scholarly journals Probiotics and Postbiotics from Food to Health: Antimicrobial Experimental Confirmation

2021 ◽  
Author(s):  
Janet Cheruiyot Kosgey ◽  
Mercy W. Mwaniki ◽  
Fengmin Zhang
Keyword(s):  
Microbial Pathogens ◽  
Screening Methods ◽  
Screening Process ◽  
Standard Methods ◽  
Screening Experiments ◽  
Microbial Cultures ◽  
To Come ◽  
Nutritional Benefits

The field of probiotics is up-and-coming, especially in management of microbial pathogens. Probiotics confer nutritional benefits, reduce inflammation and infection. Probiotics have also shown to be helpful in the management of microbial pathogens, which include bacteria, fungi, and viruses. To ernes this potential maximumly, there is a need for an elaborate screening system for new isolates. This entails; rigorous screening methods and thorough confirmatory systems. There is need also to come up with standard methods used to evaluate the probiotics mechanism of action both in vivo and in vitro. In summary, there is a need for a standard screening process for probiotic microorganisms that is reproducible. The aim is to ensure that, the candidate microbial cultures are not written off without proper investigations. This will also fasten the screening process and save time and resources wasted in pre-screening experiments.

Synthesis of Novel 3(N,N-dialkylamino)alkyl/phenyl Substituted Thieno [2,3-d]pyrimidinones as H1-Anti-Histaminic and Antimicrobial Agents

2019 ◽  
Vol 15 (1) ◽  
pp. 63-70
Author(s):  
Shiv Dev Singh ◽  
Arvind Kumar ◽  
Firoz Babar ◽  
Neetu Sachan ◽  
Arun Kumar Sharma
Keyword(s):  
Antimicrobial Activity ◽  
Potassium Hydroxide ◽  
Antimicrobial Agents ◽  
Pyrimidine Ring ◽  
Antimicrobial Activities ◽  
Screening Methods ◽  
Chlorpheniramine Maleate ◽  
In Vivo Screening

Background: Thienopyrimidines are the bioisoster of quinazoline and unlike quinazoline exist in three isomeric forms corresponding to the three possible types annulation of thiophene to the pyrimidine ring viz thieno[2,3-d] pyrimidine, thieno[3,2-d] pyrimidine and thieno[3,4-d]pyrimidine. Heterocyclic containing the thienopyrimidinone moiety exhibits various pronounced activities such as anti-hypertensive, analgesic and anti-inflammatory, antiviral, platelet aggregation inhibitory, antiprotozoal bronchodilatory, phosphodiesterase inhibitory, antihistaminic, antipsychotic and antimicrobial activity. Objective: Synthesis of novel 3(N,N-dialkylamino)alkyl/phenyl substituted thieno[2,3-d]pyrimidinones as H1-anti-histaminic and antimicrobial agents. Methods: A series of 3-[(N,N-dialkylamino)alkyl/phenyl]-2-(1H)thioxo-5,6,7,8-tetrahydrobenzo(b) thieno(2,3-d)pyrimidine-4(3H)-ones[4a-d], their oxo analogous [5a-d] and 3-[(N,N-dialkylamino)alkyl]- 2-chlorophenyl-5,6,7,8-tetrahydrobenzo(b)thieno(2,3-d)pyrimidine- 4 (3H)-ones[6a-d]derivative were synthesized from 2-amino-4,5,6,7-tetrahydrobenzo(b)thiophene-3-carboxylic acid by nucleophilic substitution of different N,N-dialkyl alkylene/phenylene diamines on activated 3-acylchloride moiety followed by cyclocondensation with carbon disulfide and ethanolic potassium hydroxide to get [4a-d] and in second reaction by condensation with 4-chlorobenzoyl chloride to get [6a-d] by single pot novel innovative route. The oxo analogous [5a-d] were prepared by treating derivatives [4a-d] with potassium permagnate in ethanolic KOH. The synthesized compound were evaluated for H1-antihistaminic and antimicrobial activities. Results: All synthesized compounds exhibited significant H1-antihistaminic activity by in vitro and in vivo screening methods and data were verified analytically and statistically. The compound 4a, 4b, 5a and 5b showed significant H1-antihistaminiic activity than the reference standard chlorpheniramine maleate. The compound 6d, 6c, 5c and 4c exhibited significant antimicrobial activity.

