| AIM Diabetic neuropathic pain(DNP)is a common and serious complication of the nervous system in patients with diabetes,characterized by pain in both distal lower limbs and high morbidity.Although some progress has been made in the treatment of DNP in recent years,the drug efficacy for DNP is still fairly limited.MicroRNAs(miRNAs)are a class of small non-coding RNAs that control post-transcriptional regulation of gene expression by binding to target mRNAs.Involved in a variety of cellular activities,including cell growth,differentiation,development and apoptosis,miRNAs have been implicated in many diseases and have potential to treat them.Studies have shown that a significant number of miRNAs are highly dysregulated in physiological processes such as nervous system,immune system and pain,and may be a key link in pain treatment.In previous studies,we found that miR-184-5p was significantly down-regulated in DNP,but whether miR-184-5p was involved in DNP was not clear.This study aims to explore the expression of miR-184-5p in DNP mouse models,and whether it is involved in DNP and its possible mechanism,so as to provide a reliable theoretical basis and basis for the future clinical treatment of DNP and the research and development of related drugs.Methods In this study,Balb/c mice aged 5-8 weeks were injected with streptozotocin(STZ)to establish a DNP model.Pain behavior was evaluated by paw withdrawal mechanical threshold(PWMT).Firstly,the expression level of miR-184-5p was detected to verify its dysregulation in DNP model.Then,bioinformatics analysis was used to predict the potential target genes of miR-184-5p.Double luciferase reporting assay was used to confirm whether there was a target relationship between miR-184-5p and predicted target genes.Fluorescence in situ hybridization(FISH)and immunofluorescence were used to detect the co-expression of miR-184-5p and its target genes.Then,real-time quantitative fluorescent polymerase chain reaction and western blot were used to analyze the expression of relevant target genes and proteins.At the animal level,DNP mice were injected intrathecally with miR-184-5p agomir to observe the expression of miR-184-5p and its target genes and the changes of pain behavior.In addition,normal mice were injected with miR-184-5p antagomir to observe the expression of miR-184-5p and its target genes,as well as the changes of pain behavior.Results(1)Mice injected intraperitoneally with STZ showed elevated blood sugar,weight loss,increased water intake on the 7th day after injection and positive reactions in pain behavior(P < 0.0001).The DNP model was successfully established.(2)In the lumbar spinal dorsal horn of DNP mouse model,miR-184-5p was significantly down-regulated(P < 0.05).(3)Bioinformatics technology was used to predict that chemokine C-C motif ligand 1(CCL1)was a potential target of miR-184-5p,and double luciferase reporting experiments showed that CCL1 was correlated with miR-184-5p.(4)In situ hybridization fluorescence results showed co-localization of miR-184-5p and CCL1.(5)In the lumbar spinal dorsal horn of DNP mouse model,the expression of CCL1 protein and C-C motif chemokine receptor 8(CCR8)gene increased(P < 0.01).(6)Overexpression of miR-184-5p significantly reduced neuropathic pain in DNP mice(P < 0.05),and decreased the expression of CCL1/CCR8 in DNP models(P <0.0001).(7)Intrathecal miR-184-5p antagomir in na(?)ve mice could increase the expression of CCL1/CCR8 in spinal dorsal horn(P < 0.01)and cause pain behavior(P < 0.0001).Conclusion This study shows that miR-184-5p regulates DNP by inhibiting the expression of CCL1/CCR8 signaling. |
