Small heat shock proteins (HSPs) regulate a variety of cell functions. Among them, HSP22 and HSP20 are recognized to be ubiquitously expressed in various tissues. With regard to hepatocellular carcinoma (HCC) cells, we previously reported that phosphorylated HSP20 plays a suppressive role in transforming growth factor (TGF)alpha-induced cell migration and invasion. In the present study, we investigated whether or not HSP22 is implicated in HCC cell migration. We detected HSP22 protein expression both in human HCC tumor (189.9 +/- 68.4 ng/mg protein) and the adjacent non-tumor liver tissues (167.9 +/- 94.6 ng/mg protein). The cases of low-quantity HSP22 protein level group (88.3 >= ng/mg protein, the optimum cut-off value of HSP22) were increased in tumor tissues compared with the adjacent non-tumor tissues. The migration of human HCC-derived HuH-7 cells stimulated by TGF-alpha or hepatocyte growth factor (HGF) was significantly enhanced by the knockdown of HSP22 expression. Down-regulation of HSP22 protein in the cells markedly strengthened the AKT phosphorylation induced by TGF-alpha or HGF. Inhibitors of the phosphoinositide 3-kinase (PI3K)/AKT pathway, which suppressed the TGF-alpha-induced migration, significantly reduced the amplification by HSP22 knockdown. PI3K but not AKT was coimmunoprecipitated with HSP22 in HuH-7 cells. In addition, in human HCC tissues, a significantly lower HSP22 protein level in tumor tissues than in adjacent non-tumor tissues was observed more frequently in cases of moderately or poorly differentiated HCC than well-differentiated HCC. Taken together, our results strongly suggest that HSP22 represses HCC progression, especially HCC cell migration, by the down-regulation of the PI3K/AKT signaling pathway.