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Embryo implantation is a complicated process involving a series of endometrial changes that depend on differential gene expression. MicroRNAs (miRNAs) are important for regulation of gene expression. Previous studies have shown that miRNAs may participate in the regulation of gene expression during embryo implantation. To explore the role of endometrial miRNAs in early murine pregnancy, we used microarrays to investigate whether miRNAs were differentially expressed in the mouse endometrium on pregnancy day 4 (D4) and day 6 (D6). The results demonstrated that 17 miRNAs were upregulated and 18 were downregulated (>2-fold) in D6 endometria compared to D4. We identified that mmu-miR-193 exhibited the highest upregulation on D6, and the upregulation of mmu-miR-193 before embryo implantation could reduce the embryo implantation rate. Further, we demonstrated that mmu-miR-193 influenced embryo implantation by regulating growth factor receptor-bound protein 7 expression. In summary, our study suggests that mmu-miR-193 plays an important role in embryo implantation.

Benzo(a)pyrene (BaP) is a ubiquitous environmental pollutant. Studies have demonstrated it to be an endocrine-disrupting chemical that can cause adverse effects on the female reproductive system. However, the effect of BaP on early pregnancy has not been reported. We investigated the effect of BaP on endometrial receptivity and embryo implantation. Pregnant mice were dosed with BaP at 0.2, 2 and 20 mg/kg/day from day 1 (D1) to day 5 (D5) of gestation. Exposure to BaP impaired the morphology of the endometrium and decreased the number of implantation sites (p0.2=0.006, p2=0.167, p20=0.003). Levels of estrodiol (p0.001, for three treatment group compare with control group) and progesterone-4 in plasma were elevated in BaP-treatment groups (p0.20.001, p20.001, p20=0.032). Expression of estrogen receptor-α was up-regulated (p0.2=0.002, p2=0.131, p20=0.024) whereas expression of the progesterone receptor was down-regulated (p0.20.001, p2=0.064, p20=0.021). Levels of receptivity-related genes HoxA10 (p0.20.001, p2=0.135, p200.001) and E-cadherin (p0.2=0.002, p2=0.624, p20=0.137) were changed by BaP. These results revealed that BaP can disrupt the balance of estrogen and progesterone, influence expression of their receptors and downstream related genes, lead to changes in endometrium receptivity, and reduce of the number of implantation sites.

To 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.The 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.IDO 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.The 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.

Successful mouse embryo implantation requires a receptive uterus and an activated blastocyst. A large number of genes, cytokines, and other factors are involved in the process. MicroRNAs (miRNAs) regulate the expression of many genes, and previous studies have investigated the relationship between miRNA expression and embryo implantation. In this study, we show that mmu-microRNA-200a (mmu-miR-200a) is expressed in a spatiatemporal manner during implantation in mouse uterus and found that phosphatase and tensin homolog (PTEN), SON, and programmed cell death 4 (Pdcd4) are the target genes of mmu-miR-200a by bioinformatics analysis. In vitro gain and loss of function experiments confirm that PTEN, a critical gene for cell proliferation and apoptosis, is the target gene of mmu-miR-200a. Our experiments also show that injection of the uterine horn with mmu-miR-200a lentivirus leads to a decreased implantation rate. Collectively, our results suggest that mmu-miR-200a affects embryo implantation by regulating PTEN protein expression. Thus, clarifying the physiological functions of uterine miRNAs will help to elucidate the embryo implantation process and may even contribute to curing infertility and inventing new contraceptives.

