Håversen Liliana, Ohlsson Bertil G, Hahn-Zoric Mirjana, Hanson Lars A, Mattsby-Baltzer Inger
Department of Clinical Bacteriology, University of Göteborg, Guldhedsgatan 10, S-41346 Göteborg, Sweden.
Cell Immunol. 2002 Dec;220(2):83-95. doi: 10.1016/s0008-8749(03)00006-6.
Lactoferrin, a glycoprotein present in milk, mucosal secretions and neutrophils contributes to host defense. We have previously shown that orally given milk lactoferrin (LF) mediates anti-infectious and anti-inflammatory activities in vivo. Moreover, we have shown that LF could inhibit the LPS-induced IL-6 secretion in a human monocytic cell line, THP-1. This observation was expanded in the present study investigating the capacity of LF to inhibit cytokine mRNA expression and the involvement of nuclear transcription factor kappa B (NF-kappa B). Cells (THP-1 and Mono Mac 6 monocytic cell lines) were stimulated with Escherichia coli LPS (5-10 ng/10(6) cells) and LF was added (50-500 microg/10(6) cells) 30 min before, or after the LPS addition. By a semiquantitative RT-PCR lower levels of TNF-alpha, IL-1 beta, IL-6, and IL-8 mRNA expression were detected at the peak of the expression in THP-1 cells treated with LF. The reduction in the cytokine expression was followed by a similar reduction in the secreted cytokines as analyzed by ELISA. LF down-regulated also the IL-10 secretion (detected only in LPS-stimulated Mono Mac 6 cells). A similar level of inhibition of these cytokines was detected regardless of the time at which LF was added to the cells in relation to LPS. In addition, LF was internalized into cells and detected in the nucleoli as determined by immunostaining and immunofluorescence. Moreover, by electrophoretic mobility shift assay (EMSA) analysis LF decreased the LPS-induced binding of NF-kappa B to the TNF-alpha promoter. The results show that LF down-regulates the LPS-induced cytokine production in monocytic cells. The inhibitory mechanism is suggested to involve the interference of LF with NF-kappa B activation.
乳铁蛋白是一种存在于牛奶、黏膜分泌物和中性粒细胞中的糖蛋白，有助于宿主防御。我们之前已经表明，口服牛奶乳铁蛋白（LF）在体内介导抗感染和抗炎活性。此外，我们还表明，LF可以抑制人单核细胞系THP-1中脂多糖（LPS）诱导的白细胞介素-6（IL-6）分泌。在本研究中，这一观察结果得到了扩展，该研究调查了LF抑制细胞因子mRNA表达的能力以及核转录因子κB（NF-κB）的参与情况。用大肠杆菌LPS（5 - 10 ng/10⁶个细胞）刺激细胞（THP-1和Mono Mac 6单核细胞系），并在添加LPS前30分钟或添加后添加LF（50 - 500 μg/10⁶个细胞）。通过半定量逆转录聚合酶链反应（RT-PCR），在用LF处理的THP-1细胞中，在表达峰值时检测到肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、IL-6和白细胞介素-8（IL-8）mRNA表达水平较低。如通过酶联免疫吸附测定（ELISA）分析所示，细胞因子表达的降低伴随着分泌的细胞因子的类似降低。LF还下调了IL-10的分泌（仅在LPS刺激的Mono Mac 6细胞中检测到）。无论LF相对于LPS添加到细胞中的时间如何，均检测到对这些细胞因子的类似抑制水平。此外，通过免疫染色和免疫荧光测定，LF被内化到细胞中并在核仁中被检测到。此外，通过电泳迁移率变动分析（EMSA），LF降低了LPS诱导的NF-κB与TNF-α启动子的结合。结果表明，LF下调单核细胞中LPS诱导的细胞因子产生。抑制机制被认为涉及LF对NF-κB激活的干扰。
