High expression of B3GALT5 suppresses the galectin-4-mediated peritoneal dissemination of poorly differentiated gastric cancer cells.

Peritoneal metastasis frequently accompanies metastatic and/or recurrent gastric cancer, leading to a poor prognosis owing to a lack of effective treatment. Hence, there is a pressing need to enhance our understanding of the mechanisms and molecules driving peritoneal metastasis. In a previous study, galectin-4 inhibition impeded peritoneal metastasis in a murine model. This study examined the glycan profiles of cell surface proteins and glycosphingolipids (GSLs) in cells with varying tumorigenic potentials to understand the intricate mechanisms underlying galectin-4-mediated regulation, particularly glycosylation. Detailed mass spectrometry analysis showed that galectin-4 knockout cells exhibit increased expression of lacto-series GSLs with ß1,3-linked galactose while showing no significant alterations in neolacto-series GSLs. We conducted real-time polymerase chain reaction (PCR) analysis to identify candidate glycosyltransferases that synthesize increased levels of GSLs. Subsequently, we introduced the candidate B3GALT5 gene and selected the clones with high expression levels. B3GALT5 gene-expressing clones showed GSL glycan profiles like those of knockout cells and significantly reduced tumorigenic ability in mouse models. These clones exhibited diminished proliferative capacity and showed reduced expression of galectin-4 and activated AKT. Moreover, co-localization of galectin-4 with flotillin-2 (a raft marker) decreased in B3GALT5-expressing cells, implicating GSLs in galectin-4 localization to lipid rafts. D-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (a GSL synthase inhibitor) also affected galectin-4 localization in rafts, suggesting the involvement of GSL microdomains. We discovered that B3GALT5 plays a crucial role in regulating peritoneal metastasis of malignant gastric cancer cells by suppressing cell proliferation and modulating lipid rafts and galectin-4 via mechanisms that are yet to be elucidated.

A comprehensive study on the multifunctional properties of galectin-4 in red-lip mullet (Planiliza haematocheilus): Insights into molecular interactions, antimicrobial defense, and cell proliferation.

Galectin-4 belongs to the galactoside-binding protein family and is a type of tandem repeat galectin. Despite previous studies indicating its importance in fish immunology, a comprehensive investigation is necessary to fully understand its role in immunomodulatory functions and cellular dynamics. Therefore, this study aimed to explore the immunomodulatory functions of galectin-4 with a particular focus on its antimicrobial and cellular proliferative properties. The open reading frame of PhGal4 spans 1092 base pairs and encodes a soluble protein of 363 amino acids with a theoretical isoelectric point (IEP) of 6.39 and a molecular weight of 39.411 kDa. Spatial expression analysis under normal physiological conditions revealed ubiquitous expression of PhGal4 across all examined tissues, with the highest level observed in intestinal tissue. Upon stimulation with poly I:C, LPS, and L. garvieae, a significant increase (p < 0.05) in PhGal4 expression was observed in both blood and spleen tissues. Subsequent subcellular localization assay demonstrated that PhGal4 was predominantly localized in the cytoplasm. The recombinant PhGal4 (rPhGal4) exhibited specific binding capabilities to pathogen-associated molecular patterns (PAMPs), including LPS and peptidoglycan, but not poly I:C. The rPhGal4 negatively affected the bacterial growth kinetics. Additionally, rPhGal4 demonstrated complete hemagglutination of fish erythrocytes, which could be inhibited by the presence of D-galactose and α-lactose. The overexpression of PhGal4 in FHM epithelial cells demonstrated a significant suppression of viral replication during VHSV infection. Furthermore, the in vitro scratch assay and WST-1 assay demonstrated a wound healing effect of PhGal4 overexpression in FHM cells, potentially achieved through the promotion of cell proliferation by activating genes involved in cell cycle regulation. In conclusion, the responsive expression to immune stimuli, antimicrobial properties, and cell proliferation promotion of PhGal4 suggest that it plays a crucial role in immunomodulation and cellular dynamics of red-lip mullet. The findings in this study shed light on the multifunctional nature of galectin-4 in teleost fish.

Exploring galectin interactions with human milk oligosaccharides and blood group antigens identifies BGA6 as a functional galectin-4 ligand.

