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The probability distribution of the fluctuating velocity in three dimensions and the near-bed shear stress (NBSS) of a channel with a deposit body were studied using mathematical statistics and the turbulent kinetic energy method, respectively. This analysis of the longitudinal and plane distributions of the NBSS under varying discharges, width ratios and lateral slopes will help further understanding of the laws of sediment incipient motion and transport in channels. It was found that the probability distribution of the near-bed vertical fluctuating velocity was closer to a normal distribution than that in the longitudinal and transversal directions. The NBSS did not change much in the upstream sections of the deposit body, but the difference between the left bank (near the deposit body) and the right bank (far away from the deposit body) was obvious in the sections near the deposit body and downstream. A high shear stress area was located in the transition zone between the main flow and the backflow. Introducing the backwater parameter β, an exponential relationship between the relative NBSS and β was determined, which will help in quick estimates of the NBSS in future practical projects.

The objective of this study was to determine if mouse bone marrow-derived mesenchymal stem cells (BMMSCs) ameliorate preterm birth and perinatal brain injury induced by intrauterine inflammation (IUI). A mouse model of IUI-induced perinatal brain injury at embryonic (E) day 17 was utilized. BMMSCs were derived from GFP-transgenic mice and phenotypically confirmed to be CD44+, Sca-1+, CD45−, CD34−, CD11b−, and CD11c− by flow cytometry and sorted by fluorescence-activated cell sorting (FACS). Dams were assigned to four groups: phosphate-buffered saline (PBS) + PBS, PBS + BMMSCs, lipopolysaccharide (LPS) + PBS, and LPS + BMMSCs. Following maternal IUI, there was a significant increase in CD8+ T cells in the placentas. Maternally administered BMMSCs trafficked to the fetal side of the placenta and resulted in significantly decreased placental CD8+ T cells. Furthermore, fetal trafficking of maternally administered BMMSCs correlated with an improved performance on offspring neurobehavioral testing in LPS + BMMSC group compared with LPS + PBS group. Our data support that maternal administration of BMMSCs can alleviate perinatal inflammation-induced brain injury and improve neurobehavioral outcomes in the offspring via CD8+ T cell immunomodulation at the feto-placental interface.

Objective. To detect the expression levels of microRNA-200a/b (miR-200a/b) in tumor tissues and serum of patients with epithelial ovarian cancer (EOC) and to explore its clinical significance. Methods. A retrospective selection of 30 cases of benign ovarian disease or healthy physical examination (control group) and 55 cases of EOC patients. Real-time quantitative PCR was used to detect the expression level of miR-200a/b in tumor tissues and serum, and the miR-200a/b expresses relevance in the two types of samples were evaluated at the same time. Receiver operating characteristic curve (ROC) and Kaplan-Meier survival analysis were used to evaluate the diagnostic value of miR-200a/b expression and its influence on prognosis, respectively. Results. The serum and tissue miR-200a/b expression levels in EOC patients were higher than those in the control group ( P < 0.001 ), and there was a significant positive correlation between serum and tissue miR-200a/b expression ( R 2 = 0.9419 , P < 0.001 and R 2 = 0.9605 , P < 0.001 ). ROC analysis showed that the expression of serum miR-200a/b can distinguish EOC patients from the control group. In addition, there were significant differences in the TNM stage, tumor differentiation, and lymph node metastasis between the miR-200a/b high- and low-expression groups ( P < 0.05 ). Kaplan-Meier survival analysis found that the overall survival and disease-free survival of patients with high miR-200a/b expression were shorter than those of patients with low miR-200a/b expression ( P < 0.05 ). Conclusion. Upregulation of miR-200a/b expression is a common molecular event in EOC patients, and miR-200a/b can be used as a noninvasive biomarker for the diagnosis and prognosis of EOC.

microRNA-290 (miR-290) clusters are highly expressed in mouse preimplantation embryos, but their specific role and regulatory mechanisms in the development of mouse preimplantation embryos remain unclear. Here, we found that miR-291a-5p and miR-291b-5p, as mature microRNA molecules of miR-290 clusters, were dynamically expressed in mouse preimplantation embryos. The expression of miR-291a-5p and miR-291b-5p in mouse embryos increased during the 2-4-cell stages and was accompanied by the decreasing expression of the autophagy-related genes Atg5 and Becn1 in mRNA. Immunofluorescence studies showed that the formation of autophagosomes and autophagic lysosomes increased in the 1-cell stage, decreased in the 2-cell stage, and rapidly decreased during the 4-8-cell stage. Transmission electron microscopy (TEM) also demonstrated that there were autophagosomes in the cytoplasm of fertilized eggs with a double-layer membrane structure, whereas this structure was not observed in the unfertilized oocyte cytoplasm. Moreover, miR-291a/b-5p inhibited the protein and mRNA expression of Atg5 and Becn1 in NIH/3T3 cells. A dual-luciferase reporter assay confirmed that miR-291a/b-5p directly targeted the Atg5 and Becn1 genes. MiR-291a/b-5p repressed rapamycin-induced autophagy-related LC3-I to LC3-II conversion, ultimately inhibiting the formation of autophagosomes. Furthermore, the microinjection of mouse zygote cytoplasm with miR-291a-5p inhibitors increased the mRNA expression of Atg5 and Becn1 in mouse embryos and facilitated the first cleavage of mouse embryos and blastocyst formation. Our results suggest the important role of miR-291a/b-5p during mouse preimplantation embryo development.

