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 Table of Contents  
ORIGINAL ARTICLE
Year : 2017  |  Volume : 13  |  Issue : 4  |  Page : 707-714

Expression and significance of Twist, estrogen receptor, and E-cadherin in human breast cancer cells and tissues


1 Research Center of Combine Traditional Chinese and Western Medicine, Affiliated Traditional Medicine Hospital, Southwest Medical University, Luzhou, Sichuan, China
2 The Institute for Cancer Medicine, Research Center for Preclinical Medicine and College of Basic Medical Sciences, Southwest Medical University, Luzhou, Sichuan, China

Date of Web Publication13-Sep-2017

Correspondence Address:
Tao He
Institute of Ontology, Southwest Medical University, Luzhou 646000, Sichuan
China
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jcrt.JCRT_1396_16

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 > Abstract 

Objectives: Breast cancer is one of the most common malignancies in women, and the tumor cells' invasion and metastasis is the main cause of death. Recent reports showed that Twist, a transcription factor, plays multiple roles in breast cancer initiation, progress, and metastasis. However, the underlying mechanisms of Twist in tumor invasion and metastasis of breast cancer still remain unclear. Here, we examined the correlation between Twist, E-cadherin, and estrogen receptor (ER) in promoting invasion and metastasis in breast cancer cells and tissues.
Materials and Methods: The mRNA and protein expression of Twist, E-cadherin, and ER in breast cancer cell lines (MCF-7, MDA-MB-435, MDA-MB-231, and ZR-75-30) and human invasive ductal carcinoma (IDC) tissues from 32 patients were detected by reverse transcription-polymerase chain reaction and immunohistochemistry (IHC), respectively.
Results: Expression of Twist in cells with high ability of invasion and metastasis was higher than that in MCF-7 cell line which has low ability of invasion and metastasis, while the expression of ER and E-cadherin was much more higher in MCF-7 cell line than in other cells. IHC showed that the expression rate of Twist in IDC tissues and adjacent tissues was 84.38% and 31.25% and the positive expression of E-cadherin and ER was 21.88% and 40.63% in IDC tissues and 81.25% and 84.38% in adjacent tissues, respectively. Interestingly, overexpression of Twist promoted cellular invasion and metastasis and decreased the expression of E-cadherin, ER, AKT, and p-AKT in HEK-293 cells.
Conclusions: Taken together, these findings demonstrated that Twist was upregulated in high invasion and metastasis cell lines as well as IDC tissues companioned with downregulated expression of E-cadherin and ER, which provides important clues for the deeper study of breast cancer.

Keywords: Breast cancer, E-cadherin, estrogen receptor, invasion, metastasis, Twist


How to cite this article:
Tan R, Wang L, Song J, Li J, He T. Expression and significance of Twist, estrogen receptor, and E-cadherin in human breast cancer cells and tissues. J Can Res Ther 2017;13:707-14

How to cite this URL:
Tan R, Wang L, Song J, Li J, He T. Expression and significance of Twist, estrogen receptor, and E-cadherin in human breast cancer cells and tissues. J Can Res Ther [serial online] 2017 [cited 2018 May 22];13:707-14. Available from: http://www.cancerjournal.net/text.asp?2017/13/4/707/214462


 > Introduction Top


Tumor metastasis is the most common cause of death in cancer patients, especially breast cancer in woman.[1],[2] At the late stage of breast cancer, it frequently presents distant metastases with poor prognosis, which increases the difficulty of cancer therapy. Therefore, it is important to reveal the undergoing mechanisms of cancer metastasis including in breast cancer. Twist, a member of the basic helix-loop-helix (bHLH) transcription factor family with a bHLH domain, mediates heterodimerization or homodimerization.[3] Previous studies have shown that the patients with poor prognosis had increased Twist expression and knockdown of Twist lowered the tumor cells' metastasis and invasiveness.[4] During the development of metastasis, Twist plays a key role in the regulation of cancer cells metastasis through downregulating the E-cadherin and β-catenin as well as promoting the epithelial-mesenchymal transition (EMT).[5],[6],[7],[8],[9] Moreover, the high levels of Twist are associated with the high expression of endothelial growth factor, which accelerates the angiogenesis and associates the instability of chromosome in breast cancers.[10] However, the molecular mechanism of Twist in the metastasis of breast cancer remains unclear.

