Evolution of higher mesenchymal CD44 expression in the human lineage: A gene linked to cancer malignancy.

CD44 is an extracellular matrix receptor implicated in cancer progression. CD44 increases the invasibility of skin (SF) and endometrial stromal fibroblasts (ESF) by cancer and trophoblast cells. We reasoned that the evolution of CD44 expression can affect both, the fetal-maternal interaction through CD44 in ESF as well as vulnerability to malignant cancer through expression in SF. We studied the evolution of CD44 expression in mammalian SF and ESF and demonstrate that in the human lineage evolved higher CD44 expression. Isoform expression in cattle and human is very similar suggesting that differences in invasibility are not due to the nature of expressed isoforms. We then asked whether the concerted gene expression increase in both cell types is due to shared regulatory mechanisms or due to cell type-specific factors. Reporter gene experiments with cells and cis-regulatory elements from human and cattle show that the difference of CD44 expression is due to cis effects as well as cell type-specific trans effects. These results suggest that the concerted expression increase is likely due to selection acting on both cell types because the evolutionary change in cell type-specific factors requires selection on cell type-specific functions. This scenario implies that the malignancy enhancing effects of elevated CD44 expression in humans likely evolved as a side-effect of positive selection on a yet unidentified other function of CD44. A possible candidate is the anti-fibrotic effect of CD44 but there are no reliable data showing that humans and primates are less fibrotic than other mammals.

Delayed Implantation Induced by Letrozole in Mice.

Implantation timing is critical for a successful pregnancy. A short delay in embryo implantation caused by targeted gene ablation produced a cascading problem in the later stages of the pregnancy. Although several delayed implantation models have been established in wild mice, almost none of them is suitable for investigating the early delay's effects on the late events of pregnancy. Here, we report a new delayed implantation model established by the intraperitoneal administration of letrozole at 5 mg/kg body weight on day 3 of pregnancy. In these mice, initiation of implantation was induced at will by the injection of estradiol (E2). When the estradiol (3 ng) was injected on day 4 of pregnancy (i.e., without delay), the embryo implantation restarted, and the pregnancy continued normally. However, 25 ng estrogen caused compromised implantation. We also found that 67% of the female mice could be pregnant normally and finally gave birth when the estradiol injection (3 ng) was on day 5 of pregnancy (i.e., 1-day delay). Most failed pregnancies had impaired decidualization, decreased serum progesterone levels, and compromised angiogenesis. Progesterone supplementation could rescue decidualization failure in the mice. Collectively, we established a new model of delayed implantation by letrozole, which can be easily applied to study the effect and mechanisms of delay of embryo implantation on the progression of late pregnancy events.

Ventilation/perfusion imaging predicts response to balloon pulmonary angioplasty in patients with chronic thromboembolic pulmonary hypertension.

OBJECTS: Although balloon pulmonary angioplasty (BPA) has emerged as an alternative treatment option for chronic thromboembolic pulmonary hypertension (CTEPH), it is followed in some patients by residual PH. We studied the efficacy of BPA on pulmonary blood flow and the predictive value of ventilation/perfusion (V/Q) scanning. METHODS: We retrospectively reviewed the clinical database, which included patients diagnosed with CTEPH who had received BPA. All patients undergone V/Q scanning to quantify the extent of pulmonary perfusion abnormality before and after BPA. Pulmonary hemodynamics were assessed by right heart catheterization, and cardiac function and exercise capacity were evaluated at baseline and post-BPA. A total of 120 CTEPH patients were included for analysis. RESULTS: BPA significantly alleviated mean pulmonary arterial pressure (mPAP: 48.0 ± 12.9 mmHg vs 34.7 ± 10.3 mmHg, P < 0.001) and pulmonary vascular resistance (PVR: 8.8 ± 4.1 Wood units vs 5.2 ± 3.0 Wood units, P < 0.001), and improved cardiac function (N-terminal pro B-type natriuretic peptide: 1628.7 ± 2887.2 pg/mL vs 400.4 ± 669.3 pg/mL, P < 0.001) and exercise capacity (6-minute walking distance: 386 ± 122 m vs 461 ± 86 m, P < 0.001). The extent of pulmonary perfusion abnormality represented by the percentage of perfusion defects (PPDs%) was improved after BPA (50.1 ± 13.6 vs 35.6 ± 14.2, P < 0.001), with the right and inferior lung lobes benefitting the most. PPDs% < 35.5 at baseline and greater restoration of PPDs% after BPA (∆PPDs% > 20.6) were associated with a better response to BPA (PPDs% < 35.5: odds ratio [OR] 10.857, 95% confidence interval [95%CI] 1.393-84.635, P = 0.023; ∆PPDs% > 20.6: OR 1.035, 95% CI 1.002-1.068, P = 0.036). CONCLUSION: BPA significantly restored pulmonary blood flow, predominantly in the right and inferior lobes. V/Q scanning has the potential to predict the therapeutic response to BPA for CTEPH.