Aggregation behaviors in mixtures of an anionic gemini surfactant 1,3-bis(N-dodecyl-N-propanesulfonate sodium)-propane (C(12)C(3)C(12)(SO(3))(2)) and a cationic single-chain surfactant cetyltrimethylammonium bromide (CTAB) have been investigated in aqueous solutions at pH 9.5 by turbidity, rheology, isothermal titration microcalorimetry (ITC), cryogenic transmission electron microscopy, and dynamic light scattering. Reversible aggregate transitions from spherical micelles to wormlike micelles, vesicles, and back to wormlike micelles and spherical micelles are successfully realized through fine regulation over the mixing ratio of surfactants, i.e., the anionic/cationic charge ratio. The five aggregate regions display distinguished phase boundaries so that the aggregate regions can be well controlled. From thermodynamic aspect, the ITC curves clearly reflect all the aggregate transitions and the related interaction mechanism. The self-assembling ability of the C(12)C(3)C(12)(SO(3))(2)/CTAB mixtures are significantly improved compared with both individual surfactants. Micelle growth from spherical to long wormlike micelles takes place at a relative low total concentration, i.e., 2.0 mM. The wormlike micelle solution at 10 mM or higher shows high viscosity and shear thinning property. Moreover, the C(12)C(3)C(12)(SO(3))(2)/CTAB mixtures do not precipitate even at 1:1 charge ratio and relative high concentration. It suggests that applying gemini surfactant should be an effective approach to improve the solubility of anionic/cationic surfactant mixtures and in turn may promote applications of the surfactant mixtures.

During the reproductive cycle, a number of genes controlling endometrial changes are regulated by DNA methylation, a common epigenetic modification. Because dietary folate affects DNA methylation, we determined whether a folate-deficient diet (FDD) alters DNA methylation in endometria of pseudopregnant mice, focusing on the homeobox A10 (Hoxa10) promoter. Mice were given an FDD or control diet for 40 to 45 days and examined on day 5 of pseudopregnancy. Compared to control mice, FDD mice had lower folate levels in liver and serum (P = .004). However, the FDD did not significantly affect DNA methylation within the cytosine-guanine dinucleotide (CpG)-rich Hoxa10 promoter, even when specific CpG sites were examined (P > .05). In endometrial tissue sections, the localization of anti-Hoxa10 staining was unchanged in FDD mice. Therefore, folate deficiency did not significantly affect promoter methylation or expression of Hoxa10.

Background Folate, one of the B vitamins, provides the one-carbon units required for methylation. Folate deficiency has been associated with many pathologies. However, much less is known about the effect of it on human reproduction, especially on implantation. The establishment of uterine receptivity is crucial for successful embryo implantation. Gene expression can be influenced by both heredity and epigenetics such as DNA methylation. However, it is not known whether the methylation and expression of genes related to uterine receptivity can be affected by folate levels. To explore whether folate deficiency affected the epigenetic regulation of genes related to uterine receptivity, and their influence on implantation, we investigated the methylation and expression of cadherin 1 (Cdh1), progesterone receptor (Pgr) and estrogen receptor 1 (Esr1) genes during implantation and the implantation efficiency using a folate-deficient pregnant mouse model. Methods Serum folate levels of pregnant mice were measured using the electro-chemiluminescence immunoassay. The methylation status of Cdh1, Pgr and Esr1 promoter regions was determined by methylation-specific PCR and bisulfite sequencing. The expression of Cdh1, Pgr and Esr1 in the implantation-site endometrium was examined by real-time PCR, western blot and immunohistochemistry. The number and the morphology of pinopodes, important morphological markers of endometrial receptivity, were examined using scanning electron microscopy. The number of implantation sites demarcated by distinct blue bands was recorded. Results Serum folate levels of the folate-deficient group were lower (5.42 ± 1.35 ng/ml, n= 42) than those of the normal group (24.13 ± 4 .26 ng/ml, n= 37; P = 0.003). Here we show that the methylation status and mRNA levels of Esr1 were decreased (P= 0.021, P= 0.045, respectively), while the Cdh1 and Pgr expression levels were slightly but not significantly elevated and the methylation status did not vary in the folate-deficient mice compared with the wild type. Neither the number nor morphology of pinopodes was affected by folate deficiency. Furthermore, folate deficiency did not affect the number of implantation sites in mice. Conclusions This study demonstrates for the first time that, unlike the effects on Esr1, folate deficiency in mice does not influence the methylation and expression of Pgr and Cdh1, two genes shown to be essential for uterine receptivity and embryo implantation. Embryo implantation in mice appears to be unaffected by a deficiency in folate, suggesting that abnormalities in a pregnancy caused by folate deficiency start to develop after implantation.

