Insulin-like growth factor binding protein-4 accumulation is negatively correlated with growth rate in TM-3 cells

Cellular growth is controlled by multiple regulators, including the insulin-like growth factors (IGFs). In some cells, the IGF binding proteins (IGFBPs) are thought to be inhibitory molecules for cell growth and may be related to the process of contact inhibition. In the TM-3 (mouse Leydig) cell line, IGFBP-4 is the major IGFBP secreted into conditioned media (CM), as we have reported. In this study, we investigated cell growth, the peptide levels of IGFBP-4 in CM, and the inverse relationship between IGFBP-4 accumulation and cell growth rate. Quantification of TM-3 growth in serum-containing media demonstrated that TM-3 cell number gradually rose after plating, and plateaued when cells became confluent. The rate of cell growth fell gradually, and net cell growth stopped when cells reached confluency. IGFBP-4 peptide levels in CM, as measured by Western ligand blot, rose gradually during the culture period and plateaued when cells reached confluency. The amount of IGFBP-4 peptide level in CM correlated for cell number (IGFBP-4 accumulation rate) also rose gradually during the course of culture and plateaued. The IGFBP-4 accumulation rate was strongly negatively correlated with the rate of cell growth (r = 0.98, P < 0.001). In conclusion, our data suggest that in TM-3 cells, cell growth is related to IGFBP-4 accumulation. The negative correlation between IGFBP-4 accumulation and the rate of cell growth suggests that IGFBP-4 may be a primary regulator of TM-3 cell growth and possibly participate in the process of contact inhibition.