Fluorogenic labeling is a potential technique in biology that allows for direct detection and tracking of cells undergoing various biological processes. Compared to traditional genetic modification approaches, labeling cells with nanoparticles has advantages, especially for the additional safety they provide by avoiding genomic integration. However, it remains a challenge to determine whether nanoparticles interfere with cell traits and provide long-lasting signals in living cells. We employed an amphiphilic fluorophore-derived nanoparticle (denoted by TPE-11) bearing a tetraphenylethene (TPE) moiety and two ionic heads; this nanoparticle has an aggregation-induced emission (AIE) effect and the ability to self-assemble. TPE-11 exhibited the property of higher or longer fluorescence intensities in cell imaging than the other two nanomaterials under the same conditions. We used this nanomaterial to label human embryonic stem (hES) cells and monitor their differentiation. Treatment with low concentrations of TPE-11 (8.0 μg/mL) resulted in high-intensity labeling of hES cells, and immunostaining analysis and teratoma formation assays showed that at this concentration, their pluripotency remained unaltered. TPE-11 nanoparticles allowed for long-term monitoring of hES cell differentiation into neuron-like cells; remarkably, strong nanoparticle signals were detected throughout the nearly 40-day differentiation process. Thus, these results demonstrate that the TPE-11 nanoparticle has excellent biocompatibility for hES cells and is a potential fluorogen for labeling and tracking the differentiation of human pluripotent stem cells. Statement of Significance This study uses a nanoparticle-based approach to label human embryonic stem (hES) cells and monitor their differentiation. hES cells are distinguished by two distinctive properties: the state of their pluripotency and the potential to differentiate into various cell types. Thus, these cells will be useful as a source of cells for transplantation or tissue engineering applications. We noticed the effect of aggregation-induced emission, and the ability to self-assemble could enhance the persistence of signals. Treatment with low concentrations of TPE-11 nanoparticles showed high-intensity labeling of hES cells, and immunostaining analysis and teratoma formation assays showed that at this concentration, their pluripotency remained unaltered. Additionally, these nanoparticles allowed for long-term monitoring of hES cell differentiation into neuron-like cells lasting for 40 days.

Comprehensive epidemiological surveys for Lyme disease have not been conducted for the Bactrian camel in China. In this study, a total of 138 blood specimens collected from Bactrian camels from Zhangye City in Gansu Province and Yili and Aksu in Xinjiang Province, China, were examined for the presence of Borrelia spp. Species-specificity nested PCR based on the 5S-23S rRNA, OspA, flaB and 16S rRNA genes revealed that the total positive rate of Borrelia spp. was 3.6% (5/138, 95% CI = 0.2–17.9). These results were confirmed by sequence analysis of the positive PCR products or positive colonies. This is the first report of Borrelia pathogens in camels in China. Two Borrelia species that cause Lyme disease and one that causes relapsing fever were identified in the camel blood samples by sequencing. The findings of this study indicate that the Bactrian camel may serve as a potential natural host of Lyme disease and/or relapsing fever in China., Highlights • This is the first report of Borrelia pathogens using molecular tools in Bactrian camel. • Bactrian camel may play as potential natural host of Lyme disease in China. • The results revealed that the prevalence of Borrelia spp. were 3.6% based on sequences of four genes. • A novel Borrelia genespecies belong to Relapsing fever Borrelia spp. was found.

Fetal brain injury induced by intrauterine inflammation is a major risk factor for adverse neurological outcomes, including cerebral palsy, cognitive dysfunction, and behavioral disabilities. There are no adequate therapies for neuronal protection to reduce fetal brain injury, especially new strategies that may apply promptly and conveniently. In this study, we explored the effect of maternal glucose administration in a mouse model of intrauterine inflammation at term. Our results demonstrated that maternal glucose supplementation significantly increased survival birth rate and improved the neurobehavioral performance of pups exposed to intrauterine inflammation. Furthermore, we demonstrated that maternal glucose administration improved myelination and oligodendrocyte development in offspring exposed to intrauterine inflammation. Though the maternal blood glucose concentration was temporally prevented from decrease induced by intrauterine inflammation, the glucose concentration in fetal brain was not recovered by maternal glucose supplementation. The adenosine triphosphate (ATP) level and autophagy in fetal brain were regulated by maternal glucose supplementation, which may prevent dysregulation of cellular metabolism. Our study is the first to provide evidence for the role of maternal glucose supplementation in the cell survival of fetal brain during intrauterine inflammation and further support the possible medication with maternal glucose treatment.