Di-(2-ethylhexyl)-phthalate (DEHP) is a ubiquitous environmental pollutant and endocrine disruptor (ED) that causes serious adverse effects on animal and human health. The harmful effects of DEHP on human reproduction are increasingly recognized, especially in women. However, it is not known how endometrial receptivity and embryo implantation, which play important roles in the establishment of pregnancy, are affected by DEHP. This study was aimed towards investigating the effects of DEHP on endometrial receptivity and embryo implantation in pregnant mice. The pregnant mice received DEHP at 0, 250, 500 and 1000 mg/kg/day from day 1 (D1) of gestation until sacrifice. Administration of DEHP led to compromised endometrial receptivity and decreased number of implantation sites. The mRNA and protein expression levels of ERα, PR and E-cadherin, but not those of HoxA10 and MMP-2, were up-regulated by DEHP in the mouse endometrium. The results further suggested that DEHP disrupts the MAPK and NF-κB signaling pathways. This was maybe one of paths which influenced the E-cadherin expression. In conclusion, DEHP reduced endometrial receptivity and impaired embryo implantation by influencing the expression of hormone receptors and E-cadherin. Therefore, determining the full extent of the hazards of DEHP to human reproduction will be vital to developing and implementing effective protective measures.

The expression of IK cytokine was investigated in the mouse endometrium during early pregnancy (D1-D7 of pregnancy) and pseudopregnancy using real-time PCR, western blotting and immunohistochemical analysis, and the effects of IK cytokine on embryo implantation were observed by injection with antisense IK cytokine oligodeoxynucleotides in the uterine horn. Our data showed that the expression of IK cytokine mRNA increased gradually from D1 to D4 of pregnancy and reached a peak level at D4 of pregnancy (P

Manganese is a trace element and a cofactor of many enzymes, so it is essential for physiologic functioning, but it is also a neurotoxin at high doses. Manganism is most often caused by occupational exposure. It is manifested by a myriad of signs and symptoms ranging from the neurasthenia syndrome, such as headache and dizziness, to the Parkinson-like syndrome, depending on the blood manganese levels as well as the duration of exposure. We are reporting a case of manganism using both clinical and occupational hygiene investigation methods. The patient presented the neurasthenia syndrome accompanied by hypertonicity of arm muscles and was diagnosed to have mild chronic manganism. Finally, the patient was discharged from the hospital after the treatment had improved her condition. In China, there are many chronic manganese cases, partly due to a rapid industrial development with great use of Mn and the low self-protection awareness among the workers and the factories management that cannot catch up with the speed of the economical development. Therefore, factories are responsible for improving the conditions at the workplace.

Uterine decidualization, characterized by stromal cell proliferation and differentiation into polyploid decidual cells, is critical to the establishment of pregnancy in mice, although the mechanism underlying this process remains poorly understood. This study is the first to investigate the expression of gamma-amino butyric acid (GABA) and the GABA A-type receptor π subunit (GABPR) in the early-pregnancy mouse uterus and their roles in decidualization. The expression of GABRP was detected from Day 4 to 8 of pregnancy. The effects of GABA and GABA A-type receptor on cell proliferation and apoptosis were investigated using the Cell Titer 96® AQueous One Solution Cell Proliferation Assay and flow cytometry. The levels of cyclin D3 protein were measured in cultured stromal cells artificially induced to undergo decidualization, and treated with GABA and a GABA A-type receptor agonist or antagonist, respectively, at the same time. mRNA expression of gabrp in implantation sites was lower than that in inter-implanted sites. GABA and GABRP protein were localized in the luminal and glandular epithelium, stromal cells, and decidual cells. In vitro, GABPR protein level was decreased in cultured stromal cells during the decidualization process. The addition of GABA and the GABA A-type receptor agonist Muscimol inhibited stromal cell proliferation, promoted apoptosis, and arrested cells in S-phase, followed by decreased expression of cyclin D3. These results show that in mice, GABA was actively involved in inhibiting stromal cell proliferation and suppresses decidualization progress through GABA A-type receptors by down-regulating cyclin D3 level.

