Anticancer Effects of Propolis Extracts Obtained Using the Cold Separation Method on Breast Cancer Cell Lines.

Propolis and its extracts show a wide spectrum of biological activity. Due to the necessity to use high temperatures and high polarity in the eluent, the obtained extracts are depleted of active compounds. The new, cold separation method allows obtaining a qualitatively better product containing a number of chemical compounds absent in extracts obtained using high-temperature methods. The purpose of our study was to evaluate the biological activity of propolis extracts produced with the cold separation method in four female breast cancer cell lines: MDA-MB-231, MDA-MB-468, MCF-7, and T-47D. The results of the breast cancer cell viability were obtained using the MTT test. Propolis extracts at 75 and 80% showed similar cytotoxicity against cancer cells, with the polyphenol fraction 75% being slightly more negative for cells. Propolis extracts at concentrations of 50, 75, and 100 µg/mL significantly reduced cell viability. With the exception of the MDA-MB-231 line, cell viability was also decreased after incubation with a concentration of 25 µg/mL. Our results suggest that propolis extracts obtained with the cold separation method may be considered as promising compounds for the production of health-promoting supplements.

Anticancer Effects of Propolis Extracts Obtained with the Cold Separation Method on PC-3 and DU-145 Prostate Cancer Cell Lines.

Plant extracts are increasingly tested for their biological activity and interactions with neoplastic cells. One of such sources of biologically active substances is propolis. This product has been known for thousands of years and is widely used in alternative, folk medicine. Articles describing its effects on the metabolism and cell signaling pathways of neoplastic cells derived from different organs are also published more and more frequently. The purpose of our study was to evaluate the biological activity of propolis extract produced with the cold separation method into hormone-dependent and hormone-independent prostate cancer cell lines. In our study, the propolis extracts showed at least an inhibitory effect on the growth of PC-3 and DU-145 neoplastic cells. Our results suggest that propolis extracts obtained with the cold separation method may be considered as promising compounds for the production of health-promoting supplements.

A Successful New Case of Twin Pregnancy in a Patient with Swyer Syndrome-An Up-to-Date Review on the Incidence and Outcome of Twin/Multiple Gestations in the Pure 46,XY Gonadal Dysgenesis.

BACKGROUND: The aim of the present study is to report a rare occurrence of a successful twin pregnancy in a woman with pure 46,XY gonadal dysgenesis. RESULT(S): A patient with Swyer syndrome (pure 46,XY gonadal dysgenesis) presented with a twin pregnancy after in vitro fertilization. Due to unidentified conditions, the patient developed selective intrauterine growth restriction in one of the fetuses. Twins were born at 33 weeks of pregnancy due to the risk of asphyxia. Nonetheless, the patient did not develop gonadal malignancies before the pregnancy and, despite receiving estrogen, remained amenorrheic. CONCLUSION(S): The aim of this case report is to show the course of twin pregnancy in patients with Swyer syndrome through assisted reproduction. Due to certain disorders in the development of their reproductive organs, such as the less mature uterus, such pregnancies may be associated with an increased risk. The above case report demonstrates the need to systematize methods of pregnancy management in patients with Swyer syndrome, such as: preparation for the pregnancy, assessment of the uterus, medications used, and necessary checkups. Capsule: This case report and review shows clinicians that patients with Swyer syndrome may become pregnant. Twin pregnancies may occur without any major problems through assisted reproduction.

The State of Health and the Quality of Life in Women Suffering from Endometriosis.

