Zinc(II) niflumato complex effects on MMP activity and gene expression in human endometrial cell lines.

Endometriosis is a chronic inflammatory disease which increasingly affects young women under 35 years of age and leads to subfertility even infertility. Analysis of the cytotoxic effect of zinc(II) niflumato complex with neocuproine ([Zn(neo)(nif)2] or Zn-Nif) on immortalized human endometriotic cell line (12Z) and on control immortalized human endometrial stromal cell line (hTERT) was performed using xCELLigence technology for approximately 72 h following the treatment with Zn-Nif as well as cell viability Trypan Blue Assay. 12Z cell line proliferated more slowly compared to unaffected cells, whereas hTERT cells did not show similar behavior after treatment. The complex probably reduces the effect of pro-inflammatory pathways due to the effect of NSAID, while presence of zinc might reduce the level of ROS and regulate ER2 levels and MMP activity. The observed effects and high selectivity for rapidly proliferating cells with increased inflammatory activity suggest a good prognosis of successful decrease of endometriosis stage with this complex.

Supplementation of oleic acid, stearic acid, palmitic acid and ß-hydroxybutyrate increase H3K9me3 in endometrial epithelial cells of cattle cultured in vitro.

There is growing evidence that greater than homeostatic blood concentrations of nonesterified fatty acids (NEFAs) and ß-hydroxybutyrate (BHBA) have negative consequences on dairy cow's fertility, but effects on cell homeostasis in the reproductive system is not completely understood. In this study, lipids accumulation, reactive oxygen species (ROS) concentrations, abundance of gene transcripts, and immunofluorescence signal of H3K4me3 and H3K9me3 were evaluated in endometrial epithelial cells of cattle cultured with NEFAs (Oleic (OA), Stearic (SA) and Palmitic (PA) acids), BHBA, NEFAs + BHBA or each of the three NEFAs alone. The cellular lipids were in greater concentrations as a result of NEFAs + BHBA, NEFAs, SA or OA supplementation, but not by BHBA or PA. The ROS concentrations were greater when there were treatments with NEFAs + BHBA, NEFAs or BHBA. The relative mRNA abundance for genes involved in the regulation of apoptosis (XIAP), glucose transport (GLUT3), and DNA methylation (DNMT1) were greater when there were NEFAs + BHBA, but not NEFAs, BHBA, OA, SA or PA treatments. The immunofluorescence signal for H3K9me3 was greater when there were NEFAs + BHBA, NEFAs or PA, but not by BHBA, OA or SA treatments. These findings indicate that NEFAs and BHBA have an additive effect on endometrial cells of cattle by altering epigenetic markers and the expression of genes controlling important cellular pathways. Furthermore, there was cellular lipid accumulation and increased H3K9me3 in cultured bovine endometrial cells that was mainly induced by OA and PA treatments, respectively.

A novel human endometrial epithelial cell line for modeling gynecological diseases and for drug screening.

Endometrium-related malignancies including uterine endometrioid carcinoma, ovarian clear cell carcinoma and ovarian endometrioid carcinoma are major types of gynecologic cancer, claiming more than 13,000 women's lives annually in the United States. In vitro cell models that recapitulate "normal" endometrial epithelial cells and their malignant counterparts are critically needed to facilitate the studies of pathogenesis in endometrium-related carcinomas. To achieve this objective, we have established a human endometrial epithelial cell line, hEM3, through immortalization and clonal selection from a primary human endometrium culture. hEM3 exhibits stable growth in vitro without senescence. hEM3 expresses protein markers characteristic of the endometrial epithelium, and they include PAX8, EpCAM, cytokeratin 7/8, and ER. hEM3 does not harbor pathogenic germline mutations in genes involving DNA mismatch repair (MMR) or homologous repair (HR) pathways. Despite its unlimited capacity of in vitro proliferation, hEM3 cells are not transformed, as they are not tumorigenic in immunocompromised mice. The cell line is amenable for gene editing, and we have established several gene-specific knockout clones targeting ARID1A, a tumor suppressor gene involved in the SWI/SNF chromatin remodeling. Drug screening demonstrates that both HDAC inhibitor and PARP inhibitor are effective in targeting cells with ARID1A deletion. Together, our data support the potential of hEM3 as a cell line model for studying the pathobiology of endometrium-related diseases and for developing effective precision therapies.

