Initial binding of human immunodeficiency virus-1 (HIV-1) to its susceptible CD4(+) cells is the limiting step for the establishment of infection as the avidity of viral envelope gp120 for CD4 is not high and the number of viral envelope spikes on the surface is found to be low compared to highly infectious viruses. Several host factors, such as C-type lectins, are listed as being able to enforce or facilitate the crucial interaction of HIV-1 to the susceptible cell. Recent works suggest that a host soluble beta-galactoside-binding lectin, galectin-1, also facilitates both virion binding and the infection of target cells in a manner dependent on lactose but not mannose, suggesting that this soluble galectin can be considered as a host factor that influences HIV-1 pathogenesis. In this chapter, we describe methods used to investigate the potential role of the galectin family in HIV-1-mediated disease progression.

Tropheryma whipplei, is an actinobacterium that causes different infections in humans, including Whipple's disease. The bacterium infects and replicates in macrophages, leading to a Th2-biased immune response. Previous studies have shown that T. whipplei harbors complex surface glycoproteins with evidence of sialylation. However, the exact contribution of these glycoproteins for infection and survival remains obscure. To address this, we characterized the bacterial glycoprofile and evaluated the involvement of human β-galactoside-binding lectins, Galectin-1 (Gal-1) and Galectin-3 (Gal-3) which are highly expressed by macrophages as receptors for bacterial glycans. Tropheryma whipplei glycoproteins harbor different sugars including glucose, mannose, fucose, β-galactose and sialic acid. Mass spectrometry identification revealed that these glycoproteins were membrane- and virulence-associated glycoproteins. Most of these glycoproteins are highly sialylated and N-glycosylated while some of them are rich in poly-N-acetyllactosamine (Poly-LAcNAc) and bind Gal-1 and Gal-3. In vitro, T. whipplei modulates the expression and cellular distribution of Gal-1 and Gal-3. Although both galectins promote T. whipplei infection by enhancing bacterial cell entry, only Gal-3 is required for optimal bacterial uptake. Finally, we found that serum levels of Gal-1 and Gal-3 were altered in patients with T. whipplei infections as compared to healthy individuals, suggesting that galectins are also involved in vivo. Among T. whipplei membrane-associated proteins, poly-LacNAc rich-glycoproteins promote infection through interaction with galectins. T. whipplei modulates the expression of Gal-1 and Gal-3 both in vitro and in vivo. Drugs interfering with galectin-glycan interactions may provide new avenues for the treatment and diagnosis of T. whipplei infections.

Galectins (Gals) are evolutionarily conserved proteins that bind to β-galactoside containing glycans. Abnormal expression of Gals is associated with the development, progression, and metastasis of different types of cancer. Among the 11 Gals identified in humans, the roles of Gal-1 and Gal-3 have been extensively investigated in various tumors. Here, we summarize the roles of overly expressed Gal-1 and Gal-3 in the pathogenesis of hepatocellular carcinoma (HCC). The overexpression of Gal-1 and Gal-3 correlates with tumor growth, HCC cell migration and invasion, tumor aggressiveness, metastasis, and poor prognosis. A potentially promising future treatment strategy for HCC may include the combination of immunotherapy with Gal-1 inhibition. Additional research is warranted to investigate targeting Gal-1 and Gal-3 for HCC treatment.

Galectins are known to regulate cell differentiation and growth as well as cell adhesion and apoptosis. Galectins have been discussed as possible prognosticators for survival in renal cell cancer (RCC) and other urological tumors. They might also play an emerging role as possible new marker-proteins for RCC. In this study, we analyzed the expression of galectin-1 and galectin-3 mRNA in order to further investigate their clinical significance in RCC.