BACKGROUND: The DNA demethylating agent 5-aza-2'-deoxycytidine (5-aza-CdR) incorporates into DNA and decreases DNA methylation, sparking interest in its use as a potential therapeutic agent. We aimed to determine the effects of maternal 5-aza-CdR treatment on embryo implantation in the mouse and to evaluate whether these effects are associated with decreased levels of DNA methyltransferases (Dnmts) and three genes (estrogen receptor α [Esr1], progesterone receptor [Pgr], and homeobox A10 [Hoxa10]) that are vital for control of endometrial changes during implantation. METHODS AND PRINCIPAL FINDINGS: Mice treated with 5-aza-CdR had a dose-dependent decrease in number of implantation sites, with defected endometrial decidualization and stromal cell proliferation. Western blot analysis on pseudo-pregnant day 3 (PD3) showed that 0.1 mg/kg 5-aza-CdR significantly repressed Dnmt3a protein level, and 0.5 mg/kg 5-aza-CdR significantly repressed Dnmt1, Dnmt3a, and Dnmt3b protein levels in the endometrium. On PD5, mice showed significantly decreased Dnmt3a protein level with 0.1 mg/kg 5-aza-CdR, and significantly decreased Dnmt1 and Dnmt3a with 0.5 mg/kg 5-aza-CdR. Immunohistochemical staining showed that 5-aza-CdR repressed DNMT expression in a cell type-specific fashion within the uterus, including decreased expression of Dnmt1 in luminal and/or glandular epithelium and of Dnmt3a and Dnmt3b in stroma. Furthermore, the 5' flanking regions of the Esr1, Pgr, and Hoxa10 were hypomethylated on PD5. Interestingly, the higher (0.5 mg/kg) dose of 5-aza-CdR decreased protein expression of Esr1, Pgr, and Hoxa10 in the endometrium on PD5 in both methylation-dependent and methylation-independent manners. CONCLUSIONS: The effects of 5-aza-CdR on embryo implantation in mice were associated with altered expression of endometrial Dnmts and genes controlling endometrial changes, suggesting that altered gene methylation, and not cytotoxicity alone, contributes to implantation defects induced by 5-aza-CdR.

Translationally controlled tumor protein (TCTP) is a highly conserved, growth-related protein. Previous studies showed that TCTP is involved in many biological processes and it is essential for early embryo development and proliferation of embryonic stem cells. However, whether TCTP plays a role during embryo implantation remains unclear. This paper examines the expression and the role of TCTP in the uterus of mice during early pregnancy.The expression of TCTP in the uterus of mice during early pregnancy was examined by quantitative real-time PCR, immunohistochemistry and western blot. A functional study of TCTP in embryo implantation of mice was also performed by intrauterine injection with antisense oligodeoxynucleotides (A-ODNs) of TCTP on day 3 (D3) of pregnancy.The TCTP mRNA levels were significantly upgraded from D3 to D5 of pregnancy and reached maximum levels on D5, then dramatically decreased on D6 and D7. The levels of the TCTP protein detected by western blot were consistent with those of the mRNA. Immunohistochemistry analysis showed that the TCTP protein was mainly located in the luminal and the glandular epithelium on D1 and D2 of pregnancy and reached maximum levels on D5 in the luminal and glandular epithelium and in the stromal cells. The levels of TCTP in the pseudo-pregnant uterus of mice were lower than those of pregnant mice on D4 and D6. Furthermore, inhibiting the TCTP expression by intrauterine injection with A-ODNs of TCTP on D3 of pregnancy significantly reduced the number of the implanted embryos compared with the control.This study demonstrated that TCTP may play a significant role in embryo implantation in mice.

