Biomedical subjects

A M Bamberger

Publications and source records attributed to A M Bamberger.

At least 19 recordsLinked to original sources

PubMed2005

Osteopontin expression in gestational trophoblastic diseases: correlation with expression of the adhesion molecule, CEACAM1.

The human placenta is a complex tissue with multiple endocrine and nutritional functions and a unique capacity for rapid proliferation but tightly controlled invasion, differentiating it from malignant tumors. Osteopontin (OPN) is a glycoprotein of the extracellular matrix, which has been shown to mediate cellular migration and invasion and to contribute to tumorigenesis in several types of cancers. OPN also could be implicated in regulating implantation and placentation by promoting cellular migration and invasion in a placenta-specific fashion. We could demonstrate the expression pattern of OPN in the normal human placenta in which it is localized in the extravillous (intermediate) trophoblast and the villous cytotrophoblast. CEACAM1 is an adhesion molecule, which we have recently found to be expressed at the maternal-fetal interface of the normal placenta with a localization to the extravillous (invasive) trophoblast and in gestational trophoblastic disease (GTD) and also to be potentially implicated in trophoblast invasion and tumorigenesis. Both OPN and CEACAM1 have been shown to interact with integrin beta3. The purpose of this study was to investigate the expression pattern of OPN in GTD and to correlate it with the expression of CEACAM1. To analyze the expression of OPN, we performed immunohistochemistry on a total of 27 cases of GTD, including 21 hydatidiform moles and 6 choriocarcinomas, which had previously been characterized with respect to their CEACAM1 expression. Hydatidiform moles showed a positivity for OPN in villous cytotrophoblast and in the trophoblast proliferations on the villous surface. The strongest OPN expression could be observed in the choriocarcinomas with a heterogenous OPN expression pattern. CEACAM1 had shown similar results and was found to be expressed in choriocarcinoma. The expression pattern of osteopontin in gestational trophoblastic diseases indicates that it might play a role in the pathogenesis of GTD (possibly as a functional complex with CEACAM1 and integrin beta3) and might be useful as an additional diagnostic marker for such lesions.

Antigens, CD↗

PubMed2001

Expression of Rb2/p130 in breast and endometrial cancer: correlations with hormone receptor status.

Rb2/p130 is a member of the retinoblastoma family of proteins, consisting of Rb, Rb2 and p107, which are important negative regulators of cell cycle progression and differentiation. While Rb2 downregulation was observed in several malignant tumours including endometrial cancer, the role of p130 in breast carcinomas is still unknown. We investigated Rb2 protein expression in tumour tissue from 68 mammary and 41 endometrial carcinomas, 4 mammary cell lines, and normal tissue samples. Therefore, we performed Western blot experiments for Rb2, Rb, and the oestrogen and progesterone receptors (ER, PR-A, PR-B). Weak or absent Rb2 expression was more often found in endometrial (59%) than in mammary carcinomas (24%). We found significant positive correlations of Rb2 expression with Rb, ER, and PR-B in breast cancer samples, and of Rb2 with Rb, PR-A, PR-B, and younger age in endometrial carcinomas. No significant associations with histological grading, stage, nodal involvement, or Ki67 staining were detected. Rb2 mRNA expression was studied by semi-quantitative RT-PCR in 56 endometrial or mammary tissue samples and correlated significantly with Western blot results. Our results indicate that loss of Rb2 expression, mostly by transcriptional down-regulation, may be associated with the development and dedifferentiation of most endometrial and a subset of mammary carcinomas.

Adult↗

PubMed2001

Expression pattern of the AP-1 family in endometrial cancer: correlations with cell cycle regulators.

