scholarly journals A Bioengineered Three-Dimensional Cell Culture Platform Integrated with Microfluidics To Address Antimicrobial Resistance in Tuberculosis

mBio ◽  
2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Magdalena K. Bielecka ◽  
Liku B. Tezera ◽  
Robert Zmijan ◽  
Francis Drobniewski ◽  
Xunli Zhang ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Immune Cells ◽  
Three Dimensional ◽  
Host Cells ◽  
Bacterial Survival ◽  
Dimensional Matrix ◽  
Concentration Changes ◽  
Model Drug ◽  
Global Threat ◽  
New Treatments

ABSTRACT Antimicrobial resistance presents one of the most significant threats to human health, with the emergence of totally drug-resistant organisms. We have combined bioengineering, genetically modified bacteria, longitudinal readouts, and fluidics to develop a transformative platform to address the drug development bottleneck, utilizing Mycobacterium tuberculosis as the model organism. We generated microspheres incorporating virulent reporter bacilli, primary human cells, and an extracellular matrix by using bioelectrospray methodology. Granulomas form within the three-dimensional matrix, and mycobacterial stress genes are upregulated. Pyrazinamide, a vital first-line antibiotic for treating human tuberculosis, kills M. tuberculosis in a three-dimensional culture but not in a standard two-dimensional culture or Middlebrook 7H9 broth, demonstrating that antibiotic sensitivity within microspheres reflects conditions in patients. We then performed pharmacokinetic modeling by combining the microsphere system with a microfluidic plate and demonstrated that we can model the effect of dynamic antibiotic concentrations on mycobacterial killing. The microsphere system is highly tractable, permitting variation of cell content, the extracellular matrix, sphere size, the infectious dose, and the surrounding medium with the potential to address a wide array of human infections and the threat of antimicrobial resistance. IMPORTANCE Antimicrobial resistance is a major global threat, and an emerging concept is that infection should be studied in the context of host immune cells. Tuberculosis is a chronic infection that kills over a million people every year and is becoming progressively more resistant to antibiotics. Recent major studies of shorter treatment or new vaccination approaches have not been successful, demonstrating that transformative technologies are required to control tuberculosis. We have developed an entirely new system to study the infection of host cells in a three-dimensional matrix by using bioengineering. We showed that antibiotics that work in patients are effective in this microsphere system but not in standard infection systems. We then combined microspheres with microfluidics to model drug concentration changes in patients and demonstrate the effect of increasing antibiotic concentrations on bacterial survival. This system can be widely applied to address the threat of antimicrobial resistance and develop new treatments. IMPORTANCE Antimicrobial resistance is a major global threat, and an emerging concept is that infection should be studied in the context of host immune cells. Tuberculosis is a chronic infection that kills over a million people every year and is becoming progressively more resistant to antibiotics. Recent major studies of shorter treatment or new vaccination approaches have not been successful, demonstrating that transformative technologies are required to control tuberculosis. We have developed an entirely new system to study the infection of host cells in a three-dimensional matrix by using bioengineering. We showed that antibiotics that work in patients are effective in this microsphere system but not in standard infection systems. We then combined microspheres with microfluidics to model drug concentration changes in patients and demonstrate the effect of increasing antibiotic concentrations on bacterial survival. This system can be widely applied to address the threat of antimicrobial resistance and develop new treatments.

scholarly journals D936Y and Other Mutations in the Fusion Core of the SARS-CoV-2 Spike Protein Heptad Repeat 1: Frequency, Geographical Distribution, and Structural Effect

Molecules ◽  
2021 ◽  
Vol 26 (9) ◽  
pp. 2622
Author(s):  
Romina Oliva ◽  
Abdul Rajjak Shaikh ◽  
Andrea Petta ◽  
Anna Vangone ◽  
Luigi Cavallo
Keyword(s):  
Three Dimensional ◽  
Salt Bridge ◽  
Structural Effect ◽  
Host Cells ◽  
Heptad Repeat ◽  
Genomic Sequences ◽  
Strong Interactions ◽  
Virus Infectivity ◽  
S Protein ◽  
Global Threat

