Vaccines

15 pages, 7976 KiB

Open AccessArticle

Impact of COVID-19 on Immunization Services for Maternal and Infant Vaccines: Results of a Survey Conducted by Imprint—The Immunising Pregnant Women and Infants Network

by Anja Saso, Helen Skirrow and Beate Kampmann

Vaccines 2020, 8(3), 556; https://doi.org/10.3390/vaccines8030556 - 22 Sep 2020

Cited by 63 | Viewed by 13380

Abstract

The COVID-19 pandemic response has caused disruption to healthcare services globally, including to routine immunizations. To understand immunization service interruptions specifically for maternal, neonatal and infant vaccines, we captured the local experiences of members of the Immunising Pregnant Women and Infants Network (IMPRINT) [...] Read more.

The COVID-19 pandemic response has caused disruption to healthcare services globally, including to routine immunizations. To understand immunization service interruptions specifically for maternal, neonatal and infant vaccines, we captured the local experiences of members of the Immunising Pregnant Women and Infants Network (IMPRINT) by conducting an online survey over 2-weeks in April 2020. IMPRINT is a global network of clinicians and scientists working in maternal and neonatal vaccinology. The survey included discrete questions to quantify the extent of disruption as well as free-text options to explore the reasons behind reported disruptions. Of the 48 responses received, the majority (75%) were from low-and-middle-income countries (LMICs). Of all respondents, 50% or more reported issues with vaccine delivery within their country. Thematic analysis identified three key themes behind immunization disruption: “access” issues, e.g., logistical barriers, “provider” issues, e.g., staff shortages and user “concern” about attending immunization appointments due to COVID-19 fear. Access and provider issues were more commonly reported by LMIC respondents. Overall, respondents reported uncertainty among parents and healthcare providers regarding routine immunization. We conclude that further quantification of routine vaccination disruption is needed, alongside health service prioritization, logistical support and targeted communication strategies to reinforce routine immunizations during the COVID-19 response. Full article

(This article belongs to the Special Issue Immune Ontogeny and Vaccination in Early Life)

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18 pages, 1093 KiB

Open AccessReview

Advances in the Development of Anti-Haemonchus contortus Vaccines: Challenges, Opportunities, and Perspectives

by Muhammad Ehsan, Rui-Si Hu, Qin-Li Liang, Jun-Ling Hou, Xiaokai Song, Ruofeng Yan, Xing-Quan Zhu and Xiangrui Li

Vaccines 2020, 8(3), 555; https://doi.org/10.3390/vaccines8030555 - 22 Sep 2020

Cited by 30 | Viewed by 13716

Abstract

The gastrointestinal nematode parasite Haemonchus contortus (H. contortus) is a resident of tropical and subtropical regions worldwide that imposes significant production losses, economic losses, and animal health issues in the small ruminant industry, particularly sheep and goats. Considerable efforts have been [...] Read more.

The gastrointestinal nematode parasite Haemonchus contortus (H. contortus) is a resident of tropical and subtropical regions worldwide that imposes significant production losses, economic losses, and animal health issues in the small ruminant industry, particularly sheep and goats. Considerable efforts have been made to understand how immunity is elicited against H. contortus infection. Various potential vaccine antigens have been tested by different methods and strategies applied in animal models, and significant progress has been made in the development of vaccines against H. contortus. This review highlighted and shared the knowledge about the current understanding of host immune responses to H. contortus and ongoing challenges in the development of a protective, effective, and long-lasting vaccine against H. contortus infection. We have also pinpointed some achievements and failures in the development and testing of vaccines, which will establish a road map for future research directions to explore new effective vaccine candidates for controlling and preventing H. contortus infection. Full article

(This article belongs to the Special Issue Infectious Diseases Immunology)

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25 pages, 1731 KiB

Open AccessReview

Inflammasome-Mediated Immunogenicity of Clinical and Experimental Vaccine Adjuvants

by Sören Reinke, Aneesh Thakur, Cillian Gartlan, Jelena S. Bezbradica and Anita Milicic

Vaccines 2020, 8(3), 554; https://doi.org/10.3390/vaccines8030554 - 22 Sep 2020

Cited by 45 | Viewed by 7210

Abstract

In modern vaccines, adjuvants can be sophisticated immunological tools to promote robust and long-lasting protection against prevalent diseases. However, there is an urgent need to improve immunogenicity of vaccines in order to protect mankind from life-threatening diseases such as AIDS, malaria or, most [...] Read more.

In modern vaccines, adjuvants can be sophisticated immunological tools to promote robust and long-lasting protection against prevalent diseases. However, there is an urgent need to improve immunogenicity of vaccines in order to protect mankind from life-threatening diseases such as AIDS, malaria or, most recently, COVID-19. Therefore, it is important to understand the cellular and molecular mechanisms of action of vaccine adjuvants, which generally trigger the innate immune system to enhance signal transition to adaptive immunity, resulting in pathogen-specific protection. Thus, improved understanding of vaccine adjuvant mechanisms may aid in the design of “intelligent” vaccines to provide robust protection from pathogens. Various commonly used clinical adjuvants, such as aluminium salts, saponins or emulsions, have been identified as activators of inflammasomes - multiprotein signalling platforms that drive activation of inflammatory caspases, resulting in secretion of pro-inflammatory cytokines of the IL-1 family. Importantly, these cytokines affect the cellular and humoral arms of adaptive immunity, which indicates that inflammasomes represent a valuable target of vaccine adjuvants. In this review, we highlight the impact of different inflammasomes on vaccine adjuvant-induced immune responses regarding their mechanisms and immunogenicity. In this context, we focus on clinically relevant adjuvants that have been shown to activate the NLRP3 inflammasome and also present various experimental adjuvants that activate the NLRP3-, NLRC4-, AIM2-, pyrin-, or non-canonical inflammasomes and could have the potential to improve future vaccines. Together, we provide a comprehensive overview on vaccine adjuvants that are known, or suggested, to promote immunogenicity through inflammasome-mediated signalling. Full article

(This article belongs to the Special Issue Immunological Mechanisms of Vaccines and Adjuvants)

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Vaccine adjuvants can activate inflammasomes. Clinically approved (black) and experimental (grey) vaccine adjuvants induce (or possibly induce) assembly and activation of canonical- (NLRP3, NLRC4, AIM2, Pyrin) and non-canonical (caspase-11 in mice, caspase-4/5 in humans) inflammasomes. Canonical inflammasomes recruit caspase-1 via the adaptor molecule ASC, which leads to proximity-induced auto-processing and activation of caspase-1, resulting in cleavage of pro-IL-1β and pro-IL-18 as well as the secretion of their mature forms. Activated caspase-1 also cleaves and activates GSDMD, resulting in pore formation and pyroptosis, which mediates the release of pro-inflammatory DAMPs such as IL-1α or HMGB1. Caspase-11 cleaves GSDMD and induces pyroptosis, but it does not process pro-IL-1β or pro-IL-18. However, non-canonical inflammasomes activate the NLRP3 inflammasome, which indirectly induces the maturation and secretion of IL-1β and IL-18 via the non-canonical route. Abbreviations: apoptosis-associated speck-like protein containing a CARD (ASC), caspase recruitment domain (CARD), pyrin domain (PYD), high-mobility group box 1 (HMGB1), leucine-rich repeat (LRR), domain present in NAIP, CIITA, HET-E, and TP1 (NACHT), gasdermin D (GSDMD), reactive oxygen species (ROS), adenosine triphosphate (ATP), coiled-coil (CC), exon B30.2 domain (B30.2), B-box-type zinc finger domain (BB); immunostimulating complex (ISCOM); ISCOMATRIX (IMX). Created with BioRender.com (Toronto, Canada). Full article ">Figure 2
Caspase-1-dependent cytokines activate adaptive immunity. IL-1α and IL-1β signalling via IL-1R1 activates the pro-inflammatory transcription factor NFκB via the MyD88 pathway. In addition, caspase-1 induces NFκB activation through the cleavage of MAL [<a href="#B102-vaccines-08-00554" class="html-bibr">102</a>]. NFκB activation results in the production of various pro-inflammatory cytokines such as IL-6 or TNF-α, which (I) enhances innate and adaptive immune responses and (II) provides a further priming signal for robust inflammasome-mediated signalling. Cleavage and inactivation of the potent Th2 driver IL-33 by active caspase-1 blocks Th2 polarisation. Mature IL-1α and IL-1β directly affect lymphoid cells by promoting differentiation from naïve T-cells into Th17 cells, T-cell survival, or increased antibody production of B-cells through prolonged T-cell help. After caspase-1-mediated activation, IL-18 binds IL-18R and thereby initiates the production of IFN-γ eventually inducing enhanced differentiation into Th1 type immune cells. Abbreviations: antigen presenting cells (APCs), danger-associated molecular pattern (DAMP), Toll-like receptor (TLR), interleukin 1 receptor 1 (IL-1R1), myeloid differentiation primary response 88 (MyD88), Myd88 adapter-like (MAL), nuclear factor kappa-light-chain-enhancer of activated B-cells (NFκB), nucleotide-binding oligomerisation domain, leucine rich repeat and pyrin domain containing protein 3 (NLRP3). Created with BioRender.com (Toronto, Canada). Full article ">

15 pages, 321 KiB

Open AccessReview

Recent Progress in the Development of Liver Fluke and Blood Fluke Vaccines

by Donald P. McManus

Vaccines 2020, 8(3), 553; https://doi.org/10.3390/vaccines8030553 - 22 Sep 2020

Cited by 38 | Viewed by 6365

Abstract

Liver flukes (Fasciola spp., Opisthorchis spp., Clonorchis sinensis) and blood flukes (Schistosoma spp.) are parasitic helminths causing neglected tropical diseases that result in substantial morbidity afflicting millions globally. Affecting the world’s poorest people, fasciolosis, opisthorchiasis, clonorchiasis and schistosomiasis cause severe [...] Read more.

Liver flukes (Fasciola spp., Opisthorchis spp., Clonorchis sinensis) and blood flukes (Schistosoma spp.) are parasitic helminths causing neglected tropical diseases that result in substantial morbidity afflicting millions globally. Affecting the world’s poorest people, fasciolosis, opisthorchiasis, clonorchiasis and schistosomiasis cause severe disability; hinder growth, productivity and cognitive development; and can end in death. Children are often disproportionately affected. F. hepatica and F. gigantica are also the most important trematode flukes parasitising ruminants and cause substantial economic losses annually. Mass drug administration (MDA) programs for the control of these liver and blood fluke infections are in place in a number of countries but treatment coverage is often low, re-infection rates are high and drug compliance and effectiveness can vary. Furthermore, the spectre of drug resistance is ever-present, so MDA is not effective or sustainable long term. Vaccination would provide an invaluable tool to achieve lasting control leading to elimination. This review summarises the status currently of vaccine development, identifies some of the major scientific targets for progression and briefly discusses future innovations that may provide effective protective immunity against these helminth parasites and the diseases they cause. Full article

(This article belongs to the Special Issue Vaccine Candidates against Tropical Diseases)

15 pages, 5089 KiB

Open AccessArticle

Enhancement of Immune Response and Anti-Infection of Mice by Porcine Antimicrobial Peptides and Interleukin-4/6 Fusion Gene Encapsulated in Chitosan Nanoparticles

by Junjie Peng, Yongle Xiao, Xiaoping Wan, Qian Chen, Huan Wang, Jiangling Li, Jianlin Chen and Rong Gao

Vaccines 2020, 8(3), 552; https://doi.org/10.3390/vaccines8030552 - 21 Sep 2020

Cited by 9 | Viewed by 3536

Abstract

In order to develop a novel and effective immunoregulator to enhance both the immune response and antimicrobial function, a recombinant eukaryotic expression plasmid-pVAX1 co-expressing fusion cathelicidin antimicrobial peptides (CAMPs) and fusion porcine interleukin-4/6 gene (IL-4/6) was constructed and encapsulated in chitosan nanoparticles (CS-VAP4/6), [...] Read more.

In order to develop a novel and effective immunoregulator to enhance both the immune response and antimicrobial function, a recombinant eukaryotic expression plasmid-pVAX1 co-expressing fusion cathelicidin antimicrobial peptides (CAMPs) and fusion porcine interleukin-4/6 gene (IL-4/6) was constructed and encapsulated in chitosan nanoparticles (CS-VAP4/6), prepared by the ionotropic gelation method. Four-week-old female Kunming mice were divided into three groups and intramuscularly injected, respectively, with CS-VAP, CS-VAP4/6, and CS-pVAX1. On 28 days post-inoculation, the mice were challenged by intraperitoneal injection with Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922); IgG, IgG1 and IgG2a, CD4+, and CD8+ T cells increased significantly in the VAP- and VAP4/6- treated mice, detected by ELISA and flow cytometry, correspondingly (p < 0.05). As analyzed by qPCR, expression levels of Toll-like receptor (TLR) 1, TLR4, TLR6, TLR9, IL-1, IL-2, IL-4, IL-6, IL-7, IL-12, IL-15, IL-23, Tumor Necrosis Factor (TNF)-α, and Interferon-gamma (IFN-γ) genes were also significantly up-regulated in comparison with those of the control mice (p < 0.05). Their immunological markers were elevated significantly to different degrees in CS-VAP4/6-treated mice compared with CS-VAP in different days post-inoculation (p < 0.05). After challenge with E. coli and Staphylococcus aureus, most of the VAP- and VAP4/6- treated mice survived, and no symptoms of bacterial infection were observed. In contrast, 80% of control mice died of infection. Among the treated groups, VAP4/6 had a stronger resistance against challenge with E. coli infection. These results demonstrated that the fusion gene of antimicrobial peptide and interleukin-4/6 has the promising potential as a safe and effective immunomodulator for the control of bacterial infections. Full article

(This article belongs to the Special Issue Vaccines against Antimicrobial-Resistant Infections)

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16 pages, 4970 KiB

Open AccessArticle

Solid Lipid Nanoparticle Carrier Platform Containing Synthetic TLR4 Agonist Mediates Non-Viral DNA Vaccine Delivery

by Jasmine E. Francis, Ivana Skakic, Chaitali Dekiwadia, Ravi Shukla, Aya C. Taki, Anna Walduck and Peter M. Smooker

Vaccines 2020, 8(3), 551; https://doi.org/10.3390/vaccines8030551 - 21 Sep 2020

Cited by 28 | Viewed by 5682

Abstract

There is a growing demand for better delivery systems to improve the stability and efficacy of DNA vaccines. Here we report the synthesis of a non-viral DNA vaccine delivery system using a novel adjuvanted solid lipid nanoparticle (SLN-A) platform as a carrier for [...] Read more.

