Human Toll-like receptor 2 mediates induction of the antimicrobial peptide human beta-defensin 2 in response to bacterial lipoprotein.

Recognition of pathogens by Drosophila Toll or human Toll-like receptors results in translocation of Dorsal or its human homologue NF-kappaB, respectively; in Drosophila, this is followed by the production of antimicrobial peptides serving as antimicrobial effector system of the innate immune response. We investigated whether human Toll-like receptors also mediate induction of the synthesis of antimicrobial peptides. We found that HEK293 cells transfected with Toll-like receptor 2, but not wild-type cells responded to stimulation with bacterial lipoprotein by production of human beta-defensin 2. Furthermore, the human lung epithelial cell line A549 was found to constitutively express Toll-like receptor 2 and to produce beta-defensin 2 in response to bacterial lipoprotein. This response was abrogated by blocking the signaling pathway activated through Toll-like receptors by transfecting the A549 cells with a dominant-negative form of IRAK-2. Thus, exposure of human cells to bacterial lipoprotein elicits production of the antimicrobial peptide beta-defensin 2 through Toll-like receptor 2.     

CC chemokine receptor 5 (CCR5) mRNA expression in pulmonary sarcoidosis

The CC chemokine receptor 5 (CCR5) binds the chemokine ligands RANTES (CCL5) and MIP-1 (CCL3), which have been implicated in the development of alveolitis in sarcoidosis. We have, therefore, investigated CCR5 mRNA expression in bronchoalveolar lavage fluid (BALF) cells from patients with sarcoidosis. Further, we explored whether there was any association between CCR5 mRNA expression and the presence of the CCR5Δ32 DNA polymorphism. Semiquantitative RT-PCR was used to determine CCR5 mRNA expression from BALF cells from 16 control subjects (C) and 39 patients with sarcoidosis (S). The data on the CCR5Δ32 polymorphism, determined by PCR-SSP, were available for 37 patients. CCR5 mRNA expression was significantly upregulated in sarcoidosis (median±SEM, C, 0.00±0.07; S, 0.12±0.07; P<0.05). When patients were evaluated according to their CCR5Δ32 genotype, an interesting trend emerged with Δ32 positive patients (wt, mt) expressing less mRNA than the patients with both wild-type alleles (wt, wt): 0.00±0.09, and 0.26±0.09, respectively; P>0.05). In conclusion, upregulation of CCR5 mRNA in BALF of patients with sarcoidosis is consistent with its chemokine ligands RANTES and MIP-1 playing a pivotal role in inflammatory cell recruitment to disease sites. Though the data from this pilot study had no clinical correlations we suggest that further studies are warranted on the role of this Th1 subset marker in the pathogenesis of sarcoidosis.     

Functional expression of chemokine receptor 2 by normal human eosinophils

BACKGROUND: Within the granulocytes, the CC chemokines preferentially activate basophils and eosinophils on binding to chemokine receptors (CCRs). In vivo administration of neutralizing anti-monocyte chemoattractant protein 1 (MCP-1) antibodies can block accumulation of eosinophils in the lungs of antigen-challenged animals. OBJECTIVE: We studied a panel of chemokines for chemotactic activity in normal human eosinophils from healthy donors with a special focus on MCP-1, identified the respective receptor required for the biological response of eosinophils, and investigated mediators used for signal transduction. METHODS: Cells were enriched by magnetic cell sorting. Receptor expression in eosinophils was shown by RT-PCR and fluorescence-activated cell sorting. The biological response was tested in chemotaxis and calcium mobilization assays. RESULTS: Eosinophils have detectable mRNA for CCR2, and the receptor protein is expressed on cell surfaces. MCP-1 induces chemotaxis and calcium mobilization in eosinophils. The chemotactic activity of MCP-1 revealed a double-peaked dose-response curve; one of the peaks is abolished by addition of a blocking antibody to CCR2, but it is insensitive to blocking of CCR1 or CCR3. Specific enzyme inhibitors ruled out signaling characteristics of CCR2 in eosinophils. CONCLUSION: Normal human eosinophils express functional CCR2 on cell surfaces.     