Galectins (Gals), a family of multifunctional glycan-binding proteins, have been traditionally defined as ß-galactoside binding lectins. However, certain members of this family have shown selective affinity toward specific glycan structures including human milk oligosaccharides (HMOs) and blood group antigens. In this work, we explored the affinity of human galectins (particularly Gal-1, -3, -4, -7, and -12) toward a panel of oligosaccharides including HMOs and blood group antigens using a complementary approach based on both experimental and computational techniques. While prototype Gal-1 and Gal-7 exhibited differential affinity for type I versus type II Lac/LacNAc residues and recognized fucosylated neutral glycans, chimera-type Gal-3 showed high binding affinity toward poly-LacNAc structures including LNnH and LNnO. Notably, the tandem-repeat human Gal-12 showed preferential recognition of 3-fucosylated glycans, a unique feature among members of the galectin family. Finally, Gal-4 presented a distinctive glycan-binding activity characterized by preferential recognition of specific blood group antigens, also validated by saturation transfer difference nuclear magnetic resonance experiments. Particularly, we identified oligosaccharide blood group A antigen tetraose 6 (BGA6) as a biologically relevant Gal-4 ligand, which specifically inhibited interleukin-6 secretion induced by this lectin on human peripheral blood mononuclear cells. These findings highlight unique determinants underlying specific recognition of HMOs and blood group antigens by human galectins, emphasizing the biological relevance of Gal-4-BGA6 interactions, with critical implications in the development and regulation of inflammatory responses.

Different roles of the heterodimer architecture of galectin-4 in selective recognition of oligosaccharides and lipopolysaccharides having ABH antigens.

The dimeric architecture of tandem-repeat type galectins, such as galectin-4 (Gal-4), modulates their biological activities, although the underlying molecular mechanisms have remained elusive. Emerging evidence show that tandem-repeat galectins play an important role in innate immunity by recognizing carbohydrate antigens present on the surface of certain pathogens, which very often mimic the structures of the human self-glycan antigens. Herein, we have analyzed the binding preferences of the C-domain of Gal-4 (Gal-4C) toward the ABH-carbohydrate histo-blood antigens with different core presentations and their recognition features have been rationalized by using a combined experimental approach including NMR, solid-phase and hemagglutination assays, and molecular modeling. The data show that Gal-4C prefers A over B antigens (two-fold in affinity), contrary to the N-domain (Gal-4N), although both domains share the same preference for the type-6 presentations. The behavior of the full-length Gal-4 (Gal-4FL) tandem-repeat form has been additionally scrutinized. Isothermal titration calorimetry and NMR data demonstrate that both domains within full-length Gal-4 bind to the histo-blood antigens independently of each other, with no communication between them. In this context, the heterodimeric architecture does not play any major role, apart from the complementary A and B antigen binding preferences. However, upon binding to a bacterial lipopolysaccharide containing a multivalent version of an H-antigen mimetic as O-antigen, the significance of the galectin architecture was revealed. Indeed, our data point to the linker peptide domain and the F-face of the C-domain as key elements that provide Gal-4 with the ability to cross-link multivalent ligands, beyond the glycan binding capacity of the dimer.

Galectin-4 Antimicrobial Activity Primarily Occurs Through its C-Terminal Domain.

Although immune tolerance evolved to reduce reactivity with self, it creates a gap in the adaptive immune response against microbes that decorate themselves in self-like antigens. This is particularly apparent with carbohydrate-based blood group antigens, wherein microbes can envelope themselves in blood group structures similar to human cells. In this study, we demonstrate that the innate immune lectin, galectin-4 (Gal-4), exhibits strain-specific binding and killing behavior towards microbes that display blood group-like antigens. Examination of binding preferences using a combination of microarrays populated with ABO(H) glycans and a variety of microbial strains, including those that express blood group-like antigens, demonstrated that Gal-4 binds mammalian and microbial antigens that have features of blood group and mammalian-like structures. Although Gal-4 was thought to exist as a monomer that achieves functional bivalency through its two linked carbohydrate recognition domains, our data demonstrate that Gal-4 forms dimers and that differences in the intrinsic ability of each domain to dimerize likely influences binding affinity. While each Gal-4 domain exhibited blood group-binding activity, the C-terminal domain (Gal-4C) exhibited dimeric properties, while the N-terminal domain (Gal-4N) failed to similarly display dimeric activity. Gal-4C not only exhibited the ability to dimerize but also possessed higher affinity toward ABO(H) blood group antigens and microbes expressing glycans with blood group-like features. Furthermore, when compared to Gal-4N, Gal-4C exhibited more potent antimicrobial activity. Even in the context of the full-length protein, where Gal-4N is functionally bivalent by virtue of Gal-4C dimerization, Gal-4C continued to display higher antimicrobial activity. These results demonstrate that Gal-4 exists as a dimer and exhibits its antimicrobial activity primarily through its C-terminal domain. In doing so, these data provide important insight into key features of Gal-4 responsible for its innate immune activity against molecular mimicry.

