scholarly journals Strategies for the Production of Soluble Interferon-Alpha Consensus and Potential Application in Arboviruses and SARS-CoV-2

Life ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 460
Author(s):  
Felipe Grabarz ◽  
Alexandre Paulo Yague Lopes ◽  
Flávia Ferreira Barbosa ◽  
Giovana Cappio Barazzone ◽  
Jademilson Celestino Santos ◽  
...  
Keyword(s):  
Interferon Alpha ◽  
Culture Media ◽  
High Growth ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Virus Multiplication ◽  
Clinical Approach ◽  
Biopharmaceutical Production ◽  
Vitro Model

Biopharmaceutical production is currently a multibillion-dollar industry with high growth perspectives. The research and development of biologically sourced pharmaceuticals are extremely important and a reality in our current healthcare system. Interferon alpha consensus (cIFN) is a non-natural synthetic antiviral molecule that comprises all the most prevalent amino acids of IFN-α into one consensus protein sequence. For clinical use, cIFN is produced in E. coli in the form of inclusion bodies. Here, we describe the use of two solubility tags (Fh8 and DsbC) to improve soluble cIFN production. Furthermore, we analyzed cIFN production in different culture media and temperatures in order to improve biopharmaceutical production. Our results demonstrate that Fh8-cIFN yield was improved when bacteria were cultivated in autoinduction culture medium at 30 °C. After hydrolysis, the recovery of soluble untagged cIFN was 58% from purified Fh8-cIFN molecule, fourfold higher when compared to cIFN recovered from the DsbC-cIFN, which achieved 14% recovery. The biological activity of cIFN was tested on in vitro model of antiviral effect against Zika, Mayaro, Chikungunya and SARS-CoV-2 virus infection in susceptible VERO cells. We show, for the first time, that cIFN has a potent activity against these viruses, being very low amounts of the molecule sufficient to inhibit virus multiplication. Thus, this molecule could be used in a clinical approach to treat Arboviruses and SARS-CoV-2.

scholarly journals Investigation of the effects of six medicinal plants with antiviral effects against COVID-19

2021 ◽  
Author(s):  
Harun ALP ◽  
Hasan ASİL ◽  
Demet Duman
Keyword(s):  
Medicinal Plants ◽  
Nigella Sativa ◽  
Water Extract ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Glycyrrhiza Glabra ◽  
Crocus Sativus ◽  
Medicinal And Aromatic Plants ◽  
The World

Abstract Today, the coronavirus epidemic, which caused the death of 79 million cases and 1743 thousand people in 218 countries around the world, continues to increase its impact all over the world. Researchers are still trying to develop an effective solution against covid-19, including vaccines and drugs. However, there are few studies that determine the effect of natural products obtained from plants on covid-19. Medicinal and aromatic plants have been used for therapeutic purposes since the existence of humanity. In this study, the effects of some important medicinal plants including Licorice (Glycyrrhiza glabra), Saffron (Crocus sativus L.), Nigella (Nigella sativa L.), Laurel (Lauris nobilis), Karabaş (Lavandula stoechas), and Zahter (Thymbra spicata L. var. Spicata) against Covid-19 were investigated in vitro conditions. The six plants were evaluated for cytotoxic effect on Vero cells and determining inhibition of viral replication in Vero-E6 cells at concentrations of broad-spectrum antiviral non-cytotoxic against Covid-19 in cell culture and an additional antiviral effect against Covid-19. According to the results, the five examined plants (Saffron, Nigella, Laurel, Karabaş, Zahter) were ineffective against Covid-19 in vitro conditions. Interisingly, the water extract obtained from the root of the licorice plant (Glycyrrhiza glabra) inhibited Covid-19 in vitro conditions in the 2nd dilution (1: 4) following the initial concentration in Vero-E6 cells.

scholarly journals IFN-λ1 Displays Various Levels of Antiviral Activity In Vitro in a Select Panel of RNA Viruses

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1602
Author(s):  
Marina Plotnikova ◽  
Alexey Lozhkov ◽  
Ekaterina Romanovskaya-Romanko ◽  
Irina Baranovskaya ◽  
Mariia Sergeeva ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Rna Viruses ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Genome Replication ◽  
Type I ◽  
Protective Effects ◽  
Cellular Models ◽  
Viral Genome Replication

