Elevated histone demethylase KDM5C increases recurrent miscarriage risk by preventing trophoblast proliferation and invasion.

KDM5C is a histone H3K4-specific demethylase, which has been shown to play a key role in biological disease and development. However, the role of KDM5C in trophoblasts at early pregnancy is currently unknown. Here, we showed that KDM5C was upregulated in placental trophoblasts from recurrent miscarriage (RM) patients compared with healthy controls (HCs). Trophoblast proliferation and invasion was inhibited by KDM5C overexpression and was promoted by KDM5C knockdown. Transcriptome sequencing revealed that elevated KDM5C exerted anti-proliferation and anti-invasion effects by repressing the expression of essential regulatory genes. The combination analysis of RNA-seq, ChIP-seq and CUT&Tag assay showed that KDM5C overexpression leads to the reduction of H3K4me3 on the promoters and the corresponding downregulation of expression of several regulatory genes in trophoblasts. Among these genes, TGFß2 and RAGE are essential for the proliferation and invasion of trophoblasts. Importantly, overexpression of KDM5C by a systemically delivered KDM5C adenovirus vector (Ad-KDM5C) promoted embryo resorption rate in mouse. Our results support that KDM5C is an important regulator of the trophoblast function during early pregnancy, and suggesting that KDM5C activity could be responsible for epigenetic alterations seen RM disease.

Fabp4 contributes toward regulating inflammatory gene expression and oxidative stress in Ctenopharyngodon idella.

Fatty acid-binding protein (Fabp)-4 is a member of the FABP family. Mammalian fabp4 has been demonstrated to involve in inflammation and immunity, whereas the related data of fish fabp4 remain limited. Therefore, we further investigated the effects of fabp4 on immunity in Ctenopharyngodon idella. The fabp4 sequence spanned 405 bp was cloned first, sharing high identity to fabp4 from other fish and mammals. Fabp4 expression was the highest in the adipose tissue, followed by the heart, muscle, and liver. In vivo, lipopolysaccharide (LPS) triggered the expression of fabp4, toll-like receptor (tlr)-22, interleukin (il)-1ß, and tumor necrosis factor (tnf)-α in the kidney and spleen. In vitro, exposing C. idella CIK cells to LPS decreased their viability, and the expression of fabp4 was also increased by LPS. However, BMS309403, an inhibitor of FABP4, mitigated these effects. Furthermore, treating the cells with LPS or fabp4 overexpression plasmids resulted in reactive oxygen species (ROS) generation and upregulation of inflammatory genes expression, including tlr22, type-I interferon (ifn-1), interferon regulatory factor (irf)-7, tnfα, il-1ß, and interferon-ß promoter stimulator 1. These effects were ameliorated by preincubation with BMS309403. Moreover, incubating the cells with glutathione reduced the production of ROS and the expression of inflammatory genes that were evoked by LPS and plasmid treatments. These results showed that fabp4 acts as a pro-inflammatory molecule via elevating ROS levels, providing a novel understanding of the molecular regulation of innate immunity in teleosts.

Next-Generation Sequencing Is More Efficient at Detecting Mosaic Embryos and Improving Pregnancy Outcomes than Single-Nucleotide Polymorphism Array Analysis.

We compared chromosomal mosaicism, detected by next-generation sequencing (NGS), during preimplantation genetic testing (PGT) with that detected by single-nucleotide polymorphism (SNP) array-based PGT to assess the pregnancy outcomes associated with both platforms in a retrospective cohort study of patients undergoing in vitro fertilization in a single university-based assisted reproduction center. In total, 6427 blastocysts biopsied from 1513 patients who underwent 2833 oocyte retrievals from January 2017 to February 2019 were identified. The incidence of mosaicism was significantly higher in the NGS-based PGT group than in the SNP array-based PGT group. Furthermore, some aneuploid specimens were affected by mosaicism. The total mosaicism detection rate with NGS-based PGT (23.3%) was significantly higher than that with SNP array-based PGT (7.7%). Mosaicism rates were similar when stratified by maternal age or PGT type. The SNP array cohort showed a significantly higher spontaneous abortion rate than the NGS cohort (10.07% versus 6.33%; P = 0.0403). The ongoing pregnancy/live birth rate was higher in the NGS cohort (44.1%) than in the SNP array cohort (42.28%). Our results confirm that NGS-based PGT can detect mosaicism more frequently than SNP array-based PGT in trophectoderm specimens. Therefore, clinical application of NGS for PGT may improve pregnancy outcomes compared with that of SNP array-based PGT. More detailed blastocyst detection and classification is necessary to prioritize embryo transfers.

