Study on the role of integrin alpha4beta1 in bone metastasis

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1.Transfection of integrin alpha4beta1 cDNA to the cell : cDNA for integrin alpha4 (or alpha4 and beta1 both) was transfected into Chinese hamster ovary (CHO) cell. The expression of integrin alpha4beta1 on the transfected cell surface was confirmed by flow cytometry and immunoprecipitation.2.Biological analysis of the transfectant : Adhesion assay to VCAM-1 or fbronectin CS-1 peptide was examined in the transfected CHO cell (alpha4-CHO). As a result, alpha4-CHO cell was adhered both to VCAM-1 and CS-1 sufficiently.3.Metastasis experiment : alpha4-CHO or parent CHO cells were inoculated into nude mice. Only lung metastasis was observed in the nude mice in which CHO cells were inoculated, while bone metastasis was seen as well as lung metastasis in the nude mice in which alpha4-CHO cells were injected. Histological examination showed the proliferation of alpha4-CHO cells in the bone marrow and the invasion outside the bone in some cases. Clusters of tumor cells were sometimes found in the bone marrow, suggesting that bone marrow is the first emtastasis site. Bone metastasis was inhibited by the combination of anti-VCAM-1 and anti-alpha4 subunit. Since stroma cells in the bone marrow express VCAM-1 constitutively on the cell surface, alpha4-CHO cells might be adhered to the stroma cells and proliferated in the bone marrow. The tumor cells expressing integrin alpha4beta1 are supposed to metastasize to bone clinically, and the inhibition of metastasis will be expected by using neutralizing substances against integrin alpha4beta1 in the future.