LPCAT1-TERT fusions are uniquely recurrent in epithelioid trophoblastic tumors and positively regulate cell growth.

Gestational trophoblastic disease (GTD) is a heterogeneous group of lesions arising from placental tissue. Epithelioid trophoblastic tumor (ETT), derived from chorionic-type trophoblast, is the rarest form of GTD with only approximately 130 cases described in the literature. Due to its morphologic mimicry of epithelioid smooth muscle tumors and carcinoma, ETT can be misdiagnosed. To date, molecular characterization of ETTs is lacking. Furthermore, ETT is difficult to treat when disease spreads beyond the uterus. Here using RNA-Seq analysis in a cohort of ETTs and other gestational trophoblastic lesions we describe the discovery of LPCAT1-TERT fusion transcripts that occur in ETTs and coincide with underlying genomic deletions. Through cell-growth assays we demonstrate that LPCAT1-TERT fusion proteins can positively modulate cell proliferation and therefore may represent future treatment targets. Furthermore, we demonstrate that TERT upregulation appears to be a characteristic of ETTs, even in the absence of LPCAT1-TERT fusions, and that it appears linked to copy number gains of chromosome 5. No evidence of TERT upregulation was identified in other trophoblastic lesions tested, including placental site trophoblastic tumors and placental site nodules, which are thought to be the benign chorionic-type trophoblast counterpart to ETT. These findings indicate that LPCAT1-TERT fusions and copy-number driven TERT activation may represent novel markers for ETT, with the potential to improve the diagnosis, treatment, and outcome for women with this rare form of GTD.

BRCA1 promoter hypermethylation in human placenta: a hidden link with ß-hCG expression.

Gestational trophoblastic diseases (GTD) are group of pregnancy-related tumors characterized by abnormal levels of 'ß-hCG' with higher incidence in South-East Asia, especially India. Our laboratory has reported that wild-type BRCA1 transcriptionally regulates ß-hCG in triple negative breast cancers (TNBCs). These factors culminated into analysis of BRCA1 status in GTD, which would emanate into elucidation of BRCA1- ß-hCG relationship and unraveling etio-pathology of GTD. BRCA1 level in GTD is down-regulated due to the over-expression of DNMT3b and subsequent promoter hypermethylation, when compared to the normal placentae accompanied with its shift in localization. There is an inverse correlation of serum ß-hCG levels with BRCA1 mRNA expression. The effects of methotrexate (MTX), which is the first-line chemotherapeutic used for GTD treatment, when analyzed in comparison with plumbagin (PB) revealed that PB alone is efficient than MTX alone or MTX-PB in combination, in showing selective cytotoxicity against GTD. Interestingly, PB increases BRCA1 levels post-treatment, altering DNMT3b levels and resultant BRCA1 promoter methylation. Also, cohort study analyzed the incidence of GTD at Sree Avittom Thirunal (SAT) Hospital, Thiruvananthapuram, which points out that 11.5% of gestational trophoblastic neoplasia (GTN) cases were referred to Regional Cancer Centre, Thiruvananthapuram, for examination of breast lumps. This has lend clues to supervene the risk of GTD patients towards BRCA1-associated diseases and unveil novel therapeutic for GTD, a plant-derived naphthoquinone, PB, already reported as selectively cytotoxic against BRCA1 defective tumors.

Metastatic epithelioid trophoblastic tumor of the lung: A case report.

RATIONALE: Epithelioid trophoblastic tumor (ETT) is a very rare form of gestational trophoblastic disease (GTD) which arises from neoplastic proliferation of intermediate trophoblasts. Metastatic ETT of the lung is extremely rare in postmenopausal women. PATIENT CONCERNS: Here we describe a 50-year-old woman with a metastatic ETT of the lung showing increasing tracer uptake at PET/CT. DIAGNOSIS: Hematoxylin-eosin staining showed a tumor composed of nests of epithelioid cells with necrotic debris and peritumoral hyaline-like material. Immunohistochemical staining of the tumor cells was positive for human chorionic gonadotropin (HCG) and cytokeratin 18. INTERVENTIONS: The patient underwent thoracoscopic lower left lobectomy combined with mediastinal lymphadenectomy. At surgery, a solid mass (size 3.0 × 3.0 cm) was found in the left lower lung. OUTCOMES: The patient was discharged on the tenth day postsurgery, following an uneventful recovery. Three months postsurgery, the patient was asymptomatic and is currently being managed with close follow-up. LESSONS: Metastatic ETT of lung is a very rare disease. Complete surgical resection and chemotherapy may be the critical therapeutic option.

