Objective To investigate the correlations of serum eukaryotic translation promoter 2α (eIF2α) and activated transcription factor 4 (ATF4) levels with the degree of renal tissue injury and renal function in patients with diabetic nephropathy (DN). Methods A total of 102 patients with DN (DN group) and 102 patients with simple diabetes (control group) were selected. According to the severe degree of renal tissue damage, the patients with DN were divided into microalbuminuria group (MG group, 35 cases) and dominant albuminuria group (PG group, 41 cases) and renal dysfunction group (RIG group, 26 cases). Serum levels of eIF2α, ATF4, urea nitrogen (BUN), creatinine (Scr), cystatin C (CysC) and estimated glomerular filtration rate (eGFR) were measured. Pearson correlation analysis was used to explore the correlations of eIF2α and ATF4 with BUN, Scr, CysC and eGFR; multivariate Logistic regression analysis was used to explore the risk factors of DN; receiver operating characteristic (ROC) curve was used to analyze the value of eIF2α and ATF4 in diagnostic of DN. Results Serum levels of eIF2α, ATF4, BUN, Scr and CysC in the DN group were higher than those in control group, eGFR was lower than that in control group (P < 0.05). Serum eIF2α and ATF4levels in the RIG group were higher than those in PG and MG groups (P < 0.05). Serum eIF2α and ATF4 levels in DN patients were positively correlated with BUN, Scr and CysC, and negatively correlated with eGFR (P < 0.05). Long duration of diabetes, higher body mass index, high level of eIF2α and high level of ATF4 were risk factors for DN (P < 0.05). The area under the curve of eIF2α and ATF4 in diagnosis DN was 0.770 and 0.799, respectively, and the area under the curve of their combined diagnosis was 0.879, which was higher than that of single diagnosis (P < 0.05). Conclusion Serum levels of eIF2α and ATF4 are increased in patients with DN, which is related to the severity of renal injury and renal dysfunction in DN. The combination of eIF2α and ATF4 is of high value in the diagnosis of DN.