Modulation of oestrone sulphate formation and hydrolysis in breast cancer cells by breast cyst fluid from British and Hungarian women

Women with gross cystic breast disease may have an increased risk of breast cancer. In this study the ability of breast cyst fluid (BCF), obtained from British or Hungarian women, to modulate oestrone sulphate (E1S) formation or hydrolysis, has been examined. For this, oestrogen receptor-positive (ER+) MCF-7 or MDA-MB-231 (ER-) breast cancer cells were employed. The formation and hydrolysis of E1S was measured using radiometric techniques. BCF from British and Hungarian women mainly inhibited E1S hydrolysis in MCF-7 cells while stimulating hydrolysis in MDA-MB-231 cells. The extent of inhibition or stimulation of E1S hydrolysis in these cells was related to the Na+/K+ ratio of the BCF. There was a significant inverse relationship between the extent to which BCF samples inhibited hydrolysis in MCF-7 cells and stimulated it in MDA-MB-231 cells. BCF stimulated E1S formation in MCF-7 cells while inhibiting formation in MDA-MB-231 cells. No difference in the ability of BCF from British or Hungarian women to inhibit or stimulate E1S hydrolysis was detected in ER+ or ER- breast cancer cells. In contrast, BCF from British women stimulated E1S formation in ER+ cells (median 82%) to a significantly greater extent (P < 0.01) than BCF from Hungarian women (median 33%). The role that E1S has in breast cancer development remains unclear. The greater stimulation of E1S formation by BCF from British women, who have a higher risk of breast cancer than Hungarian women, suggests that it may act as a storage form of oestrogen within cells that can be activated by oestrone sulphatase.     

Effect of retinoic acid and palm oil carotenoids on oestrone sulphatase and oestradiol-17beta hydroxysteroid dehydrogenase activities in MCF-7 and MDA-MB-231 breast cancer cell lines

Oestrogen is important in the development of breast cancer. Oestrogen receptor positive breast cancers are associated with a better prognosis than oestrogen-receptor negative breast cancers since they are more responsive to hormonal treatment. Oestrone sulphate acts as a huge reservoir for oestrogens in the breast. It is converted to the potent oestrogen, oestradiol (E(2)) by the enzymes oestrone sulphatase and oestradiol-17beta hydroxysteroid dehydrogenase (E(2)DH). Retinoic acid and carotenoids have been shown to have chemopreventive activity against some cancers. The aim of our study was to determine and compare the effects of retinoic acid and palm oil carotenoids on growth of and oestrone sulphatase and E(2)DH activities in the oestrogen receptor positive, MCF-7 and oestrogen receptor negative, MDA-MB-231 breast cancer cell lines. Retinoic acid and carotenoids inhibited MCF-7 cell growth but had no effect on MDA-MB-231 cell growth. Both retinoic acid and carotenoids stimulated oestrone sulphatase activity in the MCF-7 cell line. E(1) to E(2) conversion was inhibited by 10(-7) M carotenoids but was stimulated at 10(-6) M in the MCF-7 cell line. Retinoic acid had no effect on E(1) to E(2) conversion at 10(-7) M but stimulated E(1) to E(2) conversion at 10(-6) M. Retinoic acid and carotenoids had no effect on E(2) to E(1) conversion in the MCF-7 cell line. Retinoic acid stimulated E(1) to E(2) conversion in the MDA-MB-231 cell line but had no effect on oestrone sulphatase activity or E(2) to E(1) conversion in this cell line. Both oestrone sulphatase and E(2)DH activity were not affected by carotenoids in the MDA-MB-231 cell line. In conclusion, retinoic acid and carotenoids may prevent the development of hormone-dependent breast cancers since they inhibit the growth of the MCF-7 cell line.     

Effect of transforming growth factor-beta1 on oestrogen metabolism in MCF-7 and MDA-MB-231 breast cancer cell lines.