Lowered reference limits for hCG improve follow-up of patients with hCG-producing tumors.

BACKGROUND: Human Chorionic Gonadotropin (hCG) is produced by germ cell tumors, but can also be elevated in benign conditions such as primary hypogonadism, where hCG is produced by the pituitary gland. In our experience, the reference limits for hCG (Elecsys hCG+ß-assay, Roche Diagnostics), were unnecessarily high and did not reflect levels encountered in clinical practice. We wanted to establish new reference limits to increase the clinical utility of the hCG-assay. METHODS: We analysed hCG in serum samples from a healthy adult population and in a cohort of testicular cancer survivors. The gonadotropins LH and FSH were measured in the cohort and in a selection of the reference population to assess gonadal function. RESULTS: We found low hCG levels for all men and women <45years (97.5 percentiles 0.1 and 0.2IU/L, respectively) from the healthy population (n=795) having normal FSH and LH. Due to assay limitations, we suggest a common reference limit of <0.3IU/L. For the age group ≥45, the 97.5 percentiles in the healthy population were 0.5IU/L for men and 6.0IU/L for women. In all subjects from both the reference population and the cohort (n=732), hCG levels exceeding the reference limit could be fully explained by reduced gonadal function indicated by elevated LH and FSH levels. CONCLUSION: The Elecsys hCG+ß-assay should have lower reference limits than recommended by the manufacturer, with important implications for tumor follow-up. Elevated hCG is rare with intact gonadal function, both in a normal population and among survivors of testicular cancer, and should lead to further investigations when encountered in clinical practice.

Effects of Phytoestrogen Extracts Isolated from Elder Flower on Hormone Production and Receptor Expression of Trophoblast Tumor Cells JEG-3 and BeWo, as well as MCF7 Breast Cancer Cells.

Hereinwe investigated the effect of elderflower extracts (EFE) and of enterolactone/enterodiol on hormone production and proliferation of trophoblast tumor cell lines JEG-3 and BeWo, as well as MCF7 breast cancer cells. The EFE was analyzed by mass spectrometry. Cells were incubated with various concentrations of EFE. Untreated cells served as controls. Supernatants were tested for estradiol production with an ELISA method. Furthermore, the effect of the EFE on ER/ER/PR expression was assessed by immunocytochemistry. EFE contains a substantial amount of lignans. Estradiol production was inhibited in all cells in a concentration-dependent manner. EFE upregulated ER in JEG-3 cell lines. In MCF7 cells, a significant ER downregulation and PR upregulation were observed. The control substances enterolactone and enterodiol in contrast inhibited the expression of both ER and of PR in MCF7 cells. In addition, the production of estradiol was upregulated in BeWo and MCF7 cells in a concentration dependent manner. The downregulating effect of EFE on ER expression and the upregulation of the PR expression in MFC-7 cells are promising results. Therefore, additional unknown substances might be responsible for ER downregulation and PR upregulation. These findings suggest potential use of EFE in breast cancer prevention and/or treatment and warrant further investigation.

Role of P57KIP2 Immunohistochemical Expression in Histological Diagnosis of Hydatidiform Moles.