Characterization of Copy-Number Variations and Possible Candidate Genes in Recurrent Pregnancy Losses.

It is well established that embryonic chromosomal abnormalities (both in the number of chromosomes and the structure) account for 50% of early pregnancy losses. However, little is known regarding the potential differences in the incidence and distribution of chromosomal abnormalities between patients with sporadic abortion (SA) and recurrent pregnancy loss (RPL), let alone the role of submicroscopic copy-number variations (CNVs) in these cases. The aim of the present study was to systematically evaluate the role of embryonic chromosomal abnormalities and CNVs in the etiology of RPL compared with SA. Over a 3-year period, 1556 fresh products of conception (POCs) from miscarriage specimens were investigated using single nucleotide polymorphism array (SNP-array) and CNV sequencing (CNV-seq) in this study, along with further functional enrichment analysis. Chromosomal abnormalities were identified in 57.52% (895/1556) of all cases. Comparisons of the incidence and distributions of chromosomal abnormalities within the SA group and RPL group and within the different age groups were performed. Moreover, 346 CNVs in 173 cases were identified, including 272 duplications, 2 deletions and 72 duplications along with deletions. Duplications in 16q24.3 and 16p13.3 were significantly more frequent in RPL cases, and thereby considered to be associated with RPL. There were 213 genes and 131 signaling pathways identified as potential RPL candidate genes and signaling pathways, respectively, which were centered primarily on six functional categories. The results of the present study may improve our understanding of the etiologies of RPL and assist in the establishment of a population-based diagnostic panel of genetic markers for screening RPL amongst Chinese women.

Transcriptome analysis of golden pompano (Trachinotus ovatus) liver indicates a potential regulatory target involved in HUFA uptake and deposition.

Promoting highly unsaturated fatty acid (HUFA) uptake and deposition can improve nutritional value of farmed fish and reduce dietary fish oil addition. Previously, we found that the golden pompano Trachinotus ovatus liver HUFA content increased with the increasing of dietary HUFA. Therefore, we examined the common genes and pathways responsible for HUFA uptake and deposition in T. ovatus liver using transcriptome sequencing technology after feeding with either 1.0% or 2.1% HUFA for 8 weeks. Results showed that a total of 140 and 147 genes were significantly upregulated and downregulated, respectively. Five bile acid synthesis-related genes (CYP7A1, CYP8B1, AKR1D1, SCP2 and ACOT8), which are related to dietary fat emulsification were downregulated in 2.1% HUFA group, implying that the cholate synthesized through the classical pathway might be the main bile acid form in fat emulsification. Moreover, fatty acid transport protein (FATP)-6, fatty acid binding protein (FABP)-1, -4, and -6 increased with HUFA deposition, especially FATP6 and FABP4, suggesting that the two genes may be important mediators involved in HUFA uptake and deposition. KEGG analysis showed that most of the differential genes described above were involved in peroxisome proliferator activator receptor (PPAR) signaling pathway, and PPARγ increased with HUFA deposition, indicating that PPARγ might be a key regulator of HUFA uptake and deposition by regulating the genes involved in fatty acid emulsification and transport. This study focused on the liver, which is the center of intermediary metabolism, providing a comprehensive understanding of the molecular regulation of HUFA uptake and deposition in T. ovatus, which should be further investigated to develop potential measures to improve HUFA content.

