The neural ELAVL protein HuB enhances endogenous proto-oncogene activation.

The cytoplasmic distribution of the HuR/ELAVL1 (embryonic lethal abnormal vision 1) protein is recognized as an important prognostic factor of malignant tumors. However, the previous study suggests that exogenous over-expression of HuR is not sufficient for nuclear export. Conversely, the predominantly cytosolic distribution of neuron-specific human ELAV members, including HuB/ELAVL2, HuC/ELAVL3, and HuD/ELAVL4, has been reported. In the present study, we demonstrated the expression of HuB in several types of cancer cells, but expression of HuC and HuD was not observed. In addition, our results indicated that HuR and HuB formed a complex in the cytosolic fraction of cancer cells via the RRM3 region. Ectopic expression of HuB was capable of initiating the cytosolic translocation of HuR from the nucleus to the cytosol. Furthermore, HuB-transduced cancer cells displayed significant nuclear export of HuR, with quantitative PCR experiments revealing the simultaneous upregulation of HIF-1α, c-Fos, c-MYC, and Ets2 basal mRNA expression. Phorbol 12-myristate 13-acetate (PMA)-stimulated HuB-transduced cells demonstrated significantly enhanced activation of endogenous c-Fos and CREB dependent cascades. Finally, co-transfection of HuB with the E1 region of type 5 human adenovirus significantly enhanced E1 transformation activities but that of HuR with the E1 region did not. Collectively, our findings suggest that the neural Hu family protein HuB plays a major role in the activation of memory-related proto-oncogenes.

Adenovirus infection induces HuR relocalization to facilitate virus replication.

HuR is an RNA-binding protein of the embryonic lethal abnormal vision (ELAV) family, which binds to the AU-rich element (ARE) in the 3'-untranslated region (UTR) of certain mRNAs and is involved in the nucleo-cytoplasmic export and stabilization of ARE-mRNAs. The cytoplasmic relocalization of ARE-mRNAs with several proteins such as HuR and pp32 increases in cells transformed by the adenovirus oncogene product E4orf6. Additionally, these ARE-mRNAs were stabilized and acquired the potential to transform cells. Although, the relocalization of HuR and the stabilization of ARE-mRNAs are crucial for cell transformation, evidence regarding the relationship of HuR and ARE-mRNAs with adenovirus replication is lacking. In this report, we demonstrate that adenovirus infection induces the relocation of HuR to the cytoplasm of host cells. Analysis using the luciferase-ARE fusion gene and the tetracycline (tet)-off system revealed that the process of stabilizing ARE-mRNAs is activated in adenovirus-infected cells. Heat shock treatment or knockdown-mediated depletion of HuR reduced adenovirus production. Furthermore, expression of ARE-including viral IVa2 mRNA, decreased in HuR-depleted infected cells. These results indicate that HuR plays an important role in adenovirus replication, at least in part, by up-regulating IVa2 mRNA expression and that ARE-mRNA stabilization is required for both transformation and virus replication.

Role of ATF5 in the invasive potential of diverse human cancer cell lines.

Activating transcription factor 5 (ATF5) is a member of the ATF/cAMP response element-binding protein family. Our research group recently revealed that ATF5 expression increases the invasiveness of human lung carcinoma cells. However, the effects of ATF5 on the invasive potential of other cancer cells lines remain unclear. Therefore, in this study, we investigated the role of ATF5 in the invasive activity of diverse human cancer cell lines. Invasiveness was assessed using Matrigel invasion assays. ATF5 knockdown resulted in decreased invasiveness in seven of eight cancer cell lines tested. These results suggest that ATF5 promotes invasiveness in several cancer cell lines. Furthermore, the roles of ATF5 in the invasiveness were evaluated in three-dimensional (3D) culture conditions. In 3D collagen gel, HT-1080 and MDA-MB-231 cells exhibited high invasiveness, with spindle morphology and high invasion speed. In both cell lines, knockdown of ATF5 resulted in rounded morphology and decreased invasion speed. Next, we showed that ATF5 induced integrin-α2 and integrin-ß1 expression and that the depletion of integrin-α2 or integrin-ß1 resulted in round morphology and decreased invasion speed. Our results suggest that ATF5 promotes invasion by inducing the expression of integrin-α2 and integrin-ß1 in several human cancer cell lines.

Cell type-specific reciprocal regulation of HIF1A gene expression is dependent on 5'- and 3'-UTRs.

