Supporting data for "Phosphorylated SOX9 Drives Neuroinflammatory Astrocyte Subtypes by Boosting Hexokinase 1 in Neuropathic Pain"

Neuropathic pain (NP) is elicited by multiple etiologies, including injuries or diseases, casing spontaneous and persistent pain, which exerts adverse impacts on patients’ quality of life. Currently, there is no effective treatments. Deleterious activity of astrocytes in the spinal cord dorsal horn (SDH) upon the nerve injury is thought to be one of the central mechanisms in driving long-lasting neuropathic pain. Exploring the molecular mechanisms underlying dynamic changes of astrocyte activities is critical for resolving the chronification of NP. Due to heavily relying on glycolysis of astrocytes for energy production, glycolysis represents a central metabolic pathway in astrocytes under physiological conditions, and the regulation of glycolysis is essential for the conversion between neuroprotective and neurotoxic outcomes under pathological status. Here, by using scRNA-sequencing and in vivo functional analysis, we found that HK1 in astrocytes, the first rate-limiting enzyme of glycolytic pathway, enhances glycolytic flux and contributes its reactivity, leading to the production of pro-inflammatory cytokines and chemokines, thus promoting the central sensitizations in the SDH under NP status. Additionally, SOX9 is the key regulator for gliogenesis and is also identified as a nuclei marker for astrocytes. Our results suggested that phosphorylation of SOX9 at the site of S181 positively correlated with SDH's pathological changes during pain development. Interestingly, Genetic manipulation of SOX9 phosphorylation regulates its transcriptional activities by increasing the nucleus localization of SOX9 proteins. Phosphorylated SOX9 (p-SOX9) promotes the HK1 expression in transcriptional level and accelerates HK1-mediated astrocytes transformation and inflammatory response to aggravate NP development. Therefore, specifically inhibiting HK1 or SOX9 phosphorylation to reduce the deleterious reactive astrocytes and thereafter neuroinflammation is the potential therapeutic strategy for the treatment of NP.