scholarly journals The Anticoagulant Nafamostat Potently Inhibits SARS-CoV-2 S Protein-Mediated Fusion in a Cell Fusion Assay System and Viral Infection In Vitro in a Cell-Type-Dependent Manner

Viruses ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 629 ◽  
Author(s):  
Mizuki Yamamoto ◽  
Maki Kiso ◽  
Yuko Sakai-Tagawa ◽  
Kiyoko Iwatsuki-Horimoto ◽  
Masaki Imai ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Assay System ◽  
Dependent Manner ◽  
Cell Type ◽  
S Protein ◽  
Candidate Drug ◽  
A Cell ◽  
Transmembrane Serine Protease ◽  
Nafamostat Mesylate

Although infection by SARS-CoV-2, the causative agent of coronavirus pneumonia disease (COVID-19), is spreading rapidly worldwide, no drug has been shown to be sufficiently effective for treating COVID-19. We previously found that nafamostat mesylate, an existing drug used for disseminated intravascular coagulation (DIC), effectively blocked Middle East respiratory syndrome coronavirus (MERS-CoV) S protein-mediated cell fusion by targeting transmembrane serine protease 2 (TMPRSS2), and inhibited MERS-CoV infection of human lung epithelium-derived Calu-3 cells. Here we established a quantitative fusion assay dependent on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) S protein, angiotensin I converting enzyme 2 (ACE2) and TMPRSS2, and found that nafamostat mesylate potently inhibited the fusion while camostat mesylate was about 10-fold less active. Furthermore, nafamostat mesylate blocked SARS-CoV-2 infection of Calu-3 cells with an effective concentration (EC)50 around 10 nM, which is below its average blood concentration after intravenous administration through continuous infusion. On the other hand, a significantly higher dose (EC50 around 30 μM) was required for VeroE6/TMPRSS2 cells, where the TMPRSS2-independent but cathepsin-dependent endosomal infection pathway likely predominates. Together, our study shows that nafamostat mesylate potently inhibits SARS-CoV-2 S protein-mediated fusion in a cell fusion assay system and also inhibits SARS-CoV-2 infection in vitro in a cell-type-dependent manner. These findings, together with accumulated clinical data regarding nafamostat’s safety, make it a likely candidate drug to treat COVID-19.

scholarly journals The anticoagulant nafamostat potently inhibits SARS-CoV-2 infection in vitro: an existing drug with multiple possible therapeutic effects

2020 ◽  
Author(s):  
Mizuki Yamamoto ◽  
Maki Kiso ◽  
Yuko Sakai-Tagawa ◽  
Kiyoko Iwatsuki-Horimoto ◽  
Masaki Imai ◽  
...  
Keyword(s):  
Continuous Infusion ◽  
Disseminated Intravascular Coagulation ◽  
Blood Concentration ◽  
Human Lung ◽  
Therapeutic Effects ◽  
Lung Epithelium ◽  
S Protein ◽  
Candidate Drug ◽  
Nafamostat Mesylate

AbstractAlthough infection by SARS-CoV-2, the causative agent of COVID-19, is spreading rapidly worldwide, no drug has been shown to be sufficiently effective for treating COVID-19. We previously found that nafamostat mesylate, an existing drug used for disseminated intravascular coagulation (DIC), effectively blocked MERS-CoV S protein-initiated cell fusion by targeting TMPRSS2, and inhibited MERS-CoV infection of human lung epithelium-derived Calu-3 cells. Here we established a quantitative fusion assay dependent on SARS-CoV-2 S protein, ACE2 and TMPRSS2, and found that nafamostat mesylate potently inhibited the fusion while camostat mesylate was about 10-fold less active. Furthermore, nafamostat mesylate blocked SARS-CoV-2 infection of Calu-3 cells with an EC50 around 10 nM, which is below its average blood concentration after intravenous administration through continuous infusion. These findings, together with accumulated clinical data regarding its safety, make nafamostat a likely candidate drug to treat COVID-19.

scholarly journals Trypsin promotes porcine deltacoronavirus mediating cell-to-cell fusion in a cell type-dependent manner

