RESUMEN
The primary immune response against Staphylococcus aureus is mediated by neutrophils. In response to S. aureus and its proteins, neutrophil shows two different kinds of NETosis, viz. suicidal and vesicular NETosis. Glucose is the major energy source of neutrophils for performing NETosis. However, NETosis was found altered in response to high glucose levels. Growth of S. aureus was also found modulated in response to high glucose and they behave differently at different glucose levels. This work was attempted to study NET release in response to S. aureus cell-free culture supernatant at different glucose concentrations. Freshly isolated neutrophils were treated with different concentrations of glucose along with S. aureus cell-free culture supernatant and were analyzed for neutrophil extracellular trap formation, ROS production, and peptidylarginine deiminase 4 activities. Influence of calcium on NETosis was analyzed using calcium chelator (EDTA) and calcium inhibitor (TMB-8). With increasing glucose levels, NET release in response to S. aureus cell-free culture supernatant was increased. Oxidant level was also increased dose-dependently with increasing concentrations of glucose. At very high glucose concentrations (> 15 mM), vesicular NETosis was predominantly observed. At these glucose concentrations, peptidylarginine deiminase activity was found to be decreased. Furthermore, calcium quenching in the medium facilitated vesicular mode of NET release. In conclusion, calcium depletion occurring at high glucose concentrations can reduce peptidylarginine deiminase 4 activity and can thereby promote the vesicular NET release.
Asunto(s)
Calcio/metabolismo , Trampas Extracelulares/metabolismo , Glucosa/metabolismo , Neutrófilos/inmunología , Staphylococcus aureus/inmunología , Células Cultivadas , Trampas Extracelulares/inmunología , Humanos , Vesículas Secretoras/metabolismo , Infecciones Estafilocócicas/inmunologíaRESUMEN
Citrullination is a post-translation modification of proteins, where the proteinaceous arginine residues are converted to non-coded citrulline residues. The immune tolerance to such citrullinated protein can be lost, leading to inflammatory and autoimmune diseases. Citrullination is a chemical reaction mediated by peptidylarginine deiminase enzymes (PADs), which are a family of calcium-dependent cysteine hydrolase enzymes that includes five isotypes: PAD1, PAD2, PAD3, PAD4, and PAD6. Each PAD has specific substrates and tissue distribution, where it modifies the arginine to produce a citrullinated protein with altered structure and function. All mammalian PADs have a sequence similarity of about 70-95%, whereas in humans, they are 50-55% homologous in their structure and amino acid sequences. Being calcium-dependent hydrolases, PADs are inactive under the physiological level of calcium, but could be activated due to distortions in calcium homeostasis, or when the cellular calcium levels are increased. In this article, we analyze some of the currently available data on the structural properties of human PADs, the mechanisms of their calcium-induced activation, and show that these proteins contain functionally important regions of intrinsic disorder. Citrullination represents an important trigger of multiple physiological and pathological processes, and as a result, PADs are recognized to play a number of important roles in autoimmune diseases, cancer, and neurodegeneration. Therefore, we also review the current state of the art in the development of PAD inhibitors with good potency and selectivity.