Prognostic value of circulating regulatory T cells for worsening heart failure in heart failure patients with reduced ejection fraction.

Regulatory T cells (Tregs) play a crucial role in the negative regulation of immune responses. Recent studies suggest that Tregs are involved in the pathogenesis of atherosclerosis and myocarditis. Here, we investigated the involvement of Tregs on worsening heart failure (HF) in patients with reduced ejection fraction (HF-REF). The study population consisted of 32 HF-REF patients who were hospitalized for worsening HF, and 18 control subjects. Cardiac function was evaluated by echocardiography. A single venous blood sample was collected before discharge. Circulating T cells were evaluated by flow cytometry. Tregs were defined as CD4(+)CD25(+)Foxp3(+)T cells, and the correlations between the frequency of Tregs and CRP, IL-6 and several echoparameters were analysed. Furthermore, all HF-REF patients were followed up to 12 months from discharge to examine the predictors of recurrent hospitalization.In HF-REF patients, Tregs were significantly decreased (5.9 ± 1.4 versus 8.0 ± 2.2%, P < 0.01), while CD4(+)HLADR(+)T cells were increased (10.1 ± 5.4 versus 7.3 ± 3.1%, P < 0.05), compared with controls. Tregs were negatively correlated with left ventricular end-diastolic dimension, and levels of CRP and IL-6. Eleven of 32 HF-REF patients were rehospitalized for worsening HF within 12 months. Multivariate Cox regression analysis showed that CD4/CD8 and frequency of Tregs were independent predictors for recurrent hospitalization. Furthermore, HF-REF patients expressing under 6% Treg/CD4(+)T cells showed a significantly higher incidence of recurrent hospitalization for worsening HF within 12 months.Our data suggest that Tregs might be involved in the pathogenesis of decompensated HF, and may be a novel predictor of poor prognosis in HF-REF patients.

Preventive effect of galectin-9 on double-stranded RNA-induced airway hyperresponsiveness in an exacerbation model of mite antigen-induced asthma in mice.

BACKGROUND: Viral respiratory infection is the most common cause of acute asthma exacerbation in patients with stable asthma. The replication of most respiratory viruses requires the generation of double-stranded RNA (dsRNA), resulting in the activation of host immune responses. Synthetic dsRNA, polyinosinic-polycytidylic acid (PolyIC), mimics the effects of viruses in various cell types. To evaluate new therapies for mite antigen-induced chronic asthma, we developed an acute exacerbation model of mouse chronic asthma using mite antigen and PolyIC. We also examined the preventive effects of recombinant galectin-9 (Gal-9) on acute asthma exacerbation in this model. METHODS: Airway hyperresponsiveness (AHR) was examined to evaluate the exacerbation of chronic asthma. To analyze airway inflammation, the numbers of inflammatory cells and concentrations of cytokines in the bronchoalveolar lavage fluid (BALF) were estimated by flow cytometry and enzyme-linked immunosorbent assay, respectively. RESULTS: AHR was accelerated by intranasal administration of PolyIC in addition to mite antigen. Levels of cytokines that contribute to AHR, including interferon-γ, tumor necrosis factor-α, and RANTES (CCR5), and of Gal-9 in the BALF were elevated in this acute asthma exacerbation mouse model. Intranasal administration of recombinant Gal-9 reduced the PolyIC-induced AHR and levels of these cytokines in the BALF. Further, Gal-9 suppressed the production of cytokines induced by PolyIC in the alveolar macrophages. CONCLUSIONS. Our findings demonstrated that exogenous Gal-9 suppressed dsRNA-induced AHR in an acute exacerbation model of chronic asthma in mice, and suggest that recombinant Gal-9 could be therapeutically effective for preventing acute asthma exacerbation.

Self-association of the galectin-9 C-terminal domain via the opposite surface of the sugar-binding site.

