Limits...
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.

Show MeSH

Related in: MedlinePlus

Down-regulation of USP39 inhibits cell proliferation and colony formation ability of SMMC-7721 cells. (A) The growth curves of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. **, P < 0.01. (B) Representative images of colony formation assays of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. From top to bottom: crystals violet staining, bright field and GFP of single colony, and crystals violet staining of six-well plate. Scale bar, 25 μm. (C) Colony numbers of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. ***, P < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig2: Down-regulation of USP39 inhibits cell proliferation and colony formation ability of SMMC-7721 cells. (A) The growth curves of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. **, P < 0.01. (B) Representative images of colony formation assays of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. From top to bottom: crystals violet staining, bright field and GFP of single colony, and crystals violet staining of six-well plate. Scale bar, 25 μm. (C) Colony numbers of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. ***, P < 0.001.

Mentions: To study whether USP39 was related with SMMC-7721 cell proliferation, we performed 5-day MTT assay. Lv-shUSP39 infected SMMC-7721 showed slower growth rate compared with control and Lv-shCon infected cells (Figure 2A). On day 5, OD595 of Lv-shUSP39 infected cell was only 3.51 ± 0.12, while that of control and Lv-shCon infected cells were 5.31 ± 0.10 and 5.24 ± 0.53, respectively. We then analyzed the colony formation ability of SMMC-7721 cells after lentiviral infection using crystal violet staining. The cell number in a single colony was significantly reduced after Lv-shUSP39 infection (Figure 2B). Furthermore, we calculated the number of colons formed after lentivirus infection. The colony number of LvshUSP39 infected SMMC-7721 cells was only 46 ± 8, compared with that of 207 ± 5 in control cells and 203 ± 5 in Lv-shCon infected cells (Figure 2C). Furthermore, these results suggested that suppression of USP39 could inhibit cell proliferation and colony formation of HCC cells.Figure 2


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 inhibits cell proliferation and colony formation ability of SMMC-7721 cells. (A) The growth curves of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. **, P < 0.01. (B) Representative images of colony formation assays of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. From top to bottom: crystals violet staining, bright field and GFP of single colony, and crystals violet staining of six-well plate. Scale bar, 25 μm. (C) Colony numbers of Con, Lv-shCon and Lv-shUSP39 infected 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

Fig2: Down-regulation of USP39 inhibits cell proliferation and colony formation ability of SMMC-7721 cells. (A) The growth curves of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. **, P < 0.01. (B) Representative images of colony formation assays of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. From top to bottom: crystals violet staining, bright field and GFP of single colony, and crystals violet staining of six-well plate. Scale bar, 25 μm. (C) Colony numbers of Con, Lv-shCon and Lv-shUSP39 infected SMMC-7721 cells. ***, P < 0.001.
Mentions: To study whether USP39 was related with SMMC-7721 cell proliferation, we performed 5-day MTT assay. Lv-shUSP39 infected SMMC-7721 showed slower growth rate compared with control and Lv-shCon infected cells (Figure 2A). On day 5, OD595 of Lv-shUSP39 infected cell was only 3.51 ± 0.12, while that of control and Lv-shCon infected cells were 5.31 ± 0.10 and 5.24 ± 0.53, respectively. We then analyzed the colony formation ability of SMMC-7721 cells after lentiviral infection using crystal violet staining. The cell number in a single colony was significantly reduced after Lv-shUSP39 infection (Figure 2B). Furthermore, we calculated the number of colons formed after lentivirus infection. The colony number of LvshUSP39 infected SMMC-7721 cells was only 46 ± 8, compared with that of 207 ± 5 in control cells and 203 ± 5 in Lv-shCon infected cells (Figure 2C). Furthermore, these results suggested that suppression of USP39 could inhibit cell proliferation and colony formation of HCC cells.Figure 2

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