scholarly journals ModelingIn VitroCellular Responses to Silver Nanoparticles

2014 ◽  
Vol 2014 ◽  
pp. 1-13 ◽  
Author(s):  
Dwaipayan Mukherjee ◽  
Steven G. Royce ◽  
Srijata Sarkar ◽  
Andrew Thorley ◽  
Stephan Schwander ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Cell Types ◽  
Engineered Nanoparticles ◽  
Mrna Levels ◽  
Cellular Interactions ◽  
Cellular Mechanisms ◽  
Nanoparticle Transport ◽  
Cell Culture Systems

Engineered nanoparticles (NPs) have been widely demonstrated to induce toxic effects to various cell types.In vitrocell exposure systems have high potential for reliable, high throughput screening of nanoparticle toxicity, allowing focusing on particular pathways while excluding unwanted effects due to other cells or tissue dosimetry. The work presented here involves a detailed biologically based computational model of cellular interactions with NPs; it utilizes measurements performed in human cell culture systemsin vitro, to develop a mechanistic mathematical model that can support analysis and prediction ofin vivoeffects of NPs. The model considers basic cellular mechanisms including proliferation, apoptosis, and production of cytokines in response to NPs. This new model is implemented for macrophages and parameterized usingin vitromeasurements of changes in cellular viability and mRNA levels of cytokines: TNF, IL-1b, IL-6, IL-8, and IL-10. The model includesin vitrocellular dosimetry due to nanoparticle transport and transformation. Furthermore, the model developed here optimizes the essential cellular parameters based onin vitromeasurements, and provides a “stepping stone” for the development of more advancedin vivomodels that will incorporate additional cellular and NP interactions.

Control of dorsoventral pattern in vertebrate neural development: induction and polarizing properties of the floor plate

Development ◽  
1991 ◽  
Vol 113 (Supplement_2) ◽  
pp. 105-122 ◽  
Author(s):  
Marysia Placzek ◽  
Toshiya Yamada ◽  
Marc Tessier-Lavigne ◽  
Thomas Jessell ◽  
Jane Dodd
Keyword(s):  
Neural Tube ◽  
Neural Development ◽  
Cell Types ◽  
Floor Plate ◽  
Neural Cells ◽  
Dorsoventral Patterning ◽  
Cellular Mechanisms ◽  
Cell Position

Distinct classes of neural cells differentiate at specific locations within the embryonic vertebrate nervous system. To define the cellular mechanisms that control the identity and pattern of neural cells we have used a combination of functional assays and antigenic markers to examine the differentiation of cells in the developing spinal cord and hindbrain in vivo and in vitro. Our results suggest that a critical step in the dorsoventral patterning of the embryonic CNS is the differentiation of a specialized group of midline neural cells, termed the floor plate, in response to local inductive signals from the underlying notochord. The floor plate and notochord appear to control the pattern of cell types that appear along the dorsoventral axis of the neural tube. The fate of neuroepithelial cells in the ventral neural tube may be defined by cell position with respect to the ventral midline and controlled by polarizing signals that originate from the floor plate and notochord.

Advances and Challenges in Kidney Organoids

2021 ◽  
Vol 16 ◽  
Author(s):  
Vikram Sabapathy ◽  
Gabrielle Costlow ◽  
Rajkumar Venkatadri ◽  
Murat Dogan ◽  
Sanjay Kumar ◽  
...  
Keyword(s):  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Complex Structures ◽  
Cell Culture Systems ◽  
Epigenetic Signatures ◽  
Kidney Organoids

: The advent of organoids has renewed researcher's interest in in vitro cell culture systems. A wide variety of protocols, primarily utilizing pluripotent stem cells, are under development to improve organoid generation to mimic organ development. The complexity of organoids generated is greatly influenced based on the method used. Understanding the process of kidney organoid formation gives developmental insights into how renal cells form, mature, and interact with the adjacent cells to form specific spatiotemporal structural patterns. This knowledge can bridge the gaps in understanding in vivo renal developmental processes. Evaluating genetic and epigenetic signatures in specialized cell types can help interpret the molecular mechanisms governing cell fate. In addition, development in single-cell RNA sequencing and 3D bioprinting and microfluidic technologies has led to better identification and understanding of a variety of cell types during differentiation and designing of complex structures to mimic the conditions in vivo. While several reviews have highlighted the application of kidney organoids, there is no comprehensive review of various methodologies specifically focusing on the kidney organoids. This review summarizes the updated differentiation methodologies, applications, and challenges associated with kidney organoids. Here we have comprehensively collated all the different variables influencing the organoid generation.

