H19-7/IGF-IR (ATCC® CRL-2526)

Organism: Rattus norvegicus, rat  /  Cell Type: fibroblast  /  Tissue: brain/hippocampus  / 

Organism Rattus norvegicus, rat
Tissue brain/hippocampus
Cell Type fibroblast
Product Format frozen
Morphology fibroblast
Culture Properties adherent
Biosafety Level 2 [Cells contain SV40 DNA Viral Sequences]
Age embryo; 17 day gestation
Strain Holtzman
Applications This cell line can be used for analysis and comparison of mitogenesis (34°C) and differentiation (39°C).
Storage Conditions liquid nitrogen vapor phase
Derivation
The H19-7 cell line was derived from hippocampi dissected from embryonic day 17 (E17) Holtzman rat embryos and immortalized by retroviral transduction of temperature sensitive tsA58 SV40 large T antigen. H19-7/IGF-IR cells were established by infecting H19-7 cells with a retroviral vector expressing the human type I insulin-like growth factor receptor (IGF-IR). The cells were selected in medium containing puromycin.
Receptor Expression
human type I insulin-like growth factor receptor (IGF-IR)
Comments
H19-7 cells grow at the permissive temperature (34°C) in epidermal growth factor or serum.

They differentiate to a neuronal phenotype at the nonpermissive temperature (39°C) when induced by basic fibroblast growth factor (bFGF) in N2 medium (DMEM-high glucose medium with supplements).

At 39°C, expression of the human IGF-IR in H19-7 cells induces an insulin-like growth factor (IGF) I dependent differentiation. The cells extend neurites and show increased expression of NF68.

H19-7/IGF-IR cells express the IGF-IR protein. IGF-IR is known to send two seemingly contradictory signals inducing either cell proliferation or cell differentiation, depending on cell type and/or conditions.

This cell line does not express detectable levels of the SV40 T antigen.

Complete Growth Medium The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: 0.2 mg/ml G418
  • 0.001 mg/ml puromycin
  • fetal bovine serum to a final concentration of 10%

  • Subculturing Volumes used in this protocol are for 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
    Note: Flasks used to propagate these cells must be coated with 0.015 mg/mL poly-l-lysine. Cover the surface of the vessel with poly-l-lysine solution for 5 minutes, remove, and let dry.

    1. Remove and discard culture medium.
    2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
    3. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
      Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
    4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
    5. Add appropriate aliquots of the cell suspension to new culture vessels.
    6. Incubate cultures at 34°C.

    Subculture Ratio: 1:3 to 1:5
    Medium Renewal: Every 2 to 3 days.
    Note: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 10 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 3rd edition, published by Alan R. Liss, N.Y., 1994.

    Cryopreservation
    Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO
    Storage temperature: liquid nitrogen vapor phase
    Culture Conditions
    Atmosphere: air, 95%; carbon dioxide (CO2), 5%
    Temperature: 34°C
    Growth Conditions: Flasks used to propagate these cells must be coated with 0.015 mg/mL poly-l-lysine. Cover the surface of the vessel with poly-l-lysine solution for 5 minutes, remove and let dry.
    Name of Depositor R Baserga, A Morrione
    References

    Morrione A, et al. Insulin-like growth factor I receptor signaling in differentiation of neuronal H19-7 cells. Cancer Res. 60: 2263-2272, 2000. PubMed: 10786694

    Eves EM, et al. Immortal rat hippocampal cell lines exhibit neuronal and glial lineages and neurotrophin gene expression. Proc. Natl. Acad. Sci. USA 89: 4373-4377, 1992. PubMed: 1316607

    Basic Documentation
    References

    Morrione A, et al. Insulin-like growth factor I receptor signaling in differentiation of neuronal H19-7 cells. Cancer Res. 60: 2263-2272, 2000. PubMed: 10786694

    Eves EM, et al. Immortal rat hippocampal cell lines exhibit neuronal and glial lineages and neurotrophin gene expression. Proc. Natl. Acad. Sci. USA 89: 4373-4377, 1992. PubMed: 1316607