Transforming growth factor-beta (TGF-beta) has been shown to inhibit the growth of mammary epithelial cells and may play a protective role in mammary carcinogenesis. In contrast, oestrogens promote the development of breast cancer. Oestrone sulphate (E1S) is a huge reservoir of active oestrogens in the breast being converted to the weak oestrogen, oestrone (E1), by oestrone sulphatase. E1 is reversibly converted by oestradiol-17beta hydroxysteroid dehydrogenase to the potent oestrogen, oestradiol (E2). The aim of this study was to assess the effect of the TGF-beta1 isoform on growth and oestrogen metabolism in the hormone-dependent MCF-7 and hormone-independent MDA-MB-231 human breast cancer cell lines. The results showed that TGF-beta1 significantly inhibited cell growth and stimulated the conversion of E1S to E1 and E1 to E2 in the MCF-7 cell line. In the MDA-MB-231 cell line TGF-beta1 significantly stimulated cell growth and inhibited the interconversions between E1 and E2. In conclusion, the growth inhibitory effect of TGF-beta1 on the MCF-7 cell line would appear to confer a protective effect in breast cancer. However, its ability to increase the amount of E2 would increase the risk of breast cancer. Which of these effects predominates in vivo remains to be explored. The growth stimulatory effect of TGF-beta1 on the MDA-MB-231 cell line probably acts through a mechanism independent of the effect of TGF-beta1 on oestrogen concentrations since this cell line is hormone unresponsive.     

Changes in Radiation Sensitivity and Steroid Receptor Content Induced by Hormonal Agents and Ionizing Radiation in Breast Cancer Cells in Vitro

Possible influences of tamoxifen and estradiol on in vitro radiation sensitivity and cellular receptor content after irradiation and/or tamoxifen treatment were studied in breast cancer cell lines; estrogen receptor (ER) and progesterone receptor (PgR) positive cell lines MCF-7 and MCF-7/TAM^R-1 and the ER and PgR negative cell line MDA-MB-231. The tamoxifen resistant MCF-7/TAM^R-1 cells were more resistant to ionizing radiation than the MCF-7 and MDA-MB-231 cells. Exposure to tamoxifen made the MCF-7 cells more radiation resistant, while estradiol made the MDA-MB-231 cells more radiation sensitive. A radiation dose of 6 Gy reduced the ER content in cytosol in both MCF-7 and MCF-7/TAM^R-1 cells, but brought no alterations to the PgR content. In MCF-7/TAM^R-1 cells tamoxifen exposure significantly increased the ER and reduced the PgR content, an effect not observed in the MCF-7 cells. To conclude, the present study indicates that irradiation and tamoxifen may modify the ER and PgR content in cytosol in breast cancer cells. Hormonal treatment may alter the radiation sensitivity, even in ER negative cells, suggesting that hormonal agents may act both via receptor and non-receptor binding mechanisms.     

Modulation of the insulin-like growth factor-I system by N-(4-hydroxyphenyl)-retinamide in human breast cancer cell lines.

The potent mitogenic activity of insulin-like growth factor I (IGF-I) on breast epithelium is inhibited by retinoic acid in oestrogen receptor-positive (ER+) breast cancer cell lines. We studied and compared the effects of N-(4-hydroxyphenyl)-retinamide (4-HPR) in terms of growth inhibition and modulation of the IGF-I system in ER+ (MCF-7) and oestrogen receptor-negative (ER-) (MDA-MB231) breast cancer cell lines. Treatment with 1-10 microM 4-HPR for up to 96 h induced a dose- and time-dependent inhibition of proliferation in both breast cancer cell lines. Induction of apoptosis was much more evident in MCF-7 than in MDA-MB231 cells (30-40% compared with 0-5% respectively at 5 microM for 48 h). Exogenous human recombinant IGF-I (hr-IGF-I)-stimulated cell proliferation was abolished by 1 microM 4-HPR in MCF-7 cells. Immunoreactive IGF-I-like protein concentration in conditioned medium was reduced by 38% in MCF-7 and by 90% in MDA-MB231 cell lines following treatment for 48 h with 5 microM 4-HPR. Western ligand blot analysis showed a reduction of IGF-binding protein 4 (BP4) and BP5 by 67% and 87%, respectively, in MCF-7, whereas IGF-BP4 and -BP1 were reduced by approximately 20% in MDA-MB231 cells. Exposure to 5 microM 4-HPR for 48 h inhibited [125I]IGF-I binding and Scatchard analysis revealed a decrease of more than 50% in maximum binding capacity (Bmax) and a reduced receptor number/cell in both cancer cell lines. Steady-state type I IGF-receptor mRNA levels were reduced by approximately 30% in both tumour cell lines. We conclude that 4-HPR induces a significant down-regulation of the IGF-I system in both ER+ (MCF-7) and ER- (MDA-MB231) breast cancer cell lines. These findings suggest that, in our model, interference with the ER signalling pathway is not the only mechanism of breast cancer growth inhibition by 4-HPR.     

