Exponential Concentration Gradients in Microfluidic Devices for Cell Studies

2011 ◽  
Author(s):  
Šeila Selimović ◽  
Woo Young Sim ◽  
Sang Bok Kim ◽  
Yun Ho Jang ◽  
Won Gu Lee ◽  
...  
Keyword(s):  
Protein Crystallization ◽  
Experimental Studies ◽  
Cellular Responses ◽  
Analyte Concentration ◽  
Concentration Gradients ◽  
Cell Microenvironment ◽  
Biological Responses ◽  
Native Cell ◽  
High Throughput Assay ◽  
Concentration Levels

Microscale technologies are a powerful tool in many biological and chemical applications, as they utilize only small reagent volumes. Microfluidics is especially well compatible with biological materials and applications, for example protein crystallization, high throughput assay analysis, and various cell studies. In that context, non-linear gradients of particles and molecules as well as efficient mixing of the components inside the lab-on-a-chip are crucial for many experimental studies: testing of and analyzing biological responses to different analyte concentration levels, studying the native cell microenvironment or cellular responses during different growth and proliferation stages. Thus, a microfluidic approach that allows for generation of different concentration gradients and specifically exponential gradients emerges as a helpful technology, and is also compatible with cells.

scholarly journals GSH Modification as a Marker for Plasma Source and Biological Response Comparison to Plasma Treatment

2020 ◽  
Vol 10 (6) ◽  
pp. 2025 ◽  
Author(s):  
Pietro Ranieri ◽  
Hager Mohamed ◽  
Brayden Myers ◽  
Leah Dobossy ◽  
Keely Beyries ◽  
...  
Keyword(s):  
Treatment Time ◽  
Surface Display ◽  
Chemical Species ◽  
Biological Response ◽  
Cell Surface Display ◽  
Cellular Responses ◽  
Mitochondrial Activity ◽  
Plasma Parameters ◽  
Plasma Sources ◽  
Biological Responses

This study investigated the use of glutathione as a marker to establish a correlation between plasma parameters and the resultant liquid chemistry from two distinct sources to predefined biological outcomes. Two different plasma sources were operated at parameters that resulted in similar biological responses: cell viability, mitochondrial activity, and the cell surface display of calreticulin. Specific glutathione modifications appeared to be associated with biological responses elicited by plasma. These modifications were more pronounced with increased treatment time for the European Cooperation in Science and Technology Reference Microplasma Jet (COST-Jet) and increased frequency for the dielectric barrier discharge and were correlated with more potent biological responses. No correlations were found when cells or glutathione were exposed to exogenously added long-lived species alone. This implied that short-lived species and other plasma components were required for the induction of cellular responses, as well as glutathione modifications. These results showed that comparisons of medical plasma sources could not rely on measurements of long-lived chemical species; rather, modifications of biomolecules (such as glutathione) might be better predictors of cellular responses to plasma exposure.

scholarly journals Be Cautious with Crystal Structures of Membrane Proteins or Complexes Prepared in Detergents

Crystals ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 86 ◽  
Author(s):  
Youzhong Guo
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Crystal Structures ◽  
Structure Determination ◽  
Protein Crystallization ◽  
Energy Coupling ◽  
Careful Examination ◽  
Cubic Phase ◽  
Native Cell ◽  
Membrane Protein Crystallization