Dietary arachidonic acid decreases the expression of transcripts related to adipocyte development and chronic inflammation in the adipose tissue of juvenile grass carp, Ctenopharyngodon idella.

Overdevelopment of adipose tissue in cultured fish is one of the biggest issues plaguing current aquaculture industry, leading to unhealthy status of fishes and production losses. Diet supplemented with 0.30% arachidonic acid (ARA) has been found to reduce adipogenesis and inflammation in grass carp, but the potential mechanism is not comprehensively understood. To fully reveal the effects of dietary ARA on the mRNA profiles of adipose tissue, transcriptome techniques were applied in this study. A 10-weeks feeding experiment was performed using two isonitrogenous and isoenergetic purified diets, namely ARA-free (control) and 0.30% ARA (ARA group). Results showed increased ARA content and decreased intraperitoneal fat index and adipocyte size in the adipose tissue of fish fed ARA (P < 0.05). A total of 611 and 973 genes of the adipose tissue were significantly up-regulated and down-regulated, respectively, in fish fed ARA (P < 0.05). Dietary ARA upregulated LOX pathway but downregulated CYP450 pathway annotated genes expression. A total of 65 cell development annotated genes including 30 adipocyte proliferation, 21 adipocyte differentiation, and 14 cell apoptosis annotated genes were down-regulated in the ARA group. In addition, 19 lipid catabolism annotated genes were increased. The mRNA expression levels of 5 chemokines, 10 cytokines, 26 cytokine and chemokine receptors, 15 cell adhesion, 6 oxidative stress, and 6 angiogenesis annotated genes were all down-regulated in fish fed ARA. Finally, dietary ARA also decreased the expression of transcripts annotated with glucose transportation, glycolysis and gluconeogenesis. Overall, our results demonstrate that dietary ARA has a fat reducing role, and tends to retard adipocyte development and attenuate chronic inflammation based on these adipose transcript expression results in grass carp.

Lipid droplets participate in modulating innate immune genes in Ctenopharyngodon idella kidney cells.

Lipid droplets (LDs) are increasingly being recognized as important immune modulators in mammals, in additional to their function of lipid ester deposition. However, the role of LDs in fish immunity remains poorly understood. In this study, the function of LDs in the innate immune response of Ctenopharyngodon idella kidney (CIK) cells, which are the equivalent of myeloid cells in vertebrates, was investigated. LD number and TG content significantly increased in the CIK cells following exposure to lipopolysaccharide (LPS), peptidoglycan (PGN), and polyriboinosinic-polyribocytidylic acid (Poly [I: C]) for 24 h, accompanied by increases in the relative expression of several innate immune genes. However, fatty acid compositions of the triglycerides were not changed after treatment with these three pathogenic mimics. LPS, PGN, and Poly (I: C) did not alter the relative expressions of lipogenic (FAS, SCD, and DGAT) and lipid catabolic (PPARα, ATGL, and CPT-1) genes. However, these treatments did increase the mRNA levels of lipid transportation genes (FATP/CD36, ACSL1, and ACSL4), and also decreased the non-esterified fatty acid level in the medium. To further explore the role of LDs in the immune response, CIK cells were incubated with different concentrations (0, 100, 200, 300, 400, 500 µM) of exogenous lipid mix (LM; oleic acid [OA]:linoleic acid [LA]:linolenic acid [LNA] = 2:1:1), and were then transferred to a lipid-free medium and incubated for 24 h. LD size and number increased with the increase in lipid levels, and this was accompanied by increased expression of innate immune genes, including MyD88, IRF3, and IL-1ß, which were expressed at their highest levels in 300 µM exogenous lipid mix. Interestingly, after incubating with different fatty acids (LM, OA, LA, LNA, arachidonic acid [ARA], and docosahexaenoic acid [DHA]; 300 µM), ARA and DHA were more potent in inducing LD formation and innate immune gene expression in the CIK cells. Finally, atglistatin, an ATGL inhibitor, effectively attenuated the expression of most genes upregulated by ARA or DHA, suggesting that lipolysis may be involved in the regulation of immune genes at the transcriptional level. Overall, the findings of this study demonstrate that LDs are functional organelles that could act as modulators in the innate immune response of CIK cells. Additionally, long-chain polyunsaturated fatty acid enriched LDs play a unique role in regulating this process.

