Limits...
Expression of CD24 is associated with HER2 expression and supports HER2-Akt signaling in HER2-positive breast cancer cells.

Hosonaga M, Arima Y, Sugihara E, Kohno N, Saya H - Cancer Sci. (2014)

Bottom Line: We found that expression of CD24 was increased by stable overexpression of HER2.Knockdown of CD24 also suppressed the phosphorylation of Akt, which functions downstream of HER2 and PI3K to promote cell survival.Our results thus indicate that CD24 supports both the expression of HER2 and the consequent activation of PI3K-Akt signaling.

View Article: PubMed Central - PubMed

Affiliation: Division of Gene Regulation, Institute for Advanced Medical Research, School of Medicine, Keio University, Tokyo, Japan; Department of Breast Oncology, Tokyo Medical University, Tokyo, Japan.

Show MeSH

Related in: MedlinePlus

Effects of overexpression of human epidermal growth factor receptor 2 (HER2) on cancer stem cell (CSC) properties. (a) Tumorsphere formation by 231-Luc and HER2-90 cells. Tumorspheres were photographed after culture of the cells for 6 days. Scale bar, 100 μm. (b) Quantitation of tumorsphere formation by 231-Luc and HER2-90 cells. The number of tumorspheres with a diameter of >100 μm was counted after cell culture for 6 days. Data are means ± SD for triplicate experiments. **P < 0.01 (Student's t test). (c) Aldehyde dehydrogenase (ALDH) activity of 231-Luc and HER2-90 cells as determined with an ALDEFLUOR kit. Cells were incubated with the ALDEFLUOR substrate in the absence or presence of the specific ALDH inhibitor diethylaminobenzal-dehyde (DEAB) and then analyzed by flow cytometry. The percentage of ALDEFLUOR-positive cells is indicated. (d) Comparison of tumor-initiating activity between 231-Luc and HER2-90 cells. Tumor formation was evaluated 8 weeks after injection of the indicated numbers of cells into a mammary fat pad of NOD/SCID mice. (e,f) Quantitative RT-PCR analysis of mRNA derived from epithelial-mesenchymal transition-related (e) or ES cell–related (f) genes in 231-Luc and HER2-90 cells. Data are normalized by the amount of HPRT1 mRNA, expressed relative to the corresponding normalized value for 231-Luc cells, and are presented as means ± SD for triplicate experiments. *P < 0.05, **P < 0.01 (Student's t-test). NS, not significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4317915&req=5

fig02: Effects of overexpression of human epidermal growth factor receptor 2 (HER2) on cancer stem cell (CSC) properties. (a) Tumorsphere formation by 231-Luc and HER2-90 cells. Tumorspheres were photographed after culture of the cells for 6 days. Scale bar, 100 μm. (b) Quantitation of tumorsphere formation by 231-Luc and HER2-90 cells. The number of tumorspheres with a diameter of >100 μm was counted after cell culture for 6 days. Data are means ± SD for triplicate experiments. **P < 0.01 (Student's t test). (c) Aldehyde dehydrogenase (ALDH) activity of 231-Luc and HER2-90 cells as determined with an ALDEFLUOR kit. Cells were incubated with the ALDEFLUOR substrate in the absence or presence of the specific ALDH inhibitor diethylaminobenzal-dehyde (DEAB) and then analyzed by flow cytometry. The percentage of ALDEFLUOR-positive cells is indicated. (d) Comparison of tumor-initiating activity between 231-Luc and HER2-90 cells. Tumor formation was evaluated 8 weeks after injection of the indicated numbers of cells into a mammary fat pad of NOD/SCID mice. (e,f) Quantitative RT-PCR analysis of mRNA derived from epithelial-mesenchymal transition-related (e) or ES cell–related (f) genes in 231-Luc and HER2-90 cells. Data are normalized by the amount of HPRT1 mRNA, expressed relative to the corresponding normalized value for 231-Luc cells, and are presented as means ± SD for triplicate experiments. *P < 0.05, **P < 0.01 (Student's t-test). NS, not significant.

Mentions: To determine if HER2 overexpression might affect CSC properties, we examined sphere formations of HER2-90 and 231-Luc cells. The numbers of spheres formed with a diameter of >100 μm were counted. HER2-90 cells formed significantly fewer spheres than did 231-Luc cells (Fig. 2a,b). Cells were also cultured using 1% methylcellulose containing a medium to avoid cell aggregation. In contrast to 231-Luc cells, HER2-90 cells did not form sphere-like colonies in the presence of methylcellulose (Fig. 2a,b). An examination of ALDH1 activity, a functional marker for breast cancer CSC,(17) revealed that overexpression of HER2 decreased the proportion of ALDEFLUOR-positive CSC (Fig. 2c). To determine the ability of 231-Luc and HER2-90 cells to initiate tumors, we injected cells into a mammary fat pad of female NOD/SCID mice, which revealed that there was no difference in the frequency of tumor initiation (Fig. 2d). These results suggested that overexpression of HER2 decreased representative CSC properties, such as the sphere-forming ability and ALDH1 activity, and did not affect the tumorigenicity of 231-Luc triple-negative breast cancer cells.