In the present study, we demonstrated the reciprocal regulation of hypoxia-inducible factor 1 alpha (HIF1A) gene expression via untranslated region-(UTR) dependent mechanisms. A 151 nucleotide sequence found in the HIF1A 5'-UTR is sufficient for significant translational up-regulation. On the other hand, the 3'-UTR of HIF1A has been implicated in mRNA degradation. In the non-metastatic breast cancer cell line MCF7, the 3'-UTR-dependent down-regulatory machinery predominates over the 5'-UTR-dependent up-regulation of HIF1A. However, 5'-UTR-dependent up-regulation is dominant among metastatic cell lines (MDA-MB453, U87MG). It is therefore likely that the predominance of 5'-UTR-dependent translational enhancement of HIF1A is critical for the malignant phenotype of cancer cells. PTBP-1, but not HuR, is a candidate RNA binding protein for the translational control of HIF1A.

Substrate stiffness regulates temporary NF-κB activation via actomyosin contractions.

Physical properties of the extracellular matrix (ECM) can control cellular phenotypes via mechanotransduction, which is the process of translation of mechanical stresses into biochemical signals. While current research is clarifying the relationship between mechanotransduction and cytoskeleton or adhesion complexes, the contribution of transcription factors to mechanotransduction is not well understood. The results of this study revealed that the transcription factor NF-κB, a major regulator for immunoreaction and cancer progression, is responsive to substrate stiffness. NF-κB activation was temporarily induced in H1299 lung adenocarcinoma cells grown on a stiff substrate but not in cells grown on a soft substrate. Although the activation of NF-κB was independent of the activity of integrin ß1, an ECM-binding protein, the activation was dependent on actomyosin contractions induced by phosphorylation of myosin regulatory light chain (MRLC). Additionally, the inhibition of MRLC phosphorylation by Rho kinase inhibitor Y27632 reduced the activity of NF-κB. We also observed substrate-specific morphology of the cells, with cells grown on the soft substrate appearing more rounded and cells grown on the stiff substrate appearing more spread out. Inhibiting NF-κB activation caused a reversal of these morphologies on both substrates. These results suggest that substrate stiffness regulates NF-κB activity via actomyosin contractions, resulting in morphological changes.

Human adenovirus oncolytic properties and the inhibitory role of E4 orf4 and E4 orf6/7 on endogenously activated NF-κB.

Human adenovirus is a promising tool for cancer therapy as an oncolytic virus. To predict which region of the oncolytic adenovirus E4 gene could be deleted, we investigated the relationship between the E4 proteins and NF-κB. Here, we report that TLR2-dependent NF-κB activation in Ad5-infected cells was significantly inhibited 24 h post-infection. Among the six E4 proteins, E4 orf4 and E4 orf6/7 exhibited notable suppressive effects on NF-κB activation. However, only E4 orf4 was co-immunoprecipitated with the RelA protein, also known as p65. It appears likely that E4 orf6/7 represses NF-κB activation via E2F-dependent pathways. Our results suggest that both E4 orf4 and E4 orf6/7 are novel inhibitors of NF-κB activation. The inhibition of endogenous NF-κB activation by E4 proteins during the late phase of infection also appears to elucidate the previously reported suppression of E1A expression in the late phase of infection. These redundant suppressive effects of E4 orf4 and E4 orf6/7 on NF-κB suggest that these proteins may play a major role in the anticancer properties of oncolytic adenovirus.

Adenovirus infection controls processing bodies to stabilize AU-rich element-containing mRNA.

In the adenovirus-infected cells, virus mRNAs are selectively exported to the cytoplasm by virus early gene products to facilitate virus replication. We previously showed AU-rich elements (AREs) containing mRNAs are exported to the cytoplasm and stabilized in infected cells. Here, we analyzed ribonucleoprotein (RNP) granules in the cytoplasm that are involved in mRNA degradation to elucidate the mechanism of ARE-mRNA stabilization in adenovirus infected cells. Our findings showed that processing bodies (PBs) aggregate, then almost all PBs are translocated to aggresomes formed by adenoviral gene products during the late phase of infection. Furthermore, E4orf3 was required for the PBs translocation, and the same domains of E4orf3-mutants required to change the form of promyelocytic leukemia bodies were also needed for PBs translocation. Luciferase activity showed that these domains were critical for miRNA- and ARE-mediated mRNA decay. These findings suggest that adenovirus changes the behavior of PBs to prevent ARE-mRNA downregulation.

