胚胎干细胞诱导分化的研究进展

胚胎干细胞(embryonic stem cell,ESC)因其具有自我更新能力和发育的多能性,成为当前医学研究的热点。ESC不但可以自发分化,而且在诱导因素作用下可以定向分化为某一种特定的成熟细胞。因此,ESC在移植医学、发育生物学等领域有着广阔的应用前景。本文对几种定向诱导ESC分化的策略进行了综述。     

Factors from Human Embryonic Stem Cell-derived Fibroblast-like Cells Promote Topology-dependent Hepatic Differentiation in Primate Embryonic and Induced Pluripotent Stem Cells

The future clinical use of embryonic stem cell (ESC)-based hepatocyte replacement therapy depends on the development of an efficient procedure for differentiation of hepatocytes from ESCs. Here we report that a high density of human ESC-derived fibroblast-like cells (hESdFs) supported the efficient generation of hepatocyte-like cells with functional and mature hepatic phenotypes from primate ESCs and human induced pluripotent stem cells. Molecular and immunocytochemistry analyses revealed that hESdFs caused a rapid loss of pluripotency and induced a sequential endoderm-to-hepatocyte differentiation in the central area of ESC colonies. Knockdown experiments demonstrated that pluripotent stem cells were directed toward endodermal and hepatic lineages by FGF2 and activin A secreted from hESdFs. Furthermore, we found that the central region of ESC colonies was essential for the hepatic endoderm-specific differentiation, because its removal caused a complete disruption of endodermal differentiation. In conclusion, we describe a novel in vitro differentiation model and show that hESdF-secreted factors act in concert with regional features of ESC colonies to induce robust hepatic endoderm differentiation in primate pluripotent stem cells.     

A multiscale simulation framework for the manufacturing facility and supply chain of autologous cell therapies

Background aimsAutologous cell therapy (AuCT) is an emerging therapeutic treatment that is undergoing transformation from laboratory- to industry-scale manufacturing with recent regulatory approvals. Various challenges facing the complex AuCT manufacturing and supply chain process hinder the scale out and broader application of this highly potent treatment.MethodsWe present a multiscale logistics simulation framework, AuCT-Sim, that integrates novel supply chain system modeling algorithms, methods, and tools. AuCT-Sim includes a single facility model and a system-wide network model. Unique challenges of the AuCT industry are analyzed and addressed in AuCT-Sim. Decision-supporting tools can be developed based on this framework to explore “what-if” manufacturing and supply chain scenarios of importance to various cell therapy stakeholder groups.ResultsTwo case studies demonstrate the decision-supporting capability of AuCT-Sim where one investigates the optimal reagent base stocking level, and the other one simulates a reagent supply disruption event. These case studies serve as guidelines for designing computational experiments with AuCT-Sim to solve specific problems in AuCT manufacturing and supply chain.DiscussionThis simulation framework will be useful in understanding the impact of possible manufacturing and supply chain strategies, policies, regulations, and standards informing strategies to increase patient access to AuCT.     

Fostering diffusion of scientific contents of National Society Cardiovascular Journals:the new ESC search engine

European Society of Cardiology (ESC) National Society Cardiovascular Journals (NSCJs) are high-quality biomedical journals focused on cardiovascular diseases. The Editors’ Network of the ESC devises editorial initiatives aimed at improving the scientific quality and diffusion of NSCJ. In this article we will discuss on the importance of the Internet, electronic editions and open access strategies on scientific publishing. Finally, we will propose a new editorial initiative based on a novel electronic tool on the ESC web-page that may further help to increase the dissemination of contents and visibility of NSCJs.     