Background: Despite the great potential of utilizing human embryonic stem cells (hESCs)-derived cells as cell source for transplantation, these cells were often rejected during engraftment by the immune system due to adaptive immune response. Methods: We first evaluated HLA-G expression level in both hESCs and differentiated progenitor cells. After that, we generated modified hESC lines that over-express HLA-G1 using lentiviral infection with the construct contains both HLA-G1 and GFP tag. The lentivirus was first produced by co-transfecting HLA-G1 expressing lentiviral vector together with packaging vectors into packaging cell line 293T. Then the produced virus was used for the infection of selected hESC lines. We characterized the generated cell lines phenotype, including pluripotency and self-renewal abilities, as well as immune tolerance ability by mixed lymphocyte reaction (MLR) and cytotoxicity assays. Results: Although the hESCs do not express high levels of HLA-G1, over-expression of HLA-G1 in hESCs still retains their stem cell characteristics as determined by retaining the expression levels of OCT4 and SOX2, two critical transcriptional factors for stem cell function. Furthermore, the HLA-G1 overexpressing hESCs retain the self-renewal and pluripotency characteristics of stem cells, which can differentiate into different types of cells, including pigment cells, smooth muscle cells, epithelia-like cells, and NPCs. After differentiation, the differentiated cells including NPCs retain the high levels of HLA-G1 protein. In comparison with conventional NPCs, these HLA-G1 positive NPCs have enhanced immune tolerance ability. Conclusions: Ectopic expression of HLA-G1, a non-classical major histocompatibility complex class I (MHC I) antigen that was originally discovered involving in engraftment tolerance during pregnancy, can enhance the immunological tolerance in differentiated neural progenitor cells (NPCs). Our study shows that stably overexpressing HLA-G1 in hESCs might be a feasible strategy for enhancing the engraftment of NPCs during transplantation.

The objective of this study was to determine if mouse bone marrow-derived mesenchymal stem cells (BMMSCs) ameliorate preterm birth and perinatal brain injury induced by intrauterine inflammation (IUI). A mouse model of IUI-induced perinatal brain injury at embryonic (E) day 17 was utilized. BMMSCs were derived from GFP-transgenic mice and phenotypically confirmed to be CD44

This paper proposes a real-time, continuous pedestrian tracking and positioning method with multiple coordinated overhead-view cameras. First, a discriminative correlation filter with detection and space reliability (DSR-DCF) is presented for continuous, real-time pedestrian tracking with single cameras. Then, the frame difference method is employed to detect moving pedestrians when they walk from one camera’s view to another’s. Motion-area images are cropped to establish a pedestrian image library (PIL). Finally, this PIL is relied upon to reidentify pedestrians. Pedestrian positioning is realized based on the pinhole imaging model. Tracking and positioning experiments showed the following. DSR-DCF is stable with a high success rate. Its tracking speed exceeds 24 fps. It produces a success rate higher than 90% for cross-camera pedestrian tracking and misses no pedestrians. It yields stable pedestrian positioning results with an accuracy better than 0.2 m. The proposed algorithm will be used in airport terminals.

Bovine theileriosis, a tick-borne protozoan disease caused by Theileria annulata, Theileria orientalis and Theileria sinensis, is widespread in China and is a serious economic problem for the Chinese livestock industry. In this study, recombinant major piroplasma surface proteins (MPSP) of T. annulata, T. orientalis and T. sinensis based on MPSP genes were expressed in Escherichia coli BL21(DE3). The immunogenicity and specificity of the three purified recombinant MPSP proteins were evaluated with the reference positive sera of T. annulata, T. orientalis, T. sinensis, Babesia bovis, B abesia bigemina, Babesia major, Babesia motasi, Theileria luwenshuni, Theileria uilenbergi and Anaplasma ovis using an enzyme-linked immunosorbent assay (ELISA) or western blotting. The results showed that all three of the rMPSP proteins had a strong reaction with the sera from cattle infected with T. annulata, T. orientalis and T. sinensis via western blotting but not with other piroplasma and Anaplasma species. Then, the rMPSP protein of T. sinensis was used to develop an iELISA for detecting the three Theileria species infections. The specificity and sensitivity were 95.7 and 95.5 %, respectively, with a threshold of 28.8 % of the specific mean antibody rate (AbR). Finally, 2473 field-collected bovine sera, from 42 prefectures of 17 provinces in China, were tested using the ELISA to evaluate the prevalence of bovine theileriosis, and the average positive rate was 43.6 %. The developed iELISA could be a suitable tool to detect the three bovine Theileria species, and the data also provided important information regarding the current prevalence of bovine theileriosis in China.