Mammaglobin 1 mediates progression of trastuzumab-resistant breast cancer cells through regulation of cyclins and NF-κB.

Overexpression of human epidermal growth factor receptor 2 (HER2) in various cancers is correlated with poor patient survival. Trastuzumab, a recombinant humanized monoclonal antibody against HER2, has been considered to be a first-line therapy for HER2-positive breast cancer patients, but its usefulness is limited by the development of resistance. In this study, we established resistant cells by long-term treatment with trastuzumab. These cells showed higher proliferation, invasion, and migration abilities than the wild-type cells. Mammaglobin 1 (MGB1), cyclin D1, E1, A2, and phosphorylated NF-κB (p-p65) were upregulated in resistant cells. These proteins regulate cell proliferation, migration, and invasion of resistant cells. Depletion of MGB1 decreased cyclin and p-p65 expression. Cyclin D1 and A2, but not E1 expression, were affected by p-p65 downregulation. In summary, our results indicate that MGB1 expression is increased in breast cancer cells that have gained resistance to trastuzumab, and suggest that MGB1 promotes aggressiveness through cyclin and NF-κB regulation.

Radiation-induced cancer cell repopulation: a possible mechanism implied by experiments using transplantable mouse-derived sarcoma cell line.

PURPOSE: Treatment with any cytotoxic agent can trigger surviving cells in a tumor to divide faster than before. This phenomenon is widely recognized as "repopulation". To better clarify the mechanism, gene expression profiling and pathological experiments were performed. MATERIALS AND METHODS: A mouse fibrosarcoma cell line, QRsP, was used. Cells were irradiated with 10 Gy. Colony assay and cloning were performed. Six clones were established. cDNA analysis was performed on the clone that showed the largest number of colonies on the 2nd 10 Gy irradiation. Mouse transplantation experiment was then carried out. RESULTS: cDNA analysis showed that cyclin-dependent kinase inhibitors, p16 and p57 were down-regulated; 14.8- and 12.0-fold, respectively for the tolerant clone. Matrix metalloproteinase 3 and 13 were up-regulated; 22.5- and 25.8-fold, respectively. Transplantation ratio was 100% (5/5) for the tolerant clone whereas it was 40% (2/5) for the parent. Under light microscope, the mean mitotic cell number was 4.0+/-3.9 for the parent, and 12.8+/-3.4 for the tolerant clone (p<0.01, Student's t-test). CONCLUSIONS: This study implies that repopulation is not a temporary reaction to irradiation. It is caused probably by "clonal" gene-expression changes, though it remains unknown whether the changes are attributable to tolerant cell selection or to gene mutation/modification.

High prevalence of multiple human papillomavirus infection in Japanese patients with invasive uterine cervical cancer.

OBJECTIVE: Multiple human papillomavirus (HPV) infection of the uterine cervix has been suggested as a risk factor for persistent HPV infection, resulting in the development of invasive cervical cancer. The aim of this study was to reveal the actual state of multiple HPV infection in Japanese patients with invasive cervical cancer. METHODS: Sixty fresh-frozen invasive cervical cancer tissues were examined for genotyping of HPV. The presence of HPV genotypes was determined with an HPV-DNA array, which can discriminate 25 different HPV genotypes with high sensitivity and specificity. RESULTS: Among 60 samples, 59 (96.7%) were positive for HPV. The three common genotypes were HPV-16 (83.3%), HPV-18 (45.0%) and HPV-52 (28.3%). Multiple HPV infection was observed in 47 of 60 samples (78.3%), among which 42 were infected with more than one high-risk genotype (70.0%). Multiple high-risk HPV infection was significantly more prevalent in patients below 40 years old (14/15, 93.3%) than in patients 40 years of age and over (28/45, 62.2%). CONCLUSION: The HPV-DNA array is the preferred method to detect HPV genotypes. Multiple HPV infection in Japanese patients with invasive cervical cancer seemed to be more frequent than reported in the literature.

Integrin beta1-dependent invasive migration of irradiation-tolerant human lung adenocarcinoma cells in 3D collagen matrix.