Predictive value of SPECT myocardial perfusion imaging in patients with unrevascularized coronary chronic total occlusion.

OBJECTIVE: Data involved the association between myocardial ischaemia and the outcome for unrevascularized coronary chronic total occlusion (CTO) patients were limited. The purpose of this study was to evaluate the predictive value of ischaemia detected by single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI) for the adverse events in unrevascularized CTO patients. We further explored whether ischaemia generated from CTO vessel can independently predict the outcome. METHODS: Patients with at least one unrevascularized CTO on coronary angiography were enrolled in this study. Exercise stress/rest SPECT MPI was performed in all patients. All patients were then followed by telephone interview and reviewing of medical records. RESULTS: Patients with ischaemia experienced significantly higher rate of adverse events than non-ischaemia patients (40.7% vs 7.1%, P = 0.002). Ischaemia demonstrated on MPI [odds ratio (OR) = 7.656; 95% confidence interval (CI) 1.598-36.677; P = 0.011] was an independent predictor for adverse events. Moreover, CTO-ischaemia (OR = 5.466; 95% CI 1.015-29.420; P = 0.048), non-CTO ischaemia (OR = 29.174; 95% CI 3.245-262.322; P = 0.003), mixed-ischaemia (OR = 7.130, 95% CI 1.257-40.445; P = 0.027) were all independent predictors for outcome. CONCLUSION: Ischaemia demonstrated on MPI, especially CTO-ischaemia were independent predictors for the adverse events. SPECT MPI can aid to identify patients at risk of adverse events, who may benefit from subsequent CTO percutaneous coronary intervention.

The value of ventilation/perfusion scanning and CT pulmonary angiography in predicting chronic thromboembolic pulmonary hypertension after acute pulmonary embolism: a one-year follow-up study.

To investigate the value of ventilation/perfusion (V/Q) scanning and CT pulmonary angiography (PA) in predicting CTEPH development after acute pulmonary embolism (APE). This study was performed in APE patients who had undergone both V/Q and CT PA after 3-month anticoagulation. The residual pulmonary obstructions were assessed based on V/Q and CT PA, and then recorded as pulmonary perfusion detect score (PPDs) and CT pulmonary artery obstruction index (PAOI). The predictive performance of PPDs and CT PAOI for CTEPH were determined and risk factors for predicting CTEPH development were identified. A total of 235 patients with initial diagnosis of APE were included in this study. ROC analysis showed that the AUCs of the PPDs and CT PAOI were 0.957 and 0.895, with corresponding cut-off values of 20.50% and 17.50% for predicting CTEPH development. Neither sensitivity nor specificity differed significantly between PPDs and CT PAOI (Sensitivity: 92.00% vs. 80.00%, P = 0.25; Specificity: 88.10% vs. 89.52%, P = 0.69). The univariable and multivariable logistic regression analysis demonstrated that pulmonary arterial hypertension confirmed by echocardiography at initial APE diagnosis (OR: 6.16, 95%CI: 1.31-29.02, P = 0.02), a PPDs of > 20.50% (OR: 22.95, 95%CI: 2.37-222.19, P = 0.007), and a CT PAOI of > 17.50% (OR: 9.98, 95%CI: 2.06-48.49, P = 0.004) were associated with CTEPH development. Both V/Q and CT PA after 3-month anticoagulation for APE showed great performance in predicting CTEPH development, and V/Q scanning has a tendency to be more sensitive but less specific than CT PA. The residual pulmonary embolism detected by V/Q and CT PA was associated with an increased risk of CTEPH development.

