Cell-environment Interactions Research Articles

Gasotransmitters: novel regulators of ion channels and transporters

More than 25 years ago, it was a big surprise for physiologists that nitric oxide (NO) was identified as the endothelium derived relaxing factor which is responsible for endothelium-induced smooth muscle relaxation (Ignarro et al., 1987). Until then, small gaseous molecules were simply regarded as byproducts of cellular metabolism which were unlikely to be of any physiological relevance. The discovery that NO was synthesized by specific enzymes (NO-synthases), upon stimulation by specific, physiologically relevant stimuli (e.g., acetylcholine stimulation of endothelial cells), as well as the fact that it acted on specific cellular targets (e.g., soluble guanylate cyclase), set the course for numerous studies which investigated the physiological roles of gaseous signaling molecules—in other words, gasotransmitters (Wang, 2002). Aside from NO, there are two more gases which subsequently have been identified as gasotransmitters: carbon monoxide (CO) (Wu and Wang, 2005) and hydrogen sulfide (H2S) (Abe and Kimura, 1996; Wang, 2002). Although the concept of a gas being involved in cellular signaling processes was already accepted since the discovery of NO, the new candidates CO and H2S were initially met with skepticism—especially since they were well-known for their high toxicity and regarded as environmental chemical threats. However, in concert with the principle of Paracelsus that “everything is toxic- it just depends on the dose,” it is nowadays accepted that small amounts of CO and H2S are of physiological relevance and are even endogenously produced in human cells (Wu and Wang, 2005; Wang, 2012). All three gasotransmitters are produced in human cells by specific enzymes: NO is produced by NO-synthases (NOS), originating from the amino acid L-arginine. There are three NOS-isoforms, NOS1-3, which have been originally termed neuronal, inducible and endothelial NOS, respectively (Garvin et al., 2011). CO is generated by heme oxygenases (HO) within heme degradation (Wu and Wang, 2005). There are also three heme oxygenases (HO-1-3) (Yoshida et al., 1974; Maines et al., 1986; McCoubrey et al., 1997), including an inducible HO-1 and a constitutively active HO-2. H2S is mainly produced within the metabolism of L-cysteine by the enzymes cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE or CTH) and 3-mercaptopyruvate sufurtransferase (3-MST) (Wang, 2012). Recently, Kimura's group described an additional pathway for endogenous H2S production, which involves D-cysteine, 3-MST, and D-amino acid oxidase (Shibuya et al., 2013). Despite the knowledge of the substrates and specific enzymes which are involved in gasotransmitter production, the precise endogenous concentrations of NO, CO, and H2S have still not been sufficiently determined. The measurement of endogenous gas concentrations is limited by the accuracy and specificity of currently available methods (Olson, 2013), the reactivity and short half-life of some gases (NO) (Wall et al., 2012), and simply the fact that gases are volatile and rapidly disappear during the preparation procedures of biological samples. As highlighted within this research topic issue, it is crucial—and probably most challenging—to precisely quantify endogenous concentrations of NO, CO, and H2S (Kimura, 2012; Peers, 2012; Olson, 2013). Although precise endogenous concentrations of gasotransmitters remain to be determined, numerous studies investigated the physiological effects of those gaseous signaling molecules in almost every organ system [as summarized in excellent review articles such as Olson (2011), Wu and Wang (2005), and Wang (2012)]. Common molecular targets for all gasotransmitters are ion channels and transporters. Changes in the activity of membrane-located ion channels/transporters are involved in the majority of physiological processes which are regulated by gasotransmitters. This allows the gases to specifically act at the interface of cell-environment interactions, electrolyte homeostasis and electrochemical communication—thus allowing the regulation of numerous physiological processes in cells and tissues. This research topic summarizes currently available data on the regulation of ion channels and transporters by NO, CO, and H2S. Excellent review and research articles highlight the importance of gasotransmitter/ion channel interactions for vegetative physiology (Althaus, 2012; Peers, 2012; Pouokam and Diener, 2012), neurophysiology (Njie-Mbye et al., 2012; Peers, 2012; Takahashi et al., 2012; Wang et al., 2012), and explain gasotransmitter-induced signaling mechanisms (Wall et al., 2012). Furthermore, opinion articles from pioneers in gasotransmitter research (Kimura, 2012; Peers, 2012; Olson, 2013) give perspectives on important routes which should be followed in this field. With the articles included in this issue we wish to highlight the importance of ion channel/transporter regulation by gaseous signaling molecules and wish to stimulate future research in this exciting area of physiology.

