Fig. 2

Phenotype and irreversible de-differentiation of Immortomouse^® CPECs. a Primary Immortomouse^® choroid plexus epithelial cells (p0) grown at 37 °C for 7 days showed the same staining patterns for cytokeratin (CK) and claudin-1 (Cldn-1) as pmCPECs isolated from wild-type mice. b When Immortomouse^® CPECs were grown under permissive conditions (33 °C, 10U IFNγ) the heterogenous de-differentiation process started in areas with low cell density, whereas CPECs kept their cuboidal shape longer in areas with high cell density. The contrast of the pictures in a and b was enhanced using Adobe Photoshop software. c Vimentin staining, rather than CK staining was observed at 33 °C. Cell death took place at 33 °C upon Ara-C addition to culture. d The irreversible loss of CPEC specific markers Cldn-1 and CK and an increasing proliferation rate of de-differentiated Immortomouse^® CPECs was observed with increased passage. The first row is passage 2 (p2), the second row is p3, the third row is p4. e The cells failed to form confluent monolayers or display the correct expression pattern of epithelial markers upon shift to non-permissive temperature and IFNγ withdrawal. Immortomouse^® CPECs in e were stained after 7 days of non-permissive growth; the passage numbers (p) were E-cad/DAPI: p8, ZO1/β-Cat/DAPI: p5, Cldn11/N-Cad/DAPI: p6, Cldn-1/CK/DAPI: p4. Scale bar in all immunofluorescent and phase contrast pictures = 100 μm. p0 primary culture, E-Cad E-cadherin, N-Cad N-cadherin, Cldn-1 Claudin 1, Cldn-11 Claudin 11, CK cytokeratin, β-Cat β-Catenin, ZO1 zona occludens protein 1