scholarly journals Therapeutic Potential of CUDC-907 (Fimepinostat) for Hepatocarcinoma Treatment Revealed by Tumor Spheroids-based Drug Screening

2020 ◽  
Author(s):  
Wei Liao ◽  
Wanren Yang ◽  
Yue Zhang ◽  
Fanhong Zeng ◽  
Jiecheng Xu ◽  
...  
Keyword(s):  
Cell Lines ◽  
Drug Screening ◽  
Screening Method ◽  
Screening Methods ◽  
Spheroid Culture ◽  
3D Print ◽  
Culture Plate ◽  
Hcc Cells

Abstract Background: Cancer is the second leading cause of death globally. However, most of the new anti-cancer agents screened by traditional drug screening methods fail in the clinic because of lack of efficacy. One of the reasons for this dilemma is that the two-dimensional (2D) culture cancer cell lines could not represent the in vivo cancer cells well. Fortunately, the development of a three-dimensional (3D) culture technique helps in this problem. Methods: The high-throughput spheroid culture plate was fabricated by using 3D print technique and agarose. 4 hepatocarcinoma (HCC) cell lines were 3D cultured to screen 19 small molecular agents based on the spheroid culture plate. 3D cultured primary HCC cells and tumor-bearing mice model were established to verify the candidate anti-hepatocarcinoma agent. Cell function experiments and western blotting were conducted to explore the anti-hepatocarcinoma mechanism of the candidate agent. Results: Based on the previous study, we established an in vitro 3D drug screening method by using our invented spheroid culture device and found that CUDC-907 can serve as a potent anti-hepatocarcinoma agent. The study data show that CUDC-907 (fimepinostat), a novel dual acting inhibitor of phosphoinositide 3-kinase (PI3K) and histone deacetylase (HDAC), has potent inhibitory effects on HCC cell lines and primary HCC cells in vitro, Animal studies have shown that CUDC-907 can also suppress HCC cells in vivo. Furthermore, we investigated the antitumor mechanism of CUDC-907 in HCC cells. We found that it inhibits the PI3K/AKT/mTOR pathway and downregulates the expression of c-Myc, leading to the suppression of HCC cells. Conclusion: Our results suggest that CUDC-907 can be a candidate anti-HCC drug, and the 3D in vitro drug screening method based on our novel spheroid culture device is promising for drug screening.

scholarly journals Tetraspanin immunocapture phenotypes extracellular vesicles according to biofluid source but may limit identification of multiplexed cancer biomarkers

2021 ◽  
Author(s):  
Rachel R. Mizenko ◽  
Terza Brostoff ◽  
Tatu Rojalin ◽  
Hanna J. Koster ◽  
Hila S. Swindell ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Extracellular Vesicles ◽  
Fluorescence Detection ◽  
Single Particle ◽  
Cancer Biomarkers ◽  
Surface Markers ◽  
Isolation Method ◽  
Standard Methods

AbstractTetraspanin expression of extracellular vesicles (EVs) is often used as a surrogate for their general detection and classification from background contaminants. This common practice typically assumes a consistent expression of tetraspanins across EV sources, thus obscuring subpopulations of variable or limited tetraspanin expression. While some recent studies indicate differential expression of tetraspanins across bulk isolated EVs, here we present analysis of single EVs isolated using various field-standard methods from a variety of in vitro and in vivo sources to identify distinct patterns in colocalization of tetraspanin expression. We report an optimized method for the use of antibodycapture single particle interferometric reflectance imaging sensing (SP-IRIS) and fluorescence detection to identify subpopulations according to tetraspanin expression and compare our findings with nanoscale flow cytometry. Using SP-IRIS and immunofluorescence, we report that tetraspanin profile is consistent from a given EV source regardless of isolation method, but that tetraspanin profiles are distinct across various sources. Tetraspanin profiles as measured by flow cytometry do not share similar trends, suggesting that limitations in subpopulation detection significantly impact apparent protein expression. We further analyzed tetraspanin expression of single EVs captured non-specifically, revealing that tetraspanin capture can bias the apparent multiplexed tetraspanin profile. Finally, we demonstrate that this bias can have significant impact on diagnostic sensitivity for tumor-associated EV surface markers. Our findings may reveal key insights into the complexities of the EV biogenesis and signaling pathways and better inform EV capture and detection platforms for diagnostic or other downstream use.

