Generation of controllable gradients in cell density.
Ontology highlight
ABSTRACT:
SUBMITTER: Liu W
PROVIDER: S-EPMC3535515 | biostudies-literature | 2013 Jan
REPOSITORIES: biostudies-literature
Publications
Generation of controllable gradients in cell density.
Liu Wenying W Zhang Yu Y Thomopoulos Stavros S Xia Younan Y
Angewandte Chemie (International ed. in English) 20120905 1
Making the grad(ient): a gradient in cell density was generated on a substrate that was inserted into a homogeneous suspension of cells at a specific tilt angle by taking advantage of the gradual change in the number of cells available for sedimentation. Reverse gradients were also fabricated on the same substrate using multiple sedimentation procedures. ...[more]
Similar Datasets
Project description:The ability to rapidly generate concentration gradients of diffusible molecules has important applications in many chemical and biological studies. Here we established spatially and temporally controllable concentration gradients of molecules (i.e. proteins or toxins) in a portable microfluidic device in an easy and rapid manner. The formation of the concentration gradients was initiated by a passive-pump-induced forward flow and further optimized during an evaporation-induced backward flow. The centimeter-long gradients along the microfluidic channel were shown to be spatially and temporally controlled by the backward flow. The gradient profile was stabilized by stopping the flow. Computational simulations of this dynamic process illustrated the combined effects of convection and diffusion on the gradient generation, and fit well with the experimental data. To demonstrate the applications of this methodology, a stabilized concentration gradient of a cardiac toxin, alpha-cypermethrin, along the microchannel was used to test the response of HL-1 cardiac cells in the micro-device, which correlated with toxicity data obtained from multi-well plates. The approach presented here may be useful for many biological and chemical processes that require rapid generation of long-range gradients in a portable microfluidic device.
| S-EPMC2790062 | biostudies-other
Project description:Here we report controlled formation of sustained, soluble protein concentration gradients within hydrated polymer networks. The approach involves spatially localizing proteins or biodegradable, protein-loaded microspheres within hydrogels to form a protein-releasing "depot". Soluble protein concentration gradients are then formed as the released protein diffuses away from the localized source. Control over key gradient parameters, including maximum concentration, gradient magnitude, slope, and time dynamics, is achieved by controlling the release of protein from the depot and subsequent transport through the hydrogel. Results demonstrate a direct relationship between the amount of protein released from the depot and the source concentration, gradient magnitude, and slope of the concentration gradient. In addition, an inverse relationship exists between the diffusion coefficient of protein within the hydrogel and the slope of the concentration gradient. The time dynamics of the concentration gradient profile can be directly correlated to protein release from the localized source, providing a mechanism for temporally controlling gradient characteristics. Therefore, each key, biologically relevant parameter associated with the protein concentration gradient can be controlled by defining protein release and diffusion. We anticipate that the resulting materials may be useful in three-dimensional cell culture systems, and in emerging tissue engineering approaches that aim to regenerate complex, functional tissues.
| S-EPMC2900850 | biostudies-literature
Project description:Glioblastoma (GBM) is among the most aggressive cancers. Despite aggressive radiotherapy and treatment with the alkylating agent temozolomide (TMZ), patients ultimately succumb to the disease. Although much interest has focused on highly tumorigenic GBM stem cells (GSCs), adaption of a concept from microbial research proposes that a minor population of dormant “persister” cells in cancer evade current therapies. To separate dormant and treatment-resistant tumor cells in human GBM tumorspheres, we have refined density gradient protocols previously used for separation of neurosphere-forming neural stem cells (NSCs). We find that a minor cell population in human GBM tumorsphere cultures and patient-derived tumor biopsies display increased cell density. These high-density GBM cells (HDGCs) display dormancy, variable expression of proposed GSC markers, and 10-100 fold higher levels of reprogramming gene expression compared to low-density GBM cells (LDGCs). Transcriptional profiling data confirmed the slow-cycling state of HDGCs. As a result, HDGCs show decreased tumorsphere formation capacity in vitro and reduced tumorigenicity in vivo. Using tumorspheres and xenografts, we demonstrated that HDGCs show increased treatment-resistance to ionizing radiation (IR) and temozolomide treatment compared to LDGCs. Similar to the NSC lineage, our data suggest that dormant HDGCs become increasingly sensitive to anti-proliferative therapies as they become activated and further differentiate. In conclusion, density gradients represents a marker-independent approach to separate dormant and treatment-resistant tumor cells in human GBMs and other solid cancers.
