| Chronic inflammatory pain is a persistent pain state caused by central or peripheral tissue caused by a variety of reasons.Opioids or non-steroidal anti-inflammatory drugs are often used in clinic,but these drugs have many side effects or poor efficacy,and chronic pain has seriously reduced the patients' quality of life.therefore,it is urgent for us to seek other safe and effective drugs.Spalt-like transcription factor 1 is the tagged gene of MG and one of the most important transcriptional suppressors in MG.Existing studies have confirmed that a variety of transcription factors regulate the phenotype of MG through signal-specific ways and play an important role in the occurrence and development of pain.Some studies have found that Sall1 may have an effect on pain threshold in SNI mouse model.Therefore,it is speculated whether Sall1 also plays an important role in chronic inflammatory pain.Objective:(1)to construct Sall1 overexpression adenovirus vector and verify its transfection efficiency,so as to provide a reliable experimental basis for follow-up experiments.(2)to establish the model of chronic inflammatory hyperalgesia in mice,observe the expression of Sall1 in the model of inflammatory hyperalgesia in mice,and analyze the relationship between the changes of Sall1 and pain threshold in mice.(3)Intrathecal injection of adenovirus overexpressing Sall1 to study the changes of pain threshold in mice with high expression of Sall1 in inflammatory pain model,and to further clarify the relationship between Sall1 and pain threshold,so as to further explore whether Sall1 affects pain and its mechanism by regulating MG.Methods:(1)the gene sequence of Sall1 was obtained by GenBank,and the target gene was connected with the tool vector(pSB50)to get the shuttle plasmid,and then co-transfected with the auxiliary packaging plasmid(pBHG lox ?E1,3 Cre)to obtain the recombinant adenovirus.finally,the transfection efficiency was verified by RT-PCR detection of Sall1.(2)the inflammatory pain model of complete Freund's adjuvant(complete Freund's aduvant,CFA was established in male BALB/c mice,and 30 ? l CFA was subcutaneously injected into the left posterior toe of mice.Von Frey cilia was used to measure the change of mechanical withdraw threshold in mice before and after operation.(3)after Von Frey cilia detection on the second day of CFA model,adenoviruses upregulating the expression of Sall1 in spinal cord were injected intrathecally,and the changes of mechanical pain threshold of mice in CFA group,O-Sall1 group and E-Sall1 group were measured before and after administration.RT-qPCR and Western-blot were used to detect the changes of Sall1 in the spinal cord of CFA mice after adenovirus treatment.Results:(1)the overexpression plasmid PSE3940 of Sall1 was transfected into HEK293 cells.The results showed that the overexpression of Sall11 gene was obvious,and the overexpression plasmid was constructed successfully.(2)the red and swollen foot could be seen on the affected side of CFA mice.After modeling,the mechanical pain threshold of the affected side of mice decreased significantly compared with the normal group and sham operation group,and decreased most significantly one day after modeling,and then increased slowly;the expression of Sall1 protein in CFA group showed an upward trend on the 7th day of modeling,and that of Sall1 mRNA on the 13 th day of modeling.When the pain threshold recovered on the 17 th day of modeling,the expression of Sall1 mRNA and protein returned to the level before modeling.(3)the mechanical pain threshold of CFA mice treated with intrathecal adenovirus was higher than that of CFA group at 3 d,5 d,7 d and 9 d after administration,and there was no difference between O-Sall1 group and CFA group from 11 d after administration,indicating that intrathecal injection of adenovirus could relieve pain in mice,and the expression of SallmRNA and protein in O-Sall1 group was higher than that in E-Sall1 group at the same time point after intrathecal injection of Sall1 overexpressed adenovirus.Conclusion:The MWT of male mice injected subcutaneously with complete Freund's adjuvant decreased significantly,indicating that the model of chronic inflammatory pain was established successfully,and the expression level of Sall1 decreased,and the change of Sall1 expression was consistent with the change of pain threshold.Intrathecal injection of Sall1 overexpression adenovirus can increase the expression of Sall1 in chronic inflammatory pain mice to participate in the regulation of chronic inflammatory pain. |
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