Comparative analysis of xanafide cytotoxicity in breast cancer cell lines

Xanafide, a DNA-intercalating agent and topoisomerase II inhibitor, has previously demonstrated comparable cytotoxicity to the parent drug amonafide (NSC 308847). The current study was conducted to investigate further the anti-proliferative effects of xanafide in human breast cancer cell lines, in vitro and in vivo. The in vitro activity of xanafide against MCF-7, MDA-MB-231, SKBR-3 and T47D cell lines was compared to that of paclitaxel, docetaxel, gemcitabine, vinorelbine and doxorubicin. In MCF-7, xanafide demonstrated comparable total growth inhibition (TGI) concentrations to the taxanes and lower TGI values than gemcitabine, vinorelbine and doxorubicin. MCF-7 (oestrogen receptor (ER)+/p53 wild-type) was the most sensitive cell line to xanafide. MDA-MB-231 and SKBR-3 exhibited similar sensitivity to xanafide. T47 D (ER+/p53 mutated), showed no response to this agent. The in vivo activity of xanafide was further compared to that of docetaxel in MCF-7 and MDA-MB-231 cell lines using the hollow fibre assay. Xanafide was slightly more potent than docetaxel, at its highest dose in MCF-7 cell line, whereas docetaxel was more effective than xanafide in MDA-MB-231 cell line. Our results show that there is no relationship between sensitivity of these cell lines to xanafide and cellular levels of both isoforms of topoisomerase II and suggest that ER and p53 status and their crosstalk may predict the responsiveness or resistance of breast cancer patients to xanafide.     

Differential expression of CYP1A1 and CYP1B1 in human breast epithelial cells and breast tumor cells

Human cytochromes P450 1A1 (CYP1A1) and P450 1B1 (CYP1B1) catalyze the metabolic activation of a number of procarcinogens and the hydroxylation of 17beta-estradiol (E2) at the C-2 and C-4 positions, respectively. The aromatic hydrocarbon receptor (AhR) agonist 2,3,7,8- tetrachlorodibenzo-p-dioxin (TCDD) has a marked effect on estrogen metabolism in MCF-7 breast-tumor cells by induction of these two enzymes. To investigate whether induction of CYP1A1 and CYP1B1 by AhR agonists and the associated increase in E2 metabolism are common to all breast epithelial cells and breast-tumor cells, we determined the effects of TCDD on E2 metabolism, and CYP1A1 and CYP1B1 mRNA levels in a series of non-tumor-derived breast epithelial (184A1 and MCF-10A) and breast-tumor (MCF-7, T-47D, ZR-75-1, BT-20, MDA-MB-157, MDA-MB-231 and MDA-MB-436) cell lines. In 184A1 cells, which did not express detectable estrogen receptor (ER) alpha mRNA, CYP1A1 mRNA and activity were induced by TCDD, and enhanced E2 metabolism in TCDD-treated cells was predominantly E2 2-hydroxylation. In MCF-10A, MCF-7, T-47D, ZR-75-1 and BT-20 cells, which expressed varying levels of ER alpha mRNA, both CYP1A1 and CYP1B1 mRNA levels and rates of both E2 2- and 4- hydroxylation were highly elevated following exposure to TCDD. In MDA- MB-157, MDA-MB-231 and MDA-MB-436 cells, which did not express detectable ER alpha mRNA and generally displayed fibroblastic or mesenchymal rather than epithelial morphology, CYP1B1 induction was favored, and the rate of E2 4-hydroxylation exceeded that of 2- hydroxylation in TCDD-treated cells. These results show that breast epithelial cells and tumor cells vary widely with regard to AhR- mediated CYP1A1 and CYP1B1 induction, suggesting that factors in addition to the AhR regulate CYP1A1 and CYP1B1 gene expression. In these cell lines, significant CYP1A1 inducibility was restricted to cultures displaying epithelial morphology, whereas CYP1B1 inducibility was observed in cells of both epithelial and mesenchymal morphology.      