Transcriptomic responses to hypoxia in endometrial and decidual stromal cells.

Human reproductive success depends on a properly decidualized uterine endometrium that allows implantation and the formation of the placenta. At the core of the decidualization process are endometrial stromal fibroblasts (ESF) that differentiate to decidual stromal cells (DSC). As variations in oxygen levels are functionally relevant in endometrium both upon menstruation and during placentation, we assessed the transcriptomic responses to hypoxia in ESF and DSC. In both cell types, hypoxia-upregulated genes in classical hypoxia pathways such as glycolysis and the epithelial mesenchymal transition. In DSC, hypoxia restored an ESF-like transcriptional state for a subset of transcription factors that are known targets of the progesterone receptor, suggesting that hypoxia partially interferes with progesterone signaling. In both cell types, hypoxia modified transcription of several inflammatory transcription factors that are known regulators of decidualization, including decreased transcription of STATs and increased transcription of CEBPs. We observed that hypoxia-upregulated genes in ESF and DSC had a significant overlap with genes previously detected to be upregulated in endometriotic stromal cells. Promoter analysis of the genes in this overlap suggested the hypoxia-upregulated Jun/Fos and CEBP transcription factors as potential drivers of endometriosis-associated transcription. Using immunohistochemistry, we observed increased expression of JUND and CEBPD in endometriosis lesions compared to healthy endometria. Overall, the findings suggest that hypoxic stress establishes distinct transcriptional states in ESF and DSC and that hypoxia influences the expression of genes that contribute to the core gene regulation of endometriotic stromal cells.

Aggregation Behavior of Acylated Pepsin-Solubilized Collagen Based on Fluorescence Spectrum Technology.

The aggregation behavior of collagen-based materials plays an important role in their processing because it could affect their physicochemical properties. Based on the intrinsic fluorescence characteristic of tyrosine, fluorescence spectrum technology was used to investigate the aggregation state of the acylated collagen molecules in aqueous solution. The results showed that the aggregate degree of the acylated collagen was higher than that of the native collagen due to the hydrophobic interaction. With the increase of concentrations of the acylated collagen or at NaCl higher than 40 mmol/L, the aggregate degree of the acylated collagen molecules increased. When the pH was close to the isoelectric point of the acylated collagen, the hydrophobic interaction and the hydrogen bond helped to increase the aggregation degree. However, with the increase of temperature (10-70 ℃), the aggregation state of the acylated collagen decreased gradually due to the quenching, the molecular collision, and the broken of hydrogen bonds. Furthermore, two-dimensional correlation spectroscopy (2D-COS) showed that the response order was 360 > 305 nm at various acylated collagen and NaCl (>40 mmol/L) concentrations, while the response order was 305 > 360 nm when the pH value was increased from 5.0 to 9.0. Temperature-dependent 2D-COS showed there were four bands that occurred and the response order was listed as follows: 293 > 305 > 360 > 420 nm. In brief, the results might provide an important guide for molding processes of the acylated collagen.

Quantitative assessment of right ventricular glucose metabolism in idiopathic pulmonary arterial hypertension patients: a longitudinal study.

