OBJECTIVE: Pulmonary arterial hypertension (PAH) is a serious consequence of congenital heart disease (CHD). PAH is characterized by a cancer-like pro-proliferative and anti-apoptotic phenotype of pulmonary artery smooth muscle cells (PASMCs). Never in mitosis a-related kinase 2 (NEK2) has recently been identified as a key factor in tumor cell proliferation and migration whlie the functional importance of NEK2 in PAH associated with CHD (CHD-PAH) has not been elucidated yet. METHODS: NEK2 expression in the pulmonary arterioles of rats with CHD-PAH and shunt-related PAH was evaluated. For this purpose, human PASMCs (hPASMCs) were transfected with lentiviruses for NEK2 knockdown or overexpression, and changes in expression of phenotypic markers in hPASMCs were determined. The proliferation, migration and apoptosis abilities of hPASMCs were respectively detected. Changes in vascular remodeling following NEK2 suppression were also observed in the shunt-related PAH rat model. RESULTS: NEK2 was found to be highly expressed both in the PASMCs of the middle pulmonary arterioles of patients with CHD-PAH and shunt-related PAH rats. Additionally, overexpression of NEK2 enhanced phenotypic switch, proliferation, migration, and apoptosis resistance of hPAMSCs by activating the nuclear factor kappa B pathway. Moreover, hemodynamic parameters and pulmonary vascular remodeling were both found to be improved considerably following suppression of NEK2 expression by intratracheal instillation of adeno-associated virus in shunt-related PAH rats. CONCLUSION: We demonstrated for the first time that NEK2 is a potential regulator of PASMCs function. Targeting NEK2 may be an effective strategy for the treatment of CHD-PAH.