There is a growing demand for better delivery systems to improve the stability and efficacy of DNA vaccines. Here we report the synthesis of a non-viral DNA vaccine delivery system using a novel adjuvanted solid lipid nanoparticle (SLN-A) platform as a carrier for a DNA vaccine candidate encoding the Urease alpha (UreA) antigen from Helicobacter pylori. Cationic SLN-A particles containing monophosphoryl lipid A (adjuvant) were synthesised by a modified solvent-emulsification method and were investigated for their morphology, zeta potential and in vitro transfection capacity. Particles were found to bind plasmid DNA to form lipoplexes, which were characterised by electron microscopy, dynamic light scattering and fluorescence microscopy. Cellular uptake studies confirmed particle uptake within 3 h, and intracellular localisation within endosomal compartments. In vitro studies further confirmed the ability of SLN-A particles to stimulate expression of pro-inflammatory cytokine tumor necrosis factor alpha (TNF-α) in human macrophage-like Tohoku Hospital Pediatrics-1 (THP-1) cells. Lipoplexes were found to be biocompatible and could be efficiently transfected in murine immune cells for expression of recombinant H. pylori antigen Urease A, demonstrating their potential as a DNA vaccine delivery system. Full article

(This article belongs to the Special Issue Vaccines for Infectious and Chronic Diseases)

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TEM micrographs under 100k accelerating voltage of adjuvanted solid lipid nanoparticle (SLN-A) nanoparticles at (a) 50 k magnification and (b) 20 k magnification; (c) a schematic representation of SLN-A particles binding DNA to form lipoplex-A; (d) lipoplex-A imaged at 50 k magnification and (e) 20 k magnification. SLN-A nanoparticles were found to be heterogenous in size and cuboid in shape with an average diameter of 75.63 ± 31.99 nm, while lipoplex-A complexes were heterogenous in size with undefined edges and had an average diameter of 74.67 ± 39.93 nm. Full article ">Figure 2
Characterisation by dynamic light scattering (DLS) of stability of SLN-A (a) particle size; (b) zeta potential at −20 °C, 4 °C or 22 °C for up to 4 weeks. The effect of temperature on particle size and zeta potential was determined not to be statistically significant by two-way ANOVA (F(2,6) = 4.686, p = 0.0595). However, the effect of storage time was determined to be statistically significant for both particle size (F(2.163, 12.98) = 19.02, p = 0.0001) and zeta potential (F(1.679, 10.07) = 4.780, p = 0.0393). Full article ">Figure 3
Agarose gel electrophoresis of a range of DNA:SLN-A weight ratios. (a) DNA binding assay: Lane 1—Hyperladder I, lane 2—plasmid DNA only, lane 3: SLN-A only, lane 4–8: 1:1, 1:5, 1:10, 1:50, 1:100 DNA:SLN weight ratio lipoplexes; (b) DNase I protection assay: Lane 1—Hyperladder I, lane 2—digested plasmid DNA, lane 3: SLN-A only, lane 4–8: 1:1, 1:5, 1:10, 1:50, 1:100 DNA:SLN weight ratio lipoplexes treated with DNase I and subsequently released from particles by SDS treatment. Full article ">Figure 4
Expression of tumor necrosis factor alpha (TNF-α) by THP-1 cells stimulated with SLN-A particles. One-way ANOVA confirmed that a significant difference in TNF-α expression was observed between cells treated with media and cells treated with SLN-A (F(2,6) = 39.35, p = 0.0009). *** denotes p ≤ 0.001. Full article ">Figure 5
Viability of human embryonic kidney (HEK)293T cells treated with (a) SLN-A nanoparticles; (b) lipoplex-A (SLN-A with loaded plasmid DNA) for 24, 48 or 72 h. Cell viability increased across both treatments in a dose dependent manner across all three timepoints. Cell viability was lowest for cells treated with lipoplex-A for 72 h. Two-way ANOVA confirmed that a significant difference in cell viability was observed between timepoints for cells treated with both SLN-A (F(1.279, 15.35) = 21.24, p = 0.0002) and lipoplex-A (F(1.222, 14.66) = 32.56, p = < 0.0001). *** denotes p ≤ 0.001, and **** denotes p ≤ 0.0001. Full article ">Figure 6
TEM micrographs of SLN-A nanoparticle uptake by DC2.4 cells. (a) Cells treated with SLN-A nanoparticles; (b) Control cells without nanoparticle treatment. Nanoparticle uptake was evident within cells treated with SLN-A nanoparticles (image a), with aggregation in endosomes (inset). No nanoparticle uptake was seen in the control cell sample (image b), where empty endosomes were observed (inset). Scale bar 2 µm. Full article ">Figure 7
Fluorescence micrographs of DC2.4 cells internalising SLN-A nanoparticles within 24 h. Cell nuclei counterstained with DAPI (blue) and SLN-A nanoparticles (red) are displayed. The speed of cell uptake of SLN-A nanoparticles by DC2.4 cells was visualised at 0, 1, 3, 6, 12 and 24 h timepoints. Particles were internalised from 3 h and were no longer visible within the cell by 24 h. Scale bar 100 µm. Full article ">Figure 8
Fluorescence micrograph of DAPI-stained DC2.4 cells (blue; nuclei) expressing Urease alpha (UreA) labelled with AlexaFluor 488 conjugated anti-His monoclonal antibody (green). Cells were transfected with lipoplex-A (SLN-A nanoparticles loaded with pcDNA3.1-UreA) and were found to be expressing UreA protein indicated by the binding of anti-6XHis antibody. Scale bar 20 µm. Full article ">

31 pages, 1494 KiB

Open AccessReview

The Role of Birds of Prey in West Nile Virus Epidemiology

by Beatriz Vidaña, Núria Busquets, Sebastian Napp, Elisa Pérez-Ramírez, Miguel Ángel Jiménez-Clavero and Nicholas Johnson

Vaccines 2020, 8(3), 550; https://doi.org/10.3390/vaccines8030550 - 21 Sep 2020

Cited by 47 | Viewed by 7336

Abstract

Reported human cases of West Nile virus (WNV) in Europe increased dramatically in 2018. Lineage 1 strains had been circulating in Euro-Mediterranean countries since the early 1990s. The subsequent introduction of WNV lineage 2 has been responsible for the remarkable upsurge of European [...] Read more.

Reported human cases of West Nile virus (WNV) in Europe increased dramatically in 2018. Lineage 1 strains had been circulating in Euro-Mediterranean countries since the early 1990s. The subsequent introduction of WNV lineage 2 has been responsible for the remarkable upsurge of European WNV outbreaks since 2004, including the dramatic increase in human cases observed since 2018. The virus exists in a natural cycle between mosquitoes and wild birds, with humans and horses acting as dead-end hosts. As the key vertebrate hosts in the transmission cycle of WNV, avian species have been the focus of surveillance across many countries. Raptors appear particularly susceptible to WNV infection, resulting in higher prevalence, and in some cases exhibiting neurological signs that lead to the death of the animal. In addition, birds of prey are known to play an important role as WNV reservoir and potentially amplifying hosts of infection. Importantly, raptor higher susceptibility/prevalence may indicate infection through predation of infected prey. Consequently, they are considered important target species when designing cost-effective surveillance for monitoring both seasonal WNV circulation in endemic countries and its emergence into new areas, where migrating raptors may play a critical role in virus introduction. This review summarizes the different aspects of the current knowledge of WNV infection in birds of prey and evaluates their role in the evolution of the epizootic that is spreading throughout Europe. Full article

(This article belongs to the Special Issue West Nile Virus Disease)

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18 pages, 1461 KiB

Open AccessArticle

African Swine Fever Circulation among Free-Ranging Pigs in Sardinia: Data from the Eradication Program

by Giulia Franzoni, Silvia Dei Giudici, Federica Loi, Daria Sanna, Matteo Floris, Mariangela Fiori, Maria Luisa Sanna, Paola Madrau, Fabio Scarpa, Susanna Zinellu, Monica Giammarioli, Stefano Cappai, Gian Mario De Mia, Alberto Laddomada, Sandro Rolesu and Annalisa Oggiano

Vaccines 2020, 8(3), 549; https://doi.org/10.3390/vaccines8030549 - 21 Sep 2020

Cited by 27 | Viewed by 5330

Abstract

African swine fever virus (ASFV), the cause of a devastating disease affecting domestic and wild pigs, has been present in Sardinia since 1978. In the framework of the regional ASF eradication plan, 4484 illegal pigs were culled between December 2017 and February 2020. [...] Read more.

African swine fever virus (ASFV), the cause of a devastating disease affecting domestic and wild pigs, has been present in Sardinia since 1978. In the framework of the regional ASF eradication plan, 4484 illegal pigs were culled between December 2017 and February 2020. The highest disease prevalence was observed in the municipality with the highest free-ranging pig density, and culling actions drastically reduced ASFV circulation among these animals. ASFV-antibody were detected in 36.7% of tested animals, which were apparently healthy, thus, the circulation of low-virulence ASFV isolates was hypothesized. ASFV genome was detected in 53 out of 2726 tested animals, and virus isolation was achieved in two distinct culling actions. Two ASFV haemadsorbing strains were isolated from antibody-positive apparently healthy pigs: 55234/18 and 103917/18. Typing analysis revealed that both isolates belong to p72 genotype I, B602L subgroup X; phylogenetic analysis based on whole genome sequencing data showed that they were closely related to Sardinian ASFV strains collected since 2010, especially 22653/Ca/2014. Our data suggested the absence of immune-escaped ASFV variants circulating among free-ranging pigs, indicating that other elements contributed to virus circulation among these animals. Understanding factors behind disease persistence in endemic settings might contribute to developing effective countermeasures against this disease. Full article

(This article belongs to the Special Issue African Swine Fever Virus Prevention and Control)

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Map representing the Sardinian municipalities of the free-ranging pig culling actions (blue and red dots). The 55234/18 and the 103917/18 were isolated from samples collected in Desulo and Talana (red dots), both inside the wild boar infected zone (red line). Full article ">Figure 2
Growth kinetic in macrophages of the two ASFV strains collected from illegal free-ranging pigs. moMΦ were infected with the strains under study (103917/14 or 55234/18) or the virulent 26544/OG10 ASFV strains using a multiplicity of infection (MOI) of 0.01. At 0, 24, 48, and 72 h pi, duplicate samples were collected, and infectious viral progeny in culture supernatants were assessed by titration (TCID50/mL). The sensitivity of this assay was 1.8 log10 50% haemadsorbing doses/mL (TCID50/mL). The mean values and the standard deviation from three independent experiments utilizing different animals are shown. At each time-point, values of 103917/14 and 55234/18 and 26544/OG10 viral titres were compared using the analysis of variance (ANOVA) for repeated measures. Full article ">Figure 3
Effect of 103917/14 and 55234/18 ASFV infection on MHC class I expression and viability of macrophages. moMΦ were infected with the low-virulence NH/P68, the virulent 26544/OG10, or the isolates under study 103917/18 and 55234/18 using an MOI of 1, alongside mock-infected controls. At 21 hpi, surface expression of MHC class I and moMФ viability were assessed. (A) MHC class I expression (MFI) and intracellular levels of ASFV p72 were assessed by flow cytometry; MFI data are presented as fold change relative to the mock-infected condition. Values of ASFV-infected or bystander macrophages were compared to the corresponding mock-infected control. (B) moMФ viability was assessed using a non-radioactive cytotoxic assay, which quantifies lactate dehydrogenase (LDH) levels in culture supernatants. For both panels, mean values and standard deviation from three (A) or four (B) independent experiments utilizing different animals are shown as green and red bars with whiskers lines, respectively. Analysis of variance and relative Bonferroni method for multiple comparisons were applied to evaluate difference between samples; *** p < 0.001, * p < 0.05. Full article ">Figure 4
Bayesian phylogenetic tree based on ASFV complete genome sequences. The main nodes of the tree are fully supported by values of posterior probabilities = 1. The clade, including the 16 Sardinian ASFV genomes, was collapsed in the main tree and indicated with red font. In the inset are the phylogenetic relationships within the Sardinian group. The sequences isolated in the present study are indicated in red font. Full article ">

23 pages, 4057 KiB

Open AccessReview

Influenza–Host Interplay and Strategies for Universal Vaccine Development

by Hye Suk Hwang, Mincheol Chang and Yoong Ahm Kim

Vaccines 2020, 8(3), 548; https://doi.org/10.3390/vaccines8030548 - 20 Sep 2020

Cited by 8 | Viewed by 8261

Abstract

Influenza is an annual epidemic and an occasional pandemic caused by pathogens that are responsible for infectious respiratory disease. Humans are highly susceptible to the infection mediated by influenza A viruses (IAV). The entry of the virus is mediated by the influenza virus [...] Read more.

