Overexpression of copper zinc superoxide dismutase impairs human trophoblast cell fusion and differentiation.: SOD-1 and Human Trophoblast Differentiation

The syncytiotrophoblast is the major component of the human placenta, involved in feto-maternal exchanges and secretion of pregnancy-specific hormones. Multinucleated syncytiotrophoblast arises from fusion of mononuclear cytotrophoblast cells. In trisomy 21-affected placentas, we recently have shown that there is a defect in syncytiotrophoblast formation and a decrease in the production of pregnancy-specific hormones. Due to the role of oxygen free radicals in trophoblast cell differentiation, we investigated the role of the key antioxidant enzyme, copper/zinc superoxide dismutase, encoded by chromosome 21 in in vitro trophoblast differentiation. We first observed that overexpression of superoxide dismutase in normal cytotrophoblasts impaired syncytiotrophoblast formation. This was associated with a significant decrease in mRNA transcript levels and secretion of hCG and other hormonal markers of syncytiotrophoblast. We confirmed abnormal cell fusion by overexpression of green fluorescence protein-tagged superoxide dismutase in cytotrophoblasts. In addition, a significant decrease in syncytin transcript levels was observed in superoxide dismutase-transfected cells. We then examined superoxide dismutase expression and activity in isolated trophoblast cells from trisomy 21-affected placentas. Superoxide dismutase mRNA expression (P < 0.05), protein levels (P < 0.01), and activity (P < 0.05) were significantly higher in trophoblast cells isolated from trisomy 21-affected placentas than in those from normal placentas. These results suggest that superoxide dismutase overexpression may directly impair trophoblast cell differentiation and fusion, and superoxide dismutase overexpression in Down's syndrome may be responsible at least in part for the failure of syncytiotrophoblast formation observed in trisomy 21-affected placentas

Biochemical characterization and modulation of LH/CG-receptor during human trophoblast differentiation.: LH/CG-R in human trophoblast differentiation.

Due to the key role of the human chorionic gonadotropin hormone (hCG) in placental development, the aim of this study was to characterize the human trophoblastic luteinizing hormone/chorionic gonadotropin receptor (LH/CG-R) and to investigate its expression using the in vitro model of human cytotrophoblast differentiation into syncytiotrophoblast. We confirmed by in situ immunochemistry and in cultured cells, that LH/CG-R is expressed in both villous cytotrophoblasts and syncytiotrophoblasts. However, LH/CG-R expression decreased during trophoblast fusion and differentiation, while the expression of hCG and hPL (specific markers of syncytiotrophoblast formation) increased. A decrease in LH/CG-R mRNA during trophoblast differentiation was observed by means of semi-quantitative RT-PCR with two sets of primers. A corresponding decrease ( approximately 60%) in LH/CG-R protein content was shown by Western-blot and immunoprecipitation experiments. The amount of the mature form of LH/CG-R, detected as a 90-kDa band specifically binding (125)I-hCG, was lower in syncytiotrophoblasts than in cytotrophoblasts. This was confirmed by Scatchard analysis of binding data on cultured cells. Maximum binding at the cell surface decreased from 3,511 to about 929 molecules/seeded cells with a kDa of 0.4-0.5 nM. Moreover, on stimulation by recombinant hCG, the syncytiotrophoblast produced less cyclic AMP than cytotrophoblasts, indicating that LH/CG-R expression is regulated during human villous trophoblast differentiation. J. Cell. Physiol. 212: 26-35, 2007. (c) 2007 Wiley-Liss, Inc

Implication d'une forme d'hCG anormale et de son récepteur (R-LH/CG) dans le développement placentaire lors d'une grossesse associée à une trisomie 21

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Dynamics of transcriptional (re)-programming of syncytial nuclei in developing muscles