E-cadherin, a single-span transmembrane glycoprotein, has been widely considered to be a marker of the EMT process that is the basic mechanism of tumor progression.[11] The E-cadherin/catenin adhesion system is crucial for cell adhesion, which maintains the structural integrity and cell polarity.[12],[13] In breast cancers, E-cadherin has been found to be loss of expression or reduced expression, which positively correlates with the ongoing metastasis and poor prognosis.[14] On the other hand, the estrogen receptor (ER) has been reported decreased in breast cancer, which was implicated in the initiation and/or development of the malignancy.[15] Clinical studies have showed that loss of expression or reduced expression of ER gene has been closely associated with the poor differentiation of cancer cells and the poor prognosis.[16],[17],[18],[19] Furthermore, protein kinase B, also known as Akt, plays a key role in multiple cellular processes, which has recently been reported to suppress tumorigenesis, acting anti-oncogenically by suppressing invasion and metastasis.[20] Although a number of studies have reported the association of E-cadherin and ER in breast cancer, the association of Twist and these two genes with tumor invasion and migration remains need to be explored.

The present study was performed to investigate the association of Twist, E-cadherin, and ER with breast cancer invasion and migration. To detect mRNA and protein levels of Twist, E-cadherin, and ER in breast cancer cell lines and cancer tissues, we used reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) to test four kinds of cell lines and tissues from 32 patients. In addition, Twist was overexpressed by transfecting pcDNA5/FRT/TO-Twist vector to HEK-293 cells to reveal the underlying mechanism of Twist promoting cancer invasion and migration.


 > Materials and Methods Top


Sample collection

The human invasive ductal carcinoma (IDC) and adjacent tissues (cancer edge distance at least 3 cm) were obtained from the affiliated hospital of Southwest Medical University. All patients were females and aged from 37 to 69 years. All patients had not received preoperative radiation therapy and chemotherapy before the breast cancer operation, and the diagnosis of breast cancer was made by two pathologists review, independently.

Cell culture

The breast carcinoma cell lines MDA-MB-435, MDA-MB-231, ZR-75-30, and MCF-7 were purchased from the Type Culture Collection of the Chinese Academy of Sciences, Shanghai, China. Flp-In-T-REx-293 cell was purchased from Invitrogen. All cell lines were cultured in H-Dulbecco's Modified Eagle's Medium (DMEM) medium with 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 0.1 mg/mL streptomycin at 37°C in a 5% CO2 humidified atmosphere.

Construction of HEK-293-Vector and HEK-293-Twist cells

The plasmid pcDNA5/FRT/TO and pcDNA3-Twist were gifts from Infection and Immunology Laboratory of Southwest Medical University in China. These two plasmids were digested with restriction enzyme NotI and BamHI to release the vector backbone and hTwist fragment and recombined to construct the vector pcDNA5/FRT/TO-Twist. HEK-293-Twist cells were generated by cotransfecting 7.5 μg of pOG44 with 0.75 μg of pcDNA5/FRT/TO-Twist to Flp-In-T-REx-293 cells, and HEK-293-Vector cells were obtained by cotransfecting 7.5 μg of pOG44 with 0.75 μg of pcDNA5/FRT/TO to Flp-In-T-REx-293 cells. Before performing IHC and cell migration test, HEK-293-Twist and HEK-293-Vector cells were cultured in medium containing 0.1 μg/ml tetracycline.

In vitro scratch assay

HEK-293-Vector and HEK-293-Twist cells on logarithmic growth phase were digested by 2.5 g/L trypsin and plated into a 6-well plate. When cell culture reached approximately 80% confluence, the medium was replaced with non-FBS DMEM. After culture reached approximately 90% confluence, scraping the cell monolayer in a straight line to create a “scratch” with a 200 μl pipet tip. Remove the debris by washing the cells once with PBS, and keep culturing cell in blank DMEM at 37°C in a 5% CO2 humidified atmosphere. Take photographs at the time point of 0 h, 7 h, and 24 h. Experiments were done in triplicate.