scholarly journals Selective induction of endothelial P2Y6 nucleotide receptor promotes vascular inflammation

Blood ◽  
2011 ◽  
Vol 117 (8) ◽  
pp. 2548-2555 ◽  
Author(s):  
Ann-Kathrin Riegel ◽  
Marion Faigle ◽  
Stephanie Zug ◽  
Peter Rosenberger ◽  
Bernard Robaye ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Vascular Inflammation ◽  
In Vivo Studies ◽  
Nucleotide Receptors ◽  
Transcriptional Responses ◽  
Lps Challenge ◽  
Screening Experiments ◽  
P2y6 Receptor

Abstract During a systemic inflammatory response endothelial-expressed surface molecules have been strongly implicated in orchestrating immune responses. Previous studies have shown enhanced extracellular nucleotide release during acute inflammatory conditions. Therefore, we hypothesized that endothelial nucleotide receptors could play a role in vascular inflammation. To address this hypothesis, we performed screening experiments and exposed human microvascular endothelia to inflammatory stimuli, followed by measurements of P2Y or P2X transcriptional responses. These studies showed a selective induction of the P2Y6 receptor (> 4-fold at 24 hours). Moreover, studies that used real-time reverse transcription–polymerase chain reaction, Western blot analysis, or immunofluorescence confirmed time- and dose-dependent induction of P2Y6 with tumor necrosis factor α or Lipopolysaccharide (LPS) stimulation in vitro and in vivo. Studies that used MRS 2578 as P2Y6 receptor antagonist showed attenuated nuclear factor κB reporter activity and proinflammatory gene expression in human microvascular endothelial cells in vitro. Moreover, pharmacologic or genetic in vivo studies showed attenuated inflammatory responses in P2Y6−/− mice or after P2Y6 antagonist treatment during LPS-induced vascular inflammation. These studies show an important contribution of P2Y6 signaling in enhancing vascular inflammation during systemic LPS challenge and implicate the P2Y6 receptor as a therapeutic target during systemic inflammatory responses.

scholarly journals Granulocyte aggregometry: a sensitive technique for the detection of C5a and complement activation

Blood ◽  
1980 ◽  
Vol 55 (6) ◽  
pp. 898-902 ◽  
Author(s):  
DE Hammerschmidt ◽  
TK Bowers ◽  
CJ Lammi-Keefe ◽  
HS Jacob ◽  
PR Craddock
Keyword(s):  
Lupus Erythematosus ◽  
Cytochalasin B ◽  
Normal Subjects ◽  
Standard Methods ◽  
Sensitive Technique ◽  
Fresh Serum ◽  
Induced Aggregation ◽  
Transfusion Reactions

Abstract We have previously shown that complement (C) activated plasma causes granulocyte (PMN) aggregation in vitro and that C5a is responsible. The C-induced aggregation of PMNs treated with cytochalasin-B (CB) is markedly enhanced and irreversible, and the magnitude of the response is proportional to the log (concentration of activated plasma), allowing use of this technique to detect C5a and hence C-activation. To compare the sensitivity of granulocyte aggregometry to that of more standard methods of detecting C-activation, we produced graded C- activation in vitro by treating fresh serum with varying amounts of zymosan. Aggregometry was the most sensitive index of C-activation, detecting C-activation, produced by 0.02 mg zymosan/ml of serum--1/10 that required to produce C-activation detectable by C3 immunoelectrophoresis (the next most sensitive technique). Granulocyte aggregometry may also be used to detect in vivo C-activation. We have found aggregating activity in plasmas from patients with systemic lupus erythematosus, immune vasculitis, transfusion reactions, and other conditions associated with in vivo C-activation, but not in the plasmas of normal subjects.

scholarly journals A Unique Panel of Patient-Derived Cutaneous Squamous Cell Carcinoma Cell Lines Provides a Preclinical Pathway for Therapeutic Testing

2019 ◽  
Vol 20 (14) ◽  
pp. 3428 ◽  
Author(s):  
Sakinah Hassan ◽  
Karin J. Purdie ◽  
Jun Wang ◽  
Catherine A. Harwood ◽  
Charlotte M. Proby ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Cell Lines ◽  
Squamous Cell ◽  
Drug Screening ◽  
Cutaneous Squamous Cell Carcinoma ◽  
Model Systems ◽  
Screening Methods

