scholarly journals Long-lasting and responsive DNA/enzyme-based programs in serum-supplemented extracellular media

2021 ◽  
Author(s):  
Jean-Christophe Galas ◽  
Andre Estevez-Torres ◽  
Marc Van Der Hofstadt
Keyword(s):  
Chemical Reactivity ◽  
In Vitro Models ◽  
Extracellular Dna ◽  
In Vitro Assays ◽  
Division Rate ◽  
Cellular Division ◽  
Biomolecular Sensing ◽  
Letter Code

DNA molecular programs are emerging as promising pharmaceutical approaches due to their versatility for biomolecular sensing and actuation. However, the implementation of DNA programs has been mainly limited to serum-deprived in vitro assays due to the fast deterioration of the DNA reaction networks by the nucleases present in the serum. Here, we show that DNA/enzyme programs are functional in serum for 24h but are latter disrupted by nucleases that give rise to parasitic amplification. To overcome this, we implement 3-letter code networks that suppress autocatalytic parasites while still conserving the functionality of DNA/enzyme programs for at least 3 days in the presence of 10% serum. In addition, we define a new buffer that further increases the biocompatibility and conserves responsiveness to changes in molecular composition across time. Finally, we demonstrate how serum-supplemented extracellular DNA molecular programs remain responsive to molecular inputs in the presence of living cells, having responses 6-fold faster than cellular division rate and are sustainable for at least 3 cellular divisions. This demonstrates the possibility of implementing in situ biomolecular characterization tools for serum-demanding in vitro models. We foresee that the coupling of chemical reactivity to our DNA programs by aptamers or oligonucleotide conjugations will allow the implementation of extracellular synthetic biology tools, which will offer new biomolecular pharmaceutical approaches and the emergence of complex and autonomous in vitro models.

In vitro degradability of feed proteins in the rumen: use of non-rumen proteases

2005 ◽  
Vol 56 (8) ◽  
pp. 797 ◽  
Author(s):  
M. Aslam Mirza ◽  
E. L. Miller
Keyword(s):  
Correlation Coefficient ◽  
Soybean Meal ◽  
Streptomyces Griseus ◽  
Correlation Coefficients ◽  
In Vitro Assays ◽  
Vitro Assay ◽  
Bacterial Protease ◽  
Ruminal Protein Degradability

Various feed proteins were incubated independently with bacterial protease from Streptomyces griseus (SGP), papain (Corica papaya), and ficin (Ficus glabrata) in a simple laboratory assay to predict ruminal protein degradability. The estimates obtained from in vitro assays were compared with those obtained from an in situ analysis using synthetic fibre bags. The rate and extent of degradation in vitro using proteases from non-rumen sources differed among substrates used. A high correlation coefficient (r2 = 0.99) was observed between N-degradability from the in vitro method using SGP and in situ estimates when soybean meal was the substrate. Soybean meal nitrogen (N) was almost completely hydrolysed (0.99) in vitro. The correlation coefficients were low and variable with assays using other enzymes. The correlation coefficient was also high (r2 = 0.77–0.84) with in vitro methods using either SGP, papain, or ficin when incubated with fish meal. The N disappearance from barley in vitro was slow to moderate. The ‘b’ estimate of barley obtained with the in vitro assay was significantly (P < 0.01) lower than that observed in situ. Slower proteolysis observed in barley may possibly be linked to poor accessibility of structural proteins rather than the degradability of N per se. None of the enzymes could rank barley in the same order as the in situ method.

Transport Properties of Puerarin and Effect of Extract of Radix Angelicae dahuricae on Puerarin Intestinal Absorption Using In Situ and In Vitro Models

2014 ◽  
Vol 28 (9) ◽  
pp. 1288-1294 ◽  
Author(s):  
Zheng-Gen Liao ◽  
Xin-Li Liang ◽  
Jing-Yun Zhu ◽  
Guo-Wei Zhao ◽  
Yong-Mei Guan ◽  
...  
Keyword(s):  
Transport Properties ◽  
Intestinal Absorption ◽  
In Vitro Models

scholarly journals Airway-On-A-Chip: Designs and Applications for Lung Repair and Disease

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1602
Author(s):  
Tanya J. Bennet ◽  
Avineet Randhawa ◽  
Jessica Hua ◽  
Karen C. Cheung
Keyword(s):  
In Vitro Models ◽  
Chronic Obstructive ◽  
Obstructive Pulmonary Disease ◽  
Characterization Methods ◽  
Composition And Structure ◽  
Challenges And Opportunities ◽  
On Chip ◽  
Respiratory Conditions

