Low microvascular density at the tumor center is related to the expression of metalloproteases and their inhibitors and with the occurrence of distant metastasis in breast carcinomas.

BACKGROUND: The aims of this study were to evaluate the microvascular density (MVD) at the center of breast carcinomas, its relationship with the expression of metalloproteases (MMPs) and their inhibitors (TIMPs), and its connection with the distant metastasis rate. METHODS: An immunohistochemical study of four MMPs and two TIMPs was performed on cancer specimens from 97 women with a histological confirmed diagnosis of early invasive breast cancer. RESULTS: Expressions of MMP-9 by cancerous cells, or MMP-11 and TIMP-2 by stromal cells, were all negative and significantly associated with MVD, whereas MMP-7 score values were positive and also significantly associated with MVD. However, positive expression of MMP-1 by mononuclear inflammatory cells was significantly associated with MVD. Multivariate analysis demonstrated a significant and inverse relationship between MVD and the occurrence of distant metastasis. CONCLUSIONS: Our data point out the clinical importance of low MVD at the tumor center as an independent prognostic factor of distant metastasis development in breast cancer.

Impact of CD68/(CD3+CD20) ratio at the invasive front of primary tumors on distant metastasis development in breast cancer.

Tumors are infiltrated by macrophages, T and B-lymphocytes, which may favor tumor development by promoting angiogenesis, growth and invasion. The aim of this study was to investigate the clinical relevance of the relative amount of macrophages (CD68⁺), T-cells (CD3⁺ and B-cells (CD20⁺) at the invasive front of breast carcinomas, and the expression of matrix metalloproteases (MMPs) and their inhibitors (TIMPs) either at the invasive front or at the tumor center. We performed an immunohistochemical study counting CD3, CD20 and CD68 positive cells at the invasive front, in 102 breast carcinomas. Also, tissue sections were stained with MMP-2, -9, -11, -14 and TIMP-2 antibodies, and immunoreactivity location, percentage of reactive area and intensity were determined at the invasive front and at the tumor center. The results showed that an increased CD68 count and CD68/(CD3+CD20) ratio were directly associated with both MMP-11 and TIMP-2 expression by mononuclear inflammatory cells at the tumor center (p = 0.041 and p = 0.025 for CD68 count and p = 0.001 and p = 0.045 for ratio, respectively for MMP-11 and TIMP-2). In addition, a high CD68/(CD3+CD20) ratio (>0.05) was directly associated with a higher probability of shortened relapse-free survival. Multivariate analysis revealed that CD68/(CD3+CD20) ratio was an independent factor associated with distant relapse-free survival (RR: 2.54, CI: (1.23-5.24), p<0.01). Therefore, CD68/(CD3+CD20) ratio at the invasive front could be used as an important prognostic marker.

Study of TLR3, TLR4 and TLR9 in breast carcinomas and their association with metastasis.

BACKGROUND: Toll-like receptors (TLRs) have garnered an extraordinary amount of interest in cancer research due to their role in tumor progression. By activating the production of several biological factors, TLRs induce type I interferons and other cytokines, which drive an inflammatory response and activate the adaptive immune system. The aim of this study was to investigate the expression and clinical relevance of TLR3, 4 and 9 in breast cancer. METHODS: The expression levels of TLR3, TLR4 and TLR9 were analyzed on tumors from 74 patients with breast cancer. The analysis was performed by immunohistochemistry. RESULTS: Samples of carcinomas with recurrence exhibited a significant increase in the mRNA levels of TLR3, TLR4 and TLR9. Tumors showed high expression of TLRs expression levels by cancer cells, especially TLR4 and 9. Nevertheless, a significant percentage of tumors also showed TLR4 expression by mononuclear inflammatory cells (21.6%) and TLR9 expression by fibroblast-like cells (57.5%). Tumors with high TLR3 expression by tumor cell or with high TLR4 expression by mononuclear inflammatory cells were significantly associated with higher probability of metastasis. However, tumours with high TLR9 expression by fibroblast-like cells were associated with low probability of metastasis. CONCLUSIONS: The expression levels of TLR3, TLR4 and TLR9 have clinical interest as indicators of tumor aggressiveness in breast cancer. TLRs may represent therapeutic targets in breast cancer.

Comparative analysis and clinical value of the expression of metalloproteases and their inhibitors by intratumour stromal mononuclear inflammatory cells and those at the invasive front of breast carcinomas.