Influenza is an annual epidemic and an occasional pandemic caused by pathogens that are responsible for infectious respiratory disease. Humans are highly susceptible to the infection mediated by influenza A viruses (IAV). The entry of the virus is mediated by the influenza virus hemagglutinin (HA) glycoprotein that binds to the cellular sialic acid receptors and facilitates the fusion of the viral membrane with the endosomal membrane. During IAV infection, virus-derived pathogen-associated molecular patterns (PAMPs) are recognized by host intracellular specific sensors including toll-like receptors (TLRs), C-type lectin receptors, retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs), and nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) either on the cell surface or intracellularly in endosomes. Herein, we comprehensively review the current knowledge available on the entry of the influenza virus into host cells and the molecular details of the influenza virus–host interface. We also highlight certain strategies for the development of universal influenza vaccines. Full article

(This article belongs to the Special Issue Infectious Diseases Immunology)

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Influenza A virus endocytosis and hemagglutinin proteins (HA) conformational change. (A) Process of the entry of influenza virus into host cell. The virus binds to sialic acid-containing proteins on the cell surface receptors by association with the viral hemagglutinin proteins (HA1, HA2). HAs also bind to the sialic acid-containing Ca2+ channel to trigger intracellular Ca2+ oscillations. The virus is then internalized by endocytosis. Acidification of the endosome causes a conformational change in the HA proteins that leads to a fusion between the viral membrane and the endosomal membrane. This allows the escape of the viral RNA and proteins into the cytoplasm. (B) Structure of the HA of IAV. The trimeric complex of HA is shown with one monomer highlighted in color (HA1; red, HA2; blue, and the receptor binding pocket; green). (C) The pre- and post-fusion conformations of HA [<a href="#B4-vaccines-08-00548" class="html-bibr">4</a>]. This figure was created using BioRender (Toronto, ON, Canada). Full article ">Figure 2
The intracellular cytoplasmic pattern-recognition receptor RIG-I is essential for the control of RNA virus infection. Upon IAV recognition, RIG-I recruits the adaptor MAVS protein to activate the IKKα–IKKβ and TBK1–IKKϵ complexes, which are responsible for the activation of the IRF 3 and IRF7 transcription factors. These transcription factors then translocate into the nucleus and cooperatively induce IRF dependent type I IFNs and NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) dependent pro-inflammatory cytokines and chemokines. This is followed by the binding of the IFNAR1 and IFNAR2 to their cognate receptor, which leads to the transcriptional activation of ISGs by the JAK/STAT signaling pathway. The products of ISGs are key factors limiting pathogen spreading. Moreover, ssRNA from IAVs can prime the inflammasome by activating a TLR inducing NF-κB activation and the expression of NLRP3, ASC, and preforms of IL-18 and IL-1β. A second activation signal is provided by the oligomerization of the NLRP3 complex and recruitment of ASC and procaspase-1, allowing the processing and cleavage of pro-IL-1β and pro-IL-18 precursors into their bioactive mature forms (IL-18 and IL-1β). NLRP3 can be activated by imbalances in potassium ion concentration in intracellular vesicles through the ATP-gated P2 × 7 channel and responses of mitochondrial reactive oxygen species. This figure was created using BioRender. RIG-1;retinoic acid-inducible gene-1, MAVS; mitochondrial antiviral signaling adaptor, IKKα; nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor (IκB) kinase α, IKKβ; IκB kinase β, IKKϵ; IκB kinase ϵ, TBK1; TRAF family member-associated NF-kappa-B activator (TANK)-binding kinase 1, IRF; interferon-regulatory factors, IFNs; Interferons, NF-κB; Nuclear Factor kappa-light-chain-enhancer of activated B cells, IFNAR1; Interferon Alpha And Beta Receptor Subunit 1, IFNAR2; Interferon Alpha And Beta Receptor Subunit 2, ISGs; interferon-stimulated gene, JAK/STAT; Janus kinase (JAK)/signal transducer and activator of transcription (STAT), ssRNA; single stranded RNA, TLR; Toll like receptor, NLRP3; nucleoside oligomerization domain (NOD), leucine-rich repeat (LRR), and pyrin domain (PYD) domain-containing protein 3, ASC; Apoptosis-associated speck-like protein containing a CARD, IL-18; Interleukin 18, IL-1β; Interleukin 1β, ATP-gated P2X7; Adenosine triphosphate (ATP)-gated purinergic P2X7 receptor. Full article ">Figure 3
A schematic model showing the balance between successful viral clearance and a life-threatening immunopathology following influenza infection. (A) The excessive response to influenza infection results in the development of influenza immunopathology despite efficient viral clearance. The excessive inflammation sustained by an uncontrolled host response can induce epithelial disruption and lung damage. (B) Low immune response with immune escape from host immunosurveillance may increase viral replication, which in turn induces a strong release of secretory molecules. (C) The adequate cell mediated immunity with vaccination can control lung viral load without a severe lung pathology. This figure was created using the BioRender software. Full article ">Figure 4
Universal IAV vaccination approaches. (A) Chimeric hemagglutinins (cHAs) consist of the exotic globular head domains and the conserved H2 stalk domain. (B) Mixture of virus like particles (VLPs) that express multiple subtypes of HA (C) Combination approaches with Matrix protein 1 (M1) and nucleoprotein (NP) with virus vectors or DNA vectors. (D) Vaccination strategies based on conserved M2 ectodomain (M2e). Full article ">

11 pages, 235 KiB

Open AccessArticle

Antibody Response to Canine Parvovirus Vaccination in Dogs with Hyperadrenocorticism Treated with Trilostane

by Michèle Bergmann, Monika Freisl, Katrin Hartmann, Stephanie Speck, Uwe Truyen, Yury Zablotski, Matthias Mayr and Astrid Wehner

Vaccines 2020, 8(3), 547; https://doi.org/10.3390/vaccines8030547 - 19 Sep 2020

Cited by 9 | Viewed by 4697

Abstract

It is unknown how dogs with hyperadrenocorticism (HAC) respond to vaccination. This study measured antibodies against canine parvovirus (CPV) in dogs with HAC treated with trilostane before and after CPV vaccination, and compared the immune response to that from healthy dogs. Eleven dogs [...] Read more.

It is unknown how dogs with hyperadrenocorticism (HAC) respond to vaccination. This study measured antibodies against canine parvovirus (CPV) in dogs with HAC treated with trilostane before and after CPV vaccination, and compared the immune response to that from healthy dogs. Eleven dogs with HAC, and healthy age-matched control dogs (n = 31) received a modified-live CPV vaccine. Antibodies were determined on days 0, 7, and 28 by hemagglutination inhibition. Univariate analysis was used to compare the immune response of dogs with HAC and healthy dogs. Pre-vaccination antibodies (≥10) were detected in 100% of dogs with HAC (11/11; 95% CI: 70.0–100) and in 93.5% of healthy dogs (29/31; 95% CI: 78.3–99.2). No ≥4-fold increase in antibody titer was observed in dogs with HAC while in 22.6% of healthy dogs, a ≥4-fold titer increase was observed (7/31; 95% CI: 11.1–40.1). Mild vaccine-associated adverse events (VAAEs) were detected in 54.5% of dogs with HAC (6/11; 95% CI: 28.0–78.8) and in 29.0% of healthy dogs (9/31; 95% CI: 15.9–46.8). There was neither a significant difference in presence of pre-vaccination antibodies (p = 1.000), or response to vaccination (p = 0.161), nor in the occurrence of VAAEs (p = 0.158). Immune function of dogs with HAC treated with trilostane seems comparable to that of healthy dogs. Full article

(This article belongs to the Special Issue Infectious Diseases Immunology)

21 pages, 1343 KiB

Open AccessReview

The Power of First Impressions: Can Influenza Imprinting during Infancy Inform Vaccine Design?

by Melissa Rioux, Mara McNeil, Magen E. Francis, Nicholas Dawe, Mary Foley, Joanne M. Langley and Alyson A. Kelvin

Vaccines 2020, 8(3), 546; https://doi.org/10.3390/vaccines8030546 - 19 Sep 2020

Cited by 9 | Viewed by 7971

Abstract

Influenza virus infection causes severe respiratory illness in people worldwide, disproportionately affecting infants. The immature respiratory tract coupled with the developing immune system, and lack of previous exposure to the virus is thought to synergistically play a role in the increased disease severity [...] Read more.

Influenza virus infection causes severe respiratory illness in people worldwide, disproportionately affecting infants. The immature respiratory tract coupled with the developing immune system, and lack of previous exposure to the virus is thought to synergistically play a role in the increased disease severity in younger age groups. No influenza vaccines are available for those under six months, although maternal influenza immunization is recommended. In children aged six months to two years, vaccine immunogenicity is dampened compared to older children and adults. Unlike older children and adults, the infant immune system has fewer antigen-presenting cells and soluble immune factors. Paradoxically, we know that a person’s first infection with the influenza virus during infancy or childhood leads to the establishment of life-long immunity toward that particular virus strain. This is called influenza imprinting. We contend that by understanding the influenza imprinting event in the context of the infant immune system, we will be able to design more effective influenza vaccines for both infants and adults. Working through the lens of imprinting, using infant influenza animal models such as mice and ferrets which have proven useful for infant immunity studies, we will gain a better understanding of imprinting and its implications regarding vaccine design. This review examines literature regarding infant immune and respiratory development, current vaccine strategies, and highlights the importance of research into the imprinting event in infant animal models to develop more effective and protective vaccines for all including young children. Full article

(This article belongs to the Special Issue Vaccinology of Influenza Infection)

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Figure 1

Figure 1
Trends in early-life immune development and influenza virus-associated mortality. As immune and respiratory function improve over the first few years, influenza-associated mortality decreases. Maternal immunoglobulin G (IgG) is high in full term babies but then decreases rapidly over the first few months of life. At birth, the immune system is biased toward T helper cell type 2 responses, but maturation over the first 2–3 years of life is characterized by increased T helper cell type 1 responses and increased antibody production allowing a balanced response to antigens. Full article ">Figure 2
Schematic of influenza A, B, C and D virus structure. Influenza A and B viruses express surface glycoproteins hemagglutinin (HA) and neuraminidase (NA), as well as the M2 ion channel. Both A and B viruses have 8 genomic segments coding for at least 10 proteins. Influenza C and D viruses express the surface glycoprotein hemagglutinin-esterase fusion (HEF), as well as the M2 ion channel. Both C and D viruses have 7 genomic segments coding for 9 proteins. All four types of influenza viruses express the M1 protein along the inner surface on the envelope, adjacent to the nuclear export protein (NEP). Full article ">Figure 3
Differences in antigen presenting cell (APC) function between infants and adults. Despite similar pattern recognition receptor (PRR) expression between infant and adult APCs, the downstream signaling following antigen recognition by PRRs differs. Reduced adaptor protein function (e.g., IRF-4) in neonatal APCs contributes to tolerance of self-antigens and commensal microbes, while also reducing responsiveness to pathogens and vaccine antigens, decreased inflammatory cytokine production, and skewing toward a T helper cell type 2 response. Full article ">

6 pages, 208 KiB

Open AccessEditorial

Vaccination and Vaccine Effectiveness: A Commentary of Special Issue Editors

by Claudio Costantino, Alessandra Casuccio and Vincenzo Restivo

Vaccines 2020, 8(3), 545; https://doi.org/10.3390/vaccines8030545 - 18 Sep 2020

Cited by 10 | Viewed by 4178

Abstract

The Special Issue “Vaccination and Vaccine Effectiveness”, published in the journal Vaccines, has the main aim to increase international literature data on vaccine effectiveness and safety and on vaccination strategies in order to reduce vaccine hesitancy and improve vaccination coverage rates. The [...] Read more.

The Special Issue “Vaccination and Vaccine Effectiveness”, published in the journal Vaccines, has the main aim to increase international literature data on vaccine effectiveness and safety and on vaccination strategies in order to reduce vaccine hesitancy and improve vaccination coverage rates. The main topics included in the call for papers were vaccines administered to infants, adolescents, adults, elderly people, at-risk populations (due to comorbidities and personal risk factors) and healthcare workers and strategies adopted to promote vaccination adherence among these categories. This Special Issue started from the assumption that, despite vaccination being universally recognized as one of the best strategies to increase duration and quality of life during the last centuries, vaccination coverage rates are often under the levels recommended to reduce circulation and to extinguish vaccine-preventable diseases. Vaccine hesitancy involves at least 15% of the general population, and healthcare workers also sometimes demonstrate doubts on vaccination effectiveness and safety. At the end of the six-month submission period, 16 articles (15 research article and one review) were accepted after the peer-review processes and published online. Full article

(This article belongs to the Special Issue Vaccination and Vaccine Effectiveness)

6 pages, 214 KiB

Open AccessCase Report

Pneumocystis jirevocii and SARS-CoV-2 Co-Infection: A Common Feature in Transplant Recipients?

by Maria A. De Francesco, Federico Alberici, Nicola Bossini, Francesco Scolari, Federico Pascucci, Gabriele Tomasoni and Arnaldo Caruso

Vaccines 2020, 8(3), 544; https://doi.org/10.3390/vaccines8030544 - 18 Sep 2020

Cited by 24 | Viewed by 3375

Abstract

COVID-19 might potentially give rise to a more severe infection in solid organ transplant recipients due to their chronic immunosuppression. These patients are at a higher risk of developing concurrent or secondary bacterial and fungal infections. Co-infections can increase systemic inflammation influencing the [...] Read more.