Growth inhibition and differentiation of human breast cancer cells by the PAFR antagonist WEB-2086

WEB-2086 -- an antagonist of platelet-activating factor receptor (PAFR) with known anti-inflammatory, antiangiogenic and antileukaemic properties -- also proved to inhibit the proliferation in human solid tumour cell lines of different histology, and with much higher efficacy than in normal fibroblasts. A detailed analysis of WEB-2086 anticancer activity was then performed focusing on breast adenocarcinoma MCF-7 and MDA-MB-231 cells. WEB-2086-treated cells, either expressing (MCF-7) or unexpressing (MDA-MB-231) the oestrogen receptor (ER)alpha, underwent a dose-dependent growth arrest (IC(50)=0.65+/-0.09 and 0.41+/-0.07 mM, respectively) and accumulation in G(0)-G(1) phase. WEB-2086 also induced morphological and functional changes typical of mature mammary phenotype including (i) cell enlargement and massive neutral lipid deposition (best accomplished in MCF-7 cells); (ii) decrease in motility and active cathepsin D levels (mainly observed in highly invasive MDA-MB-231 cells). The expression of ERalpha was neither increased nor reactivated in treated MCF-7 or MDA-MB-231 cells, respectively. WEB-2086-induced differentiation in breast cancer cells involved the upregulation of PTEN, a key tumour suppressor protein opposing tumorigenesis, and was apparently independent of p53, PAFR, peripheral benzodiazepine receptor and ERalpha status. Overall, WEB-2086 can be proposed as an effective antiproliferative and differentiative agent with interesting translational opportunities to treat breast cancers in support to conventional chemotherapy.     

Influence of 17β-Estradiol on 15-Deoxy-Δ12,14 Prostaglandin J2 -Induced Apoptosis in MCF-7 and MDA-MB-231 Cells

The nuclear receptor, peroxisome proliferator-activated receptor gamma (PPARγ), is expressed in variouscancer cells including breast, prostate, colorectal and cervical examples. An endogenous ligand of PPARγ,15-deoxy-Δ12,14 prostaglandin J2 (PGJ2), is emerging as a potent anticancer agent but the exact mechanismhas not been fully elucidated, especially in breast cancer. The present study compared the anticancer effects ofPGJ2 on estrogen receptor alpha (ERα)-positive (MCF-7) and ERα-negative (MDA-MB-231) human breastcancer cells. Based on the reported signalling cross-talk between PPARγ and ERα, the effect of the ERα ligand,17β-estradiol (E2) on the anticancer activities of PGJ2 in both types of cells was also explored. Here we reportthat PGJ2 inhibited proliferation of both MCF-7 and MDA-MB-231 cells by inducing apoptotic cell death withactive involvement of mitochondria. The presence of E2 potentiated PGJ2-induced apoptosis in MCF-7, but notin MDA-MB-231 cells. The PPARγ antagonist, GW9662, failed to block PGJ2-induced activities but potentiatedits effects in MCF-7 cells, instead. Interestingly, GW9662 also proved capable of inducing apoptotic cell death.It can be concluded that E2 enhances PPARγ-independent anticancer effects of PGJ2 in the presence of itsreceptor.     

Inhibition of colon and breast carcinoma cell growth by interleukin-4.