Reverse transcription-polymerase chain reaction assay

Total RNA of MDA-MB-435, MDA-MB-231, ZR-75-30, and MCF-7 cells was extracted using RNAprep Pure Cell/Bacteria Kit (Tiangen, Beijing, China), followed by synthesizing cDNA from total RNA using the PrimeScript™ RT reagent Kit (TAKARA, Dalian, China). Total RNA of breast cancer tissues and adjacent tissues was harvested using RNAprep Pure Tissue Kit (Tiangen, Beijing, China), and the cDNA synthesis was also performed using PrimeScript™ RT reagent Kit (TAKARA, Dalian, China). PCR amplification of collected cDNA (2 μl) was performed in 20 μl reaction system consisting of 1 μl forward primer, 1 μl reverse primer, 10 μl 2 × Taq PCR MasterMix, and 6 μl RNase-Free H2O at 94°C for 10 min, followed by 25 cycles of 30 s denaturation at 94°C, 30 s annealing at 58°C, and 30 s extension at 72°C. The final concentration of primers is 0.5 μM. Primers used in this reaction were as follows: Twist: Forward, 5'-GTGCGCAGTCTTACGAGGAG-3', Reverse, 5'-GCTTGAGGGTCAGAATCTTGCT-3', product size: 156 bp; E-cadherin: Forward, 5'-TGGACCGAGAGAGTTTCC-3', Reverse, 5'-AATATGGTGTATACAGCCTC-3', product size: 250 bp; ER: Forward, 5'-GTGCCTGGCTAGAGATCCTG-3', Reverse, 5'-AGAGACTTCAGGGTGCTGGA-3', product size: 265 bp; glyceraldehyde-3-phosphate dehydrogenase (GAPDH): Forward, 5'-ATGCTGGCGCTGAGTACGTC-3', Reverse, 5'-GGTCATGAGTCCTTCCACGATA-3', product size: 263 bp. The RT-PCR images were taken by ChemiDOC XRS (Bio-Rad) and analyzed by QuantityOne software (Bio-Rad). Experiments were done in triplicate.

Immunohistochemistry staining

Fresh tumor tissues and the adjacent tissues were fixed in 10% formalin and embedded in paraffin. Serial sections (4 μm thick) were taken to deparaffinized in xylene and rehydrated in a series of diluted alcohol and distilled water. Then, after antigen retrieval, the activity of endogenous peroxidase was blocked using 0.3% hydrogen peroxide in methanol for 0.5 h. To block the nonspecific background, the sections were rehydrated in 1 × PAB and incubated with 5% rabbit serum for 0.5 h. The sections were subsequently incubated with Twist (Diluted 1:500; Santa Cruz, Dallas, USA), E-cadherin (Diluted 1:200; Santa Cruz, Dallas, USA), and ER (Diluted 1:500; Bioworld, St. Louis Park, USA) antibody at 4°C overnight. After that, the slides were incubated with the secondary antibodies at dilution of 1:500 at room temperature for 0.5 h and then incubated with peroxidase-conjugated avidin-biotin complexes, counterstained with hemotoxylin. Finally, the images of IHC were taken by the standard light microscopy and analyzed by ImagePro-Plus 6.0 (Media Cybernetics, MD, USA) software.

Four kinds of cancer cells, HEK-293-Vector cells, and HEK-293-Twist cells were grown on glass coverslips with a diameter of 13 mm for immunostaining. All cells on coverslips were fixed by 2% paraformaldehyde after washing with PBS for three times. Immunostaining was performed with SABC kit (ZSGB-Bio, Beijing, China) according to the manufacturer's instruction. The primary antibodies were used as in tissue IHC. Experiments were done in triplicate.

Statistic analysis

All data are presented as means ± standard deviation, and all results were analyzed using SPSS statistical software (Version 21.0; SPSS, Chicago, IL, USA). Chi-square test was used to detect the expression differences of Twist, E-cadherin, and ER between breast cancer and adjacent cancer tissues. Spearman's rank correlation coefficient was used to assess the statistical correlation between the protein expression of Twist, E-cadherin, and ER in breast cancer tissues. P < 0.05 was considered to be statistically significant.


 > Results Top


RNA expression of Twist, E-cadherin, and estrogen receptor in low and high metastatic breast cancer cell lines