Bioengineering Methods for Analysis of Cells In Vitro

Efforts in the interdisciplinary field of bioengineering have led to innovative methods for investigating the complexities of cell responses in vitro. These approaches have emphasized the reduction of complex multicomponent cellular microenvironments into distinct individual signals as a means to both (a) better construct mimics of in vivo microenvironments and (b) better deconstruct microenvironments to study them. Microtechnology tools, together with advances in biomaterials, have been fundamental to this progress by enabling the tightly controlled presentation of environmental cues and the improved systematic analysis of cellular perturbations. In this review, we describe bioengineering approaches for controlling and measuring cell-environmental interactions in vitro, including strategies for high-throughput analysis. We also describe the mechanistic insights gained by the use of these novel tools, with associated applications ranging from fundamental biological studies, in vitro modeling of in vivo processes, and cell-based therapies.

Monitoring chemical impacts on cell cultures by means of image analyses

Many chemical processes are involved in the interactions of living cells with their environment; however, monitoring such processes often requires sophisticated analyzers. In this study, a sensing strategy based on imaging techniques has been developed to (i) enable cell discrimination based on their physical appearance such as size and shape and (ii) to build predictive models that relate the measured cell appearance to chemical parameters in their environment. Both goals aim at innovative and straightforward sensing strategies for analyzing cell–environment interactions. Image analyses offer several advantages such as the use of simpler, more robust sensors and the omission of extensive sample/sensor preparations. Imaging can analyze numerous cells and thus gains a culture representative insight rather than a potentially nonrepresentative single-cell response. As a proof-of-principle application, different species of microalgae cells have been exposed to various nutrient conditions. Microalgae are known to sensitively adapt to changing nutrient conditions and could potentially become biological “probes” for chemical shifts in ecosystems. Because of considerable spreads of cell size and shapes within one class, size and shape distributions have been derived from visible images of cell cultures. It is shown that the novel image analyses are capable of discriminating different cell species based on their cell shapes and sizes. It is also demonstrated that in conjunction with the recently introduced, nonlinear multivariate “predictor surfaces”, the nutrient availability has a quantifiable impact on the cell size distributions. In this application, predictor surfaces are somewhat more precise than partial least squares. Copyright © 2012 John Wiley & Sons, Ltd.

Combining Stem Cells and Tissue Engineering in Cardiovascular Repair - a Step Forward to Derivation of Novel Implants with Enhanced Function and Self-Renewal Characteristics

The use of stem and progenitor cells in cardiovascular therapy has been proposed as a feasible option to promote repair of tissue damage by ischemia, or to devise definitive artificial tissue replacements (valves, vessels, myocardium) to be surgically implanted in patients. Whereas in other medical branches such as dermatology and ophthalmology the use of ex vivo grown tissues is already accessible to a large degree, the use of bio-artificial implants in cardiovascular surgery is still marginal. This represents a major limitation in cardiovascular medicine at present. In fact, the limited durability and the lack of full compatibility of current implantable devices or tissues prevent a long-term resolution of symptoms and often require re-intervention thereby further increasing the economic burden of the cardiovascular disease. Stem cell technology can be of help to derive tissues with improved physiologic function and permanent durability. Specifically, the intrinsic ability of stem cells to produce tissue-specific "niches", where immature cells are perpetuated while differentiated progenitors are continuously produced, makes them an ideal resource for bioengineering approaches. Furthermore, recent advancements in biocompatible material science, designing of complex artificial scaffolds and generation of animal or human-derived natural substrates have made it feasible to have ex vivo reproduction of complex cell environment interactions - a process necessary to improve stem cells biological activity. This review focuses on current understanding of cardiovascular stem cell biology as well as tissue engineering and explores their interdisciplinary approach. By reviewing the relevant recent patents which have enabled this field to advance, it concentrates on various design substrates and scaffolds that grow stem cells in order to materialize the production of cardiovascular implants with enhanced functional and self-renewal characteristics.

