scholarly journals Novel Immunodominant Peptide Presentation Strategy: a Featured HLA-A*2402-Restricted Cytotoxic T-Lymphocyte Epitope Stabilized by Intrachain Hydrogen Bonds from Severe Acute Respiratory Syndrome Coronavirus Nucleocapsid Protein

2010 ◽  
Vol 84 (22) ◽  
pp. 11849-11857 ◽  
Author(s):  
Jun Liu ◽  
Peng Wu ◽  
Feng Gao ◽  
Jianxun Qi ◽  
Ai Kawana-Tachikawa ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Hydrogen Bonds ◽  
Mononuclear Cells ◽  
Vaccine Development ◽  
Computer Assisted ◽  
Antigenic Peptides ◽  
Peripheral Blood Mononuclear ◽  
Presentation Strategy ◽  
Peptide Presentation

ABSTRACT Antigenic peptides recognized by virus-specific cytotoxic T lymphocytes (CTLs) are presented by major histocompatibility complex (MHC; or human leukocyte antigen [HLA] in humans) molecules, and the peptide selection and presentation strategy of the host has been studied to guide our understanding of cellular immunity and vaccine development. Here, a severe acute respiratory syndrome coronavirus (SARS-CoV) nucleocapsid (N) protein-derived CTL epitope, N1 (QFKDNVILL), restricted by HLA-A*2402 was identified by a series of in vitro studies, including a computer-assisted algorithm for prediction, stabilization of the peptide by co-refolding with HLA-A*2402 heavy chain and β2-microglobulin (β2m), and T2-A24 cell binding. Consequently, the antigenicity of the peptide was confirmed by enzyme-linked immunospot (ELISPOT), proliferation assays, and HLA-peptide complex tetramer staining using peripheral blood mononuclear cells (PBMCs) from donors who had recovered from SARS donors. Furthermore, the crystal structure of HLA-A*2402 complexed with peptide N1 was determined, and the featured peptide was characterized with two unexpected intrachain hydrogen bonds which augment the central residues to bulge out of the binding groove. This may contribute to the T-cell receptor (TCR) interaction, showing a host immunodominant peptide presentation strategy. Meanwhile, a rapid and efficient strategy is presented for the determination of naturally presented CTL epitopes in the context of given HLA alleles of interest from long immunogenic overlapping peptides.

scholarly journals Peptides derived from gp43, the most antigenic protein from Paracoccidioides brasiliensis, form amyloid fibrils in vitro: implications for vaccine development

2021 ◽  
Author(s):  
Thyago R Cardim-Pires ◽  
Sant'Anna Ricardo ◽  
Debora Foguel
Keyword(s):  
In Silico ◽  
Mononuclear Cells ◽  
Vaccine Development ◽  
Paracoccidioides Brasiliensis ◽  
Public Health Problem ◽  
Proteinase K ◽  
Important Public Health Problem ◽  
Peripheral Blood Mononuclear ◽  
Potential Vaccine

Fungal infection is an important public health problem afflicting more than a billion people worldwide. Mycoses are especially important in Latin America, and in Brazil in particular. Paracoccidioides is the genus of fungi responsible for paracoccidioidomycosis comprising two species, P. brasiliensis and P. lutzii. The lungs are the primary infection site, but oral mucosa and airways can also be affected. The glycoprotein gp43 is involved in fungi adhesion to epithelial cells and is the most studied protein of P. brasiliensis. Seminal work identified a specific stretch of 15 amino acids that spans the region 181-195 (called P10) as an important epitope of gp43, being recognized by T lymphocytes in peripheral blood mononuclear cells of mice and humans and is envisioned as a potential vaccine component. Here, we show by using thioflavin T (ThT), transmission electron microscopy and other methods that synthetic P10 forms typical amyloid aggregates in solution in very short times, a property that could hamper vaccine development. In silico, aggregation analysis reveals several aggregation-prone regions (APR) in the P10 sequence that are capable of forming amyloid cores with steric zipper architecture. Seeds of P10 obtained by fibril mechanical fragmentation were able to induce the aggregation of P4 but not P23, as evidenced by ThT binding and mass spectrometry. These two peptides, also derived from gp43, are potent modulators of local and systemic inflammation. In-silico proteolysis studies with gp43 revealed that aggregation-prone, P10-like peptides could be generated by the action of several proteases such as proteinase K, trypsin and pepsin, which suggests that P10 could be formed upon gp43 digestion in a physiological condition. Considering our data in the context of a potential vaccine development, we redesigned the sequence of the P10 peptide, maintaining the antigenic region (HTLAIR), but drastically reducing its aggregation propensity.

