Research On Electrotaxis Of Three Type Lung Adenocarcinoma Cell Lines With Different Source

Metastasis is the crucial factor for most cancer-related deaths and low survival.The directional migration of tumor cells plays an important role in tumor,metastasis.The endogenous electric fields are ubiquitous in nature.Numerous investigations suggested that tumor metastasis was the resulted from abnormal change of endogenous electric fields.The electrotaxis of cells is closely asscociated with tumor metastasis.The electrical signal dominates the directional migration.Lung cancer now is the leading cause of death for cancer-related death in worldwide,while China and Yunnan province holds the highest incidence of Lung adenocarcinoma around the world and in China,respectively.Three different sources of lung adenocarcinoma cell lines,namely PG49,A549,Calu-3 and human bronchial epithelial cells(HBE)as control were chosen to study the different response to applied small direct current field through Time-lapse microscopy.Meanwhile,we also investigated influencing factors for the eletrotaxis of lung adenocarcinoma cells,including the cell cluster,nutritional supply status and pseudopod.Our findings may provide detailed data for tumor metastasis mechanisms and theoretical basis for the application of cancer electro-diagnosis and treatment.1)Electrotaxis is founded in all of four cell lines.The response of four cell lines to electric fields of 0,2,4,6v/cm were analyzed quantitatively by four parameters.All of four cell lines changed model from random migration to cathode directed migration after electric fields applied.There was a significant electrotaxis of HBE,Calu-3,PG49 when electric fields was 4 v/cm,and A549 had a significant directedness when 6 v/cm.In applied electric fields,HBE and Calu-3 had a higher stability of directed migration toward cathode than no electric field.At 2v/cm,the persistence index of A549 and PG49 reached to the highest.The migration speed of cells also could be increase when electric fields were applied.The highest migration speed of Calu-3 and PG49 was at 2 v/cm and 4 v/cm,respectively,while both of HBE and A549 move with fastest speed at 6 v/cm.The orientation of directed migration among four different cells was same,but the track speed anddisplacement speed of PG49,A549,Calu-3 were significantly higher than HBE..These results showed that the electrotaxis could be common in lung adenocarcinoma cell lines.The higher speed of tumor cells may be the base of their abnormal metastasis ability.2)Both of isolated cell and clustered cells of Calu-3 were sensitive to electric field,but there were different between their electrotaxis ability.We analyzed the effect of direct current electric fields(dc EFs)on lung adenocarcinoma cells,Calu-3 through Time-lapse microscopy.The cells without electric field were taken as the control group.The cells of experimental group were exposed to dc EFs with intensity of 2v/cm,4v/cm and 6v/cm,respectively.The cells of Calu-3 migrated through two way,including isolated cell and clustered cells.Our data showed that dc EFs had important effect on the migrations of Calu-3.Both isolated cells and clustered cells of Calu-3 moved randomly in the absence of an EF.In the presence of dc EFs with different strengths,both the two type cells migrated toward the cathode electrode of electric field.The parameters of migration ability,including directedness index,persistence index,track speed and displacement speed had significant differences between control and experimental groups.Generally,the dc EFs make Calu-3 cells possessed more obvious directional ability and faster speed.Both isolated and clustered cells achieved maximal electrotaxis and movement speed in 4v/cm EFs.The clustered cells of Calu-3 exhibited greater sensitivity to dc EFs but ran more slowly than the isolated cells under EFs with same strength.3)A549 in serum-free medium showed higher electrotaxis with more obvious directedness and faster speedTo explore the effect of serum on electrotaxis of A549,we applied the electric field with strength 0?2?4?6v/cm to A549 cells in serum-containing and serum-free medium,respectively.The results showed that serum didn't affect the electrotaxis orientation of A549.Both of them changed model from random migration to cathode directed migration after electric fields applied.When the electric field intensity was6v/cm,the electrotaxis index and speed of them reached peak value.However,A549 in serum-free medium exhibited more sensitive to cathode with higher electrotaxis index and faster speed(displacement speed and track speed)than that of A549 inserum-containing medium at same electric strength.These results suggest that the absence of the serum does not result in the loss of the ability of the cells to migrate,but promotes the migration of the cells in the electric field.4)Electric field had effect on pseudopod of A549We did the comparisons on pseudopod of A549 cells in absence of electric field and in 6 v/cm electric field.The results were as followed: there were no significant difference in pseudopod number and formation speed of A549 under electric field with different strength.However,pseudopod of A549 obviously extended toward the cathode of electric field.The ratio of splitting pseudopod significantly increased.Furthermore,we found that the ratio of splitting pseudopod in the cell area closed to cathode of electric field was significantly higher than that closed to anode of electric field.The same trends were also detected in number and formation speed of pseudopod when A549 cells were exposed in the electric field.Our results accorded with the predictions from pseudopod-centered model.It was indicated that electric signal influenced formation way,distribution,spread orientation of pseudopod instead of creating new pseudopod to keep their electrotaxis sensitivity.