Interleukin-30 subverts prostate cancer-endothelium crosstalk by fostering angiogenesis and activating immunoregulatory and oncogenic signaling pathways.

BACKGROUND: Cancer-endothelial interplay is crucial for tumor behavior, yet the molecular mechanisms involved are largely unknown. Interleukin(IL)-30, which is expressed as a membrane-anchored cytokine by human prostate cancer (PC) cells, promotes PC vascularization and progression, but the underlying mechanisms have yet to be fully explored. METHODS: PC-endothelial cell (EC) interactions were investigated, after coculture, by flow cytometry, transcriptional profiling, western blot, and ELISA assays. Proteome profiler phospho-kinase array unveiled the molecular pathways involved. The role of tumor-derived IL30 on the endothelium's capacity to generate autocrine circuits and vascular budding was determined following IL30 overexpression, by gene transfection, or its deletion by CRISPR/Cas9 genome editing. Clinical value of the experimental findings was determined through immunopathological study of experimental and patient-derived PC samples, and bioinformatics of gene expression profiles from PC patients. RESULTS: Contact with PC cells favors EC proliferation and production of angiogenic and angiocrine factors, which are boosted by PC expression of IL30, that feeds autocrine loops, mediated by IGF1, EDN1, ANG and CXCL10, and promotes vascular budding and inflammation, via phosphorylation of multiple signaling proteins, such as Src, Yes, STAT3, STAT6, RSK1/2, c-Jun, AKT and, primarily CREB, GSK-3α/ß, HSP60 and p53. Deletion of the IL30 gene in PC cells inhibits endothelial expression of IGF1, EDN1, ANG and CXCL10 and substantially impairs tumor angiogenesis. In its interaction with IL30-overexpressing PC cells the endothelium boosts their expression of a wide range of immunity regulatory genes, including CCL28, CCL4, CCL5, CCR2, CCR7, CXCR4, IL10, IL13, IL17A, FASLG, IDO1, KITLG, TNFA, TNFSF10 and PDCD1, and cancer driver genes, including BCL2, CCND2, EGR3, IL6, VEGFA, KLK3, PTGS1, LGALS4, GNRH1 and SHBG. Immunopathological analyses of PC xenografts and in silico investigation of 1116 PC cases, from the Prostate Cancer Transcriptome Atlas, confirmed the correlation between the expression of IL30 and that of both pro-inflammatory genes, NOS2, TNFA, CXCR5 and IL12B, and cancer driver genes, LGALS4, GNRH1 and SHBG, which was validated in a cohort of 80 PC patients. CONCLUSIONS: IL30 regulates the crosstalk between PC and EC and reshapes their transcriptional profiles, triggering angiogenic, immunoregulatory and oncogenic gene expression programs. These findings highlight the angiostatic and oncostatic efficacy of targeting IL30 to fight PC.

Galectin-4 is associated with diabetes and obesity in a heart failure population.