Quality of life is related to good health, family relations, feeling of self-esteem, and ability to cope with difficult situations. Endometriosis is a chronic condition which affects different areas of life. The lack of satisfaction in everyday life is mainly due to constant pain. The process of adjusting to a life with illness is associated with negative emotions. The aim of the article is to review the current state of knowledge concerning the impact of social and medical factors on a population of women affected by endometriosis. Women with endometriosis have an impaired quality of life compared to the general female population. Psychological consequences of endometriosis include: depression, anxiety, powerlessness, guilt, self-directed violence, and deterioration of interpersonal relations. It may contribute to lower productivity at work and less satisfying intimate life. A multi-disciplinary, evidence-based care is needed. The disease can take away the ability to be physically active, obtain an education, work continuously, and interact with friends. Social support and cognitive-behavioral therapy are extremely important for healing.

LC-ESI-MS/MS Identification of Biologically Active Phenolics in Different Extracts of Alchemilla acutiloba Opiz.

Liquid chromatography electrospray ionization tandem mass spectrometric (LC-ESI-MS/MS) qualitative and quantitative analysis of different extracts from the aerial parts and roots of Alchemilla acutiloba led to the identification of phenolic acids and flavonoids. To the best of our knowledge, isorhamnetin 3-glucoside, kaempferol 3-rutinoside, narcissoside, naringenin 7-glucoside, 3-O-methylquercetin, naringenin, eriodictyol, rhamnetin, and isorhamnetin were described for the first time in Alchemilla genus. In addition, the antioxidant, anti-inflammatory and cytotoxic activity of all extracts were evaluated. The results clearly showed that among analyzed extracts, the butanol extract of the aerial parts exhibited the highest biological activity comparable with the positive controls used.

Brief Review of Endometriosis and the Role of Trace Elements.

Endometriosis is a chronic, estrogen-dependent, inflammatory condition that is defined as the presence of endometrial glands and stroma outside the uterine cavity. Despite the progress in research into the mechanisms leading to the development of endometriosis, its cause has not yet been established. It seems to be possible that the formation of oxidative stress may be one of the main causes of the development of endometriosis. There is much research that studies the potential role of trace elements in the appearance of endometrial-like lesions. Most studies focus on assessing the content of selected trace elements in the blood, urine, or peritoneal fluid in women with endometriosis. Meanwhile, little is known about the content of these elements in endometrial-like implants, which may be helpful in developing the theory of endometriosis. Investigations that are more comprehensive are needed to confirm a hypothesis that some trace elements play a role in the pathomechanism of endometriosis.

Thrombocytopenia in pregnant women.

Thrombocytopenia is one of the two most common hematological problems in pregnant women. It is defined as the platelet (PLT) count below 150 × 103/µL. Gestational incidental thrombocytopenia (GIT) represents about 75% of thrombocytopenia cases in pregnancy and it is believed that GIT is secondary to accelerated platelet destruction and increased plasma volume associated with pregnancy. The pregnancy complications such as preeclampsia and its most severe form - HELLP syndrome account for 20% cases of thrombocytopenia in pregnancy and primary immune thrombocytopenic purpura (ITP) - for 3-4 percent. During ITP, maternal antiplatelet antibodies can pass through the placenta and bind to fetal thrombocytes leading to the development of fetal thrombocytopenia which occurs in about 50% cases. Even if the maternal platelet count stabilizes, the estimated fetal and neonatal risk of thrombocytopenia in ITP is approximately 30%. Other types of thrombocytopenia in pregnant women constitute 1-2% of cases (disseminated intravascular coagulation, autoimmunological diseases, congenital, infection and drug-related, concomitant with blood neoplastic diseases). Although thrombocytopenia in pregnant women usually has a mild course, in case of a significant decrease in PLT count may lead to dangerous bleeding, especially when the platelet count falls below 20 × 103/µL. Since it is important to identify the cause of thrombocytopenia and to determine the risk for both the mother and the child, this paper presents the influence of maternal thrombocytopenia on the pregnancy course as well as its etiology and diagnostics. The treatment principles are discussed.

An association of circulating Tregs and Th17 cells producing IL-21 and IL-22 with the ROMA in ovarian cancer patients.

