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PLoS One ; 16(6): e0252302, 2021.
Article in English | MEDLINE | ID: covidwho-1278172

ABSTRACT

A potent therapy for the infectious coronavirus disease COVID-19 is urgently required with, at the time of writing, research in this area still ongoing. This study aims to evaluate the in vitro anti-viral activities of combinations of certain commercially available drugs that have recently formed part of COVID-19 therapy. Dual combinatory drugs, namely; Lopinavir-Ritonavir (LOPIRITO)-Clarithromycin (CLA), LOPIRITO-Azithromycin (AZI), LOPIRITO-Doxycycline (DOXY), Hydroxychloroquine (HCQ)-AZI, HCQ-DOXY, Favipiravir (FAVI)-AZI, HCQ-FAVI, and HCQ-LOPIRITO, were prepared. These drugs were mixed at specific ratios and evaluated for their safe use based on the cytotoxicity concentration (CC50) values of human umbilical cord mesenchymal stem cells. The anti-viral efficacy of these combinations in relation to Vero cells infected with SARS-CoV-2 virus isolated from a patient in Universitas Airlangga hospital, Surabaya, Indonesia and evaluated for IC50 24, 48, and 72 hours after viral inoculation was subsequently determined. Observation of the viral load in qRT-PCR was undertaken, the results of which indicated the absence of high levels of cytotoxicity in any samples and that dual combinatory drugs produced lower cytotoxicity than single drugs. In addition, these combinations demonstrated considerable effectiveness in reducing the copy number of the virus at 48 and 72 hours, while even at 24 hours, post-drug incubation resulted in low IC50 values. Most combination drugs reduced pro-inflammatory markers, i.e. IL-6 and TNF-α, while increasing the anti-inflammatory response of IL-10. According to these results, the descending order of effective dual combinatory drugs is one of LOPIRITO-AZI>LOPIRITO-DOXY>HCQ-AZI>HCQ-FAVI>LOPIRITO-CLA>HCQ-DOX. It can be suggested that dual combinatory drugs, e.g. LOPIRITO-AZI, can potentially be used in the treatment of COVID-19 infectious diseases.


Subject(s)
Anti-Bacterial Agents/pharmacology , Antiviral Agents/pharmacology , COVID-19/drug therapy , Hydroxychloroquine/pharmacology , SARS-CoV-2/drug effects , Animals , Anti-Bacterial Agents/therapeutic use , Antiviral Agents/therapeutic use , COVID-19/virology , Cell Survival/drug effects , Cells, Cultured , Chlorocebus aethiops , Drug Combinations , Hospitalization , Host-Pathogen Interactions/drug effects , Humans , Hydroxychloroquine/therapeutic use , Indonesia , Inhibitory Concentration 50 , Inpatients , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/drug effects , SARS-CoV-2/isolation & purification , SARS-CoV-2/physiology , Time Factors , Vero Cells , Viral Load/drug effects
2.
Cytotherapy ; 22(8): 458-472, 2020 08.
Article in English | MEDLINE | ID: covidwho-209852

ABSTRACT

BACKGROUND AIMS: Human platelet lysate can replace fetal bovine serum (FBS) for xeno-free ex vivo expansion of mesenchymal stromal cells (MSCs), but pooling of platelet concentrates (PCs) increases risks of pathogen transmission. We evaluated the feasibility of performing nanofiltration of platelet lysates and determined the impact on expansion of bone marrow-derived MSCs. METHODS: Platelet lysates were prepared by freeze-thawing of pathogen-reduced (Intercept) PCs suspended in 65% storage solution (SPP+) and 35% plasma, and by serum-conversion of PCs suspended in 100% plasma. Lysates were added to the MSC growth media at 10% (v/v), filtered and subjected to cascade nanofiltration on 35- and 19-nm Planova filters. Media supplemented with 10% starting platelet lysates or FBS were used as the controls. Impacts of nanofiltration on the growth media composition, removal of platelet extracellular vesicles (PEVs) and MSC expansion were evaluated. RESULTS: Nanofiltration did not detrimentally affect contents of total protein and growth factors or the biochemical composition. The clearance factor of PEVs was >3 log values. Expansion, proliferation, membrane markers, differentiation potential and immunosuppressive properties of cells in nanofiltered media were consistently better than those expanded in FBS-supplemented media. Compared with FBS, chondrogenesis and osteogenesis genes were expressed more in nanofiltered media, and there were fewer senescent cells over six passages. CONCLUSIONS: Nanofiltration of growth media supplemented with two types of platelet lysates, including one prepared from pathogen-reduced PCs, is technically feasible. These data support the possibility of developing pathogen-reduced xeno-free growth media for clinical-grade propagation of human cells.


Subject(s)
Blood Platelets/cytology , Cell Culture Techniques/methods , Filtration , Mesenchymal Stem Cells/cytology , Nanotechnology , Adipogenesis/drug effects , Biomarkers/metabolism , Cell Differentiation/drug effects , Cell Lineage/drug effects , Cell Proliferation/drug effects , Cells, Cultured , Cellular Senescence/drug effects , Culture Media/pharmacology , Extracellular Vesicles/metabolism , Gene Expression Profiling , Humans , Immunophenotyping , Intercellular Signaling Peptides and Proteins/metabolism , Intercellular Signaling Peptides and Proteins/pharmacology , Mesenchymal Stem Cells/drug effects , Osteogenesis/drug effects , Particle Size , Serum/chemistry
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