To investigate the differences of RNA expression between Twist, E-cadherin, and ER in low metastatic cell line of MCF7 and high metastatic cell lines of MDA-MB-435, MDA-MB-231, and ZR-75-30, semi-quantitative PCR was performed, respectively. The results showed that the expression of Twist in high metastatic cell lines was higher than that in low metastatic cell line. In contrast, expression of E-cadherin and ER in high metastatic cells was lower than those in low metastatic cells [Figure 1]a, suggesting that higher expression of Twist and lower expression of E-cadherin and ER were related with promoting metastasis in breast cancer. The relative intensity of RNA expression of Twist, E-cadherin, and ER to GAPDH is illustrated in [Figure 1]b.
Figure 1: The mRNA expression of Twist, E-cadherin, and estrogen receptor in MDA-MB-435, MDA-MB-231, ZR-75-30, and MCF-7 cells. (a) Reverse transcription-polymerase chain reaction was performed to detect the mRNA levels of Twist, E-cadherin, and estrogen receptor in three kinds of high invasion and metastasis breast cancer cell lines and one of low invasion and metastasis cells. (b) Relative intensity of Twist, E-cadherin, and estrogen receptor mRNA levels to glyceraldehyde-3-phosphate dehydrogenase. Compared within MCF-7 cells, mRNA level of Twist was significantly increased in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells. *P < 0.05 versus mRNA expression in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells. Moreover, the mRNA expression of Twist in MCF-7 cells was significantly lower than that in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells. **P < 0.01 versus mRNA expression of E-cadherin and estrogen receptor in same cell line

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Immunocytochemistry analysis for Twist, E-cadherin, and estrogen receptor in breast cancer cell lines

The results showed that the protein expression of Twist was significantly higher in high invasive and metastatic breast cancer cell lines MDA-MB-435, MDA-MB-231, ZR-75-30 than those in low-invasive and low-metastatic breast carcinoma cell line MCF-7. However, the protein expression level of E-cadherin and ER was lower in the MCF-7 cell line than that in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells. Negative correlation was observed between the expression of Twist and E-cadherin/ER protein, while positive correlation was gained between the expression of ER and E-cadherin protein. Data are illustrated in [Figure 2].
Figure 2: The protein expression of Twist, E-cadherin, and estrogen receptor in MDA-MB-435, MDA-MB-231, ZR-75-30, and MCF-7 cells. The immunohistochemistry results (×200) showed that protein expression of Twist was elevated in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells, which was opposite in MCF-7 cell. However, the protein expression of E-cadherin and estrogen receptor were decreased in MDA-MB-435, MDA-MB-231, and ZR-75-30 cells with higher ability of invasion and metastasis, these were consistent with the reverse transcription polymerase chain reaction results

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RNA expression of Twist, E-cadherin, and estrogen receptor in breast cancer tissues and adjacent tissues

To examine the expression of Twist, ER, and E-cadherin in breast cancer tissues and adjacent tissues, RT-PCR was performed with the mRNA from eight patients' tissues [Figure 3]a. In [Figure 3], C presents cancer tissue and A presents adjacent tissue. As shown in [Figure 3]b, the relative expression of Twist (Twist/GAPDH) in cancer tissues was higher than that in adjacent tissues (P < 0.01). In contrast, the expression of ER [Figure 3]c, P < 0.05] and E-cadherin [Figure 3]d, P < 0.01] in cancer tissues was lower than those in adjacent tissues, which was consistent with the results from cell test. Compared with the expression of ER and E-cadherin, the expression of Twist was higher in cancer tissues and lower in adjacent tissues [Figure 3]e. This also keeps in line with the results in tumor cells. In summary, expression of Twist was raised and expression of ER and E-cadherin was downregulated in breast cancer cells and tissues. Meanwhile, expression of Twist was also higher than ER and E-cadherin in vivo and in vitro.
Figure 3: The mRNA expression of Twist, E-cadherin, and estrogen receptor in breast cancer tissues and adjacent tissues. (a) The reverse transcription polymerase chain reaction results of Twist, E-cadherin, and estrogen receptor expression in breast cancer tissues and adjacent tissues. “C” presents cancer tissue and “A” presents adjacent tissue. (b) The relative intensity of Twist to glyceraldehyde-3-phosphate dehydrogenase. “P” presents patient. The data indicated higher expression of Twist in cancer tissues than that in adjacent tissues. (c) The relative intensity of estrogen receptor to glyceraldehyde-3-phosphate dehydrogenase. (d) The relative intensity of E-cadherin to glyceraldehyde-3-phosphate dehydrogenase. (e) The relative intensity of Twist, E-cadherin, and estrogen receptor to glyceraldehyde-3-phosphate dehydrogenase. *P < 0.05 versus mRNA expression of estrogen receptor in adjacent tissue. **P < 0.01 versus mRNA expression in adjacent tissue

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Protein expression of Twist, E-cadherin, and estrogen receptor in breast cancer tissues