The purpose of the present study was to assess the association of regulatory T cells (Tregs; CD4+ FOXP3+) and helper T lymphocytes (Th17) releasing interleukin (IL)-21 and IL-22, with the Risk of Ovarian Malignancy Algorithm (ROMA). Similar association was made with two additional tumour markers, human epididymis protein 4 (HE4) and carbohydrate antigen 125 (CA125) from patients serum. The presence of Tregs and Th17 was determined both in the peripheral blood and in the tissue of epithelial ovarian tumors. Mononuclear cells obtained from patient's peripheral blood (PBMCs) and from ovarian tissue were isolated by density gradient centrifugation. As a control group patients who had undergone surgery for infertility without ovarian pathology were selected. The percentage of Tregs and Th17 releasing IL-21 or IL-22 cells from both peripheral blood and tumor tissue was measured by flow cytometry. No differences in demographic parameters like body mass index, age, or gravidity were observed among the studied groups. However, an increased concentration of marker HE4 and value of ROMA was identified in individuals with ovarian cancer when compared with women with cystadenomas. Furthermore, a negative correlation between the ROMA value in the serum and Tregs from the peripheral blood of patients with cystadenoma ovarian tumors was detected. The presented work documents, for the first time, the negative association between peripheral blood Tregs and ROMA evaluation based on the tumour markers present in the serum of women with ovarian cystadenoma. Such an effect might result from the negative impact of Tregs on the inflammation process and on tumorigenesis caused by the persistent inflammation.

Th17 Cells and IL-17 As Novel Immune Targets in Ovarian Cancer Therapy.

Ovarian cancer (OC) is usually diagnosed at an advanced stage and is related with poor prognosis. Despite numerous studies, the pathogenesis of OC is still unknown. Recent studies indicate the role of the immune system in the development and spread of OC. The identification of factors and mechanisms involved in that process and their modulation is crucial for creating effective antitumor therapy. We investigated the potential role of Th17 cells in OC patients (n = 71) by analyzing the frequencies of Th17 cells in three different environments, i.e., peripheral blood (PB), peritoneal fluid (PF), and tissue (Th17 infiltrating cells), and the concentration of IL-17A in plasma and PF of patients in terms of their clinical and prognostic significance. Th17 cells were analyzed by flow cytometry as a percentage of CD4+ lymphocytes that expressed intracellular expression of IL-17A. The level of IL-17A in plasma and PF were determined by ELISA. Our results showed accumulation of Th17 cells among tumor-infiltrating CD4+ lymphocytes (p < 0.001 in relation to PB). Moreover, the percentage of Th17 cells in both PB and PF of OC patients was significantly lower than that in benign tumors group (n = 35). There were no significant differences in the percentage of Th17 cells in PB, PF, and tissue in relation to clinicopathological characteristics of OC patients and survival. The lower percentage of Th17 cells in the PB and PF of OC patients may promote evasion of host immune response by cancer cells. The concentration of IL-17A in plasma of OC patients was higher (p < 0.0001) than that in both benign tumors and control group (n = 10). The PF IL-17A level in OC patients was higher (p < 0.0001) than that in women with benign ovarian tumors, indicating its synthesis in OC microenvironment. Higher IL-17A level in PF is correlated with longer (median: 36.5 vs. 27 months) survival of OC patients.

Atypical Endometrial Hyperplasia Arising in a Cesarean Section Scar: A Mechanism of Malignant Transformation.

The incidence of scar endometriosis in Cesarean sections varies between 0.03 and 0.4%. However, the recently increased rate of Cesarean sections worldwide may be causing an increase in occurrence of scar endometriosis. This report presents anatomopathological evidence of an early-stage malignant transformation in endometriotic tissue from a post-Cesarean scar and briefly reviews possible underlying mechanisms. A 40-year-old woman with a body mass index of 42.7 was referred to the gynecological department with recurrent pain and presence of a palpable mass in her Cesarean section scar. She had undergone this procedure 7 years earlier and began experiencing discomfort and pain at the incision site 6 months postoperatively. Surgical treatment was instituted with complete removal of the lesion. Anatomopathological examination revealed endometriotic tissue intertwined with atypical endometrial hyperplasia and fibrosis. At 2 years' follow-up, she was asymptomatic, both clinically and based on ultrasound examination. Endometriotic foci inoculated within an abdominal scar may undergo malignant transformation. Long-lasting abdominal scar endometriosis, in morbidly obese women, requires special attention from the physician.

