A receptor-electromigration-based model for cellular electrotactic sensing and migration

Directed cell migration in tissues mediates various physiological processes and is guided by complex cellular factors such as chemoattractant gradients and electric fields. Direct current (DC) electric fields can be generated in physiological settings and the electric field guided migration of various cell types (i.e., electrotaxis) has been demonstrated both in vitro and in vivo. Although several mechanisms have been proposed for electrotaxis, there are so far very few quantitative models. Furthermore, because chemoattractant gradients and electric fields co-exist in tissues, it is important to understand how chemotaxis and electrotaxis interact for mediating cell migration and trafficking. In this study, we developed a mathematical model to investigate the role of electromigration of cell surface chemoattractant receptors in mediating electrochemical sensing and migration of cells. Our results show that electromigration of chemoattractant receptors enables cell electrotactic sensing and migration in the presence of a uniform chemoattractant field. Furthermore, in the physiologically-relevant range of receptor electromigration rates, application of electric fields overcomes chemical guiding signals for directional sensing and migration of cells in co-existing competing electric fields and chemoattractant gradients.     

Physiological similarity and bioreactor scale-up

An economically effective transfer of biological processes from laboratory to production scale is the main task of microbial process engineering. In contrast to the principle of geometrical, chemical, thermal, hydrodynamic or chemical similarity, recommended for scale-up of chemical reactors we propose the principle of physiological similarity. According to this principle same the microenvironment of the living cell must be established to reproduce the same physiological function (e.g. growth, product formation or substrate consumption rates) in the large scale bioreactor as in the laboratory one.     

Organ-on-Chip platforms to study tumor evolution and chemosensitivity

Despite tremendous advancements in oncology research and therapeutics, cancer remains a primary cause of death worldwide. One of the significant factors in this critical challenge is a precise diagnosis and limited knowledge on how the tumor microenvironment (TME) behaves to the treatment and its role in chemo-resistance. Therefore, it is critical to understand the contribution of a heterogeneous TME in cancer drug response in individual patients for effective therapy management. Micro-physiological systems along with tissue engineering have facilitated the development of more physiologically relevant platforms, known as Organ-on-Chips (OoC). OoC platforms recapitulate the critical hallmarks of the TME in vitro and subsequently abet in sensitivity and efficacy testing of anti-cancer drugs before clinical trials. The OoC platforms incorporating conventional in vitro models enable researchers to control the cellular, molecular, chemical, and biophysical parameters of the TME in precise combinations while analyzing how they contribute to tumor progression and therapy response. This review discusses the application of OoC platforms integrated with conventional 2D cell lines, 3D organoids and spheroid models, and the organotypic tissue slices, including patient-derived and xenograft tumor slice cultures in cancer treatment responses. We summarize the relevance and drawbacks of conventional in vitro models in assessing cancer treatment response, challenges and limitations associated with OoC models, and future opportunities enabled by the OoC technologies towards developing personalized cancer diagnostics and therapeutics.     

Cellular and molecular mechanisms controlling the migration of melanocytes and melanoma cells

During embryonic development in vertebrates, the neural crest‐derived melanoblasts migrate along the dorsolateral axis and cross the basal membrane separating the dermis from the epidermis to reach their final location in the interfollicular epidermis and epidermal hair follicles. Neoplastic transformation converts melanocytes into highly invasive and metastatic melanoma cells. In vitro, these cells extend various types of protrusions and adopt two interconvertible modes of migration, mesenchymal and amoeboid, driven by different signalling molecules. In this review, we describe the major contributions of natural mouse mutants, mouse models generated by genetic engineering and in vitro culture systems, to identification of the genes, signalling pathways and mechanisms regulating the migration of normal and pathological cells of the melanocyte lineage, at both the cellular and molecular levels.     

