Recombinant human plasma gelsolin reverses increased permeability of the blood-brain barrier induced by the spike protein of the SARS-CoV-2 virus.

BACKGROUND: Plasma gelsolin (pGSN) is an important part of the blood actin buffer that prevents negative consequences of possible F-actin deposition in the microcirculation and has various functions during host immune response. Recent reports reveal that severe COVID-19 correlates with reduced levels of pGSN. Therefore, using an in vitro system, we investigated whether pGSN could attenuate increased permeability of the blood-brain barrier (BBB) during its exposure to the portion of the SARS-CoV-2 spike protein containing the receptor binding domain (S1 subunit). MATERIALS AND METHODS: Two- and three-dimensional models of the human BBB were constructed using the human cerebral microvascular endothelial cell line hCMEC/D3 and exposed to physiologically relevant shear stress to mimic perfusion in the central nervous system (CNS). Trans-endothelial electrical resistance (TEER) as well as immunostaining and Western blotting of tight junction (TJ) proteins assessed barrier integrity in the presence of the SARS-CoV-2 spike protein and pGSN. The IncuCyte Live Imaging system evaluated the motility of the endothelial cells. Magnetic bead-based ELISA was used to determine cytokine secretion. Additionally, quantitative real-time PCR (qRT-PCR) revealed gene expression of proteins from signaling pathways that are associated with the immune response. RESULTS: pGSN reversed S1-induced BBB permeability in both 2D and 3D BBB models in the presence of shear stress. BBB models exposed to pGSN also exhibited attenuated pro-inflammatory signaling pathways (PI3K, AKT, MAPK, NF-κB), reduced cytokine secretion (IL-6, IL-8, TNF-α), and increased expression of proteins that form intercellular TJ (ZO-1, occludin, claudin-5). CONCLUSION: Due to its anti-inflammatory and protective effects on the brain endothelium, pGSN has the potential to be an alternative therapeutic target for patients with severe SARS-CoV-2 infection, especially those suffering neurological complications of COVID-19

Bactericidal and immunomodulatory properties of magnetic nanoparticles functionalized by 1,4-dihydropyridines

Background: 1,4-Dihydropyridine (1,4-DHP) and its derivatives are well-known calcium channel blockers with antiarrhythmic and antihypertensive activities. These compounds exhibit pleiotropic effects including antimicrobial activities that rely on their positive charge and amphipathic nature. Use of magnetic nanoparticles (MNPs) as carriers of 1,4-DHP modulates their properties and enables improved formulations with higher efficacy and less toxicity. Methods: In this study, the antimicrobial and immunomodulatory activities of novel 1,4-DHP derivatives in free form and immobilized on MNPs were determined by evaluating pathogen outgrowth and proinflammatory cytokine release in experimental settings that involve incubation of various 1,4-DHPs with clinical isolates of bacteria or fungi as well as mammalian cell culture models. Results: Conventional immobilization of 1,4-DHP on aminosilane-coated MNPs markedly enhances their antimicrobial activity compared to nonimmobilized molecules, in part because of the higher affinity of these nanosystems for bacterial cell wall components in the presence of human body fluids. Conclusion: Optimized nanosystems are characterized by improved biocompatibility and higher anti-inflammatory properties that provide new opportunities for the therapy of infectious diseasesThis study was supported by the National Science Center, Poland, under grant UMO-2015/17/B/NZ6/03473 (to RB). In 2016, KN-L was awarded a fellowship from the Foundation for Polish Science (FNP). This study was conducted with the use of equipment purchased by the Medical University of Białystok as a part of the RPOWP 2007–2013 funding, Priority I, Axis 1.1, contract no UDA-RPPD.01.01.00-20-001/15-00 dated June 26, 2015.Robert Bucki: [email protected] Niemirowicz-Laskowska - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokKatarzyna Głuszek - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokEwelina Piktel - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokKarlis Pajuste - Laboratory of Membrane Active Compounds and β-Diketones, Latvian Institute of Organic Synthesis, Riga, LatviaBonita Durnaś - Department of Microbiology and Immunology, The Faculty of Health Sciences of the Jan Kochanowski University in KielceGrzegorz Król - Department of Microbiology and Immunology, The Faculty of Health Sciences of the Jan Kochanowski University in KielceAgnieszka Z. Wilczewska - Institute of Chemistry, University of BialystokPaul A Janmey - Department of Physiology, Institute for Medicine and Engineering, University of PennsylvaniaAiva Plotniece - Laboratory of Membrane Active Compounds and β-Diketones, Latvian Institute of Organic Synthesis, Riga, LatviaRobert Bucki - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokYang J, Jiang C, Yang J, Qian C, Fang D. A clean procedure for the synthesis of 1,4-dihydropyridines via Hantzsch reaction in water. Green Chem Lett Rev. 2013;6(3):262–267.Filipan-Litvić M, Litvić M, Cepanec I, Vinković V. Hantzsch synthesis of 2,6-dimethyl-3,5-dimethoxycarbonyl-4-(o-methoxyphenyl)-1,4-dihydropyridine; a novel cyclisation leading to an unusual formation of 1-amino-2-methoxy-carbonyl-3,5-bis(o-methoxyphenyl)-4-oxacyclohexan-1-ene. Molecules. 2007;12(11):2546–2558.Khedkar SA, Auti PB. 1, 4-Dihydropyridines: a class of pharmacologically important molecules. 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Inhibition of inflammatory response in human keratinocytes by magnetic nanoparticles functionalized with PBP10 peptide derived from the PIP2-binding site of human plasma gelsolin