Therapeutic targeting of CC ligand 21 or CC chemokine receptor 7 abrogates pulmonary fibrosis induced by the adoptive transfer of human pulmonary fibroblasts to immunodeficient mice

Idiopathic interstitial pneumonias (IIPs) are a collection of pulmonary fibrotic diseases of unknown etiopathogenesis. CC chemokine receptor 7 (CCR7) is expressed in IIP biopsies and primary fibroblast lines, but its role in pulmonary fibrosis was not previously examined. To study the in vivo role of CCR7 in a novel model of pulmonary fibrosis, 1.0 x 10(6) primary fibroblasts grown from idiopathic pulmonary fibrosis/usual interstitial pneumonia, nonspecific interstitial pneumonia, or histologically normal biopsies were injected intravenously into C.B-17 severe combined immunodeficiency (SCID)/beige (bg) mice. At days 35 and 63 after idiopathic pulmonary fibrosis/usual interstitial pneumonia fibroblast injection, patchy interstitial fibrosis and increased hydroxyproline were present in the lungs of immunodeficient mice. Adoptively transferred nonspecific interstitial pneumonia fibroblasts caused a more diffuse interstitial fibrosis and increased hydroxyproline levels at both times, but injected normal human fibroblasts did not induce interstitial remodeling changes in C.B-17SCID/bg mice. Systemic therapeutic immunoneutralization of either human CCR7 or CC ligand 21, its ligand, significantly attenuated the pulmonary fibrosis in groups of C.B-17SCID/bg mice that received either type of IIP fibroblasts. Thus, the present study demonstrates that pulmonary fibrosis is initiated by the intravenous introduction of primary human fibroblast lines into immunodeficient mice, and this fibrotic response is dependent on the interaction between CC ligand 21 and CCR7.     

CC chemokine receptor 4 ligand production by bronchoalveolar lavage fluid cells in cigarette-smoke-associated acute eosinophilic pneumonia

We examined the production of macrophage-derived chemokine (MDC/CCL22) and thymus- and activation-regulated chemokine (TARC/CCL17) by bronchoalveolar lavage fluid (BALF) cells in cigarette-smoke-associated acute eosinophilic pneumonia (CS-AEP). The CC Chemokine Receptor 4 (CCR4) ligand levels in BALF from patients with CS-AEP were considerably higher than those in healthy volunteers and correlated well with Th2 cytokine levels. Interleukin-4 enhanced CCR4 ligand production. MDC expression was observed in CD68-positive cells from patients with CS-AEP and in healthy control smokers. In contrast, TARC expression in CD68- or CD1a-positive cells was detected only in CS-AEP. An in vivo cigarette smoke challenge test induced increases in CCR4 ligands in the BALF and in the cultured supernatant of BALF adherent cells. These results suggest that alveolar macrophages and dendritic cells contribute to the pathogenesis of CS-AEP by generating CCR4 ligands, probably in response to cigarette smoke.     

Differential expression of immunoregulatory genes in monocytes in response to Porphyromonas gingivalis and Escherichia coli lipopolysaccharide

Porphyromonas gingivalis lipopolysaccharide (LPS) (strain W50) interacts with Toll‐like receptor 2 (TLR‐2) leading to cytokine expression and inflammation, and thereby plays a key role in the pathogenesis of periodontal disease. The aims of this study were to investigate gene expression of key regulatory mediators of innate immune responses in a human monocytic cell line (THP‐1) to P. gingivalis LPS and to compare these results with those obtained using the TLR‐4 ligand, Escherichia coli LPS. Custom‐made Taqman low‐density arrays were used for expression profiling of 45 different cytokine‐related genes. Both types of LPS highly up‐regulated interleukin (IL)‐1α and IL‐1β, IL‐18 receptor (IL‐18R), IL‐18R accessory protein and IL‐1 family (IL‐1F)9. Expression levels of IL‐1F6, IL‐1F7 and caspase‐1 were unaltered by either LPS. Genes for tumour necrosis factor‐α, IL‐6, leukaemia inhibitory factor and IL‐32 were also highly induced by both LPS. For a subset of genes, including CXC chemokine ligand 5 (CXCL5), expression was induced only by E. coli LPS or was up‐regulated more highly by E. coli compared with P. gingivalis LPS in THP‐1 monocytes. A similar expression pattern was also observed in dendritic cells. Analysis of signalling pathways which lead to CXCL5 expression indicated that the mechanisms underpinning the differential responses did not involve the recruitment of different adaptor proteins by TLR‐2 and TLR‐4, and therefore occur downstream of the receptor–adaptor complex. We conclude that differences in signalling pathways activated by TLR‐2 and TLR‐4 ligands lead to differential innate immune responses which may be important in polymicrobial diseases such as periodontal disease.     

