Peripheral nerve injury often causes neuropathic pain and is associated with changes in the expression of numerous proteins in the dorsal horn of the spinal cord. To date, proteomic analysis method has been used to si...Peripheral nerve injury often causes neuropathic pain and is associated with changes in the expression of numerous proteins in the dorsal horn of the spinal cord. To date, proteomic analysis method has been used to simultaneously analyze hundreds or thousands of proteins differentially expressed in the dorsal horn of the spinal cord in rats or dorsal root ganglion of rats with certain type of peripheral nerve injury. However, a proteomic study using a mouse model of neuropathic pain could be attempted because of abundant protein database and the availability of transgenic mice. In this study, whole proteins were extracted from the ipsilateral dorsal half of the 4th-6th lumbar spinal cord in a mouse model of spared nerve injury（SNI）-induced neuropathic pain. In-gel digests of the proteins size-separated on a polyacrylamide gel were subjected to reverse-phase liquid-chromatography coupled with electrospray ionization ion trap tandem mass spectrometry（MS/MS）. After identifying proteins, the data were analyzed with subtractive proteomics using ProtAn, an in-house analytic program. Consequently, 15 downregulated and 35 upregulated proteins were identified in SNI mice. The identified proteins may contribute to the maintenance of neuropathic pain,and may provide new or valuable information in the discovery of new therapeutic targets for neuropathic pain.展开更多
Epigenetic changes in the spinal cord play a key role in the initiation and maintenance of nerve injury-induced neuro pathic pain.N6-methyladenosine(m6A)is one of the most abundant internal RNA modifications and plays...Epigenetic changes in the spinal cord play a key role in the initiation and maintenance of nerve injury-induced neuro pathic pain.N6-methyladenosine(m6A)is one of the most abundant internal RNA modifications and plays an essential function in gene regulation in many diseases.However,the global m6A modification status of mRNA in the spinal cord at different stages after neuropathic pain is unknown.In this study,we established a neuropathic pain model in mice by preserving the complete sural nerve and only damaging the common peroneal nerve.High-throughput methylated RNA immunoprecipitation sequencing res ults showed that after spared nerve injury,there were 55 m6A methylated and diffe rentially expressed genes in the spinal cord.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway results showed that m6A modification triggered inflammatory responses and apoptotic processes in the early stages after spared nerve injury.Over time,the diffe rential gene function at postoperative day 7 was enriched in "positive regulation of neurogenesis" and "positive regulation of neural precursor cell prolife ration." These functions suggested that altered synaptic morphological plasticity was a turning point in neuropathic pain formation and maintenance.Results at postoperative day 14 suggested that the persistence of neuropathic pain might be from lipid metabolic processes,such as "very-low-density lipoprotein particle clearance," "negative regulation of choleste rol transport" and "membrane lipid catabolic process." We detected the expression of m6A enzymes and found elevated mRNA expression of Ythdf2 and Ythdf3 after spared nerve injury modeling.We speculate that m6A reader enzymes also have an important role in neuropathic pain.These results provide a global landscape of mRNA m6A modifications in the spinal cord in the spared nerve injury model at diffe rent stages after injury.展开更多
Objective: To investigate the role of spinal glial cells activation in neuropathic pain in a recently developed spared nerve injury (SNI) animal model by Decosterd and Woolf. Methods: A lesion was made to two of the t...Objective: To investigate the role of spinal glial cells activation in neuropathic pain in a recently developed spared nerve injury (SNI) animal model by Decosterd and Woolf. Methods: A lesion was made to two of the three terminal branches of the sciatic nerve of rats (tibial and common peroneal nerves) leaving the sural nerve intact. Continuous intrathe-cal administration of propentofylline, a glial modulating agent, 1 d before and 5 d after operation, was performed to disrupt spinal cord glia function. The vehicle was intrathecally administrated as control. The paw withdrawal threshold to mechanical stimulation (paw withdrawal mechaical threshold PWMT), body mass and motor function were determined pre- and post-surgery. Results: It produced a prolonged mechanical allodynia in the medial and lateral part of the ipsilateral hind paw in SNL models. The treatment with propentofylline significantly prevented the development of mechanical allodynia located in either medial or lateral plantar surface. Rats in two groups showed normal motor function and body weight increase. Conclusion: SNI model can be applied as a useful method with little variance in searching the mechanism of neuropathic pain. These study suggest that spinal glia activation may contribute to mechanical allodynia induced by SNI.展开更多
