Aim To investigate the expression of microRNA-129 (miR-129) in serum of patients with chronic systolic heart failure (CSHF) and analyze its relationship with cardiac function indicators. Methods 103 patients with CSHF who were treated in Wuhan Sixth Hospital from May 2016 to October 2018 were selected as the observation group. According to the New York Heart Disease Assocation (NYHA) standard, the observation group was divided into three groups:NYHA class Ⅱ group (33 cases), NYHA class Ⅲ group (28 cases), NYHA class Ⅳ group (42 cases). In addition, 110 healthy people from Wuhan Sixth Hospital at the same time were selected as the control group. The left ventricular end systolic diameter (LVESD), left ventricular end diastolic diameter (LVEDD), left ventricular ejection fraction (LVEF) were measured by Philips iEElite type echocardiography, real-time fluorescence quantitative PCR (qRT-PCR) was used to detect the expression of miR-129 in serum, and the correlation between miR-129 and LVESD, LVEDD, LVEF and NYHA classification was analyzed. Results Compared with the control group, the LVESD and LVEDD increased significantly in NYHA class Ⅱ group, NYHA class Ⅲ group and NYHA class Ⅳ group (P＜0.05), while the LVEF and level of miR-129 decreased in turn (P＜0.05). There was no significant difference in serum miR-129 expression among CSHF patients with coronary heart disease, hypertension heart disease and senile heart valve disease(P＞0.05). The serum miR-129 expression of CSHF patients was negatively correlated with LVESD, LVEDD and NYHA grade (P＜0.05), and positively correlated with LVEF (P＜0.05). Conclusion miR-129 is low expressed in the serum of patients with CSHF. It is negatively related to the LVESD, LVEDD and NYHA grade, and positively related to LVEF, which may provide reference for the evaluation of CSHF.