Cisplatin (CDDP) is a major chemotherapeutic drug used in the treatment of human ovarian cancer. Tubeimoside I (TBMS1) has also shown potent antitumor and antitumor-promoting effects, and may offer a promising new approach in the effective treatment of CDDP-resistant human ovarian cancers. This study aimed to investigate the effect of TBMS1 in sensitizing CDDP in CDDP-resistant human ovarian cancer cells (A2780/DDP). A variety of methods were employed to measure cell apoptosis, p38, ERK1/2 and glutathione S-transferase (GST)-π expressions. It was found that TBMS1 combined with CDDP promoted cell apoptosis, decreased proliferation activity and increased cytosolic Ca2+ levels. Bcl-2 protein expression was down-regulated but Bax was up-regulated. Moreover, GST-π mRNA and protein expression were decreased. TBMS1 reduced the resistance of the cells to CDDP-induced cytotoxicity. Both the p38 inhibitor (SB203580) and the ERK1/2 inhibitor (PD98059) effectively blocked this effect. These results suggest that TBMS1 can effectively sensitize CDDP in CDDP-resistant human ovarian cancer cells through the down-regulation of the ERK1/2 and the up-regulation of the p38 signaling pathways.

Mitochondria play important roles in the intrinsic pathways that trigger apoptosis. Anticancer chemotherapies eliminate cancer cells mainly through the induction of apoptosis. In the present study, we investigated the mechanism of the cytotoxic effects of tubeimoside I (TBMS1) on the human choriocarcinoma cell line (JEG-3). Choriocarcinoma is one of the most common malignant tumors in the reproductive system. TBMS1, a triterpenoid saponin, isolated from the tubers of Bolbostemma paniculatum (Maxim) Franquet (Cucurbitaceae), showed potent antitumor effects. However, the potential roles of TBMS1 in the treatment of choriocarcinoma remain unknown. In the present study, we examined the effects of TBMS1 on JEG-3 cells. TBMS1 displayed strong growth inhibitory effects on JEG-3 cell growth. In addition, TBMS1 treatment with TBMS1 led to marked cell apoptosis, significant cell cycle arrest at G2 phase and decrease in mitochondrial transmembrane potential (ΔΨm). Cytochrome c was released from the mitochondria and caspase-3 expression was enhanced. Furthermore, TBMS1 induced the up-regulation of Bcl-2 associated X protein (Bax) expression, down-regulation of Bcl-2 expression, inhibition of nuclear factor-κ-B (NF-κB) function and impacted the phosphorylation of p38 mitogen-activated protein kinase (p38/MAPK), extracellular signal-regulated kinases (ERK)1/2 and protein kinase B (Akt). Taken together, our findings suggest that TBMS1 is an efficient apoptosis-inducing agent for choriocarcinoma cells, which exerts its effects, at least partially, by the induction of mitochondrial dysfunction and regulation of the p38/MAPK, ERK1/2 and PI3K/Akt signaling pathways.

The Leydig cell tumor, derived from interstitial cells, is a rare neoplasm. In most cases, Leydig cell tumors are benign, however, if the tumor is malignant, no effective treatments are currently available. In this study, we aimed to evaluate the effects of icariin on the growth of the mouse Leydig tumor cell line MLTC-1 and to examine its underlying mechanism. Icariin caused a dose-dependent decrease in the viability of MLTC-1 cells, which coincided with an increase in cell apoptosis through regulation of the expression of Bcl-2/Bax and cytochrome c, activation of caspase-9 and -3. Moreover, the pro-apoptotic effect of icariin on MLTC-1 cells is related to piwil4, since icariin induced a decrease in piwil4 protein expression and piwil4 silencing significantly enhanced the cytotoxic effects of icariin in MLTC-1 cells. These findings suggest a novel anticancer effect of icariin in Leydig cell tumor through activation of the mitochondrial pathway and down-regulation of the expression of piwil4.

