scholarly journals A New Model of Sensorial Neuron-Like Cells for HTS of Novel Analgesics for Neuropathic Pain

2018 ◽  
Vol 24 (2) ◽  
pp. 158-168 ◽  
Author(s):  
Antón L. Martínez ◽  
José Brea ◽  
Xavier Monroy ◽  
Manuel Merlos ◽  
Javier Burgueño ◽  
...  
Keyword(s):  
Neuropathic Pain ◽  
Intracellular Calcium ◽  
High Throughput Screening ◽  
Cell Model ◽  
Chemical Library ◽  
Differentiated Cells ◽  
Drg Neuron ◽  
Undifferentiated Cells ◽  
Vitro Model

In this study we developed a new translational phenotypic in vitro model for high-throughput screening (HTS) of novel analgesics for treating neuropathic pain, in order to address the poor translation of traditional recombinant models. The immortalized dorsal root ganglia (DRG) neuron-like F11 cell line was selected based on its phenotype after differentiation. The acquisition of neuronal characteristics was evaluated by measuring the expression of TrkA as a DRG neuron marker ( p < 0.01) as well as by measuring the global neurite length ( p < 0.001). The response of F11 cells to ATP and KCl was obtained by measuring intracellular calcium concentration, dynamic mass redistribution, and membrane potential. A KCl-induced increase of intracellular calcium levels was chosen as the readout because of the better signal quality, higher reproducibility, and greater compatibility with HTS assay requirements compared with other methods. The response to KCl differed significantly between differentiated and undifferentiated cells ( p < 0.05), with an EC50 value of 5 mM in differentiated cells. The model was validated by screening the Prestwick Chemical Library. Five hits already proposed for neuropathic-related pain were identified, with IC50 values between 1 and 7 µM. This cell model provides a new tool for screening novel analgesics for the relief of neuropathic pain.

Preincubation with Sn-complexes causes intensive intracellular retention of 99mTc in thyroid cells in vitro

2012 ◽  
Vol 51 (05) ◽  
pp. 179-185 ◽  
Author(s):  
M. Wendisch ◽  
D. Aurich ◽  
R. Runge ◽  
R. Freudenberg ◽  
J. Kotzerke ◽  
...  
Keyword(s):  
Cell Model ◽  
Low Energy ◽  
Thyroid Cells ◽  
Low Energy Electrons ◽  
A Cell ◽  
Vitro Model ◽  
Uptake Studies ◽  
Radiobiological Effects ◽  
Radioactivity Retention

SummaryTechnetium radiopharmaceuticals are well established in nuclear medicine. Besides its well-known gamma radiation, 99mTc emits an average of five Auger and internal conversion electrons per decay. The biological toxicity of these low-energy, high-LET (linear energy transfer) emissions is a controversial subject. One aim of this study was to estimate in a cell model how much 99mTc can be present in exposed cells and which radiobiological effects could be estimated in 99mTc-overloaded cells. Methods: Sodium iodine symporter (NIS)- positive thyroid cells were used. 99mTc-uptake studies were performed after preincubation with a non-radioactive (cold) stannous pyro - phosphate kit solution or as a standard 99mTc pyrophosphate kit preparation or with pure pertechnetate solution. Survival curves were analyzed from colony-forming assays. Results: Preincubation with stannous complexes causes irreversible intracellular radioactivity retention of 99mTc and is followed by further pertechnetate influx to an unexpectedly high 99mTc level. The uptake of 99mTc pertechnetate in NIS-positive cells can be modified using stannous pyrophosphate from 3–5% to >80%. The maximum possible cellular uptake of 99mTc was 90 Bq/cell. Compared with nearly pure extracellular irradiation from routine 99mTc complexes, cell survival was reduced by 3–4 orders of magnitude after preincubation with stannous pyrophosphate. Conclusions: Intra cellular 99mTc retention is related to reduced survival, which is most likely mediated by the emission of low-energy electrons. Our findings show that the described experiments constitute a simple and useful in vitro model for radiobiological investigations in a cell model.