Type III interferons (lambda IFNs) are a quite new, small family of three closely related cytokines with interferon-like activity. Attention to IFN-λ is mainly focused on direct antiviral activity in which, as with IFN-α, viral genome replication is inhibited without the participation of immune system cells. The heterodimeric receptor for lambda interferons is exposed mainly on epithelial cells, which limits its possible action on other cells, thus reducing the likelihood of developing undesirable side effects compared to type I IFN. In this study, we examined the antiviral potential of exogenous human IFN-λ1 in cellular models of viral infection. To study the protective effects of IFN-λ1, three administration schemes were used: ‘preventive’ (pretreatment); ‘preventive/therapeutic’ (pre/post); and ‘therapeutic’ (post). Three IFN-λ1 concentrations (from 10 to 500 ng/mL) were used. We have shown that human IFN-λ1 restricts SARS-CoV-2 replication in Vero cells with all three treatment schemes. In addition, we have shown a decrease in the viral loads of CHIKV and IVA with the ‘preventive’ and ‘preventive/therapeutic’ regimes. No significant antiviral effect of IFN-λ1 against AdV was detected. Our study highlights the potential for using IFN-λ as a broad-spectrum therapeutic agent against respiratory RNA viruses.

scholarly journals In vitro Assessment of Antiviral Effect of Natural Compounds on Porcine Epidemic Diarrhea Coronavirus

2021 ◽  
Vol 8 ◽  
Author(s):  
Manuel Gómez-García ◽  
Héctor Puente ◽  
Héctor Argüello ◽  
Óscar Mencía-Ares ◽  
Pedro Rubio ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Formic Acid ◽  
Antiviral Agents ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Optical Microscope ◽  
Dependent Manner ◽  
Diarrhea Virus ◽  
Porcine Epidemic Diarrhea

Organic acid and essential oils (EOs), well-known antimicrobials, could also possess antiviral activity, a characteristic which has not been completely addressed up to now. In this study, the effect of two organic acids (formic acid and sodium salt of coconut fatty acid distillates) and two single EO compounds (thymol and cinnamaldehye) was evaluated against porcine epidemic diarrhea virus (PEDV). The concentration used for each compound was established by cytotoxicity assays in Vero cells. The antiviral activity was then evaluated at three multiplicities of infection (MOIs) through visual cytopathic effect (CPE) evaluation and an alamarBlue assay as well as real-time reverse-transcription PCR (RT-qPCR) and viral titration of cell supernatants. Formic acid at at a dose of 1,200 ppm was the only compound which showed antiviral activity, with a weak reduction of CPE caused by PEDV. Through the alamarBlue fluorescence assay, we showed a significant anti-CPE effect of formic acid which could not be observed by using an inverted optical microscope. RT-qPCR and infectivity analysis also showed that formic acid significantly reduced viral RNA and viral titers in a PEDV MOI-dependent manner. Our results suggest that the antiviral activity of formic acid could be associated to its inhibitory effect on viral replication. Further studies are required to explore the anti-PEDV activity of formic acid under field conditions alone or together with other antiviral agents.

scholarly journals FDA Approved Drugs Efavirenz, Tipranavir, and Dasabuvir Inhibit Replication of Multiple Flaviviruses In Vitro

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 6
Author(s):  
Michal Stefanik ◽  
Fortunatus C Ezebuo ◽  
Jan Haviernik ◽  
Ikemefuna C. Uzochukwu ◽  
Martina Fojtikova ◽  
...  
Keyword(s):  
Yellow Fever Virus ◽  
Antiviral Drug ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Significant Inhibition ◽  
Structural Protein ◽  
Inhibition Effect ◽  
Tick Borne Encephalitis Virus ◽  
Approved Drugs