The crown of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is constituted by its spike (S) glycoprotein. S protein mediates the SARS-CoV-2 entry into the host cells. The “fusion core” of the heptad repeat 1 (HR1) on S plays a crucial role in the virus infectivity, as it is part of a key membrane fusion architecture. While SARS-CoV-2 was becoming a global threat, scientists have been accumulating data on the virus at an impressive pace, both in terms of genomic sequences and of three-dimensional structures. On 15 February 2021, from the SARS-CoV-2 genomic sequences in the GISAID resource, we collected 415,673 complete S protein sequences and identified all the mutations occurring in the HR1 fusion core. This is a 21-residue segment, which, in the post-fusion conformation of the protein, gives many strong interactions with the heptad repeat 2, bringing viral and cellular membranes in proximity for fusion. We investigated the frequency and structural effect of novel mutations accumulated over time in such a crucial region for the virus infectivity. Three mutations were quite frequent, occurring in over 0.1% of the total sequences. These were S929T, D936Y, and S949F, all in the N-terminal half of the HR1 fusion core segment and particularly spread in Europe and USA. The most frequent of them, D936Y, was present in 17% of sequences from Finland and 12% of sequences from Sweden. In the post-fusion conformation of the unmutated S protein, D936 is involved in an inter-monomer salt bridge with R1185. We investigated the effect of the D936Y mutation on the pre-fusion and post-fusion state of the protein by using molecular dynamics, showing how it especially affects the latter one.

scholarly journals Nonlinear active control of a cancerous tumour

2018 ◽  
Vol 7 (2.21) ◽  
pp. 72 ◽  
Author(s):  
Bhabani Shankar Dey ◽  
Manas Kumar Bera ◽  
Binoy Krishna Roy
Keyword(s):  
Active Control ◽  
Immune Cells ◽  
Normal Cell ◽  
Tumour Growth ◽  
Three Dimensional ◽  
Tumour Cells ◽  
Host Cells ◽  
Cancer Model ◽  
Control Objective ◽  
Simulation Results

This paper deals with the control of a cancerous tumour growth. The model used is a Three-Dimensional Cancer Model (TDCM). The competition terms include tumour cells, healthy cells, and immune cells. Nature of the competition among the populations of tumour cells, healthy host cells, and immune cells results in a chaotic behaviour. In this paper, a nonlinear active control has been used to control the growth of a tumour. Effect of chemotherapy drug on the different cell populations has been studied. Our control objective is to control the tumour growth and minimize its population to a small value which can be considered as harmless.Along with the above objective, the normal cell population is also be maintained at a particular level. This work has been done completely inin-sillico environment. The simulation results are shown extensively to support the theoretical analysis and confirmed that the preliminary objectives of the paper are attained.  

scholarly journals D936Y and Other Mutations in the Fusion Core of the SARS-Cov-2 Spike Protein Heptad Repeat 1 Undermine the Post-Fusion Assembly

2020 ◽  
Author(s):  
Luigi Cavallo ◽  
Romina Oliva
Keyword(s):  
Three Dimensional ◽  
Point Mutations ◽  
Protein Sequences ◽  
Salt Bridge ◽  
Host Cells ◽  
Heptad Repeat ◽  
Genomic Sequences ◽  
Protein Conformations ◽  
S Protein ◽  
Global Threat

AbstractThe iconic “red crown” of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is made of its spike (S) glycoprotein. The S protein is the Trojan horse of coronaviruses, mediating their entry into the host cells. While SARS-CoV-2 was becoming a global threat, scientists have been accumulating data on the virus at an impressive pace, both in terms of genomic sequences and of three-dimensional structures. On April 21st, the GISAID resource had collected 10,823 SARS-CoV-2 genomic sequences. We extracted from them all the complete S protein sequences and identified point mutations thereof. Six mutations were located on a 14-residue segment (929-943) in the “fusion core” of the heptad repeat 1 (HR1). Our modeling in the pre- and post-fusion S protein conformations revealed, for three of them, the loss of interactions stabilizing the post-fusion assembly. On May 29th, the SARS-CoV-2 genomic sequences in GISAID were 34,805. An analysis of the occurrences of the HR1 mutations in this updated dataset revealed a significant increase for the S929I and S939F mutations and a dramatic increase for the D936Y mutation, which was particularly widespread in Sweden and Wales/England. We notice that this is also the mutation causing the loss of a strong inter-monomer interaction, the D936-R1185 salt bridge, thus clearly weakening the post-fusion assembly.