Arthropod abundance in an Australian apple orchard under mating disruption and supplementary insecticide treatments for codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae)

The impacts of three codling moth management strategies (i, mating disruption alone; ii, mating disruption plus azinphos-methyl; iii, mating disruption plus fenoxycarb) on some secondary pests and their natural enemies in an apple orchard were compared over two growing seasons: 1993/1994 and 1994/1995. In the absence of azinphos-methyl (strategies i and iii), two-spotted mite ( Tetranychus urticae ) was controlled by Typhlodromus occidentalis and populations of generalist predators (e.g. ladybirds, lacewings and earwigs) increased. The populations of a parasitoid of woolly aphid ( Eriosoma lanigerum ), Aphelinus mali, also increased but not enough to provide adequate control of the aphid. Combined damage caused by lightbrown apple moth ( Epiphyas postvittana ), budworms ( Helicoverpa spp.) and San José scale ( Quadraspidiotus perniciosus ) was significantly higher in the absence of azinphos-methyl in 1994/1995. Beneficial insect populations were not suppressed by fenoxycarb. In 1994/1995, mating disruption plus fenoxycarb produced better control of E. postvittana than mating disruption alone. During transition to an apple integrated pest management program based on codling moth mating disruption, fenoxycarb was shown to be less disruptive to any natural control of secondary pests than azinphos-methyl.     

Immune system of channel catfish: An overture on immunity to Edwardsiella ictaluri

Edwardsiella ictaluri is a gram negative rod that causes enteric septicemia of catfish (ESC). The E. ictaluri disease complex includes acute and chronic ESC. One of the most economically important diseases of channel catfish, Ictalurus punctatus, ESC is difficult to control. Although the bacterium is usually susceptible to antibiotics, resistant strains are emerging. Immunization is a more attractive approach to control. Vaccine efficacy is not well documented, in part due to a lack of basic information about immune system components and their interactions after E. ictaluri infection. Channel catfish vaccinated by immersion in E. ictaluri bacterin are only partly protected against ESC. Specific antibody responses are poor in the vaccinates. E. ictaluri infection causes both humoral and cell-mediated immune responses. A vaccine that stimulates both types of immunity may provide better ESC immunity. Diagnosis of E. ictaluri infection is accomplished by detection of specific antibody. Recently the discovery of an immunodominant E. ictaluri exoantigen is allowing the development of accurate and rapid diagnostic tests that can detect infection before clinical ESC occurs. Health management programs for food animals depend on serological tests to detect early infections and carriers; the value of these tests is now realized in the channel catfish industry. This review documents factors that influence the immune responses of channel catfish. These factors include the influence of water temperature, seasons, stressful conditions, diet, and E. ictaluri carrier status.     

Ovine (Ovis aries) blastula from an in vitro production system and isolation of primary embryonic stem cells

Livestock embryo production in in vitro systems has been highlighted due to the emergence of interest in embryo stem cells (ESC). ESC potency and their wide potential applications have been recognized in medicine, fundamental research fields and commercial markets due to ESC totipotency or pluripotency and self-renewal. Ovine ESC probably is a useful technical platform for transgenic livestock and animal cloning, but ESC lines have not yet been founded because of difficulties in ESC isolation and the lack of blastula materials. We have established an IVP (in vitro production) system in our laboratory, including in vitro maturation, in vitro fertilization and in vitro culture, to produce sheep blastula using fresh ovaries and testes collected from livestock production. This system can achieve rates of mature eggs and blastulas of 65 and 50% respectively, and can provide enough blastulas for ICM (inner cell mass) isolation. Furthermore, ESC-like clones were isolated from the ICM on ovine embryonic fibroblast (OEF) feeder cells and in ES-DMEM supplemented with the cell factors LIF and SCF, and these survived to the third passage, which was primarily identified by AKP staining and morphology. This work provides a basis for ovine ESC isolation and foundation of ESC lines.     

Relaxed 3D genome conformation facilitates the pluripotent to totipotent-like state transition in embryonic stem cells

The 3D genome organization is crucial for gene regulation. Although recent studies have revealed a uniquely relaxed genome conformation in totipotent early blastomeres of both fertilized and cloned embryos, how weakened higher-order chromatin structure is functionally linked to totipotency acquisition remains elusive. Using low-input Hi-C, ATAC-seq and ChIP-seq, we systematically examined the dynamics of 3D genome and epigenome during pluripotent to totipotent-like state transition in mouse embryonic stem cells (ESCs). The spontaneously converted 2-cell-embryo-like cells (2CLCs) exhibited more relaxed chromatin architecture compared to ESCs, including global weakening of both enhancer-promoter interactions and TAD insulation. While the former correlated with inactivation of ESC enhancers and down-regulation of pluripotent genes, the latter might facilitate contacts between the putative new enhancers arising in 2CLCs and neighboring 2C genes. Importantly, disruption of chromatin organization by depleting CTCF or the cohesin complex promoted the ESC to 2CLC transition. Our results thus establish a critical role of 3D genome organization in totipotency acquisition.     