An association between high Galectin-4 (Gal-4) and prevalence of diabetes in subjects with heart failure (HF) has previously been reported. The purpose of this study was to confirm these findings, as well as to further investigate this association, in a Swedish HF population. In addition, a second aim was to explore Gal-4's association with obesity and biomarkers of metabolism and heart failure. Gal-4 was measured using a proximity extension array technique in 324 hospitalized HF patients within the Swedish HeArt and bRain failure investigation trial cohort. Obesity was defined as BMI ≥ 30. Multivariable logistic regression models were used to explore associations between Gal-4 and diabetes/obesity, and linear regression models were used to explore the associations between Gal-4 and biomarkers. A total of 309 participants (29.1% female; mean age 74.8 years) provided complete data for the analysis of associations between Gal-4 and diabetes. Additionally, for the analysis of heart failure phenotype, complete data was available for 230 subjects. Gal-4 was positively associated with prevalent diabetes (OR 2.60; CI 95% 1.56-4.32). In multivariable models, Gal-4 levels were significantly associated with obesity, but only for subjects with diabetes (OR 2.48; 1.09-5.62). Additionally, Gal-4 demonstrated a significant association with the incretin Glucose-dependent insulinotropic polypeptide (GIP), as well as with biomarkers of HF. In the stratified analyses, the association between Gal-4 and diabetes was prominent in patients with reduced ejection fraction (n = 160, OR 3.26; 95%CI 1.88-5.66), while it was not observed in those without (n = 70, 1.96 (0.75-5.10)). In this cross-sectional, observational study, higher Gal-4 levels in HF patients were associated with higher GIP levels. Further, increased levels of Gal-4 were associated with increased likelihood of diabetes, and obesity. This association was particularly pronounced in individuals with HF characterized by reduced ejection fraction. Additionally, Gal-4 levels were significantly elevated in heart failure patients with diabetes and obesity.

Galectin-4 Is Involved in the Structural Changes of Glycosphingolipid Glycans in Poorly Differentiated Gastric Cancer Cells with High Metastatic Potential.

Gastric cancer with peritoneal dissemination is difficult to treat surgically, and frequently recurs and metastasizes. Currently, there is no effective treatment for this disease, and there is an urgent need to elucidate the molecular mechanisms underlying peritoneal dissemination and metastasis. Our previous study demonstrated that galectin-4 participates in the peritoneal dissemination of poorly differentiated gastric cancer cells. In this study, the glycan profiles of cell surface proteins and glycosphingolipids (GSLs) of the original (wild), galectin-4 knockout (KO), and rescue cells were investigated to understand the precise mechanisms involved in the galectin-4-mediated regulation of associated molecules, especially with respect to glycosylation. Glycan analysis of the NUGC4 wild type and galectin-4 KO clones with and without peritoneal metastasis revealed a marked structural change in the glycans of neutral GSLs, but not in N-glycan. Furthermore, mass spectrometry (MS) combined with glycosidase digestion revealed that this structural change was due to the presence of the lacto-type (ß1-3Galactosyl) glycan of GSL, in addition to the neolacto-type (ß1-4Galactosyl) glycan of GSL. Our results demonstrate that galectin-4 is an important regulator of glycosylation in cancer cells and galectin-4 expression affects the glycan profile of GSLs in malignant cancer cells with a high potential for peritoneal dissemination.

TGM2, HMGA2, FXYD3, and LGALS4 genes as biomarkers in acquired oxaliplatin resistance of human colorectal cancer: A systems biology approach.

Acquired resistance to oxaliplatin is considered as the primary reason for failure in colorectal cancer (CRC) therapy. Identifying the underlying resistance mechanisms may improve CRC treatment. The present study aims to identify the key genes involved in acquired oxaliplatin-resistant in CRC by confirming the oxaliplatin resistance index (OX-RI). To this aim, two public microarray datasets regarding oxaliplatin-resistant CRC cells with different OX-RI, GSE42387, and GSE76092 were downloaded from GEO database to identify differentially expressed genes (DEGs). The results indicated that the OX-RI affects the gene expression pattern significantly. Then, 54 common DEGs in both datasets including 18 up- and 36 down-regulated genes were identified. Protein-protein interaction (PPI) analysis revealed 13 up- (MAGEA6, TGM2, MAGEA4, SCHIP1, ECI2, CD33, AKAP12, MAGEA12, CALD1, WFDC2, VSNL1, HMGA2, and MAGEA2B) and 12 down-regulated (PDZK1IP1, FXYD3, ALDH2, CEACAM6, QPRT, GRB10, TM4SF4, LGALS4, ALDH3A1, USH1C, KCNE3, and CA12) hub genes. In the next step, two novel up-regulated hub genes including ECI2 and SCHIP1 were identified to be related to oxaliplatin resistance. Functional enrichment and pathway analysis indicated that metabolic pathways, proliferation, and epithelial-mesenchymal transition may play dominant roles in CRC progression and oxaliplatin resistance. In the next procedure, two in vitro oxaliplatin-resistant sub-lines including HCT116/OX-R4.3 and HCT116/OX-R10 cells with OX-IR 3.93 and 10.06 were established, respectively. The results indicated the up-regulation of TGM2 and HMGA2 in HCT116/OX-R10 cells with high OX-RI and down-regulation of FXYD3, LGALS4, and ECI2 in both cell types. Based on the results, TGM2, HMGA2, FXYD3, and LGALS4 genes are related to oxaliplatin-resistant CRC and may serve as novel therapeutic targets.