Galectin-9 is a lectin, which has various biological functions such as T-cell differentiation and apoptosis. Multivalency of carbohydrate binding is required for galectin-9 to function. Although galectin-1 (a proto-type galectin) forms an oligomer to obtain its multivalency, galectin-9 (a tandem-repeat-type one) has two carbohydrate recognition domains (CRD) in one polypeptide. However, a single CRD of galectin-9, especially the C-terminal one, exhibited pro-apoptotic activity suggesting oligomer formation capability. In this study, we monitored the nuclear magnetic resonance (NMR) signals of the backbone atoms of the galectin-9 C-terminal CRD (G9CCRD). Protein concentration dependence of the signals suggested that a region (F1-F4 strands) opposite to the ligand-binding site was involved in the self-association of G9CCRD. Site-directed mutagenesis in this region (Leu210, Trp277 and Leu279 to Thr; G9CCRD-3T) inhibited the self-association of G9CCRD, and improved the solubility, whereas it reduced its pro-apoptotic activity towards T cells. The high pro-apoptotic activity of G9CCRD seems to be due to the ability to form an oligomer. In addition, the same substitution in two-CRD-containing galectin-9 (G9Null-3T) also diminished the self-association and improved its solubility, although it hardly reduced the anti-proliferative and pro-apoptotic activities. G9CCRD contributes the self-association of full-length galectin-9 at high protein concentrations.

Cell surface galectin-9 expressing Th cells regulate Th17 and Foxp3+ Treg development by galectin-9 secretion.

Galectin-9 (Gal-9), a ß-galactoside binding mammalian lectin, regulates immune responses by reducing pro-inflammatory IL-17-producing Th cells (Th17) and increasing anti-inflammatory Foxp3(+) regulatory T cells (Treg) in vitro and in vivo. These functions of Gal-9 are thought to be exerted by binding to receptor molecules on the cell surface. However, Gal-9 lacks a signal peptide for secretion and is predominantly located in the cytoplasm, which raises questions regarding how and which cells secrete Gal-9 in vivo. Since Gal-9 expression does not necessarily correlate with its secretion, Gal-9-secreting cells in vivo have been elusive. We report here that CD4 T cells expressing Gal-9 on the cell surface (Gal-9(+) Th cells) secrete Gal-9 upon T cell receptor (TCR) stimulation, but other CD4 T cells do not, although they express an equivalent amount of intracellular Gal-9. Gal-9(+) Th cells expressed interleukin (IL)-10 and transforming growth factor (TGF)-ß but did not express Foxp3. In a co-culture experiment, Gal-9(+) Th cells regulated Th17/Treg development in a manner similar to that by exogenous Gal-9, during which the regulation by Gal-9(+) Th cells was shown to be sensitive to a Gal-9 antagonist but insensitive to IL-10 and TGF-ß blockades. Further elucidation of Gal-9(+) Th cells in humans indicates a conserved role of these cells through evolution and implies the possible utility of these cells for diagnosis or treatment of immunological diseases.

Possible regulatory role of galectin-9 on Ascaris suum-induced eosinophilic lung inflammation in mice.

BACKGROUND: Galectin-9 (Gal-9) is a member of the galectin family of lectins that exhibit binding affinity for ß-galactosides. We found a T cell line-derived Gal-9 with novel eosinophil chemoattractant activity, but its role in eosinophilic inflammation of the lung is unknown. We evaluated the role of Gal-9 in Ascaris suum-induced eosinophilic lung inflammation in mice. METHODS: To evaluate the role of Gal-9 in Ascaris suum-induced eosinophilic lung inflammation, we developed a mouse model of eosinophilic pneumonia induced by the Ascaris suum antigen, and analyzed eosinophilic inflammation in Gal-9-deficient mice. The therapeutic effects of recombinant Gal-9 on lung inflammation were also examined in this mouse model. To evaluate lung inflammation, numbers of inflammatory cells and cytokine levels in the bronchoalveolar lavage fluid (BALF) were estimated by flow cytometry and enzyme-linked immunosorbent assay, respectively. RESULTS: The BALF of this mouse model of eosinophilic pneumonia induced by the Ascaris suum antigen contained increased numbers of inflammatory cells and elevated Gal-9 levels. Compared with wild-type mice, the BALF of Gal-9-deficient mice contained higher numbers of both eosinophils and T helper type 2 (Th2) cells. Th2 cytokines and eotaxin levels were also higher, and levels of CD4+CD25+Foxp3+ regulatory T cells were lower in Gal-9-deficient mice than in wild-type mice. Intranasal administration of recombinant Gal-9 prevented eosinophilic inflammation of the lung and upregulated the release of endogenous Gal-9. CONCLUSIONS: Our findings suggest that Gal-9 negatively regulates Th2-mediated eosinophilic inflammation of the lung and that Foxp3+ regulatory T cells might be involved in suppressing allergic inflammation.