The lungs are affected by illnesses including asthma, chronic obstructive pulmonary disease, and infections such as influenza and SARS-CoV-2. Physiologically relevant models for respiratory conditions will be essential for new drug development. The composition and structure of the lung extracellular matrix (ECM) plays a major role in the function of the lung tissue and cells. Lung-on-chip models have been developed to address some of the limitations of current two-dimensional in vitro models. In this review, we describe various ECM substitutes utilized for modeling the respiratory system. We explore the application of lung-on-chip models to the study of cigarette smoke and electronic cigarette vapor. We discuss the challenges and opportunities related to model characterization with an emphasis on in situ characterization methods, both established and emerging. We discuss how further advancements in the field, through the incorporation of interstitial cells and ECM, have the potential to provide an effective tool for interrogating lung biology and disease, especially the mechanisms that involve the interstitial elements.

scholarly journals Regional Evaluation of Fungal Pathogen Incidence in Colombian Cocoa Crops

Agriculture ◽  
2019 ◽  
Vol 9 (3) ◽  
pp. 44
Author(s):  
Raquel Villamizar-Gallardo ◽  
Johann Osma ◽  
Oscar Ortíz-Rodriguez
Keyword(s):  
Fungal Pathogens ◽  
Morphological Variability ◽  
Phytopathogenic Fungi ◽  
Phytophthora Palmivora ◽  
In Vitro Assays ◽  
Post Conflict ◽  
Solid Media ◽  
Black Pod

The production of cocoa (Theobroma cacao L.) in Colombia has a significant environmental and socioeconomic importance as a promissory crop in the post-conflict process. The department of Norte de Santander has cocoa crops that are dramatically affected by fungal pathogens causing important losses during harvest and post-harvest. Therefore, the current study focused on the determination of the incidence of diseases caused by phytopathogenic fungi in cocoa crops, and the identification of primary phytopathogenic fungi found in biological material from different farms of the region. The study was conducted in four municipalities of the department, by sampling fruits infected with frosty pod rot (FPR) and black pod rot (BPR) that presented in situ incidence ranging from 0.37 to 21.58% and from 1.75 to 35.59%, respectively. The studied hybrid materials, together with clone TSH 65, were found to be the most susceptible, while the remaining clones were more tolerant, especially CCN 51, IMC 67, and ICS95. Fifteen strains were isolated using in vitro assays and then morphologically characterized both in solid media and by microscopy. Nine of them corresponded to the pathogen Moniliophthora roreri, and other six to Phytophthora palmivora. The isolated agents showed in vitro morphological variability, as well as the ability to adapt to different environments when growing in situ.

In Vitro Models That Support Adhesion Specificity in Biofilms of Oral Bacteria

1997 ◽  
Vol 11 (1) ◽  
pp. 33-42 ◽  
Author(s):  
R.P. Ellen ◽  
G. Lépine ◽  
P.-M. Nghiem
Keyword(s):  
Matrix Protein ◽  
Protein S ◽  
Specific Protein ◽  
In Vitro Models ◽  
Oral Bacteria ◽  
Host Cells ◽  
In Vitro Assays ◽  
Covalent Bonds ◽  
Enzyme Substrate Recognition

Adhesion to adsorbed pellicles and interspecies co-adhesion to form plaque biofilms involve selective interactions of bacterial adhesins with specific receptors. Our laboratory has devised in vitro assays for co-adhesion between Actinomyces naeslundii and Streptococcus oralis or Porphyromonas gingivalis on saliva-coated mineral and hexadecane droplet substrata. P. gingivalis structures significant for co-adhesion with A. naeslundii include surface vesicles and fimbriae. A family of arginine-specific cysteine proteinases in vesicles may be involved in adherence to bacteria, to host cells, and to matrix proteins. New research from several laboratories has found that such proteinases are processed from genes encoding polyproteins containing both proteinase and hemagglutinin domains. In addition to enzyme-substrate recognition, bacterial adhesion is often determined by specific protein-peptide and lectin-carbohydrate recognition. A. naeslundii - salivary proline-rich protein, S. gordonii - salivary a-amylase, and Treponema denticola - matrix protein recognition are examples of the former. Co-adhesion of A. naeslundii and S. oralis is an example of the latter. Lactose can selectively desorb A. naeslundii cells from mixed biofilms with S. oralis, a demonstration of the significance of specificity. Although non-specific forces are probably secondary to stereochemical fit in determining the selective range of surfaces that bacteria have evolved to recognize and bind, they probably help stabilize non-covalent bonds within aligned, complementary domains.

scholarly journals Natural Transformation of Pseudomonas fluorescens and Agrobacterium tumefaciens in Soil

2001 ◽  
Vol 67 (6) ◽  
pp. 2617-2621 ◽  
Author(s):  
Sandrine Demanèche ◽  
Elisabeth Kay ◽  
François Gourbière ◽  
Pascal Simonet
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Pseudomonas Fluorescens ◽  
Natural Transformation ◽  
Physiological State ◽  
Extracellular Dna ◽  
Soil Microcosms ◽  
Do So

