基于BV2小胶质细胞极化探索片仔癀抗炎机制

目的：从小鼠小胶质瘤细胞（BV2）极化角度探索片仔癀（PTH）的抗炎作用机制。

方法：将对数生长期的BV2细胞分为空白组，M1模型组[脂多糖（LPS）100 μg·L⁻¹ + 干扰素-γ（IFN-γ）10 μg·L⁻¹]，M1片仔癀组[LPS 100 μg·L⁻¹ + IFN-γ 10 μg·L⁻¹ + PTH 0.4 g·kg⁻¹]，M2模型组[白细胞介素-4（IL-4）20 μg·L⁻¹]，M2片仔癀组[IL-4 20 μg·L⁻¹ + PTH 0.4 g·kg⁻¹]，给以相应处理。通过酶联免疫吸附测定（ELISA）检测细胞上清中肿瘤坏死因子-α（TNF-α）、一氧化氮（NO）、白细胞介素-10（IL-10）、转化生长因子-β1（TGF-β1）的含量，使用实时荧光定量聚合酶链式反应（Real-time PCR）检测细胞中一氧化氮诱导合酶（iNOS）、精氨酸-1（Arg-1）mRNA的水平，蛋白免疫印迹法（Western blot）检测细胞中 iNOS、p-STAT1、p-STAT3、Arg-1、p-STAT6 蛋白表达情况。

结果：与空白组比较，M1模型组上清 TNF-α、NO 含量显著升高（P＜0.01），细胞 iNOS mRNA、iNOS、p-STAT1、p-STAT3 蛋白水平显著升高（P＜0.01），而与 M1 模型组比较，M1片仔癀组上清 TNF-α 及 NO 的含量显著下调（P＜0.01），细胞 iNOS mRNA 显著下调（P＜0.01），iNOS、p-STAT1、p-STAT3 蛋白水平也明显下调（P＜0.05，P＜0.01）。与空白组比较，M2 模型组细胞培养上清中 IL-10 及 TGF-β1 含量显著升高（P＜0.01），细胞 Arg-1 mRNA、Arg-1、p-STAT6 蛋白水平显著升高（P＜0.01）。与 M2 模型组比较，M2片仔癀组上清中 IL-10 及 TGF-β1 含量明显上调（P＜0.05，P＜0.01），细胞 Arg-1 mRNA 水平、Arg-1 及 p-STAT6 蛋白水平明显上调（P＜0.05，P＜0.01）。

结论：片仔癀可以通过调节 BV2 的极化发挥抗炎作用。

Objective: To investigate the anti-inflammation mechanism of Pien Tze Huang (PTH) via regulating microglia polarization.

Method: The experiment was divided into five groups, Blank, M1[lipopolysaccharide (LPS) 100μg·L-1+interferon-γ (IFN-γ) 10μg·L^-1], M1-PTH group[LPS 100μg·L^-1+IFN-γ10μg·L^-1+PTH 0. 4 g·kg^-1], M2 group[interleukin-4 (IL-4) 20μg·L^-1], and M2-PTH group[IL-420μg·L^-1+PTH 0. 4 g·kg^-1]. The concentration of nitric oxide (NO) , tumor necrosis factor-α (TNF-α) , interleukin-10 (IL-10) , and transforming growth factor-β1 (TGF-β1) in the culture supernatant were measured by enzyme-linked immunosorbent assay (ELISA) , the levels of inducible nitric oxide synthase (i NOS) and arginine-1 (Arg-1) mRNA were detected by real-time fluorescent quantitative polymerase chain reaction technique (Realtime PCR) , and the expression levels of p-STAT1, p-STAT3, i NOS, p-STAT6, and Arg-1 were detected by Western blot.

Result: The concentration of NO and TNF-αof the culture supernatant, the level of i NOS mRNA, as well as the level of p-STAT1, p-STAT3 and i NOS in M1 group, which were significantly increased (P＜0. 01) . Compared with blank group, but the concentration of NO and TNF-αwere down-regulated (P＜0. 01) , and i NOS mRNA (P＜0. 05) , as well as the expression of i NOS, p-STAT1, and p-STAT3 was decreased (P＜0. 05, P＜0. 01) after the invention of PTH in M1-PTH group compared with M1 group. The concentration of IL-10 and TGF-β1in the culture supernatant, the mRNA level of Arg-1, as well as the levels of p-STAT6 and Arg-1 were significantly increased in M2 group when compared with Blank group, addition to the concentration of IL-10 and TGF-β1were up-regulated (P＜0. 05, P＜0. 01) , and the expression of Arg-1 mRNA, the level of Arg-1, pSTAT6 were enhanced (P＜0. 05, P＜0. 01) in M2-PTH group compared with M2 group.

Conclusion: PTH plays an anti-inflammatory role via regulating microglia polarization.