MAPK phosphatase-1 represents a novel antiinflammatory target of glucocorticoids in the human endothelium

Glucocorticoids are well-established anti- inflammatory drugs thought to mainly act by inhibition of proinflammatory transcription factors like NF-κB. In recent years, however, transcription factorindependent mechanisms of glucocorticoid action have been proposed, namely the influence on MAPK pathways. Here we identify MAPK phosphatase-1 (MKP-1) as a pivotal mediator of the anti-inflammatory action of glucocorticoids in the human endothelium. We applied dexamethasone (Dex) to TNF-α-activated human endothelial cells and used the adhesion molecule E-selectin as inflammatory read-out parameter. Dex is known to reduce the expression of E-selectin, which is largely regulated by NF-κB. Here, we communicate that Dex at low concentrations (1–100 nM) markedly attenuates E-selectin expression without affecting NF-κB. Importantly, Dex is able to increase the expression of MKP-1, which causes an inactivation of TNF-α-induced p38 MAPK and mediates inhibition of E-selectin expression. In endothelial MKP-1ˉ/ˉ cells differentiated from MKP-1ˉ/ˉ embryonic stem cells and in MKP-1-silenced human endothelial cells, Dex did not inhibit TNF-α-evoked E-selectin expression. Thus, our findings introduce MKP-1 as a novel and crucial mediator of the anti-inflammatory action of glucocorticoids at low concentrations in the human endothelium and highlight MKP-1 as an important and promising antiinflammatory drug target

The ePetri dish, an on-chip cell imaging platform based on subpixel perspective sweeping microscopy (SPSM)

We report a chip-scale lensless wide-field-of-view microscopy imaging technique, subpixel perspective sweeping microscopy, which can render microscopy images of growing or confluent cell cultures autonomously. We demonstrate that this technology can be used to build smart Petri dish platforms, termed ePetri, for cell culture experiments. This technique leverages the recent broad and cheap availability of high performance image sensor chips to provide a low-cost and automated microscopy solution. Unlike the two major classes of lensless microscopy methods, optofluidic microscopy and digital in-line holography microscopy, this new approach is fully capable of working with cell cultures or any samples in which cells may be contiguously connected. With our prototype, we demonstrate the ability to image samples of area 6 mm × 4 mm at 660-nm resolution. As a further demonstration, we showed that the method can be applied to image color stained cell culture sample and to image and track cell culture growth directly within an incubator. Finally, we showed that this method can track embryonic stem cell differentiations over the entire sensor surface. Smart Petri dish based on this technology can significantly streamline and improve cell culture experiments by cutting down on human labor and contamination risks

Multiscale mechanisms of cell migration during development: theory and experiment

Long-distance cell migration is an important feature of embryonic development, adult morphogenesis and cancer, yet the mechanisms that drive subpopulations of cells to distinct targets are poorly understood. Here, we use the embryonic neural crest (NC) in tandem with theoretical studies to evaluate model mechanisms of long-distance cell migration. We find that a simple chemotaxis model is insufficient to explain our experimental data. Instead, model simulations predict that NC cell migration requires leading cells to respond to long-range guidance signals and trailing cells to short-range cues in order to maintain a directed, multicellular stream. Experiments confirm differences in leading versus trailing NC cell subpopulations, manifested in unique cell orientation and gene expression patterns that respond to non-linear tissue growth of the migratory domain. Ablation experiments that delete the trailing NC cell subpopulation reveal that leading NC cells distribute all along the migratory pathway and develop a leading/trailing cellular orientation and gene expression profile that is predicted by model simulations. Transplantation experiments and model predictions that move trailing NC cells to the migratory front, or vice versa, reveal that cells adopt a gene expression profile and cell behaviors corresponding to the new position within the migratory stream. These results offer a mechanistic model in which leading cells create and respond to a cell-induced chemotactic gradient and transmit guidance information to trailing cells that use short-range signals to move in a directional manner

Spatiotemporal endothelial cell-pericyte association in tumors as shown by high resolution 4D intravital imaging

Endothelial cells and pericytes are integral cellular components of the vasculature with distinct interactive functionalities. To study dynamic interactions between these two cells we created two transgenic animal lines. A truncated eNOS (endothelial nitric oxide synthase) construct was used as a GFP tag for endothelial cell evaluation and an inducible Cre-lox recombination, under control of the Pdgfrb (platelet derived growth factor receptor beta) promoter, was created for pericyte assessment. Also, eNOStag-GFP animals were crossed with the already established Cspg4-DsRed mice expressing DsRed fluorescent protein in pericytes. For intravital imaging we used tumors implanted in the dorsal skinfold of these transgenic animals. This setup allowed us to study time and space dependent complexities, such as distribution, morphology, motility, and association between both vascular cell types in all angiogenetic stages, without the need for additional labeling. Moreover, as fluorescence was still clearly detectable after fixation, it is possible to perform comparative histology following intravital evaluation. These transgenic mouse lines form an excellent model to capture collective and individual cellular and subcellular endothelial cell-pericyte dynamics and will help answer key questions on the cellular and molecular relationship between these two cells