Galectin-9 suppresses Th17 cell development in an IL-2-dependent but Tim-3-independent manner.

Galectin-9 (Gal-9) ameliorates autoimmune reactions by suppressing Th17 cells while augmenting Foxp3(+) regulatory T cells (Tregs). However, the exact mechanism of Gal-9-mediated immune modulation has been elusive. In a MOG-induced experimental allergic encephalomyelitis model using Gal-9(-/-) mice, we observed exacerbated inflammation and an increase in IL-17-producing Th17 cells balanced by a decrease in Foxp3+ Tregs. During in vitro Th17 skewing using TGF-ß1 and IL-6, exogenous Gal-9 suppressed Th17 cell development and expanded Foxp3(+) Tregs from naïve CD4 T cells in an IL-2-dependent manner. Although Gal-9 induced cell death in Tim3-expressing differentiated Th17 cells, Gal-9 suppressed Th17 development in a Tim-3-independent. Benzyl-α-GalNAc (an O-glycan biosynthesis inhibitor), but not swainsonine (a complex-type N-glycan biosynthesis inhibitor) abrogated Gal-9-mediated inhibition of Th17 development indicating that there is a linkage between Gal-9 and an unidentified glycoprotein(s) with O-linked ß-galactosides that suppress Th17 development.

Galectin-9 signaling prolongs survival in murine lung-cancer by inducing macrophages to differentiate into plasmacytoid dendritic cell-like macrophages.

Galectin-9 (Gal-9) expanded plasmacytoid dendritic cell-like macrophages (pDC-Mϕs) in lung cancer-bearing mice and prolonged the survival. Gal-9 increased the frequency of CD11c(high) cells in M-CSF- but not GM-CSF-induced Mϕs in vitro in a Tim-3 independent manner. CD11c(high) cells differentiated with M-CSF+Gal-9 expressed pDC-Mϕ markers, such as PDCA-1 and F4/80. These cells expressed high TLR7, TLR8 and TLR9, although they exhibited decreased IFN-α mRNA levels. LPS or LLC stimulation further elevated pDC-Mϕ markers, indicating that M-CSF+Gal-9-induced Mϕs were pDC-Mϕ precursors. Moreover, LPS stimulation resulted in the increased IRF7 and E2-2 levels, suggesting that the pDC-Mϕ precursors matured into pDC-Mϕs. These matured pDC-Mϕs augmented NK cell-mediated cytotoxicity though they did not produce IFN-α upon TLR7 or TLR9 stimulation. The present results suggest that Gal-9 induces Mϕs to differentiate to pDC-Mϕs, and that this switch in differentiation favors the activation of NK cells that are able to prolong the survival of tumor-bearing mice.

Galectin-9 attenuates acute lung injury by expanding CD14- plasmacytoid dendritic cell-like macrophages.

RATIONALE: Galectin (Gal)-9 plays a crucial role in the modulation of innate and adaptive immunity. OBJECTIVES: To investigate whether Gal-9 plays a role in a murine acute lung injury (ALI) model. METHODS: C57BL/6 mice were pretreated with Gal-9 by subcutaneous injection 24 and 48 hours before intranasal LPS inoculation. MEASUREMENTS AND MAIN RESULTS: Gal-9 suppressed pathological changes of ALI induced by LPS. Gal-9 reduced levels of proinflammatory cytokines and chemokines, such as tumor necrosis factor (TNF)-α, IL-1ß, IL-6, and keratinocyte-derived cytokine; decreased neutrophils; and increased IL-10 and CD11b(+)Gr-1(+) macrophages in the bronchoalveolar lavage fluid of ALI mice. In Gal-9-deficient mice, pathological changes of ALI were exaggerated, and the number of neutrophils and the TNF-α level were increased. CD11b(+)Gr-1(+) cells were increased in the spleen of both Gal-9-treated and phosphate-buffered saline (PBS)-treated ALI mice, but only Gal-9 increased the ability of CCR2-expressing macrophages to migrate toward monocyte chemoattractant protein-1. Transfer of CD11b(+)Gr-1(+) macrophages obtained from Gal-9-treated mice ameliorated ALI. CD11b(+)Gr-1(+) macrophages obtained from Gal-9-treated but not PBS-treated mice suppressed TNF-α and keratinocyte-derived cytokine production from LPS-stimulated macrophages, and down-regulated Toll-like receptor-4 (TLR4) and TLR2 expression on thioglycollate-elicited macrophages. Fluorescence-activated cell-sorting analysis revealed that CD14 is negligible on CD11b(+)Gr-1(+) macrophages obtained from Gal-9-treated mice, although those from both groups resembled plasmacytoid dendritic cells (pDCs). Gal-9 down-regulated CD14 on pDC-like macrophages from PBS-treated mice independently of Gal-9/Tim-3 (T-cell immunoglobulin- and mucin domain-containing molecule-3) interaction, resulting in the acquisition of suppressive function, suggesting that the loss of CD14 by Gal-9 is critical for the suppression of pDC-like macrophages. CONCLUSIONS: Gal-9 attenuates ALI by expanding CD14(-)CD11b(+)Gr-1(+) pDC-like macrophages by preferentially suppressing macrophage functions to release proinflammatory cytokines through TLR4 and TLR2 down-regulation.