PURPOSE: To determine the significance of P57KIP2 immunohistochemistry expression in the histopathological diagnosis of hydatidiform mole. MATERIALS AND METHODS: Hydatidiform mole patients at King Chulalongkorn Memorial Hospital between January 1999 and December 2011 were recruited. Two gynecologic pathologists reviewed histopathologic slides to confirm diagnosis. Formalin-fixed, paraffin-embedded tissue sections were stained using a bstandard immunostaining system with monoclonal antibodies against P57KIP2 protein. Correlations among pathological features, immunohistochemical expression and clinical data were analyzed. RESULTS: One hundred and twenty-seven hydatidiform mole patients were enrolled. After consensus review, 97 cases were diagnosed as complet (CHM) and 30 cases as partial (PHM). Discordance between the first and final H and E diagnoses was found in 19 cases (14.9%, k= 0.578). Significant pathological features to classify the type of hydatidiform mole are central cisterns, trophoblastic proliferation, trophoblastic atypia, two populations of villi, fetal vessels and scalloped borders. After performing immunohistochemistry for P57KIP2, 107 cases were P57KIP2 negative and 20 cases positive. Discordant diagnoses between final H and E diagnosis and P57KIP2 immunohistochemistry was identified in 12 cases (9.4%). Sensitivity of final H and E diagnosis for CHM was 89.7%; specificity was 95.0%. PHM sensitivity and specificity of final H and E diagnosis was 95.0% and 89.7%, respectively. CONCLUSIONS: Histopathological diagnosis alone has certain limitations in accurately defining types of hydatidiform mole; P57KIP2 immunohistochemistry is practical and can be a useful adjunct to histopathology to distinguish CHM from non-CHM.

Epithelioid trophoblastic tumor presenting as an ovarian mass in a postmenopausal woman.

Epithelioid trophoblastic tumor (ETT) is a rare neoplasm derived from chorionic-type intermediate trophoblastic cells. Most cases of ETT are intrauterine and present during reproductive age. We report a case of ovarian ETT developing 47 yr after the patient's last pregnancy. A 75-yr-old woman transferred to our hospital because of multiple pulmonary masses which was diagnosed as sqaumous cell carcinoma in another hospital. PET-CT revealed a huge solid mass in the pelvic cavity, suspicious for ovarian malignancy. Serum ß-hCG was 57,971 mIU/mL. Hysterectomy and bilateral salpingo-oophorectomy were performed. Gross examination showed an enlarged right ovary, measuring 17×14×7 cm. The cut surface was yellow-tan and solid with extensive areas of necrosis. The uterus was unremarkable. The histologic finding was the same as the previous lung biopsy. The tumor consisted of monomorphic cells with abundant eosinophilic cytoplasm, forming solid sheets and nests. There was geographic tumor cell necrosis with hyaline materials. Immunohistochemically, cytokeratin 7 and p63 showed diffuse reactivity in the tumor cells. There was focal staining for ß-hCG. Ki-67 proliferative index was about 80%. This case indicates that ETT can rarely occur in postmenopausal women and to the best of our knowledge, our patient is the oldest reported case of ETT to date.

The multidrug-resistance transporter ABCB5 is expressed in human placenta.

ATP-binding cassette (ABC) transporters in placenta protectively transport drugs and xenobiotics. ABCB5 [subfamily B (MDR/TAP)] is a novel ABC multidrug-resistance transporter that also mediates cell fusion, stem cell function, and vasculogenic plasticity. Immunohistochemistry and double-labeling immunofluorescence staining for ABCB5 and ABCB5/CD200, respectively, was performed on formalin-fixed, paraffin-embedded placental tissue from 5 first trimester, 5 second trimester, and 5 term pregnancies as well as 5 partial moles, and 5 complete moles. In addition, tumor cells from 5 choriocarcinoma and 5 placental site trophoblastic tumor cases were examined. ABCB5 staining was observed in villous trophoblasts in 100% (5/5) of first trimester placentas (with progressive decrease in term placentas); 100% of partial moles (5/5); and 100% of complete moles (5/5). Notably, reactivity was discretely restricted to the inner trophoblast layer, with no staining of overlying syncytiotrophoblast. Antibody specificity and localization was confirmed further by in situ hybridization. ABCB5 expression was retained in 20% of choriocarcinomas (1/5) and 40% of placental site trophoblastic tumors (2/5). Prior studies have localized expression of multidrug-resistance-1, also known as ABCB1, within the syncytiotrophoblast of early placentas, where it serves a protective function as an efflux transporter. Our results show that ABCB5 is preferentially expressed in the cytotrophoblast layer of placental villi. The expression of this novel biomarker at the maternal-fetal interface raises questions on its role in placental structure and function as well as on its potential contribution to the protective efflux provided by other P-glycoprotein transporters.

Peroxisome proliferator-activated receptor-gamma (PPARγ) is down regulated in trophoblast cells of gestational diabetes mellitus (GDM) and in trophoblast tumour cells BeWo in vitro after stimulation with PPARγ agonists.