Abstract Background A stereotyped array of body wall muscles enables precision and stereotypy of animal movements. In Drosophila, each syncytial muscle forms via fusion of one founder cell (FC) with multiple fusion competent myoblasts (FCMs). The specific morphology of each muscle, i.e. distinctive shape, orientation, size and skeletal attachment sites, reflects the specific combination of identity transcription factors (iTFs) expressed by its FC. Here, we addressed three questions: Are FCM nuclei naive? What is the selectivity and temporal sequence of transcriptional reprogramming of FCMs recruited into growing syncytium? Is transcription of generic myogenic and identity realisation genes coordinated during muscle differentiation? Results The tracking of nuclei in developing muscles shows that FCM nuclei are competent to be transcriptionally reprogrammed to a given muscle identity, post fusion. In situ hybridisation to nascent transcripts for FCM, FC-generic and iTF genes shows that this reprogramming is progressive, beginning by repression of FCM-specific genes in fused nuclei, with some evidence that FC nuclei retain specific characteristics. Transcription of identity realisation genes is linked to iTF activation and regulated at levels of both transcription initiation rate and period of transcription. The generic muscle differentiation programme is activated independently. Conclusions Transcription reprogramming of fused myoblast nuclei is progressive, such that nuclei within a syncytial fibre at a given time point during muscle development are heterogeneous with regards to specific gene transcription. This comprehensive view of the dynamics of transcriptional (re)programming of post-mitotic nuclei within syncytial cells provides a new framework for understanding the transcriptional control of the lineage diversity of multinucleated cells

Angiogenin Expression during Early Human Placental Development; Association with Blood Vessel Formation.

17 pagesInternational audienceThe placenta is a transient organ essential for fetal development. During human placental development, chorionic villi grow in coordination with a large capillary network resulting from both vasculogenesis and angiogenesis. Angiogenin is one of the most potent inducers of neovascularisation in experimental models in vivo. We and others have previously mapped angiogenin expression in the human term placenta. Here, we explored angiogenin involvement in early human placental development. We studied, angiogenin expression by in situ hybridisation and/or by RT-PCR in tissues and primary cultured trophoblastic cells and angiogenin cellular distribution by coimmunolabelling with cell markers: CD31 (PECAM-1), vascular endothelial cadherin (VE-cadherin), vascular endothelial growth factor receptor-2 (VEGF-R2), Tie-2, von Willebrand factor, CD34, erythropoeitin receptor (Epo-R), alpha-smooth muscle actin, CD45, cytokeratin 7, and Ki-67. Extravillous and villous cytotrophoblasts, isolated and differentiated in vitro, expressed and secreted angiogenin. Angiogenin was detected in villous trophoblastic layers, and structured and nascent fetal vessels. In decidua, it was expressed by glandular epithelial cells, vascular cells and macrophages. The observed pattern of angiogenin expression is compatible with a role in blood vessel formation and in cross-talk between trophoblasts and endothelial cells. In view of angiogenin properties, we suggest that angiogenin may participate in placental vasculogenesis and organogenesis

Human Trophoblast in Trisomy 21: A Model for Cell–Cell Fusion Dynamic Investigation

International audienceTrophoblastic cell fusion is one essential step of the human trophoblast differentiation leading to formation of the syncytiotrophoblast, site of the numerous placental functions. This process is multifactorial and finely regulated. Using the physiological model of primary culture of trophoblastic cells isolated from human placenta, we have identified different membrane proteins directly involved in trophoblastic cell fusion: connexin 43, ZO-1 and recently syncytins. These fusogenic membrane retroviral envelop glycoproteins: syncytin-1 (encoded by the HERV-W gene) and syncytin-2 (encoded by the FRD gene) and their receptors are major factors involved in human placental development. Disturbances of syncytiotrophoblast formation are observed in trisomy 21-affected placentas. Overexpression of the copper/zinc superoxide dismutase (SOD-1), encoded by chromosome 21 as well as an abnormal hCG signaling are implicated in the defect of syncytiotrophoblast formation. This abnormal trophoblast fusion and differentiation in trisomy 21-affected placenta is reversible in vitro by different ways