Proliferin-related protein (PRP) was originally identified as an angiogenesis inhibitor in mouse placentas. Indeed, the tissue expression of PRP has mainly been documented in placentas. We report herein for the first time that PRP is expressed in male rat testes. Immunocytochemical and in situ hybridization results showed positive PRP immunostaining in Leydig cells. Immunofluorescent staining of PRP in the TM3 Leydig cell line indicates that PRP is located within the cytoplasm. The expression pattern of PRP in rat testis exhibited an age-related increase. HCG significantly up-regulated the level of expression of PRP in TM3 cells via the PKA pathway. To elucidate the function of PRP, experiments were conducted to examine the consequences of lentiviral-mediated RNA interference (RNAi) of PRP on testosterone production and expression of several genes involved in steroidogenesis. PRP silencing caused a decrease in HCG-stimulated testosterone production. In addition, PRP silencing attenuated the increase in PRLR mRNA following HCG stimulation. Moreover, the enhanced effect of PRL on HCG-induced testosterone production was also weakened following PRP silencing, indicating that PRP may be involved in PRL function through an effect on PRL receptor expression in response to stimuli. Taken together, these data suggest that PRP is regulated by HCG and plays roles in male reproduction, such as testosterone production.

The effects of anionic surfactants on the aggregation-induced emission (AIE) feature of cationic M-silole molecules have been studied. The electrostatic binding of M-silole with the surfactants greatly promotes the aggregation of the mixtures. The M-silole/surfactant aggregates at 1:1 charge ratio exhibit the maximum fluorescence intensity. Excess surfactant molecules will distribute the M-silole molecules into different micelles and weaken the fluorescence. The fluorescence intensity of the mixed M-silole/surfactant aggregates can be effectively modulated by choosing different surfactants. The gemini surfactants display a much stronger ability of enhancing fluorescence intensity than do the single-chain surfactants. Especially, the gemini surfactant with benzene rings shows the best performance in enhancing fluorescence of M-silole due to both the strongest aggregation ability and the pi-pi interaction with M-silole.

As a highly conserved nuclear protein, death domain-associated protein (Daxx) plays an important role in transcriptional control, carcinogenesis, and resistance to virus infection and so on. In order to further investigate the mechanism of Daxx, the yeast two-hybrid technique was used to screen the intra-cellular proteins interacting with Daxx. And 13 positive colonies and three proteins interacting with Daxx were obtained. One of the candidate proteins was identified as ferritin, heavy polypeptide 1(FTH1). The interaction between Daxx and FTH1 was further supported by GST pull-down and co-immunoprecipitation respectively. Then Daxx was determined to induce apoptosis and FTH1 can inhibit Daxx-mediated apoptosis. Besides, it is found that Daxx mediated apoptosis through the Fas-Daxx-ASK1-JNK1 signaling pathway, while FTH1 can inhibit the activation of JNK signaling pathway. We present evidence to demonstrate the FTH1 and Daxx are able to participate in apoptosis pathway through JNK signal molecule and FTH1 can inhibit this pathway.

An improved method, which exhibited simultaneously clearly kinetochores and the nucleolar organizer regions of human chromosomes by pretreating of human metaphase chromosomes with HCl and NaOH, followed by staining with silver nitrate and visualizing using ammoniacal silver, is described in the present communication. It has important role for analysis of kinetochore variation, mechanism of chromosomal non-disjunction as well as identification of fuctional active centromeres.

