Neuraminidases 1 and 3 Trigger Atherosclerosis by Desialylating Low-Density Lipoproteins and Increasing Their Uptake by Macrophages.

Background Chronic vascular disease atherosclerosis starts with an uptake of atherogenic modified low-density lipoproteins (LDLs) by resident macrophages, resulting in formation of arterial fatty streaks and eventually atheromatous plaques. Increased plasma sialic acid levels, increased neuraminidase activity, and reduced sialic acid LDL content have been previously associated with atherosclerosis and coronary artery disease in human patients, but the mechanism underlying this association has not been explored. Methods and Results We tested the hypothesis that neuraminidases contribute to development of atherosclerosis by removing sialic acid residues from glycan chains of the LDL glycoprotein and glycolipids. Atherosclerosis progression was investigated in apolipoprotein E and LDL receptor knockout mice with genetic deficiency of neuraminidases 1, 3, and 4 or those treated with specific neuraminidase inhibitors. We show that desialylation of the LDL glycoprotein, apolipoprotein B 100, by human neuraminidases 1 and 3 increases the uptake of human LDL by human cultured macrophages and by macrophages in aortic root lesions in Apoe-/- mice via asialoglycoprotein receptor 1. Genetic inactivation or pharmacological inhibition of neuraminidases 1 and 3 significantly delays formation of fatty streaks in the aortic root without affecting the plasma cholesterol and LDL levels in Apoe-/- and Ldlr-/- mouse models of atherosclerosis. Conclusions Together, our results suggest that neuraminidases 1 and 3 trigger the initial phase of atherosclerosis and formation of aortic fatty streaks by desialylating LDL and increasing their uptake by resident macrophages.

Neuraminidase 1 activates insulin receptor and reverses insulin resistance in obese mice.

OBJECTIVES: Neuraminidase 1 (NEU1) cleaves terminal sialic acids of glycoconjugates during lysosomal catabolism. It also modulates the structure and activity of cellular surface receptors affecting diverse pathways. Previously we demonstrated that NEU1 activates the insulin receptor (IR) and that NEU1-deficient CathAS190A-Neo mice (hypomorph of the NEU1 activator protein, cathepsin A/CathA) on a high-fat diet (HFD) develop hyperglycaemia and insulin resistance faster than wild-type animals. The major objective of the current work was to reveal the molecular mechanism by which NEU1 desialylation activates the IR and to test if increase of NEU1 activity in insulin target tissues reverses insulin resistance and glucose intolerance. METHODS: To test if desialylation causes a conformational change in the IR dimer we measured interaction between the receptor subunits by Bioluminescence Resonance Energy Transfer in the HEK293T cells either overexpressing NEU1 or treated with the NEU1 inhibitor. The influence of NEU1 overexpression on insulin resistance was studied in vitro in palmitate-treated HepG2 cells transduced with NEU1-expressing lentivirus and in vivo in C57Bl6 mice treated with HFD and either pharmacological inducer of NEU1, Ambroxol or NEU1-expressing adenovirus. NEU1-deficient CathAS190A-Neo mice were used as a control. RESULTS: By desialylation of IR, NEU1 induced formation of its active dimer leading to insulin signaling. Overexpression of NEU1 in palmitate-treated HepG2 cells restored insulin signaling, suggesting that increased NEU1 levels may reverse insulin resistance. Five-day treatment of glycemic C57Bl6 mice receiving HFD with the activator of the lysosomal gene network, Ambroxol, increased NEU1 expression and activity in muscle tissue, normalized fasting glucose levels, and improved physiological and molecular responses to glucose and insulin. Ambroxol did not improve insulin sensitivity in obese insulin-resistant CathAS190A-Neo mice indicating that the Ambroxol effect is mediated through NEU1 induction. Sustained increase of liver NEU1 activity through adenovirus-based gene transfer failed to attenuate insulin resistance most probably due to negative feedback regulation of IR expression. CONCLUSION: Together our results demonstrate that increase of NEU1 activity in insulin target tissues reverses insulin resistance and glucose intolerance suggesting that a pharmacological modulation of NEU1 activity may be potentially explored for restoring insulin sensitivity and resolving hyperglycemia associated with T2DM.

Identification of selective inhibitors for human neuraminidase isoenzymes using C4,C7-modified 2-deoxy-2,3-didehydro-N-acetylneuraminic acid (DANA) analogues.

In the past two decades, human neuraminidases (human sialidases, hNEUs) have been found to be involved in numerous pathways in biology. The development of selective and potent inhibitors of these enzymes will provide critical tools for glycobiology, help to avoid undesired side effects of antivirals, and may reveal new small-molecule therapeutic targets for human cancers. However, because of the high active site homology of the hNEU isoenzymes, little progress in the design and synthesis of selective inhibitors has been realized. Guided by our previous studies of human NEU3 inhibitors, we designed a series of C4,C7-modified analogues of 2-deoxy-2,3-didehydro-N-acetylneuraminic acid (DANA) and tested them against the full panel of hNEU isoenzymes (NEU1, NEU2, NEU3, NEU4). We identified inhibitors with up to 38-fold selectivity for NEU3 and 12-fold selectivity for NEU2 over all other isoenzymes. We also identified compounds that targeted NEU2 and NEU3 with similar potency.