PURPOSE: To study the expression pattern and the role of the AP-1 (activating protein-1) family of transcription factors in endometrial carcinogenesis. METHODS: We performed Western blot experiments with specific antibodies for each of the AP-1 proteins (c-jun, junB, junD, c-fos, fosB, fra-1, fra-2) with 41 endometrial carcinomas. Expression levels of the AP-1 factors were correlated with clinico-pathologic tumor parameters, steroid receptor status, Ki-67 expression and the expression levels of eight cell cycle regulatory proteins (cyclin D1, cyclin E, cyclin B1, cdk2, cdk4, p16, p21, and Rb). RESULTS: Of the seven AP-1 factors, three (c-fos, fra2, and junB) clearly showed higher expression levels in tumors when compared to matched normal endometrial samples. These factors also correlated significantly with cell cycle promoters (c-fos with cyclin E, cyclin B1, cdk2, and cdk4; fra-2 with cyclin B1; and junB with cyclin D1). Furthermore, high c-fos expression correlated with low ER and PR immunoreactivity and high grading (G3). On the other hand, correlations with classic cell cycle inhibitors (Rb, p16, p21) have also been observed for all AP-1 factors except c-jun and junD. CONCLUSIONS: Our results indicate that the AP-1 family of transcription factors is probably implicated in the regulation of cell cycle progression and control in endometrial carcinomas. In particular, c-fos might be an additional negative prognostic factor and/or implicated in tumor progression in endometrial cancer.

Blotting, Western↗

PubMed2001

Expression of cell-cycle regulatory proteins in endometrial carcinomas: correlations with hormone receptor status and clinicopathologic parameters.

PURPOSE: The normal human endometrium is characterized by hormone-dependent variations in the levels of cell-cycle regulatory proteins during the menstrual cycle. As this tightly controlled system is disturbed in endometrial carcinomas, we analyzed which cell-cycle regulators are involved in endometrial carcinogenesis. METHODS: We performed Western blot analysis of five cell-cycle stimulating (cyclins D1, E, B1, cdk2, cdk4) and three cell-cycle inhibiting (p16(INK4a), p21(WAF1), Rb) proteins in 41 endometrial carcinoma specimens. In addition, expression of the estrogen and progesterone receptors (ER, PR), Ki67, and, in selected cases, p16, cyclin E, and cyclin B1 was studied by immunohistochemistry. RESULTS: We found upregulation of all analyzed cell-cycle regulators in most tumors compared to normal endometrial tissue samples. Overexpression of cyclin E, cyclin B1, and p21 was associated with a less differentiated phenotype. In addition, high levels of cyclin E, cdk2, and cdk4 correlated with weak/absent ER expression, and p16 and p21 overexpression was significantly associated with low PR immunoreactivity. Cyclin B1 expression correlated with cyclin E, cdk2, cdk4, p21, Rb, and Ki67, and cyclin E expression with cyclin D1 and Rb. CONCLUSIONS: We conclude that cyclin E and cyclin B1 might be the major cell-cycle regulators involved in proliferation and reduced differentiation of endometrial carcinomas. In addition, p16, p21, and Rb appear to be uncoupled from their normal cell-cycle inhibiting function in many endometrial carcinomas.

Blotting, Western↗

PubMed2001

cis Interaction of the cell adhesion molecule CEACAM1 with integrin beta(3).

CEACAM1 is a cell adhesion molecule that has been implicated in a number of physiological processes (eg, tumor suppressor in epithelial tissues, potent angiogenic factor in microvessel formation, microbial receptor in human granulocytes and epithelial cells). The mechanism of CEACAM1 action is still largely unresolved but recent findings demonstrated that the cytoplasmic CEACAM1 domain is linked indirectly to the actin-based cytoskeleton. We have isolated integrin beta(3) as an associated protein using CEACAM1 tail affinity purification. This association depends on phosphorylation of Tyr-488 in the CEACAM1 cytoplasmic domain. Confocal laser scanning microscopy confirmed in vivo colocalization of both molecules in human granulocytes and epithelial cells. Furthermore, the concentrated colocalization at the tumor-stroma interface of invading melanoma masses suggests a functional role of CEACAM1-integrin beta(3) interaction in melanoma invasion. Moreover, colocalization of the two adhesion molecules is also found at the apical surface of glandular cells of pregnancy endometrium. Colocalization of CEACAM1 and integrin beta(3) at the transitional zone from proliferative to invasive extravillous trophoblast of the maternal-fetal interface supports a role for CEACAM1/integrin beta(3) complexes in cell invasion.

