Cell Biology

There are two ways how the emergence of the Biosphere as a single living being could be understood. One, that it is the latest stage of the evolution of life on Earth that began some 4 billion years ago, from smallest and simplest organisms to the larger and more complex. Another would be, that entire development of life was just an embryonic growth of the largest the most complex living being on Earth. Since the Biosphere is and will be quite a complex organism, it is almost certain that will acquire some kind of intelligence as a single being (entity). Although it will be dominated by the human component of its being, it will include some other living beings as well. However, how this intelligence will manifest itself is hard to anticipate or imagine at this point. In addition to possible self-awareness, it might be able to acquire some other properties unknown even unimaginable to us.

Parasitic protozoa of the genus Leishmania infect mammalian mononuclear phagocytic cells causing a potentially fatal disease with a broad spectrum of clinical manifestations. The drugs of choice used in the leishmaniasis therapy are significantly toxic, expensive and faced with a growing frequency of refractory infections. Thus the search for new leishmanicidal compounds is urgently required. In order to perform a proper drug design and to understand the modes of action of such compounds it is necessary to elucidade the intrincate cellular and molecular events that orchestrate the parasite biology. In order to invade the host cell Leishmania are able to recruit different surface receptors which may assist engaging the microbicidal responses. In the intracellular millieu these pathogens can deactivate and/or subvert the phagocyte signal transduction machinery rendering these cells permissive to infection. In the present review we attempted to approach some of the most relevant cellular and biochemical invasion and evasion strategies employed by Leishmania parasites.

Plants deploy a finely tuned balance between growth and defence responses for better fitness. Crosstalk between defence signalling hormones such as salicylic acid (SA) and jasmonates (JAs) as well as growth regulators plays a significant role in mediating the trade-off between growth and defence in plants. Here, we specifically discuss how the mutual antagonism between the signalling of auxin and SA impacts on plant growth and defence. Furthermore, the synergism between auxin and JA benefits a class of plant pathogens. JA signalling also poses growth cuts through auxin. We discuss how the effect of cytokinins (CKs) is multifaceted and is effective against a broad range of pathogens in mediating immunity. The synergism between CKs and SA promotes defence against biotrophs. Reciprocally, SA inhibits CK-mediated growth responses. Recent reports show that CKs promote JA responses; however, in a feedback loop, JA suppresses CK responses. We also highlight crosstalk between auxin and CKs ...

X-ray microcomputed tomography (mCT) allows nondestructive visualisation of plant root systems within their soil environment and thus offers an alternative to the commonly used destructive methodologies for the examination of plant roots and their interaction with the surrounding soil. Various methods for the recovery of root system information from X-ray computed tomography (CT) image data have been presented in the literature. Detailed, ideally quantitative, evaluation is essential, in order to determine the accuracy and limitations of the proposed methods, and to allow potential users to make informed choices among them. This, however, is a complicated task. Three-dimensional ground truth data are expensive to produce and the complexity of X-ray CT data means that manually generated ground truth may not be definitive. Similarly, artificially generated data are not entirely representative of real samples. The aims of this work are to raise awareness of the evaluation problem and to propose experimental approaches that allow the performance of root extraction methods to be assessed, ultimately improving the techniques available. To illustrate the issues, tests are conducted using both artificially generated images and real data samples.

Sox2 has yet to be determined. Sox2 can act synergistically with Oct3/4 in vitro to activate Oct-Sox enhancers, which regulate the expression of pluripotent stem cellspecific genes, including Nanog, Oct3/4 and Sox2 itself. These findings suggest that Sox2 is required by ES cells for its Oct-Sox enhancer activity. Using inducible Sox2-null mouse ES cells, we show that Sox2 is dispensable for the activation of these Oct-Sox enhancers. In contrast, we demonstrate that Sox2 is necessary for regulating multiple transcription factors that affect Oct3/4 expression and that the forced expression of Oct3/4 rescues the pluripotency of Sox2-null ES cells. These results indicate that the essential function of Sox2 is to stabilize ES cells in a pluripotent state by maintaining the requisite level of Oct3/4 expression.

We have investigated the developmental transitions of myosin heavy chain (MHC) gene expression in the rat extraocular musculature (EOM) at the mRNA level using Sl-nuclease mapping techniques and at the protein level by polypeptide mapping and ...

