Fig. 6.

Knockdown of NEAT1 in microglia protects primary neurons from OGD injury. a Immunofluorescence images of primary neurons (NeuN) and astrocytes (GFAP) in red under normoxia conditions. Nuclei were counterstained with DAPI in blue. b A summary of the experimental paradigm for the in vitro co-culture model. Primary microglia and astrocytes were isolated from neonatal C57BL/6 mice on postnatal day 0, whereas primary cortical neurons were obtained from E16.5 mouse embryos. The neurons were subjected to 10 h of OGD, whereas astrocytes underwent 8 h of OGD. Microglia were subjected to four conditions (normoxia, OGD, OGD + ASO scramble, OGD + ASO NEAT1), followed by co-culture with hypoxic neuorns or astrocytes. Afterwards, they were cultured under normoxic conditions for 24 h. c, d The MTT assay was performed to assess the cell survival of primary neurons and astrocytes co-cultured with microglia, which were treated in different ways. As such, microglia were treated with ASO NEAT1 knockdown constructs or with scramble constructs. All results are expressed as mean ± standard deviation and analyzed by one-way ANOVA followed by Tukey's post-hoc-test. NS, no significance, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. Abbreviations: NEAT1, nuclear paraspeckle assembly transcript 1; ASO, antisense oligonucleotide; OGD, oxygen-glucose-deprivation; PM, primary microglia; N, neuron; AS, astrocyte