OBJECTIVE: To investigate the expression of Talin1 in the fallopian tube and chorionic villi in patients with tubal pregnancy and its role in regulating invasion and migration of trophoblasts. METHODS: Immunohistochemistry and Western blotting were used to detect the localization and expression level of Talin1 in the fallopian tube and chorionic villi in patients with tubal pregnancy and in women with normal pregnancy. In the cell experiment, HTR-8/SVneo cells was transfected with Talin1 siRNA and the changes in cell invasion and migration were assessed using scratch assay and Transwell assay. The expressions of MMP-2, MMP-9, N-cadherin and Snail in the transfected cells were detected by qRT-PCR and Western blotting. RESULTS: Positive expression of Talin1 was detected in both normal fallopian tube tissues and tissues from women tubal pregnancy, and its expression was localized mainly in the cytoplasm of cilia cells. The expression level of Talin1 was significantly higher in both the fallopian tube and chorionic villi in women with tubal pregnancy than in normal fallopian tube and chorionic villi samples (P < 0.01). In HTR-8/SVneo cells, transfection with Talin1 siRNA significantly inhibited cell invasion (P < 0.01) and migration (P < 0.05), down-regulated the expression of N-cadherin, MMP-2 and Snail (P < 0.05), and up-regulated the expression of MMP-9 in the cells (P < 0.05). CONCLUSION The expression of Talin1 in the fallopian tube and chorionic villi is significantly increased in women with tubal pregnancy, suggesting the association of Talin1-regulated trophoblast cell invasion with the occurrence of tubal pregnancy.
Cadherins/metabolism* ; Cell Movement ; Chorionic Villi/metabolism* ; Fallopian Tubes/metabolism* ; Female ; Humans ; Matrix Metalloproteinase 2/metabolism* ; Matrix Metalloproteinase 9/metabolism* ; Pregnancy ; Pregnancy, Tubal/metabolism* ; RNA, Small Interfering/metabolism* ; Talin/metabolism* ; Trophoblasts/metabolism*

BACKGROUNDPlacental multidrug resistance-associated protein 2 (MRP2), encoded by ABCC2 gene in human, plays a significant role in regulating drugs' transplacental transfer rates. Studies on placental MRP2 regulation could provide more therapeutic targets for individualized and safe pharmacotherapy during pregnancy. Currently, the roles of epigenetic mechanisms in regulating placental drug transporters are still unclear. This study aimed to investigate the effect of histone deacetylases (HDACs) inhibition on MRP2 expression in the placental trophoblast cell line and to explore whether HDAC1/2/3 are preliminarily involved in this process.

METHODSThe human choriocarcinoma-derived trophoblast cell line (Bewo cells) was treated with the HDAC inhibitors-trichostatin A (TSA) at different concentration gradients of 0.5, 1.0, 3.0, and 5.0 μmol/L. Cells were harvested after 24 and 48 h treatment. Small interfering RNA (siRNA) specific for HDAC1/HDAC2/HDAC3 or control siRNA was transfected into cells. Total HDAC activity was detected by colorimetric assay kits. HDAC1/2/3/ABCC2 messenger RNA (mRNA) and protein expressions were determined by real-time quantitative polymerase chain reaction and Western-blot analysis, respectively. Immunofluorescence for MRP2 protein expression was visualized and assessed using an immunofluorescence microscopy and ImageJ software, respectively.

RESULTSTSA could inhibit total HDAC activity and HDAC1/2/3 expression in company with increase of MRP2 expression in Bewo cells. Reduction of HDAC1 protein level was noted after 24 h of TSA incubation at 1.0, 3.0, and 5.0 μmol/L (vs. vehicle group, all P < 0.001), accompanied with dose-dependent induction of MRP2 expression (P = 0.045 for 1.0 μmol/L, P = 0.001 for 3.0 μmol/L, and P < 0.001 for 5.0 μmol/L), whereas no significant differences in MRP2 expression were noted after HDAC2/3 silencing. Fluorescent micrograph images of MRP2 protein were expressed on the cell membrane. The fluorescent intensities of MRP2 in the control, HDAC2, and HDAC3 siRNA-transfected cells were week, and no significant differences were noticed among these three groups (all P > 0.05). However, MRP2 expression was remarkably elevated in HDAC1 siRNA-transfected cells, which displayed an almost 3.19-fold changes in comparison with the control siRNA-transfected cells (P < 0.001).