生物沼气生产利用系统建模分析及可持续性评价

生物质废弃物通过厌氧发酵方式生产生物沼气是生物质高效资源化利用的重要手段之一，具有经济和环境的双重效益，是可再生能源领域的研究热点。现有研究主要针对生物质的厌氧发酵及生物沼气的利用等关键单元技术，从全系统角度的研究较为缺乏，而该体系涉及多种技术、多个单元过程以及多种影响评价指标，是一个复杂拓扑网络系统评价问题。因此，需要借助系统工程的方法对该复杂系统进行深入剖析，了解各单元及变量对系统性能影响的机制，并对影响系统效率的多种技术采用多种指标进行综合评价，筛选出具有代表性的可持续发展路线为生物沼气技术的大规模应用提供指导。基于上述背景，本文对生物沼气全系统进行了能量、环境及经济综合评价，并对系统中的沼气提纯及厌氧发酵关键单元进行了建模与能耗分析。主要研究内容与成果如下：（1）针对粗沼气提纯分离单元，开展了变压吸附法（PSA）粗生物沼气提纯制备生物甲烷工艺的动态模拟与评价研究。分别以13X沸石（Zeolite 13X）、 3K碳分子筛（CMS-3K）和508b金属有机骨架材料（MOF-508b）为吸附剂，建立了两塔-六步的改进 Skarstrom 动态变压吸附模拟流程，对变压吸附过程的关键参数如吸脱附压力、进料吹扫比等进行了灵敏度分析，确定了优化工艺参数； 考察了吸附塔内的压力、CH4 及CO2 组分的浓度随塔高及循环时间的变化；对三种吸附剂装填条件下工艺过程的能耗、设备尺寸及吸附剂装填量进行了计算与比较。结果表明，采用MOF-508b和CMS-3K作为吸附剂时的工艺能耗比Zeolite 13X作为吸附剂时分别低56%和50%；MOF-508b及CMS-3K填充的吸附塔塔径比 Zeolite 13X 填充的吸附塔塔径分别小13%和27%。（2）针对生物质厌氧发酵单元，建立了容积产气率及单元的热平衡模型。对中温 35?C 和高温 55?C 厌氧发酵状态，根据实验数据拟合了三种二元共发酵体系（牛粪与秸秆、鸡粪与秸秆、人粪与秸秆）的容积产气率模型，模型预测结果与实验结果平均偏差在 7%以内。 对热平衡模型， 考察了沼液低温余热的回收对减少过程外供热量的影响。结果表明，发酵过程所需热量的约 89%用于加热进口的冷物料，而约 11%用于维持发酵罐的恒温。 因此， 对沼液的低温余热加以回收以减少过程热量供给是十分必要的。以沼气锅炉供热发酵罐方式为例，通过沼液余热回收， 在中温发酵和高温发酵状态下，每天可以分别减少42%和49%的沼气消耗。（3）针对生物甲烷生产全系统，开展了物流与能量分析研究。在单元过程模拟基础上，建立了该系统的能耗模型和能量效率评价指标。考虑了2种发酵技术、4种粗沼气提纯技术、2 种系统热量供给及是否进行沼液低温余热的利用等因素，设计了32种情景路线；考察了发酵温度、系统热量供给方式及沼液低温余热回收模式对系统能效的影响。结果显示，采用高温发酵技术， 粗沼气及生物甲烷转化率比中温发酵分别提高120%及110%，能量的转化效率提高1倍；高温发酵比中温发酵减少了约3.1 wt%沼液及沼渣的处理量及26%的能量损失； 对中温及高温发酵情景下系统的能效分析与比较，结果表明，采用高温发酵及加压水洗技术，系统所需的热量由外部热源供给且沼液与发酵原料换热回收低温余热，系统的能效在 32 种情景中最高（46.5%）；采用中温发酵及变压吸附技术，系统所需的热量由燃烧发酵过程自产沼气供给且沼液与发酵原料不进行换热回收低温余热，系统的能量效率在 32 种情景中最低（15.8%）；在系统的热量供给方式及沼液与发酵原料换热模式相同的情况下，高温条件下系统的能效约为中温条件下的2倍。在对系统能效影响的三个因素中，发酵温度是对系统能效影响最大的因素，其次是系统热量的供给方式，最后是沼液与发酵原料的换热模式。（4）针对三种不同沼气利用方式（提纯制备生物甲烷、热电联产、固体燃料电池）构成的生物沼气生产及利用系统，对其进行了概念设计，并分别对其能效、绿色度及净现值等指标进行了综合评价与比较。结果表明， 对于系统的能量效率，为沼气提纯 &gt;沼气 SOFCs &gt; 沼气 CHP，提纯利用方式系统能效最高，SOFCs利用方式系统的能效比 CHP 利用方式系统的能效高 2.5%；对于系统绿色度变化量，沼气SOFCs &gt; 沼气 CHP &gt; 沼气提纯；对于系统的投资回收期，沼气 CHP &gt; 沼气 SOFCs &gt;沼气提纯。</p