Theileria annulata is a tick-borne intracellular protozoan parasite that causes tropical theileriosis, a fatal bovine lymphoproliferative disease. The parasite predominantly invades bovine B lymphocytes and macrophages and induces host cell transformation by a mechanism that is not fully comprehended. Analysis of signaling pathways by quantitative real-time PCR (qPCR) could be a highly efficient means to understand this transformation mechanism. However, accurate analysis of qPCR data relies on selection of appropriate reference genes for normalization, yet few papers on T. annulata contain evidence of reference gene validation. We therefore used the geNorm and NormFinder programs to evaluate the stability of 5 candidate reference genes; 18S rRNA, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), ACTB (β-actin), PRKG1 (protein kinase cGMP-dependent, type I) and TATA box binding protein (TBP). The results showed that 18S rRNA was the reference gene most stably expressed in bovine PBMCs transformed and non-transformed with T. annulata, followed by GAPDH and TBP. While 18S rRNA and GAPDH were the best combination, these 2 genes were chosen as references to study signaling pathways involved in the transformation mechanism of T. annulata.

Trophoblast cells are important in embryo implantation and fetomaternal tolerance. HLA-G is specifically expressed at the maternal–fetal interface and is a regulator in pregnancy. The aim of the present study was to detect the effect of HLA-G1 on trophoblast cell proliferation, adhesion, and invasion. Human trophoblast cell lines (JAR and HTR-8/SVneo cells) were infected with HLA-G1-expressing lentivirus. After infection, HLA-G1 expression of the cells was detected by western blotting. Cell proliferation was detected by the BrdU assay. The cell cycle and apoptosis of JAR and HTR-8/SVneo cells was measured by flow cytometry (FCM). The invasion of the cells under different conditions was detected by the transwell invasion chamber assay. HLA-G1 didn't show any significant influence on the proliferation, apoptosis, adhesion, and invasion of trophocytes in normal culture conditions. However, HLA-G1 inhibited JAR and HTR-8/SVneo cells invasion induced by hepatocyte growth factor (HGF) under normal oxygen conditions. In conditions of hypoxia, HLA-G1 couldn't inhibit the induction of cell invasion by HGF. HLA-G1 is not an independent factor for regulating the trophocytes. It may play an indirect role in embryo implantation and formation of the placenta.

Preeclampsia is major cause of maternal and fetal morbidity and mortality. Currently, the etiology of preeclampsia is unclear. In this study, we investigated differences in gene expression between preeclampsia patients and controls using partial least squares-based analysis, which is more suitable than routine analysis. Expression profile data were downloaded from the Gene Expression Omnibus database. A total of 503 genes were found to be differentially expressed, including 248 downregulated genes and 255 overexpressed genes. Network analysis identified 5 hub genes, including UBB, PIK3R1, MAPRE1, VEGFA, and ITGB1. Three of these, PIK3R1, VEGFA, and ITGB1, are known to be associated with preeclampsia or preeclampsia-related biological processes. Our findings shed light on expression signatures of preeclampsia patients that can be used as theoretical support in future therapeutic studies.

PROBLEM: We investigated the mechanisms by which CD8(+) T cell trafficking in placenta contributes to perinatal brain injury by studying effects of maternal CD8(+) T cell depletion (DEP) in a mouse model of intrauterine inflammation (IUI). METHOD OF STUDY: Maternal CD8(+) T cells were depleted with anti-CD8(+) antibodies. IUI was induced with lipopolysaccharide (LPS). DEP was confirmed using flow cytometry. Preterm birth rate was evaluated. Offspring neurologic sequelae were assessed by Nissl staining, immune arrays, confirmatory individual TaqMan(®) gene assays, and neurobehavioral tests. RESULTS: DEP did not significantly prevent LPS-induced preterm birth, but improved neurobehavioral performance (p