AIMS: Right ventricular (RV) glucose metabolism disorder in pulmonary arterial hypertension (PAH) has been studied using (18)F-fluorodeoxyglucose positron emission tomography (FDG-PET) imaging with inconsistent results. We aimed to quantitatively assess RV glucose metabolism and further identify its role of monitoring RV function in idiopathic PAH (IPAH) patients in a longitudinal study. METHODS AND RESULTS: Twenty-seven treatment-naïve IPAH patients and 21 healthy control subjects performed FDG-PET dynamic scan for quantification of the rate of myocardium glucose utilization (rMGU) and echocardiography for assessment of cardiac function. Right heart catheterization was conducted for IPAH patients for haemodynamic measurement. A subgroup of 14 patients repeated FDG-PET and echocardiography after 6-month treatment. RV rMGU was significantly increased compared with controls; while the rMGU in left ventricle showed no difference. RV rMGU was significantly correlated with pulmonary artery pressure, pulmonary vascular resistance, RV Tei index, and right atrial area, and negatively correlated with RV ejection fraction (RVEF) and tricuspid annular plane systolic excursion. Six of 14 patients with increased RV rMGU after 6-month treatment showed no change in RVEF, 6-min walk distance (6MWD), and RV Tei index; however, the other 8 patients with decreased RV rMGU demonstrated significantly increased RVEF and 6MWD and decreased RV Tei index. Notably, the change in RV rMGU of 14 patients was significantly correlated with the change in 6MWD and RV Tei index. CONCLUSION: Increased RV rMGU of IPAH correlates with RV dysfunction and RV pressure overload. The change in RV glucose metabolism may help monitor RV function after treatment.

The characterization and prognostic significance of right ventricular glucose metabolism in non-ischemic dilated cardiomyopathy.

AIMS: In dilated cardiomyopathy (DCM), there are limited data on right ventricular (RV) glucose metabolism assessed by [(18)F]fludeoxyglucose positron emission tomography ((18)F-FDG PET) imaging. We aimed to characterize RV glucose metabolism and investigate the prognostic significance of RV FDG uptake in DCM. METHODS AND RESULTS: (18)F-FDG PET imaging and cardiac magnetic resonance imaging (MRI) were performed in 63 consecutive DCM patients within an interval of 3-7 days. There was a significant correlation between RVEF and RV FDG uptake whether corrected RV standard uptake value (cRVSUV) (r = -0.571, P < .001) or the relative RV FDG uptake determined as the ratio of RV to left ventricular (LV) corrected SUV (cR/L) (r = -0.405, P < .001) was used. During a median follow-up period of 804 days, 15 patients (23.8%) reached the primary endpoint of all-cause mortality or heart transplantation. On univariate Cox analysis, cRVSUV > 7.01 and cR/L > 0.795 were significantly associated with the overall survival (hazard ratio [HR] 5.415, 95% confidence interval [CI] 1.945-15.078, P < .001; HR 6.422, 95% CI 2.250-18.332, P < .001). Patients with increased RV FDG uptake had a worse outcome (cRVSUV > 7.01 vs cRVSUV ≤ 7.01, log-rank 13.085, P < .001; cR/L > 0.795 vs cR/L ≤ 0.795, log-rank 15.695, P < .001). On multivariate analysis, cR/L > 0.795 remained a significant independent predictor of the endpoint (HR 5.001, 95% CI 1.641-15.239, P = .004), while cRVSUV > 7.01 showed no significance (HR 2.611; 95% CI 0.797-8.558; P = .113). CONCLUSIONS: Increased RV FDG uptake was associated with RV dysfunction and may be a prognostic predictor of all-cause mortality or heart transplantation in patients with DCM.

Control of regional decidualization in implantation: Role of FoxM1 downstream of Hoxa10 and cyclin D3.

Appropriate regulation of regional uterine stromal cell decidualization in implantation, at the mesometrial triangle and secondary decidual zone (SDZ) locations, is critical for successful pregnancy, although the regulatory mechanisms remain poorly understood. In this regard, the available animal models that would specifically allow mechanistic analysis of site-specific decidualization are strikingly limited. Our study found that heightened expression of FoxM1, a Forkhead box transcription factor, is regulated during decidualization, and its conditional deletion in mice reveals failure of implantation with regional decidualization defects such as a much smaller mesometrial decidua with enlarged SDZ. Analysis of cell cycle progression during decidualization both in vivo and in vitro demonstrates that the loss of FoxM1 elicits diploid cell deficiency with enhanced arrests prior to mitosis and concomitant upregulation of polyploidy. We further showed that Hoxa10 and cyclin D3, two decidual markers, control transcriptional regulation and intra-nuclear protein translocation of FoxM1 in polyploid cells, respectively. Overall, we suggest that proper regional decidualization and polyploidy development requires FoxM1 signaling downstream of Hoxa10 and cyclin D3.