Engineering of microscale three-dimensional pancreatic islet models in vitro and their biomedical applications

Diabetes now is the most common chronic disease in the world inducing heavy burden for the people's health. Based on this, diabetes research such as islet function has become a hot topic in medical institutes of the world. Today, in medical institutes, the conventional experiment platform in vitro is monolayer cell culture. However, with the development of micro- and nano-technologies, several microengineering methods have been developed to fabricate three-dimensional (3D) islet models in vitro which can better mimic the islet of pancreases in vivo. These in vitro islet models have shown better cell function than monolayer cells, indicating their great potential as better experimental platforms to elucidate islet behaviors under both physiological and pathological conditions, such as the molecular mechanisms of diabetes and clinical islet transplantation. In this review, we present the state-of-the-art advances in the microengineering methods for fabricating microscale islet models in vitro. We hope this will help researchers to better understand the progress in the engineering 3D islet models and their biomedical applications such as drug screening and islet transplantation.     

Chemosensory behavioural response of freshwater phytoplanktonic flagellates

Unlike behavioural responses to physical gradients, active responses to chemical gradients, and their physiological and ecological implications, have rarely been studied in freshwater phytoplanktonic flagellates. This study used microscale preference chambers to investigate the population and individual cell responses of five species exposed to four chemical gradients which commonly develop with depth in lakes: phosphate, oxygen, carbon dioxide and pH. Upon exposure of nutrient‐replete and ‐depleted cells to a phosphate gradient, only nutrient‐depleted cells of the autotrophic Chlamydomonas moewusii responded, accumulating at high concentrations. In contrast, all species responded to an oxygen gradient with Ceratium furcoides, Chlamydomonas moewusii, Dinobryon sertularia and Plagioselmis nannoplanctica preferring high concentrations whereas Euglena gracilis preferred low concentrations. In addition, all species displayed a strong affinity for carbon dioxide which was not mediated by detecting pH. Analysis of the swimming trajectories of individual cells showed that directed chemotaxes, rather than speed‐dependent chemokineses, were responsible for the observed preferences. These complex and diverse species‐dependent chemosensory responses may optimize photosynthesis, facilitate nutrient retrieval during migration, increase growth rate and may influence spatial and temporal distribution, contributing to the delineation of niche separation in phytoplanktonic flagellates.     

Pollen representation of vegetational patterns along an elevational gradient

Abstract. Modern pollen assemblages from 16 small lakes (< 2.5 ha) and 11 moderate-sized lakes (4.5–19.3 ha) arrayed along an elevational gradient (300 to 1320 m) in the east-central Adirondack Mountains were studied to determine how well the pollen assemblages recorded patterns of forest composition along the gradient. Forest composition ranges from Pinus strobus/Tsuga/ hardwoods forests at low elevations through Tsuga/ hardwoods, hardwoods, and Picea/Abies forests to Abies-dominated forests at high elevations. Modern pollen percentages for 10 tree taxa were compared with lake elevation using scatter plots and correlation and regression analysis. Differential smoothing of vegetational patterns along the elevational gradient occurred among the 10 taxa owing to differences in pollen dispersibility, pollen production, and spatial pattern of taxon abundance in forests of the region. No differences were observed in pollen-elevation patterns between small and moderate-sized lakes. Pollen-elevation patterns were obscured for most taxa when the gradient was shortened (e.g. to 600–1320 m) owing to increased spatial smoothing by pollen dispersal. Design and interpretation of paleoecological studies of spatial gradients can be improved by careful attention to site spacing, gradient length, and gradient steepness in the context of pollen dispersal and representation models.     