COVID-19 might potentially give rise to a more severe infection in solid organ transplant recipients due to their chronic immunosuppression. These patients are at a higher risk of developing concurrent or secondary bacterial and fungal infections. Co-infections can increase systemic inflammation influencing the prognosis and the severity of the disease, and can in turn lead to an increased need of mechanical ventilation, antibiotic therapy and to a higher mortality. Here we describe, for the first time in Europe, a fatal case of co-infection between SARS-CoV-2 and Pneumocystis jirevocii in a kidney transplant recipient. Full article

(This article belongs to the Special Issue Infectious Diseases Immunology)

11 pages, 235 KiB

Open AccessArticle

Meningococcal Disease and Related Vaccinations: Knowledge, Attitudes, and Practices among Healthcare Workers Who Provide Care to Patients with Underlying High-Risk Medical Conditions

by Gabriella Di Giuseppe, Concetta P. Pelullo, Giorgia Della Polla and Maria Pavia

Vaccines 2020, 8(3), 543; https://doi.org/10.3390/vaccines8030543 - 18 Sep 2020

Cited by 12 | Viewed by 3342

Abstract

This cross-sectional study assessed knowledge, attitudes, and practices regarding meningococcal disease and related vaccinations among healthcare workers (HCWs) who provided care to patients with underlying high-risk medical conditions. A total of 411 HCWs returned the survey. Only 35% of the respondents had a [...] Read more.

This cross-sectional study assessed knowledge, attitudes, and practices regarding meningococcal disease and related vaccinations among healthcare workers (HCWs) who provided care to patients with underlying high-risk medical conditions. A total of 411 HCWs returned the survey. Only 35% of the respondents had a good knowledge about the incidence and lethality of meningococcal disease, the most frequent serogroups in Italy and the diseases or conditions that expose patients to a high-risk of severe complications caused by meningococcal disease. Vaccination against meningococcal disease was perceived to be highly effective by 38.4% of participants, very safe by 36.2%, and 82% agreed or strongly agreed that HCWs should promote adherence to recommended vaccinations even in hesitant patients. Moreover, 34.1% recommended meningococcal vaccinations to all eligible patients and the results of the multivariate analysis showed that older HCWs, who work in pediatric/neonatal wards, have good knowledge about meningococcal vaccinations, have a favourable attitude towards vaccinations, and do not need additional information about meningococcal vaccinations, were more likely to recommend meningococcal vaccinations to all eligible patients. Interventions aimed at the enhancement of knowledge and awareness of HCWs who provide care to these patients on the benefits of meningococcal vaccinations are warranted. Full article

(This article belongs to the Special Issue Vaccination and Public Health: Optimizing Vaccine Uptake through the Application of Social and Behavioral Science Theory, Principles, and Strategies)

20 pages, 4998 KiB

Open AccessArticle

Control of Cytoskeletal Dynamics by β-Arrestin1/Myosin Vb Signaling Regulates Endosomal Sorting and Scavenging Activity of the Atypical Chemokine Receptor ACKR2

by Alessandro Vacchini, Cinzia Cancellieri, Samantha Milanesi, Sabrina Badanai, Benedetta Savino, Francesco Bifari, Massimo Locati, Raffaella Bonecchi and Elena Monica Borroni

Vaccines 2020, 8(3), 542; https://doi.org/10.3390/vaccines8030542 - 17 Sep 2020

Cited by 9 | Viewed by 3542

Abstract

The atypical chemokine receptor ACKR2, formerly named D6, is a scavenger chemokine receptor with a non-redundant role in the control of inflammation and immunity. The scavenging activity of ACKR2 depends on its trafficking properties, which require actin cytoskeleton rearrangements downstream of a β-arrestin1-Rac1-PAK1-LIMK1-cofilin-dependent [...] Read more.

The atypical chemokine receptor ACKR2, formerly named D6, is a scavenger chemokine receptor with a non-redundant role in the control of inflammation and immunity. The scavenging activity of ACKR2 depends on its trafficking properties, which require actin cytoskeleton rearrangements downstream of a β-arrestin1-Rac1-PAK1-LIMK1-cofilin-dependent signaling pathway. We here demonstrate that in basal conditions, ACKR2 trafficking properties require intact actin and microtubules networks. The dynamic turnover of actin filaments is required to sustain ACKR2 constitutive endocytosis, while both actin and microtubule networks are involved in processes regulating ACKR2 constitutive sorting to rapid, Rab4-dependent and slow, Rab11-dependent recycling pathways, respectively. After chemokine engagement, ACKR2 requires myosin Vb activity to promote its trafficking from Rab11-positive recycling endosomes to the plasma membrane, which sustains its scavenging activity. Other than cofilin phosphorylation, induction of the β-arrestin1-dependent signaling pathway by ACKR2 agonists also leads to the rearrangement of microtubules, which is required to support the myosin Vb-dependent ACKR2 upregulation and its scavenging properties. Disruption of the actin-based cytoskeleton by the apoptosis-inducing agent staurosporine results in impaired ACKR2 internalization and chemokine degradation that is consistent with the emerging scavenging-independent activity of the receptor in apoptotic neutrophils instrumental for promoting efficient efferocytosis during the resolution of inflammation. In conclusion, we provide evidence that ACKR2 activates a β-arrestin1-dependent signaling pathway, triggering both the actin and the microtubule cytoskeletal networks, which control its trafficking and scavenger properties. Full article

(This article belongs to the Special Issue Investigating Gametogenesis and Cancer for Discovering New Immunotherapeutic Targets)

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Figure 1

Figure 1
Atypical Chemokine Receptor 2 (ACKR2) constitutive cycling is regulated by cytoskeletal dynamics. (A,B) Confocal microscopy analysis of Chinese Hamster Ovary (CHO)-K1/ACKR2 cells incubated (30 min) with indicated inhibitors of (A) actin and (B) microtubule dynamics. Top panels show nuclear staining (DAPI, blue) merged with double staining of ACKR2 (red) and actin (phalloidin staining in panel A; green) or microtubules (α-tubulin staining in panel B; green). Bottom panels show colocalization channels of ACKR2 with actin filaments or microtubules, with mean ± SEM of Pearsons’ Correlation Coefficient (PCC) evaluated in at least n = 20 cells. Results are normalized over vehicle-treated cells. (C–F) CHO-K1/ACKR2 cells incubated (30 min) with vehicle (DMSO), 1 µM cytochalasin D (CD), 1 µM latrunculin A (LA), 1 µM jasplakinolide (J), 10 µM nocodazole (N), and 1 µM paclitaxel (P) and analyzed for ACKR2 membrane expression (A–C,E) and ACKR2 constitutive internalization after indicated time points (D,F). Results are normalized over vehicle-treated cells (panel C and E) or over 30 min-treated cells with vehicle or inhibitors (panel D and F), and shown as mean ± SEM of at least n = 3 experiments. In panels D and F, data were analyzed by two-way ANOVA with Tukey’s post hoc test and only statistical analysis of inhibitor versus corresponding vehicle is shown. *: p ≤ 0.05, **: p ≤ 0.01, ***: p ≤ 0.005. Full article ">Figure 2
Alteration of cytoskeletal dynamics causes ACKR2 missorting into recycling compartments. (A,B) Flow cytometry analysis of ACKR2 membrane expression in CHO-K1/ACKR2 cells transiently transfected with the indicated pEGFP-tagged plasmids after incubation (1 h) with vehicle or 1 µM latrunculin A (panel A,) and 10 µM nocodazole (panel B). Transfected but pEGFP-negative cells (pEGFPneg) served as internal control of cells transfected and overexpressing dominant negative Rab4 (Rab4 S22N) and Rab11 (Rab11 S25N) pEGFP plasmids (pEGFPpos). Results are representative of mean ± SEM of n = 3 experiments and are shown as percentage of MFI of pEGFPneg/pEGFPpos cells. (C,D) Confocal microscopy analysis of CHO-K1/ACKR2 cells incubated (1 h) with vehicle (DMSO), 1 µM latrunculin A (panel C), or 10 µM nocodazole (panel D). Left panels show ACKR2 staining (red) merged in panel C with Rab4 (blue) and actin (phalloidin staining in green), and in panel D with Rab11 (blue) and microtubules (α-tubulin staining in green). Panels on the right show Nomarski interference contrast merged with the colocalization channel. Quantification of ACKR2 colocalization with Rab4 or Rab11 is shown as the mean ± SEM of PCC evaluated in at least n = 20 cells. Data were analyzed by two-way ANOVA with Tukey’s post hoc test (panel A and B), or unpaired Student’s t-test (panel C and D). *: p ≤ 0.05, ***: p ≤ 0.005 vehicle versus inhibitor-treated cells, ###: p ≤ 0.005 pEGFPpos versus pEGFPneg cells. Full article ">Figure 3
The chemokine scavenger function of ACKR2 requires ligand-induced microtubule rearrangement. (A,B) Confocal microscopy analysis of CHO-K1/ACKR2 (A) and CHO-K1/CCR5 cells (B) stimulated with 100 nM CCL3L1 (1 h) and double stained for ACKR2 or CCR5 (red) and microtubule networks (α-tubulin staining; green). Nuclear staining (DAPI) is in blue. Merged images are shown in left panels, right panels show colocalization channels, with quantification of ACKR2 or CCR5 colocalization with microtubules expressed as mean ± SEM of PCC evaluated in at least n = 20 cells. (C) Confocal microscopy analysis of CHO-K1/ACKR2 upon stimulation with indicated chemokines (100 nM, 1 h). Colors for staining are as in panel A. Colocalization channels are enclosed in the corresponding panels. (D) Quantification of ACKR2 colocalization with microtubules in response to treatment of the indicated chemokines. Colocalization results are expressed as mean ± SEM of PCC evaluated in at least n = 20 cells. (E) Flow cytometry analysis of ACKR2 membrane expression in CHO-K1/ACKR2 cells pre-treated (30 min) with vehicle, 10 µM nocodazole (N), or 1 µM paclitaxel (P) and stimulated with 100 nM CCL3L1 (1 h). (F) Effect of microtubule inhibitors (10 µM nocodazole; 1 µM paclitaxel) on ACKR2 scavenging activity in CHO-K1/ACKR2 cells incubated with 0.1 nM 125I-CCL2 and indicated concentrations of CCL2. Results are shown as mean ± SEM of at least n = 3 experiments, whereas in panel F of n = 2 experiments for paclitaxel treatment. Data were analyzed by unpaired Student’s t-test (panel A and B), or two-way ANOVA with Tukey’s post hoc test (panel F). *: p ≤ 0.05, **: p ≤ 0.01, ***: p ≤ 0.005 stimulated versus unstimulated cells (panels A to E) or vehicle versus inhibitor-treated cells (panel F). Full article ">Figure 4
Myosin Vb sustains ACKR2 upregulation and chemokine degradation. (A,B) Confocal microscopy analysis of CHO-K1/ACKR2 or CHO-K1/CCR5 cells stimulated with 100 nM CCL3L1 (1 h) and double stained for ACKR2 or CCR5 (red; panels A and B, respectively) and myosin Vb (green). Nuclear staining (DAPI) is in blue. Merged images are shown in left panels, right panels show colocalization channels, with quantification of ACKR2 or CCR5 colocalization with myosin Vb expressed as mean ± SEM of PCC evaluated in at least n = 30 cells. (C,D) Confocal microscopy analysis of CHO-K1/ACKR2 cells transiently transfected with a pEGFP-tagged myosin Vb tail and stimulated with 100 nM CCL3L1 (1 h) and double stained for ACKR2 (red) and microtubules (α-tubulin staining; green; panel C) or Rab11 (green; panel D). Nuclear staining (DAPI) is in blue. Left and right images refer to cells negative and positive for myosin Vb tail transfection, respectively. Graphs on the right report quantification of ACKR2 colocalization with microtubules (panel C) and Rab11 (panel D) shown as mean ± SEM of PCC evaluated in at least n = 15 cells (open bar: untreated cells, black bar: stimulated cells). (E) Flow cytometry analysis of ACKR2 constitutive internalization in transiently transfected pEGFP-negative CHO-K1/ACKR2 cells (open symbols) or expressing a pEGFP-tagged myosin Vb tail (closed symbols), evaluated as membrane expression after incubation with anti-ACKR2 antibody at 4 °C, and finally shifted to 37 °C for the indicated time, followed by incubation with a secondary antibody at 4 °C. (F) Flow cytometry analysis of ACKR2 membrane expression in unstimulated (open bars) or stimulated cells for 1 h with 100 nM of CCL3L1. (G) ACKR2 scavenging activity evaluated in sorted CHO-K1/ACKR2 cells not expressing (open symbols) or expressing a pEGFP-tagged myosin Vb tail (closed symbols) following incubation with 10 nM CCL3L1 at indicated time points. In panels E and F, results are shown as mean ± SEM of n = 3 experiments, whereas in panel G of n = 2 experiments. Data were analyzed by two-way ANOVA with Tukey’s post hoc test (panel C–F), or unpaired Student’s t-test (panel A and B). ***: p ≤ 0.005 stimulated versus untreated cells; ##: p ≤ 0.01, ###: p ≤ 0.005 negative versus positive, or untransfected versus myosin Vb tail cells. Full article ">Figure 5
ACKR2 promotes microtubule network rearrangement through a β-arrestin1–Rac1–PAK1–LIMK1 dependent pathway. Confocal microscopy analysis of CHO-K1/ACKR2 cells pretreated with 100 ng/mL PTX (16 h; panel A) or 200 µM NSC23766 (1 h; panel C), or transfected with scrambled or specific siRNA for β-arrestin1, PAK1, or LIMK1 (50 nM for 72 h in panels B, D, and E, respectively) and stimulated with 100 mM CCL3L1 (1 h). The siRNA-transfected cells were analyzed by western blotting for β-arrestin1, PAK1, and LIMK1 content, as shown in the top right of corresponding panels. Figures show nuclear staining (DAPI) in blue, ACKR2 in red, and microtubules (α-tubulin staining) in green. Quantification of ACKR2 colocalization with microtubules is shown as the mean ± SEM of PCC evaluated in at least n = 20 cells. Data were analyzed by two-way ANOVA with Tukey’s post hoc test. *: p ≤ 0.05, ***: p ≤ 0.01, **: p ≤ 0.005 stimulated versus unstimulated cells; ###: p ≤ 0.005 vehicle-treated versus NSC23766-treated cells. Full article ">Figure 6
Apoptosis-induced alteration of actin dynamics impairs ACKR2 internalization and scavenging. (A) Confocal microscopy analysis of CHO-K1/ACKR2 incubated for 6 h with vehicle (DMSO) or 1 µM staurosporine and double stained for ACKR2 (red) and phallodin (green). Staining was performed in the absence of triton permabilization. Nuclear staining (DAPI) is in blue. CHO-K1/ACKR2 cells were incubated for 6 h with vehicle (DMSO) or 1 µM staurosporine and analyzed for ACKR2 membrane expression (B) and ACKR2 constitutive internalization after indicated time points (C). Effect of staurosporine (1 µM, 6 h) on ACKR2 scavenging activity in CHO-K1/ACKR2 cells following incubation with 1 nM CCL3L1 for 3 h. Results are shown as mean ± SEM of at least n = 3 experiments. Data were analyzed by unpaired Student’s t-test (panel D) or two-way ANOVA with Tukey’s post hoc test and only statistical analysis of inhibitor versus corresponding vehicle is shown (panel C). **: p ≤ 0.01, ***: p ≤ 0.005. Full article ">