Expression of p53 and selected proliferative markers (Ki-67, MCM3, PCNA, and topoisomerase IIα) in borderline ovarian tumors: Correlation with clinicopathological features.

BACKGROUND: The expression of p53 has been studied not only in primary human ovarian carcinomas, but also in borderline ovarian tumors, however, the results were discordant. Expression patterns of proteins involved in cell proliferation and apoptosis have been investigated in various human neoplasms, including female genital tract neoplasms. OBJECTIVE: The aim of this investigation was to assess the staining pattern and immunolocalization of p53 and selected proliferative markers (Ki-67, MCM3, PCNA, and topoisomerase IIα) in borderline ovarian tumors (BOTs). DESIGN: The study group consisted of 42 women who underwent pelvic surgery between 2006-2015. The median patients' age was 46 years. The immunoperoxidase technique was employed using antibodies against p53, Ki-67, MCM3, PCNA, and topoisomerase IIα. RESULTS: For p53, nuclear expression was observed in BOTs, however, cytoplasmatic immunoreactivity was also detected. Altogether, 25 (60%) tumors demonstrated positive p53 immunostaining, including overexpression found in 6 (14%). There were no significant differences in p53 expression between subgroups of clinicopathological variables. Immunoexpression of Ki-67, MCM3, PCNA, and topoisomerase IIα was nuclear. Ki-67 expression was positive in 12 (29%) cases and there was a trend towards a relationship between patients' age and Ki-67 staining (P=0.08). Interestingly, a significantly higher Ki-67 expression was found in tumors of ≥10 cm in diameter compared to smaller tumors (P=0.008). MCM3 expression was detected in 38 (90%) tumors, and PCNA expression in 28 (67%), yet none of clinicopathological factors was related to them. Topoisomerase IIα expression was present in 14 (33%) cases and, interestingly, its significantly higher expression was observed in BOTs of ≥10 cm in diameter compared to smaller tumors (P=0.008). Moreover, Spearman's correlation revealed highly significant positive associations between Ki-67 and topoisomerase IIα (R=0.403, P=0.008) and Ki-67 and MCM3 (R=0.469, P=0.001). CONCLUSIONS: We report a high positive immunostaining rate for p53, suggesting a role of TP53 alterations in the development of BOTs in humans. The new finding of higher topoisomerase IIα immunostaining positivity in BOTs of ≥10 cm may be clinically relevant and requires further studies on larger patient groups.

Fas-Related Apoptosis of Peritoneal Fluid Macrophages in Endometriosis Patients: Understanding the Disease.

Recent studies of the peritoneal cavity environment in endometriosis demonstrate quantitative and qualitative changes in the cells responsible for cell-mediated immunity. Such changes may have led to disturbances in the surveillance, recognition, and destruction of misplaced endometrial cells and might have, in fact, brought about the disease. The aim of the study was to assess CD95 (Fas) expression on (activated) peritoneal fluid (PF) macrophages, as well as to ascertain soluble Fas (sFas) concentration in the PF of endometriosis patients, as compared to the nonendometriotic group. The concentration of leukocytes in the PF, the percentage of cells expressing CD45+/CD14+, and the percentage of PF macrophages expressing the HLA-DR antigen were significantly higher in patients with stages I and II endometriosis. The percentage of Fas- (CD95+-) expressing macrophages was significantly higher in all stages of the disease, in comparison with controls. Moreover, the concentration of sFas in the PF of patients with moderate and severe endometriosis was significantly higher, as compared to the reference group. The high number of immune cells in PF in early stage endometriosis and their increased susceptibility to apoptosis confirm the role of the impaired peritoneal environment and immune defects in the development and progression of the disease.