Cryopreservation and quality control of mouse embryonic feeder cells

Stem cell research is a highly promising and rapidly progressing field inside regenerative medicine. Embryonic stem cells (ESCs), reprogrammed “induced pluripotent” cells (iPS), or lately protein induced pluripotent cells (piPS) share one inevitable factor: mouse embryonic feeder cells (MEFs), which are commonly used for ESC long term culture procedures and colony regeneration. These MEFs originate from different mouse strains, are inactivated by different methods and are differently cryopreserved. Incomprehensibly, there are to date no established quality control parameters for MEFs to insure consistency of ESC experiments and culture. Hence, in this work, we developed a bench-top quality control for embryonic feeder cells.According to our findings, MEFs should be inactivated by irradiation (30 Gy) and cryopreserved with optimal 10% DMSO at 1 K/min freezing velocity. Thawed cells should be free of mycoplasma and should have above 85 ± 13.1% viability. Values for the metabolic activity should be above 150 ± 10.5% and for the combined gene expression of selected marker genes above 225 ± 43.8% compared to non-irradiated, cryopreserved controls. Cells matching these criteria can be utilized for at least 12 days for ESC culture without detaching from the culture dish or disruption of the cell layer.     

Managing to harvest? Perspectives on the potential of aquaculture

Aquaculture has been one of the most rapid and technically innovative of food production sectors globally, with significant investment, scientific and technical development and production growth in many parts of the world over the past two decades. While this has had a significant effect on the global supply of aquatic food products and had an important impact in rural and urban food supply and employment in many developing economies, growth and increasing internationalization has not been without concern for natural resource use, environmental impact and social disruption. The expectations for production and diversification are now significant and while the scientific and technical means are already available to meet much of the intended targets, practical constraints of investment, profitability, resource access and system efficiency are likely to become far more important constraints for the future. This review offers a contemporary perspective on the ways in which the sector might develop, its interactions with constraints and the strategies that may be required to ensure that future development is both positive and sustainable.     

Neural differentiation of embryonic stem cells is induced by signalling from non-neural niche cells.

BACKGROUND/AIMS: Embryonic stem cell (ESC) transplantation offers new therapeutic strategies for neurodegenerative diseases and injury. However, the mechanisms underlying integration and differentiation of engrafted ESCs are poorly understood. This study elucidates the influence of exogenous signals on ESC differentiation using in vitro modelling of non-stem/stem cell interactions. METHODS: Murine ESCs were co-cultured with endothelial cells and astrocytes or conditioned medium obtained from endothelial or astrocyte cultures. After 7 days of co-culture isolated RNA was analysed using RT-PCR for the expression of pluripotency marker oct-4, neural progenitor marker nestin, and neurofilament (NFL), an early marker of neuronal lineage commitment. The presence of the glial cell surface marker A2B5 was determined in ESCs by flow cytometry. RESULTS: Neuronal differentiation was inhibited in ESCs when grown in close vicinity to cerebral endothelial or glial cells. Under these conditions, ESC differentiation was predominantly directed towards a glial fate. However, treatment of ESCs with endothelial cell- or astrocyte-conditioned medium promoted neuronal as well as glial differentiation. CONCLUSION: Our results indicate that ESC fate is determined by endothelial and glial cells that comprise the environmental niche of these stem cells in vivo. The direction of differentiation processes appears to be dependent on humoral factors secreted by adjacent cell lines.     

