Immortalized Slc4a11^-/- Mouse Corneal Endothelial Cells

Immortalized Slc4a11^-/- Mouse Corneal Endothelial Cells were isolated from the cornea of progeny mice produced by the intercrossing of Slc4a11þ/þ and Slc4a11/C57BL/6 mice with H-2Kb-tsA58 Immortomouse, which carries a temperature-sensitive mutant form of the simian virus-40 large T antigen. Immortalized Slc4a11^-/- Mouse Corneal Endothelial Cells also demonstrates temperature sensitive and IFN-c dependent proliferation. This cell line represents a useful model for the investigation of pathology and therapies for SLC4A11 associated congenital hereditary endothelial dystrophy (CHED) and Fuchs’ endothelial corneal dystrophy.

Immortalization Method: Intercrossed the Slc4a11þ/þ and Slc4a11/C57BL/6 mice with H-2Kb-tsA58 Immortomouse, which carries a temperature-sensitive mutant form of the simian virus-40 large T antigen

BioSafety Level: II

Organism: Mouse

Species: Mouse

Source Organs: Eye

Organ Type: Eye

Growth Properties: Adherent

Morphology: Fibroblast-like

Population Doubling: 18.96 hours

Propagation: Use of FNC coating is required for cell adhesion to the culture vessels. Grow cells in FNC-coated culture vessels with the following conditions. The base medium for this cell line is Opti-MEM I (Invitrogen). To make the complete growth medium, add the following components to the base medium: heat-inactivated fetal bovine serum (TM999) to a final concentration of 8%, Antibiotic/Antimycotic (G273) to a final concentration of 1%, chondroitin sulfate (Sigma) to a final concentration of 0.08%, CaCl₂ (Sigma) to a final concentration of 200 μg/mL, pituitary extract (Biomedical Technologies. Inc.) to a final concentration of 100 μg/mL, gentamycin (Invitrogen) to a final concentration of 0.5%, EGF to a final concentration of 5 ng/ml (Millipore), and IFN-γ (R&D Systems) to a final concentration of 200 ng/mL.Change media every other day.Carbon dioxide (CO₂): 5%, Temperature: +33.0°C.Growing cells at +37.0°C will decrease tsTAg levels and stop replication.

Quality Control: 1) Western blot; 2) Intracellular pH (pHi) measurement; 3) RT-PCR; 4) Real-time quantitative PCR

Shipping Condition: Dry Ice

Storage Condition: liquid nitrogen or -180°C

Reference: Zhang, W., Ogando, D. G., Kim, E. T., Choi, M.-J., Li, H., Tenessen, J. M., & Bonanno, J. A. (2017). Conditionally Immortal Slc4a11−/− Mouse Corneal Endothelial Cell Line Recapitulates Disrupted Glutaminolysis Seen in Slc4a11−/− Mouse Model. Investigative Opthalmology & Visual Science, 58(9), 3723. https://doi.org/10.1167/iovs.17-21781