CONCLUSIONSHDACs inhibition could up-regulate placental MRP2 expression in vitro, and HDAC1 was probably to be involved in this process.

Cell Line ; Histone Deacetylase 1 ; metabolism ; Histone Deacetylase 2 ; metabolism ; Histone Deacetylase Inhibitors ; pharmacology ; Histone Deacetylases ; metabolism ; Humans ; Hydroxamic Acids ; pharmacology ; Microscopy, Fluorescence ; Multidrug Resistance-Associated Proteins ; genetics ; metabolism ; RNA, Messenger ; Trophoblasts ; cytology ; metabolism

Antigens, Neoplasm ; metabolism ; Antigens, Surface ; metabolism ; Colorectal Neoplasms ; immunology ; pathology ; Humans ; Prognosis ; Trophoblasts ; immunology

BACKGROUNDPreeclampsia is a multifactorial disease during pregnancy. Dysregulated lipid metabolism may be related to some preeclampsia. We investigated the relationship between triglycerides (TGs) and liver injury in different preeclampsia-like mouse models and their potential common pathways.

METHODSPreeclampsia-like models (Nw-nitro-L-arginine-methyl ester [L-NAME], lipopolysaccharide [LPS], apolipoprotein C-III [Apo] transgnic mice + L-NAME, β2 glycoprotein I [βGPI]) were used in four experimental groups: L-NAME (LN), LPS, Apo-LN and βGPI, respectively, and controls received saline (LN-C, LPS-C, Apo-C, βGPI-C). The first three models were established in preimplantation (PI), early-, mid- and late-gestation (EG, MG and LG). βGPI and controls were injected before implantation. Mean arterial pressure (MAP), 24-hour urine protein, placental and fetal weight, serum TGs, total cholesterol (TC) and pathologic liver and trophocyte changes were assessed.

RESULTSMAP and proteinuria were significantly increased in the experimental groups. Placenta and fetal weight in PI, EP and MP subgroups were significantly lower than LP. Serum TGs significantly increased in most groups but controls. TC was not different between experimental and control groups. Spotty hepatic cell necrosis was observed in PI, EG, MG in LN, Apo-LN and βGPI, but no morphologic changes were observed in the LPS group. Similar trophoblastic mitochondrial damage was observed in every experimental group.

CONCLUSIONSEarlier preeclampsia onset causes a higher MAP and urine protein level, and more severe placental and fetal damage. Preeclampsia-like models generated by varied means lead to different changes in lipid metabolism and associated with liver injury. Trophoblastic mitochondrial damage may be the common terminal pathway in different preeclampsia-like models.

Animals ; Cholesterol ; blood ; Disease Models, Animal ; Female ; Fetal Weight ; physiology ; Liver ; injuries ; Male ; Mice ; Mice, Inbred C57BL ; Mitochondrial Diseases ; blood ; pathology ; Placenta ; metabolism ; Pre-Eclampsia ; blood ; pathology ; Pregnancy ; Triglycerides ; blood ; Trophoblasts ; pathology

OBJECTIVETo investigate the clinical utility of short tandem repeats(STR) genotyping technique for diagnosis of partial hydatidiform moles (PHM).

METHODSTen cases with the original diagnosis of PHM and six cases diagnosed as "favour PHM" or "abnormal villous, PHM not excluded" were selected for the study. The clinical information and follow-up data were reviewed. Histopathologic features were evaluated along with p57 immunohistochemistry. After DNA extraction from each sample, genotyping was performed by AmpFlSTR(®) Identifiler™ PCR kit to amplify 15 STR polymorphism loci plus the amelogenin gender-determining in a single robust PCR.

RESULTSThe age of patients ranged from 18 to 49 years (mean=29 years, median=29 years). Two villous populations (7/16), irregular villous contour (13/16), at least moderate trophoblastic hyperplasia (2/16), cistern formation (8/16), syncytiotrophoblastic knuckles (14/16), trophoblastic pseudoinclusions (6/16) and nucleated fetal red blood cells (8/16) were presented in these cases. Of the cases in the study, STR genotyping identified 4 monospermic complete hydatidiform moles (MCM), 3 dispermic partial hydatidiform moles (DPM) and 9 hydropic abortions (HA). The misdiagnosis rate was 13/16 only relied on morphology evaluation. Immunostaining of p57 showed 3/4 of MCM were focally positive (<5%-20%+), 1/4 of MCM were diffusely positive (70%+), 3/3 of DPM were diffusely positive (≥50%+), 7/9 of HA were diffusely positive (≥50%+), and 2/9 of HA were focally positive (10%+).