AIMS: Peroxisome proliferator-activated receptor-gamma (PPARγ) plays an important role in insulin metabolism, trophoblast differentiation and anti-inflammatory circuits. The aim of this study was to investigate the expression of PPARγ in the placenta of patients with gestational diabetes mellitus (GDM) and the regulation of PPARγ by its agonists in trophoblast tumour cells BeWo in vitro. METHODS: PPARγ expression in a total of 80 placentas (40 GDM/40 controls) was analysed by immunohistochemistry using the semi-quantitative immunoreactive score. Furthermore, a quantitative reverse transcription-polymerase chain reaction (PCR) was performed to determine the PPARγ mRNA-expression in both groups. We used a fused and a non-fused BeWo cell culture model for the stimulation with arachidonic acid and 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2). Afterwards PPARγ mRNA-expression was analysed by quantitative real-time PCR (RT-PCR) (TaqMan). RESULTS: Using immunohistochemistry we identified a decreased expression of PPARγ in the syncytiotrophoblast and the extravillous trophoblast of GDM placentas compared to normal controls. Furthermore, PPARγ mRNA-expression was reduced in GDM placentas. Stimulation of BeWo cells with arachidonic acid and 15d-PGJ2 caused a downregulation of PPARγ expression. CONCLUSION: As PPARγ is down regulated by arachidonic acid and 15d-PGJ2, the reduced PPARγ expression in GDM placentas may be due to an altered concentration of fatty acid derivates.

HSD3B1 is a specific trophoblast-associated marker not expressed in a wide spectrum of tumors.

OBJECTIVE: Hydroxyl-δ-5-steroid dehydrogenase (HSD3B1) is an enzyme that catalyzes the oxidative conversion of δ-5-3 ß-hydroxyl steroids to the δ-4-3-keto configuration and is involved in steroid hormone synthesis. It has been shown to be expressed in normal trophoblastic tissue and benign and neoplastic trophoblastic lesions. HSD3B1 has not been detected in a large number of lung, breast, and uterine carcinomas; however, its expression has not been studied in a wide variety of other nontrophoblastic neoplasms. To test if HSD3B1 is highly specific for normal trophoblasts and trophoblast-associated lesions, we examined the expression of HSD3B1 in a wide spectrum of tumors. METHODS: Tissue microarrays containing 473 carcinomas from the lung, breast, ovary, uterus, liver, pancreas, stomach, and colon; 32 ovarian granulose cell tumors; and 12 adrenocortical adenomas were studied by immunohistochemistry using a commercially available monoclonal antibody, HSD3B1. One tissue microarray containing normal tissues was also included. Positive staining of intermediate trophoblasts and syncytiotrophoblasts in normal placental tissue served as a positive control. RESULTS: Normal tissues and tumors from the various sites were all negative for HSD3B1 except for 2 adrenocortical adenomas with weak focal immunoreactivity. CONCLUSIONS: Our study further confirmed that HSD3B1 is a highly specific trophoblast-associated marker that can be used in the distinction of trophoblastic tumorlike lesions and tumors from nontrophoblastic lesions and tumors.

Leukemia inhibitory factor increases the invasiveness of trophoblastic cells through integrated increase in the expression of adhesion molecules and pappalysin 1 with a concomitant decrease in the expression of tissue inhibitor of matrix metalloproteinases.

OBJECTIVE: To identify the invasion-associated molecules during leukemia inhibitory factor-(LIF-)mediated increase in the invasion of trophoblast cells. DESIGN: Experimental study. SETTING: Research institution. PATIENT(S): None. INTERVENTION(S): Cultured trophoblastic HTR-8/SVneo cells were treated with LIF. MAIN OUTCOME MEASURE(S): Matrigel matrix-based invasion assay of HTR-8/SVneo cells. Signaling molecules associated with LIF-mediated increase in invasion were investigated by Western blot, cDNA microarray, quantitative reverse transcriptase polymerase chain reaction, immunofluorescence, immunohistochemistry, and gene silencing by siRNA. RESULT(S): Treatment of HTR-8/SVneo cells with LIF (50 ng/mL) led to a significant increase in invasion. Treatment with LIF also led to an increase in nuclear localization of activated STAT1 and STAT3. Among 237 differentially expressed genes after LIF treatment, expression of pappalysin 1, SERPINB3, podoplanin, integrin ß3, ID1, ICAM1, and so on went up, while tissue inhibitor of matrix metalloproteinase 1 (TIMP1), TIMP2, and TIMP3 went down significantly. The presence of several of these proteins has also been demonstrated in human trophoblast cells. Silencing of pappalysin 1 led to a significant reduction in basal as well as LIF-mediated invasiveness of HTR-8/SVneo cells. CONCLUSION(S): Identification of novel molecules associated with a LIF-mediated increase in trophoblastic cell invasion may facilitate our understanding of implantation biology.