Involvement of connexin 43 in human trophoblast cell fusion and differentiation.: Cx43and human trophoblast differentiation

The syncytiotrophoblast is the principal component of the human placenta involved in feto-maternal exchanges and hormone secretion. The syncytiotrophoblast arises from the fusion of villous cytotrophoblasts. We recently showed that functional gap junctional intercellular communication (GJIC) is an important prerequisite for syncytiotrophoblast formation and that connexin 43 (Cx43) is present in cytotrophoblasts and in the syncytiotrophoblast. To determine whether Cx43 is directly involved in trophoblast fusion, we used an antisense strategy in primary cultures of human villous cytotrophoblasts that spontaneously differentiate into the syncytiotrophoblast by cell fusion. We assessed the morphological and functional differentiation of trophoblasts by desmoplakin immunostaining, by quantifying hCG (human chorionic gonadotropin) production and by measuring the expression of specific trophoblast genes (hCG and HERV-W). Furthermore, we used the gap-FRAP (fluorescence recovery after photobleaching) method to investigate functional GJIC. Cytotrophoblasts treated with Cx43 antisense aggregated and fused poorly. Furthermore, less HERV-W env mRNA, hCGbeta mRNA and hCG secretion were detected in Cx43 antisense-treated cytotrophoblasts than in cells treated with scrambled antisense. Treatment with Cx43 antisense dramatically reduced the percentage of coupled trophoblast cells. Taken together, these results suggest that Cx43 is directly involved in human trophoblast cell-cell communication, fusion and differentiation

Requirement of Gap Junctional Intercellular Communication for Human Villous Trophoblast Differentiation 1

International audienceDuring pregnancy, the villous trophoblast develops from the fusion of cytotrophoblastic cells (CT) into a syncytiotrophoblast (ST), supporting the main physiological functions of the human placenta. Connexin43 (Cx43) is demonstrated in situ and in vitro in the villous trophoblast between CT and between CT and ST. Moreover, the presence of a gap junctional intercellular communication (GJIC) during in vitro trophoblast differentiation was previously demonstrated. Because the exchange of molecules through gap junctions is considered to play a major role in the control of cell and tissue differentiation, we studied the effects of a gap junctional uncoupler, heptanol, on morphological and functional trophoblast differentiation and on GJIC measured by the fluorescence recovery after photobleaching method. We found that when the GJIC was interrupted, CT still aggregated but fused poorly. This morphological effect was associated with a significant decrease of trophoblastic-specific gene expression (␤ human chorionic gonadotropin and human chorionic soma-tomammotropin). This blocking action was reversible as demonstrated by recovery of GJIC and trophoblast differentiation process after heptanol removal. Moreover, the inhibition of the trophoblast differentiation did not affect Cx43 transcript expression and Cx43 protein expression. These data suggest that the molecular exchanges through gap junctions preceding cellular fusion are essential for trophoblast differentiation generating the multifunctional syncytiotrophoblast. human chorionic gonadotropin, placenta, signal transduction, syncytiotrophoblast, trophoblas