双膦酸盐纳米乳在大鼠实验性自身免疫性神经炎中的应用

Bisphosphonate nanoemulsion attenuates inflammatory response in rat experimental autoimmune neuritis

目的:探讨双膦酸盐(bisphosphonate nanoemulsion,BP)纳米乳对实验性自身免疫性神经炎(experimental autoimmune neuritis,EAN)的治疗作用。方法:BP纳米乳体外处理巨噬细胞系RAW264.7,实时定量PCR(real-time PCR,RT-PCR)检测炎症因子白介素(interleukin,IL)-1β、IL-6、诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)及抑炎因子IL-10、巨噬细胞M2型标记物CD206的表达。构建EAN大鼠模型,给予BP纳米乳处理,测大鼠体重,分析机械疼痛阈值及临床评分,劳克坚牢蓝(luxol fast blue,LFB)染色检测坐骨神经髓鞘病变,免疫组化观察其炎性细胞浸润及病理改变;RT-PCR检测EAN大鼠坐骨神经IL-1β、IL-17、i NOS及基质金属蛋白酶9(matrix metalloproteinase 9,MMP-9)等炎症因子表达。结果:BP与BP纳米乳体外处理巨噬细胞,均可降低IL-1β、iNOS,同时提高IL-10和CD206的表达,促进M1型巨噬细胞向M2型转化。BP纳米乳干预的EAN大鼠体重下降减轻、临床评分降低,机械性疼痛程度缓解,病程明显缩短。同时坐骨神经炎性细胞浸润及髓鞘病变显著减轻,IL-1β、IL-17、i NOS、MMP-9表达降低。结论:BP纳米乳可能通过减轻大鼠坐骨神经炎性细胞浸润及抑制促炎因子表达,来缓解EAN的临床病理变化,提示BP纳米乳可作为神经炎性病变的潜在治疗药物。

Objective:To test potential therapeutic effects of bisphosphonate nanoemulsion(BP nanoemulsion)in the experimental autoimmune neuritis(EAN). Methods:In vitro BP nanoemulsion treatment was performed in macrophage cell line RAW264.7,realtime PCR(RT-PCR)was used to evaluate the expression of inflammatory factors interleukin(IL)-1β,IL-6,inducible nitric oxide synthase(iNOS),anti-inflammatory factor IL-10 and macrophage M2 marker CD206. After successfully establishing EAN rats model,BP nanoemulsion treatment was given,weight change of rats was assessed. The mechanical pain threshold and clinical score were analyzed. Luxol fast blue(LFB)staining was applied to show sciatic myelinopathy. Inflammatory cell infiltration and pathological changes in the sciatic nerve were evaluated by immunohistochemistry. The expressions of inflammatory cytokines IL-1β,IL-17,iNOS and matrix metalloproteinase 9(MMP-9)m RNA level in sciatic nerve were detected by RT-PCR. Results:BP and BP nanoemulsion treatments inhibited IL-1β and iNOS,increased the expressions of IL-10 and CD206,and induced phenotypic switch of macrophage polarization from M1 to M2 subtype in cell culture. In EAN rats:BP nanoemulsion ameliorated body weight loss and neurological signs,ameliorated mechanical allodynia,shortened disease duration,reduced myelin lesions. Meanwhile,it inhibited accumulation of macrophages and other immune cells,and decreased expressions of inflammatory cytokines IL-1β,IL-17,iNOS,and MMP-9 mRNA level in sciatic nerves of EAN rats. Conclusion:BP nanoemulsion can reduce accumulation of immune cells and attenuated inflammatory cytokines in sciatic nerves,and improve the clinical pathological changes of EAN rats. BP nanoemulsion may therefore be considered a potential therapeutic option for neuroinflammatory diseases.