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ANGPTL3 is a novel biomarker as it activates ERK/MAPK pathway in oral cancer.

Koyama T, Ogawara K, Kasamatsu A, Okamoto A, Kasama H, Minakawa Y, Shimada K, Yokoe H, Shiiba M, Tanzawa H, Uzawa K - Cancer Med (2015)

Bottom Line: Significant (P < 0.05) ANGPTL3 upregulation was detected in the cell lines and most primary OSCCs (60%) compared with the normal counterparts.In vitro, cellular growth in ANGPTL3 knockdown cells significantly (P < 0.05) decreased with inactivated extracellular regulated kinase (ERK) and cell-cycle arrest at the G1 phase resulting from upregulation of the cyclin-dependent kinase inhibitors, including p21(Cip1) and p27(Kip1) .The current data indicated that ANGPTL3 may play a role in OSCCs via MAPK signaling cascades, making it a potentially useful diagnostic/therapeutic target for use in patients with OSCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Science, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8670, Japan.

No MeSH data available.


Related in: MedlinePlus

Expression of ANGPTL3 and ANGPTL3 knockdown inhibits ERK activation and promotes G1 arrest. (A) qRT-PCR shows that ANGPTL3 mRNA expression in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly (*P < 0.05, Mann–Whitney U-test) lower than in the shMock-transfected cells. (B) Immunoblotting analysis shows that the ANGPTL3 protein levels in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) also are decreased markedly compared with the shMock-transfected cells. (C) To determine the effect of shANGPTL3 on cellular proliferation, shANGPTL3-, and shMock-transfected cells were seeded in six-well plates at a density of 1 × 104 viable cells/well. Both transfectants were counted on seven consecutive days. The cellular growth of shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly inhibited compared with the shMock-transfected cells after 7 days (168 h). The results are expressed as the means ± SEM of values from three assays. The asterisks indicate significant (*P < 0.05, Mann–Whitney U-test) differences between the shANGPTL3 and shMock cells. (D) Immunoblotting analysis shows that ANGPTL3 knockdown results in decreased levels of pERK compared with the shMock-transfected cells (HSC-3- and Sa3-derived transfectants). Densitometric pERK/ERK protein data are normalized to GAPDH protein levels. (E) Immunoblotting analysis shows upregulation of p21Cip1 and p27Kip1 and downregulation of cyclin D1, cyclin E, CDK2, CDK4, and CDK6 in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) compared with the shMock-transfected cells. (F) Flow cytometric analysis was performed to investigate cell-cycle progression in the shANGPTL3- and shMock-transfected cells after synchronization at the G2/M phase to treatment with nocodazole. The percentage of cells at the G1 phase in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is increased markedly compared with the shMock- transfected cells.
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fig03: Expression of ANGPTL3 and ANGPTL3 knockdown inhibits ERK activation and promotes G1 arrest. (A) qRT-PCR shows that ANGPTL3 mRNA expression in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly (*P < 0.05, Mann–Whitney U-test) lower than in the shMock-transfected cells. (B) Immunoblotting analysis shows that the ANGPTL3 protein levels in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) also are decreased markedly compared with the shMock-transfected cells. (C) To determine the effect of shANGPTL3 on cellular proliferation, shANGPTL3-, and shMock-transfected cells were seeded in six-well plates at a density of 1 × 104 viable cells/well. Both transfectants were counted on seven consecutive days. The cellular growth of shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly inhibited compared with the shMock-transfected cells after 7 days (168 h). The results are expressed as the means ± SEM of values from three assays. The asterisks indicate significant (*P < 0.05, Mann–Whitney U-test) differences between the shANGPTL3 and shMock cells. (D) Immunoblotting analysis shows that ANGPTL3 knockdown results in decreased levels of pERK compared with the shMock-transfected cells (HSC-3- and Sa3-derived transfectants). Densitometric pERK/ERK protein data are normalized to GAPDH protein levels. (E) Immunoblotting analysis shows upregulation of p21Cip1 and p27Kip1 and downregulation of cyclin D1, cyclin E, CDK2, CDK4, and CDK6 in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) compared with the shMock-transfected cells. (F) Flow cytometric analysis was performed to investigate cell-cycle progression in the shANGPTL3- and shMock-transfected cells after synchronization at the G2/M phase to treatment with nocodazole. The percentage of cells at the G1 phase in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is increased markedly compared with the shMock- transfected cells.

Mentions: Since frequent upregulation of ANGPTL3 was observed in OSCC-derived cells (Fig.1), the OSCC-derived cells (HSC-3 and Sa3) were transfected with ANGPTL3 shRNA and shMock as controls. To confirm that shANGPTL3 transfection works and ANGPTL3 mRNA and protein decrease, we performed qRT-PCR and Western blotting (Fig.3A and B, respectively). The ANGPTL3 mRNA expression in shANGPTL3 cells was significantly (P < 0.05) lower than in shMock cells (Fig.3A). The ANGPTL3 protein level in the shANGPTL3 cells also was decreased compared with shMock cells (Fig.3B).