Dietary docosahexaenoic acid decreased lipid accumulation via inducing adipocytes apoptosis of grass carp, Ctenopharygodon idella.

The purpose of this study was to explore the mechanism of by which docosahexaenoic acid (DHA) inhibit the accumulation of adipose tissue lipid in grass carp (Ctenopharyngodon idella). We therefore designed two semi-purified diets, namely DHA-free (control) and DHA-supplemented, and fed them to grass carp (22.19 ± 1.76 g) for 3 and 6 weeks. DHA supplementation led to a significantly lower intraperitoneal fat index (IPFI) than that in the control group by reducing the number of adipocytes but significantly higher adipocyte size (P < 0.05). In the intraperitoneal adipose tissue, the DHA-fed group showed significantly higher peroxisome proliferator-activated receptor (PPAR)γ, CCAAT enhancer-binding protein (C/EBP)α, and sterol regulatory element-binding protein (SREBP)1c mRNA expression levels at both 3 and 6 weeks (P < 0.05). However, the ratio of the expression levels of B cell leukemia 2 (Bcl-2) and Bcl-2-associated X protein (Bax) was significantly lower in the DHA-fed group than in the control group (P < 0.05), and the protein expression levels of the apoptosis-related proteins caspase 3, caspase 8, and caspase 9 were also significantly higher (P < 0.05). Overall, although DHA promotes lipid synthesis, it is more likely that DHA could suppress the lipid accumulation in adipocytes of grass carp by inducing adipocyte apoptosis.

Comparative analysis of effects of dietary arachidonic acid and EPA on growth, tissue fatty acid composition, antioxidant response and lipid metabolism in juvenile grass carp, Ctenopharyngodon idellus.

Four isonitrogenous and isoenergetic purified diets containing free arachidonic acid (ARA) or EPA (control group), 0·30 % ARA, 0·30 % EPA and 0·30 % ARA+EPA (equivalent) were designed to feed juvenile grass carp (10·21 (sd 0·10) g) for 10 weeks. Only the EPA group presented better growth performance compared with the control group (P<0·05). Dietary ARA and EPA were incorporated into polar lipids more than non-polar lipids in hepatopancreas but not intraperitoneal fat (IPF) tissue. Fish fed ARA and EPA showed an increase of serum superoxide dismutase and catalase activities, and decrease of glutathione peroxidase activity and malondialdehyde contents (P<0·05). The hepatopancreatic TAG levels decreased both in ARA and EPA groups (P<0·05), accompanied by the decrease of lipoprotein lipase (LPL) activity in the ARA group (P<0·05). Fatty acid synthase (FAS), diacylglycerol O-acyltransferase and apoE gene expression in the hepatopancreas decreased in fish fed ARA and EPA, but only the ARA group exhibited increased mRNA level of adipose TAG lipase (ATGL) (P<0·05). Decreased IPF index and adipocyte sizes were found in the ARA group (P<0·05). Meanwhile, the ARA group showed decreased expression levels of adipogenic genes CCAAT enhancer-binding protein α, LPL and FAS, and increased levels of the lipid catabolic genes PPAR α, ATGL, hormone-sensitive lipase and carnitine palmitoyltransferase 1 (CPT-1) in IPF, whereas the EPA group only increased PPAR α and CPT-1 mRNA expression and showed less levels than the ARA group. Overall, dietary EPA is beneficial to the growth performance, whereas ARA is more potent in inducing lipolysis and inhibiting adipogenesis, especially in IPF. Meanwhile, dietary ARA and EPA showed the similar preference in esterification and the improvement in antioxidant response.