Porcine endometrial 3D co-culture: Morphological changes in 3D endometrium tissues according to hormonal changes.

Cells cultured as monolayers proliferate well, but do not sustain their differentiation characteristics. Previous studies have investigated the interactions between cells and growth factors or cytokines by establishing either in vivo or in vitro three-dimensional (3D) cultures. Using porcine uterine epithelial cells and endometrial cells, the current study was designed to develop a 3D uterine culture system and investigate the response to hormone treatment. Formation of the 3D uterine model was similar to that of uterus from the group supplemented with calcium and magnesium, and the addition of these ions altered the spectrum of basement membrane degrading enzyme expression and activity. In particular, the epithelial cell junctions in the 3D model most closely resembled those of an actual uterus when the medium was supplemented with calcium and magnesium; the intercellular basement membrane structure was also tall under these conditions. The study confirmed that Casp-3 expression was lowest in the P4 (progesterone) treatment group, and this hormone was the most potent stimulus for formation of the endometrial cell layer. Therefore, the addition of calcium and magnesium plays an important role in the formation of a 3D uterine model, and the addition of P4 hormone mimics uterine thickening by stimulating growth of the epithelial cell layer.

Comparative biochemical profiles, utero-ovarian function, and fertility of the postpartum buffalo with and without subclinical endometritis.

In postpartum buffaloes, the process of uterine involution and changes in blood metabolic profile has not been studied in relation to development of subclinical endometritis (SCE). In this study, buffaloes (n = 100) approaching calving were identified. Weekly blood samples were collected on the day of calving up to 6 weeks post-calving. The diameter of uterine horns and onset of ovarian cyclicity (corpus luteum) were recorded through ultrasonography. On the basis of polymorphonuclear cell (PMN) cell count in endometrial cytology at days 45-50 postpartum, buffaloes were divided into two groups, viz., with SCE (> 5% PMN; n = 38) and without SCE (≤ 5% PMN; n = 62). Buffaloes with SCE took longer (P < 0.05) time to complete uterine involution and had larger (P < 0.05) uterine horn diameter between 3rd and 6th weeks postpartum and lower prostaglandin F2α metabolite (PGFM) concentration on the day of calving (P < 0.05) and 1 week (P < 0.001) post-calving than without SCE group. Buffaloes with SCE had lower (P < 0.001) concentration of glucose at weeks 2 and 3, higher (P < 0.001) ß-hydroxybutyric acid (BHBA) at week 3, and lower serum albumin concentration throughout the sampling period (P < 0.05 to 0.001) except at 1 week post-calving as compared to without SCE group. The urea concentration was significantly lower (P < 0.05 to 0.001) in buffaloes with SCE from 4 weeks post-calving onwards than without SCE group. The calcium concentration was lower in buffaloes with SCE at weeks 5 (P < 0.001) and 6 (P < 0.05) postpartum, whereas the concentration of magnesium and phosphorus was uniform between the two groups. No significant (P > 0.05) difference in onset of ovarian cyclicity between the 2 groups was observed, whereas buffaloes with SCE had longer (P = 0.001) median days open (141 days) than their counterpart (117 days). The first service conception rate, cumulative pregnancy rate, and pregnancy rate at 150 days postpartum were lower (P < 0.05) in buffaloes with SCE than without SCE group. In summary, higher BHBA and lower serum concentrations of glucose, albumin, urea, and calcium control onset of subclinical endometritis which in turn has negative impact on fertility of buffaloes.

