Aim To investigate whether reactive oxygen species (ROS) were involved in cobalt chloride (CoCl₂) induced cellular proliferation in human pulmonary artery smooth muscle cells (HPASMC). Methods HPASMC were exposed to a chemical hypoxia agent CoCl₂ to establish a cellular model of pulmonary arterial hypertension. An exogenous ROS donor, hydrogen peroxide (H₂O₂), was administered to examine the direct effect of ROS on HPASMC proliferation. Before the exposure of HPASMC to CoCl₂ or H₂O₂, a ROS scavenger, N-acetylcysteine (NAC), was used to assess the effect of inhibitory oxidative stress on the cellular proliferation induced by the two agents above. Cell proliferation was measured by cell counter kit 8, expression of hypoxia inducible factor-1α (HIF-1α) was tested by Western blot assay and intercellular ROS were observed by 2′,7′-dichlorfluorescein-diacetate staining followed by photofluorography. Results Treatment of HPASMC with CoCl₂ for 24 h at concentrations ranging from 25 to 50 μmol/L induced significant cellular proliferation, and treatment with 50 μmol/L CoCl₂ for 24 h obviously increased intercellular HIF-1α level. Similarly, treatment with H₂O₂ for 24 h at concentrations ranging from 12 to 25 μmol/L triggered cellular proliferation, however, treatment with H₂O₂ didn’t alter HIF-1α level in HPASMC. Pretreatment with NAC, prior to the treatment with CoCl₂ or H₂O₂, statistically attenuated H₂O₂-induced HPASMC proliferation (P<0.05), but not CoCl₂-induced cellular proliferation (P>0.05). Exposure of HPASMC to 50 μmol/L CoCl₂ for 6-24 h didn’t alter intracellular ROS content. Conclusion ROS may not be involved in CoCl₂-induced cellular proliferation in HPASMC.