Preliminary Report of In vitro and In vivo Effectiveness of Dornase Alfa on SARS-CoV-2 Infection (Preprint)

2020 ◽  
Author(s):  
Hacer Kuzu Okur ◽  
Koray Yalcin ◽  
Cihan Tastan ◽  
Sevda Demir ◽  
Bulut Yurtsever ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Respiratory Tract ◽  
Mononuclear Cells ◽  
Multiple Organ ◽  
Peripheral Blood Mononuclear ◽  
Dornase Alfa ◽  
Tract Infections ◽  
Adult Peripheral Blood

UNSTRUCTURED Dornase alfa, the recombinant form of the human DNase I enzyme, breaks down neutrophil extracellular traps (NET) that include a vast amount of DNA fragments, histones, microbicidal proteins and oxidant enzymes released from necrotic neutrophils in the highly viscous mucus of cystic fibrosis patients. Dornase alfa has been used for decades in patients with cystic fibrosis to reduce the viscoelasticity of respiratory tract secretions, to decrease the severity of respiratory tract infections, and to improve lung function. Previous studies have linked abnormal NET formations to lung diseases, especially to acute respiratory distress syndrome (ARDS). Coronavirus disease 2019 (COVID-19) pandemic affected more than two million people over the world, resulting in unprecedented health, social and economic crises. The COVID-19, viral pneumonia that progresses to ARDS and even multiple organ failure, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). High blood neutrophil levels are an early indicator of SARS-CoV-2 infection and predict severe respiratory diseases. A similar mucus structure is detected in COVID-19 patients due to the accumulation of excessive NET in the lungs. Here, we show our preliminary results with dornase alfa that may have an in-vitro anti-viral effect against SARS-CoV-2 infection in a bovine kidney cell line, MDBK without drug toxicity on healthy adult peripheral blood mononuclear cells. In this preliminary study, we also showed that dornase alfa can promote clearance of NET formation in both an in-vitro and three COVID-19 cases who showed clinical improvement in radiological analysis (2-of-3 cases), oxygen saturation (SpO2), respiratory rate, disappearing of dyspnea and coughing.

scholarly journals Potential Role and Impact of Peripheral Blood Mononuclear Cells in Radiographic Axial Spondyloarthritis-Associated Endothelial Dysfunction

Diagnostics ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1037
Author(s):  
Patricia Ruiz-Limon ◽  
Maria L. Ladehesa-Pineda ◽  
Clementina Lopez-Medina ◽  
Chary Lopez-Pedrera ◽  
Maria C. Abalos-Aguilera ◽  
...  
Keyword(s):  
Endothelial Dysfunction ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Axial Spondyloarthritis ◽  
Endothelial Injury ◽  
In Vitro Studies ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

Endothelial dysfunction (ED) is well known as a process that can lead to atherosclerosis and is frequently presented in radiographic axial spondyloarthritis (r-axSpA) patients. Here, we investigated cellular and molecular mechanisms underlying r-axSpA-related ED, and analyzed the potential effect of peripheral blood mononuclear cells (PBMCs) in promoting endothelial injury in r-axSpA. A total of 30 r-axSpA patients and 32 healthy donors (HDs) were evaluated. The endothelial function, inflammatory and atherogenic profile, and oxidative stress were quantified. In vitro studies were designed to evaluate the effect of PBMCs from r-axSpA patients on aberrant endothelial activation. Compared to HDs, our study found that, associated with ED and the plasma proatherogenic profile present in r-axSpA, PBMCs from these patients displayed a pro-oxidative, proinflammatory, and proatherogenic phenotype, with most molecular changes noticed in lymphocytes. Correlation studies revealed the relationship between this phenotype and the microvascular function. Additional in vitro studies confirmed that PBMCs from r-axSpA patients promoted endothelial injury. Altogether, this study suggests the relevance of r-axSpA itself as a strong and independent cardiovascular risk factor, contributing to a dysfunctional endothelium and atherogenic status by aberrant activation of PBMCs. Lymphocytes could be the main contributors in the development of ED and subsequent atherosclerosis in this pathology.

