Transcriptional responses to deep cerebellar nuclei (DCN) and entorhinal cortex (EC) protein homogenates in cultured microglia

We hypothesised that enriched cytokines in the microenvironment of the deep cerebellar nuclei (DCN) contribute to the inflammatory gene expression profiles of DCN microglia and thus might induce a similar response in cultured microglia. To test this hypothesis, we treated BV2 microglia cell lines with DCN or entorhinal cortex (EC) protein homogenates and analysed RNA expression 4h post-stimulation, the peak timepoint of LPS-induced inflammatory gene expression (Das et al, 2016). To enrich for extracellular secreted proteins and to prevent effects of extraction buffer components on cell cultures, proteins were gently homogenised in DPBS on ice in the absence of detergent and protease inhibitors. As a positive control for inflammatory response gene induction, we concurrently treated BV2 cultures with LPS. Gene expression profiling of BV2 microglia 4h after treatment with protein homogenates from the DCN or EC, with LPS, or with D-PBS negative control (N=3)