scholarly journals Development of an In Vitro System to Study the Interactions of Aerosolized Drugs with Pulmonary Mucus

Pharmaceutics ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 145 ◽  
Author(s):  
Safar Alqahtani ◽  
Clive J. Roberts ◽  
Snjezana Stolnik ◽  
Cynthia Bosquillon
Keyword(s):  
Drug Disposition ◽  
Fluorescent Dyes ◽  
Thin Layers ◽  
Inhaled Drugs ◽  
Upper Airways ◽  
In Vitro System ◽  
Unique Parameter ◽  
Airway Mucus ◽  
The Impact

Mucus is the first biological component inhaled drugs encounter on their journey towards their pharmacological target in the upper airways. Yet, how mucus may influence drug disposition and efficacy in the lungs has been essentially overlooked. In this study, a simple in vitro system was developed to investigate the factors promoting drug interactions with airway mucus in physiologically relevant conditions. Thin layers of porcine tracheal mucus were prepared in Transwell® inserts and initially, the diffusion of various fluorescent dyes across those layers was monitored over time. A deposition system featuring a MicroSprayer® aerosolizer was optimized to reproducibly deliver liquid aerosols to multiple air-facing layers and then exploited to compare the impact of airway mucus on the transport of inhaled bronchodilators. Both the dyes and drugs tested were distinctly hindered by mucus with high logP compounds being the most affected. The diffusion rate of the bronchodilators across the layers was in the order: ipratropium ≈ glycopyronnium > formoterol > salbutamol > indacaterol, suggesting hydrophobicity plays an important role in their binding to mucus but is not the unique parameter involved. Testing of larger series of compounds would nevertheless be necessary to better understand the interactions of inhaled drugs with airway mucus.

scholarly journals SELECTIVE SYSTEM IN VITRO «FUNGI COLLETOTRICHUM LINI - LINEN» AS AN EFFICIENT WAY TO FIND OUT GENOTYPES RESISTANT TO THE POD SPOT

2019 ◽  
pp. 42-50
Author(s):  
N. V. Prolyotova
Keyword(s):  
Practical Application ◽  
In Vitro System ◽  
Linum Usitatissimum L ◽  
Regenerated Plants ◽  
Resistant Lines ◽  
Selective System ◽  
The Impact ◽  
Initial Forms ◽  
Resistant Genotypes

The research aims at development of an effective selective agent in vitro system for founding linseed genotypes resistant to the pod spot. The authors see the object of research as varieties and lines of flax Linum usitatissimum L., which differ in their resistance to the pod spot. Fungi strains differed in their virulence. The authors applied methods of such scientists as Dospekhov and Kurchakova, methodological guidance on foundation, maintenance, storage and practical application of microorganisms, i.e. flax pathogens. This results in creation of selective in vitro system “Colletotrichum lini Manns et Bolley fungus – flax”. This system selects in vitro flax cells resistant to culture filtrate, from which it is possible to obtain regenerated plants resistant to the pathogen with greater efficiency. The authors enumerate the aminoacids that were found in the culture filtrates of the investigated pathogen strains; they are alanine, glycine, asparagine, cysteine, asparagine and glutamic acids, arginine and threonine. The authors outline the observed relationship between flax cell responsiveness and fungi pathogen in the environment of the fungius - anthracnose pathogen - on the value of the explant. Anthers cells in selection conditions were less resistant than those of immature embryos. The researchers observed the impact of flax genotype on cells ability to morphogenesis under selection conditions. Genotype cells L 957-8-7, Alexim, Pendzhab, Zaryanka had high morphogenetic activity. Morphogenetic capacities of genotypes L 1506-8-4 and Rosinka were rather low by the 2nd-3rd passages. When designing the scheme of flax selection in vitro for resistance to anthracnotism, 86 shoots were obtained, the check of which on the artificial infectious-provocative background showed that the genotypes differed in their resistance. The authors observed forms less resistant to the disease as well as resistant and medium resistant lines (at the level of 50 - 75%). The parameters of resistance in resistant and medium resistant genotypes were 12 - 37% higher than in the initial forms.

scholarly journals In vitro investigation on the impact of airway mucus on drug dissolution and absorption at the air-epithelium interface in the lungs