Arthropod-borne flaviviruses such as tick-borne encephalitis virus (TBEV), West Nile virus (WNV), Zika virus (ZIKV), Dengue virus (DENV), and yellow fever virus (YFV) cause several serious life-threatening syndromes (encephalitis, miscarriages, paralysis, etc.). No effective antiviral therapy against these viruses has been approved yet. We selected, via in silico modeling, 12 U.S. Food and Drug Administration (FDA)-approved antiviral drugs (paritaprevir, dolutegravir, raltegravir, efavirenz, elvitegravir, tipranavir, saquinavir, dasabuvir, delavirdine, maraviroc, trifluridine, and tauroursodeoxycholic acid) for their interaction with ZIKV proteins (NS3 helicase and protease, non-structural protein 5 (NS5) RNA-dependent RNA polymerase, and methyltransferase). Only three of them were active against ZIKV, namely, dasabuvir (ABT-333), efavirenz, and tipranavir. These compounds inhibit virus replication of ZIKV (MR-766 and Paraiba_01) in Vero cells; therefore, we tested these compounds against other medically important flaviviruses WNV (13-104 and Eg101) and TBEV (Hypr). Dasabuvir was originally developed as an antiviral drug against hepatitis C virus (HCV); tipranavir and efavirenz are used for treating human immunodeficiency virus (HIV) infection. The antiviral effects of efavirenz, tipranavir, and dasabuvir were tested for ZIKV in HUH-7, astrocytes (HBCA), and UKF-NB-4 cells, where we also identified a significant inhibition effect of these compounds. For Vero cells, efavirenz inhibited all investigated viruses with EC50 ranging from 9.70 to 29.26 µM; the tipranavir inhibition effect was from 16.19 (WNV 13-104) to 27.47 µM (TBEV), while the strongest and most robust antiviral effect was demonstrated in the case of dasabuvir (EC50 values ranging from 9.09 (TBEV) to 10.85 µM (WNV 13-104)). These results warrant further research of these drugs, either individually or in combination, as possible pan-flavivirus inhibitors.

scholarly journals Viral RNA-Dependent RNA Polymerase Inhibitor 7-Deaza-2′-C-Methyladenosine Prevents Death in a Mouse Model of West Nile Virus Infection

2019 ◽  
Vol 63 (3) ◽  
Author(s):  
Luděk Eyer ◽  
Martina Fojtíková ◽  
Radim Nencka ◽  
Ivo Rudolf ◽  
Zdeněk Hubálek ◽  
...  
Keyword(s):  
West Nile Virus ◽  
West Nile Virus Infection ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Modified Nucleosides ◽  
West Nile ◽  
Brain Infection ◽  
Highly Effective ◽  
Time Of Infection

ABSTRACTWest Nile virus (WNV) is a medically important emerging arbovirus causing serious neuroinfections in humans and against which no approved antiviral therapy is currently available. In this study, we demonstrate that 2′-C-methyl- or 4′-azido-modified nucleosides are highly effective inhibitors of WNV replication, showing nanomolar or low micromolar anti-WNV activity and negligible cytotoxicity in cell culture. One representative ofC2′-methylated nucleosides, 7-deaza-2′-C-methyladenosine, significantly protected WNV-infected mice from disease progression and mortality. Twice daily treatment at 25 mg/kg starting at the time of infection resulted in 100% survival of the mice. This compound was highly effective, even if the treatment was initiated 3 days postinfection, at the time of a peak of viremia, which resulted in a 90% survival rate. However, the antiviral effect of 7-deaza-2′-C-methyladenosine was absent or negligible when the treatment was started 8 days postinfection (i.e., at the time of extensive brain infection). The 4′-azido moiety appears to be another important determinant for highly efficient inhibition of WNV replicationin vitro. However, the strong anti-WNV effect of 4′-azidocytidine and 4′-azido-aracytidine was cell type dependent and observed predominantly in porcine kidney stable (PS) cells. The effect was much less pronounced in Vero cells. Our results indicate that 2′-C-methylated or 4′-azidated nucleosides merit further investigation as potential therapeutic agents for treating WNV infections as well as infections caused by other medically important flaviviruses.

scholarly journals Viral RNA-dependent RNA polymerase inhibitor 7-deaza-2′-C-methyladenosine prevents death in a mouse model of West Nile virus infection

2018 ◽  
Author(s):  
Luděk Eyer ◽  
Martina Fojtíková ◽  
Radim Nencka ◽  
Ivo Rudolf ◽  
Zdeněk Hubálek ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Post Infection ◽  
West Nile Virus Infection ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Modified Nucleosides ◽  
West Nile ◽  
Brain Infection ◽  
Highly Effective