scholarly journals Genetic neuroscience of mammalian learning and memory

2003 ◽  
Vol 358 (1432) ◽  
pp. 787-795 ◽  
Author(s):  
Susumu Tonegawa ◽  
Kazu Nakazawa ◽  
Matthew A. Wilson
Keyword(s):  
Cell Types ◽  
Genetically Engineered ◽  
Mouse Strains ◽  
Cellular Mechanisms ◽  
Genetic Manipulations ◽  
Adult Mice ◽  
Multiple Levels ◽  
Primary Research Interest

Our primary research interest is to understand the molecular and cellular mechanisms on neuronal circuitry underlying the acquisition, consolidation and retrieval of hippocampus-dependent memory in rodents. We study these problems by producing genetically engineered (i.e. spatially targeted and/or temporally restricted) mice and analysing these mice by multifaceted methods including molecular and cellular biology, in vitro and in vivo physiology and behavioural studies. We attempt to identify deficits at each of the multiple levels of complexity in specific brain areas or cell types and deduce those deficits that underlie specific learning or memory. We will review our recent studies on the acquisition, consolidation and recall of memories that have been conducted with mouse strains in which genetic manipulations were targeted to specific types of cells in the hippocampus or forebrain of young adult mice.

scholarly journals CARPOOL: A library-based platform to rapidly identify next generation chimeric antigen receptors

2021 ◽  
Author(s):  
Taeyoon Kyung ◽  
Khloe S Gordon ◽  
Caleb R Perez ◽  
Patrick V Holec ◽  
Azucena Ramos ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Cell Types ◽  
Tumor Control ◽  
Antigen Receptors ◽  
Cell Therapies ◽  
Promising Tool ◽  
Signaling Domain ◽  
Screening Platform

CD19-targeted CAR therapies have successfully treated B cell leukemias and lymphomas, but many responders later relapse or experience toxicities. CAR intracellular domains (ICDs) are key to converting antigen recognition into anti-tumor effector functions. Despite the many possible immune signaling domain combinations that could be included in CARs, almost all CARs currently rely upon CD3𝛇, CD28, and/or 4-1BB signaling. To explore the signaling potential of CAR ICDs, we generated a library of 700,000 CD19 CAR molecules with diverse signaling domains and developed a high throughput screening platform to enable optimization of CAR signaling for anti-tumor functions. Our strategy identifies CARs with novel signaling domain combinations that elicit distinct T cell behaviors from a clinically available CAR, including enhanced proliferation and persistence, lower exhaustion, potent cytotoxicity in an in vitro tumor rechallenge condition, and comparable tumor control in vivo. This approach is readily adaptable to numerous disease models, cell types, and selection conditions, making it a promising tool for rapidly improving adoptive cell therapies and expanding their utility to new disease indications.

scholarly journals Graded regulation of cellular quiescence depth between proliferation and senescence by a lysosomal dimmer switch

2019 ◽  
Vol 116 (45) ◽  
pp. 22624-22634 ◽  
Author(s):  
Kotaro Fujimaki ◽  
Ruoyan Li ◽  
Hengyu Chen ◽  
Kimiko Della Croce ◽  
Hao Helen Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Signature ◽  
Cell Types ◽  
The Body ◽  
Cellular Mechanisms ◽  
Lysosomal Function ◽  
Quiescent Cells ◽  
Common Transition

The reactivation of quiescent cells to proliferate is fundamental to tissue repair and homeostasis in the body. Often referred to as the G0 state, quiescence is, however, not a uniform state but with graded depth. Shallow quiescent cells exhibit a higher tendency to revert to proliferation than deep quiescent cells, while deep quiescent cells are still fully reversible under physiological conditions, distinct from senescent cells. Cellular mechanisms underlying the control of quiescence depth and the connection between quiescence and senescence are poorly characterized, representing a missing link in our understanding of tissue homeostasis and regeneration. Here we measured transcriptome changes as rat embryonic fibroblasts moved from shallow to deep quiescence over time in the absence of growth signals. We found that lysosomal gene expression was significantly up-regulated in deep quiescence, and partially compensated for gradually reduced autophagy flux. Reducing lysosomal function drove cells progressively deeper into quiescence and eventually into a senescence-like irreversibly arrested state; increasing lysosomal function, by lowering oxidative stress, progressively pushed cells into shallower quiescence. That is, lysosomal function modulates graded quiescence depth between proliferation and senescence as a dimmer switch. Finally, we found that a gene-expression signature developed by comparing deep and shallow quiescence in fibroblasts can correctly classify a wide array of senescent and aging cell types in vitro and in vivo, suggesting that while quiescence is generally considered to protect cells from irreversible arrest of senescence, quiescence deepening likely represents a common transition path from cell proliferation to senescence, related to aging.