Role of cyclooxygenase-mediated metabolites in lipid metabolism and expression of some immune-related genes in juvenile grass carp (Ctenopharyngodon idellus) fed arachidonic acid.

Cyclooxygenase (COX) catalyzes the conversion of arachidonic acid (ARA) to prostaglandins, and COX-mediated metabolites play important roles in the regulation of lipid metabolism and immunity in mammals. However, such roles of COX in fish remain largely unknown. In this study, we designed three semi-purified diets, namely ARA-free (control), ARA, and ARA + acetylsalicylic acid (ASA; a COX inhibitor), and used them to feed grass carp (27.65 ± 3.05 g) for 8 weeks. The results showed that dietary ARA significantly increased the amount of ARA in the hepatopancreas, muscle, and kidney (P < 0.05), whereas this increase was reduced by dietary ASA. The hepatopancreatic prostaglandin E2 content increased in the ARA group, and this increase was inhibited by ASA (P < 0.05). ARA decreased the lipid content in the hepatopancreas, whereas ASA recovered lipid content to a significant level (P < 0.05). ARA significantly decreased the messenger RNA (mRNA) expression levels of fatty acid synthase and stearoyl-CoA desaturase in the hepatopancreas (P < 0.05). However, ASA did not rescue the mRNA expression of these genes (P > 0.05). Interestingly, ARA significantly enhanced the level of peroxisome proliferator-activated receptor α gene expression, and this increase was attenuated by ASA (P < 0.05). Finally, ARA significantly enhanced the mRNA expression of myeloid differentiation factor 88 (MyD88) in the kidney, and ASA attenuated the expression of toll-like receptor 22 and MyD88 (P < 0.05). In conclusion, our findings suggest that COX metabolites play important roles in the inhibition of lipid accumulation in the hepatopancreas of grass carp fed with ARA and that regulation of gene expression promotes lipid catabolism rather than lipogenic activities. Additionally, these eicosanoids might participate in the upregulation of immunity-related genes in the kidney.

Dietary Arachidonic Acid Has a Time-Dependent Differential Impact on Adipogenesis Modulated via COX and LOX Pathways in Grass Carp Ctenopharyngodon idellus.

In this study, we explored the function of arachidonic acid (ARA) in adipogenesis in the grass carp (Ctenopharyngodon idellus) using in vivo and in vitro models. An 8-week feeding trial was performed using three isonitrogenous and isoenergetic purified diets: ARA-free, ARA, and ARA + acetylsalicylic acid [ASA, a cyclooxygenase (COX) inhibitor]. Fish were sampled after 4 and 8 weeks of feeding. Results showed that ARA-fed fish had a significantly lower intraperitoneal fat index (IPFI) and smaller adipocytes; these decreases were reversed by ASA after 8 weeks of feeding. Nevertheless, at week 4, the IPFI and adipocyte size were higher in the ARA group, and they were comparable to those of fish fed ARA + ASA. To further investigate the influence of ARA on adipocyte differentiation, confluent pre-adipocytes of grass carp were incubated with ARA for 3 days. This in vitro experiment demonstrated that ARA promoted adipogenesis in a dose-dependent manner. Pre-treatment with the lipoxygenase (LOX) inhibitor nordihydroguaiaretic acid attenuated the pro-adipogenic function of ARA. However, after treatment with ARA for 8 days, adipocytes had a lower lipid content than cells treated with oleic acid, and ASA could suppress this effect. We thus revealed the dual function of ARA in adipogenesis in grass carp. The LOX pathway may play a key role in pro-adipogenesis after short-term treatment with ARA, whereas the COX pathway is possibly responsible for the inhibition of adipogenesis after long-term treatment.