AIMS: Matrix metalloproteases (MMPs) and their inhibitors (TIMPs) play an essential role in the degradation of stromal connective tissue and basement membrane components. The aim of this study was to determine whether the dynamic analysis of these components can help to predict tumour aggressiveness. METHODS AND RESULTS: An immunohistochemical study was performed using tissue arrays and specific antibodies against MMPs -1, -2, -7, -9, -11, -13 and -14 and TIMPs -1, -2 and -3. More than 5000 determinations on cancer specimens from 124 patients with invasive breast cancer were performed on the tumour centre core as well as on the invasive front. Immunostaining for MMPs/TIMPs on mononuclear inflammatory cells (MICs) was evaluated. To identify specific groups of tumours with distinct expression profiles, data obtained from both MICs populations were analysed by unsupervised hierarchical cluster analysis. When compared with MICs at the invasive front, intratumour MICs more frequently showed expression of MMP-7 and -1 and TIMP-3, but less frequently expression of MMP-9 and -11 and TIMP-2. CONCLUSIONS: Our data led us to consider the need of further studies in order to identify subsets of MICs and other protein elements of the microenvironment as attractive targets for new therapeutic strategies against cancer.

An adult myometrial pluripotential precursor that promotes healing of damaged muscular tissues.

The use of adult stem cells for tissue and organ regeneration constitutes a promising alternative therapy in many human diseases that are currently not treatable. We have isolated a new cell type from mouse adult uterine biopsies (murine adult myometrial precursors or mAMPs) by means of using a simple and non-invasive approach. These cells have been characterized by surface markers, being positive for CD31, CD34, CD44, CD117, Stro-1 and Sca-1. A similar cell population (hAMPs) was isolated from human biopsies. AMPs can differentiate in vitro into a number of mesodermal (smooth and skeletal muscle, osteoblasts and adipocytes) as well as epidermal lineages (all neural lineages). AMPs are unusual adult stem cells as they still express some embryonic antigens and remain undifferentiated through a high number of passages before entering senescence. Importantly, when injected into animal models of muscular disease, AMPs can regenerate new muscle fibers, and promote functional muscular recovery. Moreover, these cells can regenerate the uterine lining after wound healing, reconstructing the uterine muscular architecture. In addition, these cells can form new vessels both in vitro and in vivo. We believe that these cells have superior features to other known adult stem cells and, consequently, their use holds great promise for regenerative medicine, drug development and basic research.

Androgen receptor expresion in breast cancer: relationship with clinicopathological characteristics of the tumors, prognosis, and expression of metalloproteases and their inhibitors.

BACKGROUND: In the present study we analyze, in patients with breast cancer, the tumor expression of androgen receptors (AR), its relationship with clinicopathological characteristics and with the expression of several matrix metalloproteases (MMPs) and their inhibitors (TIMPs), as well as with prognosis. METHODS: An immunohistochemical study was performed using tissue microarrays and specific antibodies against AR, MMPs -1, -2, -7, -9, -11, -13, -14, and TIMPs -1, -2 and -3. More than 2,800 determinations on tumor specimens from 111 patients with primary invasive ductal carcinoma of the breast (52 with axillary lymph node metastases and 59 without them) and controls were performed. Staining results were categorized using a score based on the intensity of the staining and a specific software program calculated the percentage of immunostained cells automatically. RESULTS: A total of 83 cases (74.8%) showed a positive immunostaining for AR, but with a wide variation in the staining score values. There were no significant associations between the total immunostaining scores for AR and any clinicopathological parameters. However, score values for MMP-1, -7 and -13, were significantly higher in AR-positive tumors than in AR-negative tumors. Likewise, when we considered the cellular type expressing each factor, we found that AR-positive tumors had a higher percentage of cases positive for MMP-1, -7, -11, and TIMP-2 in their malignant cells, as well as for MMP-1 in intratumoral fibroblasts. On the other hand, multivariate analysis demonstrated that patients with AR-positive tumors have a significant longer overall survival than those with AR-negative breast carcinomas (p = 0.03). CONCLUSION: Our results confirm that AR are commonly expressed in breast cancer, and are correlated with the expression of some MMPs and TIMP-2. Although we found a specific value of AR expression to be a prognostic indicator in breast cancer, the functional role of AR in these neoplasms is still unclear and further data are needed in order to clarify their biological signification in breast cancer.

Cytosolic levels of TFF1/pS2 in breast cancer: Their relationship with clinical-pathological parameters and their prognostic significance.