Expression of CD24 is associated with HER2 expression and supports HER2-Akt signaling in HER2-positive breast cancer cells.

Hosonaga M, Arima Y, Sugihara E, Kohno N, Saya H - Cancer Sci. (2014)

Effects of overexpression of human epidermal growth factor receptor 2 (HER2) on cancer stem cell (CSC) properties. (a) Tumorsphere formation by 231-Luc and HER2-90 cells. Tumorspheres were photographed after culture of the cells for 6 days. Scale bar, 100 μm. (b) Quantitation of tumorsphere formation by 231-Luc and HER2-90 cells. The number of tumorspheres with a diameter of >100 μm was counted after cell culture for 6 days. Data are means ± SD for triplicate experiments. **P < 0.01 (Student's t test). (c) Aldehyde dehydrogenase (ALDH) activity of 231-Luc and HER2-90 cells as determined with an ALDEFLUOR kit. Cells were incubated with the ALDEFLUOR substrate in the absence or presence of the specific ALDH inhibitor diethylaminobenzal-dehyde (DEAB) and then analyzed by flow cytometry. The percentage of ALDEFLUOR-positive cells is indicated. (d) Comparison of tumor-initiating activity between 231-Luc and HER2-90 cells. Tumor formation was evaluated 8 weeks after injection of the indicated numbers of cells into a mammary fat pad of NOD/SCID mice. (e,f) Quantitative RT-PCR analysis of mRNA derived from epithelial-mesenchymal transition-related (e) or ES cell–related (f) genes in 231-Luc and HER2-90 cells. Data are normalized by the amount of HPRT1 mRNA, expressed relative to the corresponding normalized value for 231-Luc cells, and are presented as means ± SD for triplicate experiments. *P < 0.05, **P < 0.01 (Student's t-test). NS, not significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4317915&req=5

fig02: Effects of overexpression of human epidermal growth factor receptor 2 (HER2) on cancer stem cell (CSC) properties. (a) Tumorsphere formation by 231-Luc and HER2-90 cells. Tumorspheres were photographed after culture of the cells for 6 days. Scale bar, 100 μm. (b) Quantitation of tumorsphere formation by 231-Luc and HER2-90 cells. The number of tumorspheres with a diameter of >100 μm was counted after cell culture for 6 days. Data are means ± SD for triplicate experiments. **P < 0.01 (Student's t test). (c) Aldehyde dehydrogenase (ALDH) activity of 231-Luc and HER2-90 cells as determined with an ALDEFLUOR kit. Cells were incubated with the ALDEFLUOR substrate in the absence or presence of the specific ALDH inhibitor diethylaminobenzal-dehyde (DEAB) and then analyzed by flow cytometry. The percentage of ALDEFLUOR-positive cells is indicated. (d) Comparison of tumor-initiating activity between 231-Luc and HER2-90 cells. Tumor formation was evaluated 8 weeks after injection of the indicated numbers of cells into a mammary fat pad of NOD/SCID mice. (e,f) Quantitative RT-PCR analysis of mRNA derived from epithelial-mesenchymal transition-related (e) or ES cell–related (f) genes in 231-Luc and HER2-90 cells. Data are normalized by the amount of HPRT1 mRNA, expressed relative to the corresponding normalized value for 231-Luc cells, and are presented as means ± SD for triplicate experiments. *P < 0.05, **P < 0.01 (Student's t-test). NS, not significant.
Mentions: To determine if HER2 overexpression might affect CSC properties, we examined sphere formations of HER2-90 and 231-Luc cells. The numbers of spheres formed with a diameter of >100 μm were counted. HER2-90 cells formed significantly fewer spheres than did 231-Luc cells (Fig. 2a,b). Cells were also cultured using 1% methylcellulose containing a medium to avoid cell aggregation. In contrast to 231-Luc cells, HER2-90 cells did not form sphere-like colonies in the presence of methylcellulose (Fig. 2a,b). An examination of ALDH1 activity, a functional marker for breast cancer CSC,(17) revealed that overexpression of HER2 decreased the proportion of ALDEFLUOR-positive CSC (Fig. 2c). To determine the ability of 231-Luc and HER2-90 cells to initiate tumors, we injected cells into a mammary fat pad of female NOD/SCID mice, which revealed that there was no difference in the frequency of tumor initiation (Fig. 2d). These results suggested that overexpression of HER2 decreased representative CSC properties, such as the sphere-forming ability and ALDH1 activity, and did not affect the tumorigenicity of 231-Luc triple-negative breast cancer cells.

Bottom Line: We found that expression of CD24 was increased by stable overexpression of HER2.Knockdown of CD24 also suppressed the phosphorylation of Akt, which functions downstream of HER2 and PI3K to promote cell survival.Our results thus indicate that CD24 supports both the expression of HER2 and the consequent activation of PI3K-Akt signaling.

View Article: PubMed Central - PubMed

Affiliation: Division of Gene Regulation, Institute for Advanced Medical Research, School of Medicine, Keio University, Tokyo, Japan; Department of Breast Oncology, Tokyo Medical University, Tokyo, Japan.

Show MeSH
Related in: MedlinePlus