The aim of our study was to examine the production of monocyte chemoattractant protein (MCP-4) and eotaxin-3 during the onset and progression of COPD. The expression levels of MCP-4 and eotaxin-3 were evaluated in COPD samples and healthy controls using immunostaining and ELISA. The relationship between the clinic pathological features in the participants and the expression of MCP-4 and eotaxin-3 were evaluated. The association of MCP-4/eotaxin-3 production in COPD patients was also determined. The results revealed enhanced production of MCP-4 and eotaxin-3 in COPD patients especially the cases with AECOPD in both bronchial biopsies and bronchial washing fluid samples. Furthermore, the expression signatures of MCP-4/eotaxin-3 show high AUC values in distinguishing COPD patients and healthy volunteers and AECOPD and stable COPD cases, respectively. Additionally, the number of MCP-4/eotaxin-3 positive cases was notably increased in AECOPD patients compared to those with stable COPD. Moreover, the expression of MCP-4 and eotaxin-3 was positively correlated in COPD and AECOPD cases. In addition, the levels of MCP-4 and eotaxin-3 could be increased in HBEs stimulated with LPS, which is a risk factor of COPD. Moreover, MCP-4 and eotaxin-3 may exert their regulatory functions in COPD by regulating CCR2, 3, and 5. These data indicated that MCP-4 and eotaxin-3 were potential markers for the clinical course of COPD, which could provide guidance for accurate diagnosis and treatment for this disease in future clinical practice.