The Putative Role of TP53 Alterations and p53 Expression in Borderline Ovarian Tumors - Correlation with Clinicopathological Features and Prognosis: A Mini-Review.

Borderline ovarian tumors (BOTs) represent an independent group among ovarian malignancies, being diagnosed at clinical stage earlier than invasive ovarian carcinomas (OCs) and characterized by a rather favorable outcome after careful surgical management. Data published worldwide showed a substantial discordance of p53 expression in BOTs. The purpose of this work was to present the current status of knowledge on the significance of TP53 gene and p53 protein product alterations in BOTs. In general, higher p53 expression patterns were reported for ovarian malignancies compared to BOTs. Serous, mucinous, and endometrioid BOTs differ substantially in relation to p53 immunostaining, but data concerning the relationship between the protein's immunoreactivity and other clinico-pathological variables are scarce. Finally, reports published to date support the view that TP53 alterations may not be commonly associated with the borderline phenotype of ovarian tumors but they probably occur during the development of invasive OCs. In light of these uncertainties, the impact of TP53 alterations and p53 expression on overall survival in women affected by BOTs requires further multi-institutional studies in large cohorts of patients.

Hyperprolactinaemia - a problem in patients from the reproductive period to the menopause.

Hyperprolactinaemia especially affects women in reproductive age (90/100,000) but also often is diagnosed in menopause age and leads to disturbances in functioning of LH-RH neurons and, as a consequence, to a decrease of FSH and LH, which causes inhibition of oestradiol production. Prolactin is a peptide hormone, phylogenetically one of the oldest, stimulating cells of various organs, which is produced and secreted mainly by lactotrophic acidophilic cells of the anterior lobe of the pituitary. It influences the increase in the mass of the mammary glands, and stimulation and maintenance of lactation after delivery. There are a number of factors apart of pregnancy, delivery, and lactation than can influence secretion of the hormone in other physiological and pathological circumstances, like high-protein diet, stress, REM sleep, or neoplastic tumours, inflammatory diseases, chronic systematic diseases, thyroid hormonal changes, and drug intake. The purpose of this review is to summarise the current knowledge regarding the proper diagnosis and possible influence of hyperprolactinaemia on fertility and menopause symptoms and current treatment methods.

Assessment of Th17 lymphocytes and cytokine IL­17A in epithelial ovarian tumors.

The present study was carried out to assess the percentage of T helper 17 (Th17) lymphocytes in blood and tissue and IL-17A serum concentrations in patients with epithelial ovarian tumors. Two isoforms, IL-17A and IL-17F, as well as IL-21 and IL-22, were simultaneously investigated. The study group consisted of 60 women affected by epithelial ovarian tumors (benign, borderline and malignant) and 20 women without ovarian pathology as a control group. The evaluation of the percentage of Th17 cells secreting IL-17A, IL-17F, IL-21 and IL-22 in peripheral blood and tumor tissues was performed using flow cytometry applying a Th17 cytokine staining panel. The blood serum concentration of IL-17A was determined using ELISA. We found no statistically significant differences in the subpopulations of Th17 lymphocytes, either in peripheral blood or in ovarian tissues, following comparison of the women with and without ovarian pathology. Negative correlations were found between the percentage of CD4+/IL-21+ (rs=0.8, p=0.02) and CD4+/IL-17+ (rs=-0.78, p=0.03) in the tissue and IL-17A in blood serum in the group of patients with borderline ovarian tumors. A negative correlation was also found between IL-17A and the percentage of CD4+/IL-21+ in peripheral blood (rs=­0.48, p=0.03) in the group of patients with ovarian cancer. The increased percentage of Th17 cells in tissue was not correlated with the overall survival of the ovarian cancer patients. In conclusion, we showed that more Th17 cells secreted IL-17A and IL-21 in the tissue of borderline ovarian tumors and less IL-17A in serum. We also observed that in peripheral blood of the patients with ovarian cancer, there was a higher percentage of Th17 lymphocytes and a lower concentration of IL-17A in serum indicating a negative correlation. An increased percentage of Th17 cells in ovarian tissue does not influence the time of survival of patients with ovarian cancer.

