Sirt3 deficiency does not affect venous thrombosis or NETosis despite mild elevation of intracellular ROS in platelets and neutrophils in mice.

Hideki Hayashi,Leonard P Guarente,Kimberly Martinod,Deya Cherpokova,Stephen M Cifuni,Siu Ling Wong,Thilo Witsch,Maureen Gallant,Denisa D Wagner

doi:10.1371/journal.pone.0188341

Abstract

Inflammation is a common denominator in chronic diseases of aging. Yet, how inflammation fuels these diseases remains unknown. Neutrophils are the primary leukocytes involved in the early phase of innate immunity and inflammation. As part of their anti-microbial defense, neutrophils form extracellular traps (NETs) by releasing decondensed chromatin lined with cytotoxic proteins. NETs have been shown to induce tissue injury and thrombosis. Here, we demonstrated that Sirt3, a nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylase, an enzyme linked to human longevity, was expressed in mouse neutrophils and platelets. Using Sirt3-/- mice as a model of accelerated aging, we investigated the effects of Sirt3 deficiency on NETosis and platelet function, aiming to detect enhancement of thrombosis. More mitochondrial reactive oxygen species (ROS) were generated in neutrophils and platelets of Sirt3-/- mice compared to WT, when stimulated with a low concentration of phorbol 12-myristate 13-acetate (PMA) and a high concentration of thrombin, respectively. There were no differences in in vitro NETosis, with or without stimulation. Platelet aggregation was mildly augmented in Sirt3-/- mice compared to WT mice, when stimulated with a low concentration of collagen. The effect of Sirt3 deficiency on platelet and neutrophil activation in vivo was examined by the venous thrombosis model of inferior vena cava stenosis. Elevation of plasma DNA concentration was observed after stenosis in both genotypes, but no difference was shown between the two genotypes. The systemic response to thrombosis was enhanced in Sirt3-/- mice with significantly elevated neutrophil count and reduced platelet count. However, no differences were observed in incidence of thrombus formation, thrombus weight and thrombin-antithrombin complex generation between WT and Sirt3-/- mice. We conclude that Sirt3 does not considerably impact NET formation, platelet function, or venous thrombosis in healthy young mice.

Highlights

The aging population of the world is growing rapidly
Sirtuin 3 is expressed in neutrophils and intracellular reactive oxygen species (ROS) was higher in neutrophils isolated from Sirt3-/- mice compared to WT mice
We discovered that Sirt3 is expressed in neutrophils, and that Sirt3 deficiency increases neutrophil intracellular ROS levels after stimulation

Summary

Introduction

Since aging is a general risk factor for many chronic conditions, there has been an increase in the incidence of cardiovascular disease [1, 2]. It has been generally regarded that the imbalance of innate immunity and inflammation is an important etiological mechanism that causes age-related diseases [3]. Neutrophil counts [9, 10] and ROS generation [11] in neutrophils both increase with age in humans. These are implicated in predisposition toward NETosis as intracellular ROS production is essential for NETosis [12]. The etiology underlying neutrophil function that enhances age-related chronic disease remains unknown. To uncover the link between thrombosis, an age-related disease, and age-related alteration in neutrophil function, it is necessary to investigate aging-related genes such as Sirtuin 3; its role in NETosis, and its contribution to thrombus formation

Methods

Results

Conclusion