The association of transcription factor Prox1 with the proliferation, migration, and invasion of lung cancer

Abstract

Background: The current study investigates the effect of transcription factor Prox1 on the proliferation, migration, and invasion ability of lung cancer.

Methods: Lung cancer cell lines (A549 and H446 cells) were transfected with Prox1NAD and siRNA, respectively. Thus, the A549 and H446 cells overexpressed Prox1 after transfection of Prox1NAD plasmids, and A549 and H446 cells have low expression of Prox1 after transfection with siRNA. Reverse transcriptase quantitative PCR and western blot analyses were used to detect Prox1 mRNA and protein expression in cells. Plate clone formation experiments and MTT experiments were used to detect cell proliferation. Western blot was used to detect the expression of Rho family-related proteins in cells.

Results: Compared to untransfected wild-type A549 and H446 that served as blank controls, the expression level of Prox1mRNA and protein in A549 and H446 cells overexpressing Prox1 after plasmid transfection was high, while the expression level of Prox1mRNA and protein in A549 and H446 cells with low expression of Prox1 after siRNA transfection was low. With the increase of Prox1 expression, the expression of RhoA and RhoC increased, while the expression of RhoB decreased.

Conclusion: The finding of this study may provide a new approach for the treatment of lung cancer using targeted gene therapy.

Keywords: Rho family; invasion; lung cancer; migration; transcription factor Prox1.

Figure 1

Global cancer statistics 2012. (a) The cancer types’ percentages associated with POTSFs. (b) The cancer types’ percentages documented in statistics of 2012’s global cancer. The same cancer type is marked with the same color in (a) and (b); e.g., lung is represented with pink color in both (a) and (b).

Figure 2

Agarose gel experiment to verify the expression of Prox1 in lung cancer.

Figure 3

Expression of Prox1 protein in different lung cancer cell lines, including A549 and H446.

Figure 4

Plate colony formation assay to detect the effect of Prox1 plasmid transfection on the proliferation of A549 and H446 cells. Plate cloning was used to detect the clone formation of A549 and H446 cells overexpressing Prox1 after plasmid transfection and wild-type untransfected A549 and H446 cells (b and d). Wild-type untransfected A549 and H446 cells served as a control group (a and c).

Figure 5

MTT assay to detect the growth curve of (a) A549 and (b) H446 cells after transfection with Prox1 plasmid. Data were presented as mean ± SD and analyzed using Student’s t-test (n = 3). P value; ns, no significance.

Figure 6

Transwell to detect the change of migration ability of A549 and H446 cells after changes in Prox1 expression. Wild-type untransfected A549 and H446 cells served the control group (n = 3). Data were presented as mean ± SD and analyzed using Student’s t-test (n = 3). ns, no significance; **P < 0.01; *P < 0.05.

Figure 7

Transwell to detect the change of invasive ability of A549 and H446 cells after changes in Prox1 expression. Wild-type untransfected A549 and H446 cells served as the control group. Cell invasive ability increased with the increased expression of Prox1 and decreased with the decreased expression of Prox1 (P < 0.01). Data were presented as mean ± SD and analyzed using Student’s t-test (n = 3). ns, no significance; **P < 0.01; *P < 0.05.

Figure 8

RT-PCR to detect RhoA, RhoB, and RhoC expression after changes in Prox1 expression. The expression of RhoA, RhoB, and RhoC in A549, H446 lung cancer cells was detected by transfection with plasmid Prox1 and interfering siRNA. When the Prox1 increased, the RhoA and RhoC increased, while RhoB decreased (P < 0.05); when Prox1 decreased, the RhoA and RhoC decreased, while RhoB increased (P > 0.05). ns, no significance; *P < 0.05, (n = 3).

Figure 9

Western blot to detect RhoA, RhoB, and RhoC expression after changes in Prox1 expression. The expression of RhoA, RhoB, and RhoC in A549, H446 lung cancer cells was detected by transfection with plasmid Prox1 and interfering siRNA.