Emerging evidence indicates that CXCL12/ CXCR4 signaling is involved in chronic pain. However, few studies have systemically assessed its role in direct nerve injury-induced neuropathic pain and the underlying mech- a...Emerging evidence indicates that CXCL12/ CXCR4 signaling is involved in chronic pain. However, few studies have systemically assessed its role in direct nerve injury-induced neuropathic pain and the underlying mech- anism. Here, we determined that spared nerve injury （SNI） increased the expression of CXCL12 and its cognate receptor CXCR4 in lumbar 5 dorsal root ganglia （DRG） neurons and satellite glial cells. SNI also induced long- lasting upregulation of CXCL12 and CXCR4 in the ipsi- lateral L4-5 spinal cord dorsal horn, characterized by CXCL12 expression in neurons and microglia, and CXCR4 expression in neurons and astrocytes. Moreover, SNI- induced a sustained increase in TNF-α expression in the DRG and spinal cord. Intraperitoneal injection （i.p.） of the TNF-α synthesis inhibitor thalidomide reduced the SNI-in- duced mechanical hypersensitivity and inhibited the expression of CXCL12 in the DRG and spinal cord. Intrathecal injection （i.t.） of the CXCR4 antagonist AMD3100, both 30 rain before and 7 days after SNI, reduced the behavioral signs of allodynia. Rats given an i.t. or i.p. bolus of AMD3100 on day 8 of SNI exhibited attenuated abnormal pain behaviors. The neuropathic pain established following SNI was also impaired by i.t. admin- istration of a CXCL12-neutralizing antibody. Moreover, repetitive i.t. AMD3100 administration prevented the acti- vation of ERK in the spinal cord. The mechanical hyper- sensitivity induced in nai＇ve rats by i.t. CXCL12 was alleviated by pretreatment with the MEK inhibitor PD98059. Collectively, our results revealed that TNF-α might mediate the upregulation of CXCL12 in the DRG and spinal cord following SNI, and that CXCL 12/CXCR4 sig- naling via ERK activation contributes to the development and maintenance of neuropathic pain.展开更多
Neuropathic pain is a chronic debilitating symptom characterized by spontaneous pain and mechanical allodynia. It occurs in distinct forms, including brushevoked dynamic and filament-evoked punctate mechanical allodyn...Neuropathic pain is a chronic debilitating symptom characterized by spontaneous pain and mechanical allodynia. It occurs in distinct forms, including brushevoked dynamic and filament-evoked punctate mechanical allodynia. Potassium channel 2.1(Kir2.1), which exhibits strong inward rectification, is and regulates the activity of lamina I projection neurons. However, the relationship between Kir2.1 channels and mechanical allodynia is still unclear. In this study, we first found that pretreatment with ML133, a selective Kir2.1 inhibitor, by intrathecal administration, preferentially inhibited dynamic, but not punctate, allodynia in mice with spared nerve injury(SNI).Intrathecal injection of low doses of strychnine, a glycine receptor inhibitor, selectively induced dynamic, but not punctate allodynia, not only in na¨?ve but also in ML133-pretreated mice. In contrast, bicuculline, a GABAAreceptor antagonist, induced only punctate, but not dynamic,allodynia. These results indicated the involvement of glycinergic transmission in the development of dynamic allodynia. We further found that SNI significantly suppressed the frequency, but not the amplitude, of the glycinergic spontaneous inhibitory postsynaptic currents(gly-sIPSCs) in neurons on the lamina II-III border of the spinal dorsal horn, and pretreatment with ML133 prevented the SNI-induced gly-sIPSC reduction. Furthermore, 5 days after SNI, ML133, either by intrathecal administration oracute bath perfusion, and strychnine sensitively reversed the SNI-induced dynamic, but not punctate, allodynia and the gly-sIPSC reduction in lamina IIi neurons, respectively.In conclusion, our results suggest that blockade of Kir2.1 channels in the spinal dorsal horn selectively inhibits dynamic, but not punctate, mechanical allodynia by enhancing glycinergic inhibitory transmission.展开更多
基金supported by Basic Science Research Program through the National Research Foundation of Korea(NRF)funded by the Ministry of Education(2015RIDIAIA01059432)