Effect of Dehydrocostus Lactone Isolated from the Roots of Aucklandia lappa on the Apoptosis of Endometriotic Cells and the Alternative Activation of Endometriosis-Associated Macrophages.

The roots of Aucklandia lappa have been used in traditional medicine in Asia to treat inflammation and diseases associated with pain, including endometriosis. The aim of this study was to investigate the anti-endometriotic effect of dehydrocostus lactone, an active compound in A. lappa roots, using human endometriotic cells and macrophages stimulated by these cells. Dehydrocostus lactone induced apoptotic cell death in 12Z human endometriotic cells. Dehydrocostus lactone stimulated the activation of caspase-3, -8, and -9, while caspase inhibitors significantly reversed the dehydrocostus lactone-induced cell death in 12Z cells. In addition, dehydrocostus lactone decreased the production of PGE2 and neurotrophins (BDNF, NGF, NT3, and NT4/5), which are regarded as endometriosis-associated pain factors in human endometriotic cells. Moreover, dehydrocostus lactone inhibited the expression of M2 markers (CD206, and Trem-2), IL-10, VEGF, and MMP-2/-9 in endometriosis-associated macrophages (EAMs). Furthermore, dehydrocostus lactone inhibited the Akt and NFκB pathways in both endometriotic cells and EAMs. Taken together, our findings suggest that dehydrocostus lactone, an active compound of A, lappa, has anti-endometriotic activities via induction of apoptosis and downregulation of pain factors in endometriotic cells and inhibition of the alternative activation of EAMs.

Phytoestrogen exposure alters endometrial stromal cells and interferes with decidualization signaling.

OBJECTIVE: To investigate whether phytoestrogens (genistein and daidzein) alter in vitro decidualization of human endometrial stromal cells (ESCs). DESIGN: Isolated primary ESCs were exposed to phytoestrogens and decidualized in vitro. SETTING: Academic fertility center. PATIENT(S): Twenty fertile oocyte donors attending the IVI Valencia clinic. INTERVENTION(S): Treatment of ESC with phytoestrogens at 0, 10, 20, 50, and 100 µM. MAIN OUTCOME MEASURE(S): The ESC proliferation was analyzed by MTS assay. In vitro decidualization was induced in the presence of phytoestrogens by medroxyprogesterone acetate/cyclic adenosine 3':5' monophosphate and evaluated by prolactin (PRL) ELISA and F-actin immunostaining. The Ki67 proliferative marker was analyzed by immunofluorescence. The ESC apoptosis was assessed by annexin V/propidium iodide detection using flow cytometry. Estrogen (ERß) and P receptor (PR) localization were evaluated by immunofluorescence. RESULT(S): The ESC exposed to 0, 19, 20, 50, and 100 µM of genistein, daidzein, and genistein + daidzein showed a dose-dependent proliferation decrease. After 48-96 hours of culture, this reduction was significant in the presence of 50 µM of phytoestrogens versus 10 µM untreated ESC. The ESC decidualized in the presence of phytoestrogens did not rearrange their cytoskeletons and showed a significant decrease in PRL secretion compared with untreated decidualized ESCs (dESCs). However, phytoestrogens did not alter proliferative status or the percentage of viable/apoptotic cells in dESC compared with untreated dESC. During decidualization, phytoestrogens induced the same nuclear translocation of ERß and PR as the control dESC. CONCLUSION(S): This study reveals that high doses of phytoestrogens could affect the in vitro decidualization process.

Three-dimensional culture of endometrial cells from domestic cats: A new in vitro platform for assessing plastic toxicity.