Aberrant keratinocyte differentiation is considered to be a key mechanism in the onset of hyperproliferative dermatological diseases, including basal cell carcinoma (BCC). It is, therefore, vital to understand what drives keratinocytes to develop such pathological phenotypes.

Autofluorescent protein tags represent one of the major and, perhaps, most powerful tools in modern cell biology for visualization of various cellular processes in vivo. In addition, advances in confocal microscopy and the development of autofluorescent proteins with different excitation and emission spectra allowed their simultaneous use for detection of multiple events in the same cell. Nevertheless, while autofluorescent tags are widely used in plant research, the need for a versatile and comprehensive set of vectors specifically designed for fluorescent tagging and transient and stable expression of multiple proteins in plant cells from a single plasmid has not been met by either the industrial or the academic communities. Here, we describe a new modular satellite (SAT) vector system that supports N-and C-terminal fusions to five different autofluorescent tags, EGFP, EYFP, Citrine-YFP, ECFP, and DsRed2. These vectors carry an expanded multiple cloning site that allows easy exchange of the target genes between different autofluorescence tags, and expression of the tagged proteins is controlled by constitutive promoters, which can be easily replaced with virtually any other promoter of interest. In addition, a series of SAT vectors has been adapted for high throughput Gateway recombination cloning. Furthermore, individual expression cassettes can be assembled into Agrobacterium binary plasmids, allowing efficient transient and stable expression of multiple autofluorescently tagged proteins from a single vector following its biolistic delivery or Agrobacteriummediated genetic transformation.

The present study was undertaken to examine cell cycle characteristics of endangered Goral (CITES Appendix I) adult skin fibroblasts. Seven experiments were performed, each with a one-way completely randomized design involving three replicates. Least significant difference (LSD) was used to determine variation among treatment groups. Experiment I focused on the effects of cycling, serum-starved, and fully confluent stages of Goral cells. In Experiments II and III, the effects of different antioxidants like b-mercaptoethanol (b-ME, 10 mM), cysteine (2 mM), and glutathione (2 mM) were examined after cells were fully confluent without serum starvation for 24 h and 4 h, respectively. In Experiments IV and V, three protease inhibitors, namely 6-dimethylaminopurine (6-DMAP, 2 mM), cycloheximide (7.5 mg/ml) and cytochalasin B (7.5 mg/ml), were used as in Experiment II. In Experiments VI and VII, the effect of different levels of dimethylsulphoxide (DMSO) at 0%, 0.5%, 1.0% and 2.5% were tested by flow cytometry (FACS). In Experiment I, 68.7% of Goral skin fibroblasts reached the G 0 /G 1 stage (2C DNA content) when subjected to the serum-starved medium, which was more than the cycling (64.9%) and fully confluent groups (61.0%) (P > 0.05). Among the chemically treated group, b-ME, cysteine and DMSO showed better results for synchronization of G 0 þ G 1 phases than cycling, serum-starved and fully confluent groups. It can thus be concluded that b-ME, cysteine and DMSO at certain concentrations can synchronize the cell cycle effectively, which could have a positive impact on somatic cell nuclear transfer in the goral. Ó

Molecular imaging is the visualization, characterization, and measurement of biologic processes at the molecular and cellular levels in humans and other living systems (1). It comprises an emerging set of technologies that builds on advances in imaging procedures (e.g., PET, SPECT, MRI, ultrasound, optical, and photoacoustic), improved understanding of biology, and the development of molecularly targeted agents. These continuously expanding sets of imaging methods are often used in combination, and advances in data acquisition and analyses facilitate a more complete understanding of biology. Molecular imaging aims to improve our understanding of mammalian biology and lead to advances in patient care by providing targeted therapies that will enable personalized medicine and the imaging tools to assess outcome. Implementation of these new technologies in clinical care has many educational, technical, and regulatory challenges that must be overcome before molecular imaging reaches its full potential. The impact of molecular imaging has been significant in several disciplines, because it represents a paradigm shift in how scientists and clinicians can observe biology in real time and in a relatively noninvasive manner to enable the power of repeated measures in living organisms.