Mechanisms of cancer-associated glycosylation changes

Cell membrane glycoconjugates undergo characteristic changes as a consequence of neoplastic transformation. The cancer-associated carbohydrate structures play key roles in cancer progression by altering the cell-cell and cell-environment interactions. In this review, we will discuss some of the most relevant cancer-associated carbohydrate structures, including the β1,6-branching of N-linked chains, the sialyl Lewis antigens, the α2,6-sialylated lactosamine, the Thomsen-Friedenreich-related antigens and gangliosides. We will describe the mechanisms leading to the expression of these structures and their interactions with sugar binding molecules, such as selectins and galectins. Finally, we will discuss how the glycosylation machinery of the cell is controlled by signal transduction pathways, epigenetic mechanisms and responds to hypoxia.

Biocompatible noisy nanotopographies with specific directionality for controlled anisotropic cell cultures

All cells are exposed to extra-cellular physical stimuli determined by the details of the micro-/nano-environment within which they exist. These stimuli are present in organs and tissues where specific directional signals coexist with biotopographical noise (e.g. cellular debris, residues of apoptotic cells, protein accumulation, sclerotic plaques). Here, we present a platform for the investigation of the impact of this noise based on nanostructured plastic scaffolds with a controlled level of anisotropy. Two different types of topographical noise are introduced into fully ordered nanostructures. Starting from nanogratings, we randomly introduce nanomodifications, whose density determines the overall substrate directionality. A general quantitative definition of directionality is discussed and applied to our nanostructures. Substrate biocompatibility is assayed by culturing PC12 cells and evaluating cell viability and NGF-induced neuronal-differentiation efficiency. The suitability for high-resolution microscopy on living cells is demonstrated by visualizing focal adhesion complexes by total internal reflection fluorescence (TIRF) microscopy. Finally, we show the impact of noise in modulating focal adhesion maturation in PC12 cells upon NGF-induced neuronal differentiation. Our results indicate design rules both for biocompatible textured substrates allowing the study of cell–environment interaction in vitro and for tissue engineering applications.

The human phosphotyrosine signaling network: Evolution and hotspots of hijacking in cancer

Phosphotyrosine (pTyr) signaling, which plays a central role in cell–cell and cell–environment interactions, has been considered to be an evolutionary innovation in multicellular metazoans. However, neither the emergence nor the evolution of the human pTyr signaling system is currently understood. Tyrosine kinase (TK) circuits, each of which consists of a TK writer, a kinase substrate, and a related reader, such as Src homology (SH) 2 domains and pTyr-binding (PTB) domains, comprise the core machinery of the pTyr signaling network. In this study, we analyzed the evolutionary trajectories of 583 literature-derived and 50,000 computationally predicted human TK circuits in 19 representative eukaryotic species and assigned their evolutionary origins. We found that human TK circuits for intracellular pTyr signaling originated largely from primitive organisms, whereas the inter- or extracellular signaling circuits experienced significant expansion in the bilaterian lineage through the “back-wiring” of newly evolved kinases to primitive substrates and SH2/PTB domains. Conversely, the TK circuits that are involved in tissue-specific signaling evolved mainly in vertebrates by the back-wiring of vertebrate substrates to primitive kinases and SH2/PTB domains. Importantly, we found that cancer signaling preferentially employs the pTyr sites, which are linked to more TK circuits. Our work provides insights into the evolutionary paths of the human pTyr signaling circuits and suggests the use of a network approach for cancer intervention through the targeting of key pTyr sites and their associated signaling hubs in the network.