Fibrinogen-neutrophil interactions in response to fMLP and Porphyromonas gingivalis fimbrial peptides

Porphyromonas gingivalis (P.g) is the primary bacterial agent in many forms of chronic periodontitis. Since polymorphonuclear leukocytes (PMNs) are first-line responders to P.g.- induced inflammation, and fibrinogen is important for in vivo PMN in this disease, we have studied the effect of N-formyl-methionyl-leucyl-phenylalanine (fMLP) (an inflammatory stimulus), P.g. fimbriae and fimbrial peptides (based on FimA, the main structural protein of P.g. fimbriae) on PMN-fibrinogen interactions. Freshly isolated human PMNs were allowed to react with FITC-Fibrinogen and various fimbrial peptides (denoted as FimA followed by amino acid number within whole FimA protein), and FITC-Fibrinogen binding was measured using flow cytometry. Freshly isolated neutrophils were also challenged with Fibrinogen and/or fimbrial peptides to measure IL-8 secretion using ELISA. Our studies show that fibrinogen binding to PMNs is enhanced (p < 0.01) in response to fMLP as well as fimbrial peptides (FimA 61-80) containing the motif LTTE (p < 0.01) in a dose dependent manner but not in response to peptides without that motif. We also observed that fMLP and FimA 61-80 have an additive effect on fibrinogen binding to PMNs (p < 0.05), and fMLP and FimA 171-185 significantly inhibit fMLP-induced fibrinogen binding (p < 0.01). To determine of the role of inflammatory cytokines, we examined IL-8 release from PMNs in response to combinations of P. gingivalis fimbriae, fMLP and fibrinogen. In all cases, IL-8 release increased in a dose-dependent manner (p < 0.05). fMLP-fibrinogen effect on IL-8 release from PMNs was synergistic while fimbriae-fibrinogen effect was additive. In summary, PMN priming by fimbrial peptides facilitates fibrinogen-PMN interaction and may increase inflammation.     

Immunization enhances inflammation and tissue destruction in response to Porphyromonas gingivalis

It is well established that host-bacterium interactions play a critical role in the initiation and progression of periodontal diseases. By the use of inhibitors, it has been shown that mediators associated with the innate immune response significantly contribute to the disease process. Less is known regarding the role of the acquired immune response. To investigate mechanisms by which the acquired immune response to Porphyromonas gingivalis could affect connective tissue, we used a well-documented calvarial model to study host-bacterium interactions. Injection of P. gingivalis stimulated gamma interferon, interleukin 6, macrophage inflammatory protein 2, and monocyte chemoattractant protein 1 expression as determined by real-time PCR. Prior immunization against P. gingivalis significantly enhanced the mRNA levels of these cytokines and chemokines. Similarly, immunization significantly increased and prolonged the formation of a polymorphonuclear leukocyte and mononuclear cell infiltrate (P < 0.05). In addition, the area of connective tissue destruction, osteoclastogenesis, bone loss, mRNA expression of proapoptotic genes, and degree of fibroblast apoptosis were increased in immunized mice (P < 0.05). These results indicate that activation of the acquired immunity by P. gingivalis increases the inflammatory and destructive responses which occur in part through up-regulating the innate immune response and enhancing osteoclastogenesis and fibroblast apoptosis.     

Serum antibodies to Porphyromonas gingivalis block the prostaglandin E2 response to lipopolysaccharide by mononuclear cells.

The ability of rabbit and monkey immune sera to neutralize prostaglandin E2 (PGE2) production by human monocytes stimulated with lipopolysaccharide (LPS) was examined. CD14-dependent LPS activation of PGE2 was examined under assay conditions which allowed the comparison of preimmune and immune sera. Serum obtained from rabbits immunized with formalin-fixed Porphyromonas gingivalis cells dramatically reduced the amount of PGE2 produced in response to LPS obtained from three different strains of P. gingivalis but not that from Escherichia coli or Bacteroides fragilis. In addition, a significant reduction in the mean PGE2 level was observed in the presence of sera from immunized but not control monkeys employed in a vaccine trial. Immune serum samples from five of nine immunized monkeys were able to reduce LPS-induced production of PGE2 by greater than 50% compared to that in the corresponding preimmune sera. Immune monkey serum, similar to immune rabbit serum, blocked PGE2 production in response to P. gingivalis LPS but not E. coli LPS. These data demonstrate that immunization with P. gingivalis whole cells can elicit an antibody response that is able to block the PGE2 response to LPS. Neutralization of LPS-mediated inflammatory mediator production may account in part for the observed suppression of alveolar bone loss in immunized monkeys.     