Radiotherapy is one of the effective therapies used for treating various malignant tumors. However, the emergence of tolerant cells after irradiation remains problematic due to their high metastatic ability, sometimes indicative of poor prognosis. In this study, we showed that subcloned human lung adenocarcinoma cells (A549P-3) that are irradiation-tolerant indicate high invasive activity in vitro, and exhibit an integrin beta1 activity-dependent migratory pattern. In collagen gel overlay assay, majority of the A549P-3 cells displayed round morphology and low migration activity, whereas a considerable number of A549P-3IR cells surviving irradiation displayed a spindle morphology and high migration rate. Blocking integrin beta1 activity reduced the migration rate of A549P-3IR cells and altered the cell morphology allowing them to assume a round shape. These results suggest that the A549P-3 cells surviving irradiation acquire a highly invasive integrin beta1-dependent phenotype, and integrin beta1 might be a potentially effective therapeutic target in combination with radiotherapy.

Advantages of Using Paclitaxel in Combination with Oncolytic Adenovirus Utilizing RNA Destabilization Mechanism.

Oncolytic virotherapy is a novel approach to cancer therapy. Ad-fosARE is a conditionally replicative adenovirus engineered by inserting AU-rich elements (ARE) in the 3'-untranslated region of the E1A gene. In this study, we examined the oncolytic activity of Ad-fosARE and used it in a synergistic combination with the chemotherapeutic agent paclitaxel (PTX) for treating cancer cells. The expression of E1A was high in cancer cells due to stabilized E1A-ARE mRNA. As a result, the efficiency of its replication and cytolytic activity in cancer cells was higher than in normal cells. PTX treatment increased the cytoplasmic HuR relocalization in cancer cells, enhanced viral replication through elevated E1A expression, and upregulated CAR (Coxsackie-adenovirus receptor) required for viral uptake. Furthermore, PTX altered the instability of microtubules by acetylation and detyrosination, which is essential for viral internalization and trafficking to the nucleus. These results indicate that PTX can provide multiple advantages to the efficacy of Ad-fosARE both in vitro and in vivo, and provides a basis for designing novel clinical trials. Thus, this virus has a lot of benefits that are not found in other oncolytic viruses. The virus also has the potential for treating PXT-resistant cancers.

Conditionally Replicative Adenovirus Controlled by the Stabilization System of AU-Rich Elements Containing mRNA.

AU-rich elements (AREs) are RNA elements that enhance the rapid decay of mRNAs, including those of genes required for cell growth and proliferation. HuR, a member of the embryonic lethal abnormal vision (ELAV) family of RNA-binding proteins, is involved in the stabilization of ARE-mRNA. The level of HuR in the cytoplasm is up-regulated in most cancer cells, resulting in the stabilization of ARE-mRNA. We developed the adenoviruses AdARET and AdAREF, which include the ARE of TNF-α and c-fos genes in the 3'-untranslated regions of the E1A gene, respectively. The expression of the E1A protein was higher in cancer cells than in normal cells, and virus production and cytolytic activities were also higher in many types of cancer cells. The inhibition of ARE-mRNA stabilization resulted in a reduction in viral replication, demonstrating that the stabilization system was required for production of the virus. The growth of human tumors that formed in nude mice was inhibited by an intratumoral injection of AdARET and AdAREF. These results indicate that these viruses have potential as oncolytic adenoviruses in the vast majority of cancers in which ARE-mRNA is stabilized.

Novel function of transcription factor ATF5: blockade of p53-dependent apoptosis induced by ionizing irradiation.

PURPOSE: To find a new molecule that affects p53-dependent radiosensitivity. METHODS AND MATERIALS: A mouse sarcoma cell line, QRsP(p53+/+), was used. From this cell line, we established a radiosensitive clone and a radioresistant one. Colony assay, p53 gene transfer, a luciferase assay for p53 and p21, animal transplantation experiment, and DNA array analyses were performed. RESULTS: Microarray showed marked reduction of a transcription factor, ATF5, both in vitro and in vivo for the radiosensitive clone. Interestingly, flow cytometric analysis demonstrated marked apoptosis for the radiosensitive clone by p53 cloned adenovirus infection. Luciferase reporter assay revealed that ATF5 suppressed the transactivational activity of p53 and p63. By ATF5 gene transfer, the radiosensitive clone regained resistance to both ionizing-radiation and Ad-p53 infection-induced cell death. Surprisingly, time-lapse cell migration observation revealed greater cell motility for ATF5-transfected radiosensitive clone. CONCLUSIONS: It seems likely that ATF5 is a potent repressor of p53 and elevated expression of ATF5 in a tumor may relate to enhanced malignant phenotypes, such as radioresistance or greater cell motility.

Increased motility and invasiveness in tumor cells that survive 10 Gy irradiation.