Antigens, CD↗

PubMed2001

Overexpression of the p16 cell cycle inhibitor in breast cancer is associated with a more malignant phenotype.

In order to study the role of the p16INK4A(MTS1/CDKN2a) tumor suppressor in breast cancer, we analyzed p16 protein expression in 60 breast cancer samples which were also analyzed for expression of Rb, Ki67, HER2/neu, and estrogen and progesterone receptors (ER, PR). P16 expression was investigated by two methods: western blotting (WB) followed by densitometry, and immunohistochemistry (IHC). The Rb status was studied by western blotting, and expression of Ki67, HER2/neu, ER, and PR was analyzed immunohistochemically. P16-negative results were found in 18% of the carcinomas by WB, but in only one case by IHC and were not associated with established prognostic parameters. In contrast, p16 overexpression which was detected by WB and IHC in 15% and 25% of the tumors, respectively, was significantly associated with unfavorable prognostic indicators. High p16 expression as detected by both methods correlated significantly with high grading and a negative estrogen receptor status. In addition, a significant association of p16 staining with inverse progesterone receptor status and high Ki67 expression was found with IHC. No correlation of p16 expression with clinical stage, HER2/neu immunostaining, Rb expression or Rb phosphorylation was found. Comparison of western blot results and immunohistochemistry suggests that both nuclear and cytoplasmic immunoreactivity in tumor cells is specific and due to p16 expression. We conclude that high p16 reactivity (both nuclear andcytoplasmic) is indicative of a more undifferentiated, malignant phenotype in mammary carcinomas.

Blotting, Western↗

PubMed2001

Expression pattern of the adhesion molecule CEACAM1 (C-CAM, CD66a, BGP) in gestational trophoblastic lesions.

CEACAM1 (CD66a, BGP, C-CAM) is an adhesion molecule of the carcinoembryonic antigen (CEA) family which has been shown to be normally expressed at the apical pole of epithelial cells and to show a dysregulated expression pattern in tumors derived from the latter. The purpose of the present study was to investigate the expression pattern of CEACAM1 in gestational trophoblastic lesions and to compare this expression with the one observed in the normal trophoblast. For this purpose, we performed immunohistochemistry using the 4D1/C2 monoclonal antibody which specifically recognizes CEACAM1 and does not interact with other members of the CEA family. Immunohistochemistry was performed on a total of 20 cases of gestational trophoblastic lesions including complete hydatidiform moles, one placental site trophoblastic nodule (PSN), one placental site trophoblastic tumor (PSTT), and three choriocarcinomas. Immunostaining for cytokeratin, hPL, hCG, and Ki-67 was also performed. Normal placental samples served as a control. CEACAM1 was absent from villous cyto- and syncytiotrophoblast in both normal placenta and hydatidiform molar samples. It was present in the benign extravillous trophoblast, with stronger expression in the proximal extravillous trophoblast of anchoring villi, but was also observed in interstitial and endovascular intermediate trophoblast and chorionic intermediate-like trophoblast. Partial expression was observed in the trophoblast proliferating from the surface of molar villi. In choriocarcinomas, areas of weak expression could be observed along with large areas without CEACAM1 expression. In the PSN and especially in the PSTT, CEACAM1 expression was stronger and more diffuse. The specific localization to extravillous trophoblast and its expression pattern in gestational trophoblastic lesions indicate that CEACAM1 can potentially be a helpful additional diagnostic marker in the differential diagnosis of such lesions.

Antigens, CD↗

PubMed2001

PTEN expression in breast and endometrial cancer: correlations with steroid hormone receptor status.