On the symmetry of siblings: automated single-cell tracking to quantify the behavior of hematopoietic stem cells in a biomimetic setup

The interplay between hematopoietic stem and progenitor cells (HSPC) and their local microenvironment is a key mechanism for the organization of hematopoiesis. To quantitatively study this process, a time-resolved analysis of cellular dynamics at the single-cell level is an essential prerequisite. One way to generate sufficient amounts of appropriate data is automatic single-cell tracking using time-lapse video microscopy. We describe and apply newly developed computational algorithms that allow for an automated generation of high-content data of single-cell characteristics at high temporal and spatial resolution, together with the reconstruction and statistical evaluation of complete genealogical histories. This methodology has been applied to the particular example of purified primary human HSPCs in bioengineered culture conditions. The combination of genealogical information and dynamic profiles of cellular properties identified a marked symmetry between sibling HSPCs regarding cell cycle time, but also migration speed and growth kinetics. Furthermore, we demonstrate that this symmetry of HSPC siblings can be altered by exogenous cues of the local biomimetic microenvironment. Using the example of HSPC growth in biomimetic culture systems, we show that our approach provides a valuable tool for the quantitative analysis of dynamic single-cell features under defined invitro conditions, allowing for integration of functional and genealogical data. The efficiency and accuracy of our approach pave the way for new and intriguing insights into the organizational principles of developmental patterns and the respective influence of exogenous cues not limited to the study of primary HSPCs.

Glycohydrolases β-hexosaminidase and β-galactosidase are associated with lipid microdomains of Jurkat T-lymphocytes

Growing evidence suggests the presence of active lysosomal enzymes in extra-lysosomal compartments, such as the plasma membrane. Although in the past little attention was paid to glycohydrolases acting on cellular compartments different from lysosomes, there is now increasing interest on plasma membrane-associated glycohydrolases because they should be involved, together with glycosyltransferases, in glycosphingolipids oligosaccharide modification processes regulating cell-to-cell and/or cell–environment interactions in both physiological and pathological conditions. Starting from the previous evidence of the presence of β-hexosaminidase and β-galactosidase at the plasma membrane of cultured fibroblasts, we here investigated the association of these glycohydrolases with lipid microdomains of Jurkat T-lymphocytes. Monosialoganglioside GM3 represents the major glycosphingolipid constituent of T-cell plasma membrane and its amount largely increases after T-cell stimulation. β-hexosaminidase and β-galactosidase cleave specific β-linked terminal residues from a wide range of glycoconjugates and in particular are involved in the stepwise degradation of GM1 to GM3 ganglioside. Here we demonstrated that fully processed plasma membrane-associated β-hexosaminidase and β-galactosidase co-distribute with the lipid microdomain markers and co-immunoprecipitate with the signalling protein lck in Jurkat T-cell. Furthermore, Jurkat cell stimulation up-regulates the expression and activity of lysosomal β-hexosaminidase and β-galactosidase and increases their targeting to lipid microdomains. The non-random distribution of plasma membrane-associated β-hexosaminidase and β-galactosidase and their localization within lipid microdomains, suggest a role of these enzymes in the local reorganization of glycosphingolipid-based signalling units.