Radiotherapy is an important noninvasive treatment for many types of cancer. However, it has been reported that the proliferative, invasive, and metastatic capacities of tumor cells can be increased in the repopulated tumors that survive radiotherapy. We have previously established a radiation-surviving cell model for the human non-small cell lung cancer cell line H1299 by harvesting relic cells 14 days after irradiation (IR cells). Here, we report that cell invasion, cell migration, and cell adhesion are enhanced in these surviving cancer cells. The mRNA expression levels of matrix metalloproteinases (MMPs), including mmp1, mmp2, and mmp9, were upregulated in IR cells compared with parental cells. A gelatin zymogram, wound healing assay, and invasion assay showed increased MMP activity, cell motility, and invasiveness in IR cells, respectively. Moreover, IR cells adhered more tightly to collagen-coated dishes than parental cells. Consistently, paxillin, phosphorylated FAK, integrin beta1, and vinculin were strongly localized at focal adhesions in IR cells, as visualized by immunofluorescence. In this report, we identify molecules responsible for the malignant properties of tumor cells that survive irradiation. These molecules may be important therapeutic targets for the control of repopulated tumors after radiotherapy.

Oncolytic potential of an E4-deficient adenovirus that can recognize the stabilization of AU-rich element containing mRNA in cancer cells.

AU-rich elements (AREs) are RNA elements that enhance the rapid decay of mRNA. The fate of ARE-mRNA is controlled by ARE-binding proteins. HuR, a member of the embryonic lethal abnormal vision (ELAV) family of RNA-binding proteins, is involved in the export and stabilization of ARE-mRNA. In the vast majority of cancer cells, HuR constitutively relocates to the cytoplasm, resulting in the stabilization of ARE-mRNA. Previously, we described that the adenovirus gene product, E4orf6, which is necessary for virus replication, participates in ARE-mRNA export and stabilization. In the present study, we showed the oncolytic potential of E4orf6-deleted adenovirus dl355, which is expected to be replicated selectively in cancer cells. Virus production and cytolytic activity of dl355 were higher in cancer cells than in normal cells. HuR-depletion downregulated dl355 replication, demonstrating that ARE-mRNA stabilization is required for the production of this virus. Tumor growth was inhibited in nude mice by an intratumoral injection of dl355. Furthermore, dl355 had a stronger oncolytic effect than E1B55k-deleted adenovirus. These results indicate that dl355 has potential as an oncolytic adenovirus for a large number of cancers where ARE-mRNA is stabilized.

The diacylated lipopeptide FSL-1 enhances phagocytosis of bacteria by macrophages through a Toll-like receptor 2-mediated signalling pathway.

A significant amount of evidence has been accumulated to show that Toll-like receptors (TLRs) function as sensors for microbial invasion. However, little is known about how signalling triggered by TLRs leads to the phagocytosis of pathogens. This study was designed to determine whether stimulation of TLR2 mainly with the lipopeptide FSL-1 plays a role in the phagocytosis of pathogens by macrophages. FSL-1 enhanced the phagocytosis of Escherichia coli to a markedly greater extent than it did that of Staphylococcus aureus, but did not enhance the phagocytosis of latex beads. FSL-1 stimulation resulted in enhanced phagocytosis of bacteria by macrophages from TLR2(+/+) mice but not by those from TLR2(-/-) mice. Chinese hamster ovary cells stably expressing TLR2 failed to phagocytose these bacteria, but the cells expressing CD14 did. FSL-1 induced upregulation of the expression of phagocytic receptors, including MSR1, CD36, DC-SIGN and Dectin-1 in THP-1 cells. Human embryonic kidney 293 cells transfected with DC-SIGN and MSR1 phagocytosed these bacteria. These results suggest that the FSL-1-induced enhancement of phagocytosis of bacteria by macrophages may be explained partly by the upregulation of scavenger receptors and the C-type lectins through TLR2-mediated signalling pathways, and that TLR2 by itself does not function as a phagocytic receptor.

X-ray irradiation altered chemosensitivity of a p53-null non-small cell lung cancer cell line.