Plastic polymers can be combined with additives that modify physical properties and stability of the material. However, the biocompatibility of those additives is not well known. The objective of the study was to characterize the impact of zinc stearate-a common additive-through the development of a novel three-dimensional (3-D) in vitro platform with endometrial cells from domestic cats. Epithelial and stromal cells from adult uteri were isolated and cultured in medium supplemented with 3% Matrigel for two weeks in plastic tissue culture dishes that had been identified as polystyrene with and without zinc stearate by Raman, FTIR, and X-ray fluorescence spectroscopies. Three-dimensional cell structures that were obtained were measured and categorized by shape. Cell viability, proliferation, differentiation, organization, and apoptosis then were assessed by immuno-staining. Results indicated that zinc stearate did not affect 3-D endometrial cell structure morphology, viability, or cellular composition. This first study of a new in vitro platform will be useful for studies testing the influence of other additives, drugs, or exogenous hormones.

Seminal plasma modulates expression of endometrial inflammatory meditators in the bovine†.

Seminal plasma has conventionally been viewed as a transport and survival medium for mammalian sperm; however, its role now extends beyond this process to actively targeting female tissues. Studies in rodents, swine, and humans demonstrate that seminal plasma induces molecular and cellular changes within the endometrium or cervix following insemination. Seminal-plasma-induced alterations to the maternal environment have been theorized to facilitate embryo development, modulate maternal immunity toward the conceptus, and potentially improve pregnancy success. It is unknown if bovine seminal plasma modulates the uterine environment following insemination in the cow, where routine use of artificial insemination reduces maternal exposure to seminal plasma. We hypothesize that seminal plasma modulates the expression of inflammatory mediators in the endometrium, altering the maternal environment of early pregnancy. In vitro, seminal plasma altered intact endometrial explant expression of CSF2, IL1B, IL6, IL17A, TGFB1, IFNE, PTGS2, and AKR1C4. Furthermore, endometrial epithelial cell CSF2, CXCL8, TGFB1, PTGS2, and AKR1C4 expression were increased after seminal plasma exposure, while endometrial stromal cell CSF2, IL1B, IL6, CXCL8, IL17A, TGFB1, PTGS2, and AKR1C4 expression were increased following seminal plasma exposure. Endometrial expression of IL1B was increased in the cow 24 h after uterine infusion of seminal plasma, while other evaluated inflammatory mediators remained unchanged. These data indicate that seminal plasma may induce changes in the bovine endometrium in a temporal manner. Understanding the role of seminal plasma in modulating the maternal environment may aid in improving pregnancy success in cattle.

Berberine inhibits growth and inflammatory invasive phenotypes of ectopic stromal cells: Imply the possible treatment of adenomyosis.

Adenomyosis is a common chronic gynecological disorder with some tumor-like properties, including aberrant proliferation, invasion and migration. Berberine (BBR) is an isoquinoline derivative alkaloid with diverse pharmacological activities for the treatment of a wide variety of diseases. However, the effect of BBR on adenomyosis has not been understood. This study was to evaluate the potential therapeutic effect of BBR on ectopic endometrial stromal cells (EESCs) isolated from patients with adenomyosis. Our data showed that BBR significantly inhibited the proliferation and viability of eutopic endometrial stromal cells (EuESCs) and EESCs, while slightly affected the growth of normal endometrial stromal cells (NESCs). BBR markedly exhibited a growth inhibitory effect on EESCs by triggering apoptosis and cell cycle arrest, and alleviating the expression of inflammatory invasive phenotypes (IL-6, IL-8, TGF-ß, EGF, VEGF, and MMP2). The alleviation of inflammatory invasive phenotypes partly involved nuclear translocation of NFκB/p65 and stat3 activation. Taken together, BBR markedly inhibits the growth of EESCs and might be a promising new strategy for the treatment of adenomyosis.

Pueraria mirifica inhibits 17ß-estradiol-induced cell proliferation of human endometrial mesenchymal stem cells.