ABSTRACT Little information is available concerning the occurrence of natural transformation of bacteria in soil, the frequency of such events, and the actual role of this process on bacterial evolution. This is because few bacteria are known to possess the genes required to develop competence and because the tested bacteria are unable to reach this physiological state in situ. In this study we found that two soil bacteria, Agrobacterium tumefaciens and Pseudomonas fluorescens, can undergo transformation in soil microcosms without any specific physical or chemical treatment. Moreover, P. fluorescens produced transformants in both sterile and nonsterile soil microcosms but failed to do so in the various in vitro conditions we tested. A. tumefaciens could be transformed in vitro and in sterile soil samples. These results indicate that the number of transformable bacteria could be higher than previously thought and that these bacteria could find the conditions necessary for uptake of extracellular DNA in soil.

Developing in vitro models of human ductal carcinoma in situ from primary tissue explants

2015 ◽  
Vol 153 (2) ◽  
pp. 311-321 ◽  
Author(s):  
Daniel D. Brown ◽  
David J. Dabbs ◽  
Adrian V. Lee ◽  
Kandace P. McGuire ◽  
Gretchen M. Ahrendt ◽  
...  
Keyword(s):  
Ductal Carcinoma In Situ ◽  
Carcinoma In Situ ◽  
Ductal Carcinoma ◽  
In Vitro Models ◽  
Tissue Explants ◽  
Primary Tissue

scholarly journals Synthesis and Biological Screening of New Cyano-Substituted Pyrrole Fused (Iso)Quinoline Derivatives

Molecules ◽  
2021 ◽  
Vol 26 (7) ◽  
pp. 2066
Author(s):  
Maria Cristina Al-Matarneh ◽  
Roxana-Maria Amărandi ◽  
Ionel I. Mangalagiu ◽  
Ramona Danac
Keyword(s):  
Cell Lines ◽  
Cancer Cell ◽  
Human Cancer ◽  
Cancer Cell Lines ◽  
In Vitro Assays ◽  
Biological Screening ◽  
Human Cancer Cell Lines ◽  
Breast And Prostate Cancer

Several new cyano-substituted derivatives with pyrrolo[1,2-a]quinoline and pyrrolo[2,1-a]isoquinoline scaffolds were synthesized by the [3 + 2] cycloaddition of (iso)quinolinium ylides to fumaronitrile. The cycloimmonium ylides reacted in situ as 1,3-dipoles with fumaronitrile to selectively form distinct final compounds, depending on the structure of the (iso)quinolinium salt. Eleven compounds were evaluated for their anticancer activity against a panel of 60 human cancer cell lines. The most potent compound 9a showed a broad spectrum of antiproliferative activity against cancer cell lines representing leukemia, melanoma and cancer of lung, colon, central nervous system, ovary, kidney, breast and prostate cancer. In vitro assays and molecular docking revealed tubulin interaction properties of compound 9a.

scholarly journals Comparison of in Vitro Models for the Prediction of Compound Absorption across the Human Intestinal Mucosa

2004 ◽  
Vol 9 (7) ◽  
pp. 598-606 ◽  
Author(s):  
Silvia Miret ◽  
Leo Abrahamse ◽  
Els M. de Groene
Keyword(s):  
Cell Differentiation ◽  
High Throughput ◽  
High Throughput Screening ◽  
In Vitro Models ◽  
Assay System ◽  
In Vitro Assays ◽  
Cell Models ◽  
Artificial Membrane ◽  
Permeability Assay

Several in vitro assays have been developed to evaluate the gastrointestinal absorption of compounds. Our aim was to compare 3 of these methods: 1) the bio-mimetic artificial membrane permeability assay (BAMPA) method, which offers a high-throughput, noncellular approach to the measurement of passive transport; 2) the traditional Caco-2 cell assay, the use of which as a high-throughput tool is limited by the long cell differentiation time (21 days); and 3) The BioCoat™ high-throughput screening Caco-2 Assay System, which reduces Caco-2 cell differentiation to 3 days. The transport of known compounds (such as cephalexin, propranolol, or chlorothiazide) was studied at pH 7.4 and 6.5 in BAMPA and both Caco-2 cell models. Permeability data obtained was correlated to known values of human absorption. Best correlations ( r = 0.9) were obtained at pH 6.5 for BAMPA and at pH 7.4 for the Caco-2 cells grown for 21 days. The Caco-2 BioCoat™ HTS Caco-2 Assay System does not seem to be adequate for the prediction of absorption. The overall results indicate that BAMPA and the 21-day Caco-2 system can be complementary for an accurate prediction of human intestinal absorption.

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