Hydrogen-Rich Saline Attenuates Chronic Allodynia after Bone Fractures via Reducing Spinal CXCL1/CXCR2-Mediated Iron Accumulation in Mice

Abstract

Purpose: Neuroinflammation often initiates iron overload in the pathogenesis of neurological disorders. Chemokine-driven neuroinflammation is required for central sensitization and chronic allodynia following fractures, but specific molecular modulations are elusive. This present study explored whether hydrogen-rich saline, as one potent anti-inflammatory pharmaceutical, could alleviate fracture-caused allodynia by suppressing chemokine CXCL1 expression and iron overload. Methods: A mouse model of tibial fracture with intramedullary pinning was employed for establishing chronic allodynia. Three applications of hydrogen-rich saline (1, 5 or 10 mL/kg) were administrated intraperitoneally on a daily basis from days 4 to 6 following fractures. Spinal CXCL1 and its receptor CXCR2 levels, transferrin receptor 1 (TfR1) expression and iron concentration were examined. Recombinant CXCL1, a selective CXCR2 antagonist and an iron chelator were used for verification of mechanisms. Results: Repetitive injections of hydrogen-rich saline (5 and 10 mL/kg but not 1 mL/kg) prevent fracture-caused mechanical allodynia and cold allodynia in a dose-dependent manner. Single exposure to hydrogen-rich saline (10 mL/kg) on day 14 after orthopedic surgeries controls the established persistent fracture allodynia. Furthermore, hydrogen-rich saline therapy reduces spinal CXCL1/CXCR2 over-expression and TfR1-mediated iron accumulation in fracture mice. Spinal CXCR2 antagonism impairs allodynia and iron overload following fracture surgery. Intrathecal delivery of recombinant CXCL1 induces acute allodynia and spinal iron overload, which is reversed by hydrogen-rich saline. Moreover, iron chelation alleviates exogenous CXCL1-induced acute pain behaviors. Conclusions: These findings identify that hydrogen-rich saline confers protection against fracture-caused chronic allodynia via spinal down-modulation of CXCL1-dependent TfR1-mediated iron accumulation in mice.

Annexin 1 Reduces Dermatitis-Induced Itch and Cholestatic Itch through Inhibiting Neuroinflammation and Iron Overload in the Spinal Dorsal Horn of Mice

“…Dysregulation of iron homeostasis in neurons causes neurotoxicity and neuroplasticity, which is a key factor in the neurobiology of disorders [42,43]. More strikingly, neuronal iron overload underlies neuroinflammation-induced synaptic and behavioral plasticity [15,17]. A complete complement of iron proteins mediates intracellular iron homeostasis in the central nervous system, comprising IRP1, DMT1 and TfR1.…”

“…Our previous study has highlighted the requirement of DMT1-dependent neuronal iron accumulation for spine generation and excitatory neuroplasticity in the pathogenesis of tibial fractures-associated long-term pain and remifentanil-caused hypernociception [27,45,46]. TfR1-mediated iron overload is also identified to be one crucial target of astrocyte-dependent neuroinflammation in mechanical allodynia following orthopedic surgeries [17]. However, the overall link between ANXA1, IL-17, astrocyte activation and iron overload in chronic pruritus is unknown.…”

“…By digesting the tissues, the iron content of the spinal dorsal horns was measured using a flame atomic absorption spectrophotometer (Shimadzu AA-6800, Kyoto, Japan) at 248.3 nm [17,27]. Samples (0.1-0.2 g) were dried at 60 • C for 12 h, then digested with 1 mL of 60% nitric acid at 100 • C in a water bath for 2 h. After adding 0.5 mL of hydrogen peroxide, the process was continued for an additional 0.5 h at boiling point.…”

“…Although the upregulation of keratinocytic IL-17 in dry skin and psoriatic skin has been identified [14], it is still unclear whether the spinal IL-17/IL-17RA pathway is required for chronic itch facilitation. More importantly, neuronal iron overload following neuroinflammation is pivotal a determinant of excitatory synaptic plasticity via dendritic spine morphogenesis in chronic nociceptive states [15][16][17]. However, little has been explored about the interaction between IL-17-dependent neuroinflammation, iron overload and synaptic plasticity in the central circuit mechanisms of chronic itch.…”

Glabridin Therapy Reduces Chronic Allodynia, Spinal Microgliosis, and Dendritic Spine Generation by Inhibiting Fractalkine-CX3CR1 Signaling in a Mouse Model of Tibial Fractures

“…The tibial fracture-elicited chronic nociceptive model was established as in previous reports [16,25]. Specifically, under sevoflurane (induction, 3.0%; surgery, 1.5%) anesthesia, a muscle disassociation was performed after a short incision from the knee to the midshaft of the left tibia.…”