人脐带间充质干细胞对大鼠肺泡巨噬细胞表达细胞因子的影响

Effects of the Human Umbilical Cord Mesenchymal Stem Cells Pretreatment on the Expression of Inflammatory Cytokines in Macrophages

目的研究人脐带间充质干细胞(hUCMSCs)对受脂多糖(LPS)刺激的大鼠肺泡巨噬细胞NR8383表达炎性细胞因子的影响。方法从人脐带中分离hUCMSCs细胞,与NR8383在Transwell内间接共培养,hUCMSCs接种于上室,下室LPS浓度为100μg·mL^(-1)刺激NR8383释放炎性因子。设对照组(NR8383只加培养液)、LPS组(NR8383+LPS)、hUCMSCs处理组(NR8383+LPS+hUCMSCs)、hUCMSCs预处理组(NR8383+hUCMSCs+LPS)。实时荧光定量(qRT-PCR)测定NR8383细胞中肿瘤坏死因子α(TNF-α)、白细胞介素(IL)-1β、IL-6以及IL-10的mRNA表达;ELISA法检测各组NR8383细胞培养液中四种细胞因子的动态变化。结果与对照组相比,LPS组的促炎因子TNF-α、IL-1β、IL-6的mRNA表达及蛋白水平均升高(P均<0.05);细胞培养液中,hUCMSCs处理组的IL-10含量均较LPS组升高,TNF-α、IL-6含量均降低(P<0.05);与hUCMSCs处理组比较,hUCMSCs预处理组细胞培养液中TNF-α、IL-1β蛋白表达下降(P均<0.05),抗炎因子IL-10的蛋白表达升高(P<0.05)。结论hUCMSCs可抑制LPS诱导巨噬细胞产生的过度炎性反应,且hUCMSCs预处理后巨噬细胞的抗炎作用更明显。

Objective To investigate the immunomodulatory effects of the human umbilical cord mesenchymal stem cells(hUCMSCs)on rat alveolar macrophage NR8383 which were stimulated by lipopolysaccharide(LPS).Methods hUCMSCs cells were isolated from human umbilical cord and NR8383 were co-cultured with Transwell indirectly.hUCMSCs were inoculated in the upper chamber,and LPS concentration was 100μg·mL^(-1) in the lower chamber of Transwell,stimulate NR8383 to release inflammatory factors.The control group(NR8383 with culture medium only),LPS group(NR8383+LPS),hUCMSCs treatment group(NR8383+LPS+hUCMSCs),and hUCMSCs pretreatment group(NR8383+hUCMSCs+LPS)were set up.The mRNA expression levels of tumor necrosis factors(TNF-α),interleukin(IL)-1β,IL-6 and IL-10 of NR8383 cells were determined by real-time fluorescence quantification(qRT-PCR).The dynamic changes of the four factors were detected by enzyme-linked immuno sorbent assay(ELISA).Results Compared with the control group,the expression levels of pro-inflammatory factors TNF-α,IL-1βand IL-6 in LPS group were increased(P all<0.05).Compared with LPS group,the expression level of IL-10 in hUCMSCs treated group effectively increased,and the expression levels of TNF-αand IL-6 were decreased,and the expression level of IL-10 in hUCMSCs pretreated group was significantly increased(P all<0.05).Compared with the hUCMSCs treated group,the expression level of TNF-αand IL-1βin the hUCMSCs pretreated group were decreased(P all<0.05),while the anti-inflammatory factor IL-10 was increased in the hUCMSCs pretreated group(P<0.05).Conclusion hUCMSCs can inhibit the excessive inflammatory response of macrophages induced by LPS,and the anti-inflammatory effect of hUCMSCs pretreated macrophages is more obvious.