Recent Review on Subclass B1 Metallo-β-lactamases Inhibitors: Sword for Antimicrobial Resistance

2019 ◽  
Vol 20 (7) ◽  
pp. 756-762 ◽  
Author(s):  
Aditi Kaushik ◽  
Manish Kaushik ◽  
Viney Lather ◽  
J.S. Dua
Keyword(s):  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Antimicrobial Resistance ◽  
Global Health ◽  
Human Health ◽  
Microbial Pathogens ◽  
Present Review ◽  
Drug Molecules ◽  
Global Threat ◽  
Powerful Strategy

An emerging crisis of antibiotic resistance for microbial pathogens is alarming all the nations, posing a global threat to human health. The production of the metallo-β-lactamase enzyme is the most powerful strategy of bacteria to produce resistance. An efficient way to combat this global health threat is the development of broad/non-specific type of metallo-β-lactamase inhibitors, which can inhibit the different isoforms of the enzyme. Till date, there are no clinically active drugs against metallo- β-lactamase. The lack of efficient drug molecules against MBLs carrying bacteria requires continuous research efforts to overcome the problem of multidrug-resistance bacteria. The present review will discuss the clinically potent molecules against different variants of B1 metallo-β-lactamase.

scholarly journals Complement Inactivation Strategy of Staphylococcus aureus Using Decay-Accelerating Factor and the Response of Infected HaCaT Cells

2021 ◽  
Vol 22 (8) ◽  
pp. 4015
Author(s):  
Kyoung Ok Jang ◽  
Youn Woo Lee ◽  
Hangeun Kim ◽  
Dae Kyun Chung
Keyword(s):  
Staphylococcus Aureus ◽  
Immune Cells ◽  
Respiratory Infections ◽  
Food Poisoning ◽  
Surface Expression ◽  
Host Cells ◽  
Hacat Cells ◽  
Decay Accelerating Factor ◽  
Ligand Expression ◽  
The Complement System

Staphylococcus aureus is a species of Gram-positive staphylococcus. It can cause sinusitis, respiratory infections, skin infections, and food poisoning. Recently, it was discovered that S. aureus infects epithelial cells, but the interaction between S. aureus and the host is not well known. In this study, we confirmed S. aureus to be internalized by HaCaT cells using the ESAT-6-like protein EsxB and amplified within the host over time by escaping host immunity. S. aureus increases the expression of decay-accelerating factor (CD55) on the surfaces of host cells, which inhibits the activation of the complement system. This mechanism makes it possible for S. aureus to survive in host cells. S. aureus, sufficiently amplified within the host, is released through the initiation of cell death. On the other hand, the infected host cells increase their surface expression of UL16 binding protein 1 to inform immune cells that they are infected and try to be eliminated. These host defense systems seem to involve the alteration of tight junctions and the induction of ligand expression to activate immune cells. Taken together, our study elucidates a novel aspect of the mechanisms of infection and immune system evasion for S. aureus.

Cell culturing in a three-dimensional matrix affects the localization and properties of plasma membrane cholesterol

2009 ◽  
Vol 33 (10) ◽  
pp. 1079-1086 ◽  
Author(s):  
Nadezhda Stefanova ◽  
Galya Staneva ◽  
Diana Petkova ◽  
Teodora Lupanova ◽  
Roumen Pankov ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Three Dimensional ◽  
Membrane Cholesterol ◽  
Dimensional Matrix ◽  
Cell Culturing ◽  
Plasma Membrane Cholesterol

The Use of Lectins as Tools to Combat SARS-CoV-2

2021 ◽  
Vol 27 ◽  
Author(s):  
Daniela Martinez ◽  
Diego Amaral ◽  
David Markovitz ◽  
Luciano Pinto
Keyword(s):  
Viral Infection ◽  
Host Cells ◽  
Viral Fusion ◽  
Cell Receptors ◽  
First Case ◽  
Mannose Binding ◽  
New Treatments ◽  
Viral Surface

