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The expression of CX3CL1 and CX3CR1 after subarachnoid hemorrhageObjective:To observe the expression levels of CX3CL1 and CX3CR1 in brain tissues of human and rats after subarachnoid hemorrhage(SAH).Methods:First,8 brain tissue samples from clinical patients were collected,of which 5 were in the Non-SAH group and 3 were in the SAH group.The expressions of CX3CL1 and CX3CR1 in human brain tissue were detected by Western blot.Then,84 Sprague Dawley(SD)rats were divided into 7 groups,the sham group and 3 h,6h,12h,24h,72h,7d after SAH,and 12 rast in each group.When the set time was reached after SAH,all rats were sacrificed immediately.The temporal base brain tissues of 6 rats in each group was collected for western blot,and the brain tissue of the remaining 6 rats was fixed for immunofluorescence.Results:1.The expressions of CX3CL1 and CX3CR1 in the brain tissue of clinical patients were reduced after SAH.2.After SAH,the expressions of CX3CL1 and CX3CR1 in brain tissues of SD rats were decreased and then increased,and reached the lowest level at 12 h after SAH;3.In brain tissues,CX3CL1 was expressed in neurons,while CX3CR1 was expressed in microglia.Conclusions:In rat brain tissue,CX3CL1 is mainly expressed in neurons,while CX3CR1 is only expressed in microglia.After SAH,the expression of CX3CL1 and CX3CR1 in the brain tissue of SD rats both decreased first and then increased,and the expression of both decreased in the brain tissue of clinical patients.Part ? The effect of CX3CL1/CX3CR1 axis on early brain injury after subarachnoid hemorrhageObjective:To investigate the effect of CX3CL1/CX3CR1 axis on early brain injury(EBI)after SAH.Methods:88 rats(113 rats were used,96 rats were survived,8 rats excluded)were divided into 4 groups:sham group,SAH group,SAH+Vector group and SAH+Over-CX3CL1/CX3CR1 group,and 22 rats in each group.The 10 surviving rats were randomly selected for behavioral scoring,adhesive-removal test,rotarod test and water maze test;the remaining 12 rats were sacrificed,and the brain tissues and serum were used for western blot,Fluoro-Jade C(FJC)staining and enzyme linked immunosorbent assay(ELISA).Results:1.Compared with the sham group,the number of FJC-positive cells in the brain tissue of the SAH group was significantly increased.Compared with the SAH+Vector group,the number of FJC-positive cells in the SAH+Over-CX3CL1/CX3CR1 group was significantly reduced.2.Compared with the sham group,the rats in SAH group took higher neurobehavioral score significantly,took longer to remove stickers,took less time to exercise and took longer to reach the immersion platform in the water maze significantly.Compared with the SAH+Vector group,the rats in SAH+Over-CX3CL1/CX3CR1 group took lower neurobehavioral score significantly,took less time to remove stickers,took longer to exercise and took less time to reach the immersion platform in the water maze significantly.3.Compared with sham group,the expression of CD45,major histocompatibility complex(MHC)Class II and CCAAT-enhancer binding protein ?(C/EBP-?)were significantly increased,while monocyte chemotactic protein-induced protein 1(MCPIP1)and Runx1 expression were significantly decreased;and compared with the SAH+Vector group,the expressions of CD45,MHC Class II and C/EBP-? in the SAH+Over-CX3CL1/CX3CR1 group were significantly reduced,while the expressions of MCPIP1 and Runx1 were significantly increased.4.Compared with the sham group,the serum levels of tumor necrosis factor ?(TNF-?),Interleukin 1?(IL-1?)and Complement 1q(C1q)in the SAH group rats were significantly increased;and compared with the SAH+Vector group,the levels of TNF-?,IL-1? and C1q in the SAH+Over-CX3CL1/CX3CR1 group were significantly decreased.Conclusions:After SAH,the overexpression of CX3CL1/CX3CR1 can reduce neuronal degeneration in rat brain tissue,reduce inflammation,and improve neurobehavioral function.This indicates that CX3CL1/CX3CR1 plays an important role in early brain injury after SAH in rats.Part ? CX3CL1/CX3CR1 axis promoted the delivery of exosomal microRNA-124 from neuron to microglia after subarachnoid hemorrhageObjective:To investigate the effect of CX3CL1/CX3CR1 axis on the delivery of exosomal miR-124.Methods:First,the co-cultured neuron-microglial cells were divided into 4 groups:control group,OxyHb group,OxyHb+Vector group and OxyHb+Over-CX3CL1/CX3CR1 group,and then the effect of CX3CL1/CX3CR1 axis on the delivery of miR-124 from neurons to microglia was investigated by immunofluorescence staining.Then,the co-cultured neuron-microglial cells were divided into 4 groups:control group,OxyHb group,OxyHb+Vector group,and OxyHb+Over-CX3CL1/CX3CR1 group,and then the effect of CX3CL1/CX3CR1 axis on the delivery of exosomes was investigated by immunofluorescence staining.In addition,exosomes inhibitor GW4869 and polymerase chain reaction(PCR)were used to verify that exosomes were vectors for miR-124 transport.Results:1.After SAH,neuron-derived miR-124 was reduced in microglia,and after overexpression of CX3CL1/CX3CR1,neuron-derived miR-124 was increased in microglia.2.After SAH,neuron-specific enolase(NSE)-positive exosomal-like vesicles decreased in the area of microglia;after overexpression of CX3CL1/CX3CR1,NSE-positive exosomal-like vesicles were increased in the area of microglia.3.After SAH,exosomal miR-124 markedly increased both in control and vehicle group,but the treatments with GW4869 significantly reduced the exosomal miR-124 derived from neurons.Conclusions:The CX3CL1/CX3CR1 axis promoted the delivery of miR-124 from neurons to microglia via exosomes. |
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