Fever-range hyperthermia improves the anti-apoptotic effect induced by low pH on human neutrophils promoting a proangiogenic profile

Abstract

Neutrophils have the shortest lifespan among leukocytes and usually die via apoptosis, limiting their deleterious potential. However, this tightly regulated cell death program can be modulated by pathogen-associated molecular patterns (PAMPs), danger-associated molecular pattern (DAMPs), and inflammatory cytokines. We have previously reported that low pH, a hallmark of inflammatory processes and solid tumors, moderately delays neutrophil apoptosis. Here we show that fever-range hyperthermia accelerates the rate of neutrophil apoptosis at neutral pH but markedly increases neutrophil survival induced by low pH. Interestingly, an opposite effect was observed in lymphocytes; hyperthermia plus low pH prevents lymphocyte activation and promotes the death of lymphocytes and lymphoid cell lines. Analysis of the mechanisms through which hyperthermia plus low pH increased neutrophil survival revealed that hyperthermia further decreases cytosolic pH induced by extracellular acidosis. The fact that two Na+/H+ exchanger inhibitors, 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and amiloride, reproduced the effects induced by hyperthermia suggested that it prolongs neutrophil survival by inhibiting the Na+/H+ antiporter. The neutrophil anti-apoptotic effect induced by PAMPs, DAMPs, and inflammatory cytokines usually leads to the preservation of the major neutrophil effector functions such as phagocytosis and reactive oxygen species (ROS) production. In contrast, our data revealed that the anti-apoptotic effect induced by low pH and hyperthermia induced a functional profile characterized by a low phagocytic activity, an impairment in ROS production and a high ability to suppress T-cell activation and to produce the angiogenic factors VEGF, IL-8, and the matrix metallopeptidase 9 (MMP-9). These results suggest that acting together fever and local acidosis might drive the differentiation of neutrophils into a profile able to promote both cancer progression and tissue repair during the late phase of inflammation, two processes that are strongly dependent on the local production of angiogenic factors by infiltrating immune cells.

Sepsis Induces a Dysregulated Neutrophil Phenotype That Is Associated with Increased Mortality

“…Furthermore, by manipulating the pH of the cellular environment in vitro, we could recapitulate the septic neutrophil phenotype suggesting that the disruption of cellular acid-base homeostasis may contribute to dysfunctional NETosis and perhaps other neutrophil functions. Changes in extracellular pH are common in many inflammatory diseases and lead to neutrophil activation, phagocytosis, and ROS production [ 31 , 32 ]. NET formation in relation to pH has not been studied before, and these data add to the literature of the negative impact of acid-base disturbance.…”

“…It has been proposed that the “first wave” of neutrophils to arrive at sites of infection is programmed for early NETosis in an attempt to contain the infection rapidly [ 35 ]. Those arriving later (which form the circulating pool on testing) may be activated by circulating cytokines and the septic environment and are resistant to apoptosis accounting for the attenuation of NETosis observed [ 32 , 36 ].…”

“…The changes in neutrophil phenotype may drive further local and distant organ damage and predispose patients to secondary infections by inducing a state of immunosuppression [ 32 , 43 ]. We propose a biphasic neutrophilic response to infection, with initial activation leading to migratory failure, frustrated phagocytosis, a release of large quantities of NETs, seen as cf-DNA, and a subsequent prolongation of neutrophil survival.…”

What Is the Evolutionary Fingerprint in Neutrophil Granulocytes?

“…In contrast to low temperature, fever accelerates the apoptosis rate at a neutral environment, but at the same time increases neutrophil survival if low pH is present. This is opposed to the reaction of lymphocytes to fever, as these cells die in low pH settings [231]. Fever additionally seems to maintain neutrophils inactivated as long as they are circulating.…”

Transient intracellular acidification regulates the core transcriptional heat shock response

“…More 560 plausibly, however, fever acts as a systemic signal which activates a cellular program with key 561 roles in modulating immune and inflammatory responses (Singh and Hasday, 2013). Indeed, 562 the apoptotic response of human neutrophils to fever temperatures is sharply dependent on 563 intracellular pH, with acidification promoting survival; local acidification is a hallmark of 564 inflammatory conditions and promotes neutrophil activation (Díaz et al, 2016). 565 We began by noting that the biological meaning of the longstanding association of cellular 566 stress with cytosolic acidification, observed from fungal cells to vertebrate neurons, has 567 remained unclear.…”