Differential expression of miRNAs in canine peripheral blood mononuclear cells (PBMC) exposed to Leishmania infantum in vitro

2021 ◽  
Vol 134 ◽  
pp. 58-63
Author(s):  
Matheus Fujimura Soares ◽  
Larissa Martins Melo ◽  
Jaqueline Poleto Bragato ◽  
Amanda de Oliveira Furlan ◽  
Natália Francisco Scaramele ◽  
...  
Keyword(s):  
Differential Expression ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Leishmania Infantum ◽  
Mononuclear Cells ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

scholarly journals Noble Metals for Modern Implant Materials: MOCVD of Film Structures and Cytotoxical, Antibacterial, and Histological Studies

Biomedicines ◽  
2021 ◽  
Vol 9 (8) ◽  
pp. 851
Author(s):  
Svetlana I. Dorovskikh ◽  
Evgeniia S. Vikulova ◽  
Elena V. Chepeleva ◽  
Maria B. Vasilieva ◽  
Dmitriy A. Nasimov ◽  
...  
Keyword(s):  
Vapor Deposition ◽  
Physical Vapor Deposition ◽  
Noble Metals ◽  
Mononuclear Cells ◽  
Chemical Vapor ◽  
Organic Chemical ◽  
Ti Alloy ◽  
Peripheral Blood Mononuclear ◽  
Histological Studies

This work is aimed at developing the modification of the surface of medical implants with film materials based on noble metals in order to improve their biological characteristics. Gas-phase transportation methods were proposed to obtain such materials. To determine the effect of the material of the bottom layer of heterometallic structures, Ir, Pt, and PtIr coatings with a thickness of 1.4–1.5 μm were deposited by metal–organic chemical vapor deposition (MOCVD) on Ti6Al4V alloy discs. Two types of antibacterial components, namely, gold nanoparticles (AuNPs) and discontinuous Ag coatings, were deposited on the surface of these coatings. AuNPs (11–14 nm) were deposited by a pulsed MOCVD method, while Ag films (35–40 nm in thickness) were obtained by physical vapor deposition (PVD). The cytotoxic (24 h and 48 h, toward peripheral blood mononuclear cells (PBMCs)) and antibacterial (24 h) properties of monophase (Ag, Ir, Pt, and PtIr) and heterophase (Ag/Pt, Ag/Ir, Ag/PtIr, Au/Pt, Au/Ir, and Au/PtIr) film materials deposited on Ti-alloy samples were studied in vitro and compared with those of uncoated Ti-alloy samples. Studies of the cytokine production by PBMCs in response to incubation of the samples for 24 and 48 h and histological studies at 1 and 3 months after subcutaneous implantation in rats were also performed. Despite the comparable thickness of the fibrous capsule after 3 months, a faster completion of the active phase of encapsulation was observed for the coated implants compared to the Ti alloy analogs. For the Ag-containing samples, growth inhibition of S. epidermidis, S. aureus, Str. pyogenes, P. aeruginosa, and Ent. faecium was observed.

scholarly journals Identification of Promiscuous African Swine Fever Virus T-Cell Determinants Using a Multiple Technical Approach

Vaccines ◽  
2021 ◽  
Vol 9 (1) ◽  
pp. 29
Author(s):  
Laia Bosch-Camós ◽  
Elisabet López ◽  
María Jesús Navas ◽  
Sonia Pina-Pedrero ◽  
Francesc Accensi ◽  
...  
Keyword(s):  
T Cell ◽  
Mononuclear Cells ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Cd8 T Cell ◽  
Protective Role ◽  
Full Length ◽  
T Cell Epitopes ◽  
Peripheral Blood Mononuclear