AbstractWest Nile virus (WNV) is a medically important emerging arbovirus causing serious neuroinfections in humans against which no approved antiviral therapy is currently available. In this study, we demonstrate that 2′-C- methyl- or 4′-azido-modified nucleosides are highly effective inhibitors of WNV replication, showing nanomolar or low micromolar anti-WNV activity and negligible cytotoxicity in cell culture. One representative ofC2′-methylated nucleosides, 7-deaza-2′-C- methyladenosine, significantly protected WNV-infected mice from disease progression and mortality. Twice daily treatment at 25 mg/kg starting at the time of infection resulted in 100% survival of the mice. This compound was highly effective, even if the treatment was initiated 3 days post-infection, at the time of a peak of viremia, which resulted in a 90% survival rate. However, the antiviral effect of 7-deaza-2′-C- methyladenosine was absent or negligible when the treatment was started 8 days post-infection (i.e., at the time of extensive brain infection). The 4′-azido moiety appears to be another important determinant for highly efficient inhibition of WNV replication in vitro. However, the strong anti-WNV effect of 4′-azidocytidine and 4′-azido-aracytidine was cell type-dependent and observed predominantly in PS cells. The effect was much less pronounced in Vero cells. Our results indicate that 2′-C- methylated or 4′-azidated nucleosides merit further investigation as potential therapeutic agents for treating WNV infections, as well as infections caused by other medically important flaviviruses.

scholarly journals Study of the Effect of the Earthworm (Aporrectodea molleri) Cutaneous Excreta on Bacterial Growth

2021 ◽  
Vol 2021 ◽  
pp. 1-7
Author(s):  
Lamia Yakkou ◽  
Sofia Houida ◽  
Mohammed Raouane ◽  
Souad Amghar ◽  
Abdellatif El Harti
Keyword(s):  
Bacterial Growth ◽  
Growth Rates ◽  
Culture Media ◽  
Chemical Constituents ◽  
High Growth ◽  
Maximum Growth ◽  
Plant Growth Promoting Bacteria ◽  
E Coli ◽  
Conventional Medium

The effect on the bacterial growth of Aporrectodea molleri cutaneous excreta has given very satisfactory results. The cutaneous excreta were obtained using the electrical stimulation technique, recently developed by our laboratory. This technique made it possible to collect the excreta in sufficient quantities while ruling out any possible interference with the chemical constituents of the soil. The effectiveness of cutaneous excreta, tested as culture media at different concentrations on bacterial growth, was justified not only by the high growth rates but also by the concentrations used, which were several times lower than those of the conventional medium (nutrient agar). Indeed, the maximum growth rate recorded by E. coli and P. fluorescens is, respectively, 1.32 times and 2.99 times greater than that observed on the conventional medium. These higher levels are obtained at optimal excretion concentrations which are, respectively, 8 times and 133.33 times lower than the concentration of the conventional medium. The efficacy interval that delimited the excretion concentrations tested covers 8 successive concentrations ranging from 0.0075% to 1% for P. fluorescens, while for E. coli, this interval covers only three concentrations ranging from 0.06% at 0.25%. The growth rates of P. fluorescens are always higher than those of E. coli. The higher efficiency of excretions on the growth of P. fluorescens shows that this strain is more suited to use these excretions as a source of nutrients and to react positively to the stimulation of growth. These results, obtained in vitro, prove that, in the natural environment, the production of lubricating and viscous cutaneous excretions does not only have the role of facilitating the movement and digging of galleries by earthworms but also of enriching the soil in nutrients and growth factors with high added values for the activation and development of plant growth-promoting bacteria essential for soil fertility.

scholarly journals MOXIDECTIN AND IVERMECTIN INHIBIT SARS-COV-2 REPLICATION IN VERO E6 CELLS BUT NOT IN HUMAN PRIMARY AIRWAY EPITHELIUM CELLS

2021 ◽  
Author(s):  
Nilima Dinesh Kumar ◽  
Bram M. ter Ellen ◽  
Ellen M. Bouma ◽  
Berit Troost ◽  
Denise P. I. van de Pol ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Antiviral Effect ◽  
Vero Cells ◽  
In Vitro Models ◽  
World Health ◽  
Current Evidence ◽  
High Dose ◽  
Health Organization

Antiviral therapies are urgently needed to treat and limit the development of severe COVID-19 disease. Ivermectin, a broad-spectrum anti-parasitic agent, has been shown to have anti-SARS-CoV-2 activity in Vero cells at a concentration of 5 μM. These limited in vitro results triggered the investigation of ivermectin as a treatment option to alleviate COVID-19 disease. In April 2021, the World Health Organization stated, however, the following: “the current evidence on the use of ivermectin to treat COVID-19 patients is inconclusive”. It is speculated that the in vivo concentration of ivermectin is too low to exert a strong antiviral effect. Here, we performed a head-to head comparison of the antiviral activity of ivermectin and the structurally related, but metabolically more stable, moxidectin in multiple in vitro models of SARS-CoV-2 infection, including physiologically relevant human respiratory epithelial cells. Both moxidectin and ivermectin exhibited antiviral activity in Vero E6 cells. Subsequent experiments revealed that the compounds predominantly act on a step after virus cell entry. Surprisingly, however, in human airway-derived cell models, moxidectin and ivermectin failed to inhibit SARS-CoV-2 infection, even at a concentration of 10 μM. These disappointing results call for a word of caution in the interpretation of anti-SARS-CoV-2 activity of drugs solely based on Vero cells. Altogether, these findings suggest that, even by using a high-dose regimen of ivermectin or switching to another drug in the same class are unlikely to be useful for treatment against SARS-CoV-2 in humans.

