Dilatative fetal cardiomyopathy followed by a mirror syndrome.

Mirror syndrome (Ballantyne syndrome) is a rare condition characterized by maternal edema, which often affects the lungs. It mirrors the image of fetal and placental edema; therefore, it is also called triple edema. We present the case of a 37-year-old secundigravida, referred to our clinic at 26 weeks of a pregnancy complicated by fetal dilatative restrictive cardiomyopathy and hydrops, placentomegaly, new-onset dyspnea, and maternal calf edema. Due to worsening mirror syndrome, preterm labor was induced. Labor was complicated, with soft tissue dystocia, stillbirth, and postpartum hemorrhage. The first pregnancy was also complicated by fetal right ventricular noncompaction dilatative cardiomyopathy. A eutrophic male child was born vaginally at term and died due to deterioration of the cardiac disease in the third year of life. Next-generation sequencing panel for pediatric cardiology was performed in the deceased child and parents. Two gene variants were recorded: MYOM1: c.770_771delCA (p.Thr257fs) and TPM1: c.814G>A (p.Glu272Lys). Both variants were classified as variants of uncertain significance. This case emphasizes the importance of antenatal counseling, the timing of labor induction, appropriate management of possible complications such as postpartum hemorrhage and soft tissue dystocia, and the interpretation of placental biomarkers in the context of mirror syndrome. Finally, it contributes to understanding the clinical significance of the MYOM1 and TPM1 gene variants.

Dishevelled family proteins (DVL1-3) expression in intrauterine growth restriction (IUGR) placentas.

Dishevelled family proteins (DVL1, DVL2, and DVL3) are cytoplasmic proteins that are involved in canonical and non-canonical Wnt signaling pathway during embryonic development. The role of DVL proteins in the placental tissue remains mostly unknown. In the current study, we explored the role of Dishevelled proteins in naturally invasive tissue, trophoblast. Formalin-fixed paraffin-embedded samples of 15 term placentas from physiologic term pregnancies and 15 term placentas from pregnancies complicated with intrauterine growth restrictions (IUGR) were used for the study. Expression levels of mRNA for DVL1, DVL2, and DVL3 in placentas were analyzed by quantitative real-time PCR (qRTPCR). DVL1, DVL2, and DVL3 protein expression were semi-quantitatively analyzed using immunohistochemistry. The expression of DVL2 and DVL3 proteins was significantly higher in trophoblasts in placental villi from IUGR pregnancies compared with the control group of term placentas. In contrast, DVL3 protein expression was significantly higher in endothelial cells in placental villi from IUGR pregnancies compared with normal term placentas. The observed differences at protein levels between normal and IUGR placentas were not confirmed at the mRNA levels of DVL genes. Our data indicate the active involvement of DVL proteins in IUGR-related placentas. No significant changes were observed in DVL mRNA levels between the two groups of placentas. Further studies are required to explore the clinical relevance of these observations.

The role of glycogen synthase kinase 3 (GSK3) in cancer with emphasis on ovarian cancer development and progression: A comprehensive review.

Glycogen synthase kinase 3 (GSK3) is a monomeric serine-threonine kinase discovered in 1980 in a rat skeletal muscle. It has been involved in various cellular processes including embryogenesis, immune response, inflammation, apoptosis, autophagy, wound healing, neurodegeneration, and carcinogenesis. GSK3 exists in two different isoforms, GSK3α and GSK3ß, both containing seven antiparallel beta-plates, a short linking part and an alpha helix, but coded by different genes and variously expressed in human tissues. In the current review, we comprehensively appraise the current literature on the role of GSK3 in various cancers with emphasis on ovarian carcinoma. Our findings indicate that the role of GSK3 in ovarian cancer development cannot be decisively determined as the currently available data support both prooncogenic and tumor-suppressive effects. Likewise, the clinical impact of GSK3 expression on ovarian cancer patients and its potential therapeutic implications are also limited. Further studies are needed to fully elucidate the pathophysiological and clinical implications of GSK3 activity in ovarian cancer.