Human dendritic cells respond to Porphyromonas gingivalis LPS by promoting a Th2 effector response in vitro

Understanding how mucosal pathogens modulate the immune response may facilitate the development of vaccines for disparate human diseases. In the present study, human monocyte-derived DC (MDDC)were pulsed with LPS of the oral pathogen Porphyromonas gingivalis and Escherichia coli 25922 and analyzed for: (i) production of Th-biasing/inflammatory cytokines; (ii) maturation/costimulatory molecules; and (iii) induction of allogeneic CD4+ and naive CD45RA+ T cell proliferation and release of Th1 or Th2 cytokines. We show that E. coli LPS-pulsed MDDC released Th1-biasing cytokines - consisting of high levels of IL-12 p70, IFN-gamma-inducible protein 10 (IP-10) - but also TNF-alpha, IL-10, IL-6 and IL-1beta. In contrast, no IL-12 p70 or IP-10, and lower levels of TNF-alpha and IL-10 were induced by P. gingivalis LPS. These differences were sustained at LPS doses that yielded nearly equivalent maturation of MDDC; moreover the T cell response was consistent: E. coli LPS-pulsed MDDC induced higher T cell proliferation, and T cells released more IFN-gamma and IL-2, but less IL-5 than T cells co-cultured with P. gingivalis LPS pulsed-MDDC. IL-13 was secreted by naive CD45RA+CD45RO-CD4+ T cells in response to P. gingivalisLPS-pulsed MDDC. These results suggest that human MDDC can be polarized by LPS from the mucosal pathogen P. gingivalis to induce a Th2 effector response in vitro.     

Localizing antibody-defined immunoreactivity in Porphyromonas gingivalis HtpG recognized by human serum utilizing selective protein expression

Earlier studies suggested that specific immunoreactive domains of the prokaryotic homologue of Hsp90, HtpG, might contribute to the virulence of the periodontal pathogen, Porphyromonas gingivalis (Pg) [J. Periodontol. 70 (1999) 1185]. Since serum antibodies to this protein appeared to be associated with oral health, we developed a rapid epitope-mapping system that could be tailored to detect antibodies against specific immunoreactive regions of the Pg HtpG protein. This paper describes the use of Caulobacter crescentus (Cc) and the creation of a Cc RsaA fusion protein library that defined specific regions of the Pg HtpG protein. The fusion protein library was used to identify immunoreactive regions in the Pg HtpG dominant in patient and control sera. The development of methods to rapidly localize dominant immunoreactive regions in protein antigens may prove useful for the development of screening tests, vaccines and therapeutics in periodontal and other infectious diseases.     

Reciprocal chemokine receptor and ligand expression in the human placenta: implications for cytotrophoblast differentiation.

At the onset of pregnancy, the human placenta, which forms the interface between the embryo/fetus and the mother, must rapidly develop into a life-sustaining organ. The many unusual processes entailed in placental development include the poorly understood phenomenon of maternal tolerance of the hemiallogeneic cells of the conceptus, including, most remarkably, placental trophoblasts that invade the uterine wall. To investigate whether this fetal organ exerts control over the maternal immune system at the level of leukocyte trafficking, we examined placental expression of chemokines, well-known cytokine regulators of leukocyte movements. In situ hybridization revealed abundant expression of 13 chemokines in the stromal but not the trophoblast compartment of chorionic villi. Potential roles for these molecules include recruitment of the resident macrophage (Hofbauer cell) population to the villi. In parallel, cytotrophoblast production of a panel of nine chemokine receptors was assessed by using RNase protection assays. The numerous receptors detected suggested the novel possibility that the paracrine actions of chemokine ligands derived from either the villous stroma or the decidua could mediate general aspects of placental development, with specific contributions to cytotrophoblast differentiation along the pathway that leads to uterine invasion.     