Differential Adhesion Molecule Expression during Murine Embryonic Stem Cell Commitment to the Hematopoietic and Endothelial Lineages

Mouse embryonic stem cells (ESC) make cell fate decisions based on intrinsic and extrinsic factors. The decision of ESC to differentiate to multiple lineages in vitro occurs during the formation of embryoid bodies (EB) and is influenced by cell-environment interactions. However, molecular mechanisms underlying cell-environmental modulation of ESC fate decisions are incompletely understood. Since adhesion molecules (AM) influence proliferation and differentiation in developing and adult tissues, we hypothesized that specific AM interactions influence ESC commitment toward hematopoietic and endothelial lineages. Expression of AM in the adherens, tight and gap junction pathways in ESC subpopulations were quantified. E-cadherin (E-cad), Claudin-4 (Cldn4), Connexin-43 (Cx43), Zona Occludens-1 (ZO-1) and Zona Occludens-2 (ZO-2) transcript levels were differentially expressed during early stages of hematopoietic/endothelial commitment. Stable ESC lines were generated with reduced expression of E-cad, Cldn4, Cx43, ZO-1 and ZO-2 using shRNA technology. Functional and phenotypic consequences of modulating AM expression were assessed using hematopoietic colony forming assays, endothelial sprouting assays and surface protein expression. A decrease in E-cad, Cldn4, Cx43 and ZO-1 expression was associated with less commitment to the hematopoietic lineage and increased endothelial differentiation as evidenced by functional and phenotypic analysis. A reduction in ZO-2 expression did not influence endothelial differentiation, but decreased hematopoietic commitment two-fold. These data indicate that a subset of AM influence ESC decisions to commit to endothelial and hematopoietic lineages. Furthermore, differentially expressed AM may provide novel markers to delineate early stages of ESC commitment to hematopoietic/endothelial lineages.

Laser Printing of Three-Dimensional Multicellular Arrays for Studies of Cell–Cell and Cell–Environment Interactions

Utilization of living cells for therapies in regenerative medicine requires a fundamental understanding of the interactions between different cells and their environment. Moreover, common models based on adherent two-dimensional cultures are not appropriate to simulate the complex interactions that occur in a three-dimensional (3D) cell-microenvironment in vivo. In this study, we present a computer-aided method for the printing of multiple cell types in a 3D array using laser-assisted bioprinting. By printing spots of human adipose-derived stem cells (ASCs) and endothelial colony-forming cells (ECFCs), we demonstrate that (i) these cell spots can be arranged layer-by-layer in a 3D array; (ii) any cell-cell ratio, cell quantity, cell-type combination, and spot spacing can be realized within this array; and (iii) the height of the 3D array is freely scalable. As a proof of concept, we printed separate spots of ASCs and ECFCs within a 3D array and observed cell-cell interactions in vascular endothelial growth factor-free medium. It has been demonstrated that direct cell-cell contacts trigger the development of stable vascular-like networks. This method can be applied to study complex and dynamic relationships between cells and their local environment.

Abstract 1468: MTA2 enhances breast cancer metastasis via regulation of the cytoskeleton and rho signaling pathways

Abstract Metastasis tumor associated (MTA) 2 is a component of the nucleosome remodeling and histone deacetylation complex and is implicated in regulating gene expression through its associated histone deacetylase activity. We have previously found that MTA2 enhanced the anchorage independent growth phenotype of estrogen receptor alpha (ER)-positive cells, implicating a potential role in metastasis. As MTA2-overexpression in these cells also resulted in an estrogen independent phenotype, we hypothesized that MTA2-overexpression may similarly enhance the aggressiveness of ER-negative cells. We found that MTA2-overexpressing MDA-MB-231 cells were more invasive through Matrigel, had enhanced soft-agar colony formation activity compared with vector controls, and demonstrated increased migration in wound healing assays. Furthermore, we found that MTA2 overexpression resulted in rapid systemic metastases when grown as orthotopic xenografts, while vector control cells failed to form metastases, in the same time period. Microarray analysis revealed enrichment for genes regulating the cytoskeleton and cell-environment interactions, which was validated using a previously published microarray dataset. We found that the negative regulator of the Rho pathway, Rho GDP dissociation inhibitor alpha (RhoGDI-alpha), was also decreased in expression at the protein level in the MTA2-overexpressing MDA-MB-231 cells. We found that blocking signaling through the Rho pathway by either using a Rho Kinase inhibitor or by overexpressing RhoGDI-alpha significantly reduced the soft-agar colony forming ability of MTA2-overexpressing cells. We examined the effect of MTA2 and RhoGDI-alpha expression in a retrospective breast cancer microarray dataset and found that patients classified as MTA2-high and RhoGDI-alpha-low had significantly earlier recurrence compared with all other patients. These data indicate that MTA2-overexpression results in an increase in the metastatic potential of breast cancer cells. These results have potential clinical implications as patients who have the MTA2-high/RhoGDI-alpha-low phenotype are at increased risk for early recurrence. In conclusion, our data show that MTA2 and RhoGDI-alpha enhances breast cancer metastasis possibly by promoting invasiveness and anchorage independent growth and survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1468. doi:10.1158/1538-7445.AM2011-1468

