Inhibition of mammalian target of rapamycin aggravates the respiratory burst defect of neutrophils from decompensated patients with cirrhosis

Abstract

Cirrhosis is commonly accompanied by impaired defense functions of polymorphonuclear leucocytes (PMNs), increased patient susceptibility to infections, and hepatocellular carcinoma (HCC). PMN antimicrobial activity is dependent on a massive production of reactive oxygen species (ROS) by nicotinamide adenine dinucleotide phosphate (NADPH) 2 (NADPH oxidase 2; NOX2), termed respiratory burst (RB). Rapamycin, an antagonist of mammalian target of rapamycin (mTOR), may be used in the treatment of HCC and in transplanted patients. However, the effect of mTOR inhibition on the PMN RB of patients with cirrhosis remains unexplored and was studied here using the bacterial peptide, formyl‐Met‐Leu‐Phe (fMLP), as an RB inducer. fMLP‐induced RB of PMN from patients with decompensated alcoholic cirrhosis was strongly impaired (30%‐35% of control) as a result of intracellular signaling alterations. Blocking mTOR activation (phospho‐S2448‐mTOR) with rapamycin further aggravated the RB defect. Rapamycin also inhibited the RB of healthy PMNs, which was associated with impaired phosphorylation of the NOX2 component, p47phox (phox: phagocyte oxidase), on its mitogen‐activated protein kinase (MAPK) site (S345) as well as a preferential inhibition of p38‐MAPK relative to p44/42‐MAPK. However, rapamycin did not alter the fMLP‐induced membrane association of p47phox and p38‐MAPK in patients' PMNs, but did prevent their phosphorylation at the membranes. The mTOR contribution to fMLP‐induced RB, phosphorylation of p47phox and p38‐MAPK was further confirmed by mTOR knockdown in HL‐60 cells. Finally, rapamycin impaired PMN bactericidal activity, but not bacterial uptake. Conclusion: mTOR significantly up‐regulates the PMN RB of patients with cirrhosis by p38‐MAPK activation. Consequently, mTOR inhibition by rapamycin dramatically aggravates their PMN RB defect, which may increase patients' susceptibility to infection. Thus, concerns should be raised about the use of rapamycin in immuno‐depressed patients. (HEPATOLOGY 2013)

Alterations in neutrophil extracellular traps is associated with the degree of decompensation of liver cirrhosis

“…It is known that this microbicidal mechanism is reactive oxygen species (ROS) dependent [13]; it has been demonstrated that neutrophils in patients with liver diseases present defects in the production of ROS, mainly in the production of superoxide anion and hydrogen peroxide [14]. This could be the cause of the deficient capability of NETs release in patients with LC, and it could also be involved in the high incidence of bacterial infections.…”

Impaired intracellular signaling, myeloperoxidase release and bactericidal activity of neutrophils from patients with alcoholic cirrhosis

“…Due to space constraints, this section is detailed in the Supplementary Material, and includes neutrophil isolation [8], MPO exocytosis and assay [37], bactericidal activity [8], western-blot analyses of protein phosphorylation [8] and RNA quantification.…”

“…Thus, the MPO intracellular pool was not impaired, which suggests that the deficient MPO release induced by fMLP in patients' neutrophils ( Fig 1A and B) may likely due to altered fPR-mediated signaling. To further explore this hypothesis, neutrophils were stimulated under optimal conditions with fMLP and the activated phosphorylated form of major signaling effectors was western-blotted [8,38,39]. Stimulation of healthy neutrophils induced a rapid phosphorylation of AKT, p38-MAPK and p44/42-MAPK (ERK1/2) ( Fig.…”

“…However, the impact of alcoholic cirrhosis on receptor-mediated signaling events underlying neutrophil antibacterial activities remains largely unknown. We recently showed that neutrophils from patients with decompensated alcoholic cirrhosis, exhibit impaired signaling and ROS production induced by the bacterial tripeptide fMetLeu-Phe (fMLP), which was further aggravated by the mTOR antagonist rapamycin [8]. This dysfunction was associated with a defective MAPK-dependent phosphorylation of p47phox, a major component of NADPH oxidase.…”

“…MPO release and ROS production are triggered by various pro-inflammatory mediators among which bacterial formylated peptides which also act as chemoattractants, thus alerting neutrophils in case of infection through stimulation of fPR, a G-protein coupled receptor [11]. Neutrophil antibacterial activities are tightly regulated by signaling events triggered via fPR including phospholipases, G-proteins, protein kinases such as protein kinase C (PKC), mitogen-activated protein kinases (MAPK) and mammalian target of rapamycin [6][7][8]. Signaling impairment under pathological situations or by drugs leads to neutrophil dysfunctions, which are detrimental to host-defense [9][10][11].…”