CTRP6 as a negative regulator of anti-inflammatory M2 macrophage polarization

Chronic low-grade inflammation in adipose tissue, primarily driven by macrophages, plays a central role in obesity pathophysiology. C1q/TNF-related protein 6 (CTRP6), a member of the CTRP family, has emerged as a key regulator of this inflammatory process. Here, we demonstrate that CTRP6 expression is upregulated in adipose tissue macrophages during obesity, where it acts as a potent modulator of macrophage polarization by suppressing M2 polarization.

In RAW264.7 macrophages, we distinguished M1 and M2 polarization, induced by lipopolysaccharide (LPS) + interferon-gamma (IFNγ) and interleukin (IL)-4, respectively, by selecting two marker genes for each polarization type from a set of five widely used markers, based on a time-course analysis. We then assessed the effects of recombinant CTRP6 protein treatment on M1 and M2 polarization. Finally, we validated our findings in primary bone marrow-derived macrophages (BMDMs).

In naïve RAW264.7 macrophages, recombinant CTRP6 protein upregulated M1 marker genes (Tnf, Nos2) while downregulating M2 markers (Mrc1, Pparg). During M1 polarization induced by LPS+IFNγ, CTRP6 treatment had no significant effect. However, during IL-4-induced M2 polarization, CTRP6 not only enhanced M1 markers but also strongly suppressed M2 markers by inhibiting anti-inflammatory signal transducer and activator of transcription 6 (STAT6) signaling and relieving the inhibition of pro-inflammatory ERK1/2 signaling. Additionally, CTRP6 impaired mitochondrial activity, favoring glycolysis in macrophages. Importantly, these effects were serum-independent and confirmed in BMDMs.

Since endogenous CTRP6 expression in BMDMs is upregulated by M1 polarization inducers, it may further hinder inflammation resolution, even in the presence of IL-4 during tissue repair, establishing it as a key driver of adipose tissue inflammation in obesity.