Over-Expression of HSP47 Augments Mouse Embryonic Stem Cell Smooth Muscle Differentiation and Chemotaxis

In the recent decade, embryonic stem cells (ESC) have emerged as an attractive cell source of smooth muscle cells (SMC) for vascular tissue engineering owing to their unlimited self-renewal and differentiation capacities. Despite their promise in therapy, their efficacy is still hampered by the lack of definitive SMC differentiation mechanisms and difficulties in successful trafficking of the ESC towards a site of injury or target tissue. Heat shock protein 47 (HSP47) is a 47-kDa molecular chaperone that is required for the maturation of various types of collagen and has been shown to be a critical modulator of different pathological and physiological processes. To date, the role of HSP47 on ESC to SMC differentiation or ESC chemotaxis is not known and may represent a potential molecular approach by which ESC can be manipulated to increase their efficacy in clinic. We provide evidence that HSP47 is highly expressed during ESC differentiation into the SMC lineage and that HSP47 reduction results in an attenuation of the differentiation. Our experiments using a HSP47 plasmid transfection system show that gene over-expression is sufficient to induce ESC-SMC differentiation, even in the absence of exogenous stimuli. Furthermore, HSP47 over-expression in ESC also increases their chemotaxis and migratory responses towards a panel of chemokines, likely via the upregulation of chemokine receptors. Our findings provide direct evidence of induced ESC migration and differentiation into SMC via the over-expression of HSP47, thus identifying a novel approach of molecular manipulation that can potentially be exploited to improve stem cell therapy for vascular repair and regeneration.     

生态系统服务级联效应研究进展

生态系统服务级联效应(ESC)将生态系统服务各组成部分与社会价值联系起来,构建了自然科学到社会科学跨学科研究的桥梁,可以帮助决策者更好地将生态系统服务概念融入到决策制定中。本文梳理了生态系统服务级联效应的概念,回顾了级联效应的研究进展。从理论角度看,ESC的理论研究主要集中在如何准确定义各成分及各成分间的因果关系,提高ESC模型的实用性与适用性,以及如何有效连接利益相关者与生态系统服务结构。从应用角度看,ESC在生态系统服务制图、生态系统服务评估及政策制定发挥了巨大作用。当前研究在指标筛选和互馈机制上仍存在较大不确定性。建议未来应从以下几个方面加强研究:更加关注生态系统服务结构过程及分类标准,通过多模型融合和区域调查来弥补ESC对反馈机制描述的不足,结合区域特征为人地耦合和可持续发展提供科学指导。     

Mechanical stretch upregulates IGFBP-1 secretion from decidualized endometrial stromal cells

Decidualization is an essential process of endometrial differentiation for embryo implantation and maintenance of pregnancy. Recently, uterine movement-induced mechanical stress was noticed to have possible effects on endometrial functions. In this study, we addressed the possible effect of mechanical stress on the process of decidualization of endometrial stromal cells (ESC). ESC were cultured on flexible-bottomed culture plates. After decidualization was achieved with estradiol and progesterone for 12 days, cultures were continued for 24 h with or without cyclic stretch (25% elongation) in serum-free conditions at a rate of 2 cycles/min using a computer-operated cell tension system. Concentrations of insulin-like growth factor-binding protein-1 (IGFBP-1), a marker of decidualization, in the conditioned medium were measured by specific ELISA, and IGFBP-1 mRNA expression in the ESC was measured by RT-PCR. Cyclic stretch remarkably increased IGFBP-1 secretion from decidualized ESC. It also increased IGFBP-1 mRNA in decidualized ESC. The increase in IGFBP-1 secretion was inhibited by actinomycin D but not by indomethacin, PD-98059, or H-89. Conditioned medium of decidualized ESC cultured with cyclic stretch increased IGFBP-1 secretion from decidualized ESC cultured under stationary conditions. These findings imply that uterine movement modulates decidualization of the endometrium and has a regulatory effect on reproduction.     

