Qianyue Zhang¹, Joost M. Lambooij¹, Clément Besqueut-Rougerie¹, Joeri Noteboom¹, Linh D. Nguyen¹, Arnaud Zaldumbide², Cornelis H. Hokke¹, Noortje de Haan^3*, Bruno Guigas^1*

¹Leiden University Center of Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands, ²Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden The Netherlands, and ³Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden The Netherlands. q.zhang@lumc.nl; *: shared senior authorship

Background: Obesity is a primary factor in the development of type 2 diabetes (T2D), promoting adipose tissue (AT) inflammation and insulin resistance. AT inflammation results from the crosstalk between hypertrophic adipocytes and AT immune cells, notably macrophages, which is partly driven by adipocyte-derived cytokines, metabolites and extracellular vesicles. Glycans, which are complex carbohydrate structures attached to cell-surface or secreted proteins and lipids, could also play a role in such crosstalk, although very few data are currently available. In this study, we investigated the impact of obesity on the adipocyte glycosylation machinery and cell N-glycan profiles in mice.

Method: C57BL6/J male wild-type mice were fed with a low-fat diet (LFD) or high-fat diet (HFD) for 15 weeks. Body weight, whole-body insulin sensitivity and epididymal white adipose tissue (eWAT) immune cell composition were determined. Adipocytes were isolated from eWAT, and subjected to N-glycomics and transcriptomics analyses, using MALDI mass spectrometry and bulk RNA sequencing, respectively.

Results: HFD increased body weight, adipocyte size and eWAT pro-inflammatory macrophages, and decreased insulin sensitivity, indicating whole-body insulin resistance and AT inflammation in obese mice. The N-glycomics analyses revealed distinct alterations of various types of N-glycans in adipocytes from obese mice, with increased bisecting GlcNAc structures in both hybrid (+174%; p<0.05) glycans and complex (+49%; p<0.05) di-antennary N-glycans. Adipocytes from obese mice also showed a higher abundance of tri-antennary N-glycan complex structures (+28%; p<0.05). Interestingly, the adipocyte expression of many genes related to the glycosylation machinery are significantly affected by HFD (16 upregulated, 14 downregulated; -0.5<log2FC>0.5, p<0.05), including Mgat4a, a glycosyltransferase involved in the formation of tri-antennary branching structures, that was upregulated in adipocytes from obese mice.

Conclusion: The observed alterations in adipocyte glycan profiles in obese mice suggest that some specific carbohydrate structures may play a role in cell-cell interaction. Future research should explore the functional implications of these glycan changes on macrophage activation and AT inflammation.