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Increased ARPP-19 expression is associated with hepatocellular carcinoma.

Song H, Pan J, Liu Y, Wen H, Wang L, Cui J, Liu Y, Hu B, Yao Z, Ji G - Int J Mol Sci (2014)

Bottom Line: Expression of ARPP-19 was increased in human hepatocellular carcinoma (HCC) compared to adjacent non-tumorous liver tissues in 36 paired liver samples, and the level of ARPP-19 in HCC tissues was positively correlated with the tumor size.HepG2 and SMMC-7721 cells with ARPP-19 knockdown displayed lowered cell growth rate, retarded colony formation and increased arrest at the G2/M phase transition.Therefore, ARPP-19 may play a role in HCC pathogenesis through regulating cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China. songhy@126.com.

ABSTRACT
The cAMP-regulated phosphoprotein 19 (ARPP-19) plays a key role in cell mitotic G2/M transition. Expression of ARPP-19 was increased in human hepatocellular carcinoma (HCC) compared to adjacent non-tumorous liver tissues in 36 paired liver samples, and the level of ARPP-19 in HCC tissues was positively correlated with the tumor size. To determine the interrelationship between ARPP-19 expression and HCC, we silenced ARPP-19 expression in the human hepatocarcinoma HepG2 and SMMC-7721 cells using lentivirus encoding ARPP-19 siRNA. HepG2 and SMMC-7721 cells with ARPP-19 knockdown displayed lowered cell growth rate, retarded colony formation and increased arrest at the G2/M phase transition. Silencing ARPP-19 in HCC cells resulted in decreased protein levels of phospho-(Ser) CDKs substrates and increased levels of inactivated cyclin division cycle 2 (Cdc2). Therefore, ARPP-19 may play a role in HCC pathogenesis through regulating cell proliferation.

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Down-regulating the expression of ARPP-19 in hepatocarcinoma cells. (A) The expression level of ARPP-19 in four strains of hepatocarcinoma cells and in the pool of ten pairs of HCC and the non-tumorous liver tissues (NT) was determined by qRT-PCR, with β-actin as the internal control; (B) HepG2 and SMMC-7721 cells were transfected with lentivirus with GV118-GFP vector carrying siRNA against ARPP-19 (ARPP-19-RNAi) or empty vector (scramble). The ARPP-19 mRNA level was measured by qRT-PCR, with β-actin as the internal control; (C) The protein level of ARPP-19 in HepG2 and SMMC-7721 cells transfected with ARPP-19-RNAi or scramble was measured by immunoblotting, with β-actin as the loading control; (D) The images of western blots of (C) were quantified. The data are shown as the mean ± SEM, and significant values are indicated with asterisks (*p < 0.05; **p < 0.01, versus scramble).
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ijms-16-00178-f003: Down-regulating the expression of ARPP-19 in hepatocarcinoma cells. (A) The expression level of ARPP-19 in four strains of hepatocarcinoma cells and in the pool of ten pairs of HCC and the non-tumorous liver tissues (NT) was determined by qRT-PCR, with β-actin as the internal control; (B) HepG2 and SMMC-7721 cells were transfected with lentivirus with GV118-GFP vector carrying siRNA against ARPP-19 (ARPP-19-RNAi) or empty vector (scramble). The ARPP-19 mRNA level was measured by qRT-PCR, with β-actin as the internal control; (C) The protein level of ARPP-19 in HepG2 and SMMC-7721 cells transfected with ARPP-19-RNAi or scramble was measured by immunoblotting, with β-actin as the loading control; (D) The images of western blots of (C) were quantified. The data are shown as the mean ± SEM, and significant values are indicated with asterisks (*p < 0.05; **p < 0.01, versus scramble).

Mentions: In order to study the role of ARPP-19 in HCC development in vitro, we first measured its expression in hepatocarcinoma cell lines. Figure 1A shows that more ARPP-19 was expressed in four strains of hepatocarcinoma cells than non-tumorous liver tissues, especially in HepG2 and SMMC7721 cells. Additionally, as HepG2 is well differentiated and SMMC7721 is more malignant with low differentiation, we chose these two cells for further in vitro studies by using RNA interference. We next determined the potential role that ARPP-19 plays in controlling cell growth by silencing ARPP-19 expression in HepG2 and SMMC-7721 cells using lentivirus-encoded ARPP-19-RNAi. Transfection efficiency, as determined using a viral vector encoding green fluorescent protein observed with a fluorescence microscope and compared with phase-contrast images, was nearly 100% after transfection. The ARPP-19 protein concentrations were decreased by 75.6% and 73.5%, respectively, in HepG2 and SMMC-7721 cells 72 h after virus transfection (Figure 3C,D). These results demonstrated that the expression of ARPP-19 was successfully downregulated in HepG2 and SMMC-7721 cells under these conditions.


