ObjectiveCinobufotalin (CINO), a cardiotonic steroid (CTS) or bufadienolide, is extracted from the skin secretions of giant toads and is utilized in traditional Chinese medicine (Chan Su). CINO has been used as a cardiotonic, diuretic and a hemostatic agent. Our lab is familiar with CINO and has shown it to inhibit cytotrophoblast cells function. Recently, it has been shown that CINO also inhibits the lung cancer cell function, and has been further implicated in other disease processes. In the present study, we propose to pursue this potential application of CINO using ovarian tumor cell line SK-OV-3.Study DesignWe evaluated the in-vitro effect of CINO on ovarian cancer cell line SK-OV-3. Cells were treated with 0.1, 1, 5, and 10 µM CINO. Cell proliferation was measured using a CellTiter Assay (Promega), which is a colorimetric method for determining the number of viable cells. Cell migration was measured using a CytoSelect Assay (Cell Biolabs). Cell invasion was measured using a FluoroBlok Assay (BD). Cell viability was measure using a CellTiter Assay (Promega). Cell cycle progression was evaluated by a Cell Cycle Phase Determination Kit (Cayman Chemical) and apoptosis was evaluated by an Apoptotic Blebs Assay Kit (Cayman Chemical). Cell cycle arrest and apoptotic signaling was determined by fluorescence-activated cell sorting (FACS) analysis.ResultsCINO at ≥0.5 µM inhibited SKOV-3 cell proliferation, migration, and invasion (p<0.05). There was a higher (p<0.05) percentage of S phase cells in groups treated with CINO at 0.5 µM. CINO at ≥0.5 µM down regulated expression of PCNA and caused cell death.ConclusionThis data demonstrates that CINO impairs SK-OV-3 cell function via cell cycle arrest and apoptotic signaling. These findings demonstrate the complex nature of this compound. Not only is CINO directly modulating the actions of the Na/K ATPase through classic mechanism of cardiotonic steroids, but is also directly influencing the nuclear expression of proteins involved in cell cycle progression and DNA repair. Additional investigational studies looking into the molecular pathways involved in altering cell cycle and entry into apoptosis are warranted.In conclusion, we have shown CINO to impair SK-OV3 cell function via cell cycle arrest and apoptotic signaling and suggest that CINO might be further investigated as a novel anti-ovarian cancer agent.

ObjectivePreeclampsia (PreE) is one of considerable public health threat particularly in developing countries globally affecting approximately 8% of all pregnancies. PreE is a pregnancy-specific condition that increases maternal and infant mortality and morbidity, but the etiology remains unknown. Despite numerous basic, clinical, and epidemiologic studies that have been conducted over the past half century, knowledge of the etiology and pathogenesis of preeclampsia remains elusive. It is diagnosed by new onset increased blood pressure and proteinuria during second or third trimester of gestation; key features of the preeclampsia category include a cut-off blood pressure of 140/90 mm Hg or higher and absolute requirement of proteinuria. Approximately 20% of the diabetic pregnant women develop preE. The mechanisms contributing to this effect is not well characterized. In a recent study, we have shown that hyperglycemia impairs cytotrophoblast (CTB) function via stress signaling. Several researchers demonstrate a direct link between preE and diabetes. The objective of the study was to evaluate potential linkage between the risk of developing preE and the presence of diabetes in pregnant patients in Bangladesh.MethodsThis is a cross-sectional study of 351 pregnant women performed to evaluate the prevalence of PreE with respect to different risk factors such as previous pregnancy, presence of Antiphospholipid antibodies, pre-existing diabetes (before this pregnancy), multiple gestation / singleton, family history of preE in first degree relative (mother, sister and daughters; most commonly mother), maternal age of 40 or greater. The study was conducted in selected hospitals of Dhaka city, Bangladesh during December 2013 to December 2015.ResultsThe key study findings revealed that the overall rate of development of PreE in Bangladeshi pre-gestational diabetic patients is 22.6 percent. We gave special emphasis on the occurrence of PreE in pre-gestational diabetic patients. Among 351 respondents, 145 Patients (25.5%) with either DM prior to pregnancy or developing gestational diabetes or without diabetes were older (age >35 years) pregnant women (13.1%). Prevalence of PreE is 25.5 (n=145) and 19.6 (n=199) percent among those who developed pre-gestational diabetes and without diabetes prior to pregnancy. Of the respondents 17.2 percent have both systolic and diastolic hypertension those who developed DM prior to this pregnancy and 13 percent among those who does not developed DM prior to this pregnancy. Occurrence of abortion was up to 3 percentage before this pregnancy 45 percentage who developed DM prior to this pregnancy and 35 percent who does not have diabetes among the respondents.ConclusionsThere is an association has been found between the risk of developing preE and the presence of diabetes in pregnant patients in Bangladesh.

ObjectiveCinobufotalin (CINO), a cardiotonic steroid (CTS) or bufadienolide, is extracted from the skin secretions of the traditional Chinese medicine giant toads (Chan su). CINO has been used as a cardiotonic, diuretic and a hemostatic agent. Previously we have shown that CINO inhibits the cytotrophoblast cell function. Recently other study has shown that CINO inhibits A549, a lung cancer cell function. In this study, we assessed the effect of CINO on three different ovarian cancer cell lines; SK-OV-3, CRL-1978 and CRL-11731 to confirm whether the effect of CINO is cell specific.Study DesignWe evaluated the effect of CINO on three ovarian cancer cells SK-OV-3, CRL-1978, and CRL-11731 function in vitro. Each Cell lines were treated with different concentrations of CINO (0.1, 1, 5 and 10 µM). For each cell line cell proliferation, migration and invasion were measured by using a CellTiter Assay (Promega), Cytoselect Assay (Cell Biolabs) and by using a FluoroBlock Assay (BD) respectively. Proliferating Cell Nuclear Antigen (PCNA) was also evaluated in cell lysates of CINO treated these 3 ovarian cancer cells by western blot analysis. Cell Cycle arrest and Cell viability were determined by fluorescence-activated cell sorting (FACS) analysis. We also performed Annexin V staining on CINO treated these 3 ovarian cancer cell lines by immunofluorescence to evaluate the pro-apoptotic protein expression. In addition mitochondrial membrane potential has also been measured for all these 3 ovarian cell lines after CINO treatment using MMP kit, by FACS analysis.ResultsConcentration of CINO at 0.5 µM inhibit SK-OV-3, CRL-1978, and CRL-11731 ovarian cancer cells proliferation, migration and invasion without cell death and loss of cell viability but cell viability differs for each cell line. Each cell lines differ in response to CINO doses for PCNA expression as well as Annexin V pro-apoptotic protein expression. CINO decreases mitochondrial membrane potential for SK-OV-3 but for CRL-1978 and CRL-11731 increases in response to CINO treatment.ConclusionCINO is cell specific, as each cancer cell line responds differently. These data demonstrate that the mode of action of CINO is different on these 3 types of ovarian cancer cells.