Background: Cutaneous squamous cell carcinoma (cSCC) incidence continues to rise with increasing morbidity and mortality, with limited treatment options for advanced disease. Future improvements in targeted therapy will rely on advances in genomic/transcriptomic understanding and the use of model systems for basic research. We describe here the panel of 16 primary and metastatic cSCC cell lines developed and characterised over the past three decades in our laboratory in order to provide such a resource for future preclinical research and drug screening. Methods: Primary keratinocytes were isolated from cSCC tumours and metastases, and cell lines were established. These were characterised using short tandem repeat (STR) profiling and genotyped by whole exome sequencing. Multiple in vitro assays were performed to document their morphology, growth characteristics, migration and invasion characteristics, and in vivo xenograft growth. Results: STR profiles of the cSCC lines allow the confirmation of their unique identity. Phylogenetic trees derived from exome sequence analysis of the matched primary and metastatic lines provide insight into the genetic basis of disease progression. The results of in vivo and in vitro analyses allow researchers to select suitable cell lines for specific experimentation. Conclusions: There are few well-characterised cSCC lines available for widespread preclinical experimentation and drug screening. The described cSCC cell line panel provides a critical tool for in vitro and in vivo experimentation.

scholarly journals Differential Regulation of β-Defensin Gene Expression during Cryptosporidium parvum Infection

2004 ◽  
Vol 72 (5) ◽  
pp. 2772-2779 ◽  
Author(s):  
Tarek K. Zaalouk ◽  
Mona Bajaj-Elliott ◽  
John T. George ◽  
Vincent McDonald
Keyword(s):  
Gene Expression ◽  
Cryptosporidium Parvum ◽  
Protozoan Parasite ◽  
Immune Defense ◽  
Microbial Pathogens ◽  
In Vivo Models ◽  
Defensin Gene ◽  
Intestinal Epithelia

ABSTRACT Invasion of enterocytes by pathogenic microbes evokes both innate and adaptive immune responses, and microbial pathogens have developed strategies to overcome the initial host immune defense. β-Defensins are potentially important endogenous antibiotic-like effectors of innate immunity expressed by intestinal epithelia. In this study, the interplay between the enteric protozoan parasite Cryptosporidium parvum and host epithelial β-defensin expression was investigated. Using human and murine models of infection, we demonstrated that C. parvum infection differentially regulates β-defensin gene expression. Downregulation of murine β-defensin-1 mRNA and protein was observed in both in vitro and in vivo models of infection. Infection of the human colonic HT29 cell line with the parasite resulted in differential effects on various members of the defensin gene family. Partial reduction in human β-defensin-1 (hBD-1), induction of hBD-2, and no effect on hBD-3 gene expression was observed. Recombinant hBD-1 and hBD-2 peptides exhibited significant antimicrobial activity against C. parvum sporozoites in vitro. These findings demonstrate that C. parvum infection of enterocytes may affect the expression of various defensins in different ways and suggest that the overall outcome of the effect of antimicrobial peptides on early survival of the parasite may be complex.

Toxicity assessment of nanoparticles in various systems and organs

2017 ◽  
Vol 6 (3) ◽  
pp. 279-289 ◽  
Author(s):  
Yuan Yang ◽  
Zhen Qin ◽  
Wei Zeng ◽  
Ting Yang ◽  
Yubin Cao ◽  
...  
Keyword(s):  
Predictive Models ◽  
Toxicity Assessment ◽  
Biological Applications ◽  
Standard Methods ◽  
Immune Systems ◽  
The Past ◽  
Injury Mechanisms ◽  
Histopathological Studies

AbstractIn the past decades, much attention has been paid to toxicity assessment of nanoparticles prior to clinical and biological applications. Whilein vitrostudies have been increasing constantly,in vivostudies of nanoparticles have not established a unified system until now. Predictive models and validated standard methods are imperative. This review summarizes the current progress in approaches assessing nanotoxicity in main systems, including the hepatic and renal, gastrointestinal, pulmonary, cardiovascular, nervous, and immune systems. Histopathological studies and specific functional examinations in each system are elucidated. Related injury mechanisms are also discussed.

