Department of Pharmacy, Affiliated Hospital of Inner Mongolia Medical University, Hohhot 010059, China; Mongolia Medical School, Inner Mongolia Medical University, Hohhot 010110, China.
Department of Pharmacy, Affiliated Hospital of Inner Mongolia Medical University, Hohhot 010059, China.
Chin J Nat Med. 2021 Jun;19(6):412-421. doi: 10.1016/S1875-5364(21)60040-7.
Although the etiology of sciatica remains uncertain, there is increasing evidence that the disease process of sciatica is associated with the levels of inflammatory factors. Piperine, an alkaloid isolated from Piper nigrum, has previously been demonstrated to inhibit inflammation and analgesic effects. The purpose of this study is to verify the regulatory relationship between miR-520a and p65 and to explore how miR-520a/P65 affects the level of cytokines under the action of piperine, so as to play a therapeutic role in sciatica. Through ELISA experiment, we confirmed that four inflammatory factors (IL-1β, TNF-α, IL-10, TGF-β1) can be used as evaluation indexes of sciatica. The differentially expressed miRNA was screened as miR-520a, by microarray technology, and the downstream target of miR-520a was P65 by bioinformatics. Real-time fluorescence quantitative PCR confirmed that the expression of miR-520a was negatively correlated with pro-inflammatory cytokines, positively correlated with anti-inflammatory cytokines and negatively correlated with p65 expression at mRNA level. The expression of p65 was positively correlated with pro-inflammatory cytokines and negatively correlated with anti-inflammatory cytokines at the protein level verified by ELISA and Western blot. HE staining analysis showed that the nerve fibers were repaired by piprine, the vacuoles were significantly reduced, and the degree of nerve fiber damage was also improved. Immunohistochemical analysis showed that the expression of p65 decreased after administration of piperine. Dual-luciferase reporter gene assay confirmed that the luciferase signal decreased significantly after cotransfection of miR-520a mimics and p65 3'UTR recombinant plasmid. To sum up, in the rat model of non-compressed lumbar disc herniation, piperine plays a significant role in analgesia. MiR-520a can specifically and directly target P65, and piperine can promote the expression of miR-520a, then inhibit the expression of p65, down-regulate the pro-inflammatory factors IL-1β and TNF-α, and up-regulate the effects of anti-inflammatory factors IL-10 and TGF-β1, so as to treat sciatica.
虽然坐骨神经痛的病因仍不确定，但越来越多的证据表明，坐骨神经痛的疾病过程与炎症因子水平有关。胡椒碱是从胡椒中分离出来的一种生物碱，先前已被证明具有抑制炎症和镇痛作用。本研究的目的是验证 miR-520a 和 p65 之间的调节关系，并探讨 miR-520a/P65 如何在胡椒碱作用下影响细胞因子水平，从而在坐骨神经痛中发挥治疗作用。通过 ELISA 实验，我们证实了四种炎症因子（IL-1β、TNF-α、IL-10、TGF-β1）可以作为坐骨神经痛的评价指标。通过微阵列技术筛选出差异表达的 miRNA 为 miR-520a，通过生物信息学筛选出 miR-520a 的下游靶标为 P65。实时荧光定量 PCR 证实 miR-520a 的表达与促炎细胞因子呈负相关，与抗炎细胞因子呈正相关，与 p65 在 mRNA 水平的表达呈负相关。ELISA 和 Western blot 验证了 p65 在蛋白水平上与促炎细胞因子呈正相关，与抗炎细胞因子呈负相关。HE 染色分析表明，胡椒碱修复了神经纤维，空泡明显减少，神经纤维损伤程度也得到改善。免疫组织化学分析表明，胡椒碱给药后 p65 的表达降低。双荧光素酶报告基因检测证实，miR-520a 模拟物和 p65 3'UTR 重组质粒共转染后，荧光素酶信号明显降低。综上所述，在非压迫性腰椎间盘突出症大鼠模型中，胡椒碱在镇痛方面有显著作用。miR-520a 可以特异性地直接靶向 P65，胡椒碱可以促进 miR-520a 的表达，从而抑制 p65 的表达，下调促炎因子 IL-1β 和 TNF-α，上调抗炎因子 IL-10 和 TGF-β1，从而治疗坐骨神经痛。