15 pages, 253 KiB

Open AccessReview

Cognitive and Memory Functions in Plant Immunity

by Hidetaka Yakura

Vaccines 2020, 8(3), 541; https://doi.org/10.3390/vaccines8030541 - 17 Sep 2020

Cited by 17 | Viewed by 3962

Abstract

From the time of Thucydides in the 5th century BC, it has been known that specific recognition of pathogens and memory formation are critical components of immune functions. In contrast to the immune system of jawed vertebrates, such as humans and mice, plants [...] Read more.

From the time of Thucydides in the 5th century BC, it has been known that specific recognition of pathogens and memory formation are critical components of immune functions. In contrast to the immune system of jawed vertebrates, such as humans and mice, plants lack a circulatory system with mobile immune cells and a repertoire of clonally distributed antigen receptors with almost unlimited specificities. However, without these systems and mechanisms, plants can live and survive in the same hostile environment faced by other organisms. In fact, they achieve specific pathogen recognition and elimination, with limited self-reactivity, and generate immunological memory, sometimes with transgenerational characteristics. Thus, the plant immune system satisfies minimal conditions for constituting an immune system, namely, the recognition of signals in the milieu, integration of that information, subsequent efficient reaction based on the integrated information, and memorization of the experience. In the previous report, this set of elements was proposed as an example of minimal cognitive functions. In this essay, I will first review current understanding of plant immunity and then discuss the unique features of cognitive activities, including recognition of signals from external as well as internal environments, autoimmunity, and memory formation. In doing so, I hope to reach a deeper understanding of the significance of immunity omnipresent in the realm of living organisms. Full article

(This article belongs to the Special Issue Immune Mechanisms in Plants)

16 pages, 3032 KiB

Open AccessArticle

GMMA Is a Versatile Platform to Design Effective Multivalent Combination Vaccines

by Francesca Micoli, Renzo Alfini, Roberta Di Benedetto, Francesca Necchi, Fabiola Schiavo, Francesca Mancini, Martina Carducci, Elena Palmieri, Cristiana Balocchi, Gianmarco Gasperini, Brunella Brunelli, Paolo Costantino, Roberto Adamo, Diego Piccioli and Allan Saul

Vaccines 2020, 8(3), 540; https://doi.org/10.3390/vaccines8030540 - 17 Sep 2020

Cited by 60 | Viewed by 6221

Abstract

Technology platforms are an important strategy to facilitate the design, development and implementation of vaccines to combat high-burden diseases that are still a threat for human populations, especially in low- and middle-income countries, and to address the increasing number and global distribution of [...] Read more.

Technology platforms are an important strategy to facilitate the design, development and implementation of vaccines to combat high-burden diseases that are still a threat for human populations, especially in low- and middle-income countries, and to address the increasing number and global distribution of pathogens resistant to antimicrobial drugs. Generalized Modules for Membrane Antigens (GMMA), outer membrane vesicles derived from engineered Gram-negative bacteria, represent an attractive technology to design affordable vaccines. Here, we show that GMMA, decorated with heterologous polysaccharide or protein antigens, leads to a strong and effective antigen-specific humoral immune response in mice. Importantly, GMMA promote enhanced immunogenicity compared to traditional formulations (e.g., recombinant proteins and glycoconjugate vaccines), without negative impact to the anti-GMMA immune response. Our findings support the use of GMMA as a “plug and play” technology for the development of effective combination vaccines targeting different bugs at the same time. Full article

(This article belongs to the Section Vaccines against Tropical and other Infectious Diseases)

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Figure 1

Figure 1
GMMA as a carrier for protein antigens: antigen-specific IgG response. Eight CD1 mice per group were s.c. immunized at days 0 and 28, with S. Typhimurium GMMA conjugates of Pfs25 (0.1 µg Pfs25/dose), CSP (0.1 µg CSP/dose) or fHbp (0.75 µg fHbp/dose). Protein antigens, GMMA alone or their physical mixtures were used as controls. Formulations were tested with Alhydrogel. Sera were collected at days 1, 27 and 42 and analyzed for anti-protein antigen-specific IgG response (A–C) and anti-OAg IgG response (D,E). Summary graphs of anti-antigen-specific IgG geometric mean units (bars) and individual antibody levels (dots) are reported. In graphs (D–F), no significant differences in ELISA units were observed. Datasets were analyzed using a two-tailed nonparametric Mann–Whitney test. p-values (rounded to the nearest larger number) less than 0.05 were considered statistically significant. Full article ">Figure 2
GMMA as a carrier for protein antigens: functionality of the antibody response. Eight CD1 mice per group were s.c. immunized at days 0 and 28, with S. Typhimurium GMMA conjugate of fHbp (0.75 µg fHbp/dose). Protein antigen, GMMA alone or their physical mixture were used as controls. Formulations were tested with Alhydrogel. SBA titers of pooled sera collected at day 42 from each group against a panel of meningococcal strains and the S. Typhimurium strain are reported. Full article ">Figure 3
GMMA as a carrier for polysaccharides compared to traditional CRM197 conjugates. Eight CD1 mice per group were i.m. immunized at days 0 and 28, with MenA-GMMA (MenB) (A,C) or MenC-GMMA (MenB) conjugates (B,D) and their corresponding CRM197 conjugates at 1 µg Men oligosaccharide per dose, in the presence of Alhydrogel. MenB GMMA alone and their physical mixture with Men oligosaccharides were used as controls. Sera were collected at days 1, 27, 42 and 197 and analyzed for anti-PS-specific IgG response (A,B). Summary graphs of anti-PS IgG geometric mean units (bars) and individual antibody levels (dots) are reported. Anti-PS IgG were analyzed using a two-tailed nonparametric Mann–Whitney test. p-values (rounded to the nearest larger number) less than 0.05 were considered statistically significant. SBA titers of pooled sera collected 2 weeks and 6 months after second injection against MenA (C) or MenC (D) strains are reported. Full article ">Figure 4
GMMA as a carrier for polysaccharides, formulations tested in the absence of Alhydrogel. Eight CD1 mice per group were s.c. immunized at days 0 and 28, with MenC-GMMA (MenB) or MenC-CRM197 conjugates ((A): 1 µg MenC oligosaccharide per dose). 8 adult rats per group were i.m. immunized at days 0 and 28, with Hib-GMMA (MenB) or Hib-CRM197 conjugates ((B): 0.5 µg Hib oligosaccharide per dose). Sera were collected at days 1, 27 and 42 and analyzed for anti-PS-specific IgG response. Summary graphs of anti-PS IgG geometric mean units (bars) and individual antibody levels (dots) are reported. SBA titers of pooled sera collected at day 42 from each group against MenC strain are reported in the table in panel A. Datasets were analyzed using a two-tailed nonparametric Mann–Whitney test. p-values (rounded to the nearest larger number) less than 0.05 were considered statistically significant. Full article ">Figure 5
SslE and FdeC presented on the same S. sonnei GMMA particle compared to corresponding monovalent conjugates. Eight CD1 mice per group were i.m. immunized at days 0 and 28, with 5 µg total protein, in the presence of Alhydrogel. Sera were collected at days 1, 27 and 42 and analyzed for anti-E. coli antigen-specific (A,B) and anti-S. sonnei LPS (C) IgG response. Summary graphs of IgG geometric mean units (bars) and individual antibody levels (dots) are reported. Datasets were analyzed using a two-tailed nonparametric Mann–Whitney test. p-values (rounded to the nearest larger number) less than 0.05 were considered statistically significant. Full article ">Figure 6
MenA and MenC oligosaccharides presented on the same S. Typhimurium GMMA particles compared to corresponding MenA or MenC monovalent GMMA conjugates and GMMA alone. Eight CD1 mice per group were i.m. immunized at days 0 and 28, with 1 µg MenA and MenC oligosaccharides per dose, in the presence of Alhydrogel. SBA titers of pooled sera collected 2 weeks after the second injection against MenA, MenC and S. Typhimurium strains are reported. Full article ">

15 pages, 1885 KiB

Open AccessArticle

Akt+ IKKα/β+ Rab5+ Signalosome Mediate the Endosomal Recruitment of Sec61 and Contribute to Cross-Presentation in Bone Marrow Precursor Cells

by Dan Dan Xu, Chun Fang Hu, Xiang You, Nan Nan Lu and Feng Guang Gao

Vaccines 2020, 8(3), 539; https://doi.org/10.3390/vaccines8030539 - 17 Sep 2020

Cited by 2 | Viewed by 3115

Abstract

Cross-presentation in dendritic cells (DC) requires the endosomal relocations of internalized antigens and the endoplasmic reticulum protein Sec61. Despite the fact that endotoxin-containing pathogen and endotoxin-free antigen have different effects on protein kinase B (Akt) and I-kappa B Kinase α/β (IKKα/β) activation, the [...] Read more.

Cross-presentation in dendritic cells (DC) requires the endosomal relocations of internalized antigens and the endoplasmic reticulum protein Sec61. Despite the fact that endotoxin-containing pathogen and endotoxin-free antigen have different effects on protein kinase B (Akt) and I-kappa B Kinase α/β (IKKα/β) activation, the exact roles of Akt phosphorylation, IKKα or IKKβ activation in endotoxin-containing pathogen-derived cross-presentation are poorly understood. In this study, endotoxin-free ovalbumin supplemented with endotoxin was used as a model pathogen. We investigated the effects of endotoxin-containing pathogen and endotoxin-free antigen on Akt phosphorylation, IKKα/β activation, and explored the mechanisms that the endotoxin-containing pathogen orchestrating the endosomal recruitment of Sec61 of the cross-presentation in bone marrow precursor cells (BMPC). We demonstrated that endotoxin-containing pathogen and endotoxin-free antigen efficiently induced the phosphorylation of Akt-IKKα/β and Akt-IKKα, respectively. Endotoxin-containing pathogen derived Akt+ IKKα/β+ Rab5+ signalosome, together with augmented the recruitment of Sec61 toward endosome, lead to the increased cross-presentation in BMPC. Importantly, the endosomal recruitment of Sec61 was partly mediated by the formation of Akt+ IKKα/β+ signalosome. Thus, these data suggest that Akt+ IKKα/β+ Rab5+ signalosome contribute to endotoxin-containing pathogen-induced the endosomal recruitment of Sec61 and the superior efficacy of cross-presentation in BMPC. Full article

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18 pages, 2152 KiB

Open AccessArticle

Recognition of Dimeric Lewis X by Anti-Dimeric Le^x Antibody SH2

by Sinthuja Jegatheeswaran, Ari Asnani, Adam Forman, Jenifer L. Hendel, Christopher J. Moore, Ali Nejatie, An Wang, Jo-Wen Wang and France-Isabelle Auzanneau

Vaccines 2020, 8(3), 538; https://doi.org/10.3390/vaccines8030538 - 17 Sep 2020

Cited by 4 | Viewed by 4401

Abstract

The carbohydrate antigen dimeric Lewis X (DimLe^x), which accumulates in colonic and liver adenocarcinomas, is a valuable target to develop anti-cancer therapeutics. Using the native DimLe^x antigen as a vaccine would elicit an autoimmune response against the Le^x antigen [...] Read more.