Intracerebral hemorrhage (ICH) is a devastating sub-type of stroke with no proven treatment. Given the emerging role of Galectin-1 and Galectin-3 in neuroimmune responses, the objective of the current manuscript is to elucidate hemorrhagic-injury induced modulation and cellular expression of Galectin-1 and Galectin-3 in the brain in a pre-clinical model of ICH. To address this, ICH was induced in male CD1 mice by collagenase injection method. Western blotting as well as Immunofluorescence staining was performed to characterize the temporal expression pattern as well as cellular localization of Galectin-1 and Galectin-3 after ICH. Further, genetic studies were conducted to assess the functional role of Galectin-1 and Galectin-3 in inflammatory response employing a murine macrophage cell line, RAW 264.7. Galectin-1 and Galectin-3 exhibited very profound and increased expression from day 3 to day 7-post-injury, in the perihematomal brain region after ICH in comparison to Sham. Further, Galectin-1 expression was mostly observed in GFAP-positive astrocytes whereas Galectin-3 expression was observed mostly in Iba1-positive microglia/macrophages as well as CD16/32 (M1 microglial/macrophage marker)-positive cells. Moreover, genetic studies revealed a negative regulatory role of both Galectin-1 and Galectin-3 in the release of a proinflammatory cytokine, IL-6 from RAW 264.7 cells depending on the stimulus. Altogether, the present manuscript demonstrates for the first time, increased expression as well as cellular localization of Galectin-1 and Galectin-3 in the perihematomal brain regions after ICH. In addition, the manuscript raises the potential of Galectin-1 and Galectin-3 in modulating glial responses and thereby brain injury after ICH, warranting further investigation.

Acute lymphoblastic leukemias arising from the malignant transformation of B-cell precursors (BCP-ALLs) are protected against chemotherapy by both intrinsic factors as well as by interactions with bone marrow stromal cells. Galectin-1 and Galectin-3 are lectins with overlapping specificity for binding polyLacNAc glycans. Both are expressed by bone marrow stromal cells and by hematopoietic cells but show different patterns of expression, with Galectin-3 dynamically regulated by extrinsic factors such as chemotherapy. In a comparison of Galectin-1 x Galectin-3 double null mutant to wild-type murine BCP-ALL cells, we found reduced migration, inhibition of proliferation, and increased sensitivity to drug treatment in the double knockout cells. Plant-derived carbohydrates GM-CT-01 and GR-MD-02 were used to inhibit extracellular Galectin-1/-3 binding to BCP-ALL cells in co-culture with stromal cells. Treatment with these compounds attenuated migration of the BCP-ALL cells to stromal cells and sensitized human BCP-ALL cells to vincristine and the targeted tyrosine kinase inhibitor nilotinib. Because N-glycan sialylation catalyzed by the enzyme ST6Gal1 can regulate Galectin cell-surface binding, we also compared the ability of BCP-ALL wild-type and ST6Gal1 knockdown cells to resist vincristine treatment when they were co-cultured with Galectin-1 or Galectin-3 knockout stromal cells. Consistent with previous results, stromal Galectin-3 was important for maintaining BCP-ALL fitness during chemotherapy exposure. In contrast, stromal Galectin-1 did not significantly contribute to drug resistance, and there was no clear effect of ST6Gal1-catalysed N-glycan sialylation. Taken together, our results indicate a complicated joint contribution of Galectin-1 and Galectin-3 to BCP-ALL survival, with different roles for endogenous and stromal produced Galectins. These data indicate it will be important to efficiently block both extracellular and intracellular Galectin-1 and Galectin-3 with the goal of reducing BCP-ALL persistence in the protective bone marrow niche during chemotherapy.

We have shown that galectin-1 and galectin-3 are functionally redundant splicing factors. Now we provide evidence that both galectins are directly associated with spliceosomes by analyzing RNAs and proteins of complexes immunoprecipitated by galectin-specific antisera. Both galectin antisera co-precipitated splicing substrate, splicing intermediates and products in active spliceosomes. Protein factors co-precipitated by the galectin antisera included the Sm core polypeptides of snRNPs, hnRNP C1/C2 and Slu7. Early spliceosomal complexes were also immunoprecipitated by these antisera. When splicing reactions were sequentially immunoprecipitated with galectin antisera, we found that galectin-1 containing spliceosomes did not contain galectin-3 and vice versa, providing an explanation for the functional redundancy of nuclear galectins in splicing. The association of galectins with spliceosomes was (i) not due to a direct interaction of galectins with the splicing substrate and (ii) easily disrupted by ionic conditions that had only a minimal effect on snRNP association. Finally, addition of excess amino terminal domain of galectin-3 inhibited incorporation of galectin-1 into splicing complexes, explaining the dominant-negative effect of the amino domain on splicing activity. We conclude that galectins are directly associated with splicing complexes throughout the splicing pathway in a mutually exclusive manner and they bind a common splicing partner through weak protein-protein interactions.

