Microglial HMGB2 mediates neuroinflammation and brain damage in Cx3cr1-creERT2 epileptic mice through astrocyte-derived CCL2

  • Prog Neuropsychopharmacol Biol Psychiatry. 2026 Jun 20:147:111763. doi: 10.1016/j.pnpbp.2026.111763.
Xinru Li  1 ,  Yiting Wang  1 ,  Dandan Sheng  1 ,  Jing Liu  1 ,  Bibi Yazgulyyeva  1 ,  Bo Xiao  1 ,  Luo Zhou  1 ,  Weiping Liu  2
Affiliations
  • 1. Department of Neurology, Xiangya Hospital, Central South University, Changsha, Hunan, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, China.
  • 2. Department of Neurology, Xiangya Hospital, Central South University, Changsha, Hunan, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, China. Electronic address: [email protected].
PMID: 42218973 DOI: 10.1016/j.pnpbp.2026.111763
Abstract

Background: The activation of microglia in the epileptic brain is a complex process. As a mediator of the inflammatory response of microglia, the role of High Mobility Group Box 2 (HMGB2) in epilepsy is not well understood.

Methods: We investigated the role of microglial HMGB2 in epilepsy, by knocking down HMGB2 using U6-loxP-stop-loxP-shRNA-HMGB2-EF1-ZsGreen (sh-HMGB2) in microglia-specific Cx3cr1-creERT2 mice. Behavioral tests and pathological examinations were utilized to assess brain damage in CX3CR1-Cre recombinase Enhanced by Tamoxifen 2 (Cx3cr1-creERT2) mice. Microglia from Cx3cr1-creERT2 mice in different treatment groups were extracted for transcriptome Sequencing. In vitro, microglia isolated from Cx3cr1-creERT2 mice injected with AAV-sh-NC or AAV-sh-HMGB2 were stimulated with magnesium-free medium (MGF) to induce an epileptic model, without kainic acid (KA) induction. Astrocytes were stimulated with microglial culture medium (MCM) from different treatment groups to investigate the effect of microglial HMGB2 knockdown on astrocyte derived CCL2 expression.

Results: The results showed that HMGB2 was upregulated in the epileptic Cx3cr1-creERT2 mice. Knockdown of HMGB2 in microglia attenuated the degree of epilepsy and neurological damage, and inhibited microglia activation in Cx3cr1-creERT2 mice. HMGB2 knockdown increased the CD4+T content, while decreased the levels of CD8+T, CD3-CD56+, CD11b+CD45+, and Ly6G in the spleen and brain tissue of Cx3cr1-creERT2 mice. Additionally, the expression of CCL2 was reduced and CCL2 was co-localized with the astrocyte marker GFAP in HMGB2-knockdown microglia. In vitro experiments demonstrated that knockdown of HMGB2 reduced the levels of TNF α and IL-6, and increased the levels of IL-4 in MGF-induced microglia. Meanwhile, knockdown of HMGB2 significantly inhibited the level of astrocyte-derived CCL2 and reduced the number of migrating neutrophils. Transcriptomic results showed that knockdown of HMGB2 in microglia mainly affected signaling pathways involving in metabolism, synthesis of cellular structural components, and cellular communication transduction.

Conclusion: Microglial HMGB2 knock down ameliorates brain damage in epileptic Cx3cr1-creERT2 mice by reducing the immune-inflammatory response mediated by astrocyte-derived CCL2.

Keywords
Brain damage; Epilepsy; HMGB2; Immune-inflammatory response; Microglia.
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