Comparative analysis of the hepatopancreas transcriptome of grass carp (Ctenopharyngodon idellus) fed with lard oil and fish oil diets.

n-3 highly unsaturated fatty acids (n-3 HUFAs) have been shown to suppress lipid accumulation and improve protein utilization in grass carp; however, little is known about the underlying molecular mechanism. Hence, we analyzed the hepatopancreas transcriptome of grass carp (Ctenopharyngodon idellus) fed either lard oil (LO) or fish oil (FO) diets. RNA-seq data showed that 125 genes were significantly up-regulated and 107 were significantly down-regulated in the FO group. Among them, 17 lipid metabolism related genes, 12 carbohydrate metabolism related genes, and 34 protein metabolism related genes were selected. Lipid metabolism related genes, such as very long-chain acyl-CoA synthetase (ACSVL),carnitine O-palmitoyltransferase 1 (CPT1) and carnitine-acylcarnitine translocase (CACT), were up-regulated in the FO group. But the genes of diacylglycerol O-acyltransferase 2 (DGAT2) and stearoyl-CoA desaturase (SCD) were down-regulated. Down-regulation of glycolysis related genes, such as 6-phosphofructokinase (PFK), phosphoglycerate kinase (PGK) and pyruvate dehydrogenase kinase (PDK), added with up-regulation of gluconeogenesis related genes, such as phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase), suggests lower utilization of carbohydrate of the FO group. Besides, dietary FO also influenced the protein metabolism related genes, such as up-regulation of genes involved in digestion of dietary protein, mRNA transcription, protein translation and amino acid utilization, down-regulation of genes involved in mRNA degradation and ubiquitination of protein. Interestingly, the up-regulation of mitochondrial uncoupling protein 2 (UCP2) and down-regulation of oxidative phosphorylation related genes (cytochrome c oxidase subunit 4 isoform 2 [COX4I2], HIG1 domain family member 1A [HIGD1A] and cytochrome-b5 reductase [CYB5R]) suggest that energy metabolism may be also influenced by dietary fatty acid composition. These findings presented here provide a comprehensive understanding of the molecular mechanisms governing the effects of fish oil in grass carp.

Human chorionic gonadotropin (hCG) regulation of galectin-3 expression in endometrial epithelial cells and endometrial stromal cells.

Previous investigations on galectin-3 (gal-3) have focused mainly on its role in some malignant tumors. It was believed that gal-3 plays important roles in cell proliferation, apoptosis and adhesion in many cell types. Recently, gal-3 has been recognized as a factor related to endometrial receptivity in the human endometrium and trophoblast during embryo implantation. Human chorionic gonadotropin (hCG) is a specific embryonic hormone providing a signal from the embryo involved in preparing the receptive endometrium for embryo implantation. The current study aimed to determine whether hCG regulates gal-3 expression in endometrial cells. Our results showed that expression of gal-3 in both endometrial epithelial cells (EECs) and endometrial stromal cells (ESCs) could be regulated by hCG in an intricate manner. These results indicate that gal-3 might be regulated by hCG in preparing the endometrium for embryonic implantation.

[Karyotype analysis of amniotic fluid cells and comparison of chromosomal abnormality rate during second trimester].