scholarly journals New Inhibitory Effects of Cilnidipine on Microglial P2X7 Receptors and IL-1β Release: An Involvement in its Alleviating Effect on Neuropathic Pain

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 434
Author(s):  
Tomohiro Yamashita ◽  
Sawako Kamikaseda ◽  
Aya Tanaka ◽  
Hidetoshi Tozaki-Saitoh ◽  
Jose M. M. Caaveiro ◽  
...  
Keyword(s):  
Neuropathic Pain ◽  
High Throughput Screening ◽  
Inflammatory Responses ◽  
Therapeutic Potential ◽  
Inhibitory Effect ◽  
Intrathecal Administration ◽  
Chemical Library ◽  
P2x7 Receptors ◽  
Approved Drugs ◽  
Cardinal Symptom

P2X7 receptors (P2X7Rs) belong to a family of ATP-gated non-selective cation channels. Microglia represent a major cell type expressing P2X7Rs. The activation of microglial P2X7Rs causes the release of pro-inflammatory cytokines such as interleukin-1β (IL-1β). This response has been implicated in neuroinflammatory states in the central nervous system and in various diseases, including neuropathic pain. Thus, P2X7R may represent a potential therapeutic target. In the present study, we screened a chemical library of clinically approved drugs (1979 compounds) by high-throughput screening and showed that the Ca2+ channel blocker cilnidipine has an inhibitory effect on rodent and human P2X7R. In primary cultured rat microglial cells, cilnidipine inhibited P2X7R-mediated Ca2+ responses and IL-1β release. Moreover, in a rat model of neuropathic pain, the intrathecal administration of cilnidipine produced a reversal of nerve injury-induced mechanical hypersensitivity, a cardinal symptom of neuropathic pain. These results point to a new inhibitory effect of cilnidipine on microglial P2X7R-mediated inflammatory responses and neuropathic pain, proposing its therapeutic potential.

scholarly journals A Phenylfurocoumarin Derivative Reverses ABCG2-Mediated Multidrug Resistance In Vitro and In Vivo

2021 ◽  
Vol 22 (22) ◽  
pp. 12502
Author(s):  
Shoji Kokubo ◽  
Shinobu Ohnuma ◽  
Megumi Murakami ◽  
Haruhisa Kikuchi ◽  
Shota Funayama ◽  
...  
Keyword(s):  
Multidrug Resistance ◽  
Cancer Cells ◽  
High Throughput Screening ◽  
Atp Hydrolysis ◽  
Pheophorbide A ◽  
Chemical Library ◽  
Hct 116 ◽  
Potential Inhibitors

The ATP-binding cassette subfamily G member 2 (ABCG2) transporter is involved in the development of multidrug resistance in cancer patients. Many inhibitors of ABCG2 have been reported to enhance the chemosensitivity of cancer cells. However, none of these inhibitors are being used clinically. The aim of this study was to identify novel ABCG2 inhibitors by high-throughput screening of a chemical library. Among the 5812 compounds in the library, 23 compounds were selected in the first screening, using a fluorescent plate reader-based pheophorbide a (PhA) efflux assay. Thereafter, to validate these compounds, a flow cytometry-based PhA efflux assay was performed and 16 compounds were identified as potential inhibitors. A cytotoxic assay was then performed to assess the effect these 16 compounds had on ABCG2-mediated chemosensitivity. We found that the phenylfurocoumarin derivative (R)-9-(3,4-dimethoxyphenyl)-4-((3,3-dimethyloxiran-2-yl)methoxy)-7H-furo [3,2-g]chromen-7-one (PFC) significantly decreased the IC50 of SN-38 in HCT-116/BCRP colon cancer cells. In addition, PFC stimulated ABCG2-mediated ATP hydrolysis, suggesting that this compound interacts with the substrate-binding site of ABCG2. Furthermore, PFC reversed the resistance to irinotecan without causing toxicity in the ABCG2-overexpressing HCT-116/BCRP cell xenograft mouse model. In conclusion, PFC is a novel inhibitor of ABCG2 and has promise as a therapeutic to overcome ABCG2-mediated MDR, to improve the efficiency of cancer chemotherapy.