Quantifying the Importance of Galactofuranose in Aspergillus nidulans Hyphal Wall Surface Organization by Atomic Force Microscopy

The fungal wall mediates cell-environment interactions. Galactofuranose (Galf), the five-member ring form of galactose, has a relatively low abundance in Aspergillus walls yet is important for fungal growth and fitness. Aspergillus nidulans strains deleted for Galf biosynthesis enzymes UgeA (UDP-glucose-4-epimerase) and UgmA (UDP-galactopyranose mutase) lacked immunolocalizable Galf, had growth and sporulation defects, and had abnormal wall architecture. We used atomic force microscopy and force spectroscopy to image and quantify cell wall viscoelasticity and surface adhesion of ugeAΔ and ugmAΔ strains. We compared the results for ugeAΔ and ugmAΔ strains with the results for a wild-type strain (AAE1) and the ugeB deletion strain, which has wild-type growth and sporulation. Our results suggest that UgeA and UgmA are important for cell wall surface subunit organization and wall viscoelasticity. The ugeAΔ and ugmAΔ strains had significantly larger surface subunits and lower cell wall viscoelastic moduli than those of AAE1 or ugeBΔ hyphae. Double deletion strains (ugeAΔ ugeBΔ and ugeAΔ ugmAΔ) had more-disorganized surface subunits than single deletion strains. Changes in wall surface structure correlated with changes in its viscoelastic modulus for both fixed and living hyphae. Wild-type walls had the largest viscoelastic modulus, while the walls of the double deletion strains had the smallest. The ugmAΔ strain and particularly the ugeAΔ ugmAΔ double deletion strain were more adhesive to hydrophilic surfaces than the wild type, consistent with changes in wall viscoelasticity and surface organization. We propose that Galf is necessary for full maturation of A. nidulans walls during hyphal extension.

Chapter 17 - Transplantation in Zebrafish

Tissue or cell transplantation has been an extremely valuable technique for studying developmental potential of certain cell population, dissecting cell-environment interaction relationship, identifying stem cells, and many other applications. One key technical requirement for performing transplantation assay is the capability of distinguishing the transplanted donor cells from the endogenous host cells, and tracing the donor cells over time. Zebrafish has emerged as an excellent model organism for performing transplantation assay, thanks to the transparency of embryos during development and even certain adults. Using transgenic techniques and fast-evolving imaging technology, fluorescence-labeled donor cells can be easily identified and studied in vivo. In this chapter, we will first discuss the rationale of different types of zebrafish transplantation in both embryos and adults, and then focus on detailed methods of three types of transplantation: blastula/gastrula transplantation for mosaic analysis, stem cell transplantation, and tumor transplantation.