OBJECTIVE: The PTEN (MMAC1/TEP1) tumor suppressor gene is frequently mutated and homozygously deleted in human neoplasms, but there is only sparse information about PTEN protein expression in hormone-dependent female tumors. Therefore, we investigated PTEN expression in 68 breast and 43 endometrial carcinomas. METHODS: For PTEN protein detection, we used Western blot analysis followed by densitometry and compared these data with clinicopathologic parameters, the estrogen receptor (ER) and progesterone receptor (PR) status, HER2/neu and the proliferation marker Ki67. RESULTS: We were able to show significantly decreased PTEN protein expression in endometrial carcinomas compared with normal endometrial tissue samples, especially in the endometrioid histological subtype. In contrast, PTEN downregulation was found more rarely in breast cancer. Lower PTEN expression in breast cancer correlated significantly with high ER immunoreactivity (p = 0.008) and was weakly associated with PR expression (p = 0.055) and low histological grading (p = 0.081). No correlation with any of these parameters was observed in endometrial tumors. In both tumor types, no association of PTEN expression with any other analyzed parameter was found. CONCLUSIONS: These results suggest that PTEN expression plays different roles in the pathogenesis of endometrial carcinomas and breast cancer. In mammary carcinomas, loss of PTEN expression is mainly found in more differentiated tumors and is probably not a major event in carcinogenesis.

Adenocarcinoma↗

PubMed2001

The glucocorticoid receptor is specifically expressed in the stromal compartment of the human endometrium.

The human endometrium is a classical target tissue for steroid hormones. While the expression pattern and functional roles of both the estrogen receptor (ER) and the progesterone receptor (PR) are well defined, expression of the glucocorticoid receptor (GR) in this tissue has not been described so far. In the present study, we used immunohistochemistry to analyze the expression of GR in the normal human endometrium throughout the menstrual cycle. The expression of GR was compared to that of ER and PR, which were analyzed in parallel. We show that GR is expressed in the human endometrium with a pattern that markedly differs from the expression patterns of ER and PR. ER and PR are expressed in the nuclei of endometrial glands, whereas GR is completely absent from these structures. However, GR is strongly expressed in the stromal compartment of the endometrium throughout the cycle. Both stromal fibroblasts and lymphocytes are GR-positive. In addition GR expression is also observed in the endothelium of small endometrial blood vessels, which are ER- and PR-negative. Western blot analysis performed on endometrial tumor cell lines of glandular (HEC-1B) and mesodermal (SKUT-1B) origin, respectively, showed GR expression only in the latter. In summary, we demonstrate that GR is expressed in fibroblasts, lymphocytes and endothelial cells of the human endometrial stroma, while it is absent from the glandular compartment. The specific expression pattern of GR within the human endometrium points to a possible functional role of glucocorticoids in the process of decidualization which occurs primarily in the stromal compartment.

Adult↗

PubMed2001

Expression and tissue localization of soluble guanylyl cyclase in the human placenta using novel antibodies directed against the alpha(2) subunit.

The cytoplasmic or soluble forms of guanylyl cyclase (sGC) are heme-containing heterodimeric enzymes that are regulated by nitric oxide (NO) and carbon monoxide (CO). These gaseous messenger molecules are produced in the human placenta and are potential regulators of vasodilation and trophoblast invasion. The alpha(2)-subunit of sGC has only recently been shown to naturally occur in placental extracts. In the present study, two novel antibodies directed against different epitopes of the alpha(2) subunit, were generated. Western Blot analysis confirmed the presence of a 82 kDa protein, identical with alpha(2) protein overexpressed in Sf9 cells. According to RNase protection analysis the alternatively spliced alpha(2i) variant was absent from human placenta. Immunohistochemical analysis showed the presence of alpha(2) protein in syncytiotrophoblast and villous and umbilical blood vessels, which are known sites of NO production. Strong expression was observed in the extravillous (intermediate) trophoblast, where the expression of CO-generating hemeoxygenases has recently been documented. Localization of alpha(2) subunit expression suggests a role for sGC in mediating the actions of both NO and CO. The novel antibodies characterized in the present study will be powerful tools to further elucidate the role of the NO/CO/cGMP signaling pathways in pathologic states such as preeclampsia and intrauterine growth retardation.

Amino Acid Sequence↗

PubMed2000

Expression of cell cycle-regulatory proteins rb, p16/MTS1, p27/KIP1, p21/WAF1, cyclin D1 and cyclin E in breast cancer: correlations with expression of activating protein-1 family members.