Within human carcinomas, there is often an infiltration of lymphocytes and other cells of the immune system. A variety of cytokines are produced by such cells that could have a paracrine influence on the growth of tumor epithelium. The effect of one of these cytokines, interleukin-4 (IL-4), on human breast and colon cancer cell lines was therefore examined. IL-4 inhibited the growth of human colon (HT 29) and breast [MCF-7 wild type (MCF-7 WT), MCF-7 Adriamycin-resistant (MCF-7r), MDA-MB-231, and MDA-MB-468] carcinoma cells in culture. Competitive binding of 125I-IL-4 demonstrated the presence of 2000 high affinity IL-4-binding sites on HT 29 cells. The Kd for specific binding of 125I-IL-4 to HT 29 cells was 77 pM. Further studies were conducted on the estrogen-dependent MCF-7 WT and estrogen-independent MDA-MB-231 breast carcinoma lines. Concentrations of IL-4 of 10-100 nM were required to significantly inhibit growth of these carcinoma cell lines; e.g., with MCF-7 WT cells, half-maximal inhibition of growth occurred at 20 nM IL-4. Specific binding of 125I-IL-4 was detected to MCF-7 WT and MDA-MB-231 cells, but the low level of binding precluded Scatchard analysis. IL-4 inhibited 90% of the 17 beta-estradiol-stimulated growth of MCF-7 WT cells in a dose-dependent manner but without a change in estrogen receptor expression. Inhibition of growth by IL-4 was less in the absence of estrogens. Combined treatment with IL-4 and other known inhibitors of breast carcinoma cell growth [transforming growth factor-beta 1 (TGF-beta 1) and the antiestrogen tamoxifen] showed additive inhibition. The hormone-independent cell lines MCF-7r and MDA-MB-231 were additively inhibited by IL-4 and TGF-beta 1. This was not the case with MDA-MB-468 cells in which inhibition by IL-4 and TGF-beta 1 was of similar magnitude but no significantly greater effect was observed on combined treatment. No secretion of IL-4 was detected from these cell lines either basally or on treatment with TGF-beta 1 or tamoxifen, and we conclude that IL-4 is a nonautocrine inhibitor of breast carcinoma cell growth.     

Novel retinoic acid metabolism blocking agents have potent inhibitory activities on human breast cancer cells and tumour growth

Antitumour effects of retinoids are attributed to their influence on cell proliferation, differentiation, apoptosis and angiogenesis. In our effort to develop useful agents for breast cancer therapy, we evaluated the effects of four representative retinoic acid metabolism blocking agents (RAMBAs, VN/14-1, VN/50-1, VN/66-1 and VN/69-1) on growth inhibition of oestrogen receptor positive (ER +ve, MCF-7 and T-47D) and oestrogen receptor negative (ER -ve, MDA-MB-231) human breast cancer cells. Additionally, we investigated the biological effects/molecular mechanism(s) underlying their growth inhibitory properties as well as their antitumour efficacies against MCF-7 and MCF-7Ca tumour xenografts in nude mice. We also assessed the effect of combining VN/14-1 and all-trans-retinoic acid (ATRA) on MCF-7 tumour xenografts. The ER +ve cell lines were more sensitive (IC(50) values between 3.0 and 609 nM) to the RAMBAs than the ER -ve MDA-MB-231 cell line (IC(50)=5.6-24.0 microM). Retinoic acid metabolism blocking agents induced cell differentiation as determined by increased expression of cytokeratin 8/18 and oestrogen receptor-alpha (ER-alpha). Similar to ATRA, they also induced apoptosis via activation of caspase 9. Cell cycle analysis indicated that RAMBAs arrested cells in the G1 and G2/M phases and caused significant downregulation (>80%) of cyclin D1 protein. In vivo, the growth of MCF-7 mammary tumours was dose-dependently and significantly inhibited (92.6%, P<0.0005) by VN/14-1. The combination of VN/14-1 and ATRA also inhibited MCF-7 breast tumour growth in vivo (up to 120%) as compared with single agents (P<0.025). VN/14-1 was also very effective in preventing the formation of MCF-7Ca tumours and it significantly inhibited the growth of established MCF-7Ca tumours, being as effective as the clinically used aromatase inhibitors, anastrozole and letrozole. Decrease in cyclin D1 and upregulation of cytokeratins, Bad and Bax with VN/14-1 may be responsible for the efficacy of this compound in inhibiting breast cancer cell growth in vitro and in vivo. Our results suggest that our RAMBAs, especially VN/14-1 may be useful novel therapy for breast cancer.     