2019 ◽  
Vol 141 ◽  
pp. 210-220 ◽  
Author(s):  
Emanuela Cingolani ◽  
Safar Alqahtani ◽  
Robyn C Sadler ◽  
David Prime ◽  
Snjezana Stolnik ◽  
...  
Keyword(s):  
Drug Dissolution ◽  
In Vitro Investigation ◽  
Airway Mucus ◽  
The Impact

scholarly journals Changes in Rumen Microbial Community Composition during Adaption to an In Vitro System and the Impact of Different Forages

PLoS ONE ◽  
2016 ◽  
Vol 11 (2) ◽  
pp. e0150115 ◽  
Author(s):  
Melanie B. Lengowski ◽  
Karin H. R. Zuber ◽  
Maren Witzig ◽  
Jens Möhring ◽  
Jeannette Boguhn ◽  
...  
Keyword(s):  
Microbial Community ◽  
Community Composition ◽  
Microbial Community Composition ◽  
In Vitro System ◽  
The Impact

scholarly journals Intermittent Hypoxia and Its Impact on Nrf2/HIF-1α Expression and ABC Transporters: An in Vitro Human Blood-Brain Barrier Model Study.

2020 ◽  
Vol 54 (6) ◽  
pp. 1231-1248
Keyword(s):  
Blood Brain Barrier ◽  
Abc Transporters ◽  
Intermittent Hypoxia ◽  
Brain Barrier ◽  
Upper Airways ◽  
Whole Cell ◽  
Blood Brain ◽  
Junction Protein ◽  
The Impact

BACKGROUND/AIMS: Obstructive sleep apnea (OSA) is characterized by repeated episodes of complete or partial obstruction of the upper airways, leading to chronic intermittent hypoxia (IH). OSA patients are considered at high cerebrovascular risk and may also present cognitive impairment. One hypothesis explored is that disturbances may be linked to blood-brain barrier (BBB) dysfunction. The BBB is a protective barrier separating the brain from blood flow. The BBB limits the paracellular pathway through tight and adherens junctions, and the transcellular passage by efflux pumps (ABC transporters). The aims of this study were to evaluate the impact of IH and sustained hypoxia (SH) on a validated in vitro BBB model and to investigate the factors expressed under both conditions. METHODS: Exposure of endothelial cells (HBEC-5i) in our in vitro model of BBB to hypoxia was performed using IH cycles: 1% O2-35 min/18% O2-25 min for 6 cycles or 6 h of SH at 1% O2. After exposure, we studied the cytotoxicity and the level of ROS in our cells. We measured the apparent BBB permeability using sodium fluorescein, FITC-dextran and TEER measurement. Whole cell ELISA were performed to evaluate the expression of tight junctions, ABC transporters, HIF-1α and Nrf2. The functionality of ABC transporters was evaluated with accumulation studies. Immunofluorescence assays were also conducted to illustrate the whole cell ELISAs. RESULTS: Our study showed that 6 h of IH or SH induced a BBB disruption marked by a significant decrease in junction protein expressions (claudin-5, VE-cadherin, ZO-1) and an increase in permeability. We also observed an upregulation in P-gp protein expression and functionality and a downregulation in BCRP. Hypoxia induced production of ROS, Nrf2 and HIF-1α. They were expressed in both sustained and intermittent conditions, but the expression and the activity of P-gp and BCRP were different. CONCLUSION: Understanding these mechanisms seems essential in order to propose new therapeutic strategies for patients with OSA

scholarly journals Development of a Traumatic Brain Injury Bioreactor

2013 ◽  
Author(s):  
Zachery E. Heller ◽  
Joseph Wyatt ◽  
Jeffery C. Wolchok
Keyword(s):  
Traumatic Brain Injury ◽  
Brain Injury ◽  
Human Brain ◽  
Full Scale ◽  
Crash Test ◽  
In Vitro System ◽  
Injury Threshold ◽  
Cells In Culture ◽  
The Impact

An in-vitro system was developed to mimic the impact deceleration and strain associated with concussive injuries. Similar in concept to a full scale crash test, the bench top concussive bioreactor can deliver decelerations from 0–250 g and biaxial strains from 5–25% to cells in culture. We are not aware of another system like this. From cellular testing, an injury threshold of 100g combined with 10% strain was identified. These values appear consistent with human brain injury data.