Membrane proteins are an important class of macromolecules found in all living organisms and many of them serve as important drug targets. In order to understand their biological and biochemical functions and to exploit them for structure-based drug design, high-resolution and accurate structures of membrane proteins are needed, but are still rarely available, e.g., predominantly from X-ray crystallography, and more recently from single particle cryo-EM — an increasingly powerful tool for membrane protein structure determination. However, while protein-lipid interactions play crucial roles for the structural and functional integrity of membrane proteins, for historical reasons and due to technological limitations, until recently, the primary method for membrane protein crystallization has relied on detergents. Bicelle and lipid cubic phase (LCP) methods have also been used for membrane protein crystallization, but the first step requires detergent extraction of the protein from its native cell membrane. The resulting, crystal structures have been occasionally questioned, but such concerns were generally dismissed as accidents or ignored. However, even a hint of controversy indicates that methodological drawbacks in such structural research may exist. In the absence of caution, structures determined using these methods are often assumed to be correct, which has led to surprising hypotheses for their mechanisms of action. In this communication, several examples of structural studies on membrane proteins or complexes will be discussed: Resistance-Nodulation-Division (RND) family transporters, microbial rhodopsins, Tryptophan-rich Sensory Proteins (TSPO), and Energy-Coupling Factor (ECF) type ABC transporters. These analyses should focus the attention of membrane protein structural biologists on the potential problems in structure determination relying on detergent-based methods. Furthermore, careful examination of membrane proteins in their native cell environments by biochemical and biophysical techniques is warranted, and completely detergent-free systems for membrane protein research are crucially needed.

VEGF gradients, receptor activation, and sprout guidance in resting and exercising skeletal muscle

2007 ◽  
Vol 102 (2) ◽  
pp. 722-734 ◽  
Author(s):  
Feilim Mac Gabhann ◽  
James W. Ji ◽  
Aleksander S. Popel
Keyword(s):  
Skeletal Muscle ◽  
Blood Vessels ◽  
Muscle Fibers ◽  
Experimental Studies ◽  
Receptor Activation ◽  
Basement Membranes ◽  
Interstitial Space ◽  
Vegf Receptor ◽  
Interstitial Flow ◽  
Concentration Gradients

Extensive experimental studies have identified vascular endothelial growth factor (VEGF) concentrations and concentration gradients as major factors in angiogenesis; however, localized in vivo measurements of these parameters have not been possible. We developed a three-dimensional computational model of skeletal muscle fibers, blood vessels, and interstitial space. Here it is applied to rat extensor digitorum longus. VEGF isoforms are secreted by myocytes, diffuse through extracellular matrix and basement membranes, and bind endothelial cell surface receptors on blood vessels. In addition, one isoform, VEGF164, binds to proteoglycans in the interstitial space. VEGF secretion rate is determined from the predicted tissue oxygen level through its effect on the hypoxia inducible factor-1α transcription factor. We estimate VEGF secretion and its concentrations and gradients in resting muscle and for different levels of exercise. The effects of low levels of inspired oxygen are also studied. We predict that the high spatial heterogeneity of muscle fiber VEGF secretion in hypoxic tissue leads to significant gradients of VEGF concentration and VEGF receptor activation. VEGF concentration gradients are predicted to be significant in both resting and exercising muscle (4% and 6–8% change in VEGF over 10 μm, respectively), sufficient for chemotactic guidance of 50-μm-long sprout tip cells. VEGF gradients also result in heterogeneity in VEGF receptor activation—a possible explanation for the stochasticity of sprout location. In the absence of interstitial flow, gradients are 10-fold steeper in the transverse direction (i.e., perpendicular to the muscle fibers) than in the longitudinal direction. This may explain observed perpendicular anastomoses in skeletal muscle.

Immune defense and biological responses induced by toxics in Annelida

2001 ◽  
Vol 79 (2) ◽  
pp. 233-253 ◽  
Author(s):  
André Dhainaut ◽  
Patrick Scaps
Keyword(s):  
Short Term Memory ◽  
Experimental Studies ◽  
Immune Defense ◽  
Short Term ◽  
Closely Related Species ◽  
Biological Responses ◽  
Activity Of Enzymes ◽  
Cellular Immune ◽  
Physical And Chemical ◽  
Induced Proteins