Background: Human plasma gelsolin (pGSN) is a multifunctional actin-binding protein involved in a variety of biological processes, including neutralization of pro-infammatory molecules such as lipopolysaccharide (LPS) and lipoteichoic acid (LTA) and modulation of host infammatory response. It was found that PBP10, a synthetic rhodamine B-conjugated peptide, based on the phosphoinositide-binding site of pGSN, exerts bactericidal activity against Grampositive and Gram-negative bacteria, interacts specifcally with LPS and LTA, and limits microbial-induced infammatory efects. The therapeutic efciency of PBP10 when immobilized on the surface of iron oxide-based magnetic nanoparticles was not evaluated, to date. Results: Using the human keratinocyte cell line HaCaT stimulated by bacterially-derived LPS and LTA as an in vitro model of bacterial infection, we examined the anti-infammatory efects of nanosystems consisting of iron oxidebased magnetic nanoparticles with aminosilane (MNP@NH2) or gold shells (MNP@Au) functionalized by a set of peptides, derived from the phosphatidylinositol 4,5-bisphosphate (PIP2)-binding site of the human plasma protein gelsolin, which also binds LPS and LTA. Our results indicate that these nanosystems can kill both Gram-positive and Gram-negative bacteria and limit the production of infammatory mediators, including nitric oxide (NO), reactive oxygen species (ROS), and interleukin-8 (IL-8) in the response to heat-killed microbes or extracted bacterial cell wall components. The nanoparticles possess the potential to improve therapeutic efcacy and are characterized by lower toxicity and improved hemocompatibility when compared to free peptides. Atomic force microscopy (AFM) showed that these PBP10-based nanosystems prevented changes in nanomechanical properties of cells that were otherwise stimulated by LPS. Conclusions: Neutralization of endotoxemia-mediated cellular efects by gelsolin-derived peptides and PBP10-containing nanosystems might be considered as potent therapeutic agents in the improved therapy of bacterial infections and microbial-induced infammation.This work was fnancially supported by the National Science Center, Poland under Grant: UMO-2015/17/B/NZ6/03473 (to RB) and Medical University of Bialystok (N/ST/ZB/18/002/1162 and N/ST/ZB/18/001/1162 (to RB) and N/ST/MN/18/002/1162 (to EP). Part of the study was conducted with the use of equipment purchased by the Medical University of Białystok as part of the RPOWP 2007-2013 funding, Priority I, Axis 1.1, contract No. UDARPPD.01.01.00-20-001/15-00 dated 26.06.2015. The physicochemical studies were performed in Centre of Synthesis and Analysis BioNanoTechno of the University of Bialystok (POPW.01.03.00-20-034/09-00 and POPW.01.03.00-20004/11 projects). EP acknowledges a doctoral scholarship from Polpharma Scientifc Foundation, Poland. PAJ and RB acknowledge support from NIH grant GM111942-01.Robert Bucki: [email protected] Piktel - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokUrszula Wnorowska - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokMateusz Cieśluk - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokPiotr Deptula - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokKatarzyna Pogoda - IInstitute of Nuclear Physics Polish Academy of SciencesIwona Misztalewska‑Turkowicz - Institute of Chemistry, University of BiałystokPaulina Paprocka - Department of Microbiology and Immunology, The Faculty of Medicine and Health Sciences of the Jan Kochanowski University in KielceKatarzyna Niemirowicz‑Laskowska - Department of Microbiological and Nanobiomedical Engineering, Medical University of BialystokAgnieszka Z. 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Recombinant Human Plasma Gelsolin Stimulates Phagocytosis while Diminishing Excessive Inflammatory Responses in Mice with Pseudomonas aeruginosa Sepsis