The activating protein-1 (AP-1) complex is a mitogen-activated composite transcription factor that leads to activation of various target genes and enhanced proliferation of many cells after stimulation by TPA, EGF, serum, etc. The molecular mechanism of cell-cycle activation by AP-1 complexes remains unclear. Therefore, we studied protein expression of 6 cell cycle-regulatory proteins (Rb, p16, p21, p27, cyclin D1, and cyclin E) in protein extracts from 53 breast cancer samples and 4 mammary cell lines and correlated the data with expression of the 7 AP-1 family members (c-jun, junB, junD, c-fos, fosB, fra-1, and fra-2) as determined in a previous study. After Western blot analysis, we found significant associations between members of both groups: whereas c-jun was associated with Rb expression (p = 0.002), strong junD and c-fos expression correlated with high cyclin E reactivity (p = 0.017 and p = 0.013, respectively). Over-expression of fosB was found mainly in tumors with strong Rb (p = 0.013) and weak p16 (p = 0.004) expression. Fra-1 expression was significantly associated with p16 and cyclin E over-expression, whereas fra-2 results correlated with both cyclin D1 and cyclin E. These results point to direct or indirect activation of some cell cycle-regulatory proteins by AP-1 complexes. In addition, our data suggest differential regulation of cell cycle-stimulating and -inhibiting factors depending on the abundance of single AP-1 family members.

Blotting, Western↗

PubMed2000

Expression of leukemia inhibitory factor (LIF) and LIF receptor (LIF-R) in the human adrenal cortex: implications for steroidogenesis.

It is well established that steroidogenesis in the adrenal cortex is regulated by extraadrenal factors, such as ACTH and angiotensin II. However, over the last years, it has become increasingly clear that paracrine and autocrine mechanisms are also important for steroid synthesis in the adrenal gland. The current study was designed to analyze whether the pleiotropic cytokine leukemia inhibitory factor (LIF) and/or its receptor (LIF-R) are expressed in the normal human adrenal cortex, and whether they may play a role in regulating steroidogenesis. Using LIF- and LIF-R-specific primers, we show by RT-PCR that both mRNAs are expressed in this tissue, as well as in the NCI-H295 adrenal carcinoma cell line. The correct sequences of the PCR products were verified by restriction enzyme analysis and DNA sequencing. Immunohistochemistry, employing specific antibodies against LIF and LIF-R, reveals expression of both proteins in the normal human adrenal cortex. Finally, we show that LIF can significantly enhance basal and ACTH-induced production of cortisol and aldosterone in NCI-H295 cells. In summary, we show for the first time that LIF and its receptor are expressed in the normal human adrenal cortex. Our stimulation experiments indicate that the intraadrenal LIF/LIF-R system may participate in regulating adrenal steroidogenesis.

Adrenal Cortex↗

PubMed2000

The adhesion molecule CEACAM1 (CD66a, C-CAM, BGP) is specifically expressed by the extravillous intermediate trophoblast.

CEACAM1 (CD66a, C-CAM, BGP) is an adhesion molecule of the carcinoembryonic antigen family which has been shown to be normally expressed at the apical pole of epithelial cells, including the apical pole of endometrial surface and glandular epithelia. The purpose of the present study was to investigate its expression pattern at the maternal-fetal interface, and thus to determine whether CEACAM1 could be implicated in the human implantation process. For this purpose, we performed immunohistochemistry using the 4D1/C2 monoclonal antibody (mAb) as well as flow cytometry and Western blot on isolated trophoblast populations. On the maternal side of the maternal-fetal interface, CEACAM1 was present in epithelial cells of pregnancy endometrium as well as in small endometrial vessels, whereas it was absent from decidual cells. On the fetal side, CEACAM1 was strongly expressed by the extravillous (intermediate) trophoblast at the implantation site, as well as by extravillous trophoblast cells with invasive phenotype in primary culture, as shown by flow cytometry and Western blot. Expression was also observed in placental villous core vessels but was absent from both villous cyto- and syncytiotrophoblasts throughout the pregnancy. We conclude that, given its specific expression pattern, CEACAM1 can be a useful marker for extravillous intermediate trophoblast and might be functionally implicated in mediating trophoblast/endometrial and/or trophoblast/endothelial interactions during the trophoblastic invasion of the endometrium.