The carbohydrate antigen dimeric Lewis X (DimLe^x), which accumulates in colonic and liver adenocarcinomas, is a valuable target to develop anti-cancer therapeutics. Using the native DimLe^x antigen as a vaccine would elicit an autoimmune response against the Le^x antigen found on normal, healthy cells. Thus, we aim to study the immunogenic potential of DimLe^x and search internal epitopes displayed by DimLe^x that remain to be recognized by anti-DimLe^x monoclonal antibodies (mAbs) but no longer possess epitopes recognized by anti-Le^x mAbs. In this context, we attempted to map the epitope recognized by anti-DimLe^x mAb SH2 by titrations and competitive inhibition experiments using oligosaccharide fragments of DimLe^x as well as Le^x analogues. We compare our results with that reported for anti-Le^x mAb SH1 and anti-polymeric Le^x mAbs 1G5F6 and 291-2G3-A. While SH1 recognizes an epitope localized to the non-reducing end Le^x trisaccharide, SH2, 1G5F6, and 291-2G3-A have greater affinity for DimLe^x conjugates than for Le^x conjugates. We show, however, that the Le^x trisaccharide is still an important recognition element for SH2, which (like 1G5F6 and 291-2G3-A) makes contacts with all three sugar units of Le^x. In contrast to mAb SH1, anti-polymeric Le^x mAbs make contact with the GlcNAc acetamido group, suggesting that epitopes extend further from the non-reducing end Le^x. Results with SH2 show that this epitope is only recognized when DimLe^x is presented by glycoconjugates. We have reported that DimLe^x adopts two conformations around the β-d-GlcNAc-(1→3)-d-Gal bond connecting the Le^x trisaccharides. We propose that only one of these conformations is recognized by SH2 and that this conformation is favored when the hexasaccharide is presented as part of a glycoconjugate such as DimLe^x-bovine serum albumin (DimLe^x-BSA). Proper presentation of the oligosaccharide candidate via conjugation to a protein or lipid is essential for the design of an anti-cancer vaccine or immunotherapeutic based on DimLe^x. Full article

(This article belongs to the Special Issue Carbohydrate Immunogens in Vaccines)

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19 pages, 2487 KiB

Open AccessReview

Electroporation as the Immunotherapy Strategy for Cancer in Veterinary Medicine: State of the Art in Latin America

by Felipe Maglietti, Matías Tellado, Mariangela De Robertis, Sebastián Michinski, Juan Fernández, Emanuela Signori and Guillermo Marshall

Vaccines 2020, 8(3), 537; https://doi.org/10.3390/vaccines8030537 - 17 Sep 2020

Cited by 19 | Viewed by 6483

Abstract

Electroporation is a technology that increases cell membrane permeability by the application of electric pulses. Electrochemotherapy (ECT), the best-known application of electroporation, is a very effective local treatment for tumors of any histology in human and veterinary medicine. It induces a local yet [...] Read more.

Electroporation is a technology that increases cell membrane permeability by the application of electric pulses. Electrochemotherapy (ECT), the best-known application of electroporation, is a very effective local treatment for tumors of any histology in human and veterinary medicine. It induces a local yet robust immune response that is responsible for its high effectiveness. Gene electrotransfer (GET), used in research to produce a systemic immune response against cancer, is another electroporation-based treatment that is very appealing for its effectiveness, low cost, and simplicity. In this review, we present the immune effect of electroporation-based treatments and analyze the results of the vast majority of the published papers related to immune response enhancement by gene electrotransfer in companion animals with spontaneous tumors. In addition, we present a brief history of the initial steps and the state of the art of the electroporation-based treatments in Latin America. They have the potential to become an essential form of immunotherapy in the region. This review gives insight into the subject and helps to choose promising research lines for future work; it also helps to select the adequate treatment parameters for performing a successful application of this technology. Full article

(This article belongs to the Special Issue Cancer Vaccines and Immunotherapy for Tumor Prevention and Treatment)

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A case of a canine sebaceous gland carcinoma in a 4-year-old female cocker spaniel. In (a), the initial presentation, with a large lesion of 15 × 6.5 cm containing multiple nodules and ulcers. Only 20% of the tumors were treated with Electrochemotherapy (ECT) due to the large extension and spread of the disease, intending to repeat more sessions in the future. In (b), the patient achieved a complete response within 12 weeks of a single ECT session. Fifteen months later, the patient remains free of disease (data not published). Full article ">Figure 2
A case of a golden retriever with a fibrosarcoma, in (a), before the ECT. In (b), the patient obtained a partial response after the treatment. In contrast to the natural evolution of the disease, it remained stable without any other treatment after two years (data not published). Full article ">Figure 3
In this case, a horse with a sarcoid in the eyelid. In (a), the day the ECT was performed. In (b), thirty days after the ECT, the tumor experiences an increase in its size due to swelling. In (c), after a single session of ECT, a complete response was obtained. The animal remained disease-free for 400 days, and up to the date of the writing of this work (data not published). Full article ">Figure 4
Map of Latin America. Orange spots indicate countries where ECT is performed as a standard of care in veterinary medicine. Full article ">Figure 5
Different electrodes for ECT in veterinary medicine. In (a), a six-needle electrode. In (b), a four-needle electrode using thin-needles. In (c), a 90 degrees handle for the thin-needles electrode. In (d), an electrode for treating nasal duct. In (e), on top, disposable-needles, at the bottom, plates electrode that can be attached to the handles. Courtesy of BIOTEX SRL. Full article ">

22 pages, 1723 KiB

Open AccessArticle

Effectiveness of a Simultaneous rHVT-F(ND) and rHVT-H5(AI) Vaccination of Day-Old Chickens and the Influence of NDV- and AIV-Specific MDA on Immune Response and Conferred Protection

by Fabienne Rauw, Eva Ngabirano, Yannick Gardin, Vilmos Palya and Bénédicte Lambrecht

Vaccines 2020, 8(3), 536; https://doi.org/10.3390/vaccines8030536 - 16 Sep 2020

Cited by 6 | Viewed by 4028

Abstract

The recombinant herpesvirus of turkey (rHVT) vaccines targeting Newcastle disease (ND) and H5Nx avian influenza (AI) have been demonstrated efficient in chickens when used individually at day-old. Given the practical field constraints associated with administering two vaccines separately and in the absence of [...] Read more.

The recombinant herpesvirus of turkey (rHVT) vaccines targeting Newcastle disease (ND) and H5Nx avian influenza (AI) have been demonstrated efficient in chickens when used individually at day-old. Given the practical field constraints associated with administering two vaccines separately and in the absence of a currently available bivalent rHVT vector vaccine expressing both F(ND) and H5(AI) antigens, the aim of this study was to investigate whether interference occurs between the two vaccines when simultaneously administered in a single shot. The studies have been designed to determine (i) the ND and AI-specific protection and antibody response conferred by these vaccines inoculated alone or in combination at day-old, (ii) the influence of maternally-derived antibodies (MDA), and (iii) the potential interference between the two vaccine. Our results demonstrate that their combined administration is efficient to protect chickens against clinical signs of velogenic Newcastle disease virus (vNDV) and H5-highly pathogenic avian influenza virus (HPAIV) infections. Viral shedding following co-vaccination is also markedly reduced, while slightly lower NDV- and AIV-specific antibody responses are observed. NDV- and AIV-specific MDA show negative effects on the onset of the specific antibody responses. However, if AIV-specific MDA reduce the protection against H5-HPAIV induced by rHVT-H5(AI) vaccine, it was not observed for ND. Full article

(This article belongs to the Special Issue Poultry Vaccines)

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11 pages, 986 KiB

Open AccessArticle

Relationship between Influenza Vaccination Coverage Rate and COVID-19 Outbreak: An Italian Ecological Study

by Mauro Amato, José Pablo Werba, Beatrice Frigerio, Daniela Coggi, Daniela Sansaro, Alessio Ravani, Palma Ferrante, Fabrizio Veglia, Elena Tremoli and Damiano Baldassarre

Vaccines 2020, 8(3), 535; https://doi.org/10.3390/vaccines8030535 - 16 Sep 2020

Cited by 107 | Viewed by 24684

Abstract

Background: The lack of specific vaccines or drugs against coronavirus disease 2019 (COVID-19) warrants studies focusing on alternative clinical approaches to reduce the spread of this pandemic disease. In this study, we investigated whether anti-influenza vaccination plays a role in minimizing the diffusion [...] Read more.

Background: The lack of specific vaccines or drugs against coronavirus disease 2019 (COVID-19) warrants studies focusing on alternative clinical approaches to reduce the spread of this pandemic disease. In this study, we investigated whether anti-influenza vaccination plays a role in minimizing the diffusion of COVID-19 in the Italian population aged 65 and over. Methods: Four COVID-19 outcomes were used: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence, hospitalizations for COVID-19 symptoms, admissions to intensive care units for reasons related to SARS-CoV-2, and deaths attributable to COVID-19. Results: At univariate analyses, the influenza vaccination coverage rates correlated negatively with all COVID-19 outcomes (Beta ranging from −134 to −0.61; all p < 0.01). At multivariable analyses, influenza vaccination coverage rates correlated independently with SARS-CoV-2 seroprevalence (Beta (95% C.I.): −130 (−198, −62); p = 0.001), hospitalizations for COVID-19 symptoms (Beta (95% C.I.): −4.16 (−6.27, −2.05); p = 0.001), admission to intensive care units for reasons related to SARS-CoV-2 (Beta (95% C.I.): −0.58 (−1.05, −0.12); p = 0.017), and number of deaths attributable to COVID-19 (Beta (95% C.I.): −3.29 (−5.66, −0.93); p = 0.010). The R² observed in the unadjusted analysis increased from 82% to 159% for all the considered outcomes after multivariable analyses. Conclusions: In the Italian population, the coverage rate of the influenza vaccination in people aged 65 and over is associated with a reduced spread and a less severe clinical expression of COVID-19. This finding warrants ad hoc studies to investigate the role of influenza vaccination in preventing the spread of COVID-19. Full article

(This article belongs to the Special Issue Vaccination and Public Health: Optimizing Vaccine Uptake through the Application of Social and Behavioral Science Theory, Principles, and Strategies)

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40 pages, 881 KiB

Open AccessReview

Vaccination into the Dermal Compartment: Techniques, Challenges, and Prospects

by Johanna Hettinga and Robert Carlisle

Vaccines 2020, 8(3), 534; https://doi.org/10.3390/vaccines8030534 - 16 Sep 2020

Cited by 45 | Viewed by 11672

Abstract

In 2019, an ‘influenza pandemic’ and ‘vaccine hesitancy’ were listed as two of the top 10 challenges to global health by the WHO. The skin is a unique vaccination site, due to its immune-rich milieu, which is evolutionarily primed to respond to challenge, [...] Read more.

In 2019, an ‘influenza pandemic’ and ‘vaccine hesitancy’ were listed as two of the top 10 challenges to global health by the WHO. The skin is a unique vaccination site, due to its immune-rich milieu, which is evolutionarily primed to respond to challenge, and its ability to induce both humoral and cellular immunity. Vaccination into this dermal compartment offers a way of addressing both of the challenges presented by the WHO, as well as opening up avenues for novel vaccine formulation and dose-sparing strategies to enter the clinic. This review will provide an overview of the diverse range of vaccination techniques available to target the dermal compartment, as well as their current state, challenges, and prospects, and touch upon the formulations that have been developed to maximally benefit from these new techniques. These include needle and syringe techniques, microneedles, DNA tattooing, jet and ballistic delivery, and skin permeabilization techniques, including thermal ablation, chemical enhancers, ablation, electroporation, iontophoresis, and sonophoresis. Full article

(This article belongs to the Special Issue Genomic Medicine and Advances in Vaccine Technology and Development in the Developing and Developed World)

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15 pages, 1122 KiB

Open AccessArticle

Evaluation of Antibody Response Directed against Porcine Reproductive and Respiratory Syndrome Virus Structural Proteins

by Hung Q. Luong, Huong T. L. Lai and Hiep L. X. Vu

Vaccines 2020, 8(3), 533; https://doi.org/10.3390/vaccines8030533 - 16 Sep 2020

Cited by 9 | Viewed by 4540

Abstract

Luciferase-immunoprecipitation system (LIPS), a liquid phase immunoassay, was used to evaluate antibody responses directed against the structural proteins of PRRSV in pigs that were experimentally infected with virulent PRRSV strains. First, the viral N protein was used as a model antigen to validate [...] Read more.