CONCLUSIONSCombination of histomorphologic evaluation and p57 immunostaining is insufficient for a definitive diagnosis of PHM. STR genotyping offers an accurate diagnosis of PHM.

Abortion, Spontaneous ; Adolescent ; Adult ; Cyclin-Dependent Kinase Inhibitor p57 ; metabolism ; Female ; Genotype ; Genotyping Techniques ; Humans ; Hydatidiform Mole ; diagnosis ; genetics ; Immunohistochemistry ; Microsatellite Repeats ; Middle Aged ; Polymerase Chain Reaction ; Pregnancy ; Trophoblasts ; pathology ; Uterine Neoplasms ; diagnosis ; genetics ; Young Adult

The placenta is a temporary fetomaternal organ capable of supporting fetal growth and development during pregnancy. In particular, abnormal development and dysfunction of the placenta due to cha nges in the proliferation, differentiation, cell death, and invasion of trophoblasts induce several gynecological diseases as well as abnormal fetal development. Autophagy is a catalytic process that maintains cellular structures by recycling building blocks derived from damaged microorganelles or proteins resulting from digestion in lysosomes. Additionally, autophagy is necessary to maintain homeostasis during cellular growth, development, and differentiation, and to protect cells from nutritional deficiencies or factors related to metabolism inhibition. Induced autophagy by various environmental factors has a dual role: it facilitates cellular survival in normal conditions, but the cascade of cellular death is accelerated by over-activated autophagy. Therefore, cellular death by autophagy has been known as programmed cell death type II. Autophagy causes or inhibits cellular death via the other mechanism, apoptosis, which is programmed cell death type I. Recently, it has been reported that autophagy increases in placenta-related obstetrical diseases such as preeclampsia and intrauterine growth retardation, although the mechanisms are still unclear. In particular, abnormal autophagic mechanisms prevent trophoblast invasion and inhibit trophoblast functions. Therefore, the objectives of this review are to examine the characteristics and functions of autophagy and to investigate the role of autophagy in the placenta and the trophoblast as a regulator of cell death.
Apoptosis ; Autophagy* ; Cell Death* ; Cell Differentiation ; Cellular Structures ; Digestion ; Fetal Development ; Fetal Growth Retardation ; Homeostasis ; Lysosomes ; Malnutrition ; Metabolism ; Placenta* ; Pre-Eclampsia ; Pregnancy ; Recycling ; Trophoblasts

BACKGROUNDPreeclampsia (PE) is associated with abnormal fatty acid beta-oxidation (FAO), especially metabolic disorders of long-chain fatty acid oxidation. The role of FAO dysfunction in inadequate invasion is unclear. The aim of this study was to explore the influence of various lengths fatty acids oxidation on invasiveness of trophoblasts.

METHODSPrimary human trophoblast cells and HTR8/SVneo cells were treated with fatty acids of various lengths. Morphological changes, lipid deposition and ultrastructure changes of trophoblast cells were detected. Cells invasiveness was determined by transwell insert. CPT1, CPT2 and long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) protein expression were analyzed. The correlation between intracellular lipid droplets deposition and cells invasiveness was evaluated.

RESULTSCells treated with long-chain fatty acids showed significant increased lipid droplets deposition, severe mitochondrial damage, decreased CPT2 and LCHAD protein expression (P < 0.05) but no significant difference in CPT1 protein expression (P > 0.05). Invasiveness of the trophoblast cells of the LC-FFA group significantly decreased (P < 0.05). Intracellular lipid droplets deposition was negatively correlated with invasivenss (R = -0.745, P < 0.05).

CONCLUSIONTrophoblast cells after stimulation with long chain fatty acids exist fatty acid oxidation disorders, and reduce the ability of trophoblastic invasion.