Myostatin knockout in bovine fetal fibroblasts by using TALEN.

Myostatin (MSTN) can negatively regulate the growth and development of skeletal muscle, and mutations of bovine MSTN gene can cause a "double-muscle" feature. To knock out MSTN gene in bovine fetal fibroblast by transcription activator-like effector nucleases (TALENs) and obtain MSTN knockout cell lines, we constructed one pair of MSTN-TALEN vector and transfected into bovine fetal fibroblast cells by PEI and electroporation. Sequencing results demonstrated that TALEN was available for MSTN knockout. T7 endonuclease 1 (T7E1) was used for the detection of mutation efficiency. The results indicated that knockout efficiency of electroporation transfection was 20.4%, and 10 MSTN(+/-) and MSTN(-/-) cell colonies were obtained via limiting dilution method. The deletion number of nucleotides ranged from 1 to 20, and some of them were frameshift mutation, which could provide the possibility in production of MSTN knockout cattle in the future.

Epigenetic changes through DNA methylation contribute to uterine stromal cell decidualization.

Embryo-uterine interaction during early pregnancy critically depends on the coordinated expression of numerous genes at the site of implantation. The epigenetic mechanism through DNA methylation (DNM) plays a major role in the control of gene expression, although this regulatory event remains unknown in uterine implantation sites. Our analysis revealed the presence of DNA methyltransferase 1 (Dnmt1) in mouse endometrial cells on the receptive d 4 of pregnancy and early postattachment (d 5) phase, whereas Dnmt3a had lower abundant expression. Both Dnmt1 and Dnmt3a were coordinately expressed in decidual cells on d 6-8. 5-Methycytosine showed a similar expression pattern to that of Dnmt1. The preimplantation inhibition of DNM by 5-aza-2'-deoxycytodine was not antagonistic for embryonic attachment, although endometrial stromal cell proliferation at the site of implantation was down-regulated, indicating a disturbance with the postattachment decidualization event. Indeed, the peri- or postimplantation inhibition of DNM caused significant abrogation of decidualization, with concomitant loss of embryos. We next identified decidual genes undergoing alteration of DNM using methylation-sensitive restriction fingerprinting. One such gene, Chromobox homolog 4, an epigenetic regulator in the polycomb group protein family, exhibited hypomethylation in promoter DNA and increased expression with the onset of decidualization. Furthermore, inhibition of DNM resulted in enhanced expression of hypermethylated genes (Bcl3 and Slc16a3) in the decidual bed as compared with control, indicating aberration of gene expression may be associated with DNM-inhibition-induced decidual perturbation. Overall, these results suggest that uterine DNM plays a major role for successful decidualization and embryo development during early pregnancy.

Overexpression of cyclin D3 improves decidualization defects in Hoxa-10(-/-) mice.

Uterine decidualization, a crucial process for implantation, is a tightly regulated process encompassing proliferation, differentiation, and polyploidization of uterine stromal cells. Hoxa (Homeobox A)-10, a homeobox transcription factor, is highly expressed in decidualizing stromal cells. Targeted gene deletion experiments have demonstrated marked infertility resulting from severely compromised decidualization in Hoxa-10(-/-) mice. However, the underlying mechanism by which Hoxa-10 regulates stromal cell differentiation remains poorly understood. Cyclin D3, a G(1) phase cell-cycle regulatory protein involved in stromal cell proliferation and decidualization, is significantly reduced in Hoxa-10(-/-) mice. The expression of cyclin D3 in the pregnant mouse uterus parallels stromal cell decidualization. Here, we show that adenovirus-driven cyclin D3 replacement in Hoxa-10(-/-) mice improves stromal cell decidualization. To address our question of whether cyclin D3 replacement in Hoxa-10(-/-) mice can improve decidualization, both in vitro and in vivo studies were completed after the addition of cyclin D3 or empty (control) viral vectors. Immunostaining demonstrated increased proliferation and decidualization in both in vitro and in vivo studies, and in situ hybridization confirmed increased expression of decidualization markers in vivo. Placentation was demonstrated as well in vivo in the cyclin D3-replaced animals. However, fertility was not restored in Hoxa-10(-/-) mice after d 10 of pregnancy. Finally, we identified several downstream targets of cyclin D3 during decidualization in vitro via proteomics experiments, and these were confirmed using in situ hybridization in vivo. Collectively, these results demonstrate that cyclin D3 expression influences a host of genes involved in decidualization and can improve decidualization in Hoxa-10(-/-) mice.

