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G-protein coupled receptor 34 knockdown impairs the proliferation and migration of HGC-27 gastric cancer cells in vitro.

Jin ZT, Li K, Li M, Ren ZG, Wang FS, Zhu JY, Leng XS, Yu WD - Chin. Med. J. (2015)

Bottom Line: We also measured expression profile of PI3K/PDK1/AKT and ERK using Western blotting.The ShRNA directed against GPR34 effectively inhibited both endogenous mRNA and protein expression levels of GPR34, and significantly down-regulated the expression of PIK3CB (P < 0.01), PIK3CD (P < 0.01), PDK1 (P < 0.01), phosphorylation of PDK1 (P < 0.01), Akt (P < 0.01), and ERK (P < 0.01).Furthermore, GPR34 knockdown resulted in an obvious reduction in HGC-27 cancer cell proliferation and migration activity (P < 0.01).

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology; Institute of Clinical Molecular Biology, Peking University People's Hospital, Beijing 100044, China.

ABSTRACT

Background: Overexpression of G-protein coupled receptor 34 (GPR34) affects the progression and prognosis of human gastric adenocarcinoma, however, the role of GPR34 in gastric cancer development and progression has not been well-determined. The current study aimed to investigate the effect of GPR34 knockdown on the proliferation, migration, and apoptosis of HGC-27 gastric cancer cells and the underlying mechanisms.

Methods: The expression of GPR34 in gastric cancer cell line HGC-27 was detected by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. HGC-27 cells were employed to construct the stable GPR34 knockdown cell model in this study. Real-time RT-PCR and Western blotting were applied to validate the effect of short hairpin RNA (ShRNA) on the expression of GPR34 in HGC-27 gastric cells. The proliferation, migration of these cells were examined by Cell Counting Kit-8 and transwell. We also measured expression profile of PI3K/PDK1/AKT and ERK using Western blotting.

Results: The ShRNA directed against GPR34 effectively inhibited both endogenous mRNA and protein expression levels of GPR34, and significantly down-regulated the expression of PIK3CB (P < 0.01), PIK3CD (P < 0.01), PDK1 (P < 0.01), phosphorylation of PDK1 (P < 0.01), Akt (P < 0.01), and ERK (P < 0.01). Furthermore, GPR34 knockdown resulted in an obvious reduction in HGC-27 cancer cell proliferation and migration activity (P < 0.01).

Conclusions: GPR34 knockdown impairs the proliferation and migration of HGC-27 gastric cancer cells in vitro and provides a potential implication for therapy of gastric cancer.

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Related in: MedlinePlus

Validation of GPR34 knockdown in HGC-27 cells. (a) Knockdown of GPR34 expression in HGC-27 cells. The vertical axis shows the GPR34 mRNA levels relative to that of GAPDH. The data represent the mean ± SD of triplicate experiments; (b) Western blot validation of GPR34 knock-down p185YR cell model construction; (c) Densitometry analysis of Western blotting of GPR34 in HGC-27 cells (ImageJ) (*P< 0.01).
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Figure 1: Validation of GPR34 knockdown in HGC-27 cells. (a) Knockdown of GPR34 expression in HGC-27 cells. The vertical axis shows the GPR34 mRNA levels relative to that of GAPDH. The data represent the mean ± SD of triplicate experiments; (b) Western blot validation of GPR34 knock-down p185YR cell model construction; (c) Densitometry analysis of Western blotting of GPR34 in HGC-27 cells (ImageJ) (*P< 0.01).

Mentions: HGC-27, a low-phosphatase and tensin homolog gastric cancer cell line, was selected and used as a cell model in vitro to determine the correlation between GPR34 expression and proliferation, apoptosis and migration of gastric cancer cells. Both western blotting and real-time RT-PCR results indicated that the GPR34 knockdown HGC-27 cell model (one clone) was successfully constructed [Figure 1a–c].


