Pulmonary arterial hypertension, a chronic lung disease, remains an unacceptable prognosis despite significant advances in conventional therapies. Stem cell therapy represents a novel and effective modality. This study was aimed to add new insight in gender differences of bone marrow-derived mesenchymal stem cells on therapy against pulmonary arterial hypertension and the underlying mechanism. By in vivo experiments, we showed for the first time female bone marrow-derived mesenchymal stem cells possessed a better therapeutic potential against monocrotaline-induced pulmonary arterial hypertension in C57BL/6J mice compared with male counterparts. In vitro experiments demonstrated superior function of female bone marrow-derived mesenchymal stem cells in cell proliferation, migration and [Ca-2](i) kinetics. Moreover, we unexpectedly found that, compared with male ones, female bone marrow-derived mesenchymal stem cells had a higher expression level of glyceraldehyde-3-phosphate dehydrogenase and manipulations of its expression in female or male bone marrow-derived mesenchymal stem cells profoundly affected their cellular behaviours and therapeutic efficacies against pulmonary arterial hypertension. Our results suggest that glyceraldehyde-3-phosphate dehydrogenase plays a critical role in determining the superior functions of female bone marrow-derived mesenchymal stem cells in cell therapy against pulmonary arterial hypertension by regulating [Ca-2](i) signal-associated cellular behaviours.