Dishevelled family proteins in serous ovarian carcinomas: a clinicopathologic and molecular study.

Dishevelled family proteins (DVL1, DVL2, and DVL3) are cytoplasmic mediators involved in canonical and non-canonical Wnt signaling that are important for embryonic development. Since Wnt signaling promotes cell proliferation and invasion, its increased activation is associated with cancer development as well. To get deeper insight into the behavior of Dishevelled proteins in cancer, we studied their expression in serous ovarian carcinomas [both low- (LGSC) and high-grade (HGSC)], and HGSC cell lines OVCAR5, OVCAR8, and OVSAHO. DVL protein expression in serous ovarian carcinomas tissues was analyzed using immunohistochemistry, while DVL protein and mRNA expressions in HGSC cell lines were analyzed using Western blot and quantitative real-time PCR. DVL1 protein expression was significantly higher in LGSC compared with normal ovarian tissue, while DVL3 was overexpressed in both LGSC and HGSC. DVL2 and DVL3 protein expression was higher in HGSC cell lines when compared with normal control cell line FNE1, while DVL1, DVL2, and DVL3 mRNA expression was significantly increased only in OVSAHO cell line. Survival analysis revealed no significant impact of DVL proteins on patients' outcome. Our data show an active involvement of Dishevelled family proteins in serous ovarian carcinomas. Further studies should confirm the clinical relevance of these observations.

Novel Epigenetic Biomarkers in Pregnancy-Related Disorders and Cancers.

As the majority of cancers and gestational diseases are prognostically stage- and grade-dependent, the ultimate goal of ongoing studies in precision medicine is to provide early and timely diagnosis of such disorders. These studies have enabled the development of various new diagnostic biomarkers, such as free circulating nucleic acids, and detection of their epigenetic changes. Recently, extracellular vesicles including exosomes, microvesicles, oncosomes, and apoptotic bodies have been recognized as powerful diagnostic tools. Extracellular vesicles carry specific proteins, lipids, DNAs, mRNAs, and miRNAs of the cells that produced them, thus reflecting the function of these cells. It is believed that exosomes, in particular, may be the optimal biomarkers of pathological pregnancies and cancers, especially those that are frequently diagnosed at an advanced stage, such as ovarian cancer. In the present review, we survey and critically appraise novel epigenetic biomarkers related to free circulating nucleic acids and extracellular vesicles, focusing especially on their status in trophoblasts (pregnancy) and neoplastic cells (cancers).

Negative regulators of Wnt signaling pathway SFRP1 and SFRP3 expression in preterm and term pathologic placentas.

OBJECTIVE: Since Wnt signaling pathway plays a pivotal role in the placental development, we explored the expression of its negative regulators, SFRP1 and SFRP3 proteins in placentas from pathological pregnancies and compared their levels with those in healthy placentas. METHODS: Placentas (n = 79) were stained for SFRP1, and SFRP3 proteins by immunohistochemistry and their expression levels were quantified by stereological variable of volume density (Vv, mm°). RESULTS: Significantly higher expressions of SFRP1 and SFRP3 were found in all investigated groups of term and preterm pathologic placentas as well as in preterm control placentas in comparison with normal-term placentas. CONCLUSIONS: Our findings indicate the active involvement of negative Wnt regulators SFRP1/SFRP3 in placental development and important role in pathology of pregnancy.

ELF5 transcription factor expression during gestation in humans and rats - an immunohistochemical analysis.