文摘Peripheral nerve injury often causes neuropathic pain and is associated with changes in the expression of numerous proteins in the dorsal horn of the spinal cord. To date, proteomic analysis method has been used to simultaneously analyze hundreds or thousands of proteins differentially expressed in the dorsal horn of the spinal cord in rats or dorsal root ganglion of rats with certain type of peripheral nerve injury. However, a proteomic study using a mouse model of neuropathic pain could be attempted because of abundant protein database and the availability of transgenic mice. In this study, whole proteins were extracted from the ipsilateral dorsal half of the 4th-6th lumbar spinal cord in a mouse model of spared nerve injury（SNI）-induced neuropathic pain. In-gel digests of the proteins size-separated on a polyacrylamide gel were subjected to reverse-phase liquid-chromatography coupled with electrospray ionization ion trap tandem mass spectrometry（MS/MS）. After identifying proteins, the data were analyzed with subtractive proteomics using ProtAn, an in-house analytic program. Consequently, 15 downregulated and 35 upregulated proteins were identified in SNI mice. The identified proteins may contribute to the maintenance of neuropathic pain,and may provide new or valuable information in the discovery of new therapeutic targets for neuropathic pain.
基金National Natural Science Foundation of China,No.819 73305 (to ZQ)Science and Technology Planning Project of Guangzhou of China,No.20190401 0487 (to ZQ)+1 种基金Natural Science Foundation of Guangdong Province,China,No.2021A1515010897 (to TT)Discipline Construction Fund of Cen tral Peoples Hospital of Zhanjiang,Nos.2020A01 (to TT) and 2020A02 (to TT)。
文摘Epigenetic changes in the spinal cord play a key role in the initiation and maintenance of nerve injury-induced neuro pathic pain.N6-methyladenosine(m6A)is one of the most abundant internal RNA modifications and plays an essential function in gene regulation in many diseases.However,the global m6A modification status of mRNA in the spinal cord at different stages after neuropathic pain is unknown.In this study,we established a neuropathic pain model in mice by preserving the complete sural nerve and only damaging the common peroneal nerve.High-throughput methylated RNA immunoprecipitation sequencing res ults showed that after spared nerve injury,there were 55 m6A methylated and diffe rentially expressed genes in the spinal cord.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway results showed that m6A modification triggered inflammatory responses and apoptotic processes in the early stages after spared nerve injury.Over time,the diffe rential gene function at postoperative day 7 was enriched in "positive regulation of neurogenesis" and "positive regulation of neural precursor cell prolife ration." These functions suggested that altered synaptic morphological plasticity was a turning point in neuropathic pain formation and maintenance.Results at postoperative day 14 suggested that the persistence of neuropathic pain might be from lipid metabolic processes,such as "very-low-density lipoprotein particle clearance," "negative regulation of choleste rol transport" and "membrane lipid catabolic process." We detected the expression of m6A enzymes and found elevated mRNA expression of Ythdf2 and Ythdf3 after spared nerve injury modeling.We speculate that m6A reader enzymes also have an important role in neuropathic pain.These results provide a global landscape of mRNA m6A modifications in the spinal cord in the spared nerve injury model at diffe rent stages after injury.
文摘Objective: To investigate the role of spinal glial cells activation in neuropathic pain in a recently developed spared nerve injury (SNI) animal model by Decosterd and Woolf. Methods: A lesion was made to two of the three terminal branches of the sciatic nerve of rats (tibial and common peroneal nerves) leaving the sural nerve intact. Continuous intrathe-cal administration of propentofylline, a glial modulating agent, 1 d before and 5 d after operation, was performed to disrupt spinal cord glia function. The vehicle was intrathecally administrated as control. The paw withdrawal threshold to mechanical stimulation (paw withdrawal mechaical threshold PWMT), body mass and motor function were determined pre- and post-surgery. Results: It produced a prolonged mechanical allodynia in the medial and lateral part of the ipsilateral hind paw in SNL models. The treatment with propentofylline significantly prevented the development of mechanical allodynia located in either medial or lateral plantar surface. Rats in two groups showed normal motor function and body weight increase. Conclusion: SNI model can be applied as a useful method with little variance in searching the mechanism of neuropathic pain. These study suggest that spinal glia activation may contribute to mechanical allodynia induced by SNI.