Developmental regulation of decidual cell polyploidy at the site of implantation.

Polyploidy has been reported in several animal cells, as well as within humans; however the mechanism of developmental regulation of this process remains poorly understood. Polyploidy occurs in normal biologic processes as well as in pathologic states. Decidual polyploid cells are terminally differentiated cells with a critical role in continued uterine development during embryo implantation and growth. Here we review the mechanisms involved in polyploidy cell formation in normal developmental processes, with focus on known regulatory aspects in decidual cells.

Decidual cell polyploidization necessitates mitochondrial activity.

Cellular polyploidy has been widely reported in nature, yet its developmental mechanism and function remain poorly understood. In the present study, to better define the aspects of decidual cell polyploidy, we isolated pure polyploid and non-polyploid decidual cell populations from the in vivo decidual bed. Three independent RNA pools prepared for each population were then subjected to the Affymetrix gene chip analysis for the whole mouse genome transcripts. Our data revealed up-regulation of 1015 genes and down-regulation of 1207 genes in the polyploid populations, as compared to the non-polyploid group. Comparative RT-PCR and in situ hybridization results indeed confirmed differential expressional regulation of several genes between the two populations. Based on functional enrichment analyses, up-regulated polyploidy genes appeared to implicate several functions, which primarily include cell/nuclear division, ATP binding, metabolic process, and mitochondrial activity, whereas that of down-regulated genes primarily included apoptosis and immune processes. Further analyses of genes that are related to mitochondria and bi-nucleation showed differential and regional expression within the decidual bed, consistent with the pattern of polyploidy. Consistently, studies revealed a marked induction of mitochondrial mass and ATP production in polyploid cells. The inhibition of mitochondrial activity by various pharmacological inhibitors, as well as by gene-specific targeting using siRNA-mediated technology showed a dramatic attenuation of polyploidy and bi-nucleation development during in vitro stromal cell decidualization, suggesting mitochondria play a major role in positive regulation of decidual cell polyploidization. Collectively, analyses of unique polyploidy markers and molecular signaling networks may be useful to further characterize functional aspects of decidual cell polyploidy at the site of implantation.

LSINCT5 is over expressed in breast and ovarian cancer and affects cellular proliferation.

More than 98% of the human genome is comprised of non-protein coding sequences. Interestingly, a considerable fraction of these sequences is transcribed into non-protein coding RNA transcripts. These transcripts range in size from very small RNAs such as the miRNAs (20-25 base pairs) to transcripts that can range up to 100 kb or more. Some longer non-coding RNAs (lncRNAs) have been found to play important regulatory roles within cells. In this report, we demonstrate that LSINCT5 is a 2.6 Kb polyadenylated, long stress-induced non-coding transcript that is on the negative strand, localized in the nucleus and potentially transcribed by RNA polymerase III. LSINCT5 is overexpressed in breast and ovarian cancer cell lines and tumor tissues, relative to their normal counterpart. In addition, knocking down the expression of LSINCT5 in cancer-derived cell lines causes a decrease in cellular proliferation. Finally, we identified 95 genes with more than 2-fold changes when knocking down LSINCT5 expression by using the Affymetrix U133 Plus 2 array. We chose a subset of these genes to validate using qPCR and found that ten of these genes were indeed significantly affected by the LSINCT5 knockdown. Interestingly, two genes that were significantly downregulated were the lncRNA NEAT-1 and a protein coding gene PSPC1. We have therefore characterized a novel lncRNA that is overexpressed in breast and ovarian cancers, enhances cellular proliferation and may play a significant role in multiple processes.

