Interleukin-15 Signaling in HIF-1α Regulation in Natural Killer Cells, Insights Through Mathematical Models

Abstract

Natural killer (NK) cells belong to the first line of host defense against infection and cancer. Cytokines, including interleukin-15 (IL-15), critically regulate NK cell activity, resulting in recognition and direct killing of transformed and infected target cells. NK cells have to adapt and respond in inflamed and often hypoxic areas. Cellular stabilization and accumulation of the transcription factor hypoxia-inducible factor-1α (HIF-1α) is a key mechanism of the cellular hypoxia response. At the same time, HIF-1α plays a critical role in both innate and adaptive immunity. While the HIF-1α hydroxylation and degradation pathway has been recently described with the help of mathematical methods, less is known concerning the mechanistic mathematical description of processes regulating the levels of HIF-1α mRNA and protein. In this work we combine mathematical modeling with experimental laboratory analysis and examine the dynamic relationship between HIF-1α mRNA, HIF-1α protein, and IL-15-mediated upstream signaling events in NK cells from human blood. We propose a system of non-linear ordinary differential equations with positive and negative feedback loops for describing the complex interplay of HIF-1α regulators. The experimental design is optimized with the help of mathematical methods, and numerical optimization techniques yield reliable parameter estimates. The mathematical model allows for the investigation and prediction of HIF-1α stabilization under different inflammatory conditions and provides a better understanding of mechanisms mediating cellular enrichment of HIF-1α. Thanks to the combination of in vitro experimental data and in silico predictions we identified the mammalian target of rapamycin (mTOR), the nuclear factor-κB (NF-κB), and the signal transducer and activator of transcription 3 (STAT3) as central regulators of HIF-1α accumulation. We hypothesize that the regulatory pathway proposed here for NK cells can be extended to other types of immune cells. Understanding the molecular mechanisms involved in the dynamic regulation of the HIF-1α pathway in immune cells is of central importance to the immune cell function and could be a promising strategy in the design of treatments for human inflammatory diseases and cancer.

Innate Cytokine Induced Early Release of IFNγ and CC Chemokines from Hypoxic Human NK Cells Is Independent of Glucose

“…We previously reported synergistic upregulation of HIF-1α by IL-15 priming and chemical hypoxia [50] as well as of glycolytic gene transcription by IL-15 priming and hypoxia in human NK cells [7]. Here, these responses did however not translate into concordant changes in cellular protein levels ( Figure 4A).…”

“…Nevertheless, such short-term IL-15 exposure and hypoxia transcriptionally upregulate hypoxia inducible factor (HIF-1α) and glycolysis pathways in a synergistic fashion [7]. In agreement with this, treatment of human NK cells with IL-15 for 4 h under chemical hypoxia augments protein levels of HIF-1α [50], the major mediator of cellular adaption to hypoxia [51]. The impact of hypoxia on NK cell glycolysis measured at low oxygen has, however, not yet been reported.…”

“…Isolated cells were maintained at 10 6 /mL in complete RPMI-1640 medium containing 10 mM glucose (Sigma-Aldrich, Munich, Germany) and supplemented with 10% fetal bovine serum (FBS) and 2 mM l-glutamine (Thermo Fisher Scientific, Paisley, UK) in a standard tissue culture incubator (normoxia) or, for experiments at 1% oxygen (hypoxia), in an oxygen-controlled Eppendorf New Brunswick Galaxy 48R CO 2 incubator (Hamburg, Germany). Alternative to low oxygen, chemical hypoxia was established in some experiments as described [50] using the pan-prolyl hydroxylase inhibitor dimethyloxalylglycine (DMOG) (Selleck Chemicals, Houston, TX, USA) at 50 µM or 1-(5-Chloro-6-(trifluoromethoxy)-1H-benzoimidazol-2-yl)-1H-pyrazole-4-carboxylic acid (JNJ) (Calbiochem, San Diego, CA, USA) at 20 µM. At these concentrations, DMOG and JNJ were equally efficient in increasing glycolytic rates in primed human NK cells (Supplemental Figure S1).…”

“…Western blotting was performed as recently described [50] with the following antibodies-anti-TPI (ab96696), anti-PGK1 (ab38007) and anti-PDK1 (EPR19573, ab207450) from Abcam (Cambridge, UK) and β-Actin (8H10D10) from Cell Signaling Technology (Danvers, MA, USA).…”

The role of hypoxia in the tumor microenvironment and development of cancer stem cell: a novel approach to developing treatment

“…Thereby hypoxia contributes to escape from innate immunity. In contrast, the supporting role of hypoxia in NK cell priming and activation, in synergism with IL-15, has been reported [ 248 , 249 ].…”