Profiling of selected angiogenesis-related genes in proliferative eutopic endometrium of women with endometriosis.

OBJECTIVE: To compare the expression level of the most relevant angiogenesis-related genes in the eutopic endometrium of women with and without endometriosis. STUDY DESIGN: 32 regularly menstruating patients (18 with endometriosis and 14 controls) underwent surgery in the proliferative phase of the cycle. Eutopic endometrium was collected by the use of aspirating biopsy prior to laparoscopy. Only patients with advanced (stage III and IV) histopathologically confirmed ovarian endometriosis were studied. Real-time PCR gene arrays were applied to examine the expression of 84 human angiogenesis-connected genes. Western-blot and enzyme-linked immunosorbent assays (ELISA) were used to confirm the expression of selected proteins. RESULTS: We found significantly higher levels of AKT1 (p=0.003), TYMP (p=0.02), JAG1 (p=0.007), LAMA5 (p=0.005) and TIMP-1 (p=0.03) in eutopic endometrium of patients with endometriosis as compared with controls. By the use of Western blot we found clearly positive expression of AKT1 whereas ELISA assays confirmed expression of AKT1, TYMP, JAG1, LAMA5 and TIMP1. CONCLUSION: Changes in the expression of selected genes might lead to or be a consequence of an early defect in the physiological activity of proliferative endometrium ultimately resulting in its overgrowth outside the uterine cavity.

Increased serum level of membrane type 1-matrix metalloproteinase (MT1-MMP/MMP-14) in patients with breast cancer.

UNLABELLED: Different types of matrix metalloproteinases, including membrane type 1 matrix metalloproteinase (MT1-MMP/MMP-14) can be easily detected in biological fluids and therefore may be contemplated as putative tumor markers. Although increased activity of MT1-MMP/MMP-14 have already been found in breast cancer, little is known about its circulating levels. The aim of the present study was therefore to evaluate serum levels of active form of membrane type 1 matrix metalloproteinase (MT1-MMP/MMP-14). A novel type of activity enzyme-linked immunosorbent assay was used to detect serum levels of MT1-MMP/MMP-14 in 18 patients with invasive ductal breast cancer and 11 healthy controls. In the breast cancer group of patients MT1-MMP/MMP-14 mean (+/-SD) concentration was 16.91+/-5.87 ng/ml which was significantly higher (p<0.0001) than the mean values obtained for the control i.e. 8.55+/-1.66 ng/ml. CONCLUSIONS: Higher levels of soluble form of MT1-MMP/MMP-14 could play a role in invasiveness and metastasis of breast cancer. Whether or not it has a potential as biochemical marker remains to be determined.

Expression of glucose transporter GLUT-1 and estrogen receptors ER-alpha and ER-beta in human breast cancer.

We attempted to describe a GLUT-1 expression in breast cancer and characterize correlation between GLUT-1 and ERs alpha and beta expression as well as correlate this with clinicopathologic features. Sixty-nine patients were involved in the study. GLUT-1, ER-alpha and ER-beta immunocytochemistry was performed using the streptavidin- biotin method. Thirty-seven (53.6%) out of total 69 were GLUT-1 positive. Of GLUT- 1 positive 45.3% were ER-alpha-positive, whereas 81.3% of ER-alpha-negative were GLUT-1 positive. Statistically significant correlation was observed between GLUT-1 and ER-alpha expression status but neither between GLUT-1 and ER-beta nor with clinicopathologic features. No statistically significant correlation was found between expression level (expressed as immunocytoreactive score) of GLUT-1, ER-alpha and ER-beta. Since most of ER-alpha-negative (81.3%) were GLUT-1 positive and significant correlation exists between the two receptors it is reasonable to assume that some functional relation might exists between the expression of two receptors.

Fine needle aspiration cytology of benign breast disease. Markers of apoptosis and proliferation.

Aim of the study was to compare the fine needle aspiration cytology findings of benign breast lesions with incidence of proliferation markers and apoptosis. This study included 37 patients with palpable breast lumps, referred for USG guided FNA. FNAC were prospectively classified as C2-benign, C4-suspicious of malignancy, and C5-malignant. The specimens were simultaneously stained for Ki-67, MPM2, Bcl2 and P53. The diagnoses in group-C2 were following: simple cyst, multiple cysts, simple cyst with apocrine metaplasia, inflammatory cyst, benign dysplasia (BD) and benign solid tumors. The final diagnoses, after histopathological verification, in cases of primary classification as C4 and C5 were as follow: proliferative fibroadenoma (FAp) and breas cancer, respectively. Great majority of C2/BD aspirates were negative for proliferative antigens Ki-67 and PCNA. These antigens were detected in part of benign solid tumors, as anticipated in suspicious solid tumor, and in all of cancer aspirates. Bcl-2 immunopositive cells were detected approximately in one quarter of C2/BD, nearly in half of C2 solid tumors and in one C4/FAp. Most of diagnosed specimens were P53-negative. Immunocytodetection of Ki67, MPM2, Bcl2, P53 might be promising, supportive method in the classification of benign breast lesions. FNAC increases the reliability of diagnosis when complemented by immunocytochemical staining. It could be helpful procedure of establishing more accurately the biology of these lesions and possibly serve as an essential factor in clinical follow-up. Nevertheless, further study on larger group of patients comparing cytological and histopathological diagnosis is required to estimate reliability of its predictive value.