Overexpressed PAK4 promotes proliferation, migration and invasion of choriocarcinoma.

Gestational trophoblastic disease (GTD) includes frankly malignant choriocarcinoma (CCA) and placental site trophoblastic tumor and potentially malignant hydatidiform mole. p21-Activated kinase (PAK) 4 promotes cell motility. This study investigated the role of PAK4 in the pathogenesis of GTD. PAK4 messenger RNA and protein expressions in clinical samples and cell lines of normal placentas and GTD were determined by quantitative real-time polymerase chain reaction and western blot, respectively. The effects of human chorionic gonadotropin (hCG) and phosphoinositide 3 kinase (PI3K) on the expression and activation of PAK4 were investigated by treating CCA JEG3 and JAR cells with anti-hCG antibody and PI3K inhibitor, respectively. The effects of PAK4 on CCA cell proliferation, migration and invasion were assessed by corresponding functional assays. We demonstrated overexpression of PAK4 in GTD and CCA cell lines at both RNA and protein level. hCG is one of the upstream regulators of PAK4 expression, whereas activation of PAK4 is PI3K/PKB dependent in JEG3 and JAR cells. Significant correlation was found between PAK4 expression and proliferation index minichromosome maintenance complex component 7 (P = 0.007). In JEG3 and JAR cells, stably transfected PAK4 increased proliferation, migration and invasion, whereas small interfering RNA knockdown of PAK4 decreased proliferation, migration and invasion along with downregulated CDK6 and membrane-type 1 matrix metalloproteinase (MT1-MMP) and upregulated p16. We further found PAK4-mediated transcription of MT1-MMP in CCA cells by luciferase reporter assay. Our results demonstrated for the first time that overexpressed PAK4 was involved in the pathogenesis of GTD, promoting proliferation and enhancing cell migration and invasion in CCA cells.

Evolution of a trophoblastic tumor from an endometrioid carcinoma--a morphological and molecular analysis.

We report the clinicopathological and molecular characteristics of a rare uterine endometrioid carcinoma with a nongestational trophoblastic neoplastic component that was composed of both choriocarcinoma and epithelioid trophoblastic tumor. Molecular genetic analysis showed a clonal evolution from endometrioid carcinoma to trophoblastic tumor. The findings from this case have both diagnostic and biological implications that may inspire future studies on the pathogenic mechanisms by which cancer stem cells assume aberrant differentiation programs and the molecular switch(es) that initiates trophoblastic differentiation in adult tumor tissues.

Downregulation of ASPP1 in gestational trophoblastic disease: correlation with hypermethylation, apoptotic activity and clinical outcome.

Gestational trophoblastic disease encompasses a spectrum of trophoblastic lesions including true neoplasms such as choriocarcinomas and the potentially malignant hydatidiform moles, which may develop persistent disease requiring chemotherapy. ASPP1, a member of apoptosis-stimulating proteins of p53 (ASPPs), is a proapoptotic protein that can stimulate apoptosis through its interaction with p53. We evaluated the promoter methylation and expression profiles of ASPP1 in different trophoblastic tissues and its in vitro functional effect on two choriocarcinoma cell lines, namely JEG-3 and JAR. Significant downregulation of ASPP1 mRNA and protein levels was demonstrated in hydatidiform moles and choriocarcinomas, when compared with normal placentas by quantitative-PCR and immunohistochemistry. The ASPP1 mRNA level was significantly correlated with its hypermethylation status, evaluated with methylation-specific PCR, in placenta and gestational trophoblastic disease samples (P=0.024). Moreover, lower ASPP1 immunoreactivity was shown in hydatidiform moles that progressed to persistent gestational trophoblastic neoplasms than in those that regressed (P=0.045). A significant correlation was also found between expression of ASPP1 and proliferative indices (assessed by Ki67 and MCM7), apoptotic activity (M30 CytoDeath antibody), p53 and caspase-8 immunoreactivities. An in vitro study showed that ectopic expression of ASPP1 could trigger apoptosis through intrinsic and extrinsic pathways as indicated by an increase in cleaved caspase-9 and Fas ligand protein expression. The latter suggests a hitherto unreported novel link between ASPP1 and the extrinsic pathway of apoptosis. Our findings suggest that downregulation of ASPP1 by hypermethylation may be involved in the pathogenesis and progress of gestational trophoblastic disease, probably through its effect on apoptosis.