Plasma Galectin-4 Levels Are Increased after Stroke in Mice and Humans.

Epidemiological studies have associated plasma galectin-4 (Gal-4) levels with prevalent and incident diabetes, and with an increased risk of coronary artery disease. To date, data regarding possible associations between plasma Gal-4 and stroke are lacking. Using linear and logistic regression analyses, we tested Gal-4 association with prevalent stroke in a population-based cohort. Additionally, in mice fed a high-fat diet (HFD), we investigated whether plasma Gal-4 increases in response to ischemic stroke. Plasma Gal-4 was higher in subjects with prevalent ischemic stroke, and was associated with prevalent ischemic stroke (odds ratio 1.52; 95% confidence interval 1.01-2.30; p = 0.048) adjusted for age, sex, and covariates of cardiometabolic health. Plasma Gal-4 increased after experimental stroke in both controls and HFD-fed mice. HFD exposure was devoid of impact on Gal-4 levels. This study demonstrates higher plasma Gal-4 levels in both experimental stroke and in humans that experienced ischemic stroke.

Oligosaccharide Ligands of Galectin-4 and Its Subunits: Multivalency Scores Highly.

Galectins are carbohydrate-binding lectins that modulate the proliferation, apoptosis, adhesion, or migration of cells by cross-linking glycans on cell membranes or extracellular matrix components. Galectin-4 (Gal-4) is a tandem-repeat-type galectin expressed mainly in the epithelial cells of the gastrointestinal tract. It consists of an N- and a C-terminal carbohydrate-binding domain (CRD), each with distinct binding affinities, interconnected with a peptide linker. Compared to other more abundant galectins, the knowledge of the pathophysiology of Gal-4 is sparse. Its altered expression in tumor tissue is associated with, for example, colon, colorectal, and liver cancers, and it increases in tumor progression, and metastasis. There is also very limited information on the preferences of Gal-4 for its carbohydrate ligands, particularly with respect to Gal-4 subunits. Similarly, there is virtually no information on the interaction of Gal-4 with multivalent ligands. This work shows the expression and purification of Gal-4 and its subunits and presents a structure-affinity relationship study with a library of oligosaccharide ligands. Furthermore, the influence of multivalency is demonstrated in the interaction with a model lactosyl-decorated synthetic glycoconjugate. The present data may be used in biomedical research for the design of efficient ligands of Gal-4 with diagnostic or therapeutic potential.

Multifaceted role of galectin-4 in cancer: A systematic review.

BACKGROUND: Galectins are ß-galactoside-binding proteins. Galectin-4 has shown an effect on cancer progression/metastasis, especially in cancers of the digestive system. This can be attributed to altered glycosylation pattern of cell membrane molecules, which is a characteristic attribute of oncogenesis. The aim of this paper is to systematically review galectin-4 in different cancers and its role in disease progression. METHODS: The study was designed on the basis of Preferred Reporting Items for Systematic Review and Meta-Analysis (PRISMA) guidelines. PubMed, Scopus, Web of Science, and Science Direct were used to search relevant literature with keywords "galectin-4 AND cancer", "galectin-4", "LGALS4", and "LGALS4 AND cancer". Inclusion criteria for study selection were availability of full-text articles, articles in English language and articles relevant to current topic, that is, galectin-4 and cancer. Exclusion criteria were studies that investigated other disease conditions, interventions unrelated to cancer or galectin-4 and bias outcome. RESULTS: A total of 73 articles were retrieved after removing duplication from databases, out of which 40 studies were included in the review that followed the inclusion criteria, including low to moderate bias. These included 23 studies in digestive system, 5 in reproductive system, 4 in respiratory system, and 2 in brain and urothelial cancers. CONCLUSIONS: A differential expression of galectin-4 was observed in different cancer stages/ and types. Furthermore, galectin-4 was found to modulate disease progression. A meta-analysis and comprehensive mechanistic studies, pertaining to different aspects of galectin-4 biology, could give statistically driven correlations, elucidating multifaceted role of galectin-4 in cancer.