The endemic headwater stream amphibians of the American Northwest: associations with environmental gradients in a large forested preserve

We used a large forested preserve (Olympic National Park, USA) to examine the habitat associations of a unique and environmentally sensitive stream amphibian fauna: Ascaphus truei Stegneger, Rhyacotriton olympicus (Gaige) and Dicamptodon copei Nussbaum. We quantified the relative abundance of stream amphibians and compared them to physical, topographic, climatic and landscape variables. All three species were associated with the south‐west to north‐east climate gradient, tending to be most abundant in the south‐west. Although a habitat generalist relative to the other two species, Dicamptodon copei was absent from the north‐eastern portion of the park. Ascaphus truei and Rhyacotriton olympicus were both associated with coarse substrates and steep gradients. Unlike studies in harvested forests, all stream amphibians were common in waters with unconsolidated surface geology (e.g. marine sediments that erode easily). Studies of ecological preserves can provide an important baseline for evaluating species associations with environmental gradients and can reveal patterns not evident in more disturbed landscapes.     

Models of Plant Disease Epidemics. II: Gradients of Bean Rust

The spread of bean rust from three types of inoculum sources (area, line, and point) was studied under field conditions in two seasons. Disease intensity (y) over time was quantified as incidence of diseased plants, incidence of diseased leaves, and disease severity at distances (d) of 0.3, 0.7, 1.5, 2.7, 4.3, 6.5, and 8.7 m from the inoculum sources. Seven curvilinear models were compared and the models that best explained the gradients of bean rust were y = ad^?b exp(cd), y = a exp(–bd^c) and y = a exp(–bd); where a was proportion of disease near the source, d was distance in metres, b was the slope, and c was a shape parameter. For general analysis to compare the slopes of the gradients, the gradient curves were linearized by ln(y) versus d and the parameters ln(a) (intercept) and b (slope) were estimated. The three measures of disease intensity and the three types of inoculum sources had estimated gradient parameters that were statistically different. The incidence of diseased plants rapidly approached y = 1.0 and the gradient slopes were near zero. Also with the ln(y) linearization, the gradients of the incidence of diseased leaves had slopes flatter than the gradients of disease severity. The gradient slopes from area sources of inoculum were flatter than from line sources, which in turn were flatter than from point sources. Late in the epidemics, the b-values became similar for all combinations of inoculum sources × assessment types. The interpretation of flattening of the gradients was confounded because aberrations in transformations occurred with most models. That is, when the intensity of disease (incidence or severity) approached the maximum, the increases in the intercepts were restricted and this restriction was responsible for most of the flattening. No flattening of gradients occurred over time when the gompit gradient model [–ln(–ln(y/y_max)) = –ln(–ln(a))?b ln(d)] was used for these same disease values. The fitting of models to the gradients served little purpose except to classify the curves in a general way. A new statistic, the area under the disease gradient curve (AUDGC), is proposed to compare epidemics.     

Advanced models of human skeletal muscle differentiation,development and disease: Three-dimensional cultures,organoids and beyond

Advanced in vitro models of human skeletal muscle tissue are increasingly needed to model complex developmental dynamics and disease mechanisms not recapitulated in animal models or in conventional monolayer cell cultures. There has been impressive progress towards creating such models by using tissue engineering approaches to recapitulate a range of physical and biochemical components of native human skeletal muscle tissue. In this review, we discuss recent studies focussed on developing complex in vitro models of human skeletal muscle beyond monolayer cell cultures, involving skeletal myogenic differentiation from human primary myoblasts or pluripotent stem cells, often in the presence of structural scaffolding support. We conclude with our outlook on the future of advanced skeletal muscle three-dimensional cultures (e.g. organoids and biofabrication) to produce physiologically and clinically relevant platforms for disease modelling and therapy development in musculoskeletal and neuromuscular disorders.     

A novel technique to evaluate interactions between Saccharomyces cerevisiae cell wall and mycotoxins: application to zearalenone

Three models based on sigmoidal plotting were tested for their ability to describe zearalenone adsorption on Saccharomyces cerevisiae cell walls in vitro. All three models closely fitted the experimental data, but Hill's equation gave the most accurate parameters, and provided information on the physical and chemical mechanisms involved in the adsorption of mycotoxin on yeast cell walls.     

