Effect Of HPD Combined With 630nm Laser On Vasculogenic Mimicry In Human Lung Adenocarcinoma A549 Cell In Vitro

Purpose: To observe whether photodynamic therapy has any effect on the formation of vasculogenic mimicry(VM)in human lung adenocarcinoma A549 cell in vitro,and to detect the expression of related genes osteopontin(OPN)and vascular endothelial growth factor(VEGF).Methods: Human lung adenocarcinoma cells A549 were cultured in vitro,and A549 cells with logarithmic growth phase and good growth status were randomly divided into4 groups: blank control group,photosensitizer group,laser group,and photodynamic group.(1)Each group of cells was cultured in three dimensions on Matrigel,and the vasculogenic mimicry generation of each group was observed and compared by inverted microscopy.(2)PAS staining was used to demonstrate that the groups of cells formed vasculogenic mimicry rather than endothelium-containing vascular structures.(3)The expression of OPN and VEGF m RNA,two genes associated with vasculogenic mimicry in each group,was detected by reverse transcription-polymerase chain reaction(RT-PCR).(4)The expression of OPN and VEGF proteins in each group was detected by Western Blot.Results:(1)Observation by inverted microscopy showed that vasculogenic mimicry could be observed in A549 cells cultured on Matrigel for about 6h,and the number of formation was significantly reduced in the photodynamic group compared with blank control group,photosensitizer group and laser group,with statistically significant differences(P<0.05,P<0.01,P<0.05).(2)PAS staining results showed that a tubular network of closely packed A549 cells surrounding each other(without endothelial cells)positive for PAS staining could be seen under the microscope.(3)RT-PCR results showed that the photodynamic group,i.e.,using hematoporphyrin derivatives + 630 nm laser applied to lung adenocarcinoma A549 cells for 24 h,downregulated the expression of genes OPN m RNA compared with the blank control group,photosensitizer group and laser group,with statistically significant differences(P<0.05,P<0.05,P<0.01),and also downregulate the expression of VEGF m RNA with statistically significant differences(P<0.01,P<0.001,P<0.001),and there was no difference between the blank control,photosensitizer and laser groups statistically significant.(4)Western Blot results showed that the photodynamic group,i.e.,using hematoporphyrin derivatives + 630 nm laser acting on lung adenocarcinoma A549 cells for 48 h,down-regulated the expression of OPN proteins compared with blank control group,photosensitizer group and laser group,and the difference was statistically significant(P<0.001,P<0.001,P<0.001),and also downregulate the expression of VEGF proteins with statistically significant differences(P<0.001,P<0.001,P<0.0001),and there was no difference between the blank control,photosensitizer and laser groups statistically significant.Conclusions: Photodynamic therapy significantly inhibited the formation of vasculogenic mimicry in human lung adenocarcinoma A549 cells in vitro.In addition,photodynamic therapy also differentially downregulated the expression of vasculogenic mimicry-related genes OPN,VEGF m RNA and protein levels.Therefore,inhibition of angiogenic mimetic formation in A549 cells in vitro by photodynamic therapy may be associated with downregulation of OPN and VEGF gene expression,and downregulation of OPN expression by photodynamic therapy may also be achieved through downregulation of VEGF,the mechanism of which needs to be further investigated.