Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5

Insulin-like Growth Factor 1, but not Insulin, Polarizes Macrophages to an anti-Inflammatory Phenotype by Jak/Stat Activation
A. Spychala1, R. Nederlof1, S. Reidel1, P. Petzsch2, K. Köhrer2, A. Gödecke1
1Institut für Herz- und Kreislaufphysiologie, Universitätsklinikum Düsseldorf, Düsseldorf; 2Biologisch-Medizinisches Forschungszentrum (BMFZ), Heinrich-Heine Universität, Düsseldorf;
Background Macrophages have a wide spectrum of activation states influenced by an array of tissue-derived growth factors, cytokines, chemokines, lipokines and hormones. They are classically divided in pro-inflammatory (M1) or anti-inflammatory (M2) state. Insulin-like growth factor 1 (Igf1) is an anabolic hormone, similar to insulin in molecular structure and signaling. Igf1 controls proliferation, differentiation and metabolism of cells. Previously we have reported that treatment with Igf1 after myocardial infarction leads to an enhanced number of anti-inflammatory macrophages in heart tissue associated with an improved heart function. Macrophage polarization by Il4 towards a M2-like state leads to high secretion of Igf1 and expression of Igf1r, suggesting an auto/paracrine function of Igf1 in macrophages. However, signal transduction via Igf1 in macrophages and resulting specific activation state is poorly understood. Here we investigate whether Igf1 and insulin can influence the activation state of bone marrow derived macrophages (BMDM) and via which signaling pathway this is transduced.

Methods
Tibia and femoral bones were harvested from mice. Bone marrow derived cells were isolated and cultivated for 5 days under mCSF treatment (10 ng/ml). The obtained macrophages were then treated with different polarizers to induce an M1-phenotype (10 ng/ml LPS and 2 ng/ml IFN-ɣ), an M2-phenotype (30 ng/ml IL4) or macrophages were treated with IGF1 (10 ng/ml) or insulin (10 ng/ml). The RNA was isolated from macrophages and expression analysis was carried out by next-generation sequencing (NGS) to reveal different expression patterns. To investigate different pathway transitions, cells were furthermore treated with the janus kinase (Jak) inhibitors Ruxolitinib (Jak1/2 inhibitor), InSolution™ (Jak2/3 inhibitor) or BMS-911543 (Jak2 inhibitor) and expression of marker genes and phosphorylation of Jak- and Stat-proteins were analyzed by realtime PCR or western blot.

Results
The expression analysis demonstrated a clear difference between the Igf1 and insulin effects on macrophage polarization. While treatment with Igf1 led to an increased expression of the M2 marker genes Arg1 (6129x-fold) and Retnla (489447x-fold), insulin showed neither polarization potential for the M2-like nor M1-like phenotype, but induced a distinct gene expression program. Of note, insulin activates genes, which are associated with the remodeling of the extracellular matrix (collagens, Has2, Bgn). The investigation of the signal transduction of Igf1 in primary macrophages showed that Igf1, like Il4, activates the tyrosine kinases Jak1 and Jak3, which results in an activation of the transcription factor Stat6, which is the major transcription factor responsible for the induction of anti-inflammatory genes during macrophage M2 polarization. Treatment of macrophages with different Jak inhibitors revealed, that the Igf1 induced Stat6 activation is mainly mediated via Jak1.

Conclusion
Despite a similar structure and signal transduction mediated by Igf1 and insulin in many cells, Igf1 revealed a clear difference in signal transition and activation of macrophages compared to insulin. The ability of Igf1 to polarize macrophages to an anti-inflammatory state via Jak1-Stat6 signaling may be a possible new target to find novel approaches for the treatment of inflammatory diseases via regulating macrophage activation states.
Funded by SFB1116/A06
 

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