In order to create a hemodynamic environment that can simulate the physiological condition of arteries, an in vitro experiment apparatus was designed whose key modules were compliance chamber and after-load. These two modules were developed based on the theories of hemodynamics. Both the normal and shear stress to which endothelial cells are exposed can be controlled with these modules, thus facilitating the research of endothelial cells subjected to stresses. Ó

Store-operated calcium entry (SOCE) is the predominant Ca2+ entry mechanism in nonexcitable cells and controls a variety of physiological and pathological processes. Although significant progress has been made in identifying the components required for SOCE, the molecular mechanisms underlying it are elusive. The present study provides evidence for a direct involvement of kinase suppressor of Ras 2 (KSR2) in SOCE. Using lymphocytes and fibroblasts from ksr2−/− mice and shKSR2-depleted cells, we find that KSR2 is critical for the elevation of cytosolic Ca2+ concentration. Specifically, our results show that although it is dispensable for Ca2+-store depletion, KSR2 is required for optimal calcium entry. We observe that KSR2 deficiency affects stromal interaction molecule 1 (STIM1)/ORAI1 puncta formation, which is correlated with cytoskeleton disorganization. Of interest, we find that KSR2-associated calcineurin is crucial for SOCE. Blocking calcineurin activity impairs STIM1/ORAI1 pun...

Cytomics aims to determine the molecular phenotype of single cells. Within the context of the -omics, cytomics allows the investigation of multiple biochemical features of the heterogeneous cellular systems known as the cytomes. Cytomics can be considered as the science of single cell-based analyses that links genomics and proteomics with the dynamics of cell and tissue function, as modulated by external inXuences. Inherent to cytomics are the use of sensitive, scarcely invasive, Xuorescence-based multiparametric methods and the event-integrating concept of individual cells to understand the complexity and behaviour of tissues and organisms. Among cytomic technologies, Xow cytometry, confocal laser scanning microscopy and laser capture microdissection are of great relevance. Other recent technologies based on single cell bioimaging and bioinformatic tools become important in drug discovery and toxicity testing, because of both highcontent and high-troughput. The multiparametric capacity of cytomics is very useful for the identiWcation, characterization and isolation of stem cell populations. In our experience, Xow cytometry is a powerful and versatile tool that allows quantitative analysis of single molecules, prokaryotic and eukaryotic cells for basic, biotechnological, environmental and clinical studies. The dynamic nature of cytomic assays leads to a real-time kinetic approach based on sequential examination of diVerent single cells from a population undergoing a dynamic process, the in Xuxo level. Finally, cytomic technologies may provide in vitro methods alternative to laboratory animals for toxicity assessment.

Recently, a novel approach to a highly sensitive and quantitative detection of rare earth element (REE) ions including La3+, Eu3+ and Tb3+, by the polymerase chain reaction (PCR) technique, has been reported. The detection of REE ions is based on the catalytic nature of REE ions targeting the deoxyribonucleic acid (DNA), thus monitoring of the ions can be achieved by reading the level of intact DNA by PCR. Despite of its high sensitivity (at ppb to ppt levels), the conventional PCR-based REE detection protocol requires certain length of time (1-2 hours). In the present study, we modified the PCR-based REE detection protocols by employing the high-speed PCR, and performed the automated and rapid detection of La3+ in small-sized aqueous samples within 5min.

The interferons (IFNs) were originally described over 50 years ago, identified by their ability to confer viral resistance to cells. We now know that they are much more than just anti-viral cytokines collectively having roles in both innate and adaptive immune responses, in tumor surveillance and defense, and modulation of immune cell function. Three types of IFN have now been described, simply referred to as type I, II and III. Distinguishable by the unique receptors that they rely on for signal transduction, the three types of IFN have specific and varied roles in the maintenance of human health and defense against pathogens. In mounting an IFN-mediated immune response, the human body has developed the ability to regulate IFN-mediated signal transduction. Like all cytokines, the ability of a cell to respond to IFN is completely dependent on the presence of its cognate receptor on the surface of the target cell. Thus, one of the major mechanisms used by the human body to regulate the strength and duration of the IFN response is through regulation of receptor levels, thereby altering the cytokine-specific responsiveness of the target cell. This review will discuss the receptor system utilized by the type I IFNs and compare it with that of the type II and III IFNs, which also regulate immune responses through controlling receptor level on the cell surface.