Background: in december 2019, china announced the first case of an infection caused by an, until then, unknown virus: sars-cov-2. since then, researchers have been looking for viable alternatives for the treatment and/or cure of viral infection. among the possible complementary solutions are lectins, and proteins that are reversibly bound to different carbohydrates. the spike protein, present on the viral surface, can interact with different cell receptors: ace2, cd147, and dc-signr. since lectins have an affinity for different carbohydrates, the binding with the glycosylated cell receptors represents a possibility of preventing the virus from binding to the receptors of host cells. Objective: in this review we discuss the main lectins that are possible candidates for use in the treatment of covid-19, highlighting those that have already demonstrated antiviral activity in vivo and in vitro, including mannose-binding lectin, griffithsin, banlec, and others. we also aim to discuss the possible mechanism of action of lectins, which appears to occur through the mediation of viral fusion in host cells, by binding of lectins to glycosylated receptors found in human cells and/or binding of these proteins with the spike glycoprotein, present in virus surface.moreover, we also discuss the use of lectins in clinical practice. Conclusion: Even with the development of effective vaccines, new cases of viral infection with the same virus, or new outbreaks with different viruses can occur; so, the development of new treatments should not be discarded. moreover, the discussions made in this work are relevant regarding the anti-viral properties of lectins.

On Extremal Positive Maps of Three-Dimensional Matrix Algebra

2021 ◽  
Vol 28 (02) ◽  
Author(s):  
Piotr Ługiewicz ◽  
Robert Olkiewicz
Keyword(s):  
Matrix Algebra ◽  
Three Dimensional ◽  
One Dimensional ◽  
Dimensional Matrix ◽  
Positive Maps

A class of bistochastic maps of three-dimensional matrix algebra which preserves a one-dimensional projector is studied.

Abstract 1772: Cardiomyocyte Damage-Released S100A1 Protein Evokes A Proinflammatory Phenotype In Cardiac Fibroblasts

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
David Rohde ◽  
Melanie Boerries ◽  
Herzog Nicole ◽  
Gang Qiu ◽  
Philipp Ehlermann ◽  
...  
Keyword(s):  
Western Blotting ◽  
Immune Cells ◽  
Nuclear Translocation ◽  
Cardiac Fibroblasts ◽  
Innate Immune ◽  
Host Cells ◽  
Boyden Chamber ◽  
Innate Immune Cells ◽  
Danger Signal

Background: S100A1, a cardiomyocyte specific inotropic calcium sensor protein, is released from infarcted human myocardium in the extracellular environment and circulation, reaching peak serum levels (1–2 μM) 8–9 hours after clinical onset. As growing evidence indicates that S100 proteins can act as pre-existing danger signals triggering the innate immune system into action upon release from injured host cells, we hypothesized that damage-released S100A1 can act as a cardiac danger signal alerting innate immune cells. Methods and Results: Here we report for the first time that necrotic cardiomyocytes release S100A1 protein in vitro, which is exclusively internalized by cardiac fibroblasts (CFs) in a clathrin- and caveolin-independent manner as shown by IF. Internalized S100A1 specifically activated MAPKs/SAPKs (p38, ERK1/2 and JNK) resulting in nuclear translocation of p65 (NF-kB) as assessed by Western blotting, EMSA and IF. In turn, S100A1 triggered an inflammatory gene program in CFs including enhanced expression of adhesion molecules, integrins, chemokines and cytokines including I-CAM, V-CAM, CD11b/18, IL1-alpha, MCP-1, TNF-alpha, SDF-1 among others as obtained by RT-PCR, Western blotting and ELISA. This resulted in enhanced chemoattraction and adhesion of monocytotic and stem cells to S100A1-activated CF as shown by Boyden-chamber and adhesion assays. In line with their proinflammatory transition, S100A1-activated CFs exhibited decreased collagen-1/-3 expression and de-novo collagen production, enhanced collagenolytic MMP-9 abundance and activity and increased levels of the antiangiogenic matricellular factor thrombospondin-2 reflecting extracellular matrix net degradation. Importantly, the immun-modulatory and antifibrotic actions of S100A1 protein in vitro were restricted to CFs, RAGE independent and occurred at concentrations (0.1–1 μM) that were found in patients after AMI. Conclusion: Our in vitro results indicate that S100A1 has the properties of a pre-exisiting endogenous cardiomyocyte danger signal transforming cardiac fibroblasts into immunmodulatory cells that might recruit innate immune cells to the site of cardiac injury and link cardiomyocyte damage to post-MI inflammation.

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