Seven bacterial response-related genes are biomarkers for colon cancer.

BACKGROUND: Colon cancer (CC) is a common tumor that causes significant harm to human health. Bacteria play a vital role in cancer biology, particularly the biology of CC. Genes related to bacterial response were seldom used to construct prognosis models. We constructed a bacterial response-related risk model based on three Molecular Signatures Database gene sets to explore new markers for predicting CC prognosis. METHODS: The Cancer Genome Atlas (TCGA) colon adenocarcinoma samples were used as the training set, and Gene Expression Omnibus (GEO) databases were used as the test set. Differentially expressed bacterial response-related genes were identified for prognostic gene selection. Univariate Cox regression analysis, least absolute shrinkage and selection operator-penalized Cox regression analysis, and multivariate Cox regression analysis were performed to construct a prognostic risk model. The individual diagnostic effects of genes in the prognostic model were also evaluated. Moreover, differentially expressed long noncoding RNAs (lncRNAs) were identified. Finally, the expression of these genes was validated using quantitative polymerase chain reaction (qPCR) in cell lines and tissues. RESULTS: A prognostic signature was constructed based on seven bacterial response genes: LGALS4, RORC, DDIT3, NSUN5, RBCK1, RGL2, and SERPINE1. Patients were assigned a risk score based on the prognostic model, and patients in the TCGA cohort with a high risk score had a poorer prognosis than those with a low risk score; a similar finding was observed in the GEO cohort. These seven prognostic model genes were also independent diagnostic factors. Finally, qPCR validated the differential expression of the seven model genes and two coexpressed lncRNAs (C6orf223 and SLC12A9-AS1) in 27 pairs of CC and normal tissues. Differential expression of LGALS4 and NSUN5 was also verified in cell lines (FHC, COLO320DM, SW480). CONCLUSIONS: We created a seven-gene bacterial response-related gene signature that can accurately predict the outcomes of patients with CC. This model can provide valuable insights for personalized treatment.

Identification of potential biomarkers in Barrett's esophagus derived esophageal adenocarcinoma.

Almost 50% of esophageal adenocarcinoma (EAC) patients progressed from Barrett's esophagus (BE). EAC is often diagnosed at late stages and is related to dismal prognosis. However, there are still no effective methods for stratification and therapy in BE and EAC. Two public datasets (GSE26886 and GSE37200) were analyzed to identify differentially expressed genes (DEGs) between BE and EAC. Then, a series of bioinformatics analyses were performed to explore potential biomarkers associated with BE-EAC. 27 up- and 104 down-regulated genes were observed between GSE26886 and GSE37200. The GO and KEGG enrichment analysis indicated that the DEGs were highly involved in tumorigenesis. Subsequently, Weighted Gene Co-Expression Network Analysis (WGCNA) were performed to explore the potential genes related to BE-EAC, which were validated in The Cancer Genome Atlas (TCGA) database, and 5 up-regulated genes (MYO1A, ACE2, COL1A1, LGALS4, and ADRA2A) and 3 down-regulated genes (AADAC, RAB27A, and P2RY14) were found in EAC. Meanwhile, ADRA2A and AADAC could contribute to EAC pathogenesis and progression. MYO1A, ACE2, COL1A1, LGALS4, ADRA2A, AADAC, RAB27A, and P2RY14 could be potential novel diagnostic and prognostic biomarkers in BE-EAC.

Galectin-4 increases the ability of M2 macrophages to enhance antiviral CD4+ T-cell responses.