Semaphorin4C 在乳腺癌子宫内膜癌及前列腺癌中的表达及意义*

目的:研究轴突导向分子Semap horin4C（Sema4C）在乳腺癌、子宫内膜癌及前列腺癌组织和细胞水平的表达情况及其与上述三种恶性肿瘤转移的关系。方法:应用免疫组化技术检测45例乳腺癌、42例子宫内膜癌及49例前列腺癌组织中Sema4C的表达情况,分别采用蛋白免疫印迹法Western Blot及细胞免疫荧光方法检测Sema4C 在三种乳腺癌细胞（MDA-MB-435S、MDA-MB-231、MCF-7）、两种子宫内膜癌细胞株（AN3CA、HEC-1-B）和两种前列腺癌细胞株（PC- 3M-1E8、PC- 3M-2B4）的蛋白表达及其亚细胞定位情况。结果:乳腺癌和子宫内膜癌中,其分别在有淋巴结转移的乳腺癌组和子宫内膜癌组的原发灶中的表达高于无淋巴结转移组,差异有统计学意义（P<0.05）。 前列腺癌中,Sema4C 的阳性表达率随着Gleason评分增加而升高（P<0.05）。 Western blot在七株肿瘤细胞均检测到Sema4C 的表达,且提示Sema4C 的表达与细胞的转移潜能呈正相关。荧光显微镜下观察到Sema4C 主要定位在细胞膜和细胞浆。结论:Sema4C 在乳腺癌、子宫内膜癌和前列腺癌组织和细胞中表达具有普遍性,且与上述三种恶性肿瘤转移关系密切,有望成为上述三种恶性肿瘤治疗的一个新的分子靶点。      

The relationship between the insulin-like growth factor-1 system and the oestrogen metabolising enzymes in breast cancer tissue and its adjacent non-cancerous tissue

Aims Previous studies have shown that oestrogen and Insulin-like Growth Factor-1 (IGF-1) act synergistically and cross-stimulatory while the oestrogen receptor (ER) and␣IGF-1R downstream signalling pathways interact at many levels. We investigate the relationship between the ER, and IGF-1R and their ligands in a series of human breast cancer tissue and adjacent non-cancerous tissue (ANCT).Methods A series of 139 pairs of breast cancer tissue and ANCT were obtained and divided into ER positive and ER negative groups based on tumour ER alpha immunostaining. All samples were processed for real-time quantitative-PCR to measure IGF-1, IGF-1R, ER alpha, STS and Cyp-19 mRNA levels. In addition, ER positive MCF-7 and ER negative MDA-MB-231 cell lines were treated separately with IGF-1 and an IGF-1R inhibitor called Tyrphostin AG1024 to see the effects of stimulating and inhibiting the IGF-1R. MCF-7 cell line was also treated with 4-hydroxytamoxifen. The mRNA levels of IGF-1, IGF-1R, ER alpha, STS and Cyp-19 of treated cell lines were measured and compared to those of non-treated controls. Data generated was normalised to Cytokeratin-19 mRNA levels.Results IGF-1R expression was higher in tumour tissue compared to ANCT (P=0.038) while IGF-1 expression was marginally higher in ANCT compared to tumour tissue only in the ER positive samples (P=0.098). ER positive tumours had a higher expression of IGF-1 compared to ER negative tumours (P=0.001) while IGF-1R, STS and Cyp-19 expression were higher in ER negative tumours (P=0.000, 0.000 and 0.006 respectively). There was no difference in STS or Cyp-19 expression in tumours or ANCT. Using Spearman’s Correlation test, IGF-1 positively correlated with STS, Cyp-19 and ER alpha in ER positive and negative groups (Coefficient=+0.497, +0.662 and +0.651 respectively, P=0.000 in all). IGF-1R correlated with IGF-1, STS, Cyp-19 and ER alpha only in the ER negative tumours (Coefficient=+0.620, +0.394, +0.692 and +0.662 respectively, P=0.000, 0.012, 0.000 and 0.000 respectively). In cell lines, IGF-1 treatment led to an increase in the mean expression of IGF-1, IGF-1R, STS and Cyp-19 in both cell lines while ER alpha expression increased only in MCF-7. IGF-1R inhibition caused a decrease in expression of all five genes in MDA-MB-231 but not in the MCF-7 cell line. Treatment with 4-hydroxytamoxifen caused a decrease in expression of all five genes.Conclusions IGF-1R is over-expressed in malignant tissue. IGF-1 is expressed at higher levels in ER positive tumours probably as a result of oestrogen stimulation while IGF-1R expression is higher in ER negative samples as an adaptation to lower local IGF-1 levels. An IGF-1 paracrine relationship may exist between tumour and ANCT but for STS and Cyp-19, there may be an autocrine–paracrine relationship. The IGF-1 ligand-receptor system is an important regulator of oestrogen production while oestrogen may be involved in stimulating IGF-1 expression. The expression of oestrogen synthesising enzymes is higher in ER negative breast cancers which may be due to the lack of oestrogen negative feedback or contribution from the overexpression of IGF-1R.     