The phylum Annelida comprises primitive coelomates that possess specially developed cellular immunity against pathogens. Active phagocytosis by coelomocytes occurs in the struggle against bacteria in Polychaeta and Oligochaeta. Encapsulation plays an important role in defense against parasites, and experimental studies have demonstrated that cooperation between different coelomocyte populations occurs in this process. Spontaneous cytotoxicity of coelomocytes against xenogenic or allogenic cells is analogous with that of vertebrate natural killer cells. Graft rejection is a model for studying the activity of these cells. Accelerated rejection following multiple transplantation reveals that the cellular immune defense system has a short-term memory. In humoral immunity, agglutinins aggregate foreign material and their level is enhanced by antigens; in Annelida, however, no specificity analogous to vertebrate antibodies has been revealed, except for weak specificity of some antigen-binding proteins. Hemolytic substances have been detected, particularly in Oligochaeta, where a fetidin possesses bactericidal activity. Lysozyme and some antibacterial proteins also occur in Polychaeta. Annelida react to physical and chemical insults by various processes. These responses are mainly due to synthesis of stress-induced proteins, inhibition of enzyme activity, and modulation (inhibition or stimulation) of the activity of enzymes involved in the detoxification of xenobiotics. Moreover, these responses frequently differ from those of vertebrates, particularly in terms of the nature of inducers. In other respects, these responses are extremely variable in Annelida, even in closely related species.

scholarly journals Interactions of nanoparticles with pulmonary structures and cellular responses

2008 ◽  
Vol 294 (5) ◽  
pp. L817-L829 ◽  
Author(s):  
Christian Mühlfeld ◽  
Barbara Rothen-Rutishauser ◽  
Fabian Blank ◽  
Dimitri Vanhecke ◽  
Matthias Ochs ◽  
...  
Keyword(s):  
Inhalation Exposure ◽  
Experimental Studies ◽  
Cellular Responses ◽  
Epidemiological Studies ◽  
Oxygen Species ◽  
Adverse Health Effects ◽  
Cardiovascular Morbidity And Mortality ◽  
Dose Metrics ◽  
Methodological Problems ◽  
Shed Light

Combustion-derived and synthetic nano-sized particles (NSP) have gained considerable interest among pulmonary researchers and clinicians for two main reasons. 1) Inhalation exposure to combustion-derived NSP was associated with increased pulmonary and cardiovascular morbidity and mortality as suggested by epidemiological studies. Experimental evidence has provided a mechanistic picture of the adverse health effects associated with inhalation of combustion-derived and synthetic NSP. 2) The toxicological potential of NSP contrasts with the potential application of synthetic NSP in technological as well as medicinal settings, with the latter including the use of NSP as diagnostics or therapeutics. To shed light on this paradox, this article aims to highlight recent findings about the interaction of inhaled NSP with the structures of the respiratory tract including surfactant, alveolar macrophages, and epithelial cells. Cellular responses to NSP exposure include the generation of reactive oxygen species and the induction of an inflammatory response. Furthermore, this review places special emphasis on methodological differences between experimental studies and the caveats associated with the dose metrics and points out ways to overcome inherent methodological problems.

scholarly journals Tumor Hypoxia as a Barrier in Cancer Therapy: Why Levels Matter

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 499
Author(s):  
Tord Hompland ◽  
Christina Sæten Fjeldbo ◽  
Heidi Lyng
Keyword(s):  
Cancer Cells ◽  
Current Knowledge ◽  
Tumor Hypoxia ◽  
Treatment Resistance ◽  
Cellular Responses ◽  
Model Systems ◽  
Major Barrier ◽  
Biological Responses ◽  
Advanced Tumor ◽  
The Individual

Hypoxia arises in tumor regions with insufficient oxygen supply and is a major barrier in cancer treatment. The distribution of hypoxia levels is highly heterogeneous, ranging from mild, almost non-hypoxic, to severe and anoxic levels. The individual hypoxia levels induce a variety of biological responses that impair the treatment effect. A stronger focus on hypoxia levels rather than the absence or presence of hypoxia in our investigations will help development of improved strategies to treat patients with hypoxic tumors. Current knowledge on how hypoxia levels are sensed by cancer cells and mediate cellular responses that promote treatment resistance is comprehensive. Recently, it has become evident that hypoxia also has an important, more unexplored role in the interaction between cancer cells, stroma and immune cells, influencing the composition and structure of the tumor microenvironment. Establishment of how such processes depend on the hypoxia level requires more advanced tumor models and methodology. In this review, we describe promising model systems and tools for investigations of hypoxia levels in tumors. We further present current knowledge and emerging research on cellular responses to individual levels, and discuss their impact in novel therapeutic approaches to overcome the hypoxia barrier.