OBJECTIVE: The purpose of this study was to measure immunohistochemically the expression of ELF5 protein in term human and rat placentas and in human placentas associated with gestational diabetes (GD) and intrauterine growth restriction (IUGR). METHODS: The results were quantitated stereologically using the stereological variable of volume density. A semiquantitative analysis was performed independently by a certified pathologist. RESULTS: Total expression of ELF5 protein was higher in pathological pregnancies than in corresponding control term placentas, with both methods of quantifications showing similar results. In addition, ELF5 expression was also higher in connective tissue and blood vessels in chorionic villi in IUGR placentas (but not in GD placentas) compared to healthy controls. ELF5 is higher in placenta as a whole and in most of its components in both pathologies. The two exceptions are chorionic plates in IUGR and decidua in GD, where its expression is lower than in healthy controls. CONCLUSIONS: We have shown that IUGR and GD are associated with significantly increased levels of ELF5 protein in placentas, which suggests that ELF5 may play an important role in normal placentation. However, these are term placentas and to study ELF5 in premature births would give better insight into human placentation in health and disease.

[CANCER PATIENTS FOLLOW-UP ­ CROATIAN SOCIETY OF MEDICAL ONCOLOGY CLINICAL GUIDELINES Part I: breast cancer, uterine cancer, cervical cancer, ovarian cancer]. / PRACENJE ONKOLOSKIH BOLESNIKA ­ KLINICKE PREPORUKE HRVATSKOG DRUSTVA ZA INTERNISTICKU ONKOLOGIJU HLZ-a 1. dio: rak dojke, rak tijela maternice, rak vrata maternice, rak jajnika.

Treatment of oncological patients must be based upon multidisciplinary approach, and takes place in specialized oncological centers. By the end of a specific oncological treatment further follow-up is managed mostly by the ­oncologists, but the role of the general practitioners becomes more important every day and therefore should be precisely defined. Nowadays, most of the existing follow-up guidelines are not based on prospective studies, but on the experts ­opinion of a particular oncological center or specialists. The aim of the Croatian Society of Medical Oncology (CSMO) with these recommendations is to standardise and rationalise the diagnostic procedures algorithm in the follow-up of oncological patients after primary treatment.

CANCER PATIENTS FOLLOW-UP ­ CROATIAN SOCIETY OF MEDICAL ONCOLOGY CLINICAL GUIDELINES Part I: breast cancer, uterine cancer, cervical cancer, ovarian cancer. / PRACENJE ONKOLOSKIH BOLESNIKA ­ KLINICKE PREPORUKE HRVATSKOG DRUSTVA ZA INTERNISTICKU ONKOLOGIJU HLZ-a 1. dio: rak dojke, rak tijela maternice, rak vrata maternice, rak jajnika

Treatment of oncological patients must be based upon multidisciplinary approach, and takes place in specialized oncological centers. By the end of a specifi c oncological treatment further follow-up is managed mostly by the oncologists, but the role of the general practitioners becomes more important every day and therefore should be precisely defi ned. Nowadays, most of the existing follow-up guidelines are not based on prospective studies, but on the experts opinion of a particular oncological center or specialists. The aim of the Croatian Society of Medical Oncology (CSMO) with these recommendations is to standardise and rationalise the diagnostic procedures algorithm in the follow-up of oncological patients after primary treatment.

Structural changes in the rat placenta during the last third of gestation discovered by stereology.

Structural changes in the rat placenta during the last third of gestation were for the first time assessed by stereology. Fischer female rats were euthanized on the day 16 or day 19 of gestation, and 35 placentas were collected. Three randomly selected placentas from each group were stereologically analyzed for the absolute volume. The proportion of the glycogenic cells and the trophoblast giant cells (TGC) in the basal part of the placenta was calculated using volume density. The absolute volume of the rat placenta on the day 16 of gestation was determined as 0.0638 cm3. The labyrinth comprised 0.0274 cm3, the basal plate 0.0271 cm3 and the decidua 0.0093 cm3. On the day 19 of gestation, the absolute volume of the placenta was 0.1627 cm3, the labyrinth occupied 0.0922 cm3, the basal plate 0.0596 cm3 and the decidua 0.0109 cm3. The volume density of trophoblast giant cells was 0.174 cm0 on the day 16 and 0.107 cm0 on the day 19 of gestation. The glycogenic cells comprised 0.379 percentage of the basal plate on the day 16 and 0.236 on the day 19 of gestation. We conclude that the absolute volume of the whole placenta and the labyrinth has increased from day 16 to the day 19 of gestation. In contrast, the volume density of glycogenic cells and trophoblast giant cells was higher on the day 16 than on the day 19 of gestation, probably due to the intensive trophoblast invasion during that time.

