Method for targeted inhibition of infiltration and transfer of glioma cells and application of method
A glioma cell and targeting technology, applied in the direction of tumor/cancer cells, animal cells, vertebrate cells, etc., can solve problems such as the unclear role of JAM2, achieve inhibition of invasion and metastasis, and expand the scope of targeted therapy , to overcome non-specific effects
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Embodiment 1
[0039] Example 1: JAM2 expression in human glioma tissue
[0040] In order to study the expression of JAM2 gene in glioma tissue, 60 surgical resection specimens of glioma patients were collected and a series of related experiments were done. First, the expression of JAM2 in glioma was analyzed by conventional immunohistochemical method, the results are shown in figure 1 .
[0041] Nuclei are stained blue by hematoxylin, showing cell number distribution. The yellow portion developed by DAB around the cells represents the expression of JAM2 molecules. The results showed that the stained positive tissue cells were arranged in disorder, with inconsistent compactness and looseness, and the shapes and sizes of the nuclei were different. They were overgrown tumor tissue cells, and the brownish yellow staining around the cells was also very rich. The yellow hollow oval part was the cross-section of blood vessels. , the tight junction molecule JAM2 was more abundantly expressed. T...
Embodiment 2
[0043] Example 2: Silencing glioma cell JAM2 expression by JAM2 siRNA gene transfection to inhibit cell migration experiment
[0044] The siRNA or shRNA plasmid targeting the JAM2 gene is mixed with a transfection reagent, transfected into glioma cells, and the expression of JAM2 is silenced, thereby inhibiting the proliferation, infiltration and metastasis of glioma cells. The transfection steps are as follows: Inoculate 106 / ml glioma cell U251 cells into a 6-well culture plate, add 2ml of 1640 culture solution and 10% fetal bovine serum to each well, and culture in a 5% CO2 incubator overnight. After attachment, when the cell attachment rate reaches over 60%, prepare a transfection mixture of transfection reagent and siRNA, such as 1 μl liposome plus 5 μl 2 μM siRNA, add it to the cells, continue to culture for 36-72 hours, and pass WESTERN The transfection efficiency was detected by BLOT, and the result shown was the knockout efficiency of JAM2 protein.
[0045] The proced...
Embodiment 3
[0048] Example 3: The experiment of inhibiting cell proliferation of glioma cell line U251 cells by lentivirus transfection of shRNA targeting JAM2.
[0049] The shRNA encoding JAM2 was coated with lentivirus and stably transfected into glioma cell U251 cells. The transfection steps were as follows: inoculate 106 / ml glioma cell U251 cells into a 6-well culture plate, and add 2ml of 1640 culture medium to each well solution and 10% fetal bovine serum, cultured overnight in a 5% CO2 incubator, after the cells adhered to the wall, when the cell adhesion rate reached more than 60%, the mixture of lentivirus and shRNA was prepared into the cells, and the culture was continued for 36 -72 hours, the transfection efficiency was detected by WESTERN BLOT, the result shown was the knockout efficiency of JAM2 protein, the effect on cell proliferation was detected by MTT method, and the effect on invasion was detected by cell streaking method. The results showed that after the JAM2 gene wa...
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