Desialylation and sialidase secretion from monocytes and macrophages upon LPS activation

The cell surface expresses a dense layer of glycans often terminated with sialic acids (Sias), known as sialylation, which represents different cellular statuses. Meanwhile, the removal of Sias from the glycans catalyzed by sialidase (desialylation) is also involved in many biological processes, such as inflammatory responses. Previous studies demonstrated that desialylation is involved in LPS/TLR4 signaling pathway and pro-inflammatory cytokine production. In this report, we systematically profiled desialylation and sialidase activity, expression and secretion from monocytes and macrophages upon LPS activation.

THP-1 monocytes and THP-1 macrophages were activated by LPS (100 ng/mL) for 24 hours. The cellular Sia changes (or desialylation) was quantified by measuring the total Sia level in cell lysates by LC-MS/MS method. The cell surface desialylation was confirmed by detecting the Gal exposure (after desialylation) of the cells by flow cytometry with PNA lectin staining. Total cellular desialylation was confirmed by lectin blot for both THP-1 monocytes and THP-1 macrophages. The sialidase activity in THP-1 monocytes and THP-1 macrophages and their culture media were determined by using the exogenous sialidase substrate 4-MU-Neu5Ac and ganglioside GM3, respectively. Neu1 and Neu3 sialidase levels in THP-1 monocytes and THP-1 were determined by western blot. Neu1 and Neu3 sialidase levels in in the supernatant of THP-1 monocytes and THP-1 macrophages were determined by ELISA.

The total Sia level reduced significantly in both THP-1 monocytes and THP-1 macrophages upon LPS activation. Meanwhile, sialidase expression and activity increased significantly in THP-1 monocytes and THP-1 macrophages upon LPS activation. Surprisingly, sialidase activities also significantly increased in the cell culture media of both THP-1 monocytes and THP-1 macrophages, indicating sialidase secretion from THP-1 monocytes and THP-1 macrophages upon LPS activation. In comparison, THP-1 macrophages are highly desialylated than THP-1 monocytes upon LPS activation.

LPS activation causes significant desialylation of both THP-1 monocytes and THP-1 macrophages. Desialylation is consistent with enhanced expression and activity of Neu1 and Neu3 sialidase and their secretion from both THP-1 monocytes and THP-1 macrophages upon LPS activation.