Leucine Reconstitutes Phagocytosis-Induced Cell Death in E. coli-Infected Neonatal Monocytes-Effects on Energy Metabolism and mTOR Signaling.

MΦ differentiate from circulating monocytes (Mo). The reduced ability of neonatal Mo to undergo apoptosis after E. coli infection (phagocytosis-induced cell death (PICD)) could contribute to sustained inflammatory processes. The objective of our study was to investigate whether immune metabolism in Mo can be modified to gain access to pro-apoptotic signaling. To this end, we supplemented Mo from neonates and from adults with the branched amino acid leucine. In neonatal Mo, we observed increased energy production via oxidative phosphorylation (Oxphos) after E. coli infection via Seahorse assay. Leucine did not change phagocytic properties. In neonatal Mo, we detected temporal activation of the AKT and mTOR pathways, accompanied with subsequent activation of downstream targets S6 Kinase (S6K) and S6. FACS analyses showed that once mTOR activation was terminated, the level of anti-apoptotic BCL-2 family proteins (BCL-2; BCL-XL) decreased. Release of cytochrome C and cleavage of caspase-3 indicated involvement of the intrinsic apoptotic pathway. Concomitantly, the PICD of neonatal Mo was initiated, as detected by hypodiploid DNA. This process was sensitive to rapamycin and metformin, suggesting a functional link between AKT, mTOR and the control of intrinsic apoptotic signaling. These features were unique to neonatal Mo and could not be observed in adult Mo. Supplementation with leucine therefore could be beneficial to reduce sustained inflammation in septic neonates.

Blocking CCL5-CXCL4 heteromerization preserves heart function after myocardial infarction by attenuating leukocyte recruitment and NETosis.

Myocardial infarction (MI) is a major cause of death in Western countries and finding new strategies for its prevention and treatment is thus of high priority. In a previous study, we have demonstrated a pathophysiologic relevance for the heterophilic interaction of CCL5 and CXCL4 in the progression of atherosclerosis. A specifically designed compound (MKEY) to block this CCL5-CXCR4 interaction is investigated as a potential therapeutic in a model of myocardial ischemia/reperfusion (I/R) damage. 8 week-old male C57BL/6 mice were intravenously treated with MKEY or scrambled control (sMKEY) from 1 day before, until up to 7 days after I/R. By using echocardiography and intraventricular pressure measurements, MKEY treatment resulted in a significant decrease in infarction size and preserved heart function as compared to sMKEY-treated animals. Moreover, MKEY treatment significantly reduced the inflammatory reaction following I/R, as revealed by specific staining for neutrophils and monocyte/macrophages. Interestingly, MKEY treatment led to a significant reduction of citrullinated histone 3 in the infarcted tissue, showing that MKEY can prevent neutrophil extracellular trap formation in vivo. Disrupting chemokine heterodimers during myocardial I/R might have clinical benefits, preserving the therapeutic benefit of blocking specific chemokines, and in addition, reducing the inflammatory side effects maintaining normal immune defence.