Effects of adiponectin on breast cancer cell growth and signaling

Obesity is a risk factor for postmenopausal breast cancer. Adiponectin/Acrp30 is lower in obese individuals and may be negatively regulating breast cancer growth. Here we determined that five breast cancer cell lines, MDA-MB-231, MDA-MB-361, MCF-7, T47D, and SK-BR-3, expressed one or both of the Acrp30 receptors. In addition, we found that the addition of Acrp30 to MCF-7, T47D, and SK-BR-3 cell lines inhibited growth. Oestrogen receptor (ER) positive MCF-7 and T47D cells were inhibited at lower Acrp30 concentrations than ER-negative SK-BR-3 cells. Growth inhibition may be related to apoptosis since PARP cleavage was increased by Acrp30 in the ER-positive cell lines. To investigate the role of ER in the response of breast cancer cells to Acrp30, we established the MDA-ERalpha7 cell line by insertion of ER-alpha into ER-alpha-negative MDA-MB-231 cells. This line readily formed tumours in athymic mice and was responsive to oestradiol in vivo. In vitro, MDA-ERalpha7 cells were growth inhibited by globular Acrp30 while the parental cells were not. This inhibition appeared to be due to blockage of JNK2 signalling. These results provide information on how obesity may influence breast cancer cell proliferation and establish a new model to examine interactions between ER and Acrp30.     

Structure-dependent induction of aryl hydrocarbon hydroxylase in human breast cancer cell lines and characterization of the Ah receptor

The structure-dependent induction of aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O-deethylase by 2,3,7,8-tetrachlorodibenzo-p-dioxin, 2,3,7,8-tetrachlorodibenzofuran, and 1,2,4,7,8-pentachlorodibenzo-p-dioxin was determined in the MCF-7, T47-D, and MDA-MB-231 human breast cancer cell lines. Both the MCF-7 and T47-D cells were responsive to the induction effects of the halogenated aryl hydrocarbons and the structure-induction relationships were comparable to the reported structure-activity (induction, receptor binding, and toxicity) relationships observed in rodents and rodent cells in culture. The induction of ethoxyresorufin O-deethylase in the T47-D cells was the most sensitive aryl hydrocarbon (Ah) receptor-mediated response in both cell lines and this enzyme activity was more inducible than aryl hydrocarbon hydroxylase. In contrast, the three congeners were inactive as monooxygenase enzyme inducers in the MDA-MB-231 cells. Despite the differential Ah responsiveness of the cell lines, incubation of the cells with [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin followed by extraction of the nuclei with high salt and velocity sedimentation analysis of the extracts showed that specifically bound nuclear Ah receptor complexes were present in the three cell lines. The sedimentation coefficients (and levels) for the nuclear receptors were 6.6 S (32.1 fmol/mg protein/mg DNA), 6.9 S (61.6 fmol/mg protein/mg DNA), and 7.4 S (38.2 fmol/mg protein/mg DNA) in the T47-D, MCF-7, and MDA-MB-231 cell lines, respectively. Cytosolic receptor was also detected in the MCF-7 and MDA-MB-231 cells. Thus, despite the differences in Ah responsiveness of the T47-D and MDA-MB-231 cells, comparable levels of nuclear receptor were detected in both cell lines. Furthermore, the elution profiles of the nuclear receptors from DNA-Sepharose columns by using a salt gradient were similar and this suggested that defects in the DNA-binding activity of MDA-MB-231 nuclear receptor complexes were not major factors associated with their failure to respond to 2,3,7,8-tetrachlorodibenzo-p-dioxin and related compounds.     