2016-07-01 | GSE71116 | GEO
Project description:Glioblastoma (GBM) is among the most aggressive cancers. Despite aggressive radiotherapy and treatment with the alkylating agent temozolomide (TMZ), patients ultimately succumb to the disease. Although much interest has focused on highly tumorigenic GBM stem cells (GSCs), adaption of a concept from microbial research proposes that a minor population of dormant âpersisterâ cells in cancer evade current therapies. To separate dormant and treatment-resistant tumor cells in human GBM tumorspheres, we have refined density gradient protocols previously used for separation of neurosphere-forming neural stem cells (NSCs). We find that a minor cell population in human GBM tumorsphere cultures and patient-derived tumor biopsies display increased cell density. These high-density GBM cells (HDGCs) display dormancy, variable expression of proposed GSC markers, and 10-100 fold higher levels of reprogramming gene expression compared to low-density GBM cells (LDGCs). Transcriptional profiling data confirmed the slow-cycling state of HDGCs. As a result, HDGCs show decreased tumorsphere formation capacity in vitro and reduced tumorigenicity in vivo. Using tumorspheres and xenografts, we demonstrated that HDGCs show increased treatment-resistance to ionizing radiation (IR) and temozolomide treatment compared to LDGCs. Similar to the NSC lineage, our data suggest that dormant HDGCs become increasingly sensitive to anti-proliferative therapies as they become activated and further differentiate. In conclusion, density gradients represents a marker-independent approach to separate dormant and treatment-resistant tumor cells in human GBMs and other solid cancers. 12 samples, no replicates, derived from 5 individual patients
2016-07-01 | E-GEOD-71116 | biostudies-arrayexpress
Project description:Hydrogen sulfide (H2S) plays an important role as an intercellular and intracellular signaling molecule, yet its targets are not well understood. As a molecule it easily evaporates and it is hard to acquire stable concentration for in vitro studies, constituting a major problem for the field to identify its downstream targets and function. Here we develop a microfluidic system that can provide consistent and controllable H2S levels in contrast to the current method of delivering large bolus doses to cells. The system relies on the permeability of H2S gas through a polydimethylsiloxane thin membrane. A hydrogen sulfide donor, sodium hydrosulfide, is perfused in the microchannels below the gas permeable membrane and gaseous H2S diffuses across the membrane, providing a stable concentration for up to 5 hours. Using electrochemical sensors within 3 ppm range, we found that H2S concentration was dependent on two parameters, the concentration of H2S donor, sodium hydrosulfide and the flow rate of the solution in the microchannels. Additionally, different H2S concentration profiles can be obtained by alternating the flow rate, providing an easy means to control the H2S concentration. Our approach constitutes a unique method for H2S delivery for in vitro and ex vivo studies and is ideally suited to identify novel biological processes and cellular mechanisms regulated by H2S.
| S-EPMC6168208 | biostudies-literature
Project description:Natural materials exhibit anisotropy with variations in soluble factors, cell distribution, and matrix properties. The ability to recreate the heterogeneity of the natural materials is a major challenge for investigating cell-material interactions and for developing biomimetic materials. Here we present a generic fluidic approach using convection and alternating flow to rapidly generate multi-centimeter gradients of biomolecules, polymers, beads and cells and cross-gradients of two species in a microchannel. Accompanying theoretical estimates and simulations of gradient growth provide design criteria over a range of material properties. A poly(ethylene-glycol) hydrogel gradient, a porous collagen gradient and a composite material with a hyaluronic acid/gelatin cross-gradient were generated with continuous variations in material properties and in their ability to regulate cellular response. This simple yet generic fluidic platform should prove useful for creating anisotropic biomimetic materials and high-throughput platforms for investigating cell-microenvironment interactions.
