Omagari D, Mikami Y, Suguro H, Sunagawa K, Asano M, Sanuki E, Moro I, Komiyama K
Department of Pathology, Nihon University School of Dentistry, Tokyo, Japan.
Clin Exp Immunol. 2009 May;156(2):294-302. doi: 10.1111/j.1365-2249.2009.03892.x. Epub 2009 Mar 4.
Intercellular adhesion molecul-1 (ICAM-1) is a transmembrane glycoprotein belonging to the immunoglobulin superfamily of adhesion molecules and plays perdominant roles in recruitment and trafficking of leucocytes to sites of inflammation. ICAM-1 expression in intestinal epithelial cells (IECs) is enhanced by several stimuli, such as proinflammatory cytokines, bacterial infections or pathogen-associated molecular patterns. One of these stimuli, double-stranded RNA (dsRNA), is a by-product of viral replication and can be recognized by its cognate receptor Toll-like receptor 3 (TLR-3). In spite of expression of both TLR-3 and ICAM-1 in IECs, correlation between TLR-3-signalling and ICAM-1 expression has never been examined in IECs. In the present study, we investigated whether poly I:C, an analogue of dsRNA, can stimulate the expression of ICAM-1 in IEC line, HT-29. Poly I:C-stimulation up-regulated the expression of ICAM-1 mRNA by real-time polymerase chain reaction. Enhanced expression of ICAM-1 was confirmed in protein level by immunofluoresense cell staining and enzyme-linked immunosorbent assay by measuring the released soluble ICAM-1 in culture supernatant. As the stimulation effect was reduced by pre-treatment of the cells with anti-TLR-3 antibody, poly I:C-binding signal was thought to be sensed by TLR-3 on the surface of HT-29. The results of luciferase assay and nuclear factor kappa-b (NF-kappaB) inhibitor treatment experiments indicated that the downstream signal was mainly transduced by transcription factor, NF-kappaB. All these results demonstrated the connection between TLR-3 signalling and ICAM-1 expression in HT-29 cells and indicated the importance of coordinated function of both innate and adaptive immunity against viral infections.
细胞间黏附分子-1(ICAM-1)是一种跨膜糖蛋白,属于黏附分子免疫球蛋白超家族,在白细胞募集和向炎症部位运输过程中起主要作用。肠道上皮细胞(IECs)中ICAM-1的表达可被多种刺激增强,如促炎细胞因子、细菌感染或病原体相关分子模式。这些刺激之一,双链RNA(dsRNA),是病毒复制的副产物,可被其同源受体Toll样受体3(TLR-3)识别。尽管IECs中同时表达TLR-3和ICAM-1,但TLR-3信号与ICAM-1表达之间的相关性从未在IECs中进行过研究。在本研究中,我们调查了dsRNA类似物聚肌胞苷酸(poly I:C)是否能刺激IEC细胞系HT-29中ICAM-1的表达。通过实时聚合酶链反应,poly I:C刺激上调了ICAM-1 mRNA的表达。通过免疫荧光细胞染色和酶联免疫吸附测定法,通过测量培养上清液中释放的可溶性ICAM-1,在蛋白质水平上证实了ICAM-1的表达增强。由于用抗TLR-3抗体预处理细胞可降低刺激效果,因此认为HT-29表面的TLR-3可感知poly I:C结合信号。荧光素酶测定和核因子κB(NF-κB)抑制剂处理实验结果表明,下游信号主要由转录因子NF-κB转导。所有这些结果都证明了HT-29细胞中TLR-3信号与ICAM-1表达之间的联系,并表明先天免疫和适应性免疫协同对抗病毒感染的重要性。