OBJECTIVE: The notion that the human endometrium may contain a population of stem cells has recently been proposed. The mesenchymal stem cells (MSCs) in the endometrium are believed to be responsible for the remarkable regenerative ability of endometrial cells. Estrogens influence the physiological and pathological processes of several hormone-dependent tissues, such as the endometrium. Pueraria mirifica (PM) is a herbal plant that contains several phytoestrogens, including isoflavones, lignans, and coumestans, and is known to exert an estrogenic effect on animal models. The present study investigated the effects of PM on the proliferation of human endometrial MSCs (hEN-MSCs). MATERIALS AND METHODS: The hEN-MSCs were isolated from human endometrial tissue. The surface markers of these hEN-MSCs were identified through reverse transcription-polymerase chain reaction analysis. The proliferation potential of hEN-MSCs was measured through a cell proliferation assay. Multilineage differentiation ability was confirmed through Oil red O and von Kossa staining. RESULTS: This study demonstrated that 17ß-estradiol-responsive MSCs with Oct-4, CD90, and CD105 gene expression can be derived from the human endometrium and that PM exerts biological effects on hEN-MSCs, specifically, enhanced cell growth rate, through the estrogen receptor. Furthermore, PM at 1500 and 2000 µg/mL significantly increased cell proliferation compared with the vehicle control, and PM concentration at 1000 µg/mL significantly inhibited the enhanced cell growth rate induced by 17ß-estradiol in hEN-MSCs. CONCLUSION: This study provides new insights into the possible biological effects of PM on the proliferation of hEN-MSCs.

A phytoestrogen supplement prevents the altered gene expression associated with pregnancy implantation induced by IL-1ß in endometrial epithelial cells.

Phytoestrogens stimulate expression of the uterine estrogen receptor and regulate uterine functions in reproductive tissues. However, comprehensive understanding of the beneficial impacts of phytoestrogens on uterine biology at the molecular level remains unexplored. Interleukin-1ß (IL-1ß) expression is increased in the inflamed decidua and is associated with first trimester pregnancy loss. AglyMax-Sup has the same composition as that of the phytoestrogen supplement AglyMax but with added vitamins and other components. Expression of genes associated with implantation may be enhanced by AglyMax-Sup compared with AglyMax. We tested the hypothesis that AglyMax-Sup has greater effects on implantation compared with AglyMax, using RT-PCR and Western blotting in the endometrial epithelial cell line. Furthermore, we investigated the protective effect of AglyMax-Sup on IL-1ßinduced changes in estrogen-responsive gene expression in endometrial epithelial cells. The purpose of this study was to compare the effects of the phytoestrogen supplement AglyMax-Sup with those of AglyMax on estrogen-responsive gene expression. AglyMax and AglyMax-Sup significantly (p<0.05) induced gene expression of glycodelin-A, HoxA10, IL-11, LIF, MEG-E8 and TGFß1. AglyMax-Sup induced high levels of these genes compared with the levels induced by AglyMax. The enhanced expression of LIF, IL-11, integrin αV, and HOXA10 induced by AglyMax-Sup was abolished by the ER antagonist fulvestrant and the ERK inhibitor PD98059. Meanwhile, IL-1ß inhibited progesterone plus estrogen-induced TGFß1, glycodelin-A, HOXA10, and integrin αV expression. IL-1ß-induced suppression of these expression was reversed by AglyMax-Sup. These results indicate that expression of genes associated with implantation may be increased by AglyMax-Sup compared with AglyMax. AglyMax-Sup might abrogate IL-1ß-mediated changes that can affect embryo implantation via the MAPK pathway.

Anti-Endometriotic Effects of Pueraria Flower Extract in Human Endometriotic Cells and Mice.