Hydrolyzed Konjac glucomannan suppresses IgE production in mice B cells.

BACKGROUND: Oral administration of pulverized Konjac glucomannan (KGM) reduces increased plasma IgE and the amount of epsilon-germline transcript (epsilonGT) in the spleen, as well as preventing the development of dermatitis in mice. To elucidate the mechanism of action of pulverized KGM, we solubilized KGM and studied its effect on IgE in vitro and in vivo. METHODS: Solubilized KGM was prepared by acid hydrolysis, and we analyzed the effective molecular size for the suppression of IgE production and epsilonGT in vitro and the level of plasma IgE induced by immunization with ovalbumin in BALB/c mice. RESULTS: The production of IgE and epsilonGT in splenic cells, but not purified B cells, was inhibited by hydrolyzed KGM (KGM hydrolyzed with 0.25 N HCl; H-KGM) at the optimal size of between 10 and 500 kDa. However, no effect was observed when H-KGM was substituted with unhydrolyzed KGM in vitro. IgE production from purified B cells cocultured with purified monocytes, but not with purified T cells, was inhibited by H-KGM. The release of IFNgamma in cultures of monocytes but in purified B cells with or without T cells was enhanced in the presence of H-KGM. Injection of mice with H-KGM also suppressed the production of plasma IgE and IgG1 but not IgG2a in vivo. CONCLUSION: KGM at an optimal size prevents germline class-switching and IgE production both in vitro and in vivo. H-KGM may be useful as a tool to study the mechanism of action of KGM and as a dietary supplement to prevent atopic diseases.

Galectin-9 expands immunosuppressive macrophages to ameliorate T-cell-mediated lung inflammation.

Galectin-9 (Gal-9) plays pivotal roles in the modulation of innate and adaptive immunity to suppress T-cell-mediated autoimmune models. However, it remains unclear if Gal-9 plays a suppressive role for T-cell function in non-autoimmune disease models. We assessed the effects of Gal-9 on experimental hypersensitivity pneumonitis induced by Trichosporon asahii. When Gal-9 was given subcutaneously to C57BL/6 mice at the time of challenge with T. asahii, it significantly suppressed T. asahii-induced lung inflammation, as the levels of IL-1, IL-6, IFN-gamma, and IL-17 were significantly reduced in the BALF of Gal-9-treated mice. Moreover, co-culture of anti-CD3-stimulated CD4 T cells with BALF cells harvested from Gal-9-treated mice on day 1 resulted in diminished CD4 T-cell proliferation and decreased levels of IFN-gamma and IL-17. CD11b(+)Ly-6C(high)F4/80(+) BALF Mphi expanded by Gal-9 were responsible for the suppression. We further found in vitro that Gal-9, only in the presence of T. asahii, expands CD11b(+)Ly-6C(high)F4/80(+) cells from BM cells, and the cells suppress T-cell proliferation and IFN-gamma and IL-17 production. The present results indicate that Gal-9 expands immunosuppressive CD11b(+)Ly-6C(high) Mphi to ameliorate Th1/Th17 cell-mediated hypersensitivity pneumonitis.

