This study was set to determine the expression of microRNA-449a in cancer tissue and serum of non-small cell lung cancer (NSCLC) patients and explore the underlying mechanisms of its tumor suppressor functions. We selected 50 NSCLC patients in our hospital as the lung cancer group and 50 healthy volunteers as the control group. RT-PCR was performed to detect the expression levels of microRNA-449a in NSCLC tissue and the plasma of NSCLC patients. Further, we transfected microRNA-449a mimic and inhibitor in lung cancer cell line A549, and used western blot to determine the expression level of apoptosis-related molecules Bcl-2 and p53. Compared with the surrounding tissue, microRNA-449a exhibited significantly reduced mRNA expression, which was statistically different (P < 0.05). microRNA-449a exhibited significantly lower expression in NSCLC patients' plasma than the healthy volunteers, which was statistically different (P < 0.05). Spearman correlation analysis showed that microRNA-449a expression levels in NSCLC tissue and plasma of NSCLC patients were reversely correlated with lung cancer differentiation (P < 0.05). But microRNA-449a expression in the surrounding tissue was not significantly correlated with lung cancer differentiation (P > 0.05). Compared with negative control, cell proliferation and p53 and Bcl-2 expression significantly decreased after microRNA-449a mimic transfection (P < 0.05). However, after transfection of microRNA-449a inhibitor, cell proliferation and p53 and Bcl-2 expression significantly increased after microRNA-449a mimic transfection (P < 0.05). microRNA-449a expression levels in NSCLC are significantly lower than those in the surrounding tissue, and its expression levels in NSCLC patients are also lower than those in healthy volunteers. The tumor suppression role of microRNA-449a could be due to its promotion of tumor cell proliferation and its inhibition of tumor cell apoptosis.