Antigens, CD↗

PubMed2000

Biphasic effect of medroxyprogesterone-acetate (MPA) treatment on proliferation and cyclin D1 gene transcription in T47D breast cancer cells.

While progesterone is a known differentiation-inducing factor in the human endometrium, for the breast epithelium both proliferation-inducing and -inhibiting effects have been described. Cyclin D1, which is required for cell cycle progression in G1 and has been shown to play an important role in the pathogenesis of breast cancer has been implicated as a possible mediator of such effects. In the present study we thus investigated the effects of the progestin agonist MPA (medroxy-progesterone acetate) on proliferation of T47D breast cancer cells. In parallel experiments, the regulation of the human cyclin D1 promoter as well as cyclin D1 protein levels under the influence of MPA were studied. Our results show an increase of proliferative activity in T47D cells after 24 and 48 h of MPA treatment followed by inhibition of proliferation after 72 h. In Western blot analysis an increased expression level of cyclin D1 protein can be observed after 24h of MPA stimulation, while at 72h the protein levels are barely detectable. Transient transfection experiments with a luciferase reporter plasmid containing the human cyclin D1 promoter showed an induction of the promoter after 24 and 36h of MPA treatment followed by a reduction in promoter activity. In conclusion, our results confirm the existence of a biphasic response of T47D cell proliferation in response to MPA treatment, consisting of stimulation of proliferation followed by inhibition, and further implicate cyclin D1 as a mediator of these effects, since the cyclin D1 promoter shows a similar biphasic response in this context.

Antineoplastic Agents, Hormonal↗

PubMed2000

The peripheral CRH/urocortin system.

The hypothalmus-pituitary-adrenal (HPA) axis and the immune system communicate at multiple levels: On the one hand, immune system-derived substances, such as interleukin-1, interleukin-6, tumor necrosis factor alpha, and leukemia inhibitory factor can stimulate the HPA axis. On the other hand, HPA axis-derived substances, most importantly glucocorticoids, can modulate the immune response. Furthermore, factors that were originally thought to be restricted to the HPA axis have been found to be expressed by immune cells. Proteins belonging to the CRH (corticotropin-releasing hormone) family represent important examples of such hormones. In the early 1990s, it was shown that immunoreactive CRH was present at sites of chemically induced inflammation. Administration of anti-CRH antibodies reduced the degree of inflammation, pointing to a pro-inflammatory role of "peripheral" CRH. We and others could show that lymphocytes are one source of immunoreactive CRH; however, the antiserum used in our study as well as in previous reports crossreacted with urocortin, a newly discovered member of the CRH family. Using RT-PCR, we could clearly demonstrate that human lymphocytes expressed urocortin but not CRH mRNA. These results were confirmed by immunocytochemistry, employing urocortin- and CRH-specific antibodies, respectively. The possible functional roles of urocortin expression in the immune system are discussed.

Animals↗

PubMed2000

Progesterone receptor isoforms, PR-B and PR-A, in breast cancer: correlations with clinicopathologic tumor parameters and expression of AP-1 factors.