Endothelio-hematopoietic relationship: getting closer to the beginnings

The close association between hematopoietic and endothelial cells during embryonic development led to the proposal that they may originate from a common ancestor - the hemangioblast. Due to a lack of unique specific markers for in vivo cell fate tracking studies, evidence supporting this theory derives mainly from in vitro differentiation studies. Teixeira and colleagues describe a novel enhancer that drives specific eGFP expression in blood islands of the electroporated chick embryo, thereby presenting a tool potentially suitable for analysis of hemangioblast differentiation and development of blood islands

Maximum parsimony analysis of gene expression profiles permits the reconstruction of developmental cell lineage trees

AbstractSpatiotemporal control of gene expression lies at the heart of generating several hundred distinct cell types required for the development of higher order animals. Different cell types within complex organs are often characterised by means of genome-wide gene expression profiling, but analogous information for early developmental as well as adult stem and progenitor cells is largely missing because their identity is commonly unknown or they are present in prohibitively small numbers. Here we show that maximum parsimony approaches previously used to reconstruct evolutionary trees from gene content of extant species can be adapted to reconstruct cellular hierarchies both during development and steady state homeostasis of complex mammalian tissues. Using haematopoiesis as a model, we show that developmental trees reconstructed from expression profiles of mature cells are not only consistent with current experimentally validated trees but also have predictive value in determining progenitor cell specific transcriptional programmes and lineage determining transcription factors. Subsequent analysis across diverse developmental systems such as neuronal development and endoderm organogenesis demonstrated that maximum parsimony-based reconstruction of developmental trees represents a widely applicable approach to infer developmental pathways as well as the transcriptional control mechanisms underlying cell fate specification

Hierarchical organization and early hematopoietic specification of the developing HSC lineage in the AGM region

A CD45-negative population of pre-HSCs develops into definitive HSCs in the AGM region of the embryo

A survey of visualisation for live cell imaging

Live cell imaging is an important biomedical research paradigm for studying dynamic cellular behaviour. Although phenotypic data derived from images are difficult to explore and analyse, some researchers have successfully addressed this with visualisation. Nonetheless, visualisation methods for live cell imaging data have been reported in an ad hoc and fragmented fashion. This leads to a knowledge gap where it is difficult for biologists and visualisation developers to evaluate the advantages and disadvantages of different visualisation methods, and for visualisation researchers to gain an overview of existing work to identify research priorities. To address this gap, we survey existing visualisation methods for live cell imaging from a visualisation research perspective for the first time. Based on recent visualisation theory, we perform a structured qualitative analysis of visualisation methods that includes characterising the domain and data, abstracting tasks, and describing visual encoding and interaction design. Based on our survey, we identify and discuss research gaps that future work should address: the broad analytical context of live cell imaging; the importance of behavioural comparisons; links with dynamic data visualisation; the consequences of different data modalities; shortcomings in interactive support; and, in addition to analysis, the value of the presentation of phenotypic data and insights to other stakeholders

Luteal angiogenesis and its control

Angiogenesis, the formation of new blood vessels from pre-existing ones, is critical to luteal structure and function; In addition, it is a complex and tightly regulated process. Not only does rapid and extensive angiogenesis occur to provide the corpus luteum (CL) with an unusually high blood flow and support its high metabolic rate, but in the absence of pregnancy the luteal vasculature must rapidly regress to enable the next cycle of ovarian activity. This review describes a number of the key endogenous stimulatory and inhibitory factors, which act in a delicate balance to regulate luteal angiogenesis and ultimately luteal function. In vitro luteal angiogenesis cultures have demonstrated critical roles for fibroblast growth factor 2 (FGF2) in endothelial cell proliferation and sprouting, whilst other factors such as vascular endothelial growth factor (VEGFA) and platelet derived growth factor (PDGF) were important modulators in the control of luteal angiogenesis. Post-transcriptional regulation by small non-coding micro-RNAs, is also likely to play a central role in the regulation of luteal angiogenesis. Appropriate luteal angiogenesis requires the coordinated activity of numerous factors expressed by several cell types at different times and this review will also describe the role of perivascular pericytes and the importance of vascular maturation and stability. It is hoped that a better understanding of the critical processes underlying the transition from follicle to CL, and subsequent luteal development will benefit the management of luteal function in the future