BACKGROUND: The Trefoil Factor 1 (TFF1/pS2), a peptide consisting of 60 amino acids, is the most abundant estrogen-induced messenger RNA present in MCF-7 breast cancer cells. The objective of this work was to evaluate the cytosolic TFF1 content in breast carcinomas, its possible relationship with different clinical-pathological parameters, and its potential prognostic significance and predictive value. METHODS: Cytosolic TFF1 levels were examined by immunoradiometric assay in 1031 patients with invasive breast cancer. The median follow-up period was of 50 months. RESULTS: There was a wide variability of cytosolic TFF1 levels in tumors (0.9-743.2 ng/mg protein). Statistical analysis showed that TFF1 levels were significantly higher in premenopausal patients (p = 0.001), as well as in tumors showing any of the following characteristics: good differentiation (p = 0.0001), ER and PgR positivity (p = 0.0001 and p = 0.001, respectively), diploidy (p = 0.045) and a high S-phase fraction (p = 0.001). In addition, the presence of high intratumoral TFF1 levels (cut-off: 2 ng/mg protein) was independently associated with a shorter overall survival in the group of patients as a whole (p = 0.001) as well as in the subgroup with node-negative breast cancer (p = 0.0004). Likewise, high intratumoral TFF1 levels were associated with a more prolonged overall survival in patients who received adjuvant tamoxifen (p = 0.004). CONCLUSIONS: In breast cancer patients, intratumoral TFF1 levels are associated with a better clinical outcome, especially in those with node-negative tumors. In addition, TFF1 levels have a low but significant predictive value in regards to response to adjuvant therapy with tamoxifen.

Expression of two androgen-induced proteins (pepsinogen C and apolipoprotein d) in epithelial skin cancers of the eyelids.

By immunohistochemistry, we investigated pepsinogen C (pepC) and apolipoprotein D (apoD) expressions in 23 basal cell (BCC) and 25 squamous cell carcinomas (SCC) of the eyelids and analyzed the possible relationship to clinicopathological parameters. A total of 3 (13%) BCC and 7 (28%) SCC stained positively for pepC, whereas 11 (47.8%) BCC and 6 (24%) SCC stained positively for apoD. No significant correlation was found between pepC and apoD expressions and patients or tumor characteristics (p > 0.05). pepC and apoD, two androgen-inducible proteins, may be expressed by BCC and SCC of the eyelids. Further studies will be necessary to determine their clinical significance.

Expression and prognostic significance of lysozyme in male breast cancer.

BACKGROUND: Lysozyme, one of the major protein components of human milk that is also synthesized by a significant percentage of breast carcinomas, is associated with lesions that have a favorable outcome in female breast cancer. Here we evaluate the expression and prognostic value of lysozyme in male breast cancer (MBC). METHODS: Lysozyme expression was examined by immunohistochemical methods in a series of 60 MBC tissue sections and in 15 patients with gynecomastia. Staining was quantified using the HSCORE (histological score) system, which considers both the intensity and the percentage of cells staining at each intensity. Prognostic value of lysozyme was retrospectively evaluated by multivariate analysis taking into account conventional prognostic factors. RESULTS: Lysozyme immunostaining was negative in all cases of gynecomastia. A total of 27 of 60 MBC sections (45%) stained positively for this protein, but there were clear differences among them with regard to the intensity and percentage of stained cells. Statistical analysis showed that lysozyme HSCORE values in relation to age, tumor size, nodal status, histological grade, estrogen receptor status, metastasis and histological type did not increase the statistical significance. Univariate analysis confirmed that both nodal involvement and lysozyme values were significant predictors of short-term relapse-free survival. Multivariate analysis, according to Cox's regression model, also showed that nodal status and lysozyme levels were significant independent indicators of short-term relapse-free survival. CONCLUSION: Tumor expression of lysozyme is associated with lesions that have an unfavorable outcome in male breast cancer. This milk protein may be a new prognostic factor in patients with breast cancer.

Vulval squamous cell carcinoma arising in localized Darier's disease.

Darier's disease (keratosis follicularis) is a rare vulval lesion and it has been associated with squamous and non-squamous tumors from different origin. A vulval squamous cell carcinoma (SCC) arising in a localized Darier's disease is presented. To our knowledge, this is the first case to report the association of a vulval SCC and a localized Darier's disease. The possibility of delaying or overlooking the diagnosis of a vulval carcinoma is also illustrated. It is suggested to resect the whole lesion of Darier's disease localized to the vulva or take multiple biopsies.