CIRBP Exerts Neuroprotection Against Ferroptosis Via ABCA1 After Traumatic Brain Injury In Mice

Background:As one of the most serious medical problems worldwide,traumatic brain injury(TBI)is the most common type of injury in the clinic and in forensic practice.There are many different causes in TBI,including blows to the head,penetration,rapid acceleration or deceleration of the head,or exposure to an explosion.As the main cause of death and disability among young people,the estimation of traumatic brain injury time and the assessment of cognitive impairment are hot and difficult points in forensic research.Ferroptosis,a regulated cell death dependent on iron and lipid oxidation products,plays an important role in brain and nervous system diseases,and this type of iron-dependent cell death is morphologically,biochemically and genetically distinct from other forms of programming cell death.Ferroptosis is primarily characterized by dysregulation of iron homeostasis,glutathione(GSH)depletion,and lipid peroxidation,including specific oxidation of adrenal-containing phosphatidylethanolamine and arachidonic acid.Our team has shown that ferroptosis is involved in the occurrence and development of traumatic brain injury.Cold-induced RNA-binding protein(CIRBP)is a cold-shock protein present in the nucleus of mammalian cells,which can bind to RNA,regulate the transcription of corresponding genes,and plays an important role in brain and nervous system diseases.Studies have shown that overexpression of CIRBP can protect cells from oxidative stress damage by reducing ROS levels.ATP-binding cassette transporter A1(ABCA1)is a transmembrane protein that is widely expressed in many tissues and plays an important role in maintaining the homeostasis of cellular lipid metabolism by removing excess intracellular phospholipids and cholesterol.ABCA1 is involved in the regulation Lipid accumulation and ROS induced by Prussian blue nanoparticles,while up-regulated ABCA1 reduces brain β-amyloid(Aβ)levels and improves functional recovery after TBI.However,the temporal expression profile of CIRBP after TBI is unclear,and whether there is a potential link between CIRBP and ABCA1 in the protective mechanism of ferroptosis after TBI has not been clearly elucidated.Methods:In this study,a mouse model of traumatic brain injury was established,and the expression changes of CIRBP and ABC A1 were detected over time(multiple time points from 12h to 14d),and then the effect of CIRBP on ferroptosis was studied in the mouse TBI model by using CIRBP overexpression or knockdown plasmids.Then,through molecular biology such as(immunofluorescence and Western Blot),morphology such as(Prussian Blue staining and immunohistochemistry)and behavioral tests such as(Wire-Grip Test,Morris Water Maze,Open Field Test)to study the effect of CIRBP on traumatic brain injury and its mechanism of ferroptosis.To further investigate whether ABCA1 mediates the function of CIRBP on TBI-induced ferroptosis,we constructed neuron-specific ABCA1 conditional knockout(ABCA1-KO)mice and used molecular biology and morphological methods to study the differences in iron deposition,oxidative stress and lipid peroxidation levels in rats under physiological and pathological conditions.Finally,we also tested the effect of CIRBP on injury of HT-22 cell line,and further explored the mechanism of ferroptosis and action of CIRBP after injury in vitro.Result:(1)CIRBP mainly reached its peak at 3d to 7d,while ABCA1 reached its peak at 3d,decreased slightly at 7d,and returned to baseline at 14d.(2)Overexpression of CIRBP attenuated the level of ferroptosis after TBI.(3)Overexpression of CIRBP attenuated the degree of iron deposition and neurological dysfunction after TBI.(4)Overexpression of CIRBP attenuated brain behavioral damage and cerebral edema after TBI.(5)Neuroprotection by CIRBP following TBI is largely abolished in ABCA1-KO mice.(6)In vitro,overexpression of CIRBP attenuated lipopolysaccharide(LPS)induced cell death and ferroptosis.Conclusion:(1)CIRBP mainly peaked at 3d to 7d post TBI,while ABCA1 peaked mainly at 3d.Overexpression of CIRBP inhibited the occurrence of ferroptosis,brain damage and improved neurological dysfunction after TBI.(2)Overexpression of CIRBP plays an important role in the development of ferroptosis,while in ABCA1-KO mice,the protective effect of CIRBP on TBI is abolished.(3)Overexpression of CIRBP inhibited LPSinduced ferroptosis,accumulation of ROS,and cell death in HT-22 cells.