scholarly journals Chinese Veterinary Medicine B307 Promotes Cardiac Performance and Skeletal Muscle Contraction via Enhancing Intracellular Calcium Levels and Neural Electrical Activity in Animal and Cell Models

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Chia-Ying Lien ◽  
Chen-Wen Lu ◽  
Chih-Hsiang Hsu ◽  
Tai-Yuan Chuang ◽  
Li-Yu Su ◽  
...  
Keyword(s):  
Veterinary Medicine ◽  
Electrical Activity ◽  
Intracellular Calcium ◽  
Cell Model ◽  
Color Doppler ◽  
Neuroblastoma Cell ◽  
Motor Functions ◽  
Electrical Activities

The study mainly investigated the effects of Chinese veterinary medicine B307 in cardiac and motor functions in animal models of pigeons and mice. Related cellular mechanisms were also studied in the neuroblastoma cell model of SH-SY5Y. Cardiac functions of pigeons and mice were examined by using moorFLPI Laser color Doppler imager and M-mode echocardiography, and motor functions were examined by using muscle electrical stimulation and force recording in the isolated breast muscle. Intracellular calcium levels and electrical activity of SH-SY5Y cells were examined by using Fura 2-AM fluorescence and MED64 system separately. Our results in vivo found that those pigeons under oral B307 treatment obviously enhanced subcutaneous microcirculation and contractile force and prolonged fatigue time in their breast muscles. Those mice under oral B307 treatment obviously elevated ejection fraction and cardiac output in their hearts. Our results in vitro showed that those SH-SY5Y cells under B307 treatment obviously increased intracellular calcium mobilization and electrical activities. These results revealed that improvement of cardiac and motor functions under B307 treatments may be caused by increasing electrical activities and intracellular calcium levels in neuromuscular cells and a similar mechanism may also occur in muscle cells. Thus, we suggested that B307 can be a functional Chinese veterinary medicine for flying pigeons.

scholarly journals Involvement of 27-Hydroxycholesterol in Mitotane Action on Adrenocortical Carcinoma

Cells ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 885
Author(s):  
Antonina Germano ◽  
Daniela Rossin ◽  
Valerio Leoni ◽  
Noemi Iaia ◽  
Laura Saba ◽  
...  
Keyword(s):  
Adrenocortical Carcinoma ◽  
Cholesterol Metabolism ◽  
Cell Model ◽  
Gas Chromatography Mass Spectrometry ◽  
Strong Increase ◽  
Vitro Model ◽  
A Minor ◽  
Cholesterol Precursors

Adrenocortical carcinoma (ACC) is a rare cancer with poor prognosis. Mitotane, the standard treatment for ACC, impairs adrenocortical steroid biosynthesis and cholesterol metabolism. In the H295R cell line, a standard ACC in vitro model, mitotane was previously reported to enhance the production of some oxysterols. To verify the possible mechanistic involvement of oxysterols in the anti-ACC effect of mitotane, a gas chromatography mass spectrometry (GC-MS) profiling of oxysterols and the main cholesterol precursors was carried out in H295R cells. Among the oxysterols detected in mitotane-treated cells, 27OHC was markedly produced, as well as lanosterol and lathosterol cholesterol precursors. In this cell model, mitotane was confirmed to affect mitochondrial transmembrane potential and induce apoptosis. Such cytotoxic effects were perfectly matched by H295R cell treatment with a single identical micromolar amount of 27OHC. The mitotane-dependent strong increase in 27OHC was confirmed in vivo, in the plasma of ACC patients under treatment with the drug. Moreover, lanosterol, lathosterol, desmosterol and, to a minor extent, 24-hydroxycholesterol and 25-hydroxycholesterol plasma levels were significantly increased in those patients. The cytotoxic effect of mitotane on ACC cells may be partly related to the increased intracellular level of 27OHC induced by the drug itself.