Pueraria flowers have been used as a vegetable and an ingredient for tea and jelly. In this study, we investigated the effects of Pueraria flower extract (PFE) on endometriosis, a common gynaecological disease characterised by local sterile inflammation of peritoneal cavity. PFE suppressed the adhesion of human endometriotic cells 11Z and 12Z to human mesothelial Met5A cells. In addition, PFE significantly inhibited the migration of 11Z and 12Z cells as shown by woundhealing and transwell migration assays. PFE reduced the protein and mRNA levels of matrix metalloproteinase (MMP)-2 and MMP-9 in endometriotic cells. Moreover, extracellular signalregulated kinase (ERK)1/2 was activated by PFE treatment, and an ERK1/2 inhibitor, PD98059, significantly inhibited PFE-inhibited cell migration in endometriotic cells. Furthermore, PFE significantly suppressed endometriotic lesion formation in a mouse model. These data suggest that Pueraria flower is a potential anti-endometriotic agent for the inhibition of endometriotic cell adhesion, migration, and MMP expression.

An experimental study on the use of icariin for improving thickness of thin endometrium.

This study aimed to investigate the effect of icariin (ICA) on thin endometrium in a rat model. To this end, 6- to 8-week-old female Sprague Dawley rats (105) were randomly divided into 7 groups: untreated, vehicle-treated (lavage with NaCl), high-dose ICA (lavage with ICA at 200 mg∙kg-1∙day-1), medium-dose ICA (lavage ICA at 100 mg∙kg-1∙day-1), low-dose ICA (lavage with ICA at 50 mg∙kg-1∙day-1), sham model (injected with NaCl at uterus horn), and sample group. To induce thin endometrium, rats of all groups (except sham-model) were injected with 95% ethanol via the uterine horn. Each group underwent its respective treatment for 3 estrous cycles, after which 5 rats from each group were sacrificed, and endometrial thickness was measured. The expression of CD31, factor VIII, vascular endothelial growth factor (VEGF), cytokeratin (CK), and vimentin were detected via immunohistochemistry. The results showed that CD31, factor VIII, and VEGF were primarily expressed in the cytoplasm of endometrial and vascular epithelial cells. No difference in the expression of these factors was detected between the ICA lavage groups and the untreated groups. However, high dose ICA-treated group exhibited significantly higher expression of CD31, factor VIII, and VEGF compared to that in the low dose and vehicle-treated groups. CK and vimentin in the endometrial tissue were significantly higher in the untreated and treatment groups compared to the vehicle-treated group. This study demonstrated that ICA increases thickness of the endometrium, and it may modulate expression of VEGF, CD31, and factor VIII.

Effects of Lycium barbarum polysaccharides on the damage to human endometrial stromal cells induced by hydrogen peroxide.

Previous studies have shown that Lycium barbarum polysaccharides (LBPs) serve an important role in antioxidant activity to protect the cells and tissues. However, the specific mechanism of LBPs in the prevention of endometrial damage remains to be elucidated. Using morphological observation, cell proliferation assay, the detection of superoxide dismutase (SOD) activity and the content of malondialdehyde (MDA) in cell culture supernatant fluid, the detection by western blot analysis and reverse transcription-quantitative polymerase chain reaction of the mRNA and protein expression levels of caspase­3 and Bcl­2 in endometrial stromal cells (ESCs), it was demonstrated that, in vitro, hydrogen peroxide (H2O2)-induced death of ESCs, increased the content of MDA and decreased the activity of SOD, and decreased the expression of Bcl-2 and increased the expression of caspase­3. LBPs can inhibit H2O2­induced cell death of ESCs, decrease the content of MDA in ESCs and increase the activity of SOD, as well as increasing the expression of Bcl­2 and decreasing the expression levels of caspase­3. These findings suggested that LBPs can inhibit H2O2­induced apoptosis of EECs and that LBPs are able to offer a significant protection against oxidative stress to ESCs.

Effect of quercetin on the expression of Bcl-2/Bax apoptotic proteins in endometrial cells of lipopolysaccharide-induced-abortion.