The technology to reprogram human somatic cells back to pluripotency allows the production of patient-specific induced pluripotent stem cells (iPSCs) and holds a great promise for regenerative medicine. Choosing the most suitable cell type for induction and reducing the risk of viral transgene activation, especially oncogene activation, are important for iPSC research. To date, human dermal fibroblasts (HDFs) are the most frequent cell source used for iPSC generation, but they have several limitations. An invasive skin biopsy must be performed to obtain HDFs, and HDFs must be cultured for a prolonged period before they can be used for experiments. Thus, in an effort to develop a suitable source for iPSC studies to avoid the limitations mentioned above, we have here identified stromal cells derived from menstrual blood (MenSCs) as suitable candidates. In the present study, we found that MenSCs can be reprogrammed to pluripotent status by doxycycline-inducible lentiviral transduction of OCT4, SOX2, and KLF4. Additionally, we found that MenSCs have a significantly higher reprogramming efficiency than HDFs. The combination of OCT4 and SOX2 is sufficient to reprogram MenSCs into iPSCs without the use of c-MYC or KLF4. The resulting MenSC-iPSCs showed the same characteristics as human embryonic stem cells with regard to morphology, pluripotent markers, gene expression, and the epigenetic status of pluripotent-cell-specific genes. These cells were able to differentiate into various cell types of all 3 germ layers both in vitro and in vivo. Therefore, MenSCs may be a preferred candidate for generation of iPSCs.

Human leukocyte antigen (HLA)-G is thought to confer fetal–maternal tolerance and play a crucial role in ensuring a successful pregnancy. There is increasing evidence that HLA-G is regulated at the post-transcriptional level. This study investigated the role of miR-133a in regulating HLA-G expression and the pathogenesis of recurrent spontaneous abortion (RSA). Twelve patients (25–30years) with RSA at 7 gestational weeks were screened by array-based comparative genome hybridization: 16.7% were found to have an abnormal karyotype and all induced abortion (IA) patients had normal karyotype. The villi of RSA and IA patients with normal karyotype were further screened by miRNA microarrays. Multi-software prediction and real-time PCR confirmed that miR-133a was most likely to bind to HLA-G 3′ untranscribed region (UTR). Relevance analysis showed that, compared with IA villi, miR-133a was greatly overexpressed in RSA villi with normal karyotype ( P Human leukocyte antigen (HLA)-G is supposed to confer fetal–maternal tolerance and plays a crucial role in successful pregnancy. Accumulating evidence indicates that HLA-G is mainly regulated at the post-transcriptional level. Our study aimed to further investigate the key role of a microRNA, miR-133a, in regulating HLA-G expression and the pathogenesis of recurrent spontaneous abortion (RSA). Our findings indicated that miR-133a regulates HLA-G expression by inhibiting its translation, and suggest that it is involved in the pathogenesis of RSA.

Triptolide, a compound extracted from the traditional Chinese medicine preparation of Tripterygium wilfordii Hook F., has been reported to have anti-inflammatory and anti-cancer activities. However, its effect on ovarian cancer invasion is unknown. We observed that MMP7 and MMP19 expression increased in ovarian cancer tissue. Triptolide treatment inhibited the migration and invasion of ovarian cancer cells SKOV3 and A2780 at the concentration of 15 nM. We also observed that triptolide suppressed MMP7 and MMP19 promoter activity in a dose-dependent manner, down-regulating the expressions of these promoters on mRNA and protein level. Moreover, triptolide enhanced E-cadherin expression in ovarian cancer cells. In vivo, triptolide inhibited tumor formation and metastasis in nude mice, and suppressed MMP7 and MMP19 expression; it also enhanced E-cadherin expression in tumor in a dose-dependent manner. Over expression of MMP7 and MMP19, or suppression of E-cadherin expression partially abolished the inhibitory effect of triptolide on invasion of ovarian cancer cells. To summarize, triptolide significantly inhibited the migration and invasion of ovarian cancer cells by suppression of MMP7 and MMP19 and up-regulation of E-cadherin expression. This study shows that triptolide is a good candidate for the treatment of ovarian cancer and reduction of metastasis.

The goals of this study were to analyze the change in the global gene expression profile of exogenous human leukocyte antigen-G1 (HLA-G1) overexpressed in human embryonic stem (hES) cells and to explore the molecular mechanism by which the overexpression of HLA-G1 modifies immunologic pathways. Microarray and quantitative real-time PCR analyses were performed to quantify the differential expression pattern of HLA-G1 + H1 hES cells. The results showed that HLA-G1 differentially regulated the expression of 425 genes with at least a twofold increase or decrease. These differentially expressed genes were classified into 13 functional groups, including cellular components, biological processes, and molecular functions. The pathways of focal adhesion, the TGF-β signaling pathway, and the immune response were the most predominantly affected. The synergism of these genes could explain the mechanism of the immunosuppression of HLA-G1 + H1 hES cells. Thus, the expression pattern reflected a broad spectrum of roles of HLA-G1 in hES cells.

