Deletion of protein FSTL3 unravels the role of activin signaling in regulating macrophage L-arginine metabolism and kidney injury.

INTRODUCTION: Macrophages play a critical role in regulating kidney injury and repair. Arginase 1 (ARG1), a key enzyme in L-arginine metabolism, is predominantly expressed in macrophages in injured kidneys. Polyamines derived from L-arginine metabolism promote the resolution of tissue injury. Follistatin-like 3 (FSTL3) is a secreted protein that acts as an antagonist of activins. However, its role in the kidney remains unknown. METHODS: To study this, serum FSTL3 levels were measured in patients with chronic kidney disease (CKD) or acute kidney injury (AKI) using ELISA. Kidney injury was induced by folic acid or cisplatin in mice. Bone marrow-derived macrophages (BMDMs) were isolated from Smad3 knock out mice. Bulk RNA-seq analysis was performed on injured kidneys and BMDMs. L-arginine metabolites were analyzed with liquid chromatography-mass spectrometry. RESULTS: Serum FSTL3 levels were elevated in patients with CKD or AKI and were negatively correlated with estimated glomerular filtration rate and positively correlated with serum creatinine levels in CKD. Deletion of Fstl3 in mice ameliorated kidney injury and fibrosis, and this protective effect was associated with increased SMAD3 phosphorylation and ARG1 expression in macrophages. RNA-seq analysis revealed that arginine metabolism was highly enriched in the injured kidneys of Fstl3 knock out mice, as well as in BMDMs treated with activin A and IL-4. In vitro, activin A drastically synergized with the Th2 cytokines IL-4 and IL-13 to stimulate the expression of Arg1, along with Slc7a2 (also known as Cat2) and glycine amidinotransferase (Gatm), two other key genes involved in L-arginine metabolism, in macrophages, leading to enhanced L-arginine metabolism and polyamine production. Following folic acid treatment, Fstl3 knock out mice exhibited reduced tubular cell apoptosis and increased tubular cell proliferation. CONCLUSIONS: Our results uncover a previously unrecognized role for activin signaling in the regulation of L-arginine metabolism in macrophages. Fstl3 deletion activates this metabolism, thereby facilitating the resolution of kidney injury.