OBJECTIVE: To explore the effect of quercetin on the expressions of Bcl-2/Bax apoptotic proteins in endometrial cells in mice with abortion induced by lipopolysaccharide. METHODS: For in vivo experiment, twenty five Kunming mice were randomly divided into five groups at day 4 of pregnancy, with 5 mice per group. The mice were treated with lipopolysaccharide (LPS) through tail vein intravenous injection at day 4 of pregnancy, followed by different concentrations of quercetin by oral gavage consecutively at days 5 to 6 of pregnancy. On day 7 of gestation, the mice were sacrificed and the histopathological changes of the uterus tissues were observed. Immunohistochemical staining was applied to the detection of Bcl-2/Bax apoptotic proteins in the endometrial cells. For in vitro experiment, the primary endometrial cells were cultured using a uterus tissue mass culturing method sampled at day 4.5 of pregnancy. The cells were treated with LPS with or without different dosages of quercetin, respectively, for 12 h after 80% confluence. The expression of Bcl-2/Bax apoptotic proteins were detected by western blotting. RESULTS: Both the in vivo and in vitro experiments showed decreased expression of Bcl-2 and enhanced expression of Bax after LPS treatment, leading to a decreased Bcl-2/Bax ratio. The expression of Bcl-2 significantly increased while the expression of Bax was significantly elevated, in the LPS plus quercetin group compared to the LPS only group. CONCLUSION: These results suggest that quercetin has protective effect by partially regulating the expression of Bcl-2/Bax proteins, which in turn inhibits endometrial cell apoptosis and benefits the embryo implantation.

Effect of warming Yang and removing blood stasis method on matrix metalloproteinases / tissue inhibitor metalloproteinases levels secreted by cultured endometrial cells from patients with endometriosis.

OBJECTIVE: To investigate the effect of Chinese medicines using the warming Yang and removing blood stasis method on levels of matrix metalloproteinases (MMPs)/tissue inhibitor metalloproteinases (TIMPs) secreted by cultured endometrial cells from patients with endometriosis. METHODS: Ectopic and eutopic endometrial cells obtaind from 15 endometriosis patients were cultured in vitro, and divided randomly into five groups: high dose; moderate dose; low dose; nemestran; blank control. The three dose groups were treated with a decoction prepared according to the principle of warming Yang and removing blood stasis; nemestran and 0.9% NaCl were administered to the nemestran group and balnk control group, respectively. Eutopic endometrial cells obtaind from 10 hysteromyoma patients were cultured in vitro, as the normal control group, 0.9% NaCl were administered to the normal control group. Cell culture supernatants were collected and levels of matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), tissue inhibitor metalloproteinase-1 (TIMP-1) and tissue inhibitor metalloproteinase-2 (TIMP-2) detected by enzyme-linked immuno sorbent assay (ELISA). RESULTS: Compared with the normal control group, levels of MMP-1, MMP-2, and MMP-9 in eutopic and ectopic endometrium cell supernatants in the blank control group were increased, whereas levels of TIMP-1 and TIMP-2 were decreased (P < 0.05). Compared with the blank control group, levels of MMP-1 and MMP-2 in ectopic and eutopic endometrium cell supernatants cultured in low-dose, middle-dose, and high-dose groups were decreased, whereas levels of TIMP-1 and TIMP-2 were increased significantly (P < 0.05). CONCLUSION: The warming Yang and removing blood stasis method affects expression of MMPs and TIMPs.

Effect of Xianziyizhen Recipe Capsule on PGI2-PPARδ Signaling Pathway in Embryo Implantation Dysfunction Mice.

