title: UCP2 - a key player in the interleukine-33 signalling pathway in macrophages?
Creators: name: Schröder, Fenja
Publisher: name: Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU)
Year: 2024

description: Background and goals Inflammatory processes as a reaction of the immune system to tissue damage, as well as to invading foreign material, are essential to prevent further spreading and overcoming the danger, as well as to subsequently eliminate the damage. They are divided into a pro- and anti-inflammatory phase. Macrophages are an essential component of both, with varying phenotypes depending on the trigger and the phase of inflammation. Recent evidence points to the fact that there is not an exchange of macrophage populations, but that the macrophages involved switch from a pro-inflammatory to an anti-inflammatory phenotype. Macrophages with anti-inflammatory properties are also called alternatively activated macrophages. Faas et al. identified interleukin-33 (IL-33) as the trigger of the anti-inflammatory macrophage phenotype termed M33. Initially, pro-inflammatory genes are expressed and the mitochondria are uncoupled in a phenomenon called "proton leak". This is followed by metabolic reprogramming to an alternatively activated phenotype, with increased phagocytosis activity via the transcription factor GATA3. The development of the IL-33-associated phenotype can be suppressed by chemical blockage of the uncoupling protein 2 (UCP2). UCP2 itself is associated with the ability to regulate levels of reactive oxygen species (ROS). The aim of this work is to further define the role of UCP2 and thus also the role of ROS in IL-33-induced macrophages. Methods Bone marrow-derived macrophages (BMDMs) were used for in vitro studies, focusing on gene expression and protein analyses. Cell metabolism and ROS levels were also measured, while phagocytic capacity was assessed to determine functionality. The in vivo relevance was then tested using a model of cardiotoxin (CTX)-induced sterile muscle injury. Results and observations In a repetition of the experiment by Faas et al., IL-33 caused the expression of alternative activation markers, with deletion of UCP2 leading to suppression of this phenotype and genipin, a UCP2-blocker, suppressing IL-33-induced proton leak. In vivo, the UCP2 knockout mouse showed increased tissue damage. However, there were no differences in regenerative capacity. Reduction of ROS levels by addition of substances that bind the radicals (ROS scavenger, e.g. vitamin C or N-acetyl-L-cysteine (NAC)) led to an increase in the alternative activation profile at the RNA and protein level of macrophages, but no influence on the phagocytic capacity of macrophages could be determined. Finally, the antioxidant NAC restored the IL-33 phenotype suppressed by genipin. Partial results of the present work were published in: Faas, Maria; Ipseiz, Natacha; Ackermann, Jochen; Culemann, Stephan; Grüneboom, Anika; Schröder, Fenja; Rothe, Tobias; Scholtysek, Carina; Eberhardt, Martin; Böttcher, Martin; Kirchner, Philipp; Stoll, Cornelia; Ekici, Arif; Fuchs, Maximilian; Kunz, Meik; Weigmann, Benno; Wirtz, Stefan; Lang, Roland; Hofmann, Joerg; Vera, Julio; Voehringer, David; Michelucci, Alessandro; Mougiakakos, Dimitrios; Uderhardt, Stefan; Schett, Georg; Krönke, Gerhard (2021): IL-33-induced metabolic reprogramming controls the differentiation of alternatively activated macrophages and the resolution of inflammation. In Immunity 54 (11), 2531-2546.e5. DOI: 10.1016/j.immuni.2021.09.010. This is noted in the figure descriptions of the results section for the corresponding data. Conclusions and discussion UCP2 and the subsequent reduction of ROS was identified to be an essential for the development of an alternative macrophage phenotype triggered by IL-33. In vivo, the deletion of UCP2 seems to lead only to slightly increased tissue damage, but to no differences in regenerative capacity after sterile muscle injury. Compensatory effects, including by UCP3, could possibly explain these results. Lowering ROS with ROS scavengers enhances the M33 phenotype in vitro and NAC is able to "rescue" the alternative macrophage phenotype suppressed by genipin. A possible signalling effect of the ROS level as a trigger of the change from a pro- to an anti-inflammatory profile, and thus for the resolution of inflammation, seems possible. However, whether the increase in ROS is the trigger or simply an amplifier, as well as the exact sequence of such a signalling cascade, remains an open question. The more precise identification of which ROS species are involved and of what origin, as well as the relevance of the results in vivo, must be analysed in future experiments. Should a lowering of the ROS level contribute to instigate the resolution of inflammatory processes, the clinically already established use of NAC and vitamin C would offer promising therapeutic approaches.