At present, one is still searching for physical and chemical methods which are suitable for the characterisation of biocompatibility of implant surfaces. In this paper, we analysed differently structured titanium surfaces (the roughness average of the various structures differs by four orders of magnitude) with a number of electrochemical and physical methods such as electrochemical impedance spectroscopy, chronoamperometry, linear sweep voltammetry, surface profiling and scanning electron microscopy. The aim of the work is to compare the specific methods regarding their significance, and to correlate the results. Beside the extraction of physical and electrochemical parameters which are typical for the above-mentioned methods, the possibility of the determination of the fractal dimension of the surface structures is also examined. Cell biological examinations concerning the mechanism of cell adhesion were used to verify the ability of the parameters measured by the referred methods to describe the biocompatibility of the analysed surfaces. D

A joint evaluation of all animals, uses all additive genetic relationships uses all data on all animal jointly It works as a linear model (correcting different effects for each other), jointly estimates animal effects and fixed effects (herds) but has Selection Index properties (Regression with " heritability ")  1 0 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 0 0 0 0 1 0 0 0 0 1

The kelch motif was discovered as a sixfold tandem element in the sequence of the Drosophila kelch ORF1 protein. The repeated kelch motifs predict a conserved tertiary structure, a β-propeller. This module appears in many different polypeptide contexts and contains multiple potential protein–protein contact sites. Members of this growing superfamily are present throughout the cell and extracellularly and have diverse activities. In this review, we discuss current information concerning the structural organization of kelch repeat proteins, their biological roles and the molecular basis of their action.

The Respiratory Syncytial Virus 2003 symposium took place from 8th–11th November 2003 in Stone Mountain, Georgia, and brought together more than 200 international investigators engaged in RSV research. RSV biology, pathogenesis, and clinical data, as well as RSV vaccines and antivirals, were addressed in the meeting, and this review will aim to briefly summarize and discuss the implications of new findings. The meeting also served as the inauguration of the Robert M. Chanock Award for lifetime achievement in RSV research, an award named in honor of the person who started the field of RSV research by recovering the first human RS virus from infants with severe bronchiolitis in 1956.

Exposure to particulate matter (PM) may alter lung homeostasis inducing changes in fluid balance and host defense. Bioavailability of soluble PM compounds like polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and transition metals has been shown to play a key role in lung injury. We have previously characterized the size, shape, and chemical components of urban air particles from Buenos Aires (UAP-BA) and their biological impact on lungs. Herein, we evaluate the possible toxic effect of UAP-BA-soluble fraction (UAP-BAsf) on pulmonary cells obtained from young (1-2 months old) and aged (9-12 months old) Wistar rats using phagocytosis, oxidant-antioxidant generation, and apoptosis as endpoints. UAP-BA were collected in downtown BA and residual oil fly ash (ROFA), employed as a positive control, was collected from Boston Edison Co., Mystic Power Plant, Mystic, CT, USA. Both particle-soluble fractions (sf) were employed at concentrations ranging from 0 to 100 m...

An effect of magnesium on calcium-induced depolarisation of mitochondrial transmembrane potential (DJ m ) was investigated. Depending on the presence of Mg 2þ , addition of Ca 2þ to suspension of isolated rat heart mitochondria induced either reversible depolarisation or irreversible collapse of succinate-driven DJ m . Irreversible collapse of DJ m , observed in the absence of Mg 2þ , was insensitive to Ca 2þ chelation, inhibition of Ca 2þ uptake and increased efflux of Ca 2þ from mitochondrial matrix. Based on these data, opening of mPTP in a high-conductance mode is considered to be a major cause of the Ca 2þ -induced irreversible collapse of DJ m in the absence of Mg 2þ . Involvement of mPTP in the process of Ca 2þ -induced collapse of DJ m was further supported by protective effect of both CsA and ADP. Reversible collapse of DJ m , observed in the presence of Mg 2þ , was sensitive to EGTA, ADP; and inhibition of Ca 2þ uptake and increased efflux of Ca 2þ from mitochondrial matrix. This may represent selective induction of a low-conductance permeability pathway. Presented results indicate important role of Mg 2þ in the process of Ca 2þ -induced depolarisation of DJ m mainly through discrimination between low-and high-conductance modes of mPTP. Minor effect of Mg 2þ on Ca 2þ -induced depolarisation of DJ m was observed at the level of stimulation of DJ m generation and inhibition of mitochondrial Ca 2þ uptake. Ó

