Supplementary Material for: ANGPTL3 in podocytes contributes to the pathogenesis of lupus nephritis by activating MSR1 in macrophages

Introduction: Lupus nephritis (LN), a severe complication of systemic lupus erythematosus (SLE), is characterized by persistent immune dysregulation and glomerular injury predominantly driven by macrophage infiltration and type I interferon (IFN‑I) signaling. Crosstalk between injured podocytes and infiltrating macrophages is increasingly recognized as a key contributor to LN progression, however, the molecular mediators and their corresponding receptors remain poorly defined. This study investigates whether ANGPTL3 upregulation in podocytes activates macrophages through MSR1 and drives interferon-biased inflammation in LN. Methods: We employed secretome-based protein–protein interaction screening to identify ANGPTL3 receptors. The ANGPTL3–MSR1 interaction was supported by co-immunoprecipitation. In vitro, bone marrow–derived macrophages (BMDMs) were stimulated with ANGPTL3-conditioned media, and gene expression was analyzed by RNA sequencing. MSR1 was silenced using siRNA. In vivo, a pristane-induced lupus mouse model was used to assess glomerular ANGPTL3/MSR1 expression, renal function, and macrophage infiltration. Clinical relevance was assessed using Nephroseq datasets and human LN kidney specimens. Results: ANGPTL3 was identified as an MSR1-interacting ligand through a cell-based membrane protein screening assay and further supported by co-immunoprecipitation. In vitro, ANGPTL3-conditioned media induced an interferon-dominant transcriptional response in BMDMs, including upregulation of Tnip3 and Isg20. These effects were attenuated upon MSR1 silencing, confirming receptor-dependent activation. Notably, ANGPTL3 had minimal effect on M1/M2 polarization markers, indicating selective regulation of interferon signaling. Analysis of Nephroseq datasets and LN biopsies demonstrated elevated ANGPTL3 and MSR1 expression, which correlated with histopathological severity. Single-cell RNA-seq confirmed MSR1 enrichment in glomerular macrophages. In vivo, pristane-induced lupus mice exhibited increased glomerular ANGPTL3, Synaptopodin loss, proteinuria, and co-localization of F4/80 and MSR1, supporting a pathogenic podocyte–macrophage axis. Conclusion: This study identifies the ANGPTL3-MSR1 axis as a critical pathway linking podocyte injury to macrophage-driven inflammation in LN, highlighting its therapeutic potential. Targeting this axis may disrupt maladaptive immune crosstalk, offering a novel strategy for LN management.