scholarly journals Construction and Analysis of Coexpression Network to Understand Biological Responses in Chickens Infected by Eimeria tenella

2021 ◽  
Vol 8 ◽  
Author(s):  
Baohong Liu ◽  
Xueting Ma ◽  
Jianping Cai
Keyword(s):  
Cellular Responses ◽  
Eimeria Tenella ◽  
System Level ◽  
Coexpression Network ◽  
Rna Transport ◽  
Eimeria Maxima ◽  
Biological Responses ◽  
High Degree ◽  
The Relationship ◽  
Coexpression Network Analysis

Coccidiosis, caused by various Eimeria species, is a major parasitic disease in chickens. Our understanding of how chickens respond to coccidian infections is highly limited at both the molecular and cellular levels. In this study, coexpression modules were identified by weighted gene coexpression network analysis in chickens infected with Eimeria tenella. A total of 15 correlation modules were identified using 5,175 genes with 24 chicken samples, 12 with primary and 12 with secondary E. tenella infection. The analysis of the interactions between these modules showed a high degree of scale independence. Gene Ontology and Kyoto Encyclopedia of Gene and Genomes enrichment analyses revealed that genes in these functional modules were involved in a broad categories of functions, such as immune response, amino acid metabolism, cellular responses to lipids, sterol biosynthetic processes, and RNA transport. Two modules viz yellow and magenta were identified significantly associating with infection status. Preservation analysis showed that most of the modules identified in E. tenella infections were highly or moderately preserved in chickens infected with either Eimeria acervulina or Eimeria maxima. These analyses outline a biological responses landscape for chickens infected by E. tenella, and also indicates that infections with these three Eimeria species elicit similar biological responses in chickens at the system level. These findings provide new clues and ideas for investigating the relationship between parasites and host, and the control of parasitic diseases.

scholarly journals Chimeric human opsins as optogenetic light sensitisers

2021 ◽  
Author(s):  
Doron G Hickey ◽  
Wayne I L Davies ◽  
Steven Hughes ◽  
Jessica Rodgers ◽  
Navamayooran Thavanesan ◽  
...  
Keyword(s):  
Coupling Efficiency ◽  
Basic Research ◽  
Cell Types ◽  
Cellular Responses ◽  
Wild Type ◽  
Long Wavelength ◽  
Native Cell ◽  
Pathway Activation ◽  
Intracellular Domains ◽  
Phototransduction Cascade

Human opsin-based photopigments have great potential as light-sensitisers, but their requirement for phototransduction cascade-specific second messenger proteins may restrict their functionality in non-native cell types. In this study, eight chimeric human opsins were generated consisting of a backbone of either a rhodopsin (RHO) or long-wavelength-sensitive (LWS) opsin and intracellular domains from Gq/11-coupled human melanopsin. Rhodopsin/melanopsin chimeric opsins coupled to both Gi and Gq/11 pathways. Greater substitution of the intracellular surface with corresponding melanopsin domains generally showed greater Gq/11 activity with a decrease in Gi activation. Unlike melanopsin, rhodopsin and rhodopsin/melanopsin chimeras were dependent upon exogenous chromophore to function. By contrast, wild type LWS opsin and LWS/melanopsin chimeras showed only weak Gi activation in response to light, whilst Gq/11 pathway activation was not detected. Immunocytochemistry demonstrated that chimeric opsins with more intracellular domains of melanopsin were less likely to be trafficked to the plasma membrane. This study demonstrates the importance of Gα coupling efficiency to the speed of cellular responses and created human opsins with a unique combination of properties to expand the range of customised optogenetic biotools for basic research and translational therapies.

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