We investigated the mechanisms by which CD8+ T-cell trafficking in placenta contributes to perinatal brain injury by studying effects of maternal CD8+ T-cell depletion (DEP) in a mouse model of intrauterine inflammation (IUI). Maternal CD8+ T cells were depleted with anti-CD8+ antibodies. IUI was induced with lipopolysaccharide (LPS). DEP was confirmed using flow cytometry. Preterm birth rate was evaluated. Offspring neurologic sequelae were assessed by Nissl staining, immune arrays, confirmatory individual TaqMan® gene assays, and neurobehavioral tests. DEP did not significantly prevent LPS-induced preterm birth but improved neurobehavioral performance (P

The equilibrium structures and relative stabilities of BN substituted fullerenes C60−2x(BN)x (x=1–3) have been studied using AM1 and MNDO semi-empirical methods. The calculation results obtained by both methods indicated that the BN substituted fullerenes are less stable than C60. The homo–lumo splitting and heat of formation suggests that the isomer of C58BN in which two neighboring carbon atoms between the six- and six- membered ring are substituted by BN units is the most stable species. The stabilities of C58BN decrease with increasing the distance between the heteroatoms. For C56(BN)2, the calculation results show that N–N and B–B bonds should be avoided and the smallest number of the C–X bonds is preferred for the stable isomer of C56(BN)2 and the isomer in which the B–N–B–N bond is formed has the lowest heat of formation, thus the most stable species. The structure of the most stable isomer of C54(BN)3 has been proposed based on the calculations on various isomers of C58BN and C56(BN)2, i.e. the most energetic favorable C54(BN)3 should have three BN units located in the same hexagon to form B–N–B–N–B–N ring. The BN substituted fullerenes C60−2x(BN)x have somewhat smaller ionization potentials and bigger affinity potentials compared with C60, which suggests that it is easier to oxidize and reduce C60−2x(BN)x relative to C60, thus the redox characteristics of C60 can be enhanced by doping. Based on the optimized geometries, the electronic spectra for these BN doped fullerenes have been calculated using INDO/CIS method

The equilibrium structures, relative stabilities of substituted fullerenes C 58 B 2 , C 58 N 2 and C 58 BN have been studied at the AM1, PM3, MNDO and INDO level. The calculation results obtained by all these semi-emperical methods show that the substituted fullerenes are less stable than C 60 , 6–6 isomers are the most stable species for C 58 B 2 , C 58 N 2 and C 58 BN, the stabilities decrease with the increase of the distance between the heteroatoms. The calculation results also show that the stability of C 58 BN is considerably higher than that of C 58 N 2 , and the stability of C 58 N 2 is considerably higher than that of C 58 B 2 . Calculations on the C 58 B 2 -(NH 3 ) 2 adducts have been employed in this paper to evaluate Smally's proposition on the substitute preference of C 58 B 2 . The elctronic spectra for these doped fullerenes have been calculated using INDO/CIS method

Summary To search for a method for treatment of bilateral temporal lobe epilepsy (BTLE), we report one patient with BTLE experienced bilateral stereotactic radiofrequency amygdalohippocampectomy (SAHE). Neuropsychological examinations were performed before and 5 days, and 6, 18, and 48 months after operation. No seizure occurred in the follow-up time, and no long-term memory and intelligence deficits were found except for a transient decline of the scores immediately after operation. Because severe damage of memory could be caused by bilateral resection surgery, bilateral SAHE should be considered as a possible approach for the treatment of BTLE. However, further studies with accumulation of cases are needed, especially in the detailed assessment of neuropsychological function.

The human leukocyte antigen G (HLA-G) is expressed on the fetal-maternal interface and plays a role in protecting fetal-derived trophoblasts from the maternal immune response, allowing trophoblasts to invade the uterus. However, HLA-G also possesses immune suppressing-independent functions. We found that HLA-G expressing BeWo choriocarcinoma cells increased cell-cell fusion compared to control BeWo cells under forskolin treatment. Regardless of forskolin treatment, the expression of fusogenic gene mRNAs, including syncytin-1, the transcription factor glial cell missing 1 (Gcm1), and beta human chorionic gonadotropin (β-hCG) were elevated. HLA-G up-regulates β-hCG production in human choriocarcinoma cells because HLA-G knockdown in JEG-3 cells induces a dramatic decrease in β-hCG compared with control cells. The defect in β-hCG production in HLA-G knocked-down cells could not be completely overcome by stimulating hCG production through increasing intracellular cAMP levels. HLA-G expressing cells have increased phosphorylation levels for extracellular signal-regulated kinase1/2 (Erk1/2) in BeWo cells. The Erk1/2 pathway is inactivated after the inhibition of HLA-G expression in JEG-3 cells. Finally, Erk1/2 inhibition was able to suppress the increased hCG production induced by HLA-G expression. Together, these data suggest novel roles for HLA-G in regulating β-hCG production via the modulation of the Erk1/2 pathway and by inducing trophoblast cell fusion.