Uterine contractility signals--an introduction to wavelet analysis.

PURPOSE: Analysis of the uterine contractility in the nonpregnant states has provided information about physiological changes during menstrual cycle. There is need to develop methods of recording uterine activity as well as mathematical interpretation of recorded time series. Wavelets are a new powerful tool for signal and image processing. The aim of this study is an introductory view of Fourier (one of the fundamental methods of investigating of biomedical signals) and wavelet transforms applications in the analysis of uterine contractions. MATERIAL AND METHODS: Spontaneous uterine activity of healthy patient and patient with dysmenorrhea was recorded by micro-tip two sensors catheter (Millar Instruments, Inc. USA). After amplification analogue signals were converted to digital. Signals were analysed using Fourier and wavelet transforms. RESULTS: Contrary to the Fourier decomposition, which is global and provides the information integrated over the whole signal, the continuous and discrete wavelet transforms allow to extract local and global variations of the recorded contractions. From the analysis of the coefficients of the wavelet transform we can assess various pattern of propagation: normal propagation, simultaneous propagation and inverted propagation. CONCLUSIONS: This study is the introduction to the wavelet analysis of the uterine contraction signals. Wavelet transform provides insight into the structure of the time series at various scales. It allows to localise changes of the signal in time, providing additional information in comparison with the Fourier transform.

Estrogenic and antiestrogenic effects of raloxifene on collagen metabolism in breast cancer MCF-7 cells.

We compared the effects of different concentrations of raloxifene (1, 4 and 10 microM) on collagen biosynthesis, gelatinolytic and prolidase activities and matrix metalloproteinase (MMP) expression (MMP-2 and MMP-9) in estradiol-stimulated (2 nM) breast cancer MCF-7 cells. Raloxifene inhibited in a dose-dependent manner the proliferation of MCF-7 cells, independently of the presence or absence of estradiol in the growth medium. Raloxifene at concentrations of 1 microM and 4 microM inhibited collagen biosynthesis by about 10-fold and prolidase activity by about 50%, while at a concentration of 10 microM it inhibited these processes by only about 25%. This phenomenon was accompanied by differences in gelatinolytic activity and MMP (MMP-2 and MMP-9) expression as demonstrated by zymography and Western immunoblot analysis, respectively. In estrogen-stimulated MCF-7 cells, cultured in the presence of 1 microM raloxifene, a dramatic increase in the activity of both collagenases was found. In contrast, addition of raloxifene at a concentration of 10 microM to the medium of the cells resulted in restoration of gelatinolytic activity to that found in control cells. Similarly, but at both doses (1 and 10 microM), raloxifene was able to reduce MMP-2 expression in the cells. However, when used alone (without estradiol) a concentration of 1 microM raloxifene strongly stimulated MMP-2 expression, while at a concentration of 10 microM the effect was not observed. In the case of MMP-9, only trace amounts of this gelatinase were detected, although in contrast to MMP-2, an increase in its expression was noticed at a concentration of 10 microM raloxifene. The data raise the possibility that in estrogen-stimulated MCF-7 cells, raloxifene at low concentrations (1 and 4 microM) evokes antiestrogenic effect on collagen biosynthesis and prolidase activity on the one hand, and an estrogenic effect on gelatinolytic activity on the other, while at higher concentrations (about 10 microM) it evokes an estrogenic effect on collagen biosynthesis and prolidase activity, and an antiestrogenic effect on gelatinolytic activity. Our data suggest that the effects of raloxifene on collagen synthesis, prolidase and metalloproteinase activities in breast cancer may explain its role in the prevention of breast cancer development.

[Biochemical mechanism of raloxifen and tamoxifen action for the prevention of breast cancer. Studies in vitro]. / Mechanizmy biochemiczne dzialania tamoxifenu i raloxifenu w prewencji raka sutka. Badania in vitro.