Accumulating evidence suggests that extracellular galectin-1 and galectin-3 promote angiogenesis. Increased expression of galectin-1 and/or galectin-3 has been reported to be associated with tumour progression. Thus, it is critical to identify their influence on angiogenesis.

Dengue virus (DENV) is an enveloped RNA virus that is mosquito-transmitted and can infect a variety of immune and non-immune cells. Response to infection ranges from asymptomatic disease to a severe disorder known as dengue hemorrhagic fever. Despite efforts to control the disease, there are no effective treatments or vaccines. In our search for new antiviral compounds to combat infection by dengue virus type 1 (DENV-1), we investigated the role of galectin-1, a widely-expressed mammalian lectin with functions in cell-pathogen interactions and immunoregulatory properties. We found that DENV-1 infection of cells in vitro exhibited caused decreased expression of Gal-1 in several different human cell lines, suggesting that loss of Gal-1 is associated with virus production. In test of this hypothesis we found that exogenous addition of human recombinant Gal-1 (hrGal-1) inhibits the virus production in the three different cell types. This inhibitory effect was dependent on hrGal-1 dimerization and required its carbohydrate recognition domain. Importantly, the inhibition was specific for hrGal-1, since no effect was observed using recombinant human galectin-3. Interestingly, we found that hrGal-1 directly binds to dengue virus and acts, at least in part, during the early stages of DENV-1 infection, by inhibiting viral adsorption and its internalization to target cells. To test the in vivo role of Gal-1 in DENV infection, Gal-1-deficient-mice were used to demonstrate that the expression of endogenous Galectin-1 contributes to resistance of macrophages to in vitro-infection with DENV-1 and it is also important to physiological susceptibility of mice to in vivo infection with DENV-1. These results provide novel insights into the functions of Gal-1 in resistance to DENV infection and suggest that Gal-1 should be explored as a potential antiviral compound.

Approximately 30-40% of the patients with early stage non-small cell lung cancer (NSCLC) will present with recurrent disease within two years of resection. Here, we performed extensive galectin expression profiling in a retrospective study using frozen and paraffin embedded tumor tissues from 87 stage I/II NSCLC patients. Our data show that galectin mRNA expression in NSCLC is confined to galectin-1, -3, -4, -7, -8, and -9. Next to stage, univariable Cox regression analysis identified galectin-1, galectin-9FL and galectin-9Δ5 as possible prognostic markers. Kaplan-Meier survival estimates revealed that overall survival was significantly shorter in patients that express galectin-1 above median levels, i.e., 23.0 (2.9-43.1) vs. 59.9 (47.7-72.1) months (p = 0.020) as well as in patients that express galectin-9Δ5 or galectin-9FL below the median, resp. 59.9 (41.9-75.9) vs. 32.8 (8.7-56.9) months (p = 0.014) or 23.2 (-0.4-46.8) vs. 58.9 (42.9-74.9) months (p = 0.042). All three galectins were also prognostic for disease free survival. Multivariable Cox regression analysis showed that for OS, the most significant prognostic model included stage, age, gal-1 and gal-9Δ5 while the model for DFS included stage, age and gal-9Δ5. In conclusion, the current study confirms the prognostic value of galectin-1 and identifies galectin-9Δ5 as novel potential prognostic markers in early stage NSCLC. These findings could help to identify early stage NSCLC patients that might benefit most from adjuvant chemotherapy.

The role of endogenous galectin-1 (Gal-1) in acute inflammation has been poorly investigated. We therefore performed the carrageenan-induced paw edema model in wild-type and Gal-1(-/-) mice. On subplantar injection of carrageenan, Gal-1(-/-) mice displayed a similar first phase of edema (≤24 hours) to wild-type mice; however, a much less pronounced second phase (48 to 96 hours) was evident in this genotype. This reduced inflammation was associated with lower paw expression of inflammatory genes and cell infiltrates. Analysis of galectin protein and mRNA expression revealed high expression of Gal-1 in wild-type paws during resolution (≥48 hours), with some expression of galectin-9 (Gal-9). Administration of stable Gal-1 to wild-type mice completely ablated the first phase of edema but was ineffective when administered therapeutically at the 24-hour time point. Conversely, Gal-9 administration did not alter the first phase of edema but significantly reduced the second phase when administered therapeutically. This suggests anti-inflammatory actions for both proteins in this model albeit at different phases of the inflammatory response. Collectively, these data indicate that the absence of endogenous Gal-1 results in an abrogated response during the second phase of the edema reaction.