Human embryonic stem (hES) cells have the capacity for self-renewal and exhibit multipotentiality. hES cells have promise for serving as an unlimited source of ideal seed cells for cell transplantation. However, the rejection that occurs between the transplant recipient and the transplanted cell poses a major challenge for therapeutic transplantation. This study was designed to devise methods to enhance immune tolerance in cell therapy. We established an hES cell line that could stably express human leukocyte antigen-G1 (HLA-G1). The established HLA-G1-H1 hES cells still retained all the characteristics of normal human embryonic stem cells. By using the SDIA method, we induced dopaminergic (DA) neurons by coculturing HLA-G1-H1 hES cells with the mouse stromal cell line PA6. Tyrosine hydroxylase (TH)+neurons were detected on the 10th day of differentiation, and 70% of the HLA-G1-H1 hES cells were TH+mature DA neurons because the differentiation time was only 3 weeks. Cells that had been differentiating for different periods of time still expressed HLA-G1, and these differentiated DA neurons released dopamine and other catecholamines in response to K+ depolarization as measured by HPLC. After careful study, we found that HLA-G1-H1 hES cells are capable of inhibiting the proliferation of mixed T-lymphocytes. DA neurons derived from HLA-G1-H1 hES attenuated the release of proinflammatory cytokines IL-1β and IFN-γ from lipopolysaccharide (LPS)-stimulated BV2 microglia. The efficiency of inhibition was significant and dose-dependent. This method might be used to treat Parkinson's patients via cell transplantation.

Induced pluripotent stem (iPS) cells have been generated from human somatic cells by ectopic expression of defined transcription factors. Application of this approach in human cells may have enormous potential to generate patient-specific pluripotent stem cells. However, traditional methods of reprogramming in human somatic cells involve the use of oncogenes c-MYC and KLF4, which are not applicable to clinical translation. In the present study, we investigated whether human fetal gut mesentery-derived cells (hGMDCs) could be successfully reprogrammed into induced pluripotent stem (iPS) cells by OCT4, SOX2, and NANOG alone. We used lentiviruses to express OCT4, SOX2, NANOG, in hGMDCs, then generated iPS cells that were identified by morphology, presence of pluripotency markers, global gene expression profile, DNA methylation status, capacity to form embryoid bodies (EBs), and terotoma formation. iPS cells resulting from hGMDCs were similar to human embryonic stem (ES) cells in morphology, proliferation, surface markers, gene expression, and epigenetic status of pluripotent cell-specific genes. Furthermore, these cells were able to differentiate into cell types of all three germ layers both in vitro and in vivo, as shown by EB and teratoma formation assays. DNA fingerprinting showed that the human iPS cells were derived from the donor cells, and are not a result of contamination. Our results provide proof that hGMDCs can be reprogrammed into pluripotent cells by ectopic expression of three factors (OCT4, SOX2, and NANOG) without the use of oncogenes c-MYC and KLF4.

Reprogramming human somatic cells to pluripotency represents a valuable resource for research aiming at the development of in vitro models for human diseases and regenerative medicines to produce patient-specific induced pluripotent stem (iPS) cells. Seeking appropriate cell resources for higher efficiency and reducing the risk of viral transgene activation, especially oncogene activation, are of significance for iPS cell research. In this study, we tested whether human amnion-derived cells (hADCs) could be rapidly and efficiently reprogrammed into iPS cells by the defined factors: OCT4/SOX2/NANOG. hADCs from normal placenta were isolated and cultured. The 3rd passage cells were infected with the lentiviral vectors for the delivery of OCT4, SOX2, and NANOG. Afterwards, the generated iPSCs were identified by morphology, pluripotency markers, global gene expression profiles, and epigenetic status both in vitro and in vivo. The results showed that we were able to reprogram hADCs by the defined factors (OCT4/SOX2/NANOG). The efficiency was significantly high (about 0.1%), and the typical colonies appeared on the 9th day after infection. They were similar to human embryonic stem (ES) cells in morphology, proliferation, surface markers, gene expression, and the epigenetic status of pluripotent cell-specific genes. Furthermore, these cells were able to differentiate into various cell types of all three germ layers both in vitro and in vivo. These results demonstrate that hADCs were an ideal somatic cell resource for the rapid and efficient generation of iPS cells by OCT4/SOX2/NANOG.