Galectin-9 ameliorates immune complex-induced arthritis by regulating Fc gamma R expression on macrophages.

Galectin-9 up-regulated Fc gamma RIIb expression of mouse peritoneal macrophages in vitro but down-regulated Fc gamma RIII expression. Galectin-9-treated macrophages stimulated with immune complexes (IC) produced less TNFalpha and IL-1 beta but more IL-10 than PBS-treated macrophages. Macrophage enhancing effects on IC-induced C5a and neutrophil chemotactic activity were also diminished for galectin-9-treated macrophages. In galectin-9-treated mice, the severity of IC-induced arthritis was reduced, as were pro-inflammatory cytokine levels in inflamed joints and serum C5a. Fc gamma RIIb expression of macrophages from galectin-9-treated mice was up-regulated, whereas Fc gamma RIII expression was down-regulated. Macrophages from galectin-9-treated mice produced less TNFalpha and IL-1 beta but more IL-10 than PBS-treated mice. Disease severity of galectin-9-transgenic mice was milder than wild-type mice, whereas that of galectin-9-deficient mice was exaggerated. Furthermore, macrophage Fc gamma RIIb expression in galectin-9-deficient mice was down-regulated, while Fc gamma RIII expression was up-regulated. These results suggest that galectin-9 suppresses IC-induced inflammation partly by regulating Fc gamma R expression on macrophages.

Peritoneal injection of fucoidan suppresses the increase of plasma IgE induced by OVA-sensitization.

We previously reported that fucoidan, a dietary fiber purified from seaweed, inhibited IgE production by B cells in vitro. In this study, we examined the effect of fucoidan on IgE production in vivo. The OVA-induced increase of plasma IgE was significantly suppressed when fucoidan was intraperitoneally, but not orally, administered prior to the first immunization with OVA. The production of IL-4 and IFN-gamma in response to OVA in spleen cells isolated from OVA-sensitized mice treated with fucoidan in vivo was lower than that from mice treated without fucoidan. Moreover, the flow cytometric analysis and ELISpot assay revealed that the administration of fucoidan suppressed a number of IgE-expressing and IgE-secreting B cells, respectively. These results indicate that fucoidan inhibits the increase of plasma IgE through the suppression of IgE-producing B cell population, and the effect of fucoidan in vivo is crucially dependent on the route and timing of its administration.

Galectin-9 accelerates transforming growth factor beta3-induced differentiation of human mesenchymal stem cells to chondrocytes.

Galectin-9 (Gal-9), a beta-galactoside binding lectin, plays a crucial role in innate and adaptive immunity. In the rat collagen-induced arthritis model, administration of Gal-9 induced repair of existing cartilage injury even when joints were already swollen with cartilage destruction. We thus attempted to explore the role of Gal-9 in chondrocyte differentiation utilizing human mesenchymal stem cell (MSC) pellet cultures. During chondrogenesis induced by transforming growth factor beta3 (TGFbeta3), MSCs strongly expressed endogenous Gal-9. Expression of Gal-9 peaked on day 14 and the neutralization of endogenous Gal-9 resulted in the reduced chondrogenesis, indicating possible involvement of Gal-9 in TGFbeta-mediated chondrogenesis. In pellets, addition of Gal-9 significantly enhanced TGFbeta3-induced chondrogenesis, as evidenced by increasing proteoglycan content, but not cell proliferation. In the absence of Gal-9, collagen expression by MSCs switched from type I to type II on 28 days after stimulation with TGFbeta3. When MSCs were co-stimulated with Gal-9, the class switch occurred on day 21. In addition, Gal-9 synergistically enhanced TGFbeta3-induced phosphorylation of Smad2, though Gal-9 did not itself induce detectable Smad2 phosphorylation. These results suggest that Gal-9 has a beneficial effect on cartilage repair in injured joints by induction of differentiation of MSCs into chondrocytes.

Galectin-9 expands unique macrophages exhibiting plasmacytoid dendritic cell-like phenotypes that activate NK cells in tumor-bearing mice.