Cancer growth follows fine balance disturbance between cell proliferation, differentiation and death. It has been shown that mutation of 4 to 5 genes controlling cellular proliferation events may be contributive to carcinogenesis. Estrogens play a central role in reproductive physiology. They are also a causative factors in the pathogenesis of neoplastic and non-neoplastic diseases, including breast cancer. The estrogen dependency of human breast cancer has been successfully exploited in the treatment of early and advanced diseases and provides a unique opportunity for chemoprevention of this common malignancy. The aim of present study was to examine the effects of tamoxifen and raloxifen on the induction of apoptosis and proliferative activity of human breast adenocarcinoma MCF-7 cells. It has been found that both tamoxifen and raloxifen decreased the speed of cell cycle in MCF-7 cells and acts as proapoptotic factors. It reduces viability of cancer cells and probability of neoplastic clone multiplification. This effect conducts to limitation of cancer expansion.

Influence of estrogen, antiestrogen and UV-light on the balance between proliferation and apoptosis in MCF-7 breast adenocarcinoma cells culture.

Studies of the mechanism of actions of estrogen, antiestrogen and physical factors may provide clues to an understanding of breast cancer growth and/or regression regulation and thus identify novel targets for therapeutic intervention. Defective control of apoptosis appears to play a central role in the pathogenesis of neoplasia. Conversely, cancer therapy and ionizing radiation can induce cancer cell death by apoptosis and/or necrosis. bcl-2 gene and p-53 gene products have been both linked to programmed cell death pathways. We have analyzed the effect of estradiol, tamoxifen and UV exposure on the induction of apoptosis, expression of p53 and bcl-2 gene products as well as the proliferative activity (expressed as [3H]thymidine incorporation and PCNA and MPM2 antigens involvement) in MCF7. It has been found that estradiol increases the speed of cell cycle in MCF7 and acts as antiapoptotic factor. Tamoxifen has multiple influence on the rate of growth of cancer cells: depends on estrogen receptor (ER), conducts reduction of proliferation rate; depends on ER and other mechanisms conducts to suppressions of Bcl-2 protein expression and induction of cell death through apoptotic pathway. Estradiol prevents the apoptotic influence of tamoxifen probably by enhancement of Bcl-2 protein expression and does not prevent the inhibition of proliferation rate. The irradiation with UV induces apoptosis by over-expression of p53 and down-regulation of bcl-2 gene.

Effect of exercise on glycogen metabolism in muscles of triiodothyronine-treated rats.

The aim of the present study was to examine the rate of glycogen mobilization during exercise and the rate of the postexercise glycogen replenishment in different muscle types [white (WG), and red (RG) gastrocnemius, soleus (S) and diaphragm (D)] in rats treated with triiodothyronine (T3, group T). Rats of the control group (C) were treated with saline. The animals were made to run on a treadmill set at 0 degree gradient and at a speed of 1200 m.h-1. The time taken to reach exhaustion in group C was 188 (SD 23) min, whereas in group T, it was only 63 (SD 12) min. The content of glycogen in all muscles of the rats from group T at rest and during exercise was significantly lower than in group C at each corresponding time. At exhaustion, the glycogen content was in WG(C) 34.79 (SD 4.65), (T) 20.10 (SD 4.10); in RG(C) 22.82 (SD 4.66), (T) 16.50 (SD 2.00); in S(C) 14.85 (SD 2.48), (T) 11.90 (SD 2.93); in D(C) 18.18 (SD 3.49), (T) 7.54 (SD 3.36) (mumol of glucosyl units. g-1). The amount of glycogen mobilized during exhausting exercise in RG, S and D was similar in both groups whereas in WG it was much higher in rats of group T than in group C. The concentration of glycogen returned to pre-exercise values in each muscle 3 h after exercise. The net amount of glycogen resynthesized during 3 h of recovery depended on the muscle type. It was in WG(C) 3.30, (T) 18.03; in RG(C) 21.34, (T) 25.88, in S(C) 34.00, (T) 17.68, and in D(C) 17.25, (T) 12.22 mumol of glucosyl units . g-1 (each number represents the difference between the means). It concluded that treatment with T3 markedly affects this exercise-induced metabolism of glycogen in each muscle type. From our study it is suggested that low muscle glycogen content may contribute to a reduction in exercise performance in hyperthyroidism.

[Carbohydrate metabolism in smooth muscles]. / Metabolizm weglowodanów w miesniach gladkich.

In smooth muscles carbohydrate metabolism is compartmented and somewhat different than in skeletal muscles. Oxidative metabolism is closely coordinated with contractile activity and aerobic glycolysis (measured as lactate production) is correlated with the activity of the Na/K-ATPase.