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CELL AGING AFFECTS GLYCOSYLATION OF IMMUNOGLOBULIN G SECRETED FROM MODEL CELL LINE FREESTYLE™ 293-F

Glycosylation of the Fc fragment of immunoglobulin G (IgG) affects the role of this antibody in the adaptive immune system. Aging is associated with changes in IgG glycosylation, primarily galactosylation, which leads to an increased proportion of proinflammatory IgG antibodies in human plasma. FreeStyle ™ 293-F is a model cell line used for production of recombinant IgG and is thus appropriate for studies of IgG glycosylation. In addition, glycome of IgG secreted from FreeStyle ™ 293-F cells is similar to IgG glycome from human plasma. The aim of this study was to investigate if the aging of the model cell line affects IgG glycome and, if so, are these changes similar to the changes observed on IgG from human plasma in older people. Ultra-high performance liquid chromatography revealed that cell aging, monitored during 90 days, indeed led to changes of IgG glycome. The most significant changes were an increase in the proportion of agalactosylated and a decrease in the proportion of fucosylated glycan structures. Proportion of high-mannose glycans also increased significantly, while proportions of sialylated glycans and glycans with bisecting N-acetylglucosamine remained stable during the time course experiment. Next, we investigated if glycan changes resulted from differential expression of glycosyltransferases responsible for individual steps in the IgG glycosylation pathway. This analysis revealed that a decrease of core fucosylation was associated with changes in FUT8 expression, while changes in galactosylation were not a direct consequence of altered B4GALT1 expression. An increase in the proportion of high-mannose glycans was in correlation with reduced MGAT1 and MGAT2 transcriptional activity, and the downregulation of these genes could also explain the decrease of complex IgG glycan structures. Overall, changes of IgG glycome caused by FreeStyle ™ 293-F cell aging were similar to those observed during human aging, most notably changes of IgG galactosylation. Interestingly, not all of the detected changes could be explained by differential expression of the corresponding glycosyltransferases.

immunoglobulin G, N-glycosylation, IgG glycome, HEK293 FreeStyle, in vitro cell aging

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