Objective To investigate the expression of NADH-Cytochrome b5 reductase 2 (CYB5R2) in diffuse large B-cell lymphoma (DLBCL) and its clinical significance. Methods GO enrichment analysis was performed on DLBCL differentially expressed genes in TCGA database by DAVID online platform, and CYB5R2 was screened. The CYB5R2 expression of DLBCL in TCGA database and GTEX database was analyzed by bioinformatics method. The correlation between CYB5R2 expression and clinicopathological features and prognosis in 59 cases of DLBCL with complete follow-up data in our hospital was retrospectively analyzed. Results The expression of CYB5R2 in DLBCL was higher than that in normal lymphoid tissue (P < 0.05). The positive expression of CYB5R2 in the cases with elevated lactate dehydrogenase (LDH) was higher than that in the cases with normal LDH (χ2=4.832, P= 028). The positive expression of Non-GCB subtype cases was higher than that of GCB subtype cases (χ2=4.468, P= 035), and the positive expression of CD10-positive cases was higher than that of CD10-negative cases (χ2=4.468, P= 035). survival analysis showed that the 5-year overall survival (OS) of patients with high CYB5R2 expression was significantly lower than that of patients with zero or low CYB5R2 expression (χ2=4.799, P=0.028). The rate of complete response (CR) in patients with high expression was significantly lower than that of CYB5R2 expression was significantly lower than that of patients with zero or low CYB5R2 expression (χ2=4.799, P=0.028). The complete response (CR) rate in patients with high expression was significantly lower than that in patients with zero/low expression of CYB5R2 (χ2=4.015, P=0.045). Conclusion The high expression of CYB5R2 is associated with poor prognosis in DLBCL patients, suggesting that CyB5R2 plays an important role in the development of DLBCL.