Cell Line ; Cells, Cultured ; Fatty Acids, Nonesterified ; pharmacology ; Humans ; Lipid Metabolism ; Long-Chain-3-Hydroxyacyl-CoA Dehydrogenase ; metabolism ; Trophoblasts ; drug effects

This paper aimed to study the ability of baicalein to block human cytomegalovirus (HCMV) infection in extravillous cytotrophoblasts (EVT) and its effect on the vasoactive intestinal peptide (VIP) expression in HCMV-infected EVT in vitro. A human trophoblast cell line (HPT-8) was chosen in this study. HCMV with 100 TCID50 was added into culture medium to infect HPT-8 cells, and then HCMV pp65 antigen was assayed by immunofluorescence staining. The infection status was determined by virus titration. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect virus DNA load in the infected cells. The expression of VIP mRNA and protein in the infected cells was measured by qRT-PCR, immunocytochemistry and Western blotting. Concentration of VIP secreted in supernatants was determined by ELISA. Red-stained HCMV pp65 antigens were found in infected HPT-8 cells 48 h after infection. HCMV replicated in large quantity in infected HPT-8 cells 4 days after infection, reaching a peak at day 6 post-infection. After treatment with baicalein, virus DNA load in infected HPT-8 cells was decreased (P<0.05), and the levels of VIP mRNA and protein, and the concentration were raised to the normal (P>0.05). Our study suggested that baicalein exerts a positive effect on the VIP expression in HCMV-infected EVT at maternal-fetal interface.
Antiviral Agents ; pharmacology ; Cell Line ; Cytomegalovirus ; drug effects ; physiology ; Flavanones ; pharmacology ; Humans ; Trophoblasts ; cytology ; drug effects ; metabolism ; virology ; Vasoactive Intestinal Peptide ; metabolism ; Virus Inactivation ; drug effects

OBJECTIVETo study the expression of Toll-like receptors (TLRs) mRNA in human trophoblast HTR-8/SVneo cells and the changes in indoleamine 2,3-dioxygenase (IDO) mRNA expression in response to TLR ligand stimulation.

METHODSThe expressions of TLRs and IDO mRNA in human HTR-8/SVneo cells were tested by RT-PCR, and the changes in IDO mRNA levels after exposure to TLR3, TLR4, TLR7/8, and TLR9 ligands were quantitatively analyzed with real-time PCR.

RESULTSIDO and TLR1-10 mRNAs were expressed in HTR-8/SVneo cells. As the cell culture time extended, IDO mRNA expression level tended to increase within 48 h. After stimulation with the TLR ligands, the expression of TLR-3 mRNA was down-regulated while the expression of TLR-4, 7, 8, and 9 mRNA up-regulated. Stimulation of the cells with poly(I:C) lowered the expression of IDO mRNA while IFN-γ increased its expression.

CONCLUSIONSThe expression of IDO mRNA is associated with the nutrition of the maternal-fetal interface. Stimulation with the TLR ligands affects the expression of IDO and TLR mRNA expressions in the cells, which verifies the functional activity of TLRs and suggests a role of IDO in TLR pathway-dependent antiviral immunity.

Cell Line ; Female ; Humans ; Indoleamine-Pyrrole 2,3,-Dioxygenase ; genetics ; metabolism ; Interferon-gamma ; pharmacology ; Ligands ; Poly I-C ; pharmacology ; RNA, Messenger ; metabolism ; Toll-Like Receptors ; genetics ; metabolism ; Trophoblasts ; cytology ; metabolism

The present study aims to investigate the effects of soluble endoglin (sEng) on invasive ability of cultured cytotrophoblasts of first trimester of pregnancy. Cytotrophoblasts of normal 6 to 8-week pregnancy were cultured by trypsin digestion method, and were incubated with cell culture medium without (control group) and with 10 μg/L sEng (sEng group), respectively for 24 h. The invasive ability was determined by transwell invasion assay, and expressions of MMP-2, MMP-9 mRNA and protein were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot, respectively. The results showed that the invasive ability of cytotrophoblasts in sEng group was lower than that in control group (P < 0.05). Compared with control group, the expressions of MMP-2 and MMP-9 mRNA and protein of cytotrophoblasts were significantly lower (P < 0.05). In conclusion, sEng may participate in the genesis of preeclampsia by affecting the invasive ability of cytotrophoblasts through regulation of the expression of MMP-2 and MMP-9.
Antigens, CD ; pharmacology ; Cell Movement ; drug effects ; physiology ; Cells, Cultured ; Endoglin ; Female ; Humans ; Matrix Metalloproteinase 2 ; genetics ; metabolism ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Placentation ; physiology ; Pre-Eclampsia ; physiopathology ; Pregnancy ; Pregnancy Trimester, First ; RNA, Messenger ; genetics ; metabolism ; Receptors, Cell Surface ; Trophoblasts ; cytology