ANGPTL3 is a novel biomarker as it activates ERK/MAPK pathway in oral cancer.

Koyama T, Ogawara K, Kasamatsu A, Okamoto A, Kasama H, Minakawa Y, Shimada K, Yokoe H, Shiiba M, Tanzawa H, Uzawa K - Cancer Med (2015)

Expression of ANGPTL3 and ANGPTL3 knockdown inhibits ERK activation and promotes G1 arrest. (A) qRT-PCR shows that ANGPTL3 mRNA expression in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly (*P < 0.05, Mann–Whitney U-test) lower than in the shMock-transfected cells. (B) Immunoblotting analysis shows that the ANGPTL3 protein levels in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) also are decreased markedly compared with the shMock-transfected cells. (C) To determine the effect of shANGPTL3 on cellular proliferation, shANGPTL3-, and shMock-transfected cells were seeded in six-well plates at a density of 1 × 104 viable cells/well. Both transfectants were counted on seven consecutive days. The cellular growth of shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly inhibited compared with the shMock-transfected cells after 7 days (168 h). The results are expressed as the means ± SEM of values from three assays. The asterisks indicate significant (*P < 0.05, Mann–Whitney U-test) differences between the shANGPTL3 and shMock cells. (D) Immunoblotting analysis shows that ANGPTL3 knockdown results in decreased levels of pERK compared with the shMock-transfected cells (HSC-3- and Sa3-derived transfectants). Densitometric pERK/ERK protein data are normalized to GAPDH protein levels. (E) Immunoblotting analysis shows upregulation of p21Cip1 and p27Kip1 and downregulation of cyclin D1, cyclin E, CDK2, CDK4, and CDK6 in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) compared with the shMock-transfected cells. (F) Flow cytometric analysis was performed to investigate cell-cycle progression in the shANGPTL3- and shMock-transfected cells after synchronization at the G2/M phase to treatment with nocodazole. The percentage of cells at the G1 phase in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is increased markedly compared with the shMock- transfected cells.
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fig03: Expression of ANGPTL3 and ANGPTL3 knockdown inhibits ERK activation and promotes G1 arrest. (A) qRT-PCR shows that ANGPTL3 mRNA expression in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly (*P < 0.05, Mann–Whitney U-test) lower than in the shMock-transfected cells. (B) Immunoblotting analysis shows that the ANGPTL3 protein levels in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) also are decreased markedly compared with the shMock-transfected cells. (C) To determine the effect of shANGPTL3 on cellular proliferation, shANGPTL3-, and shMock-transfected cells were seeded in six-well plates at a density of 1 × 104 viable cells/well. Both transfectants were counted on seven consecutive days. The cellular growth of shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is significantly inhibited compared with the shMock-transfected cells after 7 days (168 h). The results are expressed as the means ± SEM of values from three assays. The asterisks indicate significant (*P < 0.05, Mann–Whitney U-test) differences between the shANGPTL3 and shMock cells. (D) Immunoblotting analysis shows that ANGPTL3 knockdown results in decreased levels of pERK compared with the shMock-transfected cells (HSC-3- and Sa3-derived transfectants). Densitometric pERK/ERK protein data are normalized to GAPDH protein levels. (E) Immunoblotting analysis shows upregulation of p21Cip1 and p27Kip1 and downregulation of cyclin D1, cyclin E, CDK2, CDK4, and CDK6 in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) compared with the shMock-transfected cells. (F) Flow cytometric analysis was performed to investigate cell-cycle progression in the shANGPTL3- and shMock-transfected cells after synchronization at the G2/M phase to treatment with nocodazole. The percentage of cells at the G1 phase in the shANGPTL3-transfected cells (HSC-3- and Sa3-derived transfectants) is increased markedly compared with the shMock- transfected cells.
Mentions: Since frequent upregulation of ANGPTL3 was observed in OSCC-derived cells (Fig.1), the OSCC-derived cells (HSC-3 and Sa3) were transfected with ANGPTL3 shRNA and shMock as controls. To confirm that shANGPTL3 transfection works and ANGPTL3 mRNA and protein decrease, we performed qRT-PCR and Western blotting (Fig.3A and B, respectively). The ANGPTL3 mRNA expression in shANGPTL3 cells was significantly (P < 0.05) lower than in shMock cells (Fig.3A). The ANGPTL3 protein level in the shANGPTL3 cells also was decreased compared with shMock cells (Fig.3B).

Bottom Line: Significant (P < 0.05) ANGPTL3 upregulation was detected in the cell lines and most primary OSCCs (60%) compared with the normal counterparts.In vitro, cellular growth in ANGPTL3 knockdown cells significantly (P < 0.05) decreased with inactivated extracellular regulated kinase (ERK) and cell-cycle arrest at the G1 phase resulting from upregulation of the cyclin-dependent kinase inhibitors, including p21(Cip1) and p27(Kip1) .The current data indicated that ANGPTL3 may play a role in OSCCs via MAPK signaling cascades, making it a potentially useful diagnostic/therapeutic target for use in patients with OSCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Science, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8670, Japan.

No MeSH data available.


Related in: MedlinePlus