[關(guān)鍵詞]
[摘要]
目的 探討白及多糖(BSP)對巨噬細(xì)胞M1型極化過程的調(diào)節(jié)作用及其潛在機(jī)制。方法 誘導(dǎo)THP-1細(xì)胞分化為M0或M1型巨噬細(xì)胞,并給予不同質(zhì)量濃度(0.001、0.010、0.100、1.000、10.000 μg·mL-1)的BSP處理,觀察細(xì)胞貼壁程度及形態(tài),通過CCK-8法檢測BSP對細(xì)胞活力的影響。選擇合適的劑量后,運(yùn)用實(shí)時(shí)熒光定量PCR(qRT-PCR)技術(shù)分析BSP對M1型巨噬細(xì)胞標(biāo)記基因[白細(xì)胞介素(IL)-1β、IL-6、腫瘤壞死因子-α(TNF-α)、誘導(dǎo)型一氧化氮合酶(iNOS)]和M2型巨噬細(xì)胞標(biāo)記基因(IL-10、CD206)表達(dá)的影響;通過酶聯(lián)免疫吸附實(shí)驗(yàn)檢測BSP對上清液中IL-1β、IL-6和TNF-α釋放的影響;通過Western blotting法檢測BSP對核因子-κB(NF-κB)和信號(hào)轉(zhuǎn)導(dǎo)子和轉(zhuǎn)錄激活子(STAT1)信號(hào)通路相關(guān)蛋白表達(dá)的影響。結(jié)果 BSP在0.001~10.000 μg·mL-1對M0和M1型巨噬細(xì)胞沒有明顯毒性。BSP顯著抑制M1型巨噬細(xì)胞IL-1β、IL-6、TNF-α和iNOS的mRNA表達(dá)水平(P< 0.05、0.01、0.001),并顯著降低IL-1β、IL-6和TNF-α的釋放(P< 0.01、0.001),顯著抑制IL-1β、p65和STAT1的磷酸化蛋白的表達(dá)(P< 0.05)。BSP對M0型巨噬細(xì)胞無顯著影響。結(jié)論 BSP能夠有效抑制M0型巨噬細(xì)胞向M1極化的轉(zhuǎn)變,其機(jī)制主要通過抑制NF-κB和STAT1信號(hào)通路的活化實(shí)現(xiàn)。
[Key word]
[Abstract]
Objective To investigate the effect of Bletilla striata polysaccharide (BSP) on the polarization process of M1 macrophages and its potential mechanisms. Methods THP-1 cells were induced to differentiate into M0 or M1 macrophages and treated with different concentrations (0.001, 0.010, 0.100, 1.000, and 10.000 μg·mL-1) of BSP. The CCK-8 assay was employed to assess cell adhesion and viability following BSP treatment. After selecting an appropriate dose, real-time fluorescence quantitative PCR (qRT-PCR) was used to analyze the expression of M1 macrophage marker genes [Interleukin (IL) -1β, IL-6, tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS)] and M2 macrophage marker genes (IL-10, CD206). The release of inflammatory cytokines IL-1β, IL-6, and TNF-α in the supernatant was measured using an enzyme-linked immunosorbent assay, and Western blotting analysis was performed to investigate the effect of BSP on Nuclear factor kappa B (NF-κB) and signal transducer and activator of transcription 1 (STAT1) signaling pathways. Results BSP exhibited no significant toxicity to M0 and M1 macrophages within the tested concentration range. BSP significantly downregulated the mRNA expression of M1 macrophage markers (IL-1β, IL-6, TNF-α, and iNOS) and reduced the secretion of these inflammatory cytokines (P < 0.05, 0.01, and 0.001). Furthermore, BSP inhibited the protein expression of IL-1β and phosphorylation of p65 and STAT1 proteins. BSP had no significant effect on M0 macrophages. Conclusion BSP effectively suppresses the transformation of M0 macrophages into the M1 phenotype, primarily through the inhibition of NF-κB and STAT1 signaling pathways.
[中圖分類號(hào)]
R285.5
[基金項(xiàng)目]
南京市衛(wèi)生科技發(fā)展專項(xiàng)資金項(xiàng)目(YKK21203)