Immortalized Mouse Cerebellar Capillary Endothelial Cells (cerebEND)

The mammalian cerebellum is indispensable in the coordination of voluntary movement (i.e. balance and locomotion), muscular tonus and equilibrium. The cerebellar blood brain barrier (BBB) helps maintain the homeostasis of the cerebellum microenvironment and its integrity is often connected to many disorders of the central nervous system (CNS). The Immortalized Mouse Cerebellar Capillary Endothelial Cell Line (cerebEND) provides a useful model for studies involving the differentiation and regulation of BBB as it shares principal features of the BBB in vivo, specifically in that 1) it retains endothelial markers VE-Cadherin and PECAM-1; 2) it displays tight junction associated markers such as occludin, claudin-3, -5, -12, 3) it expresses the BBB marker protein Glut-1 and 4) it shows high electrical resistance, representing barrier properties. This cell line is also responsive to glucocorticoid, estrogen-treatment and pro-inflammatory mediator such as TNFα, making this cell line valuable in elucidating cellular responses of endothelial cells to different stimuli. Compared to the Immortalized Mouse Cerebral Capillary Endothelial Cell Line (cEND), the cerebEND shows a higher fragility of the cerebellar BBB to inflammatory stimuli; together the two cell lines represent important model systems for these different brain regions.

Immortalization Method: Transformation with oncoprotein of murine Polyomavirus, Polyoma middle T antigen (PymT)

BioSafety Level: II

Species: Mouse

Source Organ: Cerebellar cortex

Growth Properties: Adherent

Morphology: Spindle shaped

Population Doubling: N/A

Seeding Density: Recommended split ratio: no greater than 1:4

Markers: Claudin-5, Occuldin, Glut-1, VE-Cadherin

Donor Age: N/A

Donor Gender: N/A

Donor Ethnicity: N/A

Propagation: The base medium for this cell line is Prigrow III medium available at abm (TM003). To make the completed growth medium, add the following components to the base medium: heat-inactivated fetal bovine serum to final concentration of 10% and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%
Carbon dioxide (CO₂): 5%, Temperature: +37.0°C. The Differentiation Medium is composed of 2% heat-inactivated fetal bovine serum instead of 10%. Culturing cerebEND cells in serum-reduced medium will lead to an increase in TER as well as change in cell morphology (from spindle-shaped to cobble-stone like). TER can be further increased by the addition of 110nM hydrocortisone and/or 1µM insulin into the differentiation medium.

Quality Control: 1) Endothelial marker VE-Cadherin and PECAM-1 assessed by immunostainning; 2) Tight junction protein Claudin-5 assessed by immunostainning and claudin-1, -3 and -12 detected in mRNA levels; 2) BBB marker protein Glut-1 and tight junction-associated protein occludin measured by western blot analysis. The cell line's response to pro-inflammatory stimuli was measured by 1) TER, 2) western blot and 3) gene expression analysis using RT-PCR.

Shipping Condition: Dry Ice

Storage Condition: liquid nitrogen or -180°C