Myocardial infarction (MI) is the most serious manifestation of coronary artery disease and the cause of significant mortality and morbidity worldwide. Galectin-1(GAL-1), a divalent 14.5-kDa protein, is present both inside and outside cells, and has both intracellular and extracellular functions. Hypoxia inducible factor-1 alpha (HIF-1α) is a transcription factor mediating early and late responses to myocardial ischemia. Identification of the pattern of expression of GAL-1 and HIF-1α in the heart during the first 24 hours following acute MI will help in understanding early molecular changes in this event and may provide methods to overcome serious complications. Mouse model of MI was used and heart samples were processed for immunohistochemical and immunofluorescent labeling and Enzyme linked immunosorbent assay to identify GAL-1 and HIF 1α levels in the heart during the first 24 hours following MI. There was significant increase in left ventricular GAL-1 at 20 (p = 0.001) and 30 minutes (p = 0.004) following MI. There was also a significant increase in plasma GAL-1 at 4 hours (p = 0.012) and 24 hours (p = 0.001) following MI. A significant increase in left ventricular HIF-1 α was seen at 20 minutes (p = 0.047) following MI. In conclusion, we show for the first time that GAL-1 level in the left ventricle is increased in early ischemic period. We also report for the first time that HIF-1 α is significantly increased at 20 minutes following MI. In addition we report for the first time that mouse plasma GAL-1 level is significantly raised as early as 4 hours following MI.

Galectins are beta-galactose specific binding proteins. In human cancers, including hepatocellular carcinoma (HCC), galectin-1 (Gal-1) is often found to be overexpressed. In order to combat the dismal diagnosis and death rates of HCC, gene silencing and targeted inhibition of Gal-1 was investigated for its improved therapeutic potential.

Four generations of lactose-functionalized polyamidoamine (PAMAM) were employed to further the understanding of multivalent galectin-1 mediated interactions. Dynamic light scattering and fluorescence microscopy were used to study the multivalent interaction of galectin-1 with the glycodendrimers in solution, and glycodendrimers were observed to organize galectin-1 into nanoparticles. In the presence of a large excess of galectin-1, glycodendrimers nucleated galectin-1 into nanoparticles that were remarkably homologous in size (400-500 nm). To understand augmentation of oncologic cellular aggregation by galectin-1, glycodendrimers were used in cell-based assays with human prostate carcinoma cells (DU145). The results revealed that glycodendrimers provided competitive binding sites for galectin-1, which diverted galectin-1 from its typical function in cellular aggregation of DU145 cells.

The pathogenesis of oral-intestinal allergy syndrome (OIAS) has not been well understood. Published data indicate that galectin (Gal) 1 has immune regulatory functions. This study tests a hypothesis that Gal1 inhibits oral-intestinal allergy syndrome.

Clinical studies in glioblastoma and pancreatic carcinoma patients strongly support the further development of H-1 protoparvovirus (H-1PV)-based anticancer therapies. The identification of cellular factors involved in the H-1PV life cycle may provide the knowledge to improve H-1PV anticancer potential. Recently, we showed that sialylated laminins mediate H-1PV attachment at the cell membrane. In this study, we revealed that H-1PV also interacts at the cell surface with galectin-1 and uses this glycoprotein to enter cancer cells. Indeed, knockdown/out of LGALS1, the gene encoding galectin-1, strongly decreases the ability of H-1PV to infect and kill cancer cells. This ability is rescued by the re-introduction of LGALS1 into cancer cells. Pre-treatment with lactose, which is able to bind to galectins and modulate their cellular functions, decreased H-1PV infectivity in a dose dependent manner. In silico analysis reveals that LGALS1 is overexpressed in various tumours including glioblastoma and pancreatic carcinoma. We show by immunohistochemistry analysis of 122 glioblastoma biopsies that galectin-1 protein levels vary between tumours, with levels in recurrent glioblastoma higher than those in primary tumours or normal tissues. We also find a direct correlation between LGALS1 transcript levels and H-1PV oncolytic activity in 53 cancer cell lines from different tumour origins. Strikingly, the addition of purified galectin-1 sensitises poorly susceptible GBM cell lines to H-1PV killing activity by rescuing cell entry. Together, these findings demonstrate that galectin-1 is a crucial determinant of the H-1PV life cycle.

