microRNA (miR)-22 has been reported to be downregulated in hepatocellular, lung, colorectal, ovarian and breast cancer, acting as a tumor suppressor. The present study investigated the potential effects of miR-22 on gastric cancer invasion and metastasis and the molecular mechanism. miR-22 expression was examined in tumor tissues of in 89 gastric cancer patients by in situ hybridization (ISH) analysis. Additionally, the association between miR-22 levels and clinicopathological parameters was analyzed. A luciferase assay was conducted for target identification. The ability of invasion and metastasis of gastric cancer cells in vitro and in vivo was evaluated by cell migration and invasion assays and in a xenograft model. The results showed that miR-22 was downregulated in the gastric cancer specimens and significantly correlated with the advanced clinical stage and lymph node metastasis. In addition, metadherin (MTDH) was shown to be a direct target of miR-22 and the expression of MTDH was inversely correlated with miR-22 expression in gastric cancer. Ectopic expression of miR-22 suppressed cell invasion and metastasis in vitro and in vivo. The present study suggested that miR-22 may be a valuable prognostic factor in gastric cancer. miR-22 inhibited gastric cancer cell invasion and metastasis by directly targeting MTDH. The novel miR-22/MTDH link confirmed in the present study provided a novel, potential therapeutic target for the treatment of gastric cancer.