The Study Of Protein Tyrosine Phosphatase Receptor Type D Regulating Neuropathic Pain After Nerve Injury Via The STING/IFN-? Pathway

BackgroundNeuropathic pain(NP)is a chronic pain condition caused by injury or dysfunction of the somatosensory neuron system.It usually presents as stubborn spontaneous pain,hyperalgesia,ectopic pain,and paresthesia,which seriously affects the quality of life of patients and is classified as a disease with a high disability rate.It is usually treated with drugs such as tricyclic antidepressants,opioids,lidocaine,gabapentin,and anti-seizure medications but these tend not to be satisfactory as most of these drugs have serious side effects,for example,the long-term use of morphine results in both drug resistance and addiction.Therefore,the identification of safe and efficient therapeutic targets for NP is urgently needed.Protein tyrosine phosphatase receptor Type D(PTPRD)is a transmembrane protein that is highly expressed in the central nervous system and is involved in dephosphorylation and neuronal adhesion.Current studies have suggested PTPRD as a new target for addiction treatment.Inhibition of PTPRD activity can effectively relieve cocaine addiction,and its specific small-molecule inhibitor 7-BIA has consequently been developed.It is important to note that PTPRD is significantly upregulated in the dorsal root ganglion(DRG)in animal models of neuropathic pain,although whether high expression of PTPRD is associated with neuropathic pain remains unclear.ObjectiveThis study aimed to determine the involvement of PTPRD in the regulation of neuropathic pain and to explore the associated molecular mechanism,thus providing a new target for the clinical treatment of NP.Methods1.A chronic constriction injury(CCI)neuropathic pain model was induced in C57 mice.Behavioral tests were performed before and 1,3,5,7,14,and 21 days after the operation.These included the paw withdrawal threshold(PWT),paw withdrawal latency(PWL),and paw withdrawal frequency(PWF).2.Immunofluorescence staining and western blotting were used to detect the expression and distribution of PTPRD in the DRG of CCI mice.3.A plasmid expression vector carrying sh RNA was constructed and packaged with lentivirus.PTPRD knockdown in the DRG was performed by local injection.Alterations in the pain thresholds of the CCI mice after PTPRD knockdown were determined by measurement of the PWT,PWL,and PWF,and Rotarod and Open field tests were performed to evaluate whether PTPRD knockdown affected the motor function of the mice.4.The levels of tumor necrosis factor-?(TNF-?),interleukin-6(IL-6),interleukin-1?(IL-1?),interleukin-10(IL-10),and interferon-?(IFN-?)in the DRG of CCI mice after PTPRD knockdown were determined by real-time quantitative PCR(RT-q PCR),western blotting,and enzyme-linked immunosorbent assay(ELISA).The Stimulator of Interferon gene(STING)was found to be the downstream site of pro-inflammatory factors regulated by PTPRD.5.STING was inhibited in PTPRD-knockdown mice by intrathecal injection of H-151,followed by the behavioral tests(PWT,PWL,and PWF).IFN-?levels in the DRG were measured seven days after CCI by ELISA and RT-q PCR.6.Intraperitoneal injection of the PTPRD small-molecule inhibitor 7-BIA was used to inhibit the activity of PTPRD in CCI mice.This was followed by the behavioral tests(PWT,PWL,PWF)and Rotarod and Open field tests to determine whether 7-BIA affected the motor function of the mice.Western blotting and ELISA were used to detect the expression levels of STING and IFN-?in the DRGs of CCI mice after 7-BIA treatment.Results1.Compared with the sham group,the PWT and PWL of CCI mice were significantly decreased,while the PWF was significantly increased.2.Immunofluorescence staining showed that PTPRD co-localized with Tuj1+sensory neurons in the DRG,while minimal co-localization was observed with GFAP~+astrocytes and S100?~+satellite glia.3.In normal conditions,PTPRD was expressed at low levels in the DRG but expression increased significantly 7-14 days after CCI,corresponding to the behavioral tests of the CCI mice.4.Compared with the control group,both the PWT and PWL were significantly increased after knockdown of PTPRD in the DRG,while the PWF was significantly decreased,and there was no effect on the motor function of mice.After PTPRD knockdown,the expression of IFN-?in the DRG increased and STING expression upstream of IFN-?increased.5.After inhibition of STING by H-151 in the PTPRD-knockdown mice,the PWT and PWL decreased,the PWF increased,and IFN-?expression in the DRG decreased compared with the control group.6.After inhibition of PTPRD activity in CCI mice with 7-BIA,the PWT and PWL increased and the PWF decreased while the motor function was not affected,and the expression levels of STING and IFN-?in DRG increased.Conclusion1.PTPRD was significantly increased in the dorsal root ganglion of mice after CCI and was localized to sensory neurons.2.Knocking down PTPRD in the DRG of CCI mice could significantly improve neuropathic pain.3.PTPRD regulated neuropathic pain through the STING-IFN-I pathway.4.The PTPRD small-molecule inhibitor 7-BIA could effectively relieve hyperalgesia in CCI mice,suggesting that PTPRD may be a new clinical target for the treatment of neuropathic pain.