Galectin-9 (Gal-9) inhibits the metastasis of tumor cells by blocking their adhesion to endothelium and the extracellular matrix. In this study, we addressed the involvement of Gal-9 in anti-tumor activity. Gal-9 significantly prolonged the survival of B16F10 melanoma-bearing mice. Gal-9 increased the numbers of NK cells, CD8 T cells and macrophages in tumor-bearing mice. Gal-9-mediated anti-tumor activity was not induced in NK cell-, macrophage- and CD8 T cell-depleted mice. NK cells from Gal-9-treated mice, compared to PBS-treated mice, exhibited significantly higher cytolytic activity. Co-culture of naïve NK cells with macrophages from Gal-9-treated mice resulted in enhanced NK activity, although Gal-9 itself did not enhance the NK activity. We also found that Ly-6C(+)CD11b(+)F4/80(+) macrophages with plasmacytoid cell (pDC)-like phenotypes (PDCA-1 and B220) were responsible for the enhanced NK activity. These results provide evidence that Gal-9 promotes NK cell-mediated anti-tumor activity by expanding unique macrophages with a pDC-like phenotype.

Galectin-9 increases Tim-3+ dendritic cells and CD8+ T cells and enhances antitumor immunity via galectin-9-Tim-3 interactions.

A Tim-3 ligand, galectin-9 (Gal-9), modulates various functions of innate and adaptive immune responses. In this study, we demonstrate that Gal-9 prolongs the survival of Meth-A tumor-bearing mice in a dose- and time-dependent manner. Although Gal-9 did not prolong the survival of tumor-bearing nude mice, transfer of naive spleen cells restored a prolonged Gal-9-induced survival in nude mice, indicating possible involvement of T cell-mediated immune responses in Gal-9-mediated antitumor activity. Gal-9 administration increased the number of IFN-gamma-producing Tim-3(+) CD8(+) T cells with enhanced granzyme B and perforin expression, although it induced CD4(+) T cell apoptosis. It simultaneously increased the number of Tim-3(+)CD86(+) mature dendritic cells (DCs) in vivo and in vitro. Coculture of CD8(+) T cells with DCs from Gal-9-treated mice increased the number of IFN-gamma producing cells and IFN-gamma production. Depletion of Tim-3(+) DCs from DCs of Gal-9-treated tumor-bearing mice decreased the number of IFN-gamma-producing CD8(+) T cells. Such DC activity was significantly abrogated by Tim-3-Ig, suggesting that Gal-9 potentiates CD8(+) T cell-mediated antitumor immunity via Gal-9-Tim-3 interactions between DCs and CD8(+) T cells.

Galectin-9 suppresses tumor metastasis by blocking adhesion to endothelium and extracellular matrices.

We previously described an inverse correlation between galectin-9 (Gal-9) expression and metastasis in patients with malignant melanoma and breast cancer. This study verified the ability of Gal-9 to inhibit lung metastasis in experimental mouse models using highly metastatic B16F10 melanoma and Colon26 colon cancer cells. B16F10 cells transfected with a secreted form of Gal-9 lost their metastatic potential. Intravenous Gal-9 administration reduced the number of metastases of both B16F10 and Colon26 cells in the lung, indicating that secreted Gal-9 suppresses metastasis. Analysis of adhesive molecule expression revealed that B16F10 cells highly express CD44, integrin alpha1, alpha 4, alpha V, and beta1, and that Colon26 cells express CD44, integrin alpha2, alpha 5, alpha V, and beta1, suggesting that Gal-9 may inhibit the adhesion of tumor cells to vascular endothelium and the extracellular matrix (ECM) by binding to such adhesive molecules. Indeed, Gal-9 suppressed the binding of hyaluronic acid to CD44 on both B16F10 and Colon26 cells, and also suppressed the binding of vascular cell adhesion molecule-1 to very late antigen-4 on B16F10 cells. Furthermore, Gal-9 inhibited the binding of tumor cells to ECM components, resulting in the suppression of tumor cell migration. The present results suggest that Gal-9 suppresses both attachment and invasion of tumor cells by inhibiting the binding of adhesive molecules on tumor cells to ligands on vascular endothelium and ECM.

Galectin-9 suppresses the generation of Th17, promotes the induction of regulatory T cells, and regulates experimental autoimmune arthritis.