Searching for a model for use in vegetation analysis

Summary Indirect gradient analysis methods require an explicit vegetation model which must be based on direct gradient analysis studies. Various vegetation models are reviewed. Field evidence for the models is discussed. Experimental studies of species response to environmental gradients are reviewed and discussed. Three types of gradient are recognized as important for development of models: indirect environmental gradients where the environmental factor has no direct physiological influence on plant growth e.g. elevation; direct environmental gradients where the factor has a direct physiological effect on growth but is not an essential resource, e.g. pH; resource gradients where the factor is an essential resource for plant growth. The behaviour of the ecological carrying capacity and the role of competition along such gradients are shown to be important for developing vegetation models.     

A hybrid microfluidic-vacuum device for direct interfacing with conventional cell culture methods

Background

Microfluidics is an enabling technology with a number of advantages over traditional tissue culture methods when precise control of cellular microenvironment is required. However, there are a number of practical and technical limitations that impede wider implementation in routine biomedical research. Specialized equipment and protocols required for fabrication and setting up microfluidic experiments present hurdles for routine use by most biology laboratories.

Results

We have developed and validated a novel microfluidic device that can directly interface with conventional tissue culture methods to generate and maintain controlled soluble environments in a Petri dish. It incorporates separate sets of fluidic channels and vacuum networks on a single device that allows reversible application of microfluidic gradients onto wet cell culture surfaces. Stable, precise concentration gradients of soluble factors were generated using simple microfluidic channels that were attached to a perfusion system. We successfully demonstrated real-time optical live/dead cell imaging of neural stem cells exposed to a hydrogen peroxide gradient and chemotaxis of metastatic breast cancer cells in a growth factor gradient.

Conclusion

This paper describes the design and application of a versatile microfluidic device that can directly interface with conventional cell culture methods. This platform provides a simple yet versatile tool for incorporating the advantages of a microfluidic approach to biological assays without changing established tissue culture protocols.     

Fabricating tissues: Analysis of farming versus engineering strategies

Tissue Engineering has expanded rapidly towards target applications of tissue repair and regeneration, whilst generating surprisingly novel models to study tissue modelling. However, clinical success in producing effective engineered tissues such as bone, skin, cartilage, and tendon, have been rare and limited. Problems tend to focus on how to stimulate the replacement of initial scaffold with mechanically functional, native extracellular matrix (principally collagen). Typical approaches have been to develop perfused and mechanically active bioreactors, with the use of native collagen itself as the initial scaffold, though the idea remains that cells do the fabrication (i.e. a cultivation process). We have developed a new, engineering approach, in which the final collagen template is fabricatedwithout cell involvement. The first part of this biomimetic engineering involves a plastic compression of cellular native collagen gels to form dense, strong, collagenous neotissues (in minutes). Further steps can be used to orientate and increase collagen fibril diameter, again by non-cell dependent engineering. This allows operator control of cell or matrix density and material properties (influencing biological half life and fate). In addition, this (non-cultivation) approach can incorporate techniques to generate localised 3D structures and zones at a meso-scale. In conclusion, the use of biomimetic engineering based on native collagen, rather than cell-cultivation approaches for bulk matrix fabrication, produces huge benefits. These include speed of fabrication (minutes instead of weeks and months), possibility of fine control of composition and 3D nano-micro scale structure and biomimetic complexity.     