scholarly journals In vitro neuroprotective activities of two distinct probiotic consortia

2019 ◽  
Vol 10 (4) ◽  
pp. 437-447 ◽  
Author(s):  
D.R. Michael ◽  
T.S. Davies ◽  
K.E. Loxley ◽  
M.D. Allen ◽  
M.A. Good ◽  
...  
Keyword(s):  
Cell Model ◽  
Neuronal Cell ◽  
In Vivo Studies ◽  
Intact Cells ◽  
Bacterial Consortia ◽  
Differentiated Cells ◽  
Genes Encoding ◽  
Induced Apoptosis

Neurodegeneration has been linked to changes in the gut microbiota and this study compares the neuroprotective capability of two bacterial consortia, known as Lab4 and Lab4b, using the established SH-SY5Y neuronal cell model. Firstly, varying total antioxidant capacities (TAC) were identified in the intact cells from each consortia and their secreted metabolites, referred to as conditioned media (CM). 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Crystal Violet (CV) assays of cell viability revealed that Lab4 CM and Lab4b CM could induce similar levels of proliferation in SH-SY5Y cells and, despite divergent TAC, possessed a comparable ability to protect undifferentiated and retinoic acid-differentiated cells from the cytotoxic actions of rotenone and undifferentiated cells from the cytotoxic actions of 1-methyl-4-phenylpyridinium iodide (MPP+). Lab4 CM and Lab4b CM also had the ability to attenuate rotenone-induced apoptosis and necrosis with Lab4b inducing the greater effect. Both consortia showed an analogous ability to attenuate intracellular reactive oxygen species accumulation in SH-SY5Y cells although the differential upregulation of genes encoding glutathione reductase and superoxide dismutase by Lab4 CM and Lab4b CM, respectively, implicates the involvement of consortia-specific antioxidative mechanisms of action. This study implicates Lab4 and Lab4b as potential neuroprotective agents and justifies their inclusion in further in vivo studies.

Functional changes in sodium conductances in the human neuroblastoma cell line SH-SY5Y during in vitro differentiation

1996 ◽  
Vol 76 (6) ◽  
pp. 3920-3927 ◽  
Author(s):  
M. Toselli ◽  
P. Tosetti ◽  
V. Taglietti
Keyword(s):  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Human Neuroblastoma Cell ◽  
Sodium Currents ◽  
In Vitro Differentiation ◽  
Human Neuroblastoma ◽  
Human Neuroblastoma Cell Line ◽  
Differentiated Cells ◽  
Undifferentiated Cells

1. The electrophysiological properties of voltage-dependent sodium currents were studied in the human neuroblastoma cell line SH-SY5Y before and after in vitro differentiation with retinoic acid, with the use of the whole cell variant of the patch-clamp technique. 2. Voltage steps from a holding level of -90 mV to depolarizing potentials elicited, in both undifferentiated and differentiated cells, fast inward sodium currents that were full inactivating and tetrodotoxin sensitive. 3. In undifferentiated cells the current peaked at -10 mV, the half-activation potential was -35 mV, and the half-inactivation potential was -81 mV. In differentiated cells the current peaked at + 10 mV, the half-activation potential was -28 mV, and the half-inactivation potential was -56 mV. Moreover, the peak current amplitude was about a factor of 2 larger and inactivation kinetics was about a factor of 2 slower than in undifferentiated cells. 4. This diversity in sodium channel properties was related to differences in cell excitability. Under current-clamp conditions, intracellular injection of rectangular depolarizing current stimuli from a hyperpolarized membrane potential of about -100 mV elicited graded and weak regenerative responses in undifferentiated cells, whereas overshooting action potentials with faster rising phases could be elicited in differentiated cells.

scholarly journals Identification of Angiogenesis Inhibitors Using a Co-culture Cell Model in a High-Content and High-Throughput Screening Platform

