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NOTCH1 signaling contributes to cell growth, anti-apoptosis and metastasis in salivary adenoid cystic carcinoma.

Su BH, Qu J, Song M, Huang XY, Hu XM, Xie J, Zhao Y, Ding LC, She L, Chen J, Lin LS, Lin X, Zheng DL, Lu YG - Oncotarget (2014)

Bottom Line: We silenced the expression of NOTCH1 and overexpressed activated NOTCH1 to elucidate the effects of NOTCH1 on proliferation, migration and invasion.NOTCH1 target genes were validated by real-time PCR.Overexpression of NOTCH1 in SACC cells promoted cell growth, migration and invasion, and knockdown of NOTCH1 inhibited cell proliferation in vitro and tumorigenicity in vivo by inducing cell apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Preventive Dentistry, Affiliated Stomatological Hospital, Fujian Medical University, Fuzhou, China. These authors contributed equally to this work.

ABSTRACT

Background: Numerous studies have reported both the tumor-suppressive and oncogenic roles of the Notch pathway, indicating that Notch activity regulates tumor biology in a complex, context-dependent manner. The aim of the present study was to identify the role of NOTCH1 in the cell growth and metastasis of SACC.

Methods: We analyzed the expression of NOTCH1 in clinical SACC samples using immunohistochemical staining. We silenced the expression of NOTCH1 and overexpressed activated NOTCH1 to elucidate the effects of NOTCH1 on proliferation, migration and invasion. NOTCH1 target genes were validated by real-time PCR.

Results: Our results showed that NOTCH1 was upregulated in SACC tissues when compared with normal tissues, and this upregulation was further enhanced in SACC tissues with metastasis and recurrence when compared with SACC tissues without metastasis. Overexpression of NOTCH1 in SACC cells promoted cell growth, migration and invasion, and knockdown of NOTCH1 inhibited cell proliferation in vitro and tumorigenicity in vivo by inducing cell apoptosis.

Conclusions: The results of this study suggest that NOTCH1 plays a key role in the cell growth, anti-apoptosis, and metastasis of SACC. NOTCH1 inhibitors might therefore have potential therapeutic applications in treating SACC patients by inhibiting cancer cell growth and metastasis.

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Overexpression of NOTCH1 promotes the proliferation of SACC-83 cellsA, Western blot analysis of the overexpression of NOTCH1 in SACC-83 cells infected with adenoviral vector at an MOI of 5-10; B-C, After adenoviral infection, the proliferation of SACC-83 cells was detected by CCK-8 reagent (B, P<0.05 at days 3, 4 and 5) and colony formation assay (C, P<0.01, n=3).
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Figure 3: Overexpression of NOTCH1 promotes the proliferation of SACC-83 cellsA, Western blot analysis of the overexpression of NOTCH1 in SACC-83 cells infected with adenoviral vector at an MOI of 5-10; B-C, After adenoviral infection, the proliferation of SACC-83 cells was detected by CCK-8 reagent (B, P<0.05 at days 3, 4 and 5) and colony formation assay (C, P<0.01, n=3).

Mentions: To investigate the effect of NOTCH1 on the proliferation of cancer cells, siRNA-mediated knockdown of NOTCH-1 was employed in SACC-83 cells. As expected, both siRNAs targeting NOTCH1 (siRNA-2010 and siRNA-6150) efficiently reduced NOTCH1 expression in the SACC-83 cells compared with the negative control (NC), as shown by real-time RT-PCR (Fig. 2A) and Western blot (Fig. 2B). Additionally, these siRNAs significantly inhibited the growth of the SACC-83 cells, as measured using the CCK8 reagent (Fig. 2C, P<0.05 at day 3, and P<0.01 at days 4 and 5) and colony formation assays (Fig. 2D, P<0.05, n=3). To further verify the results of this loss-of-function study, SACC-83 cells were infected with a recombinant adenovirus (Ad-NOTCH1) carrying the correct coding sequence of the intracellular cytoplasmic domain of NOTCH1. Overexpression of NICD1 in SACC-83 cells (Fig. 3A) slightly increased cell proliferation over the short term as detected by the CCK8 assay (Fig. 3B, P<0.01 at days 3 and 5) but robustly increased cell proliferation over the long term as measured by colony formation assay (Fig. 3C, P<0.001). These collective data imply that overexpression of NOTCH1 promotes SACC proliferation in vitro, supporting the oncogenic roles of NOTCH1 in SACC.