Preconditioning influences mesenchymal stem cell properties in vitro and in vivo

Various diseases and toxic factors easily impair cellular and organic functions in mammals. Organ transplantation is used to rescue organ function, but is limited by scarce resources. Mesenchymal stem cell (MSC)‐based therapy carries promising potential in regenerative medicine because of the self‐renewal and multilineage potency of MSCs; however, MSCs may lose biological functions after isolation and cultivation for a long time in vitro. Moreover, after they are injected in vivo and migrate into the damaged tissues or organs, they encounter a harsh environment coupled with death signals due to the inadequate tensegrity structure between the cells and matrix. Preconditioning, genetic modification and optimization of MSC culture conditions are key strategies to improve MSC functions in vitro and in vivo, and all of these procedures will contribute to improving MSC transplantation efficacy in tissue engineering and regenerative medicine. Preconditioning with various physical, chemical and biological factors is possible to preserve the stemness of MSCs for further application in studies and clinical tests. In this review, we mainly focus on preconditioning and the corresponding mechanisms for improving MSC activities in vitro and in vivo; we provide a glimpse into the promotion of MSC‐based cell therapy development for regenerative medicine. As a promising consequence, MSC transplantation can be applied for the treatment of some terminal diseases and can prolong the survival time of patients in the near future.     

Ecological influences on larval chironomid communities in shallow lakes: implications for palaeolimnological interpretations

1. To correctly interpret chironomid faunas for palaeoenvironmental reconstruction, it is essential that we improve our understanding of the relative influence of ecosystem variables, biotic as well as physicochemical, on chironomid larvae. To address this, we analysed the surface sediments from 39 shallow lakes (29 Norfolk, U.K., 10 Denmark) for chironomid head capsules, and 70 chironomid taxa (including Chaoborus) were identified. 2. The shallow lakes were selected over large environmental gradients of aquatic macrophytes, total phosphorus (TP) and fish communities. Redundancy analysis (RDA) identified two significant variables that explained chironomid distribution: macrophyte species richness (P < 0.001) and TP (P < 0.005). Generalised linear models (GLM) identified specific taxa that had significant relationships with both these variables. Macrophyte percentage volume infested (PVI) and species richness were significant in classifying the lake types based on chironomid communities under twinspan analysis, although other factors, notably nutrient concentrations and fish communities, were also important, illustrating the complexities of classifying shallow lake ecosystems. Lakes with plant species richness >10 all had relatively diverse (Hill’s N2) chironomid assemblages, and lakes with Hill’s N2 >10 all had TP <250 μg L⁻¹ and total fish densities <2 fish per m². 3. Plant density (PVI), and perhaps more importantly species richness, were primary controls on the distribution of chironomid communities within these lakes. This clearly has implications for palaeoenvironmental reconstructions using zoobenthos remains (i.e. chironomids) and suggests that they could be used to track changes in benthic/pelagic production and could be used as indicators of changing macrophyte habitat. 4. Measuring key biological gradients, in addition to physicochemical gradients, allowed the major controls on chironomid distribution to be assessed more directly, in terms of plant substrate, food availability, competition and predation pressure, rather than implying indirect mechanisms through relationships with nutrients. Many of these variables, notably macrophyte abundance and species richness, are not routinely measured in such studies, despite their importance in determining zoobenthos in temperate shallow lakes. 5. When physical, chemical and ecological gradients are considered, as is often the case with palaeo‐reconstructions rather than training sets chosen to maximise one gradient, complex relationships exist, and attempting to reconstruct a single trophic variable quantitatively may not be appropriate or reliable.     

Cooperative Roles of SDF-1α and EGF Gradients on Tumor Cell Migration Revealed by a Robust 3D Microfluidic Model