2017 ◽  
Vol 23 (3) ◽  
pp. 217-225 ◽  
Author(s):  
Shuaizhang Li ◽  
Chia-Wen Hsu ◽  
Srilatha Sakamuru ◽  
Chaozhong Zou ◽  
Ruili Huang ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Cell Model ◽  
Hypoxia Inducible Factor ◽  
Angiogenesis Inhibitors ◽  
Culture Cell ◽  
Tube Formation ◽  
Tumor Formation ◽  
Human Telomerase Reverse Transcriptase

Angiogenesis is an important hallmark of cancer, contributing to tumor formation and metastasis. In vitro angiogenesis models for analyzing tube formation serve as useful tools to study these processes. However, current in vitro co-culture models using primary cells have limitations in usefulness and consistency. Therefore, in the present study, an in vitro co-culture assay system was optimized in a 1536-well format for high-throughput screening using human telomerase reverse transcriptase (hTERT)–immortalized mesenchymal stem cells and aortic endothelial cells. The National Center for Advancing Translational Sciences (NCATS) Pharmaceutical Collection (NPC) library containing 2816 drugs was evaluated using the in vitro co-culture assay. From the screen, 35 potent inhibitors (IC50 ≤1 µM) were identified, followed by 15 weaker inhibitors (IC50 1–50 µM). Moreover, many known angiogenesis inhibitors were identified, such as topotecan, docetaxel, and bortezomib. Several potential novel angiogenesis inhibitors were also identified from this study, including thimerosal and podofilox. Among the inhibitors, some compounds were proved to be involved in the hypoxia-inducible factor-1α (HIF-1α) and the nuclear factor-kappa B (NF-κB) pathways. The co-culture model developed by using hTERT-immortalized cell lines described in this report provides a consistent and robust in vitro system for antiangiogenic drug screening.

scholarly journals High-glucose 3D INS-1 cell model combined with a microfluidic circular concentration gradient generator for high throughput screening of drugs against type 2 diabetes

RSC Advances ◽  
2018 ◽  
Vol 8 (45) ◽  
pp. 25409-25416 ◽  
Author(s):  
Yong Luo ◽  
Xiuli Zhang ◽  
Yujiao Li ◽  
Jiu Deng ◽  
Xiaorui Li ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Pharmaceutical Industry ◽  
High Throughput ◽  
Concentration Gradient ◽  
High Glucose ◽  
High Throughput Screening ◽  
Cell Model ◽  
Gradient Generator

In vitro models for screening of drugs against type 2 diabetes are crucial for the pharmaceutical industry.

scholarly journals Future Directions in the Treatment of Patients with Chronic Hepatitis C Virus Infection

1999 ◽  
Vol 13 (1) ◽  
pp. 57-62 ◽  
Author(s):  
Robert G Gish
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
High Throughput Screening ◽  
In Vitro Model ◽  
Single Agent ◽  
Protein Translation ◽  
In Vitro Translation ◽  
Biotechnology Companies ◽  
Vitro Model

Hepatitis C virus (HCV) infects over 170 million people worldwide. While interferon is currently the most used single agent therapy, this drug may result in a sustained loss of virus from the blood in only up to 15% of patients; new options for treatment are needed. With the release of ribavirin in North America and Europe, a viral clearance rate or ‘cure’ may be attained in up to 40% of patients. Developing successful antiviral therapy that prevents or delays the development of cirrhosis, liver failure and liver cancer as well as decreasing the demand for liver transplantation are clearly identified goals. Unfortunately, there is no complete in vitro model of HCV replication or translation. Due to the lack of an animal or cell culture model of HCV infection, in vitro translation screening systems to identify inhibitors of HCV protein translation are being evaluated by a large number of biotechnology companies. With advancing computer technology, high throughput screening processes are now possible and can be joined to specific in vitro model testing systems. Along with examining some of the information known about HCV therapy and the HCV genome, the present review discusses potential targets for new therapies and identifies therapeutic agents that are nearing clinical application

Sign in / Sign up

Export Citation Format

Share Document