| Although cancer is a diverse set of diseases, cancer cells share a number of adaptive hallmarks. Dysregulated pH is emerging as a hallmark of cancer because cancers show a 'reversed' pH gradient with a constitutively increased intracellular pH that is higher than the extracellular pH. This gradient enables cancer progression by promoting proliferation, the evasion of apoptosis, metabolic adaptation, migration and invasion. Several new advances, including an increased understanding of pH sensors, have provided insight into the molecular basis for pH-dependent cell behaviours that are relevant to cancer cell biology. We highlight the central role of pH sensors in cancer cell adaptations and suggest how dysregulated pH could be exploited to develop cancer-specific therapeutics. PERSPECTIVES NATURE REVIEWS | CANCER VOLUME 11 | SEPTEMBER 2011 | 671

Biological organisms have taken part in a journey that has lasted the better part of the last few million years. The organisms we know today -including ourselves -are just a speck on the timeline of all evolutionary history. The origins of every mammal, bird, fish, and reptile is a subject of fascination for all in the scientific world. The two classes of cells which make up organisms are prokaryotic and eukaryotic cells. Endosymbiont Theory, first proposed by Konstantin Mereschkowski, states that the eukaryotic cells we find in ourselves today began from prokaryotes, originating from a moment of symbiosis which took place between "separate, single-celled organisms" around 1.2-1.5 billion years ago. (Waseemuddin, 2016) This paper will explore the evidence which substantiates Mereschkowski's theory and look at the counter theory of Symbiogenesis.

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During waterlogging conditions plants switch from aerobic respiration to anaerobic fermentation to cope with the lack of available oxygen. Plants have two main fermentation pathways: ethanol and lactic acid fermentation. In this paper we carry out a functional analysis of the Arabidopsis lactate dehydrogenase gene, LDH1. Our results indicate that LDH1, like some other anaerobic genes, is expressed in a root-specific manner and is affected by a variety of abiotic stresses (hypoxia, drought, cold) and mechanical wounding. Functional analysis of LDH1 was carried out using transgenic Arabidopsis overexpressing the gene (35S promoter) and a T-DNA knockout line. Overexpression of LDH1 resulted in improved survival of low oxygen stress conditions in roots but not in shoots. Increased lactic acid fermentation also resulted in significantly higher activities of pyruvate decarboxylase (PDC). Knockout mutants of LDH1 showed reduced survival under low oxygen conditions and PDC activity levels w...

In the present review we analyse the key features of cells, seen fundamentally as information processing machines. Even though the comparison between cells and machines or cells and computers provides a powerful tool for understanding how information is stored and processed by the cellular machinery, a series of misleading and simplistic assumptions are critically reviewed. Moreover, the seminal studies on understanding and artificially recreating the neuronal structure are analysed, both for exploiting its powerful computational abilities in information processing devices and for providing medical solutions to neurodegenerative diseases, neurological trauma and neuroprosthetics.

Poucos contestarão a afirmação de que a agricultura é uma das maiores invenções da humanidade. A criação sistemática de animais, em particular, permite produzir carne, lacticínios e numerosos derivados indispensáveis à nossa sobrevivência. No entanto, a procura crescente por alimento provocada pelo crescimento demográfico originou níveis de industrialização da criação animal pouco sustentáveis ambientalmente. A agricultura celular surge como uma nova área da biotecnologia que se propõe produzir produtos animais a partir de células sem necessidade de criar os animais. Esta ideia de produzir carne em laboratório seria inevitavelmente catalogada como ficção científica há poucos anos. No entanto, os avanços dos últimos anos sugerem que se comece a encarar a agricultura celular como uma solução real para muitos dos problemas da criação animal intensiva.

Hyperphosphorylated forms of the microtubule-associated protein (MAP) tau accumulate in Alzheimer's disease and related tauopathies and are thought to have an important role in neurodegeneration. However, the mechanisms through which phosphorylated tau induces neurodegeneration have remained elusive. Here, we show that tau-induced neurodegeneration is associated with accumulation of filamentous actin (F-actin) and the formation of actin-rich rods in Drosophila and mouse models of tauopathy. Importantly, modulating F-actin levels genetically leads to dramatic modification of tau-induced neurodegeneration. The ability of tau to interact with F-actin in vivo and in vitro provides a molecular mechanism for the observed phenotypes. Finally, we show that the Alzheimer's disease-linked human β-amyloid protein (Aβ) synergistically enhances the ability of wild-type tau to promote alterations in the actin cytoskeleton and neurodegeneration. These findings raise the possibility that a direct interaction between tau and actin may be a critical mediator of tau-induced neurotoxicity in Alzheimer's disease and related disorders.

