Objective. MiR-133b was dysregulated in myocardial infarction. However, the role and mechanism of miR-133b in myocardial infarction remains unclear. This study was aimed to explore the role of miR-133b in H9c2 cell injury induced by hypoxia and to investigate the underlying molecular mechanism. Methods. Cell injury was assessed by cell viability, migration, invasion, and apoptosis assays. The expression of miR-133b and nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) mRNA was determined by qRT-PCR. The levels of apoptosis-related proteins and NLRP3 were detected by western blotting. Results. Results showed that hypoxia significantly reduced cell viability, migration, and invasion, but increased apoptosis of H9c2 cells. Downregulation of miR-133b aggravated the cell injury induced by hypoxia. MiR-133b was directly targeted on NLRP3. Overexpression of NLRP3 significantly inhibited cell viability, migration, and invasion but induced cell apoptosis in H9c2 treated with hypoxia. Conclusions. Thus, miR-133b protects H9c2 against hypoxia injury via downregulation of NLRP3.