| S-EPMC2813888 | biostudies-literature
Project description:To study biological processes in vitro, biomaterials-based engineering solutions to reproduce the gradients observed in tissues are necessary. We present a platform for the 3D bioprinting of functionally graded biomaterials based on carboxylated agarose, a bioink amendable by extrusion bioprinting. Using this bioink, objects with a gradient of stiffness and gradient of cell concentration were printed. Functionalization of carboxylated agarose with maleimide moieties that react in minutes with a cysteine-terminated cell-adhesion peptide allowed us to print objects with a gradient of an immobilized peptide. This approach paves the way toward the development of tissue mimics with gradients.
| S-EPMC8207502 | biostudies-literature
Project description:1. Dried preparations of cell walls from perennial-ryegrass (Lolium perenne) and Italian-ryegrass (L. multiflorum) leaves were suspended in mixtures of carbon tetrachloride with light-petroleum (b.p. 45--50 degrees C) or alcohols and layered on density gradients formed from the same solvents. 2. On centrifugation, the cell walls become distributed throughout a suitably chosen gradient. Fractions corresponding to various regions of the gradient were separated, examined under the microscope and analysed. 3. Cell-wall preparations made from leaf material ground in liquid N2, or in a triple roll mill, showed considerable heterogeneity in particle size, and their behaviour in the density gradient was variable, although there was a general indication that walls derived from vascular bundles were less dense than those from sclerenchyma. 4 Treatment in a vibratory ball mill decreased the size of the particles and produced a more uniform material, but made it impossible to distinguish the origins of the particles. This material behaved more reproducibly in the density gradient. 5. Some fractionations were also made by successive centrifugation in media of increasing relative density. 6. Analyses of the fractions obtained by each method indicated that the less dense had a greater proportion of xylose in the polysaccharide components, and higher contents of acetyl groups and lignin, confirming the close relationship between these components in plant cell walls. 7. The results show that there are differences in polysaccharide composition between the cell-wall types in the grass leaf, the vascular tissue being richer in hemicellulose relative to cellulose than the sclerenchyma.
| S-EPMC1162665 | biostudies-other
Project description:Morphogen gradients are known to subdivide a naive cell field into distinct zones of gene expression. Here, we examine whether morphogens can also induce a graded response within such domains. To this end, we explore the role of the Dorsal protein nuclear gradient along the dorsoventral axis in defining the graded pattern of actomyosin constriction that initiates gastrulation in early Drosophila embryos. Two complementary mechanisms for graded accumulation of mRNAs of crucial zygotic Dorsal target genes were identified. First, activation of target-gene expression expands over time from the ventral-most region of high nuclear Dorsal to lateral regions, where the levels are lower, as a result of a Dorsal-dependent activation probability of transcription sites. Thus, sites that are activated earlier will exhibit more mRNA accumulation. Second, once the sites are activated, the rate of RNA Polymerase II loading is also dependent on Dorsal levels. Morphological restrictions require that translation of the graded mRNA be delayed until completion of embryonic cell formation. Such timing is achieved by large introns, which provide a delay in production of the mature mRNAs. Spatio-temporal regulation of key zygotic genes therefore shapes the pattern of gastrulation.
| S-EPMC8722393 | biostudies-literature
Project description:In this paper, we develop a microfluidic device capable of generating nitric oxide (NO) gradients for cell culture using spatially controlled chemical reactions. NO plays an essential role in various biological activities, including nervous, immune, and cardiovascular systems. The device developed in this paper can control NO gradients without utilizing expensive and hazardous high purity NO gas sources or direct addition of NO donors. Consequently, the device provides an efficient, cost-effective, robust, and stable platform to generate NO gradients for cell culture studies. In the experiments, NO gradients are first characterized using a NO-sensitive fluorescence dye, and cell experiments using aortic smooth muscle cells are conducted. The results demonstrate that the device can alter the intracellular NO concentrations and further affect the Ca(2+) concentration oscillation for the cells. The device developed in this paper provides a powerful platform for researchers better study the biological roles of NO and its spatial distribution using in vitro cell models with minimal instrumentation.
| S-EPMC3838423 | biostudies-literature