Nonsteroidal anti-inflammatory drugs (NSAIDs) can inhibit cell growth and metastasis, and induce cell apoptosis in cancerous cells. They have been shown to reduce incidence and mortality of gastric cancer by an unknown mechanism. NSAIDs often exert their effects by Cox-2 inhibition, and Cox-2 is overexpressed in gastric cancer cells. Nevertheless, when gastric cancer cells were treated with different NSAIDs, the non-Cox-2-inhibiting R-flurbiprofen was most effective at reducing proliferation of gastric cancer cells in vitro. R-Flurbiprofen prevented the metastatic characteristics of gastric cancer cells in vitro, and reduced tumor size and metastasis in vitro, when gastric cancer cells were injected into nude mice. R-Flurbiprofen also affected multidrug resistance, increasing the sensitivity of resistant gastric cancer cells to chemotherapeutic agents. Mechanistically, R-flurbiprofen was found to have pleiotropic effects, changing levels of cell cycle factors like Cyclin D1 and CKD4, apoptotic protwins like caspase3 and Bcl-2, and protwins that affect metastasis, like metalloproteases. Consistent with reports on other cancer cell types, NSAID treatment with R-flurbiprofen increased levels of the tumor suppressor neurotrophin receptor (p75(NTR)) in gastric cancer cells. The anticancer effects of R-flurbiprofen were found to require induction of p75(NTR) via the p38 signaling pathway, suggesting a possible mechanism of action.

OCT4 is a pivotal transcription factor in maintaining the pluripotency and self-renewal capacities of embryonic stem (ES) cells. Human OCT4 can generate two isoforms by alternative splicing, termed OCT4A and OCT4B. OCT4A confers the stemness properties of ES cells, whereas the function of OCT4B is unknown. We present here the diverse protein products and a novel function of OCT4 gene. A single OCT4B mRNA can encode three isoforms by alternative translation initiation at AUG and CUG start codons, respectively. A putative internal ribosome entry site (IRES) has been identified in OCT4B mRNA accounting for the translation mechanism. The OCT4B-190 is upregulated under stress conditions and it may protect cell against apoptosis under stress. This work evokes the significance to distinguish the biological function of the protein products of OCT4. The OCT4 gene, by the regulation of alternative splicing and alternative translation initiation, may carry out more crucial roles in many biological events. Disclosure of potential conflicts of interest is found at the end of this article.

Expression profile of microRNA (miRNA) in mouse oocytes and preimplantation embryos has been revealed by a novel high throughput microarray assay. A total of 97 (43 “new” and 54 known) including mouse, human, and predicted miRNAs have been discovered in the preimplantation mouse embryos which can be classified into developmental stage-dependent groups and non–stage-dependent group according to the statistical analysis of the expression patterns. Potential gene targets of each group of miRNAs are estimated by TargetsScan system and significantly changed signaling pathways and biological processes underlying these gene targets are searched by PANTHER classification system between the stage-dependent miRNAs and the non–stage-dependent miRNAs. Expression of some miRNAs is confirmed by reverse transcriptase-polymerase chain reaction. It is shown that dynamic synthesis and degradation of miRNAs coexists in the preimplantation development of mouse embryos. However, the overall quantity of miRNAs and percentage of the stage-dependent miRNAs increase as the preimplantation embryos develop. Developmental Dynamics 237:2315–2327, 2008. © 2008 Wiley-Liss, Inc.

DNA methylation disturbance is associated with defective human sperm. However, oligozoospermia (OZ) and asthenozoospermia (AZ) usually present together, and the relationship between the single-phenotype defects in human sperm and DNA methylation is poorly understood. In this study, 20 infertile OZ patients and 20 infertile AZ patients were compared with 20 fertile normozoospermic men. Bisulfate-specific PCR was used to analyze DNA methylation of the H19-DMR and the DAZL promoter in these subjects. A similar DNA methylation pattern of the H19-DMR was detected in AZ and NZ(control), with only complete methylation and mild hypomethylation(0.05). However, the methylation pattern of severe hypomethylation (>50% unmethylated CpGs ) and complete unmethylation was only detected in 5 OZ patients, and the occurrence of these two methylation patterns was 8.54±10.86% and 9±6.06%, respectively. Loss of DNA methylation of the H19-DMR in the OZ patients was found to mainly occur in CTCF-binding site 6, with occurrence of 18.15±14.71%, which was much higher than that in patients with NZ (0.84±2.05%) and AZ (0.58±1.77%) (P20% methylated clones in the DAZL promoter only in infertile patients, there was no significant difference between the AZ and OZ patients in the proportion of moderately-to-severely hypermethylated clones (p>0.05). In all cases, global sperm genome methylation analyses, using LINE1 transposon as the indicator, showed that dysregulation of DNA methylation is specifically associated with the H19-DMR and DAZL promoter. Therefore, abnormal DNA methylation status of H19-DMR, especially at the CTCF-binding site 6, is closely associated with OZ. Abnormal DNA methylation of the DAZL promoter might represent an epigenetic marker of male infertility.