characteristics of normal cells such as contact inhibition of growth and failure to form soft agar colonies or form tumors in immune compromised mice (RefKalita M, et al. Systems approaches to modeling chronic mucosal inflammation. Biomed. Res. Int. 2013: 505864, 2013. PubMed: 24228254).

TGFβ-treated HSAEC1-KT cells showed an elongated shape with markedly induced F-actin staining. This morphological change of enhanced front-rear polarity and cytoskeletal actin rearrangement are characteristic morphological changes of EMT (epithelial mesenchymal transition) (RefKalita M, et al. Systems approaches to modeling chronic mucosal inflammation. Biomed. Res. Int. 2013: 505864, 2013. PubMed: 24228254).

Storage Conditions liquid nitrogen vapor phase
Karyotype

Cytogenetic analysis was performed on G-banded metaphase cells from the human cell line HSAEC1-KT. Several abnormal male near-diploid karyotypes are found: 

Clone 1 demonstrates trisomy 5 with no other aberrations. 47,XY,+5 

Clone 2 demonstrates trisomy 5 and 20 with no other aberrations. 48,XY,+5,+20 

Clone 3 demonstrates trisomy 5, an isochromosome of the long-arm of chromosome 10, resulting in three copies of the chromosome 10 long-arm and only one copy of the short-arm, and trisomy 20. 48,XY,+5,i(10)(p10),+20

Clone 4 demonstrates a deletion on the short-arm of chromosome 10 at band p10, a deletion on the short-arm of chromosome 17 at band p13, and trisomy 20. 47,XY,del(10)(p10),del(17)(p13),+20.

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Derivation

The HSAEC1-KT cell line was established by infecting primary human small airway epithelial cell culture with human telomerase (hTERT) and mouse cyclin dependent kinase 4 (CDK4) expressing retrovirus constructs and selecting under 250 ng/mL puromycin and 30 ug/mL G418 as described in PMID: 15604268 (RefRamirez RD, Sheridan S, Girard L, et al. Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res. 64(24):9027-9034, 2004. PubMed: 15604268)

Clinical Data male
22 years
Antigen Expression Positive for p63 (TP63) and Clara cell 10 (CC10) protein
Complete Growth Medium SAGM BulletKit medium (Lonza CC-3119 and CC-4124)

To make the complete culture medium, add SAGM™ SingleQuots™ (Lonza CC-4124) which contains supplements and growth factors (BPE, Hydrocortisone, hEGF, Epinephrine, Transferrin, Insulin, Retinoic Acid, Triiodothyronine, BSA-FAF) to 500 mL bottle of SABM Basal Medium™, phenol red free basal medium (Lonza CC-3119)

Subculturing Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of dissociation solutions for culture vessels of other sizes.

Subculture when the culture is about 90% confluent.
  1. Remove and discard spent medium.
  2. Briefly rinse the cells with Dulbecco's Phosphate Buffered Saline (DPBS, ATCC 30-2200), 1 mL / 25 cm2 and discard rinse solution.
  3. Add Trypsin-EDTA, at 1 mL / 25 cm2, for Primary Cells (ATCC PCS-999-003) to the flask. Incubate at 37°C for 4-6 min (until 90% of the cells have detached).
  4. Rapt flask gently to ensure cells are detached.  Add 2% FBS in DPBS at 1 mL / 25 cm2 to neutralize the trypsin.
  5. Centrifuge cells at 1000rpm for 5 min at room temperature.
  6. Remove supernatant. Resuspend pellet in 6.0 to 8.0 mL Complete Growth Medium.
  7. Count cells, and seed 8.0 x 103 to 10.0 x 103 viable cells/cm2 to new culture vessels.
Medium Renewal: Every 2-3 days.
Cryopreservation 80% complete growth media, 10% DMSO, 10% FBS
Culture Conditions Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Temperature: 37°C
STR Profile D5S818:        12    
D13S317:      11, 12
D7S820:        8, 10
D16S539:     10, 11
vWA:             15, 18
Amelogenin:     X, Y
TPOX:            7, 8
CSF1PO:       11, 13
TH01:             7
Population Doubling Level (PDL) As part of our quality control, we have tested this cell line for its ability to grow for a minimum of 15 population doublings after recovery from cryopreservation. We have also compared its karyotype, telomerase expression level, growth rate, morphology and tissue-specific markers when first recovered from cryopreservation with that of cells at 10+ population doublings to ensure that there is no change in these parameters and that the cells are capable of extended proliferation.
Name of Depositor Boning Gao, Chunxian Huang, John Minna
References

Ramirez RD, Sheridan S, Girard L, et al. Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res. 64(24):9027-9034, 2004. PubMed: 15604268

Kalita M, et al. Systems approaches to modeling chronic mucosal inflammation. Biomed. Res. Int. 2013: 505864, 2013. PubMed: 24228254

Gazdar AF, Gao B, Minna JD. Lung cancer cell lines: Useless artifacts or invaluable tools for medical science? Lung Cancer 68(3): 309-318, 2010. PubMed: 20079948

Basic Documentation
Other Documentation
Restrictions

This material is subject to claims under U.S. Patent Nos. 6,261,836 and 6,337,200, other pending patent applications, and foreign counterparts thereof. It is required that either the Addendum for Commercial and For-Profit Organizations or the Addendum for Noncommercial and Academic Organizations, as appropriate, be signed and returned to ATCC before shipment.

References

Ramirez RD, Sheridan S, Girard L, et al. Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res. 64(24):9027-9034, 2004. PubMed: 15604268

Kalita M, et al. Systems approaches to modeling chronic mucosal inflammation. Biomed. Res. Int. 2013: 505864, 2013. PubMed: 24228254

Gazdar AF, Gao B, Minna JD. Lung cancer cell lines: Useless artifacts or invaluable tools for medical science? Lung Cancer 68(3): 309-318, 2010. PubMed: 20079948