Glycosylation pattern and axin expression in normal and IUGR placentae.

OBJECTIVE: The aim of this study was to investigate the protein glycosylation pattern and AXIN1 protein expression in human placentae of normal pregnancies and compare them with placentae of pregnancies complicated with intrauterine growth restriction (IUGR). METHODS: A total of 38 placentae (17 placentae of IUGR fetuses from singleton pregnancies and gestational age-matched 21 control placentae from normal singleton pregnancies) were collected from the Clinical Hospital Sveti Duh, Department of Gynecology and Obstetrics, Zagreb, Croatia. Gestational age was determined according to the last menstrual period (LMP) and by ultrasound measurements. Expression of glycoproteins was measured by Western blotting with SNA, UEA-I, PHA-E and DBA lectins as probes whereas expression of AXIN1 was determined by immunohistochemistry. RESULTS: Comparison of detected sugars revealed differences in protein glycosylation between normal and IUGR placentae. Higher expression of AXIN1 protein located mostly in the cytoplasm of syncytiotrophoblast and to a lesser extent in its nuclei was found in IUGR placentae. CONCLUSION: Results of our study suggest that changes in glycoprotein content may contribute to restricted placenta growth and development. Higher expression of AXIN1 protein in IUGR placentae indicates a role of Wnt/ß-catenin signaling pathway in pathology of placental development.

Epigenetic alterations of the Wnt signaling pathway in cancer: a mini review.

Epigenetic mechanisms play a crucial role in cellular proliferation, migration and differentiation in both normal and neoplastic development. One of the key signaling pathways whose components are altered through the epigenetic mechanisms is the Wnt signaling pathway. In this review, we briefly discuss the key concepts of epigenetics and focus on the recent advances in the Wnt signaling pathway research and its potential diagnostic and therapeutic implications.

Impact of 5-azacytidine on rat decidual cell proliferation.

The DNA demethylating agent 5-azacytidine (5-azaC) has a teratogenic influence during rat development influencing both the embryo and the placenta. Our aim was to investigate its impact on early decidual cell proliferation before the formation of placenta. Thus, female Fischer rats received 5-azaC (5 mg/kg, i.p.) on the 2nd, 5th or 8th day of gestation and the decidual tissues were harvested on gestation day 9. They were then analysed immunohistochemically for expression of cell proliferation marker proliferating cell nuclear antigen (PCNA) in decidual cells and for global DNA methylation using the coupled restriction enzyme digestion, random amplification and pyrosequencing assays. We found that 5-azaC administered on the 5th and 8th (but not on 2nd) day of gestation led to increased PCNA expression in decidual cells compared with untreated controls. No significant changes in DNA methylation were detected, with either method, in any of the treated rat groups compared with untreated controls. Thus, we conclude that 5-azaC can stimulate decidual cell proliferation without simultaneously changing global DNA methylation level in treated cells.

Epigenetic control of cell invasion - the trophoblast model.

Abstract Trophoblast implantation and placentation allow the survival of the young embryo and its normal development inside the uterus. In order for these processes to function properly, the trophoblast has to undergo a series of characteristic changes that lead to its adhesion and invasion of the uterus. This is achieved, among other mechanisms, by inactivation of specific tumor suppressor genes, commonly by methylation of their promoters. Cell adhesion and tissue invasion are also characteristics of malignant tumors and patterns of methylation similar to that seen in trophoblast are found in various tumor types. Another important mechanism that aids trophoblast cells invasion is their transition from epithelial to mesenchymal phenotype. Such a transition is also a common characteristic of invading malignant cells. Thus, studying tissue invasion and its control mechanisms can benefit the understanding of both the trophoblast and malignant cells behavior.

