Invadopodia Formation is a Critical Step in Cancer Cell Invasion: The Effect of Passage Number on Invadopodia Formation in MDA-MB-231 Breast Cancer Cell Line

Abstract

High invasive cancer cells are thought to recruit specialised actin-rich protrusions for invasion in metastasis process. These protrusions are termed invadopodia. To study invadopodia formation, one of the first challenges faced by researchers has been to optimise the cell line passage number in order to be used for the invadopodia assay. Therefore, this study aims to investigate the effects of the passage number on invadopodia formation in MDA-MB-231 breast cancer cell line. Invadopodia assay was used to achieve the aim of the study. The results provided evidence that invadopodia formation is affected by the high passage number. The cells were also tested with dimethyloxalylglycine (DMOG) a hypoxic mimicking agent which is known to be an invadopodia inducer, the results showed that the cells in low passage number (P7) treated with DMOG increase the cells forming invadopodia, while the cells with high passage number (P35) showed that DMOG fails to stimulate the cells to form invadopodia. Furthermore, the cells with high passage number after passage 15 are starting to lose the ability to degrade the gelatin. In conclusion, this study suggests that only cells with a low passage number, less than passage 15 should be used in the study of invadopodia formation to obtain the results in the search for molecular targets and signaling at invadopodia.

Passage Number of 4T1 Cells Influences the Development of Tumour and the Progression of Metastasis in 4T1 Orthotopic Mice

“…Previous studies have reported the role of passage number in the progression of invasion and metastasis. A study by Hamad, Kqueen and Hashim ( 18 ), reported that the difference in the passage number affects the formation of invadopodia. Invadopodias are specialised actin-rich protrusions recruited by cancer cells to perform an invasion into the surroundings ( 19 ).…”

“…Invadopodias are specialised actin-rich protrusions recruited by cancer cells to perform an invasion into the surroundings ( 19 ). Hamad, Kqueen and Hashim ( 18 ) reported that the formation of invadopodia in MDA-MB-231 cells is influenced by the passage number of the cells. The study added that the lower passage number (P7) exhibited a higher percentage of cells that formed invadopodia as compared to the cells of higher passage number (P35).…”

“…The study added that the lower passage number (P7) exhibited a higher percentage of cells that formed invadopodia as compared to the cells of higher passage number (P35). The paper concluded that cells of lower passage number (less than P15) should be an ideal choice in studies related to the formation of invadopodia ( 18 ). This statement implies that cells with higher passage number could miss their signaling and molecule components that are responsible for the formation of invadopodia formation, in order to carry out invasion.…”

“…Besides Hamad, Kqueen and Hashim ( 18 ), several other studies have also associated passage number with the invasion and metastatic capacity of cells. A study by Urtreger et al ( 20 ) highlighted that the local invasive nature as well as the spontaneous metastatic capacity of LM3 cells inoculated subcutaneously increased with the passage number and were associated with progressive loss of fibronectin.…”