[萝卜硫素对慢性阻塞性肺疾病患者单核细胞来源巨噬细胞Toll样受体4/髓样分化因子88通路的影响]

[Effects of sulforaphane on Toll-like receptor 4/myeloid differentiation factor 88 pathway of monocyte-derived macrophages from patients with chronic obstructive pulmonary disease].

作者信息

Zeng Xiaoli, Liu Xiaoju, Bao Hairong, Zhang Yi, Wang Xiaohu, Shi Kai, Pang Qi

机构信息

Department of Gerontal Respiratory Medicine, the First Hospital of Lanzhou University, Lanzhou 730000, China.

Department of Gerontal Respiratory Medicine, the First Hospital of Lanzhou University, Lanzhou 730000, China. Email:

出版信息

Zhonghua Jie He He Hu Xi Za Zhi. 2014 Apr;37(4):250-4.

PMID:24969711

Abstract

OBJECTIVE

To explore the effects of sulforaphane on Toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88) pathway and its downstream inflammatory cytokines in patients with chronic obstructive pulmonary disease (COPD).

METHODS

From Jan. 2012 to Mar. 2013, thirty-two stable COPD patients and thirty healthy donors (non-COPD group) from the First Hospital of Lanzhou University were recruited. The peripheral blood monocytes were isolated and induced to macrophages (monocyte-derived macrophages, MDMs). The MDMs of COPD patients were divided into a blank control group, a LPS group, a sulforaphane group, a sulforaphane and LPS group (combined group), while the MDMs from the non-COPD group received no drug intervention. The number of cells in each group was 3×10(6). The mRNA and protein expression of TLR4 and MyD88 were measured with real-time PCR and Western blot. The TNF-α and IL-6 levels in the culture supernatant were measured with ELISA. Oneway ANOVA and LSD-t test were used for statistical analysis.

RESULTS

The levels of mRNA and protein of TLR4 and MyD88 and the contents of TNF-α and IL-6 in the culture supernatant were higher in the blank control group [3.7 ± 0.5, 1.9 ± 0.4, 0.45 ± 0.18, 1.11 ± 0.65, (31 ± 4) and (43 ± 5) µg/L] than those in the non-COPD group [1.00, 1.00, 0.26 ± 0.14, 0.58 ± 0.40, (19 ± 2) and (29 ± 4) µg/L] (t = 2.19-12.11, P < 0.05 or P < 0.01). After LPS treatment (LPS group), the above parameters [5.5 ± 1.1, 3.4 ± 1.6, 0.65 ± 0.20, 1.66 ± 0.64, (47 ± 4) and (54 ± 5) µg/L] were increased as compared to those in the blank control group (t = 2.39-11.9, P < 0.05 or P < 0.01), but after sulforaphane treatment(Sulforaphane group), these parameters [2.2 ± 0.4, 1.0 ± 0.6, 0.25 ± 0.09, 0.62 ± 0.34, (20 ± 3) and (27 ± 4) µg/L] were decreased as compared to those in the blank control group (t = 2.13-8.46, P < 0.05 or P < 0.01). Similarly, these parameters in the combined group [3.2 ± 0.5, 1.5 ± 0.8, 0.33 ± 0.11, 0.77 ± 0.25, (31 ± 3) and (33 ± 4) µg/L] were also remarkably decreased as compared to those in the LPS group (t = 3.87-12.24, all P < 0.01).

CONCLUSIONS

The TLR4/MyD88 pathway was activated and its downstream inflammatory cytokines were increased in macrophages from COPD patients. Sulforaphane could inhibit the TLR4/MyD88 pathway and reduce the releasing of downstream inflammatory cytokines, suggesting that sulforaphane may have an anti-inflammatory effect in COPD.

摘要

目的

探讨萝卜硫素对慢性阻塞性肺疾病（COPD）患者Toll样受体4（TLR4）/髓样分化因子88（MyD88）信号通路及其下游炎症细胞因子的影响。

方法

选取2012年1月至2013年3月在兰州大学第一医院就诊的32例稳定期COPD患者和30名健康对照者（非COPD组）。分离外周血单核细胞细胞并诱导为巨噬细胞（单核细胞来源的巨噬细胞，MDMs）。COPD患者的MDMs分为空白对照组、脂多糖（LPS）组、萝卜硫素组、萝卜硫素与LPS联合组（联合组），非COPD组的MDMs不进行药物干预。每组细胞数量均为3×10⁶。采用实时荧光定量PCR和蛋白质印迹法检测TLR4和MyD88的mRNA及蛋白表达。采用酶联免疫吸附测定法检测培养上清液中肿瘤坏死因子-α（TNF-α）和白细胞介素-6（IL-6）水平。采用单因素方差分析和LSD-t检验进行统计学分析。

结果

空白对照组[3.7±0.5、1.9±0.4、0.45±0.18、1.11±0.65、（31±4）和（43±5）μg/L]TLR4和MyD88的mRNA及蛋白水平、培养上清液中TNF-α和IL-6含量高于非COPD组[1.00、1.00、0.26±0.14、0.58±0.40、（19±2）和（29±4）μg/L]（t=2.19～12.11，P<0.05或P<0.01）。LPS处理后（LPS组），上述指标[5.5±1.1、3.4±1.6、0.65±0.20、1.66±0.64、（47±4）和（54±5）μg/L]较空白对照组升高（t=2.39～11.9，P<0.05或P<0.01），而萝卜硫素处理后（萝卜硫素组），上述指标[2.2±0.4、1.0±0.6、0.25±0.09、0.62±0.34、（20±3）和（27±4）μg/L]较空白对照组降低（t=2.13～8.46，P<0.05或P<0.01）。同样，联合组上述指标[3.2±0.5、1.5±0.8、0.33±0.11、0.77±0.25、（31±3）和（33±4）μg/L]较LPS组也显著降低（t=3.87～12.24，均P<0.01）。