The polarization states and molecular signatures of macrophages in the synovium of patients with rheumatoid arthritis (RA) are not well understood. We aimed to identify specific subpopulations of macrophages and their features in RA synovium thereby providing a theoretical basis for treatment of RA. Single-cell RNA sequencing (scRNA-seq) was used to identify cell subsets and their gene signatures in synovial cells of patients with RA and osteoarthritis (OA). Spatial distribution of macrophages was visualized by deconvolving spatial transcriptomic data with scRNA-seq data. Flow cytometry and immunofluorescence were applied to investigate the expression of macrophage polarization indicators CD86 and CD206. Trajectory analysis was used to determine differentiation relationships. Transcription factor (TF) analysis was performed to find specific TFs. scRNA-seq identified three cell clusters of macrophages: M0-like MARCO+ Mϕ1, M2-like CSF1R+ Mϕ2, and M1-like PLAUR+ Mϕ3. Mϕ1 distributed widely in the synovium, whereas Mϕ2 and Mϕ3 distributed sparsely. CD86 and CD206 were both upregulated in macrophages of RA synovium, especially in lining layer. Trajectory analysis showed that Mϕ1 existed at the start of the differentiation trajectory. HOXB6, STAT1, and NFKB2 were TFs specific to Mϕ1, Mϕ2, and Mϕ3 under RA condition, respectively. Compared with in OA condition, three macrophage clusters upregulated CXCL2, CXCL1, IL1B, TNFAIP3, ICAM1, CXCL3, PLAU, CCL4L2, CCL4, and TNF in NF-kappa B signaling pathway. The identification of macrophage subsets with different polarized states and their molecular signatures provided a more precise understanding of macrophages, which may contribute to the development of novel therapeutic strategy for RA.