RESUMEN
Sialic acids are negatively charged carbohydrates that cap the glycans of glycoproteins and glycolipids. Sialic acids are involved in various biological processes including cell-cell adhesion and immune recognition. In dendritic cells (DCs), the major antigen-presenting cells of the immune system, sialic acids emerge as important regulators of maturation and interaction with other lymphocytes including T cells. Many aspects of how sialic acids regulate DC functions are not well understood and tools and model systems to address these are limited. Here, we have established cultures of murine bone marrow-derived DCs (BMDCs) that lack sialic acid expression using a sialic acid-blocking mimetic Ac53FaxNeu5Ac. Ac53FaxNeu5Ac treatment potentiated BMDC activation via toll-like receptor (TLR) stimulation without affecting differentiation and viability. Sialic acid blockade further increased the capacity of BMDCs to induce antigen-specific CD8+ T cell proliferation. Transcriptome-wide gene expression analysis revealed that sialic acid mimetic treatment of BMDCs induces differential expression of genes involved in T cell activation, cell-adhesion, and cell-cell interactions. Subsequent cell clustering assays and single cell avidity measurements demonstrated that BMDCs with reduced sialylation form higher avidity interactions with CD8+ T cells. This increased avidity was detectable in the absence of antigens, but was especially pronounced in antigen-dependent interactions. Together, our data show that sialic acid blockade in BMDCs ameliorates maturation and enhances both cognate T cell receptor-MHC-dependent and independent T cell interactions that allow for more robust CD8+ T cell responses.
Asunto(s)
Células de la Médula Ósea/inmunología , Linfocitos T CD8-positivos/inmunología , Comunicación Celular/inmunología , Células Dendríticas/inmunología , Ácido N-Acetilneuramínico/inmunología , Animales , Células de la Médula Ósea/metabolismo , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Adhesión Celular/genética , Adhesión Celular/inmunología , Comunicación Celular/genética , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Proliferación Celular/genética , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Células Cultivadas , Células Dendríticas/metabolismo , Femenino , Perfilación de la Expresión Génica/métodos , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Ratones Endogámicos C57BL , Ácido N-Acetilneuramínico/antagonistas & inhibidores , Ácido N-Acetilneuramínico/metabolismo , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunología , Receptores de Antígenos de Linfocitos T/metabolismo , Receptores Toll-Like/genética , Receptores Toll-Like/inmunología , Receptores Toll-Like/metabolismoRESUMEN
The development of a biotechnological platform for the removal of waste products (e.g. uremic toxins), often bound to proteins in plasma, is a prerequisite to improve current treatment modalities for patients suffering from end stage renal disease (ESRD). Here, we present a newly designed bioengineered renal tubule capable of active uremic toxin secretion through the concerted action of essential renal transporters, viz. organic anion transporter-1 (OAT1), breast cancer resistance protein (BCRP) and multidrug resistance protein-4 (MRP4). Three-dimensional cell monolayer formation of human conditionally immortalized proximal tubule epithelial cells (ciPTEC) on biofunctionalized hollow fibers with maintained barrier function was demonstrated. Using a tailor made flow system, the secretory clearance of human serum albumin-bound uremic toxins, indoxyl sulfate and kynurenic acid, as well as albumin reabsorption across the renal tubule was confirmed. These functional bioengineered renal tubules are promising entities in renal replacement therapies and regenerative medicine, as well as in drug development programs.