Galectin-4 (Gal-4) is a ß-galactoside-binding protein belonging to the galectin family. Although Gal-4 is known to be involved in several physiologic processes of the gastrointestinal tract, its immunomodulatory roles remain unclear. In this study, we investigated whether Gal-4 influences the function of M1 and M2 macrophages. Gal-4 treatment drove more robust changes in the gene expression of M2 macrophages compared to M1 macrophages. Antiviral immune response-related genes were significantly upregulated in Gal-4-treated M2 macrophages. Gal-4 significantly enhanced the immunostimulatory activity of M2 macrophages upon Toll-like receptor 7 stimulation or infection with lymphocytic choriomeningitis virus (LCMV). Moreover, the antibody production against LCMV infection and the antiviral CD4+ T-cell responses, but not the antiviral CD8+ T-cell responses, were greatly increased by Gal-4-treated M2 macrophages in vivo. The present results indicate that Gal-4 enhances the ability of M2 macrophages to promote antiviral CD4+ T-cell responses. Thus, Gal-4 could be used to boost antiviral immune responses.

Suppression of galectin-4 attenuates peritoneal metastasis of poorly differentiated gastric cancer cells.

BACKGROUND: Peritoneal dissemination, most often seen in metastatic and/or recurrent gastric cancer, is an inoperable condition that lacks effective treatment. The use of molecular targeted drugs is also limited; therefore, identifying novel therapeutic targets and improving our understanding of this metastatic cancer are an urgent requirement. In this study, we focused on galectin-4, which is specifically expressed in poorly differentiated cells with high potential for peritoneal dissemination. METHODS: We knocked out the galectin-4 gene in NUGC4 cells using CRISPR/Cas9-mediated genome editing. Proliferation and peritoneal cancer formation in knockout cells were compared with those in wild-type and galectin-4 re-expressing cells. Western blotting and proximity ligation assays were performed to identify associated molecules affected by the expression of galectin-4. The effect of galectin-4 knockdown on cell proliferation and peritoneal metastasis was studied using a specific siRNA. Expression of galectin-4 in peritoneal metastatic tumors from 10 patients with gastric cancer was examined by immunohistochemistry. RESULTS: Suppression of galectin-4 expression reduced proliferation and peritoneal metastasis of malignant gastric cancer cells. Galectin-4 knockout and knockdown reduced the expression of activated c-MET and CD44. Galectin-4 was found to interact with several proteins on the cell surface, including CD44 and c-MET, via its carbohydrate-binding ability. Immunohistochemistry showed galectin-4 expression in peritoneal metastatic tumor cells in all patients examined. CONCLUSIONS: We clarified the role of galectin-4 in the development of peritoneal dissemination of poorly differentiated gastric cancer cells. Our data highlight the diagnostic and therapeutic potential of galectin-4 in the peritoneal dissemination of gastric cancer.

Cytosolic galectin-4 enchains bacteria, restricts their motility, and promotes inflammasome activation in intestinal epithelial cells.

Galectin-4, a member of the galectin family of animal glycan-binding proteins (GBPs), is specifically expressed in gastrointestinal epithelial cells and is known to be able to bind microbes. However, its function in host-gut microbe interactions remains unknown. Here, we show that intracellular galectin-4 in intestinal epithelial cells (IECs) coats cytosolic Salmonella enterica serovar Worthington and induces the formation of bacterial chains and aggregates. Galectin-4 enchains bacteria during their growth by binding to the O-antigen of lipopolysaccharides. Furthermore, the binding of galectin-4 to bacterial surfaces restricts intracellular bacterial motility. Galectin-4 enhances caspase-1 activation and mature IL-18 production in infected IECs especially when autophagy is inhibited. Finally, orally administered S. enterica serovar Worthington, which is recognized by human galectin-4 but not mouse galectin-4, translocated from the intestines to mesenteric lymph nodes less effectively in human galectin-4-transgenic mice than in littermate controls. Our results suggest that galectin-4 plays an important role in host-gut microbe interactions and prevents the dissemination of pathogens. The results of the study revealed a novel mechanism of host-microbe interactions that involves the direct binding of cytosolic lectins to glycans on intracellular microbes.