Epithelioid trophoblastic tumor: a clinicopathological and immunohistochemical study of seven cases.

The objective of this study is to evaluate the clinicopathological features and immunohistochemical characteristics of epithelioid trophoblastic tumor (ETT). Seven cases of epithelioid trophoblastic tumor treated in the Women's Hospital of Zhejiang University from 2004 September to 2008 December were retrospectively analyzed. Immunohistochemical study was performed. The most common presenting symptom was vaginal bleeding. Four patients had prior evidence of molar pregnancy and three patients presented with metastases. Mean age at diagnosis was 34.7 years. Mean pregnancy interval was 3.39 years. Human chorionic gonadotropin levels were 33.25-174315.5 IU/l. One case died from metastasis in lungs. The remaining six patients survived without recurrence. Immunohistochemistry revealed diffusely positive for CK18, and focally positive for ß-hCG, HPL, Mel-CAM (CD146) and inhibin-alpha. Nuclear staining of Ki67 and p63 were seen. The confirmation of epithelioid trophoblastic tumor diagnosis is difficult before surgery. Surgical intervention is the recommended primary treatment.

p16 expression in squamous and trophoblastic lesions of the upper female genital tract.

p16, a surrogate marker for human papillomavirus (HPV) infection, is uniformly present in HPV-related carcinomas. This study aims to further characterize p16 expression in trophoblastic lesions and squamous lesions of the upper female genital tract, as little data exists. p16 immunostaining was performed on sections from ichthyosis uteri (1), primary uterine corpus squamous cell carcinoma (UCSCC) (2), primary ovarian SCC (OSCC) (5; 2 associated with a dermoid cyst), endometrial endometrioid adenocarcinoma with extensive squamous differentiation (EC-SD) (5), ovarian endometrioid adenocarcinoma with extensive squamous differentiation (OC-SD) (4), placental site nodule (5), and placental site trophoblastic tumor (PSTT) (6). We evaluated the percentage of positive cytoplasmic and nuclear staining (focal ≤10%, multifocal=10% to 50%, and diffuse ≥50%) and staining intensity (weak, moderate, and strong). HPV-DNA analysis by polymerase chain reaction was performed on 5 OSCC. Ichthyosis uteri, all UCSCC and 1 OSCC (arising in a dermoid) were negative; the other dermoid-associated OSCC showed focal moderate staining, the remaining OSCC displayed strong (100%), diffuse (2), or multifocal (1) p16 positivity. Three of the 5 EC-SD cases showed strong diffuse staining of the squamous component. The glandular component focally showed strong p16 positivity (2), with variably intense focal staining in 3 cases. The squamous component of all OC-SD showed focal moderate staining, with variable staining of the glandular component. Overall, 3 EC-SD had 80% to 90% p16 positivity. Five of the 5 placental site nodules and 4 of the 6 PSTT showed focal weak staining, whereas 2 PSTT were p16 negative. HPV-DNA analysis was negative in 3 of the 5 OSCC, the other 2 cases being technical failures. p16 is expressed in OSCC and in the squamous and glandular components of EC-SD and OC-SD. As p16 is negative in UCSCC, it may help to identify the origin of SCC diffusely involving the corpus and cervix, and suggests different pathogeneses for SCC of the upper female genital tract, likely to be unrelated to HPV infection. In contrast to earlier data, we found weak and focal p16 expression in trophoblastic lesions. Thus, when considering the differential diagnosis of cervical SCC and trophoblastic lesions, only strong diffuse p16 staining should be considered helpful.

Cytologic findings of epithelioid trophoblastic tumor of the uterus: a case report.

