| Stem cell therapy has become essential for the treatment of refractory diseases,including cancer and tumor.However,the safety of stem cells has not been confirmed and the interaction between stem cells and tumor cells remains controversial.In this paper,three different types of stem cells were co-cultured with human lung adenocarcinoma A549 cells to study the induction effects of stems cells on A549 cells.First,we establish a two-dimensional Transwell co-culture model to study the effects of three kinds of stem cells on the deterioration and metastasis of A549 cells.In order to eliminate the interference caused by serum and the additive in the medium of the co-culture system,we set up the complete medium and basic medium co-culture system respectively to study the changes of morphology,proliferation and invasion ability of A549 cells after co-cultured with stem cells,as well as the changes of the expression in genes related to EMT.Secondly,we set up a 3D tumor invasion model to further verify the induction effect of hESCs on human A549 cells.Finally,we design new driver microfluidic chip and culture four kinds of cells on it respectively to find the effects of flow.We expect that we can study the induction effects of stem cells on A549 cells under the influence of microfluidic in the future.The morphology of human A549 cells changed from polygonal to long spindle after co-cultured with hESCs.The increased expression of MMP2 and MMP9 genes indicates that the hESCs promoted the EMT process of human A549 cells.However,it was found that the proliferation ability of A549 cells was weakened after co-cultured with hESCs.Besides,in the Transwell co-culture model of hESCs and human A549 cells experiment,A549 cells were stained as irregular polygon after penetrating the membrane,we speculat that human A549 cells might aggregate into spheroids at first.Due to contact inhabitation,the proliferation rate decreased.Then A549 cells penetrate the 8μm Transwell membrane as irregular polygon.The 3D tumor invasion experiments showed that hESCs induced A549 cells to penetrate through the matrix gel and migrate to the side of hESCs.If the mimic tumor is far from the hESCs in matrigel(>1200μm),the development is from the centre and centrosymmetric,with no evident effect from the hESCs in matrigel.When the distance between hESCs in matrigel and the mimic tumor is between 600μm and 1200μm,the development centre moves to the direction of hESCs,and the mimic tumor development is eccentric on the effect of adjacent hESCs in matrigel.If the distance is shorter than 600μm,the mimic tumor A549 cells moves faster and even the hESCs break their matrigel and move to the mimic tumor.The morphology of human A549 cells changed from polygonal to long spindle after co-cultured with rat C3H10T1/2 cells and hUCMSCs.The proliferation and invasion ability of human A549 cells are improved to a certain extent.The increased expression of MMP2 genes indicates that rat C3H10T1/2 cells and hUCMSCs promoted the EMT process of human A549 cells.Furthermore,rat C3H10T1/2 cells and hUCMSCs promote the deterioration of human A549 cell metastasis.Four kinds of cells were cultured on a microfluidic chip to study the flow effects on ste cells and tumor cells respectively.At first,we further reduce velocity of liquid flow to lower than 10μm/s by using parallel U-microchannels model.In addition,we confirm that under the same channel depth condition,the liquid flow rate in the channel is proportional to the channel width(W),the distance of the U-shape ends(M)and the reciprocal of the channel length(1/L)by using series U-microchannels model,and the flow is stable.The new driver microfluidic chip independently developed by our research group can well avoid the limitations of the traditional chips.The experiments show that the four kinds of cells all grow well on the chip respectively.The growth rate of human hESCs in the flow driven by 15Hz(1min/h)grew about 1.3 times faster than that in the static state. |
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