Luciferase-immunoprecipitation system (LIPS), a liquid phase immunoassay, was used to evaluate antibody responses directed against the structural proteins of PRRSV in pigs that were experimentally infected with virulent PRRSV strains. First, the viral N protein was used as a model antigen to validate the assay. The LIPS results were highly comparable to that of the commercial IDEXX PRRS X3 ELISA. Subsequently, the assay was applied to simultaneously measure antibody reactivity against all eight structural proteins of PRRSV. The highest immunoreactivities were detected against GP3, M, and N proteins while the lowest reactivity was detected against ORF5a protein. Comparative analysis of the kinetics of antibody appearance revealed that antibodies specific to N protein appeared earlier than antibodies against GP3. Finally, the assay was applied to measure immunoreactivities of clinical serum samples against N and GP3. The diagnostic sensitivity of the LIPS with N protein was superior to that of the LIPS with GP3. Collectively, the results provide additional information about the host antibody response to PRRSV infection. Full article

(This article belongs to the Special Issue PRRSV Vaccinology and Immunology)

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Schematic representation of the luciferase-immunoprecipitation system (LIPS). (a) Generation of luciferase-tagged antigens (Ag). Each individual PRRSV structural protein antigen is cloned in-frame to the 5′ terminus of the nanoluc luciferase gene (Nluc). The resulting plasmid is transfected into HEK 293-T cells. At 60 h after transfection, cell lysate containing Nluc-tagged antigen is harvested and used for the LIPS. (b) Evaluation of immunoreactivities against the NLuc-tagged antigens. Cell lysate containing Nluc-tagged antigen is incubated with test serum samples together with protein A Sepharose beads in a filter 96-well plate. If the test samples contain antibody (IgG) specific to the Nluc-tagged antigen, the antigen-antibody complexes are formed, which are captured by the protein A Sepharose beads and retained in the well. Unbound NLuc-tagged antigen is washed away. Once the luciferase substrate is added to the well, the Nluc-tagged antigen bound to the bead will react with the substrate and produce luminescence. The light units produced by the Nluc-tagged antigen is proportional to the amount of antigen-specific antibody present in the test serum samples. Full article ">Figure 2
Optimization of the LIPS with N protein. (a) HEK-293T cells were transfected with plasmids encoding Nluc-tagged N protein (N) or Nluc only. At 60 h after transfection, cells were fixed and subjected to indirect immunofluorescence assay (IFA) using a monoclonal antibody specific to Nluc protein. The cell nuclear was stained with DAPI (blue). (b) HEK-293T cells were transfected as described previously. At 60 h post-transfection, cells were harvested and lysed in RIPA buffer and subjected to immunoblotting analysis using a monoclonal antibody specific to Nluc protein. (c) Serum samples collected from 35 pigs experimentally infected with the PRRSV-01 or FL12 and their derivative mutants at 0 dpi and 42 dpi. Serological status of the serum samples as determined by the IDEXX ELISA. Data are expressed as the sample to positive (S/P) ratios. The horizontal dotted line at S/P = 0.4 indicates the cutoff of this assay. (d) Immunoreactivity of the serum samples were measured against Nluc-tagged N protein as described in Materials and Method. Fetal bovine serum (FBS) was used as negative control. Data are expressed as sample to negative (S/N) ratios. The dotted line at S/N = 4.02 indicates the cutoff of the assay. Full article ">Figure 3
Swine antibody reactivities against PRRSV structural proteins. (a,b) Evaluation of protein expression. HEK-293T cells were transfected with plasmids encoding Nluc-tagged antigens. At 60 h post-transfection, cells were fixed and subjected to IFA and immunoblotting analysis as described in the <a href="#vaccines-08-00533-f002" class="html-fig">Figure 2</a> legend. (c) Thirty-eight serum sample collected before infection (0 dpi) and 44 samples collected at between 42 and 63 dpi from 44 pigs experimentally infected with the PRRSV strain FL12. The antibody reactivities of these serum samples were simultaneous measured against eight PRRSV structural proteins as described in Materials and Method. Fetal bovine serum (FBS) was used as negative control. Data are expressed as sample to negative (S/N) ratios. The dotted lines indicate the cutoffs of the respective assays. The numbers within parenthesis below the antigen names indicate the proportion of convalescent serum samples tested positive for that respective antigens. One-way analysis of variance (ANOVA) was used to evaluate the difference among the mean S/N ratios of the convalescent antisera, followed by Tukey’s multiple comparison test. The superscript letters at the top of the graph denote the statistical comparison of the S/N ratios among the eight antigens. Means sharing the same superscript are not significantly different from each other (p > 0.05). Full article ">Figure 4
Kinetics of antibody responses to GP3 and N protein. Serum samples were collected from 32 pigs experimentally infected with the PRRSV strain FL12 at 0, 7, 14, and 21 dpi. (a) The samples were tested with the IDEXX ELISA. Data are expressed as the sample to positive (S/P) ratios. The horizontal dotted line at S/P = 0.4 indicates the cutoff of this assay. (b) The samples were tested by the LIPS with GP3 and N protein as described in Materials and Method. Fetal bovine serum (FBS) was used as negative control. Data are expressed as sample to negative (S/N) ratios. The dotted lines indicate the cutoffs of the respective assays. Full article ">Figure 5
Measurement of antibody reactivities against GP3 and N with clinical serum samples. A set of 84 seronegative and 84 seropositive clinical serum samples were obtained from the Iowa State University Veterinary Diagnostic Laboratory. (a) The serological status of these samples was verified by using the IDEXX ELISA. Data are expressed as the sample to positive (S/P) ratios. The horizontal dotted line at S/P = 0.4 indicates the cutoff of this assay. (b) The samples were tested by LIPS with GP3 and N protein as described in Materials and Method. Fetal bovine serum (FBS) was used as negative control. Data are expressed as sample to negative (S/N) ratios. The dotted lines indicate the cutoffs of the respective assays. Full article ">

13 pages, 2584 KiB

Open AccessArticle

Potent Protective Immune Responses to Senecavirus Induced by Virus-Like Particle Vaccine in Pigs

by Suyu Mu, Shiqi Sun, Hu Dong, Manyuan Bai, Yun Zhang, Zhidong Teng, Mei Ren, Shuanghui Yin and Huichen Guo

Vaccines 2020, 8(3), 532; https://doi.org/10.3390/vaccines8030532 - 15 Sep 2020

Cited by 14 | Viewed by 3912

Abstract

Senecavirus A (SVA) is the pathogen that has recently caused porcine idiopathic vesicular disease (PIVD). The clinical symptoms of PIVD are similar to those of acute foot-and-mouth disease and also can result in the death of newborn piglets, thus entailing economic losses. Vaccine [...] Read more.

Senecavirus A (SVA) is the pathogen that has recently caused porcine idiopathic vesicular disease (PIVD). The clinical symptoms of PIVD are similar to those of acute foot-and-mouth disease and also can result in the death of newborn piglets, thus entailing economic losses. Vaccine immunization is the most effective way to prevent and control SVA. Among all SVA vaccines reported, only the SVA inactivated vaccine has been successfully developed. However, to ensure the elimination of this pathogen, safer and more effective vaccines are urgently required. A virus-like particles (VLPs)-based vaccine is probably the best alternative to inactivated vaccine. To develop an SVA VLPs vaccine and evaluate its immune effect, a prokaryotic expression system was used to produce SVA capsid protein and assemble VLPs. The VLPs were characterized by affinity chromatography, sucrose density gradient centrifugation, ZetaSizer and transmission electron microscopy. Meanwhile, the SVA CH-HB-2017 strain was used to infect pigs and to determine infection routes and dose. Experimental pigs were then immunized with the SVA VLPs vaccine emulsified in an ISA 201 adjuvant. The results showed that the VLPs vaccine induced neutralizing and specific antibodies at similar levels as an inactivated SVA vaccine after immunization. The level of INF-γ induced by the VLPs vaccine gradually decreased—similar to that of inactivated vaccine. These results indicated that VLPs vaccine may simultaneously cause both cellular and humoral immune responses. Importantly, after the challenge, the VLPs vaccine provided similar levels of protection as the inactivated SVA vaccine. In this study, we successfully obtained novel SVA VLPs and confirmed their highly immunogenicity, thus providing a superior candidate vaccine for defense and elimination of SVA, compared to the inactivated vaccine. Full article

(This article belongs to the Special Issue Controlled Clinical Evaluation of Veterinary Vaccines)

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Clinical symptoms and rectal temperature after Senecavirus A (SVA) infection in pigs. (A) Pigs infected intramuscularly (3 mL, 7 × 107.8 PFU/mL) and intranasally (1.5 mL into each nostril, 7 × 107.8 PFU/mL). Clinical symptoms of pigs were recorded during the test and scored in order. Pigs 11,12, and 15 had blisters in the nose; 13 and 14 had blisters in the hoof. Serum was collected and the virus content was detected using qRT-PCR technology; (B) clinical symptoms after SVA CH-2017-HB strain infection. Pigs developed blisters in the nose at four days and white blisters in the hoof at five and six days. Blisters ulcerated at seven days and the joints were red and swollen. The rupture of the hoof gradually returned at eight days; (C) rectal temperature 1–10 days after SVA infection in pigs (Intramuscularly (3 mL, 7 × 107.8 PFU/mL ) and intranasal (1.5 mL into each nostril, 7 × 107.8 PFU/mL) group (n = 5)). All clinical changes indicated by blue arrows. Black line represents the cutoff value of the qRT-PCR technology detection. Full article ">Figure 2
Expression and assembly of SVA capsid protein. (A) SDS-PAGE (Lanes 1, 2) and western blot (Lanes 3, 4). Lanes 1, 3: SVA purified capsid-protein complexes His6-sm-VP0, His6-sm-VP1 and His6-sm-VP3. Lanes 2, 4: Target proteins VP0, VP1, VP3 purified after protease digestion; (B) SVA VLPs purified by sucrose gradient ultracentrifugation and the OD280 fluorescence absorption measured by spectrophotometer. Negative-staining electron micrographs of SVA pentamer (right, orange arrow) and VLPs (left, orange arrow). Scale bar = 50 nm. Full article ">Figure 3
Detection of serum antibodies. (A) Specific antibodies; (B) neutralizing antibodies; (C) IgG1 antibodies; (D) IgG2a antibodies. Levels of serum antibody detected in pigs vaccinated with SVA VLPs (n = 5), inactivated vaccines (n = 5) and PBS (n = 5) at 0 days, 14 days, 21 days, 35 days and 49 days post-immunization (ns were considered no statistically significant, * p <0.05, ** p <0.01, **** p <0.0001). Full article ">Figure 4
Detection of serum INF-γ. Serum collected from pigs vaccinated with SVA VLPs (n = 5), inactivated vaccines (n = 5) and PBS (n = 5) at 0 days, 14 days, 21 days, 35 days and 49 days post-immunization. INF-γ titers represent the intensity of cellular immune response (ns were considered no statistically significant, ** p < 0.01). Full article ">

27 pages, 1099 KiB

Open AccessReview

Current State of Global African Swine Fever Vaccine Development under the Prevalence and Transmission of ASF in China

by Keke Wu, Jiameng Liu, Lianxiang Wang, Shuangqi Fan, Zhaoyao Li, Yuwan Li, Lin Yi, Hongxing Ding, Mingqiu Zhao and Jinding Chen

Vaccines 2020, 8(3), 531; https://doi.org/10.3390/vaccines8030531 - 15 Sep 2020

Cited by 95 | Viewed by 11391

Abstract

African swine fever (ASF) is a highly lethal contagious disease of swine caused by African swine fever virus (ASFV). At present, it is listed as a notifiable disease reported to the World Organization for Animal Health (OIE) and a class one animal disease [...] Read more.

African swine fever (ASF) is a highly lethal contagious disease of swine caused by African swine fever virus (ASFV). At present, it is listed as a notifiable disease reported to the World Organization for Animal Health (OIE) and a class one animal disease ruled by Chinese government. ASF has brought significant economic losses to the pig industry since its outbreak in China in August 2018. In this review, we recapitulated the epidemic situation of ASF in China as of July 2020 and analyzed the influencing factors during its transmission. Since the situation facing the prevention, control, and eradication of ASF in China is not optimistic, safe and effective vaccines are urgently needed. In light of the continuous development of ASF vaccines in the world, the current scenarios and evolving trends of ASF vaccines are emphatically analyzed in the latter part of the review. The latest research outcomes showed that attempts on ASF gene-deleted vaccines and virus-vectored vaccines have proven to provide complete homologous protection with promising efficacy. Moreover, gaps and future research directions of ASF vaccine are also discussed. Full article

(This article belongs to the Special Issue African Swine Fever Virus Prevention and Control)

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21 pages, 3498 KiB

Open AccessReview

The Molecular Interactions of ZIKV and DENV with the Type-I IFN Response

by Rosa C. Coldbeck-Shackley, Nicholas S. Eyre and Michael R. Beard

Vaccines 2020, 8(3), 530; https://doi.org/10.3390/vaccines8030530 - 14 Sep 2020

Cited by 26 | Viewed by 5557

Abstract

Zika Virus (ZIKV) and Dengue Virus (DENV) are related viruses of the Flavivirus genus that cause significant disease in humans. Existing control measures have been ineffective at curbing the increasing global incidence of infection for both viruses and they are therefore prime targets [...] Read more.