In the present study, we used Western blot analysis to determine the expression of the progesterone receptor (PR) isoforms, PR-B and PR-A, in breast tumors (n = 53), and correlated the expression patterns of the two isoforms with the clinicopathological parameters of these tumors and with expression of the AP-1 family of transcription factors. Expression of the two PR isoforms correlated significantly with each other, indicating that the expression of the two isoforms is probably regulated in a correlated fashion. Expression of both isoforms correlated significantly with expression of the estrogen receptor (ER). Furthermore, expression of PR-B was found to correlate significantly with the absence of ErbB2/neu. For the AP-1 factors, Fra-1 expression showed an inverse correlation with PR-B expression. In contrast, expression of FosB correlated significantly with expression of both isoforms, and the association was stronger with PR-B expression. An analysis of the ratio of expression of the two isoforms showed that most of the tumors expressed PR-A levels which were equal or higher than the corresponding PR-B expression levels (together 94% of the analyzed tumors) indicating that, in mammary carcinomas, a predominance of the PR-A isoform over the PR-B isoform seems to be the case. While there was no statistically significant correlation with age, staging and histological type, expression of both isoforms correlated with a more differentiated phenotype (G1/G2 grading). However, this association was stronger for PR-B. Also, a PR-A < or = PR-B expression level was associated with G1/G2 grading, while a PR-A > PR-B expression level showed an association with a more undifferentiated phenotype (G3 grading). The expression level of the two PR isoforms might prove to be of prognostic and/or predictive value, especially since the two isoforms have been shown to be functionally different and to modulate the response of tumor cells to progestins and antiprogestins differently.

Blotting, Western↗

PubMed2000

Leukemia inhibitory factor (LIF) stimulates the human HLA-G promoter in JEG3 choriocarcinoma cells.

HLA-G is a non-classic class I MHC molecule specifically expressed by human invasive cytotrophoblast cells, which has been suggested to play a role in facilitating the immune tolerance of the conceptus. So far, very little is known about the regulation of the human HLA-G gene. The present study was, thus, designed to investigate the regulation of the human HLA-G promoter. JEG3 choriocarcinoma cells, which express HLA-G endogenously, were used as a model. A 890 bp fragment of the human HLA-G promoter was amplified by nested PCR from genomic DNA, cloned into pCR-Script and, after sequencing, subcloned into pGL3-Luc in front of the luciferase reporter gene. This vector was then used in transient transfection experiments in JEG3 cells. Parallel transfection experiments were performed using an alpha subunit (alphaSU)-Luc reporter plasmid as a control. Using this system, several potential modulating substances were tested in different concentrations and for different periods of time: phorbol ester (TPA), cAMP, IFNgamma, IL-1, and leukemia inhibitory factor (LIF), with only LIF administration resulting in induction of the HLA-G promoter. LIF treatment also resulted in induction of HLA-G mRNA. JEG3 cells are shown to possess LIF receptors. LIF is a pleiotropic cytokine produced at the maternal-fetal interface which has been shown to play an essential role in implantation in mice. LIF is produced in high amounts by the human endometrium and the trophoblast itself, and LIF receptors are present on cytotrophoblast cells. LIF could, thus, play a role in modulating HLA-G production and immune tolerance at the maternal-fetal interface.

Animals↗

PubMed1999

Expression pattern of the AP-1 family in breast cancer: association of fosB expression with a well-differentiated, receptor-positive tumor phenotype.

In the present study, the expression of members of the AP-1 family of transcription factors in breast tumors (n = 53) was investigated by Western blot with antibodies specific for each of the AP-1 family members (c-jun, junB, junD and c-fos, fosB, fra1 and fra2). The tumors were characterized with regard to grading, staging, histology, steroid-receptor-expression status and c-erbB2/neu expression. For comparison, normal breast-tissue samples, human breast-cancer cell lines (T47D and MDA-MB231) and the transformed human breast epithelial cell line HBL100 were also analyzed. For c-jun, junB, c-fos and fra2, a relatively uniform expression pattern without significant differences among tumors was observed. junD-protein amounts varied strongly in the tumor specimens. fosB-expression levels also varied strongly in the tumors, weak/absent expression being found in 47%, while 45% exhibited strong/very strong levels of expression. While none of the other AP-1 family members showed significant correlations with clinico-pathological tumor parameters or receptor status, expression of fosB was found to correlate significantly with positive steroid-hormone-receptor status (in the tumors and the cell lines) and a more differentiated tumor phenotype. Expression of 2 fra-1-specific bands of 33 and 36.5 kDa showed significant negative correlation with fosB expression, as well as with estrogen-receptor status and differentiation. We conclude that strong differences in the expression pattern of AP-1 family members are present in breast tumors, and that certain members of this family, such as fosB and fra-1, might be involved in the pathogenesis of these tumors.

Breast Neoplasms↗