PPAPDC1A在体内外对乳腺癌细胞增殖、侵袭和转移能力的影响

目的:通过构建稳定过表达和干扰PPAPDC1A的乳腺癌细胞株,探讨PPAPDC1A对乳腺癌细胞增殖、侵袭和转移能力的影响。方法:利用CCK-8和Transwell实验检测PPAPDC1A稳定过表达和干扰后对乳腺癌细胞体外增殖和侵袭能力的影响。采用裸鼠皮下成瘤实验检测PPAPDC1A对乳腺癌细胞体内增殖和裸鼠致瘤性的作用。利用免疫组织化学染色法检测各组肿瘤组织中Ki-67的表达。通过裸鼠尾静脉注射实验检测PPAPDC1A对乳腺癌MCF-7细胞和MDA-MB-231细胞体内转移能力的影响。结果:成功建立稳定过表达PPAPDC1A的乳腺癌MCF-7细胞株和稳定干扰PPAPDC1A的乳腺癌MDA-MB-231细胞株;CCK-8和Transwell实验结果显示,与MCF-7和MCF-7-Vector细胞株相比,MCF-7-PPAPDC1A细胞株的生长速度显著增快,穿膜细胞数量多（P＜0.05）;与此相反,MDA-MB-231-shPPAPDC1A组细胞的生长速度和穿膜细胞数明显少于MDA-MB-231-shNC和MDA-MB-231 细胞株（P＜0.05）。动物实验结果显示,与MCF-7-Vector组相比,MCF-7-PPAPDC1A组的肿瘤生长速度较快,肿瘤的体积较大,Ki-67的阳性率高,肺转移灶的数目增多（P＜0.05）;与此相反,与MDA-MB-231-shNC组相比MDA-MB-231-shPPAPDC1A组的肿瘤生长速度较慢,肿瘤的体积较小,Ki-67的阳性率低,肺转移灶的数目减少（P＜0.05）。结论:PPAPDC1A对乳腺癌细胞的增殖、侵袭和转移有促进作用。     

Prostate-specific antigen in breast cyst fluid: Possible role of prostate-specific antigen in hormone-dependent breast cancer

Prostate-specific antigen (PSA) is a 33 kDa serine protease which is produced by many different tissues in the body and has been shown to be present in low concentrations in breast milk and in about 30% of breast cancers. The presence of PSA in breast cancers is associated with the presence of steroid-hormone receptors and may be a favourable prognostic indicator. In this study, PSA immunoreactivity was measured in breast cyst fluid obtained from women with palpable breast cysts which is the most common benign breast disease. PSA was found to be present in very low concentrations in breast cyst fluid. In an attempt to understand the possible role of PSA in the breast, the effect of PSA on growth of the hormone-dependent MCF-7 and hormone-independent MDA-MB-231 human breast cancer cell lines was studied. In addition, the effect of PSA on oestrone sulphatase activity and oestrogen 17-oxidoreductase activity in these cell lines was investigated. PSA, in low concentrations, was found to inhibit MCF-7 cell growth and to stimulate the conversion of oestradiol to the less potent oestrogen oestrone in this cell line. PSA had no effect on the MDA-MB-231 cell line. Our findings suggest that PSA may act as a negative growth regulator in hormone-dependent breast cancers. © 1996 Wiley-Liss, Inc.