CC chemokine receptor 5 and CXC chemokine receptor 6 expression by lung CD8+ cells correlates with chronic obstructive pulmonary disease severity

Chronic obstructive pulmonary disease (COPD) is a progressive disease associated with a cellular inflammatory response. CD8(+) T cells are implicated in COPD pathogenesis, and their numbers significantly correlate with the degree of airflow limitation. Dendritic cells (DCs) are important sentinel immune cells, but little is known about their role in initiating and maintaining the CD8 T-cell response in COPD. To investigate the mechanisms for CD8(+) T-cell recruitment to the lung, we used resected human lung tissue to analyze chemokine receptor expression by CD8(+) T cells and chemokine production by CD1a(+) DCs. Among 11 surveyed chemokine receptors, only CC chemokine receptor (CCR5), CXC chemokine receptor (CXCR) 3, and CXCR6 correlated with COPD severity as defined by criteria from the Global Initiative for Chronic Obstructive Lung Disease. The CD8(+) T cells displayed a Tc1, CD45RA(+) effector memory phenotype. CD1a(+) DCs produced the respective ligands for CCR5 and CXCR3, CCL3 and CXCL9, and levels correlated with disease severity. CD1a(+) DCs also constitutively expressed the CXCR6 ligand, CXCL16. In conclusion, we have identified major chemokine elements that potentially mediate CD8(+) T-cell infiltration during COPD progression and demonstrated that CD1a(+) mucosal-associated DCs may sustain CD8(+) T-cell recruitment/retention. Chemokine targeting may prove to be a viable treatment approach.     

Calprotectin expression in vitro by oral epithelial cells confers resistance to infection by Porphyromonas gingivalis

Calprotectin, an S100 calcium-binding protein with broad-spectrum antimicrobial activity in vitro, is expressed in neutrophils, monocytes, and gingival keratinocytes. In periodontitis, calprotectin appears upregulated and is detected at higher levels in gingival crevicular fluid and tissue specimens. How calprotectin contributes to the pathogenesis of periodontal diseases is unknown. To isolate the effects of calprotectin, a calprotectin-negative oral epithelial cell line was transfected with calprotectin genes to enable expression. Porphyromonas gingivalis was permitted to bind and invade transfected cells expressing calprotectin and sham transfectants. Rates of invasion into both cell lines were compared using the antibiotic protection assay. Transfected cells expressing calprotectin showed 40 to 50% fewer internalized P. gingivalis than sham transfectants. Similarly, binding to calprotectin expressing cells was reduced approximately twofold at all time points (15, 30, 45, and 60 min) as estimated by immunofluorescence analysis. Independent of invasion, however, prolonged exposure to P. gingivalis induced epithelial cell rounding and detachment from the substratum. These morphological changes were delayed, however, in cells expressing calprotectin. Using P. gingivalis protease-deficient mutants, we found that Arg-gingipain and Lys-gingipain contributed to epithelial cell rounding and detachment. In conclusion, expression of calprotectin appears to protect epithelial cells in culture against binding and invasion by P. gingivalis. In addition, cells expressing calprotectin are more resistant to detachment mediated by Arg-gingipain and Lys-gingipain. In periodontal disease, calprotectin may augment both the barrier protection and innate immune functions of the gingival epithelium to promote resistance to P. gingivalis infection.     

Expression of beta-defensin 1 and 2 mRNA by human monocytes,macrophages and dendritic cells

Human beta-defensins are broad-spectrum antimicrobial peptides known to be produced by epithelial cells. It was recently shown that beta-defensins also display chemotactic activity for dendritic cells (DC) and T cells, and thus may serve to link innate and adaptive immunity. The aim of the present study was to explore expression of mRNA for these peptides in mononuclear phagocytes and DC. The results revealed that monocytes, monocyte-derived-macrophages (MDM), and monocyte-derived-dendritic cells (DC) all express human-beta-defensin-1 (hBD-1) mRNA. hBD-1 mRNA expression by monocytes and MDM was increased after activation with interferon-gamma (IFN-gamma) and/or lipopolysaccharide (LPS) in a dose- and time-dependent fashion. Alveolar macrophages showed an intense hBD-1 expression, which could not be further increased. Expression of hBD-1 mRNA by immature DC was low, and increased considerably after maturation. Monocytes, MDM, alveolar macrophages and DC showed a limited expression of human beta-defensin-2 (hBD-2) mRNA, which could only be increased in monocytes and alveolar macrophages by IFN-gamma and/or LPS in a dose- and time-dependent fashion. Immunocytochemical stainings demonstrated the expression of hBD-2 peptide by freshly isolated blood monocytes and alveolar macrophages in cytospin preparations.     