To investigate molecular mechanism of traditional Chinese medicine Rheum offcinale against Yersinia pestis, whole genome DNA microarray that contains 4005 annotated genes of Y. pestis was used. The minimal inhibition concentration (MIC) of R. offcinale extract against Y. pestis was determined by liquid dilution method. The gene expression profile of Y. pestis was performed after exposured to R. offcinale extract at a concentration of 10 X MIC for 30 and 60 minutes. The total RNA extracted and purified from Y. pestis were reverse-transcribed to cDNA and labeled by Cy3-Cy5 dye. The labeled probes were hybridized to the microarray and the results were obtained by a laser scanner and analyzed by the SAM software. The microarray data was confirmed by RT-PCR. The platform of the DNA microarray-based bacteria transcriptional profiling was eshtablished. The results revealed general gene expression changes of Y. pestis were a global phenomenon. Down-regulation of genes encoding proteins involved in ribosome protein synthesis was a remarkable change. Genes encoding cell envelope and transport/binding proteins were the major changed genes of the Y. pestis in response to R. offcinale.

To understand the function of Mayven and investigate the pathogenesis of multiple sclerosis, the gene sequences of different truncated Mayven were amplified from the gene library of human brain. These truncated fragments, including fragment P1 (1-902 bp), fragment P2 (1-523 bp), fragment P3 (507-182 bp) and fragment P4 (887-1782 bp), were cloned into pGEX-4T-2 vector to construct recombinant plasmids. The recombinant plasmids were transformed into E. coli BL21(DE3) and induced to express by IPTG. The expressed proteins were detected by SDS-PAGE and Western blot, and were purified by GST purifying system. The results showed that recombinant express vectors of different truncated GST-Mayven were successfully constructed and were expressed in soluble form protein induced by IPTG. The fusion proteins have good reactivity to GST antibody. The construction of recombinant express vectors of different truncated GST-Mayven lays a basis for further function study on Mayven.

Calreticulin (CRT), a Ca 2+ -binding storage protein and chaperone in the endoplasmic reticulum, modulates cell adhesiveness and integrin-dependent Ca 2+ signaling. However, the role of CRT during implantation remains poorly understood. In the present study, we characterized the expression of CRT mRNA and the protein in mouse endometria from pregnancy D1 to D7. Real-Time PCR and in situ hybridization results showed that the levels of CRT mRNA in the endometria of pregnant mice were significantly higher than those of non-pregnant mice (P

Smac/DIABLO (second mitochondria-derived activator of caspase/direct IAP-binding protein with low PI) is a 29 kDa mitochondrial precursor protein, which is proteolytically processed in mitochondria into a 23 kDa mature protein. It is released from the mitochondrial intermembrane space to cytosol after an apoptotic trigger. Smac/DIABLO acts as a dimer and it contributes to caspase activation by sequestering the inhibitor of apoptosis proteins (IAPs). In order to further investigate the mechanism of Smac/DIABLO action, we used the mature form of Smac/DIABLO as a bait and screened proteins that interact with mature Smac/DIABLO in human liver cDNA library using the yeast two-hybrid system. Forty-two colonies were obtained after 5.8x10(6) colonies were screened by nutrition limitation and X-galactosidase assay. After DNA sequence analysis and homology retrieval, one of the candidate proteins was identified as TRAF domain of the TNF receptor associated factor 3 (TRAF3). The interaction site between TRAF3 and Smac/DIABLO was identified by beta-galactosidase test. The interaction between TRAF3 and Smac/DIABLO via TRAF domain was identified in vivo by co-immunoprecipitation in HepG2 cells, and the direct interaction between TRAF3 and Smac/DIABLO in vitro was identified by GST-pull down assay. Co-expression of TRAF3 and mature Smac/DIABLO in 293 cells could enhance the Smac/DIABLO-mediated apoptosis. These results suggested that TRAF3 interacted with Smac/DIABLO via TRAF domain, leading to an increased proapoptotic effect of Smac/DIABLO in cytoplasm.