The development of subunit vaccines against African swine fever (ASF) is mainly hindered by the lack of knowledge regarding the specific ASF virus (ASFV) antigens involved in protection. As a good example, the identity of ASFV-specific CD8+ T-cell determinants remains largely unknown, despite their protective role being established a long time ago. Aiming to identify them, we implemented the IFNγ ELISpot as readout assay, using as effector cells peripheral blood mononuclear cells (PBMCs) from pigs surviving experimental challenge with Georgia2007/1. As stimuli for the ELISpot, ASFV-specific peptides or full-length proteins identified by three complementary strategies were used. In silico prediction of specific CD8+ T-cell epitopes allowed identifying a 19-mer peptide from MGF100-1L, as frequently recognized by surviving pigs. Complementarily, the repertoire of SLA I-bound peptides identified in ASFV-infected porcine alveolar macrophages (PAMs), allowed the characterization of five additional SLA I-restricted ASFV-specific epitopes. Finally, in vitro stimulation studies using fibroblasts transfected with plasmids encoding full-length ASFV proteins, led to the identification of MGF505-7R, A238L and MGF100-1L as promiscuously recognized antigens. Interestingly, each one of these proteins contain individual peptides recognized by surviving pigs. Identification of the same ASFV determinants by means of such different approaches reinforce the results presented here.

scholarly journals The Effect of the Clenbuterol—β2-Adrenergic Receptor Agonist on the Peripheral Blood Mononuclear Cells Proliferation, Phenotype, Functions, and Reactive Oxygen Species Production in Race Horses In Vitro

Cells ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 936
Author(s):  
Olga Witkowska-Piłaszewicz ◽  
Rafał Pingwara ◽  
Jarosław Szczepaniak ◽  
Anna Winnicka
Keyword(s):  
Reactive Oxygen Species ◽  
Acute Exercise ◽  
Mononuclear Cells ◽  
Anabolic Steroids ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Peripheral Blood Mononuclear ◽  
Adrenoceptor Agonist ◽  
Race Horses

Clenbuterol, the β2-adrenoceptor agonist, is gaining growing popularity because of its effects on weight loss (i.e., chemical liposuction). It is also popular in bodybuilding and professional sports, due to its effects that are similar to anabolic steroids. However, it is prohibited by anti-doping control. On the other hand, it is suggested that clenbuterol can inhibit the inflammatory process. The cells from 14 untrained and 14 well-trained race horses were collected after acute exercise and cultured with clenbuterol. The expressions of CD4, CD8, FoxP3, CD14, MHCII, and CD5 in PBMC, and reactive oxygen species (ROS) production, as well as cell proliferation, were evaluated by flow cytometry. In addition, IL-1β, IL-4, IL-6, IL-10, IL-17, INF-γ and TNF-α concentrations were evaluated by ELISA. β2-adrenoceptor stimulation leads to enhanced anti-inflammatory properties in well-trained horses, as do low doses in untrained animals. In contrast, higher clenbuterol doses create a pro-inflammatory environment in inexperienced horses. In conclusion, β2-adrenoceptor stimulation leads to a biphasic response. In addition, the immune cells are more sensitive to drug abuse in inexperienced individuals under physical training.

scholarly journals A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Rachel Tanner ◽  
Andrew D. White ◽  
Charelle Boot ◽  
Claudia C. Sombroek ◽  
Matthew K. O’Shea ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mononuclear Cells ◽  
Functional Assay ◽  
Intermediate Precision ◽  
Peripheral Blood Mononuclear ◽  
Growth Inhibition Assay ◽  
Early Evaluation ◽  
Human Primate

AbstractWe present a non-human primate mycobacterial growth inhibition assay (MGIA) using in vitro blood or cell co-culture with the aim of refining and expediting early tuberculosis vaccine testing. We have taken steps to optimise the assay using cryopreserved peripheral blood mononuclear cells, transfer it to end-user institutes, and assess technical and biological validity. Increasing cell concentration or mycobacterial input and co-culturing in static 48-well plates compared with rotating tubes improved intra-assay repeatability and sensitivity. Standardisation and harmonisation efforts resulted in high consistency agreements, with repeatability and intermediate precision <10% coefficient of variation (CV) and inter-site reproducibility <20% CV; although some systematic differences were observed. As proof-of-concept, we demonstrated ability to detect a BCG vaccine-induced improvement in growth inhibition in macaque samples, and a correlation between MGIA outcome and measures of protection from in vivo disease development following challenge with either intradermal BCG or aerosol/endobronchial Mycobacterium tuberculosis (M.tb) at a group and individual animal level.

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