2020 ◽  
Vol 9 (1) ◽  
pp. 457-468 ◽  
Author(s):  
Yue-Lin Yang ◽  
Fandan Meng ◽  
Pan Qin ◽  
Georg Herrler ◽  
Yao-Wei Huang ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Dependent Manner ◽  
Cell Type ◽  
A Cell

scholarly journals PapRIV, a BV-2 microglial cell activating quorum sensing peptide

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yorick Janssens ◽  
Nathan Debunne ◽  
Anton De Spiegeleer ◽  
Evelien Wynendaele ◽  
Marta Planas ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Cell Model ◽  
Dependent Manner ◽  
Communication Signals ◽  
Gram Positive Bacteria ◽  
A Cell ◽  
Gastro Intestinal Tract

AbstractQuorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction.

scholarly journals C9orf72 Proteins Regulate Autophagy and Undergo Autophagosomal or Proteasomal Degradation in a Cell Type-Dependent Manner

Cells ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 1233 ◽  
Author(s):  
Leskelä ◽  
Huber ◽  
Rostalski ◽  
Natunen ◽  
Remes ◽  
...  
Keyword(s):  
Neuroblastoma Cells ◽  
Ubiquitin Proteasome System ◽  
Proteasomal Degradation ◽  
Serum Starvation ◽  
Dependent Manner ◽  
Cell Type ◽  
Proteasomal Activity ◽  
N2a Cells ◽  
Autophagy Induction ◽  
A Cell

Dysfunctional autophagy or ubiquitin-proteasome system (UPS) are suggested to underlie abnormal protein aggregation in neurodegenerative diseases. Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS)-associated C9orf72 is implicated in autophagy, but whether it activates or inhibits autophagy is partially controversial. Here, we utilized knockdown or overexpression of C9orf72 in mouse N2a neuroblastoma cells or cultured neurons to elucidate the potential role of C9orf72 proteins in autophagy and UPS. Induction of autophagy in C9orf72 knockdown N2a cells led to decreased LC3BI to LC3BII conversion, p62 degradation, and formation of LC3-containing autophagosomes, suggesting compromised autophagy. Proteasomal activity was slightly decreased. No changes in autophagy nor proteasomal activity in C9orf72-overexpressing N2a cells were observed. However, in these cells, autophagy induction by serum starvation or rapamycin led to significantly decreased C9orf72 levels. The decreased levels of C9orf72 in serum-starved N2a cells were restored by the proteasomal inhibitor lactacystin, but not by the autophagy inhibitor bafilomycin A1 (BafA1) treatment. These data suggest that C9orf72 undergoes proteasomal degradation in N2a cells during autophagy. Lactacystin significantly elevated C9orf72 levels in N2a cells and neurons, further suggesting UPS-mediated regulation. In rapamycin and BafA1-treated neurons, C9orf72 levels were significantly increased. Altogether, these findings corroborate the previously suggested regulatory role for C9orf72 in autophagy and suggest cell type-dependent regulation of C9orf72 levels via UPS and/or autophagy.

scholarly journals Vitamin A and bone formation. Effect of an excess of retinol on bone collagen synthesis in vitro

1985 ◽  
Vol 226 (3) ◽  
pp. 789-795 ◽  
Author(s):  
I Dickson ◽  
J Walls
Keyword(s):  
Bone Formation ◽  
Collagen Synthesis ◽  
Bone Collagen ◽  
Dependent Manner ◽  
Stimulate Bone Resorption ◽  
Dna And Rna ◽  
A Cell ◽  
Dose Dependent ◽  
Bone Collagen Synthesis

The influence of an excess of retinol on bone formation was studied by using cultures of embryonic-chick calvaria. Retinol decreased collagen synthesis in a dose-dependent manner, non-collagenous protein synthesis being relatively unaffected. Collagen synthesis was significantly inhibited after 24 h of culture with retinol and was progressively decreased, compared with control cultures containing no retinol, as the period of culture was increased. The effect of retinol on collagen synthesis could be reversed by incubation of calvaria for further periods in retinol-free medium. Incorporation of [3H]thymidine and [3H]uridine into DNA and RNA respectively was not altered by culturing calvaria with retinol for 22 h. These latter findings, and the selectivity for collagen synthesis, all suggested that the effect observed was not a cell-toxicity phenomenon. The effect of retinol on collagen synthesis by chick calvarial osteoblasts was probably direct and not mediated by osteoclasts, since a negligible number of the latter cells is present in chick calvaria. In cultures of neonatal murine calvaria, which contain many osteoclasts, retinol similarly inhibited synthesis of collagen, but not of non-collagenous protein; the concentrations of retinol necessary to produce the response were similar to those required to stimulate bone resorption in vitro.

