Biology subjects

Robinson, S.

Publications and source records attributed to Robinson, S..

6 recordsLinked to original sources

bioRxiv2018

A novel bioreactor technology for modelling fibrosis in human and rodent precision-cut liver slices.

Summary boxO_LIWhat is already known about this subject?\nO_LICurrently there are no effective anti-fibrotic drugs to treat liver fibrosis and there is an urgent unmet need to increase our knowledge of the disease process and develop better tools for anti-fibrotic drug discovery.\nC_LIO_LIPreclinical in vitro cell cultures and animal models are widely used to study liver fibrosis and test anti-fibrotic drugs, but have shortfalls; cell culture models lack the relevant complex cell-cell interactions of the liver and animal models only reproduce some features of human disease.\nC_LIO_LIPrecision Cut Liver Slices (PCLS) are structurally representative of the liver and can be used to model liver fibrosis and test anti-fibrotic drugs. However, PCLS are typically cultured in elevated, non-physiological oxygen levels and only have a healthy lifespan of 48h.\nC_LI\nC_LIO_LIWhat are the new findings?\nO_LIWe have developed a novel bioreactor culture system that increases the longevity of functional PCLS to up to 6 days under normoxic conditions.\nC_LIO_LIBioreactor cultured PCLS can be used to model fibrogenesis in both normal and fibrotic PCLS using a combination of biochemical and histological outputs.\nC_LIO_LIAdministration of an Alk5 inhibitor effectively limits fibrogenesis in normal rodent and human PCLS and in rodent PCLS with established fibrosis.\nC_LI\nC_LIO_LIHow might it impact on clinical practice in the foreseeable future?\nO_LIThe extended longevity of bioreactor cultured PCLS represent a novel pre-clinical tool to investigate the cellular and molecular mechanisms of liver fibrosis.\nC_LIO_LIBioreactor cultured human PCLS offer a clinically relevant system to test efficacy of anti-fibrotic drugs.\nC_LI\nC_LI\n\nAbstractO_ST_ABSObjectiveC_ST_ABSPrecision cut liver slices (PCLS) retain the structure and cellular composition of the native liver and represent an improved system to study liver fibrosis compared to two-dimensional mono or co-cultures. The objective of this study was to develop a bioreactor system to increase the healthy lifespan of PCLS and model fibrogenesis.\n\nDesignPCLS were generated from normal rat or human liver, or 4-week carbon tetrachloride-fibrotic rat liver and cultured in our patented bioreactor. PCLS function was quantified by albumin ELISA. Fibrosis was induced in PCLS by TGF{beta}1 and PDGF{beta}{beta} stimulation. Alk5 inhibitor therapy was used. Fibrosis was assessed by fibrogenic gene expression, Picrosirius Red and Smooth Muscle Actin staining, hydroxyproline assay and collagen 1a1, fibronectin and hyaluronic acid ELISA.\n\nResultsBioreactor cultured PCLS are viable, maintaining tissue structure and stable albumin secretion for up to 6 days under normoxic culture conditions. Conversely, standard static transwell cultured PCLS rapidly deteriorate and albumin secretion is significantly impaired by 48 hours. TGF{beta}1 and PDGF{beta}{beta} stimulation of rat or human PCLS induced fibrogenic gene expression, release of extracellular matrix proteins, activation of hepatic myofibroblasts and histological fibrosis. Fibrogenesis slowly progresses over 6-days in cultured fibrotic rat PCLS without exogenous challenge. Alk5 inhibitor limited fibrogenesis in both TGF{beta}1 and PDGF{beta}{beta} stimulated PCLS and fibrotic PCLS.\n\nConclusionWe describe a new bioreactor technology which maintains functional PCLS cultures for 6 days. Bioreactor cultured PCLS can be successfully used to model fibrogenesis and demonstrate efficacy of an anti-fibrotic therapy.

cell biology↗

bioRxiv2018

The Atypical Rho GTPase CHW-1 Works With SAX-3/Robo to Mediate Axon Guidance in Caenorhabditis elegans

During development, neuronal cells extend an axon towards their target destination in response to a cue to form a properly functioning nervous system. Rho proteins, Ras-related small GTPases that regulate cytoskeletal organization and dynamics, cell adhesion, and motility, are known to regulate axon guidance. Despite extensive knowledge about canonical Rho proteins (RhoA/Rac1/Cdc42), little is known about the Caenorhabditis elegans (C. elegans) atypical Cdc42-like family members CHW-1 and CRP-1 in regards to axon pathfinding and neuronal migration. chw-1(Chp/Wrch) encodes a protein that resembles human Chp (Wrch-2/RhoV) and Wrch-1 (RhoU), and crp-1 encodes for a protein that resembles TC10 and TCL. Here, we show that chw-1 works redundantly with crp-1 and cdc-42 in axon guidance. Furthermore, proper levels of chw-1 expression and activity are required for proper axon guidance. When examining CHW-1 GTPase mutants, we found that the native CHW-1 protein is likely partially activated, and mutations at a conserved residue (position 12 using Ras numbering, position 18 in CHW-1) alter axon guidance and neural migration. Additionally, we showed that chw-1 genetically interacts with the guidance receptor sax-3 in PDE neurons. Finally, in VD/DD motor neurons, chw-1 works downstream of sax-3 to control axon guidance. In summary, this is the first study implicating the atypical Rho GTPases chw-1 and crp-1 in axon guidance. Furthermore, this is the first evidence of genetic interaction between chw-1 and the guidance receptor sax-3. These data suggest that chw-1 is likely acting downstream and/or in parallel to sax-3 in axon guidance.