Plasma gelsolin (pGSN) is a highly conserved abundant circulating protein, characterized by diverse immunomodulatory activities including macrophage activation and the ability to neutralize pro-inflammatory molecules produced by the host and pathogen. Using a murine model of Gram-negative sepsis initiated by the peritoneal instillation of Pseudomonas aeruginosa Xen 5, we observed a decrease in the tissue uptake of IRDye®800CW 2-deoxyglucose, an indicator of inflammation, and a decrease in bacterial growth from ascitic fluid in mice treated with intravenous recombinant human plasma gelsolin (pGSN) compared to the control vehicle. Pretreatment of the murine macrophage line RAW264.7 with pGSN, followed by addition of Pseudomonas aeruginosa Xen 5, resulted in a dose-dependent increase in the proportion of macrophages with internalized bacteria. This increased uptake was less pronounced when cells were pretreated with pGSN and then centrifuged to remove unbound pGSN before addition of bacteria to macrophages. These observations suggest that recombinant plasma gelsolin can modulate the inflammatory response while at the same time augmenting host antibacterial activity.This work was supported by the National Science Center, Poland under Grant: UMO-2015/17/B/NZ6/03473 (to RB), National Institutes of Health: GM111942 (to PAJ) and Medical University of Bialystok: SUB/1/DN/19/001/1162 (to RB), N/ST/MN/18/001/1162 (to MC). Part of the study was conducted with the use of equipment purchased by the Medical University of Białystok as part of the RPOWP 2007-2013 funding, Priority I, Axis 1.1, contract No. UDA- RPPD.01.01.00-20-001/15-00 dated 26.06.2015. This work was supported by the program of the Minister of Science and Higher Education under the name “Regional Initiative of Excellence in 2019–2022”, project number: 024/RID/2018/19, financing amount: 11.999.000,00 PLN.Ewelina Piktel: [email protected] Wnorowska: [email protected] Cieśluk: [email protected] Deptuła: [email protected] V. Prasad: [email protected] Król: [email protected] Durnaś: [email protected] Namiot: [email protected] H. Markiewicz: [email protected] Niemirowicz-Laskowska: [email protected] Z. Wilczewska: [email protected] A. Janmey: [email protected] Reszeć: [email protected] Bucki: [email protected] Piktel - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokUrszula Wnorowska - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokMateusz Cieśluk - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokPiotr Deptuła - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokSuhanya V. Prasad - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokGrzegorz Król - Department of Microbiology and Immunology, the Faculty of Medicine and Health Sciences of the Jan Kochanowski University in KielceBonita Durnaś - Department of Microbiology and Immunology, the Faculty of Medicine and Health Sciences of the Jan Kochanowski University in KielceAndrzej Namiot - Department of Anatomy, Medical University of BialystokKarolina H. Markiewicz - Institute of Chemistry, University of BiałystokKatarzyna Niemirowicz-Laskowska - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of BialystokAgnieszka Z. Wilczewska - Institute of Chemistry, University of BiałystokPaul A. Janmey - Institute for Medicine and Engineering, University of PennsylvaniaJoanna Reszeć - Department of Pathology, Medical University of BialystokRobert Bucki - Department of Medical Microbiology and Nanobiomedical Engineering, Medical University of Bialystok; Department of Microbiology and Immunology, the Faculty of Medicine and Health Sciences of the Jan Kochanowski University in KielceLee, P.S.; Patel, S.R.; Christiani, D.C.; Bajwa, E.; Stossel, T.P.; Waxman, A.B. Plasma gelsolin depletion and circulating actin in sepsis: A pilot study. PLoS ONE 2008, 3, e3712.Li-ChunHsieh, K.; Schob, S.; Zeller, M.W.; Pulli, B.; Ali, M.; Wang, C.; Chiou, T.T.; Tsang, Y.M.; Lee, P.S.; Stossel, T.P.; et al. Gelsolin decreases actin toxicity and inflammation in murine multiple sclerosis. J. Neuroimmunol. 2015, 287, 36–42.Bucki, R.; Kulakowska, A.; Byfield, F.J.; Zendzian-Piotrowska, M.; Baranowski, M.; Marzec, M.; Winer, J.P.; Ciccarelli, N.J.; Górski, J.; Drozdowski, W.; et al. 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Extracellular Vimentin as a Target Against SARS-CoV-2 Host Cell Invasion