Physiologically-Based Pharmacokinetic (PBPK) Modelling of Transporter Mediated Drug Absorption, Clearance and Drug-Drug Interactions

2021 ◽  
Vol 21 ◽  
Author(s):  
Kunal S. Taskar ◽  
Isobel Harada ◽  
Ravindra V. Alluri
Keyword(s):  
Drug Interactions ◽  
Intestinal Absorption ◽  
Drug Disposition ◽  
Pbpk Modelling ◽  
Vitro Assay ◽  
Pbpk Models ◽  
In Vitro Systems ◽  
The Impact

Abstarct: Membrane transporters play an important role in intestinal absorption, distribution and clearance of drugs. Additionally transporters along with enzymes regulate tissue exposures (e.g. liver, kidney and brain), which are important for safety and efficacy considerations. Early identification of transporters involved guides generation of in vitro and in vivo data needed to gain mechanistic understanding on the role of transporters in organ clearance, tissue exposures and enables development of physiological-based pharmacokinetic (PBPK) models. A lot of progress has been made in developing several in vitro assay systems and mechanistic in silico models to determine kinetic parameters for transporters, which are incorporated into PBPK models. Although, intrinsic clearance and inhibition data from in vitro systems generally tend to underpredict in vivo clearance and magnitude of drug-drug interactions (DDIs), empirical scaling factors derived from a sizable dataset are often used to offset underpredictions. PBPK models are increasing used to predict the impact of transporters on intestinal absorption, clearance, victim and perpetrator DDIs prior to first in human clinical trials. The models are often refined when clinical data is available and are used to predict pharmacokinetics in untested scenarios such as the impact of polymorphisms, ontogeny, ethnicity, disease states and DDIs with other perpetrator drugs. The aim of this review is to provide an overview of (i) regulatory requirements around transporters, (ii) in vitro systems and their limitations in predicting transporter mediated drug disposition and DDIs, (iii) PBPK modelling tactics and case studies used for internal decision making and/or for regulatory submissions.

scholarly journals Application of the In Vitro HoxB8 Model System to Characterize the Contributions of Neutrophil–LPS Interaction to Periodontal Disease

Pathogens ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 530
Author(s):  
Maja Sochalska ◽  
Magdalena B. Stańczyk ◽  
Maria Użarowska ◽  
Natalia Zubrzycka ◽  
Susanne Kirschnek ◽  
...  
Keyword(s):  
Bacterial Biofilm ◽  
Toll Like Receptor ◽  
In Vitro System ◽  
Specific Effects ◽  
Model Cell ◽  
Toll Like Receptor 2 ◽  
Neutrophil Survival ◽  
Neutrophil Response ◽  
The Impact

(1) Background: Studying neutrophils in vitro is difficult since these cells are terminally differentiated and are easily activated during isolation. At the same time, most of the available model cell lines are associated with certain limitations, such as functional deficiency or a lack of expression of surface markers characteristic of neutrophils. P. gingivalis is a periodontopathogen that causes dysbiosis in subgingival bacterial biofilm. This triggers the accumulation of functional neutrophils in the periodontium. However, until now, the specific effects of P. gingivalis-derived lipopolysaccharide on neutrophil functions have not been analyzed. (2) Methods: The impact of two variants of commercially available P. gingivalis endotoxin on neutrophil functions was tested using the HoxB8 in vitro system that is well suited to analyze neutrophil response to different stimuli in a controlled manner. (3) Results: The Standard P. gingivalis lipopolysaccharide (LPS), known to activate cells through Toll-like receptor 2 (TLR2)- and Toll-like receptor 4 (TLR4)-dependent pathways, prolonged neutrophil survival and exhibited pro-inflammatory effects. In contrast, Ultrapure LPS, binding exclusively to TLR4, neither protected neutrophils from apoptosis, nor induced an inflammatory response. (4) Conclusion: Two variants of P. gingivalis-derived LPS elicited effects on neutrophils and, based on the obtained results, we concluded that the engagement of both TLR2 and TLR4 is required for the manipulation of survival and the stimulation of immune responses of HoxB8 neutrophils.

scholarly journals Runx2 deficiency in osteoblasts promotes myeloma resistance to bortezomib by increasing thrombospondin 1-mediated TGF-ß1 activation and suppression immunity in the bone marrow

