TNF Induces Choroid Plexus Epithelial Cell Barrier Alterations by Apoptotic and Nonapoptotic Mechanisms
Figure 8
TNF induces HMGB1 release as well as HBGB1 binding in PCPEC. TNF-stimulated PCPEC loose membrane integrity. PCPEC were stimulated with TNF (100 ng/mL) cycloheximide (CHX) (1 g/mL) for 10 hours. For a positive control PCPECs were treated with staurosporine (1 M). (a) Adherent cells were labelled according to the Live/Dead assay protocol. Vital cells are penetrated by calcein and show a green fluorescence, damaged cells were penetrated by ethidium homodimer and show red fluorescent nuclei. Images were obtained by fluorescence microscopy (400 magnification) using a digital camera. (b) Chromatin of vital and apoptotic cells binds HMGB1 (red) whereas TNF CHX and staurosporine stimulated PCPEC partly released HMGB1 from nuclei (i.e., post-apoptotic necrosis; white arrows). DAPI staining revealed also that TNF cycloheximide (CHX) resulted in chromatin condensation in PCPEC and binding of HMGB1 (i.e., apoptosis; yellow arrows). Both were detected by fluorescence microscopy (1000 magnification) and photographs were taken using a digital camera. The whole well was examined for each experimental condition in two separate experiments; results from one representative experiment are shown.
