红景天苷对脂多糖诱导大鼠肺泡巨噬细胞和Ⅱ型肺泡上皮细胞共培养炎性介质分泌的影响

Effects of salidroside on the secretion of inflammatory mediators induced by lipopolysaccharide in the co-culture of rat alveolar macrophages and type Ⅱ alveolar epithelial cells

本研究旨在探讨红景天苷(salidroside, Sal)对脂多糖(lipopolysaccharide, LPS)诱导大鼠肺泡巨噬细胞NR 8383和II型肺泡上皮细胞RLE-6TN共培养炎性活化的影响。CCK-8比色法检测细胞增殖百分率,Western blot检测磷酸化AKT (p-AKT)和总AKT蛋白表达,酶联免疫吸附法测定细胞培养上清中肿瘤坏死因子α(tumor necrosis factorα, TNF-α)、巨噬细胞炎性蛋白2(macrophage inflammatory protein-2, MIP-2)和白介素10 (interleukin-10, IL-10)的含量。结果显示:与对照组相比,32和128μg/mL Sal预处理RLE-6TN细胞或共培养RLE-6TN和NR 8383细胞1 h后继续培养24 h,细胞增殖百分率显著增加(P <0.05);与对照组相比,32和128μg/mL Sal预处理RLE-6TN细胞,p-AKT/AKT蛋白比值显著增加(P <0.05)。32μg/mL Sal预处理不仅抑制LPS诱导NR 8383细胞分泌TNF-α和MIP-2 (P <0.05),而且加强RLE-6TN和NR 8383细胞共培养对LPS诱导NR 8383细胞分泌TNF-α和MIP-2的抑制作用(P <0.05)。此外,32μg/mL Sal预处理能促进LPS诱导NR 8383细胞分泌IL-10 (P <0.05),并能加强RLE-6TN和NR 8383细胞共培养对LPS诱导NR 8383细胞分泌IL-10的促进作用(P <0.05)。以上结果提示,Sal不仅能直接抑制LPS诱导的NR 8383炎性活化,还可能通过PI3K/AKT信号通路促进RLE-6TN增殖,参与II型肺泡上皮细胞对LPS诱导肺泡巨噬细胞炎性活化的调节作用。

The aim of the present study was to investigate the effect of salidroside(Sal)on inflammatory activation induced by lipopolysaccharide(LPS)in the co-culture of rat alveolar macrophages(AM)NR 8383 and type II alveolar epithelial cells(AEC II)RLE-6 TN.CCK-8 colorimetric method was used to detect cell proliferation percentage.The enzyme-linked immunosorbent assay(ELISA)was used to determine the content of tumor necrosis factor alpha(TNF-α),macrophage inflammatory protein-2(MIP-2)and interleukin-10(IL-10)in the supernatant.Western blot was used to examine the expression levels of phosphorylated AKT(p-AKT)and total AKT protein.The results showed that pretreatment of RLE-6 TN cells or co-culture of RLE-6 TN and NR 8383 cells with 32 and 128μg/mL Sal for 1 h,followed by continuous culture for 24 h,significantly increased the cell proliferation(P<0.05).Compared with control group,32 and 128μg/mL Sal pretreatment significantly increased the ratio of p-AKT/AKT in RLE-6 TN cells(P<0.05).Pretreat-ment of 32μg/mL Sal not only inhibited the secretion of TNF-αand MIP-2 by NR 8383 cells induced by LPS(P<0.05),but also enhanced the inhibitory effect of RLE-6 TN and NR 8383 cells co-culture on the secretion of TNF-αand MIP-2 by NR 8383 cells induced by LPS(P<0.05).In addition,32μg/mL Sal pretreatment promoted LPS-induced IL-10 secretion by NR 8383 cells(P<0.05),and enhanced the promoting effect of co-culture of RLE-6 TN and NR 8383 cells on the IL-10 secretion by LPS-induced NR 8383 cells(P<0.05).In conclusion,Sal may directly inhibit LPS-induced inflammatory activation of AM(NR 8383),promote the proliferation of AEC II(RLE-6 TN)through PI3 K/AKT signaling pathway,and enhance the regulatory effect of AEC II on LPS-induced inflammatory activation of AM.