Adenoid-derived TH2 cells reactive to allergen and recall antigen express CC chemokine receptor 4

BACKGROUND: Interrupting recruitment of allergen-specific T(H)2 cells to the airway is an attractive potential therapeutic strategy for allergic disease. CC chemokine receptor 4 (CCR4) is preferentially expressed on T(H)2 cells, and CCR4-expressing cells have been described at sites of allergic inflammation. However, whether selective recruitment of allergen-specific T(H)2 cells to the airways occurs through CCR4 or other chemokine receptors remains controversial. OBJECTIVE: We investigated the expression of the T(H)2-associated chemokine receptors (CCR3, CCR4, and CCR8) by primary antigen-specific human airway T(H)2 cells. METHODS: Children undergoing elective adenoidectomy were recruited, and their atopic status was determined. Adenoid cells were cultured with allergen or recall antigen. Flow cytometric analyses permitted identification of T(H) cells proliferating in response to antigen and characterization of chemokine receptor and cytokine expression. RESULTS: An increased proportion of airway CD4(+) T cells proliferated to allergen in atopic children (n = 6, of which 4 were given diagnoses of asthma or rhinitis) compared with nonatopic children (P =.0004). These cells were 44.7% (32.6% to 50.0%) IL-4(+) and only 2.5% (0.6% to 3.3%) IFN-(gamma) and showed a greater than 5-fold upregulation of CCR4 expression to 54.0% (40.7% to 67.8%) after culture, whereas CCR3 was expressed on 9.7% (7.4% to 18.9%) of allergen-reactive cells and CCR8 on less than 1%. Interestingly, increased expansion of recall antigen-specific cells was also seen in atopic children, and these cells were also predominantly of a T(H)2 CCR4(+) phenotype. CONCLUSION: We conclude that airway allergen-specific T(H)2 cells are CCR4(+), but in the atopic child CCR4 does not distinguish between recall antigen and allergen specificity.     

Estrogen inhibits apoptosis and promotes CC motif chemokine ligand 13 expression on synovial fibroblasts in rheumatoid arthritis

Objective: Female patients have a higher prevalence of rheumatoid arthritis (RA) than male patients, suggesting that female sex hormones contribute to the disease pathogenesis. We herein report the findings of our study, which was conducted to clarify the role of estrogen in the pathogenesis of RA.

Methods: Cultured human synovial fibroblasts from a patient with RA were treated with 17β-estradiol (E₂). The effects of E₂ against cellular activation and apoptosis were evaluated. To identify the disease-related genes altered by E₂ treatment, the changes in the gene expression of the cells stimulated with and without E₂ were evaluated using a microarray analysis.

Results: We found that E₂-mediated cellular activation signaling through extracellular signal-regulated kinase (ERK)-1/2. E₂ possessed a suppressive effect for apoptosis and a promotive effect for tumor necrosis factor (TNF)-α-induced matrix metalloproteinase (MMP)-3 production on the synovial fibroblasts. A microarray analysis revealed that E₂ profoundly upregulated CC motif chemokine ligand 13 (CCL13) gene expression.

Conclusions: E₂ could mediate cellular activation signaling through ERK-1/2 on the synovial fibroblasts. The present data suggest that E₂ has adverse effects on the pathogenesis of RA as a result of unregulated cell death, increased TNF-α-induced MMP-3 production, and CCL13 overproduction, subsequently resulting in the disease progression of RA.     

Estrogen inhibits apoptosis and promotes CC motif chemokine ligand 13 expression on synovial fibroblasts in rheumatoid arthritis

Objective: Female patients have a higher prevalence of rheumatoid arthritis (RA) than male patients, suggesting that female sex hormones contribute to the disease pathogenesis. We herein report the findings of our study, which was conducted to clarify the role of estrogen in the pathogenesis of RA. Methods: Cultured human synovial fibroblasts from a patient with RA were treated with 17β-estradiol (E(2)). The effects of E(2) against cellular activation and apoptosis were evaluated. To identify the disease-related genes altered by E(2) treatment, the changes in the gene expression of the cells stimulated with and without E(2) were evaluated using a microarray analysis. Results: We found that E(2)-mediated cellular activation signaling through extracellular signal-regulated kinase (ERK)-1/2. E(2) possessed a suppressive effect for apoptosis and a promotive effect for tumor necrosis factor (TNF)-α-induced matrix metalloproteinase (MMP)-3 production on the synovial fibroblasts. A microarray analysis revealed that E(2) profoundly upregulated CC motif chemokine ligand 13 (CCL13) gene expression. Conclusions: E(2) could mediate cellular activation signaling through ERK-1/2 on the synovial fibroblasts. The present data suggest that E(2) has adverse effects on the pathogenesis of RA as a result of unregulated cell death, increased TNF-α-induced MMP-3 production, and CCL13 overproduction, subsequently resulting in the disease progression of RA.