NOTCH1 signaling contributes to cell growth, anti-apoptosis and metastasis in salivary adenoid cystic carcinoma.

Su BH, Qu J, Song M, Huang XY, Hu XM, Xie J, Zhao Y, Ding LC, She L, Chen J, Lin LS, Lin X, Zheng DL, Lu YG - Oncotarget (2014)

Overexpression of NOTCH1 promotes the proliferation of SACC-83 cellsA, Western blot analysis of the overexpression of NOTCH1 in SACC-83 cells infected with adenoviral vector at an MOI of 5-10; B-C, After adenoviral infection, the proliferation of SACC-83 cells was detected by CCK-8 reagent (B, P<0.05 at days 3, 4 and 5) and colony formation assay (C, P<0.01, n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4196170&req=5

Figure 3: Overexpression of NOTCH1 promotes the proliferation of SACC-83 cellsA, Western blot analysis of the overexpression of NOTCH1 in SACC-83 cells infected with adenoviral vector at an MOI of 5-10; B-C, After adenoviral infection, the proliferation of SACC-83 cells was detected by CCK-8 reagent (B, P<0.05 at days 3, 4 and 5) and colony formation assay (C, P<0.01, n=3).
Mentions: To investigate the effect of NOTCH1 on the proliferation of cancer cells, siRNA-mediated knockdown of NOTCH-1 was employed in SACC-83 cells. As expected, both siRNAs targeting NOTCH1 (siRNA-2010 and siRNA-6150) efficiently reduced NOTCH1 expression in the SACC-83 cells compared with the negative control (NC), as shown by real-time RT-PCR (Fig. 2A) and Western blot (Fig. 2B). Additionally, these siRNAs significantly inhibited the growth of the SACC-83 cells, as measured using the CCK8 reagent (Fig. 2C, P<0.05 at day 3, and P<0.01 at days 4 and 5) and colony formation assays (Fig. 2D, P<0.05, n=3). To further verify the results of this loss-of-function study, SACC-83 cells were infected with a recombinant adenovirus (Ad-NOTCH1) carrying the correct coding sequence of the intracellular cytoplasmic domain of NOTCH1. Overexpression of NICD1 in SACC-83 cells (Fig. 3A) slightly increased cell proliferation over the short term as detected by the CCK8 assay (Fig. 3B, P<0.01 at days 3 and 5) but robustly increased cell proliferation over the long term as measured by colony formation assay (Fig. 3C, P<0.001). These collective data imply that overexpression of NOTCH1 promotes SACC proliferation in vitro, supporting the oncogenic roles of NOTCH1 in SACC.

Bottom Line: We silenced the expression of NOTCH1 and overexpressed activated NOTCH1 to elucidate the effects of NOTCH1 on proliferation, migration and invasion.NOTCH1 target genes were validated by real-time PCR.Overexpression of NOTCH1 in SACC cells promoted cell growth, migration and invasion, and knockdown of NOTCH1 inhibited cell proliferation in vitro and tumorigenicity in vivo by inducing cell apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Preventive Dentistry, Affiliated Stomatological Hospital, Fujian Medical University, Fuzhou, China. These authors contributed equally to this work.

ABSTRACT

Background: Numerous studies have reported both the tumor-suppressive and oncogenic roles of the Notch pathway, indicating that Notch activity regulates tumor biology in a complex, context-dependent manner. The aim of the present study was to identify the role of NOTCH1 in the cell growth and metastasis of SACC.

Methods: We analyzed the expression of NOTCH1 in clinical SACC samples using immunohistochemical staining. We silenced the expression of NOTCH1 and overexpressed activated NOTCH1 to elucidate the effects of NOTCH1 on proliferation, migration and invasion. NOTCH1 target genes were validated by real-time PCR.

Results: Our results showed that NOTCH1 was upregulated in SACC tissues when compared with normal tissues, and this upregulation was further enhanced in SACC tissues with metastasis and recurrence when compared with SACC tissues without metastasis. Overexpression of NOTCH1 in SACC cells promoted cell growth, migration and invasion, and knockdown of NOTCH1 inhibited cell proliferation in vitro and tumorigenicity in vivo by inducing cell apoptosis.

Conclusions: The results of this study suggest that NOTCH1 plays a key role in the cell growth, anti-apoptosis, and metastasis of SACC. NOTCH1 inhibitors might therefore have potential therapeutic applications in treating SACC patients by inhibiting cancer cell growth and metastasis.

Show MeSH
Related in: MedlinePlus