Glycans linked to cell surface proteins encode a multitude of biological information decoded by specialized proteins termed lectins. Understanding this glycan based language and downstream signalling is of emerging significance due to research of the past two decades pointing at a large impact on regulating immune responses and maintenance of self-tolerance.Galectin-1 is a 14.5 kDa lectin conserved across species, which binds galactose ß1-4 N-acetylglucosamine containing oligosaccharides on the surface of immune and stromal cells. Its functional capability ranges from induction of T cell apoptosis, a shift of immune reactions from TH1 to TH2 and expansion of regulatory T cells (Tregs) to modulation of monocyte and macrophage functions. With its anti-inflammatory properties revealed in animal models of autoimmunity and chronic inflammation such as experimental autoimmune encephalomyelitis (EAE) and experimental autoimmune uveitis (EAU), galectin-1 has emerged as a factor of clinical significance.In view of the immune privileged status of the testis, we hypothesised that galectin-1 could play a role in maintaining the immunosuppressive phenotype of this organ. As a first step, human galectin-1 C2S was recombinantly expressed, purified, labelled with Alexa Fluor 647 and its binding was investigated by immunohistochemistry using testicular cryosections. In a second step, isolated testicular macrophages, Sertoli and peritubular cells as important cells of testicular innate immunity were used to test their binding capacity of galectin-1 by flow cytometry.It was shown that all investigated testicular cell types are able to bind galectin-1, albeit with different intensity. The fluorescence signal of Alexa Fluor 647 conjugated galectin-1 C2S binding to peritubular and Sertoli cells is comparable to that of phytohaemagglutinin (PHA) stimulated mononuclear cells as positive control. In contrast galectin-1 binding to testicular macrophages is much weaker with an intensity lower than unstimulated mononuclear cells. The specificity of binding was verified by co-incubation with lactose or sucrose, respectively.In order to examine the phenotype of cell surface glycans, which mediate the affinity of galectin-1 for the respective cells, we applied a new set of plant lectins (MAA, LEA, PNA, SNA-I) in the analysis of isolated testicular cells by flow cytometry.It was found that reduced binding of galectin-1 to testicular macrophages in comparison to Sertoli and peritubular cells could result from a high degree of terminal a2-6 sialylation in cell surface glycans of testicular macrophages a regulatory mechanism known to modulate susceptibility of TH1 and TH2 cells to galectin-1 induced apoptosis.In summation, it is concluded from these studies that peritubular cells and Sertoli cells are the primary target cells for galectin-1 amongst the investigated cell types in the testis, but surprisingly not testicular macrophages. It seems likely that galectin-1 contributes to the testicular immune privilege mainly by interaction with testicular somatic cells.
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