Increased percentage of Th17 cells in peritoneal fluid is associated with severity of endometriosis.

AIM: Th17 cells are a newly discovered T helper lymphocyte subpopulation, producing interleukin IL-17. Th17 cells are present in blood and peritoneal fluid (PF) at different stages of endometriosis. We aim to establish their potential importance in the pathogenesis and clinical features of the disease. METHODS: The percentage of Th17 cells among T helper lymphocytes was determined in the PF and peripheral blood (PB) of patients with endometriosis and in the control group by flow cytometry using monoclonal antibodies: anti-CD-4-FITC, anti-CD-3-PE/Cy5, and anti-IL-17A-PE. RESULTS: Th17 percentage is increased in PF in comparison with PB in both endometriotic patients and in the control group. In severe endometriosis, the percentage of Th17 cells in PF was higher than with early (I/II stage) endometriosis. A positive correlation between the percentage of Th17 cells in PF and the white blood cell count in PB was found in patients with endometriosis. CONCLUSION: Targeting the activity of PF Th17 cells may have an influence on the proliferation of ectopic tissue and clinical manifestations of the disease.

[Umbilical cord blood as a source of nerve and stem cells]. / Krew pepowinowa jako zródlo komórek nerwowych i macierzystych.

OBJECTIVES: The aim of the study was to assess proliferative ability of the stem cells in the umbilical cord blood and their potential to differentiate in in vitro culture. MATERIAL AND METHODS: The material consisted of 14 samples of umbilical cord blood collected from the umbilical cord vein. Mononuclear cells were isolated using the method of density gradient medium. Next, CD34 cells were isolated from the interphase with the use of the VarioMACS sorter and anti-CD34 antibodies. Long-term cultures were conducted on Iscove's modified Dulbecco medium (IMDM) with addition of granulocyte-macrophage colony stimulating factor (GM-CSF) and nerve growth factor (NGF). Qualitative identification was performed using the May-Grunwald-Giemsy staining method, taking photographs with a confocal microscope, and with the immunoenzymatic method. RESULTS: In our research, CD34+ stem cells constituted 1.16% of the mononuclear cells, and after centrifugation in medium 0.37% of leukocytes in whole umbilical cord blood. Even after 60 days of culture without addition of the growth factors, CD34+ cells were present in the fraction of adherent cells. After stimulation with GM-CSF and NGF a part of the umbilical cord blood cells were transformed into nerve cells (presence of neuron-specific enolase was shown) and into cells morphologically similar to fibroblast and dendritic cells. CONCLUSIONS: After stimulation with GM-CSF and NGF cytokines, the umbilical cord blood cells proliferate in long-term medium, transform into nerve cells and into cells similar to fibroblast and dendritic cells.

Well-differentiated mucinous uterine adenocarcinoma predominantly diagnosed as adenoma malignum: a case report with an immunohistochemical analysis.

Adenoma malignum (AM), also referred to as "minimal deviation adenocarcinoma", is an extremely uncommon variant of highly-differentiated adenocarcinoma of the uterine cervix. The study presented herein describes a case of uterine AM found out after hysteroscopy. An early-stage, well-differentiated mucinous uterine adenocarcinoma was diagnosed post-operatively. A subsequent immunohistochemical assessment of a panel of antibodies was applied, in order to distinguish between female genital tract malignancies.