The heme pathway enzyme d-aminolevulinate dehydratase is a good marker for oxidative stress and metal intoxication. This sulfhydryl enzyme is inhibited in such oxidative pathologies as lead, mercury and aluminum intoxication, exposure to selenium organic species and diabetes. Oxidative stress is a complicating factor in diabetes, inducing non-enzymatic glucose-mediated reactions that change protein structures and impair enzyme functions. We have studied the effects of high glucose, fructose and ribose concentrations on d-ALA-D activity in vitro. These reducing sugars inhibited d-ALA-D with efficacies in the order fructose Z ribose O glucose. The possible mechanism of glucose inhibition was investigated using lysine, DTT, and t-butylamine. Oxidation of the enzyme's critical sulfhydryl groups was not involved because DTT had no effect. We concluded that high concentrations of reducing sugars or their autoxidation products inhibit d-ALA-D by a mechanism not related to thiol oxidation. Also, we are not able to demonstrate that the formation of a Schiff base with the critical lysine residue of the enzyme is involved in the inhibition of d-ALA-D by hexoses. Ó

The etiology of motor neuron degeneration in amyotrophic lateral sclerosis (ALS) remains to be better understood. Based on the studies from ALS patients and transgenic animal models, it is believed that ALS is likely to be a multifactorial and multisystem disease. Many mechanisms have been postulated to be involved in the pathology of ALS, such as oxidative stress, glutamate excitotoxicity, mitochondrial damage, defective axonal transport, glia cell pathology and aberrant RNA metabolism. Mitochondria, which play crucial roles in excitotoxicity, apoptosis and cell survival, have shown to be an early target in ALS pathogenesis and contribute to the disease progression. Morphological and functional defects in mitochondria were found in both human patients and ALS mice overexpressing mutant SOD1. Mutant SOD1 was found to be preferentially associated with mitochondria and subsequently impair mitochondrial function. Recent studies suggest that axonal transport of mitochondria along microtubules and mitochondrial dynamics may also be disrupted in ALS. These results also illustrate the critical importance of maintaining proper mitochondrial function in axons and neuromuscular junctions, supporting the emerging "dying-back" axonopathy model of ALS. In this review, we will discuss how mitochondrial dysfunction has been linked to the ALS variants of SOD1 and the mechanisms by which mitochondrial damage contributes to the disease etiology.

In many eukaryotic organisms, gender is determined by a pair of heteromorphic sex chromosomes. Degeneration of the non-recombining Y chromosome is a general facet of sex chromosome evolution. Selective pressure to restore expression levels of X-linked genes relative to autosomes accompanies Y-chromosome degeneration, thus driving the evolution of dosage compensation mechanisms. This review focuses on evolutionary aspects of dosage compensation, in light of recent advances in comparative and functional genomics that have substantially increased our understanding of the molecular mechanisms of dosage compensation and how it evolved. We review processes involved in sex chromosome evolution, and discuss the dynamic interaction between Y degeneration and the acquisition of dosage compensation. We compare mechanisms of dosage compensation and the origin of dosage compensation genes between different taxa and comment on sex chromosomes that apparently lack compensation mechanisms. Finally, we discuss how dosage compensation systems can also influence the evolution of well-established sex chromosomes.

With the continuous development, in the last decades, of analytical techniques providing complex information at single cell level, the study of cell heterogeneity has been the focus of several research projects within analytical biotechnology. Nonetheless, the complex interplay between environmental changes and cellular responses is yet not fully understood, and the integration of this new knowledge into the strategies for design, operation and control of bioprocesses is far from being an established reality. Indeed, the impact of cell heterogeneity on productivity of large scale cultivations is acknowledged but seldom accounted for. In order to include population heterogeneity mechanisms in the development of novel bioprocess control strategies, a reliable mathematical description of such phenomena has to be developed. With this review, we search to summarize the potential of currently available methods for monitoring cell population heterogeneity as well as model frameworks suitable for describing dynamic heterogeneous cell populations. We will furthermore underline the highly important coordination between experimental and modeling efforts necessary to attain a reliable quantitative description of cell heterogeneity, which is a necessity if such models are to contribute to the development of improved control of bioprocesses.