G-protein coupled receptor 34 knockdown impairs the proliferation and migration of HGC-27 gastric cancer cells in vitro.

Jin ZT, Li K, Li M, Ren ZG, Wang FS, Zhu JY, Leng XS, Yu WD - Chin. Med. J. (2015)

Validation of GPR34 knockdown in HGC-27 cells. (a) Knockdown of GPR34 expression in HGC-27 cells. The vertical axis shows the GPR34 mRNA levels relative to that of GAPDH. The data represent the mean ± SD of triplicate experiments; (b) Western blot validation of GPR34 knock-down p185YR cell model construction; (c) Densitometry analysis of Western blotting of GPR34 in HGC-27 cells (ImageJ) (*P< 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4836262&req=5

Figure 1: Validation of GPR34 knockdown in HGC-27 cells. (a) Knockdown of GPR34 expression in HGC-27 cells. The vertical axis shows the GPR34 mRNA levels relative to that of GAPDH. The data represent the mean ± SD of triplicate experiments; (b) Western blot validation of GPR34 knock-down p185YR cell model construction; (c) Densitometry analysis of Western blotting of GPR34 in HGC-27 cells (ImageJ) (*P< 0.01).
Mentions: HGC-27, a low-phosphatase and tensin homolog gastric cancer cell line, was selected and used as a cell model in vitro to determine the correlation between GPR34 expression and proliferation, apoptosis and migration of gastric cancer cells. Both western blotting and real-time RT-PCR results indicated that the GPR34 knockdown HGC-27 cell model (one clone) was successfully constructed [Figure 1a–c].

Bottom Line: We also measured expression profile of PI3K/PDK1/AKT and ERK using Western blotting.The ShRNA directed against GPR34 effectively inhibited both endogenous mRNA and protein expression levels of GPR34, and significantly down-regulated the expression of PIK3CB (P < 0.01), PIK3CD (P < 0.01), PDK1 (P < 0.01), phosphorylation of PDK1 (P < 0.01), Akt (P < 0.01), and ERK (P < 0.01).Furthermore, GPR34 knockdown resulted in an obvious reduction in HGC-27 cancer cell proliferation and migration activity (P < 0.01).

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology; Institute of Clinical Molecular Biology, Peking University People's Hospital, Beijing 100044, China.

ABSTRACT

Background: Overexpression of G-protein coupled receptor 34 (GPR34) affects the progression and prognosis of human gastric adenocarcinoma, however, the role of GPR34 in gastric cancer development and progression has not been well-determined. The current study aimed to investigate the effect of GPR34 knockdown on the proliferation, migration, and apoptosis of HGC-27 gastric cancer cells and the underlying mechanisms.

Methods: The expression of GPR34 in gastric cancer cell line HGC-27 was detected by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. HGC-27 cells were employed to construct the stable GPR34 knockdown cell model in this study. Real-time RT-PCR and Western blotting were applied to validate the effect of short hairpin RNA (ShRNA) on the expression of GPR34 in HGC-27 gastric cells. The proliferation, migration of these cells were examined by Cell Counting Kit-8 and transwell. We also measured expression profile of PI3K/PDK1/AKT and ERK using Western blotting.

Results: The ShRNA directed against GPR34 effectively inhibited both endogenous mRNA and protein expression levels of GPR34, and significantly down-regulated the expression of PIK3CB (P < 0.01), PIK3CD (P < 0.01), PDK1 (P < 0.01), phosphorylation of PDK1 (P < 0.01), Akt (P < 0.01), and ERK (P < 0.01). Furthermore, GPR34 knockdown resulted in an obvious reduction in HGC-27 cancer cell proliferation and migration activity (P < 0.01).

Conclusions: GPR34 knockdown impairs the proliferation and migration of HGC-27 gastric cancer cells in vitro and provides a potential implication for therapy of gastric cancer.

Show MeSH
Related in: MedlinePlus