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Efficient inhibition of human glioma development by RNA interference-mediated silencing of PAK5.

Gu X, Wang C, Wang X, Ma G, Li Y, Cui L, Chen Y, Zhao B, Li K - Int. J. Biol. Sci. (2015)

Bottom Line: In addition, silencing PAK5 expression in U87 cells weakened their colony formation ability and in vivo tumorigenesis ability.Further studies demonstrated that PAK5 inhibition led to an increase in cleaved caspase 3 and a decrease in β-catenin.In conclusion, our results suggest that the inhibition of PAK5 by RNA interference might efficiently suppress tumor development of glioma cells with PAK5 high expression.

View Article: PubMed Central - PubMed

Affiliation: 1. Institute of Neurology, Guangdong Medical College, Zhanjiang, China. ; 2. Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Affiliated Hospital of Guangdong Medical College, Zhanjiang, China.

ABSTRACT
Glioma is the most common type of primary intracranial tumor and is highly lethal due to its pathogenetic location, high invasiveness, and poor prognosis. Even combined surgery and chemoradiotherapy do not effectively rescue glioma patients. Molecular target therapy is considered a safe and promising therapy for glioma. The identification of a novel, effective target protein in gliomas is of great interest. We found that PAK5 was highly expressed in the tumor tissues of glioma patients and human glioma cell lines. We then used a lentivirus-delivered short hairpin RNA to stably silence PAK5 expression in glioma cells and explore its influence. The results showed that the inhibition of PAK5 reduced cell viability and delayed the cell cycle at the G0/G1 phase in the glioma cells with PAK5 high expression. In addition, silencing PAK5 expression in U87 cells weakened their colony formation ability and in vivo tumorigenesis ability. Further studies demonstrated that PAK5 inhibition led to an increase in cleaved caspase 3 and a decrease in β-catenin. In conclusion, our results suggest that the inhibition of PAK5 by RNA interference might efficiently suppress tumor development of glioma cells with PAK5 high expression. This finding provides a novel, promising therapeutic target for glioma treatment.

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Inhibition of PAK5 prominently suppresses tumor development in U87 cells. (A) Colony formation assay of cells from the three groups. Cells were seeded at 500 cells/well and allowed to form colonies for 7 days. The colonies were stained with crystal violet and counted. Data represent the mean ±SEM of three independent experiments (right). **, P<0.01. (B) Cell cycle analysis of cells from the three groups as determined by PI staining and FACS analysis (left). The percentages of cell cycle phases represent the mean ± SEM of three independent experiments (right). *, P < 0.05. (C, D) Scr-shRNA-infected and PAK5-shRNA-infected groups of nude mice were injected with the corresponding infected U87 cells (106 cells per mice) into the axilla. After 3 weeks, some mice were sacrificed, and their tumors were examined (n=6) (C). The other mice were used to detected the time of visible tumors (diameter > 3 cm) (n=6) (D). **, P < 0.01.
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Figure 4: Inhibition of PAK5 prominently suppresses tumor development in U87 cells. (A) Colony formation assay of cells from the three groups. Cells were seeded at 500 cells/well and allowed to form colonies for 7 days. The colonies were stained with crystal violet and counted. Data represent the mean ±SEM of three independent experiments (right). **, P<0.01. (B) Cell cycle analysis of cells from the three groups as determined by PI staining and FACS analysis (left). The percentages of cell cycle phases represent the mean ± SEM of three independent experiments (right). *, P < 0.05. (C, D) Scr-shRNA-infected and PAK5-shRNA-infected groups of nude mice were injected with the corresponding infected U87 cells (106 cells per mice) into the axilla. After 3 weeks, some mice were sacrificed, and their tumors were examined (n=6) (C). The other mice were used to detected the time of visible tumors (diameter > 3 cm) (n=6) (D). **, P < 0.01.

Mentions: Flow cytometric analysis of cell cycle of U87 cells by PI staining revealed that PAK5 inhibition resulted in an increase in cells detained at the G0/G1 phase and a decrease in cells at the G2/M phases (Fig. 4B). These data demonstrate that PAK5 inhibition forces U87 cells into the quiescent phase. Moreover, the colony-forming ability of the PAK5-shRNA infected groups also obviously decreased, in contrast to the Scr-shRNA infected group (Fig. 4A). These data demonstrate that the inhibition of PAK5 expression decreases U87 cell growth in vitro. A nude mouse tumorigenesis test revealed that the tumorigenesis ability of U87 cells was also weakened by silencing of PAK5 expression (Fig. 4C and D). Taken together, these results suggest that the inhibition of PAK5 expression suppresses tumor growth in glioma cells with PAK5 high expression.