Chemokine-mediated directed tumor cell migration within a three dimensional (3D) matrix, or chemoinvasion, is an important early step in cancer metastasis. Despite its clinical importance, it is largely unknown how cytokine and growth factor gradients within the tumor microenvironment regulate chemoinvasion. We studied tumor cell chemoinvasion in well-defined and stable chemical gradients using a robust 3D microfluidic model. We used CXCL12 (also known as SDF-1α) and epidermal growth factor (EGF), two well-known extracellular signaling molecules that co-exist in the tumor microenvironment (e.g. lymph nodes or intravasation sites), and a malignant breast tumor cell line, MDA-MB-231, embedded in type I collagen. When subjected to SDF-1α gradients alone, MDA-MB-231 cells migrated up the gradient, and the measured chemosensitivity (defined as the average cell velocity along the direction of the gradient) followed the ligand – receptor (SDF-1α – CXCR4) binding kinetics. On the other hand, when subjected to EGF gradients alone, tumor cells increased their overall motility, but without statistically significant chemotactic (directed) migration, in contrast to previous reports using 2D chemotaxis assays. Interestingly, we found that the chemoinvasive behavior to SDF-1α gradients was abrogated or even reversed in the presence of uniform concentrations of EGF; however, the presence of SDF-1α and EGF together modulated tumor cell motility cooperatively. These findings demonstrate the capabilities of our microfluidic model in re-creating complex microenvironments for cells, and the importance of cooperative roles of multiple cytokine and growth factor gradients in regulating cell migration in 3D environments.     

IGF-1和动态微环境对脂肪干细胞向心肌细胞分化作用的研究

体外组织工程模型中,生物化学和机械信号对心肌再生起着很重要的促进作用,对人胰岛素样生长因子(IGF-1)和三维动态微环境对脂肪干细胞向心肌细胞分化过程中的促进作用进行了研究．带有IGF-1基因的质粒整合到胶原-壳聚糖支架中,脂肪干细胞接种到整合质粒的支架内,未整合质粒的支架作为对照组,心肌细胞培养基作为分化培养基,转瓶生物反应器提供动态微环境．经2周分化培养后,检测质粒在支架内释放及表达情况、细胞在支架内的活性以及心肌功能性蛋白和基因的表达．结果表明：动态微环境能促进质粒DNA的释放和转染;IGF-1可促进脂肪干细胞在胶原-壳聚糖支架内增殖以及向心肌细胞分化;动态微环境可加强IGF-1的促增殖分化作用．因此,IGF-1和动态微环境能独立或相互促进脂肪干细胞在胶原-壳聚糖支架内活性,动态微环境还可强化IGF-1对脂肪干细胞的促分化作用．对体外构建工程化心肌组织进行心肌再生研究有着重要的指导意义．     

Spatial diversity patterns of Pristimantis frogs in the Tropical Andes

Although biodiversity gradients have been widely documented, the factors governing broad‐scale patterns in species richness are still a source of intense debate and interest in ecology, evolution, and conservation biology. Here, we tested whether spatial hypotheses (species–area effect, topographic heterogeneity, mid‐domain null model, and latitudinal effect) explain the pattern of diversity observed along the altitudinal gradient of Andean rain frogs of the genus Pristimantis. We compiled a gamma‐diversity database of 378 species of Pristimantis from the tropical Andes, specifically from Colombia to Bolivia, using records collected above 500 m.a.s.l. Analyses were performed at three spatial levels: Tropical Andes as a whole, split in its two main domains (Northern and Central Andes), and split in its 11 main mountain ranges. Species richness, area, and topographic heterogeneity were calculated for each 500‐m‐width elevational band. Spatial hypotheses were tested using linear regression models. We examined the fit of the observed diversity to the mid‐domain hypothesis using randomizations. The species richness of Pristimantis showed a hump‐shaped pattern across most of the altitudinal gradients of the Tropical Andes. There was high variability in the relationship between area and species richness along the Tropical Andes. Correcting for area effects had little impact in the shape of the empirical pattern of biodiversity curves. Mid‐domain models produced similar gradients in species richness relative to empirical gradients, but the fit varied among mountain ranges. The effect of topographic heterogeneity on species richness varied among mountain ranges. There was a significant negative relationship between latitude and species richness. Our findings suggest that spatial processes partially explain the richness patterns of Pristimantis frogs along the Tropical Andes. Explaining the current patterns of biodiversity in this hot spot may require further studies on other possible underlying mechanisms (e.g., historical, biotic, or climatic hypotheses) to elucidate the factors that limit the ranges of species along this elevational gradient.