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In the context of EDC, elevated LPS levels caused by gut microbiota dysbiosis can be considered a third hit. Building upon existing central pathologies, this third hit exacerbates neuroinflammation, synergistically worsening both epileptic seizures and depressive symptoms, thereby perpetuating a vicious cycle (Arora et al., 2022)

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Figure 8 : Reaction scheme showing the generation of chemiluminescence when luminol reacts with oxygen

In vitro kinase assay To prepare the AMPK kinase complex, FLAG-AMPK was transfected into HEK293 cells which were treated with AICAR (10 mM) for 12 h to stimulate AMPK activation and then immunoprecipitated (IPed) using anti-FLAG agarose beads according to the manufacturers instructions