scholarly journals High Throughput Screening Method for Identification of New Lipofection Reagents

2001 ◽  
Vol 6 (4) ◽  
pp. 245-254 ◽  
Author(s):  
Anne E. Regelin ◽  
Erhard Fernholz ◽  
Harald F. Krug ◽  
Ulrich Massing
Keyword(s):  
High Throughput Screening ◽  
Screening Method ◽  
Genetic Material ◽  
Cell Types ◽  
Cationic Lipids ◽  
Adherent Cell ◽  
Gene Assay

Lipofection, the transfer of genetic material into cells by means of cationic lipids, is of growing interest for in vitro and in vivo approaches. In order to identify ideal lipofection reagents in a HTS, we have developed an automated lipofection method for the transfer of reporter genes into cells and for determination of the lipofection results. The method has specifically been designed and optimized for 96-well microtiter plates and can successfully be carried out by a pipetting robot with accessory equipment. It consists of two separate parts: (1) pretransfection (preparation of liposomes, formation of lipoplexes, and lipoplex transfer to the cells) and (2) posttransfection (determination of the reporter enzyme activity and the protein content of the transfected cells). Individual steps of the lipofection method were specifically optimized—for example, lipoplex formation and incubation time as well as cell lysis, cell cultivating, and the reporter gene assay. The HTS method facilitates characterization of the transfection properties (efficiency and cytotoxicity) of large numbers of (cationic) lipids in various adherent cell types.

scholarly journals Sustained hypoxia leads to the emergence of cells with enhanced growth, migratory, and promitogenic potentials within the distal pulmonary artery wall

2009 ◽  
Vol 297 (6) ◽  
pp. L1059-L1072 ◽  
Author(s):  
Maria G. Frid ◽  
Min Li ◽  
Meena Gnanasekharan ◽  
Danielle L. Burke ◽  
Miguel Fragoso ◽  
...  
Keyword(s):  
Pulmonary Artery ◽  
Mesenchymal Cell ◽  
Cell Types ◽  
Type I ◽  
Mrna Levels ◽  
Functional Characteristics ◽  
Cellular Mechanisms ◽  
Cell Markers ◽  
Type I Procollagen

All forms of chronic pulmonary hypertension (PH) are characterized by structural remodeling of the pulmonary artery (PA) media, a process previously attributed solely to changes in the phenotype of resident smooth muscle cells (SMC). However, recent experimental evidence in both systemic and pulmonary circulations suggests that other cell types, including circulating and local progenitors, contribute significantly to this process. The goal of this study was to determine if hypoxia-induced remodeling of distal PA (dPA) media involves the emergence of cells with phenotypic and functional characteristics distinct from those of resident dPA SMC and fibroblasts. In vivo, in contrast to the phenotypically uniform SMC composition of dPA media in control calves, the remodeled dPA media of neonatal calves with severe hypoxia-induced PH comprised cells exhibiting a distinct phenotype, including the expression of hematopoetic (CD45), leukocytic/monocytic (CD11b, CD14), progenitor (cKit), and motility-associated (S100A4) cell markers. Consistent with these in vivo observations, primary cell cultures isolated from dPA media of hypertensive calves yielded not only differentiated SMC, but also smaller, morphologically rhomboidal (thus termed here “R”) cells that transiently expressed CD11b, constitutively expressed the mesenchymal cell marker type I procollagen, expressed high mRNA levels of progenitor cell markers cKit, CD34, CD73, as well as for inflammatory mediators, IL-6 and MCP-1, and, with time in culture, gained expression of a myofibroblast marker, α-SM-actin. R cells exhibited highly augmented proliferative, migratory, invasive, and potent promitogenic capabilities, which were due, at least in part, to the production of PDGFs, SDF-1/CXCL12, and S100A4. These data suggest that the cellular mechanisms of dPA remodeling include the emergence of cells with phenotypic and functional characteristics markedly distinct from those of resident dPA cells.

scholarly journals Novel use of a chemically modified siRNA for robust and sustainable in vivo gene silencing in the retina

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Takazumi Taniguchi ◽  
Ken-ichi Endo ◽  
Hidetoshi Tanioka ◽  
Masaaki Sasaoka ◽  
Kei Tashiro ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Intravitreal Injection ◽  
Target Genes ◽  
Vitreous Body ◽  
Cell Types ◽  
Retinal Cell ◽  
Mrna Levels ◽  
Chemically Modified

