Figure 8

Exogenously supplied CCN3 promoted aRMS cell adhesion, migration and invasion. (a) The adhesion assay was performed by measuring the amount of RH4 cells attached to the plastic surface of microtiter plate with or without pre-coating of indicated proteins. Col1: collagen 1; FN: fibronectin; VN: vitronectin. (b) RH4 cells adhesion to CCN3-coated microplate was blocked selectively by antibodies against β1 and β5 integrins. RH4 cells binding to the control bovine serum albumin-coated plates with or without antibody treatment were similar at an absorbance below 0.05 (data not shown). Inset: western blot analysis of β-integrin and α-tubulin expression in eRMS (RD) and aRMS (RH4, RH28 and RH30) cells. (c) CCN3 enhanced migratory activity of RH4 cells as determined by wound scratch assay. Representative light microscopic photographs (100 × ) were taken 48 h after the scratch was created. (d) Matrigel invasion assay was performed on RH4 cells by using Matrigel-coated filters and invasion chambers containing low (0.5%) and high (10% FBS) serum, and low serum containing 50 μg/ml CCN3 protein.