PROBLEM: We investigated the effect of Xianziyizhen recipe capsule (XRC), a kidney-tonifying herb, on the PGI2-PPARδ signaling pathway at the maternal-fetal interface in embryo implantation dysfunction (EID) mice. METHOD OF STUDY: Intragastric administration of Progynova (estradiol) or XRC was performed in EID mouse model, following experimental induction of kidney deficiency by co-treatment with chemotherapy drug hydroxyurea and antiprogesterone mifepristone. The PPARδ and IL-11 mRNA expression in endometrium were detected by real-time relative reverse transcription-polymerase chain reaction (RT-PCR). Further, the protein expression of COX-2, PGI2, MMP-9, and TIMP-3 was detected in endometrial glandular epithelium and in stromal cells by immunohistochemical (IHC) assay. RESULTS: The results showed that hydroxyurea and mifepristone-induced EID were associated with significantly lower PPARδ and IL-11 mRNA levels in endometrium and reduced COX-2, PGI2, MMP-9, and TIMP-3 levels in endometrial glandular epithelium, compared with normal controls. However, XRC and Progynova treatment reversed these effects, leading to significant increases in PPARδ and IL-11 mRNA expression, and COX-2, PGI2, MMP-9 and TIMP-3 protein levels, when compared with the levels observed in EID mice. CONCLUSION: These results strongly suggested that XRC is beneficial in EID treatment and that XRC may mediate its effects through regulation of the PGI2-PPARδ signaling pathway.

[BDNF secretion in human mesenchymal stem cells isolated from bone marrow, endometrium and adipose tissue].

The ability of mesenchymal stem cells (MSCs) to differentiate into neuronal lineage determines the potential of these cells as a substrate for a cell replacement therapy. In this paper we compare the neurogenic potential of MSCs isolated from bone marrow (BMSC), subcutaneous adipose tissue (AD MSC) and menstrual blood (eMSC). It was found that the native eMCSs, BMSCs and AD MSCs express neuronal marker ß-III-tubulin with a frequency of 90, 50 and 14%, respectively. We also showned that eMSCs have a high endogenous level of brain-derived neurotrophic factor (BDNF), whereas the BMSCs and the AD MSCs are characterized by low basal BDNF levels. As induction of neuronal differentiation in the studied MSCs using differentiation medium containing B27 and N2 supplements, 5-azacytidine, retinoic acid, IBMX and dbcAMF caused changes in the cells morphology, the increased expression of ß-III-tubulin, and the appearance of neuronal markers GFAP, NF-H, NeuN and MAP2. BDNF secretion during differentiation was significantly enhanced in the BMSCs and decreased in the eMSCs cultures. However, no correlation between the basal and induced levels of the neuronal markers expression and BDNF secretion in the studied MSCs has been established.

A randomised controlled trial of a preconceptional dietary intervention in women undergoing IVF treatment (PREPARE trial).

BACKGROUND: In vitro fertilisation (IVF) treatment provides an opportunity to study early developmental responses to periconceptional dietary interventions. Retrospective studies have suggested links between preconception diet and fertility, and more recently, a "Mediterranean" diet has been reported to increase pregnancy rates by up to 40%. In addition, a prospective study examining increased intake of omega-3 polyunsaturated fats demonstrated a quickened rate of embryo development after IVF. However, up to now, few prospective randomised controlled trials have investigated the impact of periconceptional dietary interventions on fertility outcomes. METHODS AND DESIGN: The study is a randomised controlled trial of a dietary intervention consisting of olive oil for cooking, an olive oil based spread, and a daily supplement drink enriched with Vitamin D (10 microgram daily) and marine omega-3 fatty acids (2 g daily) for 6 weeks preconception versus a control diet of sunflower seed oil for cooking, a sunflower oil based spread, and a daily supplement drink without added Vitamin D or marine omega-3 fatty acids. Couples undergoing IVF will be randomised to either the intervention or control group (55 in each arm). The primary endpoint is embryo developmental competency in vitro, measured by validated morphokinetic markers. Secondary outcomes will include the effect of the dietary intervention on the nutritional content of the intrauterine environment. DISCUSSION: This approach will enable rigorous examination of the impact of the dietary intervention on early embryo development, together with the influence of the peri-implantation intra-uterine nutritional environment. TRIAL REGISTRATION: ISRCTN50956936.