Radiotherapy is an effective approach to treating many types of cancer. Recent progress in radiotherapy technology, such as intensity-modulated radiation therapy (IMRT) and three-dimensional (3D) radiotherapy, allow precise energy transfer to the tumor, which has improved local control rates. However, the emergence of tolerant cells during or after radiotherapy remains problematic. In the present study, we first established a cell population from H1299, the p53-null non-small cell lung cancer cell line, by 10 Gy irradiation using 6 MV X-rays. The radio- and chemosensitivity of this cell population (referred to as H1299-IR) was determined using colony formation analyses and MTS assays. Compared with the parental cell line, the radiosensitivity of H1299-IR was apparently the same. H1299 and H1299-IR were both more radio tolerant than the A549 cell line. However, H1299-IR became significantly more sensitive to cisplatin, an antitumor agent. After exposure to 25 mug/ml cisplatin for 2 h, parental cells steadily grew during the MTS assay, whereas the sensitivity of H1299-IR cells doubled both at 24 and 48 h. Microarray analysis of over 30,000 H1299-IR genes (Agilent Technology) revealed that 12 and 15 genes were up- (> 2.0) and down- (< 2.0) regulated, respectively. Rad51d (homologous recombination repair protein) gene was down-regulated 2.8-fold, whereas matrix metalloproteinase 1 (collagenase-1) gene was up-regulated 4.4-fold. These results indicated that some p53-null non-small cell lung cancers could be successfully treated when X-ray radiotherapy was administered with subsequent or concurrent cisplatin chemotherapy.

The diacylated lipopeptide FSL-1 induces TLR2-mediated Th2 responses.

The diacylated lipopeptide FSL-1 enhanced the generation of IgG antibodies in TLR2(+/+) mice, but not in TLR2(-/-) mice, when administered together with hen egg lysozyme as an antigen. Escherichia coli lipopolysaccharide enhanced the generation of antigen-specific antibodies in both TLR2(-/-) and TLR2(+/+) mice. In TLR2(+/+) mice, the level of enhancement due to FSL-1 was similar to that caused by lipopolysaccharide. Analysis of the IgG antibodies subclass demonstrated that the level of Th2-type IgG1 antibodies was higher than that of Th1-type IgG2a antibodies. Both FSL-1 and lipopolysaccharide induced production of IL-10 and IL-6 by splenocytes from TLR2(+/+) mice. Lipopolysaccharide also induced production of these cytokines by splenocytes from TLR2(-/-) mice, but FSL-1 did not. Neither FSL-1 nor lipopolysaccharide induced IL-12p70 production by splenocytes from either type of mice. FSL-1 upregulated B7.2 expression in B220(+) cells from TLR2(+/+) mice but not those from TLR2(-/-) mice, whereas lipopolysaccharide upregulated B7.2 expression in B220(+) cells from both types of mice. FSL-1 and, to a lesser extent, lipopolysaccharide activated mitogen-activated protein kinases in splenocytes. FSL-1 and, to a lesser extent, lipopolysaccharide induced the expression of c-Fos, which is known to be involved in Th2-type responses, in splenocytes. Thus, this study demonstrated that FSL-1 possessed TLR2-mediated Th2-type responses in vivo.

The synthetic analogue of mycoplasmal lipoprotein FSL-1 induces dendritic cell maturation through Toll-like receptor 2.

Granulocyte-macrophage colony-stimulating factor-differentiated bone marrow-derived dendritic cells were stimulated with the synthetic lipopeptide S-(2,3-bispalmitoyloxypropyl)-CGDPKHSPKSF (FSL-1) or the Escherichia coli lipopolysaccharide. FSL-1 induced the production of TNF-alpha and IL-12 by C57BL/6-derived bone marrow-derived dendritic cells but not by bone marrow-derived dendritic cells from Toll-like receptor 2-deficient (TLR2(-/-)) mice. Lipopolysaccharide induced the production of TNF-alpha and IL-12 by bone marrow-derived dendritic cells derived from either type of mice. FSL-1 did not induce production of IL-10 by bone marrow-derived dendritic cells from either type of mice, whereas lipopolysaccharide induced small amounts of IL-10 by bone marrow-derived dendritic cells from both types of mice. The upregulation by FSL-1 of the expression of CD80, CD86 and the MHC class II molecule IA(b) was dose- and time-dependent on the surfaces of C57BL/6-derived bone marrow-derived dendritic cells but not on the surface of TLR2(-/-)-derived bone marrow-derived dendritic cells. Lipopolysaccharide upregulated the expression of these molecules on the surfaces of bone marrow-derived dendritic cells from both types of mice. The expression of CD11c on the surfaces of C57BL/6-derived bone marrow-derived dendritic cells was upregulated by stimulation with both FSL-1 and lipopolysaccharide up to 12 h; thereafter, the expression was downregulated. The results suggest that FSL-1 can accelerate maturation of bone marrow-derived dendritic cells and this FSL-1 activity is mediated by TLR2.