Increased ARPP-19 expression is associated with hepatocellular carcinoma.

Song H, Pan J, Liu Y, Wen H, Wang L, Cui J, Liu Y, Hu B, Yao Z, Ji G - Int J Mol Sci (2014)

Down-regulating the expression of ARPP-19 in hepatocarcinoma cells. (A) The expression level of ARPP-19 in four strains of hepatocarcinoma cells and in the pool of ten pairs of HCC and the non-tumorous liver tissues (NT) was determined by qRT-PCR, with β-actin as the internal control; (B) HepG2 and SMMC-7721 cells were transfected with lentivirus with GV118-GFP vector carrying siRNA against ARPP-19 (ARPP-19-RNAi) or empty vector (scramble). The ARPP-19 mRNA level was measured by qRT-PCR, with β-actin as the internal control; (C) The protein level of ARPP-19 in HepG2 and SMMC-7721 cells transfected with ARPP-19-RNAi or scramble was measured by immunoblotting, with β-actin as the loading control; (D) The images of western blots of (C) were quantified. The data are shown as the mean ± SEM, and significant values are indicated with asterisks (*p < 0.05; **p < 0.01, versus scramble).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4307242&req=5

ijms-16-00178-f003: Down-regulating the expression of ARPP-19 in hepatocarcinoma cells. (A) The expression level of ARPP-19 in four strains of hepatocarcinoma cells and in the pool of ten pairs of HCC and the non-tumorous liver tissues (NT) was determined by qRT-PCR, with β-actin as the internal control; (B) HepG2 and SMMC-7721 cells were transfected with lentivirus with GV118-GFP vector carrying siRNA against ARPP-19 (ARPP-19-RNAi) or empty vector (scramble). The ARPP-19 mRNA level was measured by qRT-PCR, with β-actin as the internal control; (C) The protein level of ARPP-19 in HepG2 and SMMC-7721 cells transfected with ARPP-19-RNAi or scramble was measured by immunoblotting, with β-actin as the loading control; (D) The images of western blots of (C) were quantified. The data are shown as the mean ± SEM, and significant values are indicated with asterisks (*p < 0.05; **p < 0.01, versus scramble).
Mentions: In order to study the role of ARPP-19 in HCC development in vitro, we first measured its expression in hepatocarcinoma cell lines. Figure 1A shows that more ARPP-19 was expressed in four strains of hepatocarcinoma cells than non-tumorous liver tissues, especially in HepG2 and SMMC7721 cells. Additionally, as HepG2 is well differentiated and SMMC7721 is more malignant with low differentiation, we chose these two cells for further in vitro studies by using RNA interference. We next determined the potential role that ARPP-19 plays in controlling cell growth by silencing ARPP-19 expression in HepG2 and SMMC-7721 cells using lentivirus-encoded ARPP-19-RNAi. Transfection efficiency, as determined using a viral vector encoding green fluorescent protein observed with a fluorescence microscope and compared with phase-contrast images, was nearly 100% after transfection. The ARPP-19 protein concentrations were decreased by 75.6% and 73.5%, respectively, in HepG2 and SMMC-7721 cells 72 h after virus transfection (Figure 3C,D). These results demonstrated that the expression of ARPP-19 was successfully downregulated in HepG2 and SMMC-7721 cells under these conditions.

Bottom Line: Expression of ARPP-19 was increased in human hepatocellular carcinoma (HCC) compared to adjacent non-tumorous liver tissues in 36 paired liver samples, and the level of ARPP-19 in HCC tissues was positively correlated with the tumor size.HepG2 and SMMC-7721 cells with ARPP-19 knockdown displayed lowered cell growth rate, retarded colony formation and increased arrest at the G2/M phase transition.Therefore, ARPP-19 may play a role in HCC pathogenesis through regulating cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China. songhy@126.com.

ABSTRACT
The cAMP-regulated phosphoprotein 19 (ARPP-19) plays a key role in cell mitotic G2/M transition. Expression of ARPP-19 was increased in human hepatocellular carcinoma (HCC) compared to adjacent non-tumorous liver tissues in 36 paired liver samples, and the level of ARPP-19 in HCC tissues was positively correlated with the tumor size. To determine the interrelationship between ARPP-19 expression and HCC, we silenced ARPP-19 expression in the human hepatocarcinoma HepG2 and SMMC-7721 cells using lentivirus encoding ARPP-19 siRNA. HepG2 and SMMC-7721 cells with ARPP-19 knockdown displayed lowered cell growth rate, retarded colony formation and increased arrest at the G2/M phase transition. Silencing ARPP-19 in HCC cells resulted in decreased protein levels of phospho-(Ser) CDKs substrates and increased levels of inactivated cyclin division cycle 2 (Cdc2). Therefore, ARPP-19 may play a role in HCC pathogenesis through regulating cell proliferation.

Show MeSH
Related in: MedlinePlus