Cell adhesion, morphology and biochemistry on nano-topographic oxidized silicon surfaces

Manipulating an incorporated scaffold to direct cell behaviors play a key role in tissue engineering. In this study, we developed novel nano-topographic oxidized silicon nanosponges capable of being modified with various chemicals of a few nm in thickness to gain further insight into the fundamental biology of cell-environment interactions in vitro. A wet etching technique was applied to fabricate the silicon nanosponges in a high-throughput manner and was followed by vapor deposition of various organo-silane chemicals to enable self-assembly on the surfaces of the silicon nanosponges. When Chinese hamster ovary cells were cultured on these chemically modified nano-topographic structures, they displayed distinct morphogenesis, adherent responses, and biochemical properties in comparison with those of their planar oxidized silicon counterparts. There were predominant nano-actin punches and slender protrusions formed while cells were cultured on the nano-topographic structures, indicating that cell behaviors can be influenced by the physical characteristic derived from nano-topography, in addition to the hydrophobicity of contact surfaces. This study demonstrates potential applications of these nano-topographic biomaterials for controlling cell development in tissue engineering as well as in basic cell biology research.

Emergent properties of tumor microenvironment in a real-life model of multicell tumor spheroids.

Multicellular tumor spheroids are an important in vitro model of the pre-vascular phase of solid tumors, for sizes well below the diagnostic limit: therefore a biophysical model of spheroids has the ability to shed light on the internal workings and organization of tumors at a critical phase of their development. To this end, we have developed a computer program that integrates the behavior of individual cells and their interactions with other cells and the surrounding environment. It is based on a quantitative description of metabolism, growth, proliferation and death of single tumor cells, and on equations that model biochemical and mechanical cell-cell and cell-environment interactions. The program reproduces existing experimental data on spheroids, and yields unique views of their microenvironment. Simulations show complex internal flows and motions of nutrients, metabolites and cells, that are otherwise unobservable with current experimental techniques, and give novel clues on tumor development and strong hints for future therapies.

Advanced dynamic monitoring of cellular status using label-free and non-invasive cell-based sensing technology for the prediction of anticancer drug efficacy

Quantitative evaluation of anticancer drug efficacy using in vitro cell-based assays is useful for cancer patients, particularly those who show unconventional cancer development. Nevertheless, conventional chemosensitivity testing often requires widely used labeling agents and time-consuming laboratory procedures that provide low reliability. Label-free non-invasive cell-based assays are desired for dynamic monitoring of cellular status. This critical review first describes conventional chemosensitivity testing and then advanced label-free cell-based technology used to screen anticancer drugs through dynamic monitoring of cellular status, focusing on dosage and the use of drug-resistant cancer cells. Results from label-free cell-based approaches are compared with those of conventional chemosensitivity testing. The cellular statuses, addressed in terms of respective mechanisms and disadvantages, are extracellular fluxes of proton (H(+)), O(2), and anticancer drugs, cell morphology changes, cell-environment interaction, and mitochondrial membrane potential. Finally, a cell-based systems outlook is presented. This paper represents a step toward efficient and accurate initial screening of anticancer drugs and development of compounds and their combined use to achieve pharmacodynamic and pharmacokinetic interactions, and chemotherapy evaluation of particular anticancer drugs for individual patients.

Integrative multicellular biological modeling: a case study of 3D epidermal development using GPU algorithms

BackgroundSimulation of sophisticated biological models requires considerable computational power. These models typically integrate together numerous biological phenomena such as spatially-explicit heterogeneous cells, cell-cell interactions, cell-environment interactions and intracellular gene networks. The recent advent of programming for graphical processing units (GPU) opens up the possibility of developing more integrative, detailed and predictive biological models while at the same time decreasing the computational cost to simulate those models.ResultsWe construct a 3D model of epidermal development and provide a set of GPU algorithms that executes significantly faster than sequential central processing unit (CPU) code. We provide a parallel implementation of the subcellular element method for individual cells residing in a lattice-free spatial environment. Each cell in our epidermal model includes an internal gene network, which integrates cellular interaction of Notch signaling together with environmental interaction of basement membrane adhesion, to specify cellular state and behaviors such as growth and division. We take a pedagogical approach to describing how modeling methods are efficiently implemented on the GPU including memory layout of data structures and functional decomposition. We discuss various programmatic issues and provide a set of design guidelines for GPU programming that are instructive to avoid common pitfalls as well as to extract performance from the GPU architecture.ConclusionsWe demonstrate that GPU algorithms represent a significant technological advance for the simulation of complex biological models. We further demonstrate with our epidermal model that the integration of multiple complex modeling methods for heterogeneous multicellular biological processes is both feasible and computationally tractable using this new technology. We hope that the provided algorithms and source code will be a starting point for modelers to develop their own GPU implementations, and encourage others to implement their modeling methods on the GPU and to make that code available to the wider community.