基金supported by grants from the National Natural Science Foundation of China(31171070,81171060,81501070and 81571079)
文摘Emerging evidence indicates that CXCL12/ CXCR4 signaling is involved in chronic pain. However, few studies have systemically assessed its role in direct nerve injury-induced neuropathic pain and the underlying mech- anism. Here, we determined that spared nerve injury （SNI） increased the expression of CXCL12 and its cognate receptor CXCR4 in lumbar 5 dorsal root ganglia （DRG） neurons and satellite glial cells. SNI also induced long- lasting upregulation of CXCL12 and CXCR4 in the ipsi- lateral L4-5 spinal cord dorsal horn, characterized by CXCL12 expression in neurons and microglia, and CXCR4 expression in neurons and astrocytes. Moreover, SNI- induced a sustained increase in TNF-α expression in the DRG and spinal cord. Intraperitoneal injection （i.p.） of the TNF-α synthesis inhibitor thalidomide reduced the SNI-in- duced mechanical hypersensitivity and inhibited the expression of CXCL12 in the DRG and spinal cord. Intrathecal injection （i.t.） of the CXCR4 antagonist AMD3100, both 30 rain before and 7 days after SNI, reduced the behavioral signs of allodynia. Rats given an i.t. or i.p. bolus of AMD3100 on day 8 of SNI exhibited attenuated abnormal pain behaviors. The neuropathic pain established following SNI was also impaired by i.t. admin- istration of a CXCL12-neutralizing antibody. Moreover, repetitive i.t. AMD3100 administration prevented the acti- vation of ERK in the spinal cord. The mechanical hyper- sensitivity induced in nai＇ve rats by i.t. CXCL12 was alleviated by pretreatment with the MEK inhibitor PD98059. Collectively, our results revealed that TNF-α might mediate the upregulation of CXCL12 in the DRG and spinal cord following SNI, and that CXCL 12/CXCR4 sig- naling via ERK activation contributes to the development and maintenance of neuropathic pain.
基金supported by grants from the National Natural Science Foundation of China (31771188 and 31471027)the Science and Technology Commission of Shanghai Municipality, China (13DJ1400302)
文摘Neuropathic pain is a chronic debilitating symptom characterized by spontaneous pain and mechanical allodynia. It occurs in distinct forms, including brushevoked dynamic and filament-evoked punctate mechanical allodynia. Potassium channel 2.1(Kir2.1), which exhibits strong inward rectification, is and regulates the activity of lamina I projection neurons. However, the relationship between Kir2.1 channels and mechanical allodynia is still unclear. In this study, we first found that pretreatment with ML133, a selective Kir2.1 inhibitor, by intrathecal administration, preferentially inhibited dynamic, but not punctate, allodynia in mice with spared nerve injury(SNI).Intrathecal injection of low doses of strychnine, a glycine receptor inhibitor, selectively induced dynamic, but not punctate allodynia, not only in na¨?ve but also in ML133-pretreated mice. In contrast, bicuculline, a GABAAreceptor antagonist, induced only punctate, but not dynamic,allodynia. These results indicated the involvement of glycinergic transmission in the development of dynamic allodynia. We further found that SNI significantly suppressed the frequency, but not the amplitude, of the glycinergic spontaneous inhibitory postsynaptic currents(gly-sIPSCs) in neurons on the lamina II-III border of the spinal dorsal horn, and pretreatment with ML133 prevented the SNI-induced gly-sIPSC reduction. Furthermore, 5 days after SNI, ML133, either by intrathecal administration oracute bath perfusion, and strychnine sensitively reversed the SNI-induced dynamic, but not punctate, allodynia and the gly-sIPSC reduction in lamina IIi neurons, respectively.In conclusion, our results suggest that blockade of Kir2.1 channels in the spinal dorsal horn selectively inhibits dynamic, but not punctate, mechanical allodynia by enhancing glycinergic inhibitory transmission.
