The experiments were performed at least 3 times, and representative results are shown. oxidase activation, and p38 mitogen-activated protein kinase (MAPK) phosphorylation, while those specific inhibitors inhibited the HO-1 expression induced by fungal -GPs. Moreover, fungal -GPs induced Nrf2 translocation into nuclei via p38 MAPK signaling, while the HO-1 expression induced by fungal -GPs was inhibited by Nrf2-specific small interfering RNA (siRNA). Finally, knockdown of cells by HO-1- and Nrf2-specific siRNAs resulted in increased -GP-mediated ROS production compared to that in the control cells. Our results show that this HO-1 induced by fungal -GPs via ROS/p38 MAPK/Nrf2 from oral keratinocytes may have important functions in host defense against the stress caused by contamination in the oral epithelium. species, most commonly, (1, 2). Following adherence to oral mucosa, penetrates the epithelial surface at microscopic wound sites (3) and invades the oral epithelium (4). Oral keratinocytes provide the first line of host defense against contamination (5) and actively respond to live organisms by generating inflammatory mediators (6, 7). In an model, heat-killed did not enhance immune responses in the oral epithelium, whereas the contact of live organisms with the epithelium was shown to increase the expression of proinflammatory cytokines, such as interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-) (6). In contrast, heat-killed and cell wall fractions have been reported to increase the expression of inflammatory mediators, such as IL-8 and granulocyte-macrophage colony-stimulating factor, in oral keratinocytes (8). Therefore, interactions of fungal cell wall components with oral keratinocytes may regulate the stress response against contamination. and the budding yeast share similarities in regard to their cell wall structures, in both of which the cell walls are composed of an inner layer of -glucan covalently linked to a variety of cell surface mannoproteins (9,C11). -Glucan has been Syringin shown to induce phagocytosis, cytotoxic activities, and proinflammatory cytokine production in mouse macrophages (12). Furthermore, -glucan has been observed on the surface of biofilms created by in mice with oropharyngeal candidiasis showing invasion of the tongue mucosa (13). However, it is unknown whether fungal cell wall components, such as -glucan, participate in the activation of stress-mediated immune responses by oral keratinocytes. Heme oxygenase 1 (HO-1) is an enzyme that catalyzes the first rate-limiting step in the degradation of free heme to produce carbon monoxide, ferrous iron, and biliverdin (BV) (14). Furthermore, HO-1 is also thought to be a stress-inducible enzyme that mediates antioxidative and cytoprotective effects to maintain cellular redox homeostasis and provide protection against oxidative stress (14). This enzyme is usually induced by an oxidative stressor, such as hydrogen Ik3-1 antibody peroxide, and its inhibition increases hydrogen peroxide-induced oxidative damage (15,C17). On the other hand, following its induction by some bacterial components, HO-1 enhances host defense and oxidative signaling in response to bacterial infection. The Gram-negative bacterial outer membrane component lipopolysaccharide (LPS) has been shown to increase HO-1 expression in immune cells, such as macrophages and monocytes (18, 19), while HO-1 was also shown to be increased by the Gram-positive bacterial cell wall component lipoteichoic acid (LTA) in human tracheal smooth muscle mass cells (20). Even though inducer and signaling events involved in HO-1 expression in oral keratinocytes have not been completely elucidated, the HO-1 induced by microbial components in oral keratinocytes Syringin may play a role in protective intercellular stress against oral microorganism contamination. We speculated that cell wall components of participate in mediation of the stress responses against contamination in the oral epithelium. Therefore, we investigated the expression profiles of genes induced by heat-killed in oral immortalized (RT7) keratinocytes using a cDNA microarray technique and focused on the HO-1 expression induced by and the fungal cell wall component involved in its increase. Furthermore, we examined the mechanisms of the intercellular signaling pathway and antioxidative stress functions involved in induction of HO-1 expression by -glucan-containing particles (-GPs), the fungal cell wall components. RESULTS Syringin Differences in gene expression between heat-killed in comparison with their level of expression by nontreated cells. Among the 24,000 genes detected by the cDNA microarray, 33 genes were upregulated greater than 8-fold in heat-killed using quantitative reverse transcription (RT-PCR) analysis (Fig. 2). Of the 9 upregulated genes, the expression of 7 was increased by both live and heat-killed were upregulated in cells exposed to live and live organisms. Furthermore, HO-1 expression was rather.