GPR30 activation opposes estrogen-dependent uterine growth via inhibition of stromal ERK1/2 and estrogen receptor alpha (ERα) phosphorylation signals.

Although estradiol-17ß (E2)-regulated early and late phase uterine responses have been well defined, the molecular mechanisms linking the phases remain poorly understood. We have previously shown that E2-regulated early signals mediate cross talk with estrogen receptor (ER)-α to elicit uterine late growth responses. G protein-coupled receptor (GPR30) has been implicated in early nongenomic signaling mediated by E2, although its role in E2-dependent uterine biology is unclear. Using selective activation of GPR30 by G-1, we show here a new function of GPR30 in regulating early signaling events, including the inhibition of ERK1/2 and ERα (Ser118) phosphorylation signals and perturbation of growth regulation under the direction of E2 in the mouse uterus. We observed that GPR30 primarily localizes in the uterine epithelial cells, and its activation alters gene expression and mediates inhibition of ERK1/2 and ERα (Ser118) phosphorylation signals in the stromal compartment, suggesting a paracrine signaling is involved. Importantly, viral-driven manipulation of GPR30 or pharmacological inhibition of ERK1/2 activation effectively alters E2-dependent uterine growth responses. Overall, GPR30 is a negative regulator of ERα-dependent uterine growth in response to E2. Our work has uncovered a novel GPR30-regulated inhibitory event, which may be physiologically relevant in both normal and pathological situations to negatively balance ERα-dependent uterine growth regulatory functions induced by E2.

An RNA-zipcode-independent mechanism that localizes Dia1 mRNA to the perinuclear ER through interactions between Dia1 nascent peptide and Rho-GTP.

Signal-peptide-mediated ER localization of mRNAs encoding for membrane and secreted proteins, and RNA-zipcode-mediated intracellular targeting of mRNAs encoding for cytosolic proteins are two well-known mechanisms for mRNA localization. Here, we report a previously unidentified mechanism by which mRNA encoding for Dia1, a cytosolic protein without the signal peptide, is localized to the perinuclear ER in an RNA-zipcode-independent manner in fibroblasts. Dia1 mRNA localization is also independent of the actin and microtubule cytoskeleton but requires translation and the association of Dia1 nascent peptide with the ribosome-mRNA complex. Sequence mapping suggests that interactions of the GTPase binding domain of Dia1 peptide with active Rho are important for Dia1 mRNA localization. This mechanism can override the ß-actin RNA zipcode and redirect ß-actin mRNA to the perinuclear region, providing a new way to manipulate intracellular mRNA localization.

MicroRNA expression and regulation in mouse uterus during embryo implantation.

MicroRNAs (miRNAs) are 21-24-nucleotide non-coding RNAs found in diverse organisms. Although hundreds of miRNAs have been cloned or predicted, only very few miRNAs have been functionally characterized. Embryo implantation is a crucial step in mammalian reproduction. Many genes have been shown to be significantly changed in mouse uterus during embryo implantation. However, miRNA expression profiles in the mouse uterus between implantation sites and inter-implantation sites are still unknown. In this study, miRNA microarray was used to examine differential expression of miRNAs in the mouse uterus between implantation sites and inter-implantation sites. Compared with inter-implantation sites, there were 8 up-regulated miR-NAs at implantation sites, which were confirmed by both Northern blot and in situ hybridization. miR-21 was highly expressed in the subluminal stromal cells at implantation sites on day 5 of pregnancy. Because miR-21 was not detected in mouse uterus during pseudopregnancy and under delayed implantation, miR-21 expression at implantation sites was regulated by active blastocysts. Furthermore, we showed that Reck was the target gene of miR-21. Our data suggest that miR-21 may play a key role during embryo implantation.