Collagen remodelling by fibroblasts has a crucial role in organizing tissue structures that are essential to motility during wound repair, development and regulation of cell growth. However, the mechanism of collagen fibre movement in threedimensional (3D) matrices is not understood. Here, we show that fibroblast lamellipodia extend along held collagen fibres, bind, and retract them in a 'hand-over-hand' cycle, involving α2β1 integrin. Wild-type fibroblasts move collagen fibres three to four times farther per cycle than fibroblasts lacking myosin II-B (myosin II-B −/−). Similarly, myosin II-B −/− fibroblasts contract 3D collagen gels threefold less than controls. On twodimensional (2D) substrates, however, rates of collagen bead and cell movement are not affected by loss of myosin II-B. Green fluorescent protein (GFP)-tagged myosin II-B, but not II-A, restores normal function in knockout cells and localizes to cell processes, whereas myosin II-A is more centrally located. Additionally, GFP-myosin II-B moves out to the periphery and back during handover hand fibre movement, whereas on 2D collagen, myosin II-B is more centrally distributed. Thus, we suggest that cyclic myosin II-B assembly and contraction in lamellipodia power 3D fibre movements. Remodelling of extracellular matrix proteins is critical to the organization of tissues and organs; therefore, it is important to understand the cellular and molecular steps in single collagen fibre movement. Surprisingly, cell motility on glass and other 2D substrates is not necessarily relevant to the movement of fibres in 3D matrices. There is limited overlap between the proteins localized to 3D matrix contacts and those at 2D adhesions 1,2. Furthermore, the morphology of fibroblasts in a 3D collagen matrix bears little resemblance to their morphology on a 2D collagen-coated surface 3,4. Thus, there is a fundamentally different mode of motility on 3D fibres compared with 2D surfaces 5. Carefully orchestrated movements of actin and myosin underlie most cellular motile processes 6-8. This motile machinery forms the cortical cytoskeleton. Upon binding extracellular matrix proteins, integrin receptors initiate a complex signalling cascade culminating in the

We describe a protocol to rapidly and reliably visualize blood vessels in experimental animals. Blood vessels are directly labeled by cardiac perfusion using a specially formulated aqueous solution containing 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI), a lipophilic carbocyanine dye, which incorporates into endothelial cell membranes upon contact. By lateral diffusion, DiI also stains membrane structures, including angiogenic sprouts and pseudopodial processes that are not in direct contact. Tissues can be immediately examined by conventional and confocal fluorescence microscopy. High-quality serial optical sections using confocal microscopy are obtainable from thick tissue sections, especially at low magnification, for three-dimensional reconstruction. It takes less than 1 h to stain the vasculature in a whole animal. Compared with alternative techniques to visualize blood vessels, including space-occupying materials such as India ink or fluorescent dye-conjugated dextran, the corrosion casting technique, endothelial cell-specific markers and lectins, the present method simplifies the visualization of blood vessels and data analysis.

Multi-proxy approach was used to reconstruct the environmental conditions of remote lakes in the High Tatra Mountains (Slovakia) over the past few centuries (approximately 500-1000 years). Short sediment cores (*30 cm) taken from three morphologically similar glacial lakes distributed along altitudinal gradient (subalpine to alpine conditions) were analysed for organic matter content (LOI), diatoms and chironomids. Both descriptive and correlative approaches were used for analysing stratigraphical data. Predictive canonical correspondence analysis and co-correspondence analysis were applied to directly relate physical and biological proxies to each other. The relationship between LOI and biotic proxies was inconsistent across groups and lakes. Concordant patterns in diatom and chironomid composition were found in two non-acidified lakes. Common trends in those assemblages indicated major past environmental events such as the Little Ice Age, air pollution and lake acidification. In contrast, no relationship between the composition of diatom and chironomid assemblages was found in the formerly acidified lake, suggesting different responses of assemblages to acidification. While chironomids showed shifts that are attributable to recovery, diatoms assemblage remained relatively stable throughout the uppermost layers of the sediment record. On the other hand, climatic-driven changes in assemblages detected in the deeper layers were more pronounced in diatoms than in chironomids.