scholarly journals Motility patterns of Trypanosoma cruzi trypomastigotes correlate with the efficiency of parasite invasion in vitro

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Jorge A. Arias-del-Angel ◽  
Jesús Santana-Solano ◽  
Moisés Santillán ◽  
Rebeca G. Manning-Cela
Keyword(s):  
Trypanosoma Cruzi ◽  
Cell Line ◽  
Mammalian Cells ◽  
Dependent Manner ◽  
Parasite Invasion ◽  
Mammalian Cell Cultures ◽  
A Cell ◽  
Parasite Replication ◽  
Parasite Motility

Abstract Numerous works have demonstrated that trypanosomatid motility is relevant for parasite replication and sensitivity. Nonetheless, although some findings indirectly suggest that motility also plays an important role during infection, this has not been extensively investigated. This work is aimed at partially filling this void for the case of Trypanosoma cruzi. After recording swimming T. cruzi trypomastigotes (CL Brener strain) and recovering their individual trajectories, we statistically analyzed parasite motility patterns. We did this with parasites that swim alone or above monolayer cultures of different cell lines. Our results indicate that T. cruzi trypomastigotes change their motility patterns when they are in the presence of mammalian cells, in a cell-line dependent manner. We further performed infection experiments in which each of the mammalian cell cultures were incubated for 2 h together with trypomastigotes, and measured the corresponding invasion efficiency. Not only this parameter varied from cell line to cell line, but it resulted to be positively correlated with the corresponding intensity of the motility pattern changes. Together, these results suggest that T. cruzi trypomastigotes are capable of sensing the presence of mammalian cells and of changing their motility patterns accordingly, and that this might increase their invasion efficiency.

scholarly journals Ligand-dependent and -independent activation of the transcription factor γRF-1 in a cell-free system

1995 ◽  
Vol 310 (2) ◽  
pp. 461-467 ◽  
Author(s):  
C A Feghali ◽  
T M Wright
Keyword(s):  
Transcription Factor ◽  
Tyrosine Phosphatase ◽  
Cell Fractionation ◽  
Dependent Manner ◽  
Sodium Orthovanadate ◽  
Ifn Gamma ◽  
Free System ◽  
Cell Free System ◽  
A Cell

gamma RF-1 is a recently identified transcription factor induced by interferon-gamma (IFN-gamma) which binds to a unique palindromic enhancer, gamma RE-1, in the promoter of the mig gene. This paper describes the ligand-dependent and ligand-independent activation of gamma RF-1 in a cell-free system. gamma RF-1 activity was induced by IFN-gamma in a time-dependent manner from 5 to 60 min in lysates prepared from the human monocytic leukaemia line THP-1 and the human epidermoid carcinoma line A431. The activation of gamma RF-1 in vitro required both ATP and an inhibitor of tyrosine phosphatases (sodium orthovanadate or pervanadate). In the presence of limiting concentrations (micromolar) of ATP, activation was also dependent upon stimulation with IFN-gamma, whereas at millimolar concentrations of ATP, gamma RF-1 was activated by either sodium orthovanadate or pervanadate in the absence of ligand. Based on cell fractionation studies, both membrane and cytosol components were essential for activation of gamma RF-1 in vitro. Consistent with a role for one or more tyrosine kinases in the activation of gamma RF-1, its DNA binding activity was blocked by monoclonal anti-phosphotyrosine antibodies and by the tyrosine kinase inhibitors genistein, lavendustin A and herbimycin A. A comparison with recently described pathways of IFN-mediated transcription factor regulation indicates that the in vitro activation of gamma RF-1 is unique, requiring both membrane and cytosol fractions and inhibition of endogenous tyrosine phosphatase activity.

Sign in / Sign up

Export Citation Format

Share Document