Efficient inhibition of human glioma development by RNA interference-mediated silencing of PAK5.

Gu X, Wang C, Wang X, Ma G, Li Y, Cui L, Chen Y, Zhao B, Li K - Int. J. Biol. Sci. (2015)

Inhibition of PAK5 prominently suppresses tumor development in U87 cells. (A) Colony formation assay of cells from the three groups. Cells were seeded at 500 cells/well and allowed to form colonies for 7 days. The colonies were stained with crystal violet and counted. Data represent the mean ±SEM of three independent experiments (right). **, P<0.01. (B) Cell cycle analysis of cells from the three groups as determined by PI staining and FACS analysis (left). The percentages of cell cycle phases represent the mean ± SEM of three independent experiments (right). *, P < 0.05. (C, D) Scr-shRNA-infected and PAK5-shRNA-infected groups of nude mice were injected with the corresponding infected U87 cells (106 cells per mice) into the axilla. After 3 weeks, some mice were sacrificed, and their tumors were examined (n=6) (C). The other mice were used to detected the time of visible tumors (diameter > 3 cm) (n=6) (D). **, P < 0.01.
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Figure 4: Inhibition of PAK5 prominently suppresses tumor development in U87 cells. (A) Colony formation assay of cells from the three groups. Cells were seeded at 500 cells/well and allowed to form colonies for 7 days. The colonies were stained with crystal violet and counted. Data represent the mean ±SEM of three independent experiments (right). **, P<0.01. (B) Cell cycle analysis of cells from the three groups as determined by PI staining and FACS analysis (left). The percentages of cell cycle phases represent the mean ± SEM of three independent experiments (right). *, P < 0.05. (C, D) Scr-shRNA-infected and PAK5-shRNA-infected groups of nude mice were injected with the corresponding infected U87 cells (106 cells per mice) into the axilla. After 3 weeks, some mice were sacrificed, and their tumors were examined (n=6) (C). The other mice were used to detected the time of visible tumors (diameter > 3 cm) (n=6) (D). **, P < 0.01.
Mentions: Flow cytometric analysis of cell cycle of U87 cells by PI staining revealed that PAK5 inhibition resulted in an increase in cells detained at the G0/G1 phase and a decrease in cells at the G2/M phases (Fig. 4B). These data demonstrate that PAK5 inhibition forces U87 cells into the quiescent phase. Moreover, the colony-forming ability of the PAK5-shRNA infected groups also obviously decreased, in contrast to the Scr-shRNA infected group (Fig. 4A). These data demonstrate that the inhibition of PAK5 expression decreases U87 cell growth in vitro. A nude mouse tumorigenesis test revealed that the tumorigenesis ability of U87 cells was also weakened by silencing of PAK5 expression (Fig. 4C and D). Taken together, these results suggest that the inhibition of PAK5 expression suppresses tumor growth in glioma cells with PAK5 high expression.

Bottom Line: In addition, silencing PAK5 expression in U87 cells weakened their colony formation ability and in vivo tumorigenesis ability.Further studies demonstrated that PAK5 inhibition led to an increase in cleaved caspase 3 and a decrease in β-catenin.In conclusion, our results suggest that the inhibition of PAK5 by RNA interference might efficiently suppress tumor development of glioma cells with PAK5 high expression.

View Article: PubMed Central - PubMed

Affiliation: 1. Institute of Neurology, Guangdong Medical College, Zhanjiang, China. ; 2. Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Affiliated Hospital of Guangdong Medical College, Zhanjiang, China.

ABSTRACT
Glioma is the most common type of primary intracranial tumor and is highly lethal due to its pathogenetic location, high invasiveness, and poor prognosis. Even combined surgery and chemoradiotherapy do not effectively rescue glioma patients. Molecular target therapy is considered a safe and promising therapy for glioma. The identification of a novel, effective target protein in gliomas is of great interest. We found that PAK5 was highly expressed in the tumor tissues of glioma patients and human glioma cell lines. We then used a lentivirus-delivered short hairpin RNA to stably silence PAK5 expression in glioma cells and explore its influence. The results showed that the inhibition of PAK5 reduced cell viability and delayed the cell cycle at the G0/G1 phase in the glioma cells with PAK5 high expression. In addition, silencing PAK5 expression in U87 cells weakened their colony formation ability and in vivo tumorigenesis ability. Further studies demonstrated that PAK5 inhibition led to an increase in cleaved caspase 3 and a decrease in β-catenin. In conclusion, our results suggest that the inhibition of PAK5 by RNA interference might efficiently suppress tumor development of glioma cells with PAK5 high expression. This finding provides a novel, promising therapeutic target for glioma treatment.

Show MeSH
Related in: MedlinePlus