Galectin-1, a member of the beta-galactoside-binding lectin family, exists in both reduced and oxidized states. Oxidized galectin-1 (Gal-1/Ox), which lacks lectin activity, has been shown to promote axonal regeneration after injury by activating macrophages, which causes the release of factors that enhance Schwann cell migration and neurite outgrowth. However, the mechanism of macrophage activation by Gal-1/Ox remains unknown. In this study, we examined the effects of Gal-1/Ox on RAW264.7 macrophages and RT4-D6P2T Schwann cells. Gal-1/Ox stimulated migration of RT4-D6P2T Schwann cells directly and by activating RAW264.7 macrophages to release factors that promoted cell migration. Gal-1/Ox inhibited nitric oxide (NO) production induced by interferon-gamma by suppressing expression of inducible NO synthase in RAW264.7 macrophages and not by arginase activation and cell death. Furthermore, Gal-1/Ox-activated extracellular signal-regulated protein kinase 1/2 (ERK1/2) in RAW264.7 macrophages, although the mitogen-activated protein kinase (MEK)/ERK1/2 pathway was not involved in release of factors that promoted Schwann cell migration. On the other hand, Gal-1/Ox-induced RT4-D6P2T Schwann cell migration appeared to be mediated by the MEK/ERK1/2 pathway. These results suggest that Gal-1/Ox inhibits inflammatory responses in macrophages and promotes Schwann cell migration directly and by macrophage activation.

Existing data on the spatiotemporal expression patterns of a variety of galectins in murine atherosclerosis are limited. We investigated the expression levels of galectins, and their in vivo spatiotemporal expression patterns and statin responsiveness in the inflamed atherosclerotic plaques of apolipoprotein E (apoE)(-/-) mice.

Hepatic stellate cells (HSCs) induce immune privilege and promote hepatocellular carcinoma (HCC) by suppressing the immune system. On the other hand, galectin-1 and miRNA-22 (miR-22) are dysregulated in HCC and serve as prognostic indicators for patients. In this study, therefore, we measured galectin-1 and miR-22 expression in HSCs isolated from HCC tissues (Ca-HSCs), and in normal liver tissues (N-HSCs) as a control. We also investigated the apoptosis rate among T cells and the production of cytokines (IFN-γ and IL-10) in HSCs co-cultured with T cells. And we used immunohistochemical staining to tested for correlation between galectin-1 expression, CD3 expression and clinicopathological features in 162 HCC patients. Our results showed that galectin-1 expression was much higher in Ca-HSCs than in N-HSCs. Overexpression of galectin-1 promoted HSC-induced T cell apoptosis and cytokine production (IFN-γ and IL-10), while miR-22 expression inhibited it. Galectin-1 expression correlated negatively with miR-22 expression in HSCs. High galectin-1 and low CD3 expression levels were associated with poor prognosis in HCC patients. These results suggest that the immunosuppressive microenvironment promoted by HSC-derived galectin-1 in HCC can be inhibited by miR-22. Galectin-1 and miR-22 could potentially serve as prognostic markers and therapeutic targets in HCC.

Despite their abundance at gastrointestinal sites, little is known about the role of galectins in gut immune responses. We have therefore investigated the Citrobacter rodentium model of colonic infection and inflammation in Galectin-1 or Galectin-3 null mice. Gal-3 null mice showed a slight delay in colonisation after inoculation with C. rodentium and a slight delay in resolution of infection, associated with delayed T cell, macrophage and dendritic cell infiltration into the gut mucosa. However, Gal-1 null mice also demonstrated reduced T cell and macrophage responses to infection. Despite the reduced T cell and macrophage response in Gal-1 null mice, there was no effect on C. rodentium infection kinetics and pathology. Overall, Gal-1 and Gal-3 play only a minor role in immunity to a gut bacterial pathogen.