Accumulating studies have demonstrated serum exosomal microRNAs (miRNAs) represent novel biomarkers for various diseases. In this study, we aimed to explore the feasibility of using serum exosomal miRNAs as novel serological biomarkers for steroid-induced osteonecrosis of femoral head (SONFH). We identified the characters of exosomes which were obtained from fresh serum of 5 systemic lupus erythematosus (SLE) patients without SONFH, 5 SLE patients with SONFH (SLE-SONFH) and 5 healthy ones. Comprehensive exosomal miRNA sequencing was performed to profile the differentially expressed miRNAs in the three groups. We then validated the expression levels of selected miRNAs by qRT-PCR. Furthermore, KEGG pathway, GO annotation, protein-protein interaction (PPI) network, module analysis and miRNAs-mRNAs interaction network were built to analyze the potential targets and mechanism. Sequencing data conveyed that hsa-miR-135b-5p, hsa-miR-150-5p, hsa-miR-509-3-5p, hsa-miR-514a-3p and hsa-miR-708-5p were significantly differentially expressed in the three groups. The results of qRT-PCR for the first time confirmed that the expression of hsa-miR-135b-5p was strikingly up-regulated in SLE-SONFH group which were consistent with miRNA sequencing results. In addition, bioinformatics analysis indicated that the enriched functions and pathways of the most differentially expressed miRNAs including Wnt, MAPK as well as Hippo signaling pathway. The top five hub genes (FGF2, PTEN, HACE1, VAMP2, and CBL) were part of module of the PPI network, which consisted of 713 nodes and 2191 edges. In conclusion, this study provides a novel and fundamental serum exosomal miRNAs profile of SONFH and hsa-miR-135b-5p may be identified as a unique diagnostic biomarker for SONFH. AJTR Copyright © 2020.