| Objective:This paper explore the effects and related molecular mechanism of AFPm Ab-PLGA-rh DCN nanoparticles on migration and invasion of Hep G2 cells.Methods:The cultivate Hep G2 cells were randomly divided into A group, B group, C group, D group and E group. A group is the control group(untreated group) which is not any processing of Hep G2 cells. B group is the control group(blank group) without DCN gene nanoparticle processing. C group, D group and E group is the experimental group(experimental group) respectively with 5μg/ml(low concentration), 10μg/ml(concentration) and 15μg/ml(high concentration) AFPm Ab-PLGA- rh DCN nanoparticle processing.1 Effects of AFPm Ab-PLGA-rh DCN nanoparticles on migration and invasion of Hep G2 cells.1.1 The effects of migrate of Hep G2 cells is processed by AFPm Ab PLGA- rh DCN nanoparticle with scratch teste of different concentration.1.2 The effects of invasion of Hep G2 cells is processed by AFPm Ab PLGA- rh DCN nanoparticle with Transwell chambers of different concentration.2 The preliminary molecular mechanism research of AFPm Ab PLGA- rh DCN nanoparticles inhibit Hep G2 cell migration and the molecular.2.1 When Hep G2 cells processed by AFPm Ab PLGA- rh DCN nanoparticle 24 h, it detect AFP and Rho C m RNA transcription by rt-pcr method.2.2 When Hep G2 cells processed by AFPm Ab PLGA- rh DCN nanoparticle 24 h, it detect Rho C protein and Western Blot expression levels by ELISA method and Western Blot method.Results:1 Effects of AFPm Ab-PLGA-rh DCN nanoparticles on migration and invasion of Hep G2 cells.1.1 Scratches test show that the untreated group and the blank group cells scratched for 24 h, the scratch width were 308.62±5.89μm and 298.28±5.74μm, there is no significance difference(P> 0.05). The scratch width of experimental group with processed by AFPm Ab PLGA- rh DCN nanoparticles of different concentration are significantly higher than the control group.(The scrstch width of nanoparticles concentration were83.47±6.87μm 、 401.01±4.25μm 、 415.29±11.93μm),there is significant difference(P<0.001).It show that AFPm Ab-PLGA-rh DCN nanoparticles have inhibition to Hep G2 cell migration ability.1.2 Transwell Chambers experimental results show that membrane penetrating the cell numbers in Transwell cells of the untreated group and blank group were 70.67±1.16/hole and 66.67±2.08/hole. There is no significant difference(P>0.05). Compared with the untreated group and the blank group, the experimental group showed(membrane penetrating the cell numbers were62.00±3.61/hole、59.33±1.53/hole and 53.33±2.08/hole from low penetration to high penetration of nanoparticles concentration).There is significant difference(P<0.001).It showed that AFPm Ab-PLGA-rh DCN nanoparticles have inhibition to Hep G2 cell invasion ability.2 The preliminary study about molecular mechanism of AFPm Ab-PLGA-rh DCN nanoparticles inhibit migration and invasion of Hep G2 cell2.1 The m RNA transcription of AFP and Rho C molecular2.1.1 The m RNA transcription of AFP molecularThe gray analysis results show that Hep G2 cells AFP m RNA grey value of the untreated group and blank group were1.00±0.03 and 1.05±0.08.There is no significant difference(P>0.05). Experimental group(nanoparticles concentration from low to high)cells AFP m RNA grey value is lower than the control group(0.93±0.07,0.68±0.04 and0.57±0.03). Compared with low concentration group and the control group, there is no significant difference(P>0.05). Compared with medium concentration group, high concentration group and the control group, there is significant difference(P< 0.05).2.1.2 The m RNA transcription of Rho C molecularThe gray analysis results show that RhoC m RNA grey value of the untreated group and blank group cell were 1.02±0.04、1.03±0.07, there is no significant difference(P>0.05). The experimental group(nanoparticles concentration from low to high) cells Rho C m RNA grey value is lower than the control group 0.42±0.04、0.38±0.02 and 0.38±0.02),there is significant difference(P< 0.001).2.2 The protein expression of AFP and Rho C2.2.1 The results of ELISA detection showed that AFP protein concentration of untreated group and blank group were 2.40±0.25 and 2.51+ 0.31, there is no significant difference(P>0.05). The experimental group(nanoparticles concentration from low to high)cells AFP protein concentration is lower than the control group 2.01±0.21ng/ml,1.01±0.11 ng/ml and 0.49±0.07 ng/ml), there is significant difference(P< 0.05).2.2.2 The results of ELISA detection showed that Rho C protein concentration of in untreated group and blank group Rho C cells were 98.76±3.04pg/ml and 100.75±4.19pg/ml,there is no significant difference(P>0.05). The experimental group(nanoparticles concentration from low to high) cell Rho C protein concentration is lower than control group(86.63±2.60 pg/ml,47.37±2.51 pg/ml,39.78±2.47 pg/ml),there is statistically significant difference(P<0.001).2.2.3 The results of Western Blot detection Rho C showed that Rho C gray values of untreated group and blank control group cells were1.02±0.05 and 1.02±0.03, there is no significant difference(P>0.05). The experimental group(nanoparticles concentration from low to high) cell Rho C protein concentration is lower than control group(0.85±0.01,0.70±0.02, 0.62±0.02), there is significant difference(P<0.05).Conclusion:After treatment of AFPm Ab-PLGA-rh DCN nanoparticles on Hep G2 cells, the ability of cell migration and invasion decreases in different degrees, decreasing in a dependent trend with the dose. It can cut down the expression and participation of AFP and Rho C molecule. |
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