BACKGROUND: Epithelioid trophoblastic tumor (ETT) is a rare entity within trophoblastic tumors. It is difficult to recognize ETT because of its epithelioid appearance. CASE: A 35-year-old female, 5 years after pregnancy, experienced genital bleeding 2 months prior to consulting us. Preoperative laboratory data showed a slightly elevated serum level of human chorionic gonadotropin (hCG). A cytologic cervical smear revealed large, polygonal, atypical cells. These cells had mononucleate, ovoid, irregularly enlarged and hyperchromatic nuclei with 1 or 2 conspicuous nucleoli. The cytoplasm was thin and abundant, with a distinct cell membrane, and sometimes showed vacuolation. The patient was diagnosed with uterine cancer, and hysterectomy was performed. The tumor was present in the uterine corpus, measuring 3 x 2.5 x 2.5 cm. Histologically, it was composed of mainly mononuclear tumor cell nests resembling intermediate trophoblastic cells with zones of hyaline material. Immunohistochemically, the tumor was positive for cytokeratin and placental alkaline phosphatase but negative for hCG and human placental lactogen. The tumor was subsequently diagnosed as ETT. CONCLUSION: It was difficult to make a definitive diagnosis of ETT using only a cytologic specimen. The diagnosis of ETT is facilitated by a combination of cytologic, histopathologic and clinical findings.

Inhibiton of RET and JAK2 signals and upregulation of VEGFR3 phosphorylation in vitro by galectin-1 in trophoblast tumor cells BeWo.

BACKGROUND: Galectin-1 (gal-1), a member of the mammalian beta-galactoside-binding proteins, binds to cell surface glycoproteins (Mucin-1) on trophoblast cells. Although it has been demonstrated that gal-1 induces cell differentiation processes in these cells, no information on its signal transduction processes is available so far. As tyrosine phosphorylation is a major mechanism that controls multiple biological processes including cell differentiation, survival and proliferation, the aim of this study was to examine which human receptor tyrosine kinases (RTKs) were phosphorylated in trophoblast cells by gal-1. MATERIALS AND METHODS: BeWo choriocarcinoma cells were incubated for 24h in the absence (controls) and presence of 60microg/ml galectin-1. With the RayBio Human RTK Phosphorylation Antibody Array 1, the relative levels of phosphorylation of different human RTKs could be detected simultaneously. The signal intensities were compared and quantified with the Quantity One Version 4.5.2 program. Gal-1-treated and non-treated cells were incubated with antibodies against REarranged during Transfection (RET) and phosphorylated RET(Y905). Staining reaction was performed with the avidin-biotinylated peroxidase complex (ABC) reagent. RESULTS: We demonstrated that gal-1 inhibited RET and Janus Kinase 2 (JAK2) signals and upregulated Vascular endothelial growth factor receptor 3 (VEGFR3) signal in BeWo cells. We also showed the downregulation of phosphorylation on RET phosphotyrosine residue 905 in BeWo cells with phosphorylation specific antibodies and immunocytochemistry. CONCLUSION: Out of a number of 71 different RTKs, the stimulation of BeWo cells with gal-1 showed a significant alteration of signal intensity in only 3 RTKs: JAK2, RET and VEGFR3. Our data suggest that phosphorylation of these RTKs could be involved in cell differentiation processes that could be responsible for the already known effect of gal-1 on BeWo cells, the inhibition of proliferation.

Successful treatment of placental site trophoblastic tumor in twin pregnancy without hysterectomy.

To the best of the authors' knowledge, no case of placental site trophoblastic tumor (PSTT) pertinent to twin pregnancy has yet been published. There are only few case reports concerning patients with PSTT who were successfully treated by fertility-sparing methods. A 29-year-old nulliparous woman was admitted to hospital in the 36th week of a twin pregnancy due to symptoms of preterm labor. She underwent a cesarean section, during which a 4-cm uterine mass was found and resected. Histopathology revealed PSTT. Eighteen weeks after the delivery an ultrasound scan displayed another intrauterine mass, 2 cm in size. In the material resected in hysteroscopy there was necrotic decidual tissue. Another 30 months of observation revealed no abnormalities. Even though PSTT is rarely diagnosed, it may cause significant diagnostic and therapeutic problems.