AbstractDespite efficient and specific in vitro knockdown, more reliable and convenient methods for in vivo knockdown of target genes remain to be developed particularly for retinal research. Using commercially available and chemically modified siRNA so-called Accell siRNA, we established a novel in vivo gene silencing approach in the rat retina. siRNA designed for knockdown of the house keeping gene Gapdh or four retinal cell type-specific genes (Nefl, Pvalb, Rho and Opn1sw) was injected into the vitreous body, and their retinal mRNA levels were quantified using real-time PCR. Intravitreal injection of siRNA for Gapdh resulted in approximately 40–70% reduction in its retinal mRNA levels, which lasted throughout a 9-day study period. Furthermore, all the selected retinal specific genes were efficiently down-regulated by 60–90% following intravitreal injection, suggesting injected siRNA penetrated into major retinal cell types. These findings were consistent with uniform distribution of a fluorescence-labeled siRNA injected into the vitreous body. Interestingly, gene silencing of Grin1, a core subunit of NMDA receptor, was accompanied by significant prevention from NMDA-induced retinal ganglion cell death. Thus, we provide single intravitreal injection of Accell siRNA as a versatile technique for robust and sustainable in vivo retinal gene silencing to characterize their biological functions under physiological and pathophysiological conditions.

scholarly journals Under-Oil Autonomously Regulated Oxygen Microenvironments: A Goldilocks Principle-Based Approach For Microscale Cell Culture

2020 ◽  
Author(s):  
Chao Li ◽  
Mouhita Humayun ◽  
Glenn M Walker ◽  
Keon Young Park ◽  
Bryce Connors ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Types ◽  
Bacterial Cells ◽  
Intestinal Epithelial ◽  
Oxygen Levels ◽  
Disease States ◽  
Cell Culture Systems ◽  
Kinetics Of

Oxygen levels in vivo are autonomously regulated by a supply-demand balance, which can be altered in disease states. However, the oxygen levels of in vitro cell culture systems, particularly microscale cell culture, are typically dominated by either supply or demand. Further, the oxygen microenvironment in these systems are rarely monitored or reported. Here, we present a method to establish and dynamically monitor autonomously regulated oxygen microenvironments (AROM) using an oil overlay in an open microscale cell culture system. Using this method, the oxygen microenvironment is dynamically regulated via a supply-demand balance of the system. We simulate the kinetics of oxygen diffusion in multiliquid-phase microsystems on COMSOL Multiphysics and experimentally validate the method using a variety of cell types including mammalian, fungal and bacterial cells. Finally, we demonstrate the utility of this method to establish a co-culture between primary intestinal epithelial cells and a highly prevalent human gut species Bacteroides uniformis.

Cell Patterning: Interaction of Cardiac Myocytes and Fibroblasts in Three-Dimensional Culture

2008 ◽  
Vol 14 (2) ◽  
pp. 117-125 ◽  
Author(s):  
Troy A. Baudino ◽  
Alex McFadden ◽  
Charity Fix ◽  
Joshua Hastings ◽  
Robert Price ◽  
...  
Keyword(s):  
Cardiac Tissue ◽  
Three Dimensional ◽  
Normal Growth ◽  
Cell Types ◽  
Cell Interactions ◽  
Cellular Interactions ◽  
Cell Layers ◽  
Cell Cell

Patterning of cells is critical to the formation and function of the normal organ, and it appears to be dependent upon internal and external signals. Additionally, the formation of most tissues requires the interaction of several cell types. Indeed, both extracellular matrix (ECM) components and cellular components are necessary for three-dimensional (3-D) tissue formationin vitro. Using 3-D cultures we demonstrate that ECM arranged in an aligned fashion is necessary for the rod-shaped phenotype of the myocyte, and once this pattern is established, the myocytes were responsible for the alignment of any subsequent cell layers. This is analogous to thein vivopattern that is observed, where there appears to be minimal ECM signaling, rather formation of multicellular patterns is dependent upon cell–cell interactions. Our 3-D culture of myocytes and fibroblasts is significant in that it modelsin vivoorganization of cardiac tissue and can be used to investigate interactions between fibroblasts and myocytes. Furthermore, we used rotational cultures to examine cellular interactions. Using these systems, we demonstrate that specific connexins and cadherins are critical for cell–cell interactions. The data presented here document the feasibility of using these systems to investigate cellular interactions during normal growth and injury.

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