RhoA plays an important role in Ca²⁺ sensitization of bronchial smooth muscle in antigen-induced airway hyperresponsiveness (AHR). Glucocorticoids are now the most effective anti-inflammatory treatment for asthma, and inhaled corticosteroids are the most effective long-term control therapy for persistent asthma. To determine the mechanism of the inhibitory action of glucocorticoids on AHR in allergic bronchial asthma, that of prednisolone on RhoA upregulation was investigated using cultured human bronchial smooth muscle cells (hBSMCs). The upregulation of RhoA induced by interleukin (IL)-13 and tumor necrosis factor (TNF)-α, major mediators for development of AHR, was observed in hBSMCs. Prednisolone partly inhibited the IL-13-induced RhoA upregulation and RhoA promoter activity, although prednisolone had no effects on the activations of signal transducers and activators of transcription (STAT)6 and nuclear factor (NF)-κB. Increased expression and promoter activity of RhoA induced by TNF-α was completely inhibited by prednisolone, although the activation of NF-κB failed to be inhibited by prednisolone in hBSMCs. These findings suggest that prednisolone might inhibit NF-κB-induced transcription via interaction between glucocorticoid receptor (GR), resulting in an inhibition of RhoA upregulation induced by IL-13 and TNF-α.