Extracellular Galectin 4 Drives Immune Evasion and Promotes T-cell Apoptosis in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by rich deposits of extracellular matrix (ECM), affecting the pathophysiology of the disease. Here, we identified galectin 4 (gal 4) as a cancer cell-produced protein that was deposited into the ECM of PDAC tumors and detected high-circulating levels of gal 4 in patients with PDAC. In orthotopic transplantation experiments, we observed increased infiltration of T cells and prolonged survival in immunocompetent mice transplanted with cancer cells with reduced expression of gal 4. Increased survival was not observed in immunodeficient RAG1-/- mice, demonstrating that the effect was mediated by the adaptive immune system. By performing single-cell RNA-sequencing, we found that the myeloid compartment and cancer-associated fibroblast (CAF) subtypes were altered in the transplanted tumors. Reduced gal 4 expression associated with a higher proportion of myofibroblastic CAFs and reduced numbers of inflammatory CAFs. We also found higher proportions of M1 macrophages, T cells, and antigen-presenting dendritic cells in tumors with reduced gal 4 expression. Using a coculture system, we observed that extracellular gal 4 induced apoptosis in T cells by binding N-glycosylation residues on CD3ε/δ. Hence, we show that gal 4 is involved in immune evasion and identify gal 4 as a promising drug target for overcoming immunosuppression in PDAC.

Normal Cortical Myelination in Galectin-4-Deficient Mice.

Myelin, critical for the correct function of the nervous system, is organized in different patterns that can include long non-myelinated axonal segments. How myelin patterning is regulated remains unexplained. The carbohydrate-binding protein galectin-4 (Gal-4) influences oligodendrocyte differentiation in vitro and is associated with non-myelinable axon segments (NMS) in cultured neurons. In consequence, Gal-4 has been proposed as a myelin patterning regulator, although no in vivo studies have corroborated this hypothesis. We used Gal-4-deficient mice (Lgals4-KO) to study the role of Gal-4 in cortical myelination in vivo. We show that cultured neurons of Lgals4-KO mice form NMS that are regulated as in control neurons. In addition, oligodendrocyte/myelin markers expression measured by biochemical and immunochemical means, and cortical myelin microstructure studied by in-depth image analysis appear unaltered in these animals. Consistently, myelin displays an essentially normal function assessed by in vivo electrophysiology and locomotion analyses. In conclusion, cortical myelin of Lgals4-KO mice does not show any significant defect in composition, organization or function, pointing to a negligible role of Gal-4 in myelination in vivo or, as discussed, to unknown mechanisms that compensate its absence.

Construction and validation of an immune-related genes prognostic index (IRGPI) model in colon cancer.

Background: Though immunotherapy has become one of the standard therapies for colon cancer, the overall effective rate of immunotherapy is very low. Constructing an immune-related genes prognostic index (IRGPI) model may help to predict the response to immunotherapy and clinical outcomes. Methods: Differentially expressed immune-related genes (DEIRGs) between normal tissues and colon cancer tissues were identified and used to construct the co-expression network. Genes in the module with the most significant differences were further analyzed. Independent prognostic immune-related genes (IRGs) were identified by univariate and multivariate cox regression analysis. Independent prognostic IRGs were used to construct the IRGPI model using the multivariate cox proportional hazards regression model, and the IRGPI model was validated by independent dataset. ROC curves were plotted and AUCs were calculated to estimate the predictive power of the IRGPI model to prognosis. Gene set enrichment analysis (GSEA) was performed to screen the enriched KEGG pathways in the high-risk and low-risk phenotype. Correlations between IRGPI and clinical characteristic, immune checkpoint expression, TMB, immune cell infiltration, immune function, immune dysfunction, immune exclusion, immune subtype were analyzed. Results: Totally 680 DEIRGs were identified. Three independent IRGs,NR5A2, PPARGC1A and LGALS4, were independently related to survival. NR5A2, PPARGC1A and LGALS4 were used to establish the IRGPI model. Survival analysis showed that patients with high-risk showed worse survival than patients in the low-risk group. The AUC of the IRGPI model for 1-year, 3-year and 5-year were 0.584, 0.608 and 0.697, respectively. Univariate analysis and multivariate cox regression analysis indicated that IRGPI were independent prognostic factors for survival. Stratified survival analysis showed that patients with IRGPI low-risk and low TMB had the best survival, which suggested that combination of TMB and IRGPI can better predict clinical outcome. Immune cell infiltration, immune function, immune checkpoint expression and immune exclusion were different between IRGPI high-risk and low-risk patients. Conclusion: An immune-related genes prognostic index (IRGPI) was constructed and validated in the current study and the IRGPI maybe a potential biomarker for evaluating response to immunotherapy and clinical outcome for colon cancer patients.