Influence of Tamoxifen on Different Biological Pathways in Tumorigenesis and Transformation in Adipose-Derived Stem Cells, Mammary Cells and Mammary Carcinoma Cell Lines-An In Vitro Study

Abstract

Breast carcinoma is one of the most common malignant tumors in women. In cases of hormone-sensitive cells, tamoxifen as an anti-estrogenic substance is a first line medication in the adjuvant setting. The spectrum of autologous breast reconstructions ranges from fat infiltrations to complex microsurgical procedures. The influence of adipose-derived stem cells (ASC) on the tumor bed and a possibly increased recurrence rate as a result are critically discussed. In addition, there is currently no conclusive recommendation regarding tamoxifen-treated patients and autologous fat infiltrations. The aim of the present study was to investigate the effect of tamoxifen on the gene expression of a variety of genes involved in tumorigenesis, cell growth and transformation. Mammary epithelial cell line and mammary carcinoma cell lines were treated with tamoxifen in vitro as well as co-cultured with ASC. Gene expression was quantified by PCR arrays and showed increased expression in the mammary carcinoma cell lines with increasing time of treatment and concentration of tamoxifen. The data presented can be considered as an addition to the controversial discussion on the relationship between ASC and breast carcinoma cells. Further studies are needed to quantify the in vivo interaction of ASC and mammary carcinoma cells and to conclusively assess the impact of tamoxifen in reconstructive cases with fat grafting.

Keywords: ASC; PCR array; adipose derived stem cells; autologous fat grafting; breast cancer; breast reconstruction; mamma carcinoma; tamoxifen; tumorigenesis.

Figure 1

Gene expression of the examined genes of the PCR arrays of ASC, MCF-10A cells and corresponding co-cultures. (A) From left to right, ASC, MCF-10A cells and co-culture were each treated with 5 µg, 15 µg or 25 µM tamoxifen for 48 or 96 h. The color-coding (see Figure 2) indicates a relative increase (black) or reduction (red) in gene expression. The 84 genes studied and their associated pathways in tumorigenesis and transformation are listed in Table 1. (B) ASC of passage 2 after treatment with 25 µM tamoxifen after 96 h in culture. (C) MCF-10A cells of passage 29 after treatment with 25 µM tamoxifen after 96 h in culture. (D) Co-culture of ASC and MCF-10A cells after treatment with 25 µM tamoxifen after 96 h in culture.

Figure 2

Legend of the color-coding of Figure 1, Figure 3 and Figure 4. Negative values (black color-coding) indicate a relative reduction of the Ct value and thus increased gene expression. Positive values (red color-coding) indicate a relative increase in Ct values and thus reduced gene expression.

Figure 3

Gene expression of the examined genes of the PCR arrays of ASC, MCF-7 cells and corresponding co-cultures. (A) From left to right, ASC, MCF-7 cells and co-culture were each treated with 5 µg, 15 µg or 25 µM tamoxifen for 48 or 96 h. The color coding (see Figure 2) indicates a relative increase (black) or reduction (red) in gene expression. The 84 genes studied and their associated pathways in tumorigenesis and transformation are listed in Table 1. (B) ASC of passage 2 after treatment with 25 µM tamoxifen after 96 h in culture. (C) MCF-7 cells of passage 32 after treatment with 25 µM tamoxifen after 96 h in culture. (D) Co-culture of ASC and MCF-7 cells after treatment with 25 µM tamoxifen after 96 h in culture.

Figure 4

Gene expression of the examined genes of the PCR arrays of ASC, BT-474 cells and corresponding co-cultures. (A) From left to right, ASC, BT-474 cells and co-culture were each treated with 5 µg, 15 µg or 25 µM tamoxifen for 48 or 96 h. The color coding (see Figure 2) indicates a relative increase (black) or reduction (red) in gene expression. The 84 genes studied and their associated pathways in tumorigenesis and transformation are listed in Table 1. (B) ASC of passage 2 after treatment with 25 µM tamoxifen after 96 h in culture. (C) BT-474 cells of passage 15 after treatment with 25 µM tamoxifen after 96 h in culture. (D) Co-culture of ASC and BT-474 cells after treatment with 25 µM tamoxifen after 96 h in culture.

Figure 5

Expression patterns of the 14 genes with the strongest variations. For an overview of the gene abbreviations, please refer to Table 1. A positive value indicates a higher Ct value and thus down-regulated gene expression. A negative value means earlier and thus increased gene expression. (A) Expression patterns of ASC, MCF-10A cells and co-cultures. (B) Expression patterns of ASC, MCF-7 cells and co-cultures. (C) Expression patterns of ASC, BT-474 cells and co-cultures.