The effects of galectin-9 on a mouse collagen-induced arthritis (CIA) model were assessed to clarify whether galectin-9 suppresses CIA by regulating T cell immune responses. Galectin-9 suppressed CIA in a dose-dependent manner, and such suppression was observed even when treatment was started on 7 days after the booster, indicating its preventive and therapeutic effects. Galectin-9 induced the decreased levels of pro-inflammatory cytokines, IL-17, IL-12, and IFNgamma in the joint. Galectin-9 induced the decreased number of CD4(+) TIM-3(+) T cells in peripheral blood. Galectin-9-deficient mice became susceptible to CIA may be by increased number of CD4(+) TIM-3(+) T cells and decreased number of Treg cells. We further found that galectin-9 induces differentiation of naive T cells to Treg cells, and it suppresses differentiation to Th17 cells in vitro. The present results suggested that galectin-9 ameliorates CIA by suppressing the generation of Th17, promoting the induction of regulatory T cells.

Galectin-9 expression links to malignant potential of cervical squamous cell carcinoma.

PURPOSE: Galectin-9 (Gal-9) induces adhesion and aggregation of certain cell types and can be a prognostic factor in the patients with melanoma and breast cancer. We assessed the experiments to resolve whether Gal-9 expression in cervical neoplasm links to malignant potential of cervical squamous cell carcinoma (SCC) cells. METHODS: Gal-9 expression was examined with immunohistochemical techniques in 23 normal cervical squamous epithelia, 17 cervical intraepithelial neoplasia (CIN), and 38 cervical SCC compared to E-cadherin. CIN was divided into low-grade and high-grade squamous intraepithelial lesions (8 LSIL and 9 HSIL), and SCC was into well-, moderately and poorly differentiated SCC (6 WSCC, 20 MSCC and 12 PSCC). RESULTS: Gal-9 and E-cadherin were evidently detected in normal epithelium and endocervical glands, but those in CIN and SCC were significantly faint. Moreover, both the Gal-9 and E-cadherin expressions in HSIL were significant lower than those in LSIL, suggesting their association with malignant transformation. Unexpectedly, Gal-9 and E-cadherin in WSCC were significantly high compared to those in HSIL. Furthermore, those in SCC were inversely correlated with the grade of differentiation (WSCC >> MSCC >> PSCC), implying the possible involvement of Gal-9 and E-cadherin in the differentiation of SCC. In contrast, they were not different among the FIGO stage. Gal-9 expression was well correlated with E-cadherin expression in CIN and SCC but not in normal cervical epithelia. CONCLUSION: The present results suggest that decreased Gal-9 expression is inversely associated with malignant potential or differentiation of cervical CIN and SCC as a differentiation biomarker.

Beneficial effect of galectin 9 on rheumatoid arthritis by induction of apoptosis of synovial fibroblasts.

OBJECTIVE: To compare the expression of galectin 9 (Gal-9) in synovial tissue (ST) from rheumatoid arthritis (RA) patients and osteoarthritis (OA) patients and to evaluate the effects of Gal-9 on fibroblast-like synoviocytes (FLS) in these patients. METHODS: The expression of Gal-9 in ST and FLS was compared using immunohistochemical techniques. Apoptotic cells in RA and OA ST samples were detected by TUNEL assay. Apoptosis of FLS was analyzed by the sub-G(1) method in vitro. The in vivo suppressive effects of Gal-9 on collagen-induced arthritis (CIA) in a mouse model were also elucidated. RESULTS: The percentage of Gal-9-positive cells in ST samples and the amount of Gal-9 in synovial fluid samples were significantly higher in patients with RA than in patients with OA, suggesting the involvement of Gal-9 in the development of RA. Compared with the 2 wild-type Gal-9 forms, stable Gal-9, a mutant protein resistant to proteolysis, significantly induced apoptosis of FLS from RA patients. In contrast, other galectins, such as Gal-1, Gal-3, and Gal-8, did not induce apoptosis or suppress the proliferation of human RA FLS. Stable Gal-9 preferentially induced apoptosis and suppressed the proliferation of RA FLS in vitro. It also induced apoptosis of cells in RA ST implanted into SCID mice in vivo. In a mouse model of CIA, apoptotic cells were detected in the joints of stable Gal-9-treated mice, but not phosphate buffered saline-treated mice, and suppressed CIA characterized by pannus formation with inflammatory cell infiltration and bone/cartilage destruction. CONCLUSION: Gal-9-induced apoptosis of hyperproliferative RA FLS may play a critical role in the suppression of RA.