OBJECTIVE: To investigate the karyotypes of amniotic fluid cells and compare the incidence of chromosomal abnormality as well as to evaluate the clinical significance of abnormal karyotypes. METHODS: A total of 13 648 pregnant women came to Shanghai Jiai Genetics and IVF Institute, Obstetrics and Gynecology Hospital, Fudan University to do amniocentesis from September 1998 to November 2010, and 13 795 amniotic fluid specimens were successfully extracted and cultured, thus 13 795 fetuses received karyotype diagnosis. These fetuses were grouped according to different indications. If maternal age was ≥ 35, the fetuses were grouped into the advanced maternal age group (4065); and if maternal serum screening test revealed high-risk of trisomy 18 or trisomy 21, the fetuses were grouped into the high-risk serum screening group (6462); and those with abnormal signs of ultrasound screening were grouped into the abnormal ultrasound signs group (1539); and if either of the parents was with chromosome abnormalities, the fetus was grouped into the paternal/maternal abnormality group (108); whereas the remainder were grouped in other factors group (1621). The amniotic fluid cells were in-situ cultured on coverslips, harvested by conventional G-banded methods, and then analyzed by two doctors. In order to get rapid diagnosis, some pregnant women whose gestational age ≥ 26 weeks accepted fluorescense in situ hybridization (FISH). FISH was done on 78 uncultured amniotic fluid specimens using probes located at chromosome 13, 18, 21, X, Y. Some parents were required to analyze lymphocyte karyotype to help judging the origin of abnormal karyotype. RESULTS: (1) Classification and composition of abnormal karyotypes in each group: a total of 388 abnormal karyotypes were found among 13 795 fetuses, and the abnormal rate was 2.813% (388/13 795). Of the 388 fetuses, aneuploidy was the most common pattern which was up to 59.8% (232/388); autosomal structural abnormality rate was 24.7% (96/388); mosaicism was 12.4% (48/388). Other uncommon abnormal karyotypes included marker chromosome (5/388, 1.3%), sex chromosomal structural abnormality (4/388, 1.0%) and triploid (3/388, 0.8%). Aneuploidy was the most common in most groups except the paternal/maternal abnormality group. There were four cases of rare aneuploid in the advanced maternal age group, the high-risk serum screening group and the abnormal ultrasound signs group respectively. Every type of abnormality could be found in the abnormal ultrasound signs group, and autosomal structural abnormalities were concentrated in paternal/maternal abnormality group. Mosaicism mainly distributed in the high-risk serum screening group, accounting for 20.0% (29/145) of abnormalities in this group. (2) Abnormal types and the incidence: the most common type was trisomy 21 (138/388, 35.6%), followed by autosomal balanced structural rearrangements (80/388, 20.6%), mosaicism (48/388, 12.4%) and trisomy 18 (44/388, 11.3%). Others included non-balanced autosomal structural rearrangements (16/388, 4.1%), 45, X0 (16/388, 4.1%) and 47, XXY (15/388, 3.9%). (3) Lymphocyte karyotype analysis of the couples: parents of 153 fetuses were analyzed to determine the origin of abnormal karyotype. Fifty-eight familial and 95 de novo abnormalities were found. FISH results were the same with G-banding karyotype, and two of these were trisomy 21. CONCLUSIONS: Abnormal karyotype composition is different according to different maternal amniocentisis indications. There is a variety of abnormal karyotypes in the second trimester pregnancy, and the risk of fetal malformation is related with the kind of abnormal karyotype.

Interactions between galectin-3 and integrinbeta3 in regulating endometrial cell proliferation and adhesion.

BACKGROUND: Galectin-3 (gal-3) is a beta-galactoside-binding protein which can be detected in endometrium. The study was designed to investigate synergism of gal-3 and integrinbeta3 in endometrial cell proliferation and adhesion in an in vitro model of endometrial receptivity. METHODS: The RL95-2 cell line was employed as an in vitro model for receptive endometrium. Cells transfected with gal-3 siRNA or treated with exogenous gal-3 were incubated with or without function-blocking integrinbeta1/3 antibody for evaluating synergism of gal-3 and integrins on cell proliferation and adhesion. Proliferation was measured by BrdU incorporation, and adhesion to fibronectin (FN) was determined by an adhesion assay. Integrin expression was analyzed by Flow Cytometry and western blots. Bewo spheroids were co-cultured with the RL95-2 monolayer to mimic the blastocyst-endometrial interaction, and colocalization of gal-3, integrinbeta3 and FN at the interface was observed by confocal microscopy. RESULTS: The knock-down of gal-3 inhibited RL95-2 cell proliferation and adhesion. However, a reduction of proliferation and adhesion was also observed in presence of exogenous gal-3, and this was further reduced by a functional block to integrinbeta3. Moreover, gal-3 knock-down significantly increased integrinbeta3 expression, however, the colocalization of integrinbeta3 and FN was not increased. As expected, the colocalization of integrinbeta3 was decreased with the knock-down of gal-3. CONCLUSIONS: This study has provided an in vitro model for the complex interactions between gal-3 and integrinbeta3 in the regulation of endometrial cell proliferation and adhesion.