2020 ◽  
Author(s):  
Chao Zhang ◽  
Xiaoxuan Xu ◽  
Timothy N. Trotter ◽  
Pramod S. Gowda ◽  
Yun Lu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Suppressor Cells ◽  
In Vitro System ◽  
Thrombospondin 1 ◽  
Related Transcription Factor ◽  
Risk Patients ◽  
Immunosuppressive Microenvironment ◽  
Cancer Chemoresistance ◽  
The Impact

Abstract Background: Multiple myeloma (MM) is a plasma cell malignancy that thrives in the bone marrow (BM). Although the proteasome inhibitor bortezomib (BTZ) is one of the most effective front-line chemotherapeutic drugs for MM, 15–20% of high-risk patients do not respond to this drug or become resistant to treatment. The mechanisms driving this chemoresistance remain unclear. Previous studies showed that the tumor microenvironment contributes to cancer chemoresistance. Our recent studies demonstrated that Runt-related transcription factor 2 (Runx2) deficiency in osteoblasts (OBs) creates a cytokine-rich and immunosuppressive microenvironment in the BM and promotes MM progression. However, the impact of Runx2 deficiency in OBs on the efficacy of BTZ in treatment of MM is still unknown. Methods: We assessed the effects of OB-Runx2 deficiency on the outcome of BTZ treatment in OB-Runx2 +/+ and OB-Runx2 -/- mouse models with MM using bioluminescence imaging, ELISA and flow cytometry. In addition, we used a co-culturing in vitro system to explore the mechanism of BTZ resistance and assessed this system by MTT assays and Western blot analysis. Results: We discovered that OB-Runx2 deficiency induces MM cells resistance to BTZ via the suppression of immunity and increased active TGF-β1 in the BM. We further demonstrated that depletion of myeloid-derived suppressor cells (MDSCs) by gemcitabine or inhibition of TGF-ß1 activity by SRI31277, a compound that blocks thrombospondin 1 (TSP1)-mediated TGF-ß1 activation, restores anti-tumor immunity in the BM and overcomes BTZ resistance induced by OB-Runx2 deficiency. In addition, SRI31277 also directly inhibits the activity of canonical (Smad2/3) and non-canonical (Erk1/2) signaling pathways of TGF-ß1 in 5TGM1-Luc MM cells and sensitizes MM cells to BTZ, resulting in increased apoptosis of MM cells. Conclusions: OB-Runx2 deficiency promotes BTZ resistance in MM cells through the regulations of MDSCs and TSP1-mediated TGF-ß1 activation in the BM. These data identify novel mechanisms of BTZ resistance in MM and suggest new strategies to overcome BTZ resistance in treatment of MM.

In Vitro Assay of Platelet Adhesiveness with Washed and Tanned Human Red Cells

1968 ◽  
Vol 20 (03/04) ◽  
pp. 384-396 ◽  
Author(s):  
G Zbinden ◽  
S Tomlin
Keyword(s):  
Platelet Adhesion ◽  
Sodium Citrate ◽  
Blood Platelets ◽  
In Vitro Assay ◽  
Red Cells ◽  
Platelet Adhesiveness ◽  
Vitro Assay ◽  
In Vitro System ◽  
Human Red Cells

SummaryAn in vitro system is described in which adhesion of blood platelets to washed and tannic acid-treated red cells was assayed quantitatively by microscopic observation. ADP, epinephrine and TAME produced a reversible increase in platelet adhesiveness which was antagonized by AMP. With Evans blue, polyanetholsulfonate, phthalanilide NSC 38280, thrombin and heparin at concentrations above 1-4 u/ml the increase was irreversible. The ADP-induced increase in adhesiveness was inhibited by sodium citrate, EDTA, AMP, ATP and N-ethylmaleimide. EDTA, AMP and the SH-blocker N-ethylmaleimide also reduced spontaneous platelet adhesion to red cells. No significant effects were observed with adenosine, phenprocoumon, 5-HT, phthalanilide NSC 57155, various estrogens, progestogens and fatty acids, acetylsalicylic acid and similarly acting agents, hydroxylamine, glucose and KCN. The method may be useful for the screening of thrombogenic and antithrombotic properties of drugs.

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