The Complexities of Engineering Human Stem Cell-Derived Therapeutics

The aim of this special issue is to provide the scientific audience with an up-to-date review of pluripotent and tissue-specific stem cells and their differentiation in combination with cellular engineering in the quest to develop novel regenerative therapies. The field of regenerative medicine is focused on the creation of functional tissue units to either repair or replace compromised tissue or organs in vivo. The field therefore offers the promise that, in the future, scientists will be able to grow tissues in the laboratory and use them safely as extracorporeal devices to stimulate endogenous repair [1] or implant them when the body cannot heal itself, but only when cell-based therapies are deemed safe and effective [2, 3]. If successful, such an approach would have a significant impact on the problem of the shortage of donor organs available for transplantation. The collections of papers that make up this special issue provide broad-ranging coverage of the field and provide comprehensive and up-to-date reviews of human development and the relationships that exist between development, regeneration, carcinogenesis, and the role of stem cells in these processes (Kung et al., 2010). There is a focus on application of this knowledge using human embryonic stem cells which have the potential to provide novel biological models and medical devices (Sharma et al., 2010). Additionally, we discuss adult stem cell populations exploring their in vitro expansion (Wells, 2010) and plasticity (Lui et al., 2010), essential to regenerative medicine and tissue engineering. This is supported by the review article on generating the correct cell-cell and cell-environment interactions in these processes (Titushkin et al., 2010). The ability to prepare homogeneous preparations of somatic cells for regenerative medicine will necessitate the development of methods which are simple and do not expose derivative cell populations to greater stress. A highly attractive procedure, dielectrophoresis, shows great potential in population enrichment and in stem cell sorting and is discussed in this special issue (Pethig et al., 2010). It is likely that technologies which do not require cell surface labels will play an increasing role in regenerative medicine. Stem cell-based therapies have been used successfully in the past, and many more are predicted for the future, therefore, it is critical that we produce cell types which are stable and contribute to tissue homeostasis in vivo. A crucial element of tissue homeostasis and organ stability is DNA repair. This process protects stem cells in both embryonic and adult tissues from genetic damage thereby maintaining a stable genome. DNA repair is a fast and efficient process, but it can prove problematic when stem cells undergo malignant transformation (Frosina et al., 2010). In order to gain a better understanding of this process noninvasive cellular techniques have been developed to accurately determine differences in normal and transformed cell lines. Raman spectroscopy is one such example and has provided insight into changes in DNA and RNA concentrations during the lifecycle of a cell, and, as such, we have highlighted this technique as a promising approach in this issue (Downes et al., 2010). We hope that this collection of papers stimulates interest within the academic community and provides a basic and up-to-date overview of key areas in regenerative medicine, cell biology, and tissue engineering.

Physicochemical Control of Adult Stem Cell Differentiation: Shedding Light on Potential Molecular Mechanisms

Realization of the exciting potential for stem-cell-based biomedical and therapeutic applications, including tissue engineering, requires an understanding of the cell-cell and cell-environment interactions. To this end, recent efforts have been focused on the manipulation of adult stem cell differentiation using inductive soluble factors, designing suitable mechanical environments, and applying noninvasive physical forces. Although each of these different approaches has been successfully applied to regulate stem cell differentiation, it would be of great interest and importance to integrate and optimally combine a few or all of the physicochemical differentiation cues to induce synergistic stem cell differentiation. Furthermore, elucidation of molecular mechanisms that mediate the effects of multiple differentiation cues will enable the researcher to better manipulate stem cell behavior and response.