Development of placenta in a rodent--model for human placentation.

Rodents provide an excellent experimental model to study human placental development. In this review, our aim was to explain major events that underlie the placental development in mammals in general, and specifically in rodent. Those events include trophoblast cell proliferation, decidual reaction and contact between the mesenchyme of the allantois with ectoplacental cone, all orchestrated by activation of a series of genes. We also aimed to compare molecular and genetic events of rodent and human placentation. Employing the rodent placenta development model will yield better understanding of these processes in other mammals, especially in humans.

Modification of conservative treatment of heterotopic cervical pregnancy by Foley catheter balloon fixation with cerclage sutures at the level of the external cervical os: a case report.

INTRODUCTION: Conservative treatment of a heterotopic cervical pregnancy was performed with a modification of the fixation of a Foley catheter at the level of the external cervical os, followed by the ligature of the descending cervical branches of the uterine arteries and systemic methotrexate application. CASE PRESENTATION: A 34-year-old Caucasian woman was diagnosed with double gestation after 6 weeks of in vitro fertilization treatment. A gynecological examination and color Doppler ultrasound scan revealed intra-uterine and cervical gestational sacs both containing live fetuses. A Foley catheter balloon was inserted into the cervical canal, inflated and fixed by a cerclage suture at the level of the external cervical os, followed by ligation of the descending cervical branches of the uterine arteries. Systemic methotrexate was applied. Three days after removal of the Foley catheter, an evacuation of the intra-uterine gestational sac was performed. Hemorrhage from the implantation site was controlled immediately and a pregnancy termination was successfully performed. The procedure was uneventful and our patient was discharged with a preserved uterus. CONCLUSIONS: Conservative treatment of cervical pregnancy using a Foley catheter balloon is more efficacious if the Foley catheter balloon is attached in the correct position with a cerclage suture at the level of the external os, followed by ligation of the descending cervical branches of the uterine arteries, thereby exerting maximal pressure on the bleeding vessels.

[Proposal of cervical cancer early detection programme in Croatia]. / Prijedlog programa ranog otkrivanja raka vrata maternice u hrvatskoj.

Opportunistic screening by Pap smear was introduced in Croatia in 1950s, with a consequent decrease of cervical cancer incidence and mortality. Since 1990s, no further decrease has been observed, and there are still about 370 new cases and 100 deaths of cervical cancer yearly in Croatia. In scope of the proposed early detection programme, all Croatian women aged 25-64 years should be screened. In the first phase of the programme, the target population would be tested by Pap-smear every third year. In the second phase, HPV-testing would also be introduced for women over 30 years. Organization of the programme at county level is proposed, while the evaluation and monitoring would be performed both at county level and centrally. Regarding the present costs of treatment and sick-leave of cervical cancer patients, it is estimated that the introduction of cervical cancer screening programme in Croatia would be cost-effective already after the first decade.

DNA methylation as a regulatory mechanism for gene expression in mammals.

Epigenetics refers to the study of heritable changes in gene expression that occur without a change in DNA sequence. In the last decade, it has been shown that epigenetic mechanisms provide an "extra" layer of transcriptional control that regulates genes expression. Three distinct mechanisms appear intricately related in initiating and sustaining epigenetic modifications: RNA-associated silencing, DNA methylation and histone modification. These mechanisms are critical components in the normal development and cell growth. DNA methylation is involved in transcriptional silencing of genes, regulation of expression of imprinted genes, a number of tumour suppressor genes in cancer and silencing of genes located on the inactive X chromosome. In this review, we are focused on the basic principles of DNA methylation as the main epigenetic mechanism for normal embryonic development and epigenetic alterations that contribute to carcinogenesis.