From the IHC results of the cancer tissues from 32 patients, we found that the protein levels of Twist in cancer tissues were significantly higher than that in the adjacent cancer tissues. The positive rate of Twist in cancer tissues and adjacent cancer tissues was 84.38% and 31.25%, respectively (P < 0.001). However, the expression levels of E-cadherin were reduced in the cancer tissues, and the positive rate of E-cadherin in cancer tissues and adjacent tissues was 21.88% and 81.25%, respectively (P < 0.001). Consistent with E-cadherin, the cancer tissues showed the significantly lower ER protein levels than in adjacent tissues, and the positive rate of ER in cancer tissues and adjacent tissues was 40.63% and 84.38%, respectively (P = 0.001). Data are presented in [Figure 4] and [Table 1].
Figure 4: The protein expression of Twist, E-cadherin, and estrogen receptor in breast cancer tissues and adjacent tissues. The immunohistochemistry results (magnification 200×) showed that protein expression of Twist was higher in cancer tissues than that in adjacent tissues. In contrast, the protein expression of E-cadherin and estrogen receptor was lower in cancer tissues than that in adjacent tissues; these were consistent with the reverse transcription-polymerase chain reaction results in tissues

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Table 1: The positive rate of twist, E-cadherin, and estrogen receptor protein in invasive ductal carcinoma tissues and adjacent tissues

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Overexpression of Twist in HEK-293 cells promoted cell migration

We upregulated the expression of Twist in HEK-293 cells by transfecting pcDNA5/FRT/TO-Twist plasmid and scraped the cell monolayer in a straight line to observe the migration of HEK-293-Twist and HEK-293-Vector cells after scraping in 0, 7, and 24 h. As shown in [Figure 5]a, at the beginning of scraping, the cell monolayer forms an equidistant scratch. However, changes were observed after 7 h culture. The HEK-293-Twist cells moved forward and closed the gap more rapid than HEK-293-Vector cells, suggesting that HEK-293-Twist cells showed faster migration than HEK-293-Vector cells after 24 h culture, and the gap in former was almost closed. These data implied that overexpression of Twist promoted in vitro cell migration.
Figure 5: Overexpression of Twist increases migratory ability of HEK-293 cell and downregulates expression of E-cadherin, estrogen receptor, AKT, and p-AKT. (a) Scratch assay was performed to detect the migratory ability of HEK-293-Twist and HEK-293-Vector cells. HEK-293-Twist cells showed stronger migratory ability, which closed the gap faster than HEK-293-Vector cells at 7 h and 24 h. (b) The immunohistochemistry results (×200) demonstrated that overexpression Twist in HEK-293 cells down-regulated expression of E-cadherin, estrogen receptor, AKT, and p-AKT

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Overexpression of Twist in HEK-293 cells downregulated the expression of E-cadherin and estrogen receptor

To investigate the influences of the expression of ER and E-cadherin after overexpressing Twist, we performed IHC in HEK-293-Twist and HEK-293-Vector cells. The results illustrated that the expression of ER and E-cadherin was downregulated when the expression of Twist was raised [Figure 5]b, which suggested the negative relationship between Twist and ER/E-cadherin. At the same time, we found that the expression of AKT and phosphorylated AKT, which were known as important factors to inhibit infiltration and migration in cancer, was also decreased when Twist was overexpressed. These data suggested that overexpression of Twist may promote tumor metastasis through inhibiting expression of ER and E-cadherin, and AKT as well as p-AKT may also involve in the tumor metastasis induced by Twist.

Correlation between Twist, E-cadherin, and estrogen receptor in breast cancer tissues

The statistical correlation between the protein expression of Twist, E-cadherin, and ER in breast cancer tissues was analyzed by Spearman's rank correlation coefficient [Table 2] and [Table 3]. In the 32 of breast cancer tissues, three of them were Twist and E-cadherin positive, seven of them were ER and E-cadherin positive, and eight of them were ER and Twist positive [Table 2] and [Table 3]. The statistical results showed that the protein expression correlation between Twist and E-cadherin was significantly negatively related (r = −0.605, P < 0.001), the protein expression correlation between ER and E-cadherin was significantly positively related (r = 0.640, P = 0.001), and the protein expression correlation between Twist and ER was significantly negatively related (r = −0.520, P = 0.002).
Table 2: Correlation analysis of protein expression between Twist and E-cadherin, estrogen receptor, and E-cadherin in breast cancer tissues by immunohistochemistry