developmental biology↗

bioRxiv2018

Asymmetric effects of environmental filtering on the assembly of tropical bird communities along a moisture gradient

The species-sorting hypothesis (SSH) states that environmental factors influence local community assembly of metacommunities by selecting for species that are well adapted to the specific conditions of each site. Along environmental gradients, the strength of selection against individuals that are marginally adapted to the local conditions increases towards the extremes of the environment where the climate becomes harsh. In rainfall gradients, the strength of selection by the environment has been proposed to decrease with rainfall. Under this scenario SSH would predict that immigration of individuals from the metacommunity should be restricted into the dry end of the gradient creating a positive relationship between immigration and rainfall. However, if the selection is strong in both ends of the gradient, the restriction should be expected to be in both directions such that the ends behave as independent metacommunities even in the absence of geographical barriers. In this study we used models based on neutral theory to evaluate if SSH can explain the distribution of bird species along a steep rainfall gradient in Colombia. We found a strong positive relationship between immigration rates and precipitation suggesting that the dry forests impose stronger challenges for marginally adapted bird populations. However, a two-metacommunity model separating dry and wet forests was a better fit to the observed data, suggesting that both extremes impose strong selection against immigrants. The switch from the dry forest to the wet forest metacommunities occurred abruptly over a short geographic distance in the absence of any apparent geographic barrier; this apparent threshold occurs where the forest becomes mostly evergreen. The relative number of rare species in dry forest was lower than in wet forests suggesting that the selection against marginally adapted populations is stronger in the dry forests. Overall, our analyses are consistent with SSH at the regional scales, but the rarity analysis suggests that the mechanisms at the local scales are substantially different. Based on these results, we hypothesize that abiotic (climatic) factors limit immigration into dry forest communities and whereas biotic factors such as competition and predation may limit immigration into bird communities in the wet forest.

ecology↗

bioRxiv2018

Climatic effects on turnover of lowland forest bird communities across a precipitation gradient

One of the main goals of community ecology is to understand the influence of the abiotic environment on the abundance and distribution of species. It has been hypothesized that dry forests are harsher environments than wet forests, which leads to the prediction that environmental filtering should be a more important determinant of patterns of species abundance and composition than in wet forest, where biotic interactions or random assembly should be more important. We attempt to understand the influence of rainfall on the abundance and distribution of bird species along a steep precipitation gradient in an inter-Andean valley in Colombia. We gathered data on species distributions, abundance, morphological traits and phylogenetic relationships to determine the influence of rainfall on the taxonomic, functional and phylogenetic turnover of species along the Magdalena Valley. We demonstrate that there is a strong turnover of community composition at the limit of the dry forest. The taxonomic turnover is steeper than the phylogenetic turnover, suggesting that replacement of closely related species accounts for a disproportionate number of changes along the gradient. We found evidence for environmental filtering in dry forest as species tend to be more tolerant of higher temperature ranges, stronger rainfall seasonality and lower minimum rainfall. On the other hand, wet forest species tend to compete actively for nest space but not for the resources associated with the axes we measured. Our results suggest that rainfall is a strong determinant of community composition when comparing localities above and below the 2400 mm rainfall isocline.

ecology↗

bioRxiv2017

Gene expression dynamics and inter-tissue recognition mechanisms during tissue fusion at the Arabidopsis graft junction

The ability for cut tissues to join together and form a chimeric organism is a remarkable property of many plants, however, grafting is poorly characterized at the molecular level. To better understand this process we monitored genome-wide temporal and spatial gene expression changes in grafted Arabidopsis thaliana hypocotyls. Tissues above and below the graft rapidly developed an asymmetry such that many genes were more highly expressed on one side than the other. This asymmetry correlated with sugar responsive genes and we observed an accumulation of starch above the graft that decreased along with asymmetry once the sugar-transporting vascular tissues reconnected. Despite the initial starvation response below the graft, many genes associated with vascular formation were rapidly activated in grafted tissues but not in cut and separated tissues indicating that a recognition mechanism activated that was independent of functional vascular connections. Auxin which is transported cell-to-cell, had a rapidly elevated response that was symmetric, suggesting that auxin was perceived by the root within hours of tissue attachment to activate the vascular regeneration process. A subset of genes were expressed only in grafted tissues, indicating that wound healing proceeded via different mechanisms depending on the presence or absence of adjoining tissues. Such a recognition process could have broader relevance for tissue regeneration, inter-tissue communication and tissue fusion events.

plant biology↗

bioRxiv2017

An automated confocal micro-extensometer enables in vivo quantification of mechanical properties with cellular resolution.

How complex developmental-genetic networks are translated into organs with specific 3D shapes remains an open question. This question is particularly challenging because the elaboration of specific shapes is in essence a question of mechanics. In plants, this means how the genetic circuitry affects the cell wall. The mechanical properties of the wall and their spatial variation are the key factors controlling morphogenesis in plants. However, these properties are difficult to measure and investigating their relation to genetic regulation is particularly challenging. To measure spatial variation of mechanical properties, one must determine the deformation of a tissue in response to a known force with cellular resolution. Here we present an automated confocal micro-extensometer (ACME), which greatly expands the scope of existing methods for measuring mechanical properties. Unlike classical extensometers, ACME is mounted on a confocal microscope and utilizes confocal images to compute the deformation of the tissue directly from biological markers, thus providing cellular scale information and improved accuracy. ACME is suitable for measuring the mechanical responses in live tissue. As a proof of concept we demonstrate that the plant hormone gibberellic acid induces a spatial gradient in mechanical properties along the length of the Arabidopsis hypocotyl.\n\nTerms

plant biology↗