Kashin-Beck disease (KBD) is a chronic osteochondral disorder primarily associated with cartilage degeneration. The bone texture structure in KBD was also changed but it was not identical to primary knee osteoarthritis (OA). This study investigates the differences in microRNA (miRNA) profiles of subchondral bone collected from patients suffering from KBD in comparison with those with primary knee osteoarthritis (OA). Subchondral bone tissues were taken from four patients with KBD and four patients with primary knee OA undergoing total knee replacement. The miRNA array profiling was performed using an Affymetrix miRNA 4.0 Array, and then the target gene predictions and function annotations of the predicted targets were performed. Our results showed that 124 miRNAs had lower expression levels in the subchondral bone sampled from KBD patients in comparison with OA patients. Gene ontology (GO) and KEGG pathway analyses of the predicted targets demonstrated numerous significantly enriched GO terms and signal pathways essential for bone development and integrity, such as metabolic processes, PI3K-Akt, and MAPK signaling pathways. Our study confirms that a large set of miRNAs are differentially expressed in the subchondral bone of patients with KBD and OA and contributes new insights into potential pathological changes in the subchondral bone of KBD patients.