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Table 3: Correlations of protein expression between twist and estrogen receptor in breast cancer tissues by immunohistochemistry

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 > Discussion Top


In this present study, we analyzed the association of expression levels of Twist, E-cadherin, and ER in MDA-MB-435, MDA-MB-231, ZR-75-30, and MCF-7 breast cancer cell lines as well as in human IDC tissues and adjacent tissues. We found that the mRNA and protein expression of Twist were higher in high metastatic breast cancer cell lines than in low metastatic cell line, and consistent with the results in cells, the mRNA and protein expression of Twist in cancer tissues were higher than in adjacent tissues. In contrast, mRNA and protein expression of E-cadherin and ER were lower in high metastatic cells and cancer tissues. Subsequently, we demonstrated that upregulating the expression of Twist increased the ability of migration of HEK-293 cells and decreased the expression of E-cadherin, ER, AKT, and p-AKT. Furthermore, statistical results showed that there was a correlation between the expression of Twist, E-cadherin, and ER in breast cancer.

Previous studies have showed that Twist is increased in breast cancer tissue that correlates with poor prognosis and Twist promotes the metastasis of tumor cell might due to (1) the higher vascular endothelial growth factor expression and (2) the association with the instability of chromosome in cancer cells and promotes angiogenesis.[10] In the present study, we found that the Twist was increased in the breast carcinoma and in cell lines with higher ability of migration. The elevated expression of Twist was positively associated with the poor prognosis, atypia of tumor cells, and increased ability of invasion and metastasis. In line with these studies, our results suggested that Twist might serve as a promoting factor in the regulation of invasiveness and metastasis of breast cancer. In addition, during the tumor metastasis process, tumor cell needs to reduce the E-cadherin expression, which is essential to disrupt the adherence junction complex and to facilitate the invasion into the distance tissues. Numerous studies have demonstrated that loss the expression of E-cadherin has a negative association with the invasiveness and metastasis of tumor cells.[11] In our current study, with the increasing expression of Twist, the mRNA and protein expression of E-cadherin became decreased. The expression levels of E-cadherin have a significantly negatively related with the tumors' invasion and metastasis. In mammals, ER-mediated genomic or nongenomic signal transduction is essential for the breast cells proliferation and apoptosis. The dysregulation of ER pathway results in breast cancers. Then, a number of studies have shown the molecular mechanism of ER in the development of breast cancer. However, the association of Twist, E-cadherin, and ER with the invasion and metastasis of breast cancer was not elucidated yet. In the present study, we showed that the expression of ER was positively associated with the expression of E-cadherin whereas negatively associated with the expression of Twist as well as invasion and metastasis ability in breast cancer cell lines and tissues. These results suggested that in addition to in the process of breast cancer, ER might play a role in the regulation of invasiveness and metastasis of breast cancer, possibly associated with the aberrant expression of Twist and E-cadherin. Further studies are needed to clarify this association more clearly. On the other hand, after overexpressing Twist in HEK-293 cells, we found downregulated expression of AKT and phosphorylated AKT, which were also considered to actively engage with the migratory process in motile cells, including metastatic cancer cells.[21] These may suggest that Twist is a trigger to promote invasion and metastasis in breast cancer.

Limitations of the study are as follows. (1) In the clinical tissues analysis, the sample size was relatively small. Further studies are needed to evaluate these results and clarify the relationship of Twist with the high invasive and metastatic breast cancer using more samples. (2) More detailed molecular mechanisms need to be revealed in the further study.


 > Conclusions Top


The present study suggested that aberrant elevated expression of Twist in the breast cancer cell lines and tissues as well as decreased expression of E-cadherin and ER may take part in the invasion and metastasis of breast cancer cells that demonstrated a clearer connection between Twist, E-cadherin, and ER in breast cancer than previous studies. Our study provided a novel insight on the relationship between expression of Twist, E-cadherin, and ER in breast cancer cells and tissues, which may help develop new and improved treatments to reduce invasion and metastasis of breast cancer.

Financial support and sponsorship

This research was supported by grants from the Science and Technology Support Program of Sichuan Province (2013JY0076 and 14ZC0024), the Education Department Program of Sichuan Province (15TD0020 and 12ZB064).

Conflicts of interest

There are no conflicts of interest.

 
 > References Top

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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]
 
 
    Tables

  [Table 1], [Table 2], [Table 3]



 

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