OBJECTIVE: Embryo implantation is directly affected by genes related to uterine receptivity. Studies have demonstrated the important roles of miRNAs in the regulation of gene expression. Our early miRNA chip analyses revealed that the mmu-miR-141 expression in endometrial tissue is lower after embryo implantation than before it. However, the possible roles of miR-141 in embryo implantation have not yet been elucidated. Here, mmu-miR-141 was designed to detect the expression and role of miR-141 in the endometria of mice in early pregnancy following embryo implantation. METHODS: Real-time PCR and in-situ hybridization were used to study mmu-miR-141 expression in mouse uterus. Cell proliferation was detected by tetrazolium dye (MTT) assay and flow cytometry. Real-time PCR and Western blot analysis were used to confirm the mRNA and protein levels of phosphatase and tensin homolog (PTEN) to determine whether it was the target gene of mmu-miR-141. Enhanced green fluorescent protein (EGFP) fluorescence reporter vector analysis was also performed. A functional study was performed by injecting mice uteri with mmu-miR-141 inhibitor or mimic vectors. RESULTS: mmu-miR-141 expression was lower on day 6 (D6) than day 4 (D4) and could be increased by progesterone. Reduced mmu-miR-141 could decrease the proliferation activity of stromal cells and promote apoptosis. Upregulation of mmu-miR-141 inhibited PTEN protein expression but downregulation of mmu-miR-141 increased it, while the mRNA level remained unchanged. EGFP fluorescence reporter vector analysis showed that miR-141 targets the 3'-untranslated region of the PTEN mRNA. In addition, when the physiological mmu-miR-141 level was altered on D2 by injecting with inhibitor or mimic, the embryo implantation sites were significantly decreased on D7. CONCLUSIONS: This study demonstrated that mmu-miR-141 might influence cell proliferation and apoptosis in the endometrium by negatively regulating PTEN expression, and could also influence the number of embryo implantation sites. mmu-miR-141 plays an essential role in embryo implantation.

Graphene oxide obtained by the Hummers method was discovered to be an efficient and recyclable acid catalyst for the conversion of fructose-based biopolymers into 5-ethoxymethylfurfural (EMF). EMF yields of 92%, 71%, 34% and 66% were achieved when 5-hydroxymethylfurfural (HMF), fructose, sucrose and inulin were used as starting materials, respectively.

Bacterial resistance to antibiotics has become today a major public health issue. In the development of new anti-infectious therapies, antimicrobial peptides appear as promising candidates. However, their mechanisms of action against bacterial membranes are still poorly understood. We describe for the first time the interaction and penetration of plasticins into lipid monolayers and bilayers modeling the two leaflets of the asymmetrical outer membrane of Gram-negative bacteria. The lipid composition of these monolayers mimics that of each leaflet: mixtures of LPS Re 595 mutant and wild type S-form from Salmonella enterica for the external leaflet, and SOPE/SOPG/cardiolipin (80/15/5) for the inner one. The analysis of the interfacial behavior of native (PTCDA1) and modified (PTCDA1-KF) antimicrobial plasticins showed that PTCDA1-KF exhibited better surface properties than its unmodified counterpart. Both peptides could penetrate into the model monolayers at concentrations higher than 0.1 μM. The penetration was particularly enhanced for PTCDA1-KF into the mixed LPS monolayer, due to attractive electrostatic interactions. Grazing X-ray diffraction and atomic force microscopy studies revealed the changes in LPS monolayers organization upon peptide insertion. The interaction of plasticins with liposomes was also monitored by light scattering and circular dichroism techniques. Only the cationic plasticin achieved full disaggregation and structuration in α helices, whereas the native one remained aggregated and unstructured. The main steps of the penetration mechanism of the two plasticins into lipid models of the external leaflet of the outer membrane of Gram-negative bacteria have been established.