A Dynamic Study of Protein Secretion and Aggregation in the Secretory Pathway

Precise coordination of protein biogenesis, traffic and homeostasis within the early secretory compartment (ESC) is key for cell physiology. As a consequence, disturbances in these processes underlie many genetic and chronic diseases. Dynamic imaging methods are needed to follow the fate of cargo proteins and their interactions with resident enzymes and folding assistants. Here we applied the Halotag labelling system to study the behavior of proteins with different fates and roles in ESC: a chaperone, an ERAD substrate and an aggregation-prone molecule. Exploiting the Halo property of binding covalently ligands labelled with different fluorochromes, we developed and performed non-radioactive pulse and chase assays to follow sequential waves of proteins in ESC, discriminating between young and old molecules at the single cell level. In this way, we could monitor secretion and degradation of ER proteins in living cells. We can also follow the biogenesis, growth, accumulation and movements of protein aggregates in the ESC. Our data show that protein deposits within ESC grow by sequential apposition of molecules up to a given size, after which novel seeds are detected. The possibility of using ligands with distinct optical and physical properties offers a novel possibility to dynamically follow the fate of proteins in the ESC.     

Characteristics of the Early Immune Response Following Transplantation of Mouse ES Cell Derived Insulin-Producing Cell Clusters

Background

The fully differentiated progeny of ES cells (ESC) may eventually be used for cell replacement therapy (CRT). However, elements of the innate immune system may contribute to damage or destruction of these tissues when transplanted.

Methodology/Principal Findings

Herein, we assessed the hitherto ill-defined contribution of the early innate immune response in CRT after transplantation of either ESC derived insulin producing cell clusters (IPCCs) or adult pancreatic islets. Ingress of neutrophil or macrophage cells was noted immediately at the site of IPCC transplantation, but this infiltration was attenuated by day three. Gene profiling identified specific inflammatory cytokines and chemokines that were either absent or sharply reduced by three days after IPCC transplantation. Thus, IPCC transplantation provoked less of an early immune response than pancreatic islet transplantation.

Conclusions/Significance

Our study offers insights into the characteristics of the immune response of an ESC derived tissue in the incipient stages following transplantation and suggests potential strategies to inhibit cell damage to ensure their long-term perpetuation and functionality in CRT.     

2020 ESC Guidelines on acute coronary syndrome without ST-segment elevation: Recommendations and critical appraisal from the Dutch ACS and Interventional Cardiology working groups

Recently, the European Society of Cardiology (ESC) has updated its guidelines for the management of patients with acute coronary syndrome (ACS) without ST-segment elevation. The current consensus document of the Dutch ACS working group and the Working Group of Interventional Cardiology of the Netherlands Society of Cardiology aims to put the 2020 ESC Guidelines into the Dutch perspective and to provide practical recommendations for Dutch cardiologists, focusing on antiplatelet therapy, risk assessment and criteria for invasive strategy.     

Symbiotic microorganisms: untapped resources for insect pest control

Symbiotic microorganisms offer one route to meet the anticipated heightened demand for novel insect pest management strategies created by growing human populations and global climate change. Two approaches have particular potential: the disruption of microbial symbionts required by insect pests, and manipulation of microorganisms with major impacts on insect traits contributing to their pest status (e.g. capacity to vector diseases, natural enemy resistance). Specific research priorities addressed in this article include identification of molecular targets against which highly specific antagonists can be designed or discovered, and management strategies to manipulate the incidence and properties of facultative microorganisms that influence insect pest traits. Collaboration with practitioners in pest management will ensure that the research agenda is married to agricultural and public health needs.     

Connexin mutant embryonic stem cells and human diseases

Intercellular communication via gap junctions allows cells within multicellular organisms to share small molecules. The effect of such interactions has been elucidated using mouse gene knockout strategies. Although several mutations in human gap junction-encoding connexin (Cx) have been described, Cx mutants in mice do not always recapitulate the human disease. Among the 20 mouse Cxs, Cx26, Cx43, and Cx45 play roles in early cardiac or placental development, and disruption of the genes results in lethality that hampers further analyses. Embryonic stem cells (ESCs) that lack Cx43 or Cx45 have made analysis feasible in both in vitro differentiated cell cultures and in vivo chimeric tissues. The success of mouse ESCs studies is leading to the use of induced pluripotent stem cells to learn more about the pathogenesis of human Cx diseases. This review summarizes the current status of mouse Cx disruption models and ESC differentiation studies, and discusses their implication for understanding human Cx diseases.