Zika Virus (ZIKV) and Dengue Virus (DENV) are related viruses of the Flavivirus genus that cause significant disease in humans. Existing control measures have been ineffective at curbing the increasing global incidence of infection for both viruses and they are therefore prime targets for new vaccination strategies. Type-I interferon (IFN) responses are important in clearing viral infection and for generating efficient adaptive immune responses towards infection and vaccination. However, ZIKV and DENV have evolved multiple molecular mechanisms to evade type-I IFN production. This review covers the molecular interactions, from detection to evasion, of these viruses with the type-I IFN response. Additionally, we discuss how this knowledge can be exploited to improve the design of new vaccine strategies. Full article

(This article belongs to the Special Issue Virus Immune Escape and Host Immune System)

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Figure 1
A schematic representation of the Flavivirus genome. The genome is a single positive-sense single-stranded RNA (+ssRNA) molecule that is roughly 11 kb in size, capped at the 5′ terminus and flanked by 5′ and 3′ untranslated regions (UTR). The central open reading frame (ORF) encodes a polyprotein that is cleaved into individual structural and non-structural (NS) viral proteins, the text below lists some of their known functions as reviewed in [<a href="#B17-vaccines-08-00530" class="html-bibr">17</a>]. Full article ">Figure 2
Stages of the Flavivirus lifecycle. (1) Attachment and receptor-mediated endocytosis. (2) Membrane fusion and particle disassembly. (3) Genome release into the cytoplasm. (4) Polyprotein translation. (5) Replication complex (RC) formation and genome replication. (6) Virion packaging. (7) Transportation through the trans-Golgi network and virion maturation. (8) Virion egress by exocytosis. Full article ">Figure 3
Recognition of Zika Virus (ZIKV) and Dengue Virus (DENV) by the innate immune system and viral evasion of pattern recognition receptor (PRR)-mediated interferons (IFN) production. ZIKV and DENV infections are sensed by multiple PRRs inside the cell. or pathogen associated molecular pattern (PAMP) vRNA from the replication stage of the virus lifecycle activates RLRs in the cytosol (green arrows) or Toll-like receptors (TLRs) in the endosome (gold arrows). Alternatively, both ZIKV and DENV infection results in release to mitoDNA into the cytoplasm that is sensed by cyclic GMP-AMP synthase (cGAS), activating stimulator of interferon genes (STING) (purple arrows). Each of these pathways leads to phosphorylation of kinases TBK1 and IKKɛ that activate IRF3/7 and NF-κB. These transcription factors then upregulate the production of type-I IFNβ (blue arrows). However, ZIKV and DENV have evolved evasion mechanisms (red) to prevent IFN production that target multiple stages of these pathways. Full article ">Figure 4
The molecular interactions of ZIKV and DENV with the type-I IFN pathway. Once produced and secreted from infected cells type-I IFNs bind to their cognate cell surface receptor, activating janus kinase-signal transducer and activator (JAK-STAT) signaling (black arrows). This leads to the phosphorylation of STAT1 and STAT2, their heterodimerzation and subsequent complexing with IRF9 to form the complex ISGF3. ISGF3 then transcriptionally upregulates interferon stimulated genes (ISGs) that have antiviral effects against ZIKV and DENV. Several stages of the IFN pathway are inhibited by ZIKV and DENV mediated evasion mechanisms (red). Full article ">

26 pages, 974 KiB

Open AccessReview

The Role of the Tumor Microenvironment in Developing Successful Therapeutic and Secondary Prophylactic Breast Cancer Vaccines

by Benjamin Gordon and Vijayakrishna K. Gadi

Vaccines 2020, 8(3), 529; https://doi.org/10.3390/vaccines8030529 - 14 Sep 2020

Cited by 12 | Viewed by 8355

Abstract

Breast cancer affects roughly one in eight women over their lifetime and is a leading cause of cancer-related death in women. While outcomes have improved in recent years, prognosis remains poor for patients who present with either disseminated disease or aggressive molecular subtypes. [...] Read more.

Breast cancer affects roughly one in eight women over their lifetime and is a leading cause of cancer-related death in women. While outcomes have improved in recent years, prognosis remains poor for patients who present with either disseminated disease or aggressive molecular subtypes. Cancer immunotherapy has revolutionized the treatment of several cancers, with therapeutic vaccines aiming to direct the cytotoxic immune program against tumor cells showing particular promise. However, these results have yet to translate to breast cancer, which remains largely refractory from such approaches. Recent evidence suggests that the breast tumor microenvironment (TME) is an important and long understudied barrier to the efficacy of therapeutic vaccines. Through an improved understanding of the complex and biologically diverse breast TME, it may be possible to advance new combination strategies to render breast carcinomas sensitive to the effects of therapeutic vaccines. Here, we discuss past and present efforts to advance therapeutic vaccines in the treatment of breast cancer, the molecular mechanisms through which the TME contributes to the failure of such approaches, as well as the potential means through which these can be overcome. Full article

(This article belongs to the Special Issue Cancer Immunotherapy: Advances and Future Prospects)

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14 pages, 3215 KiB

Open AccessArticle

Near-Infrared Photoimmunotherapy Combined with CTLA4 Checkpoint Blockade in Syngeneic Mouse Cancer Models

by Yasuhiro Maruoka, Aki Furusawa, Ryuhei Okada, Fuyuki Inagaki, Daiki Fujimura, Hiroaki Wakiyama, Takuya Kato, Tadanobu Nagaya, Peter L. Choyke and Hisataka Kobayashi

Vaccines 2020, 8(3), 528; https://doi.org/10.3390/vaccines8030528 - 14 Sep 2020

Cited by 25 | Viewed by 4436

Abstract

Near infrared photoimmunotherapy (NIR-PIT) is a newly developed and highly selective cancer treatment that induces necrotic/immunogenic cell death. It employs a monoclonal antibody (mAb) conjugated to a photo-absorber dye, IRDye700DX, which is activated by NIR light. Tumor-targeting NIR-PIT is also at least partly [...] Read more.

Near infrared photoimmunotherapy (NIR-PIT) is a newly developed and highly selective cancer treatment that induces necrotic/immunogenic cell death. It employs a monoclonal antibody (mAb) conjugated to a photo-absorber dye, IRDye700DX, which is activated by NIR light. Tumor-targeting NIR-PIT is also at least partly mediated by a profound immune response against the tumor. Cytotoxic T-lymphocyte antigen-4 (CTLA4) is widely recognized as a major immune checkpoint protein, which inhibits the immune response against tumors and is therefore, a target for systemic blockade. We investigated the effect of combining tumor-targeted NIR-PIT against the cell-surface antigen, CD44, which is known as a cancer stem cell marker, with a systemic CTLA4 immune checkpoint inhibitor in three syngeneic tumor models (MC38-luc, LL/2, and MOC1). CD44-targeted NIR-PIT combined with CTLA4 blockade showed greater tumor growth inhibition with longer survival compared with CTLA4 blockade alone in all tumor models. NIR-PIT and CTLA4 blockade produced more complete remission in MOC1 tumors (44%) than NIR-PIT and programmed cell death protein 1 (PD-1) blockade (8%), which was reported in our previous paper. However, the combination of NIR-PIT and CTLA4 blockade was less effective in MC38-luc tumors (11%) than the combination of NIR-PIT and PD-1 blockade (70%). Nonetheless, in many cases ineffective results with NIR-PIT and PD-1 blockade were reversed with NIR-PIT and CTLA4 blockade. Full article

(This article belongs to the Section Cellular/Molecular Immunology)

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Graphical abstract

Graphical abstract
Full article ">Figure 1
Confirmation of CD44 expression as a target for NIR-PIT and evaluation of in vitro CD44-targeted NIR-PIT in MC38-luc, LL/2, and MOC1 cells. (A) Expression of cell-surface CD44 in MC38-luc, LL/2 and MOC1 cells was examined with flow cytometry. CD44-blocking antibody was added to some wells to validate specific staining. Representative histograms were shown. (B) Mean fluorescence intensity (MFI) of IR700 after labeling with anti-CD44-mAb-IR700 (n = 4, * p < 0.05, Tukey–Kramer test). (C–E) Membrane permeability as measured by PI staining, after labeling with anti-CD44-mAb-IR700 and treatment with NIR light. (C) MC38-luc; (D) LL/2; (E) MOC1 (n = 5, * p < 0.05, vs. untreated control; Mann–Whitney U test). ns, not significant. Full article ">Figure 2
CD44 expression and immune cell infiltration within MC38-luc, LL/2, and MOC1 tumors. (A) Multiplex immunohistochemistry (IHC) staining was performed to examine CD44 expression and distribution of T cells within MC38-luc, LL/2, and MOC1 tumors before treatment. The top panels show CD44 expression (shown in green). Nucleus were stained with DAPI (shown in blue). White dotted lines represent tumor edges. The middle panels show expressions of CD8 (magenta), CD4 (cyan) and FOXP3 (orange). Inset shows examples of non-regulatory CD4 T cells (membrane CD4+/nucleus FOXP3−, open arrowhead) and Treg (membrane CD4+/nucleus FOXP3+, filled arrowhead). The bottom panels show cellular phenotypes based on the antigen expressions. CD8 T cell, non-regulatory CD4 T cells, Tregs are shown as dots in magenta, cyan and orange respectively. Representative images from at least three samples are shown (×200, scale bar = 100 µm). (B) T cell count in the tumor microenvironment. Cell number of CD8 T cells, non-regulatory CD4 T cells and Tregs in stroma and tumor tissue were counted in multiplex IHC images. Data were shown as cell count per megapixel area (n = 3; * p < 0.05; Tukey’s multiple comparison test). Full article ">Figure 3
In vivo effects of CD44-targeted NIR-PIT and/or CTLA4 mAb administration for MC38-luc, LL/2, and MOC1 tumor models. (A) NIR-PIT regimen. Bioluminescence and fluorescence images were obtained at each time point as indicated. (B–D) Real-time in vivo IR700 fluorescence imaging of tumor-bearing mice before and approximately 10 min after NIR-PIT in MC38-luc (B), LL/2 (C) and MOC1 (D) tumor models. The yellow arrows and white circles indicate the tumor locations. (E) In vivo bioluminescence imaging of MC38-luc tumor-bearing mice before and after treatment at the indicated timepoints. (F) Quantitative analysis of luciferase activity before and after treatment in MC38-luc tumor-bearing mice. n ≥ 10/group, mean ± SEM; * p < 0.05, vs. the other groups; Dunnett’s test. (G–I) Tumor growth in all the groups for MC38-luc (G), LL/2 (H) and MOC1 (I) tumor models. n ≥ 9/group, mean ± SEM; * p < 0.05, vs. the other groups; Dunnett’s test; † p < 0.05, vs. combination group; Tukey–Kramer test; ‡ p < 0.05, vs. combination group; Mann–Whitney U test. (J–L) Survival curves in all the groups for MC38-luc (J), LL/2 (K) and MOC1 (L) tumor models. n ≥ 9/group; * p < 0.05; NS, not significant; Gehan–Breslow–Wilcoxon test. Full article ">Figure 4
Immune cell infiltration in MOC1 tumors 1 week after NIR-PIT. (A) Multiplex IHC images from MOC1 tumors 7 days after treatment. Control, no treatment; CTLA4 mAb, CTLA4 immune-checkpoint blockade only; CD44 PIT, CD44-targeted NIR-PIT only; combination, CD44-targeted NIR-PIT combined with CTLA4 immune-checkpoint blockade. Expressions of CD8, CD4 and FOXP3 are shown in magenta, cyan and orange respectively. The tumor tissues were stained with anti-pan-Cytokeratin, shown in white. The representative pictures from four experiments are shown. (×200, scale bar = 100 µm). (B) T cell count in the tumor microenvironment. Cell number of CD8 T cells, non-regulatory CD4 T cells and Tregs in stroma and tumor tissue were counted in multiplex IHC images. Data were shown as cell count per megapixel area. Control, no treatment; CTLA4 mAb, CTLA4 immune-checkpoint blockade only; CD44 PIT, CD44-targeted NIR-PIT only; combi, CD44-targeted NIR-PIT combined with CTLA4 immune-checkpoint blockade (n = 4 for control, CTLA4 mAb and CD44 PIT, n = 3 for combi, no statistically significant difference (One-way ANOVA). Full article ">

11 pages, 373 KiB

Open AccessArticle

Behavioral Differences in the Preference for Hepatitis B Virus Vaccination: A Discrete Choice Experiment

by Na Guo, Jian Wang, Stephen Nicholas, Elizabeth Maitland and Dawei Zhu

Vaccines 2020, 8(3), 527; https://doi.org/10.3390/vaccines8030527 - 14 Sep 2020

Cited by 16 | Viewed by 4512

Abstract

Understanding behavioral factors differences in the preferences for vaccinations can improve predictions of vaccine uptake rates and identify effective policy interventions to increase the demand for vaccinations. In this study, 353 adults in Shandong province in China were interviewed about their preferences for [...] Read more.

Understanding behavioral factors differences in the preferences for vaccinations can improve predictions of vaccine uptake rates and identify effective policy interventions to increase the demand for vaccinations. In this study, 353 adults in Shandong province in China were interviewed about their preferences for hepatitis B virus (HBV) vaccination. A discrete choice experiment (DCE) was employed to analyze the preference for HBV vaccinations, and a mixed logit model was used to estimate respondent preferences for vaccination attributes included in the DCE. While the protection rate against hepatitis B (HB), duration of protection, risk of side-effects, and vaccination cost were shown to influence adults’ preferences for HBV vaccination, adults valued “99% hepatitis B protection” above other attributes, followed by “20 years’ protection duration” and “1 in 150,000 risk of side-effects”. Individuals with lower time discount rates, non-overconfidence, or higher risk aversion were more likely to choose a vaccine. Lower risk aversion individuals showed a higher preference for lower risk of side-effects. Lower time discount rate individuals showed a higher preference for longer protection duration. Non-overconfidence individuals showed a higher preference for higher hepatitis B protection and cost. Interventions should be targeted to the behavioral determinants impeding vaccination. Full article

(This article belongs to the Special Issue Vaccination and Public Health: Optimizing Vaccine Uptake through the Application of Social and Behavioral Science Theory, Principles, and Strategies)

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Vaccines, Volume 8, Issue 3 (September 2020) – 216 articles

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