Micro- and Nanofluidics for Cell Biology, Cell Therapy, and Cell-Based Drug Testing

2009 ◽  
Author(s):  
Shuichi Takayama ◽  
Dongeun Huh ◽  
Jonathan Song ◽  
Wansik Cha ◽  
Yunseok Heo
Keyword(s):  
Cell Biology ◽  
Mammalian Cells ◽  
Cancer Metastasis ◽  
Dna Analysis ◽  
The Body ◽  
Biological Information ◽  
Screening Methods ◽  
Biological Studies

Many biological studies, drug screening methods, and cellular therapies require culture and manipulation of living cells outside of their natural environment in the body. The gap between the cellular microenvironment in vivo and in vitro, however, poses challenges for obtaining physiologically relevant responses from cells used in basic biological studies or drug screens and for drawing out the maximum functional potential from cells used therapeutically. One of the reasons for this gap is because the fluidic environment of mammalian cells in vivo is microscale and dynamic whereas typical in vitro cultures are macroscopic and static. This presentation will give an overview of efforts in our laboratory to develop microfluidic systems that enable spatio-temporal control of both the chemical and fluid mechanical environment of cells. The technologies and methods close the physiology gap to provide biological information otherwise unobtainable and to enhance cellular performance in therapeutic applications. Specific biomedical topics that will be discussed include, in vitro fertilization on a chip, microfluidic tissue engineering of small airway injuries, breast cancer metastasis on a chip, electrochemical biosensors, and development of tuneable nanofluidic systems towards applications in single molecule DNA analysis.

scholarly journals Assessment of inhibitory effects on major human cytochrome P450 enzymes by spasmolytics used in the treatment of overactive bladder syndrome

2017 ◽  
Vol 9 (7) ◽  
pp. 163-177
Author(s):  
Dominik Dahlinger ◽  
Sevinc Aslan ◽  
Markus Pietsch ◽  
Sebastian Frechen ◽  
Uwe Fuhr
Keyword(s):  
Cytochrome P450 ◽  
Liver Microsomes ◽  
Fold Increase ◽  
Pharmacokinetic Data ◽  
Cytochrome P450 Enzymes ◽  
Trospium Chloride ◽  
Screening Experiments ◽  
Human Cytochrome

Background: The objective of this study was to examine the inhibitory potential of darifenacin, fesoterodine, oxybutynin, propiverine, solifenacin, tolterodine and trospium chloride on the seven major human cytochrome P450 enzymes (CYP) by using a standardized and validated seven-in-one cytochrome P450 cocktail inhibition assay. Methods: An in vitro cocktail of seven highly selective probe substrates was incubated with human liver microsomes and varying concentrations of the seven test compounds. The major metabolites of the probe substrates were simultaneously analysed using a validated liquid chromatography tandem mass spectrometry (LC-MS/MS) method. Enzyme kinetics were estimated by determining IC50 and Ki values via nonlinear regression. Obtained Ki values were used for predictions of potential clinical impact of the inhibition using a static mechanistic prediction model. Results: In this study, 49 IC50 experiments were conducted. In six cases, IC50 values lower than the calculated threshold for drug–drug interactions (DDIs) in the gut wall were observed. In these cases, no increase in inhibition was determined after a 30 min preincubation. Considering a typical dosing regimen and applying the obtained Ki values of 0.72 µM (darifenacin, 15 mg daily) and 7.2 µM [propiverine, 30 mg daily, immediate release (IR)] for the inhibition of CYP2D6 yielded a predicted 1.9-fold and 1.4-fold increase in the area under the curve (AUC) of debrisoquine (CYP2D6 substrate), respectively. Due to the inhibition of the particular intestinal CYP3A4, the obtained Ki values of 14 µM of propiverine (30 mg daily, IR) resulted in a predicted doubling of the AUC for midazolam (CYP3A4 substrate). Conclusions: In vitro/ in vivo extrapolation based on pharmacokinetic data and the conducted screening experiments yielded similar effects of darifenacin on CYP2D6 and propiverine on CYP3A4 as obtained in separately conducted in vivo DDI studies. As a novel finding, propiverine was identified to potentially inhibit CYP2D6 at clinically occurring concentrations.

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