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Lentivirus mediated silencing of ubiquitin specific peptidase 39 inhibits cell proliferation of human hepatocellular carcinoma cells in vitro.

Pan Z, Pan H, Zhang J, Yang Y, Liu H, Yang Y, Huang G, Ni J, Huang J, Zhou W - Biol. Res. (2015)

Bottom Line: Crystal violet staining indicated that colony numbers and sizes were both reduced after knock-down of USP39.In addition, Annexin V showed that downregulation of USP39 significantly increased the population of apoptotic cells.All our results suggest that USP39 is important for HCC cell proliferation and is a potential target for molecular therapy of HCC.

View Article: PubMed Central - PubMed

Affiliation: The Third Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, 200438, Shanghai, China. zeya_pan@yeah.net.

ABSTRACT

Background: Ubiquitin Specific Peptidase 39 (USP39) is a 65 kDa SR-related protein involved in RNA splicing. Previous studies showed that USP39 is related with tumorigenesis of human breast cancer cells.

Results: In the present study, we investigated the functions of USP39 in human hepatocellular carcinoma (HCC) cell line SMMC-7721. We knocked down the expression of USP39 through lentivirus mediated RNA interference. The results of qRT-PCR and western blotting assay showed that both the mRNA and protein levels were suppressed efficiently after USP39 specific shRNA was delivered into SMMC-7721 cells. Cell growth was significantly inhibited as determined by MTT assay. Crystal violet staining indicated that colony numbers and sizes were both reduced after knock-down of USP39. Furthermore, suppression of USP39 arrested cell cycle progression at G2/M phase in SMMC-7721cells. In addition, Annexin V showed that downregulation of USP39 significantly increased the population of apoptotic cells.

Conclusions: All our results suggest that USP39 is important for HCC cell proliferation and is a potential target for molecular therapy of HCC.

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Related in: MedlinePlus

Down-regulation of USP39 impairs cell cycle progression of SMMC-7721 cells. (A) Representative graphs of flow cytometry analysis of SMMC-7721 cell cycle using PI staining. (B) Statistic analysis of percentages of SMMC-7721 cells at different cell cycle stages (G0/G1, S and G2/M). (C) Statistic analysis of percentages of sub-G1 SMMC-7721 cells. ***, P < 0.001.
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Fig3: Down-regulation of USP39 impairs cell cycle progression of SMMC-7721 cells. (A) Representative graphs of flow cytometry analysis of SMMC-7721 cell cycle using PI staining. (B) Statistic analysis of percentages of SMMC-7721 cells at different cell cycle stages (G0/G1, S and G2/M). (C) Statistic analysis of percentages of sub-G1 SMMC-7721 cells. ***, P < 0.001.

Mentions: To find out the underlying mechanisms of inhibition of cell proliferation and colony formation, we analyzed phases of cell cycle of SMMC-7721 cells after USP39 knockdown using flow cytometry with PI staining. As shown in Figure 3A and B, 43.44 ± 0.55% of cells were at G0/G1 phase in Lv-shUSP39 infected SMCC-7721 cells, which were significantly lower than those of control cells (57.78 ± 0.18%, p value) and Lv-shCon infected cells (58.28 ± 0.26%, p value). Meanwhile, there were more cells at G2/M phase after Lv-shUSP39 infection (30.20 ± 0.46%), compared with control cells (20.20 ± 0.43%) and Lv-shCon infected cells (18.52 ± 0.54%). These results indicated that cell cycle progression was impaired in USP39 knock-down HCC cells. In addition, 5.02 ± 0.09% cells were detected at sub-G1 phase in Lv-shUSP39 infected SMMC-7721 cells, much higher than those in control cells (0.79 ± 0.04%) and Lv-shCon infected cells (0.77 ± 0.05%) (Figure 3C), suggesting that there were more apoptotic cells after USP39 knock-down.Figure 3


Lentivirus mediated silencing of ubiquitin specific peptidase 39 inhibits cell proliferation of human hepatocellular carcinoma cells in vitro.

Pan Z, Pan H, Zhang J, Yang Y, Liu H, Yang Y, Huang G, Ni J, Huang J, Zhou W - Biol. Res. (2015)

Down-regulation of USP39 impairs cell cycle progression of SMMC-7721 cells. (A) Representative graphs of flow cytometry analysis of SMMC-7721 cell cycle using PI staining. (B) Statistic analysis of percentages of SMMC-7721 cells at different cell cycle stages (G0/G1, S and G2/M). (C) Statistic analysis of percentages of sub-G1 SMMC-7721 cells. ***, P < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4389921&req=5

Fig3: Down-regulation of USP39 impairs cell cycle progression of SMMC-7721 cells. (A) Representative graphs of flow cytometry analysis of SMMC-7721 cell cycle using PI staining. (B) Statistic analysis of percentages of SMMC-7721 cells at different cell cycle stages (G0/G1, S and G2/M). (C) Statistic analysis of percentages of sub-G1 SMMC-7721 cells. ***, P < 0.001.
Mentions: To find out the underlying mechanisms of inhibition of cell proliferation and colony formation, we analyzed phases of cell cycle of SMMC-7721 cells after USP39 knockdown using flow cytometry with PI staining. As shown in Figure 3A and B, 43.44 ± 0.55% of cells were at G0/G1 phase in Lv-shUSP39 infected SMCC-7721 cells, which were significantly lower than those of control cells (57.78 ± 0.18%, p value) and Lv-shCon infected cells (58.28 ± 0.26%, p value). Meanwhile, there were more cells at G2/M phase after Lv-shUSP39 infection (30.20 ± 0.46%), compared with control cells (20.20 ± 0.43%) and Lv-shCon infected cells (18.52 ± 0.54%). These results indicated that cell cycle progression was impaired in USP39 knock-down HCC cells. In addition, 5.02 ± 0.09% cells were detected at sub-G1 phase in Lv-shUSP39 infected SMMC-7721 cells, much higher than those in control cells (0.79 ± 0.04%) and Lv-shCon infected cells (0.77 ± 0.05%) (Figure 3C), suggesting that there were more apoptotic cells after USP39 knock-down.Figure 3

Bottom Line: Crystal violet staining indicated that colony numbers and sizes were both reduced after knock-down of USP39.In addition, Annexin V showed that downregulation of USP39 significantly increased the population of apoptotic cells.All our results suggest that USP39 is important for HCC cell proliferation and is a potential target for molecular therapy of HCC.

View Article: PubMed Central - PubMed

Affiliation: The Third Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, 200438, Shanghai, China. zeya_pan@yeah.net.

ABSTRACT

Background: Ubiquitin Specific Peptidase 39 (USP39) is a 65 kDa SR-related protein involved in RNA splicing. Previous studies showed that USP39 is related with tumorigenesis of human breast cancer cells.

Results: In the present study, we investigated the functions of USP39 in human hepatocellular carcinoma (HCC) cell line SMMC-7721. We knocked down the expression of USP39 through lentivirus mediated RNA interference. The results of qRT-PCR and western blotting assay showed that both the mRNA and protein levels were suppressed efficiently after USP39 specific shRNA was delivered into SMMC-7721 cells. Cell growth was significantly inhibited as determined by MTT assay. Crystal violet staining indicated that colony numbers and sizes were both reduced after knock-down of USP39. Furthermore, suppression of USP39 arrested cell cycle progression at G2/M phase in SMMC-7721cells. In addition, Annexin V showed that downregulation of USP39 significantly increased the population of apoptotic cells.

Conclusions: All our results suggest that USP39 is important for HCC cell proliferation and is a potential target for molecular therapy of HCC.

Show MeSH
Related in: MedlinePlus