Infection of human cells by pathogens, including SARS-CoV-2, typically proceeds by cell surface binding to a crucial receptor. The primary receptor for SARS-CoV-2 is the angiotensin-converting enzyme 2 (ACE2), yet new studies reveal the importance of additional extracellular co-receptors that mediate binding and host cell invasion by SARS-CoV-2. Vimentin is an intermediate filament protein that is increasingly recognized as being present on the extracellular surface of a subset of cell types, where it can bind to and facilitate pathogens’ cellular uptake. Biophysical and cell infection studies are done to determine whether vimentin might bind SARS-CoV-2 and facilitate its uptake. Dynamic light scattering shows that vimentin binds to pseudovirus coated with the SARS-CoV-2 spike protein, and antibodies against vimentin block in vitro SARS-CoV-2 pseudovirus infection of ACE2-expressing cells. The results are consistent with a model in which extracellular vimentin acts as a co-receptor for SARS-CoV-2 spike protein with a binding affinity less than that of the spike protein with ACE2. Extracellular vimentin may thus serve as a critical component of the SARS-CoV-2 spike protein-ACE2 complex in mediating SARS-CoV-2 cell entry, and vimentin-targeting agents may yield new therapeutic strategies for preventing and slowing SARS-CoV-2 infection

Bone Marrow Osteoblast Damage by Chemotherapeutic Agents

Hematopoietic reconstitution, following bone marrow or stem cell transplantation, requires a microenvironment niche capable of supporting both immature progenitors and stem cells with the capacity to differentiate and expand. Osteoblasts comprise one important component of this niche. We determined that treatment of human primary osteoblasts (HOB) with melphalan or VP-16 resulted in increased phospho-Smad2, consistent with increased TGF-β1 activity. This increase was coincident with reduced HOB capacity to support immature B lineage cell chemotaxis and adherence. The supportive deficit was not limited to committed progenitor cells, as human embryonic stem cells (hESC) or human CD34+ bone marrow cells co-cultured with HOB pre-exposed to melphalan, VP-16 or rTGF-β1 had profiles distinct from the same populations co-cultured with untreated HOB. Functional support deficits were downstream of changes in HOB gene expression profiles following chemotherapy exposure. Melphalan and VP-16 induced damage of HOB suggests vulnerability of this critical niche to therapeutic agents frequently utilized in pre-transplant regimens and suggests that dose escalated chemotherapy may contribute to post-transplantation hematopoietic deficits by damaging structural components of this supportive niche

Plasma Gelsolin: Indicator of Inflammation and Its Potential as a Diagnostic Tool and Therapeutic Target

Gelsolin, an actin-depolymerizing protein expressed both in extracellular fluids and in the cytoplasm of a majority of human cells, has been recently implicated in a variety of both physiological and pathological processes. Its extracellular isoform, called plasma gelsolin (pGSN), is present in blood, cerebrospinal fluid, milk, urine, and other extracellular fluids. This isoform has been recognized as a potential biomarker of inflammatory-associated medical conditions, allowing for the prediction of illness severity, recovery, efficacy of treatment, and clinical outcome. A compelling number of animal studies also demonstrate a broad spectrum of beneficial effects mediated by gelsolin, suggesting therapeutic utility for extracellular recombinant gelsolin. In the review, we summarize the current data related to the potential of pGSN as an inflammatory predictor and therapeutic target, discuss gelsolin-mediated mechanisms of action, and highlight recent progress in the clinical use of pGSN

Toxicity of parasites and their unconventional use in medicine

Introduction. Over 300 species of parasites can possibly be passed on humans. Most of the parasitic infections are defined based on their pathogenicity; however, some positive effects of a parasite existence within the human body have recently been suggested. Beneficial outcomes of parasite infections might result from the production and release of metabolites, modification of host immune response or products uptake of the host. Objective. The aim of the study was a comprehensive analysis of a wide range of effects of parasites on the human body, including an overview of the toxic and positive effects. State of knowledge. In the light of the latest research presenting the unconventional use of parasites in medicine, the widely understood of their impact on the human body can also be considered in a positive context. Clinical cases from diseases caused by the toxic effects of parasites, as described in recent years, indicate that the problem of parasitic infections still persists. Despite a great deal of knowledge about the toxic effects of parasites on the human organism and, above all, despite the improvement in sanitary conditions, there is a resurgence of parasitic infections, as evidenced, e.g. by the examples presented in this review. Conclusions. The examples of positive effects of parasites presented so far give hope for the future in terms of fighting many diseases for which pharmacological treatment has not yet brought a positive effect. A better understanding of those processes might lead to the development of new methods of unconventional medical treatment