scholarly journals 2586. Human Breast Milk Inhibits the Replication of Parechovirus-A3

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S899-S899
Author(s):  
Ryohei Izumita ◽  
Kazuki Kon ◽  
Yuta Aizawa ◽  
Kanako Watanabe ◽  
Akihiko Saitoh
Keyword(s):  
Breast Milk ◽  
Neutralizing Antibody ◽  
Antiviral Effect ◽  
Vero Cells ◽  
Human Breast Milk ◽  
Strong Positive Correlation ◽  
Human Breast ◽  
Mature Milk ◽  
Young Infants

Abstract Background Parechovirus-A3 (PeV-A3) is an emerging pathogen causing sepsis and meningoencephalitis in neonates and young infants. We previously reported that maternal antibodies against PeV-A3 are important to protect neonates and young infants from the infection. Recent studies showed that (1) breastfeeding had a protective effect against enterovirus, which is closely-related virus to PeV-A, and (2) human breast milk (HBM) neutralized enterovirus in vitro. Currently, no report is available related to the antiviral effect of HBM against PeV-A3. Methods HBM (colostrum, 3–5 days after childbirth; mature milk, 1 month after childbirth) and serum (within ± 1 week of child’s birthday) samples were obtained from mothers at obstetrics clinic in Niigata, Japan. Neutralizing antibody titers (NATs) against PeV-A3 were measured using the Vero cells. Results The anti-PeV-A3 NATs of colostrum (n = 32) ranged from 1:8 to 1:2048, those ≥1:32 was 59% (19/32). Whereas, the anti-PeV-A3 NATs of mature milk ranged from 1:8 to 1:96. and those ≥1:32 was 20% (2/20) (P < 0.001). The median NATs anti-PeV-A3 was higher in colostrum (1:32) compared with mature milk (1:8) (P < 0.001). There was a strong positive correlation between the NATs of colostrum and serum (r = 0.604, P < 0.001, Figure). Conclusion This study showed that HBM had high NATs against PeV-A3, which was correlated with serum NATs. Further studies are necessary to investigate which components of HBM has antiviral effects against PeV-A3. Disclosures All authors: No reported disclosures.

scholarly journals Advanced co-culture 3D breast cancer model for investigation of fibrosis induced by external stimuli: optimization study

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ilya Yakavets ◽  
Aurelie Francois ◽  
Alice Benoit ◽  
Jean-Louis Merlin ◽  
Lina Bezdetnaya ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Tumor Microenvironment ◽  
Cancer Cells ◽  
Culture Media ◽  
Luminal Breast Cancer ◽  
Radiation Toxicity ◽  
Cellular Interactions ◽  
Breast Cancer Patients ◽  
Vitro Model

AbstractRadiation-induced fibrosis (RIF) is the main late radiation toxicity in breast cancer patients. Most of the current 3D in vitro breast cancer models are composed by cancer cells only and are unable to reproduce the complex cellular homeostasis within the tumor microenvironment to study RIF mechanisms. In order to account complex cellular interactions within the tumor microenvironment, an advanced 3D spheroid model, consisting of the luminal breast cancer MCF-7 cells and MRC-5 fibroblasts, was developed. The spheroids were generated using the liquid overlay technique in culture media into 96-well plates previously coated with 1% agarose (m/v, in water). In total, 21 experimental setups were tested during the optimization of the model. The generated spheroids were characterized using fluorescence imaging, immunohistology and immunohistochemistry. The expression of ECM components was confirmed in co-culture spheroids. Using α-SMA staining, we confirmed the differentiation of healthy fibroblasts into myofibroblasts upon the co-culturing with cancer cells. The induction of fibrosis was studied in spheroids treated 24 h with 10 ng/mL TGF-β and/or 2 Gy irradiation. Overall, the developed advanced 3D stroma-rich in vitro model of breast cancer provides a possibility to study fibrosis mechanisms taking into account 3D arrangement of the complex tumor microenvironment.

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