うそはうそであると見抜ける人でないと「COVID-19」と「スパイクタンパク質」と「新型コロナワクチン」は難しい

Death汁接種したひろゆきには無理だったようだね。（爆ｗｗｗｗｗｗｗ

 

 Trends Immunol. 2021 Jan; 42(1): 3–5.

Published online 2020 Nov 2. doi: 10.1016/j.it.2020.10.012

PMCID: PMC7605819

PMID: 33214057

A Potential Role of Interleukin 10 in COVID-19 Pathogenesis

Ligong Lu,^1,⁎ Hui Zhang,² Danielle J. Dauphars,³ and You-Wen He^3,⁎

Abstract

A unique feature of the cytokine storm in coronavirus disease 2019 (COVID-19) is the dramatic elevation of interleukin 10 (IL-10). This was thought to be a negative feedback mechanism to suppress inflammation. However, several lines of clinical evidence suggest that dramatic early proinflammatory IL-10 elevation may play a pathological role in COVID-19 severity.

Potential Roles of IL-10 in COVID-19 Pathogenesis

Mortality of COVID-19 patients is caused by severe pneumonia and vital organ damage with the involvement of many proinflammatory mediators. Thus far, no definitive efficacy of reduced mortality in COVID-19 patients has been established in randomized placebo-controlled clinical trials testing blockade of IL-6, IL-1, or GM-CSF [13]. We propose that IL-10 is a potential target for reducing COVID-19 mortality. As mentioned, severe/critically ill COVID-19 patients present dramatically elevated serum IL-10 concentrations that correlate with disease severity [1,3., 4., 5., 6.]. Accordingly, recent studies also demonstrate immune activation and inflammation in COVID-19 patients [13], which supports the hypothesis that IL-10 may play a proinflammatory and immune-activating role in COVID-19 pathogenesis.

In support of this hypothesis, first, the inflammatory/immune-stimulating cytokines discussed in the previous text (IL-2Rα, IL-4, IL-7, IL-18, IFN-γ, GM-CSF, TNF-α, and chemokines IP-10 and CXCL9) are elevated in peripheral blood in severe/critically ill COVID-19 patients [1,3,4]. Second, severe/critically ill COVID-19 patients bear circulating hyperactivated and proliferating cytotoxic CD8⁺ T cells despite a total reduction in peripheral CD8⁺ T-cell count [14]. Furthermore, correlating with high serum IL-10 in severe/critically ill COVID-19 patients, the percentages of IFN-γ-producing effector CD4⁺ and CD8⁺ T cells can be increased in peripheral blood [6]. Third, as COVID-19 disease progresses, exhausted PD-1⁺TIM3⁺CD8⁺ T cells in the peripheral blood of patients have been found to increase, and these correlate with serum IL-10 concentrations in COVID-19 patients, suggesting a role of IL-10 in T cell exhaustion, presumably via overactivation and proliferation [5]. Such immune features in severe/critically ill COVID-19 patients with highly elevated systemic IL-10 have led us to speculate that IL-10 might play a pathological role in COVID-19 disease progression. However, this hypothesis remains to be rigorously tested.

In this scenario, how might elevated IL-10 contribute to COVID-19 mortality – if at all? The immunopathological pathway leading to patient death following SARS-CoV-2 infection can be divided into three stages: initiation, amplification, and consummation [13]. We propose that early induction of IL-10 upon SARS-CoV-2 infection during the initiation phase in the lung might indeed represent a negative feedback mechanism that serves as a countermeasure to inflammation caused by other proinflammatory mediators. However, as endogenous IL-10 production increases, we speculate that it might function as an immune activating/proinflammatory agent that stimulates the production of other mediators of the cytokine storm (Figure 1). As reported for human endotoxemia [12], IL-10 might also amplify the viral sepsis-related hyperinflammation observed in some severe/critically ill COVID-19 patients [15]. Because IL-10 directly expands cytotoxic effector CD8⁺ T cells in human studies, hyperactivation of adaptive immunity in COVID-19 patients might contribute to exacerbating disease severity. Although this possibility remains conjectural, we posit that the combined effects of IL-10 in promoting systemic inflammatory cytokine production and stimulating T cell activation and proliferation in COVID-19 patients might contribute to a lethal immunopathological process.

Currently, clinical trials are underway to test the therapeutic efficacy of blocking agents, either alone or in combination with more than ten inflammatory mediators reported as highly elevated in COVID-19 patients presenting a cytokine storm (discussed in [13]). Preliminary results from using blocking/neutralizing antibodies against IL-6/IL-6R, GM-CSF, and IL-1 suggest that further improvement is necessary to lower mortality in COVID-19 patients [13]. By attempting to block its pathological proinflammatory function, we suggest that IL-10 might constitute a potential target to reduce COVID-19 mortality. As such, the timing of blocking IL-10 activity in severe/critically ill COVID-19 patients might be crucial [13]. We propose that using a neutralizing antibody to block IL-10 to limit its potential immune-activating effects in the initiation phase of COVID-19 may be worth testing. Furthermore, we argue that combinatorial targeting of multiple proinflammatory mediators including IL-10, chemokines, IL-6, and IL-1 might be necessary to substantially reduce mortality in severe/critically ill COVID-19 patients. Evidently, the potential roles of systemically elevated IL-10 in COVID-19 pathogenesis and putative treatments warrant robust experimental validation, but certainly merit further attention.

Interleukin 10 (IL-10)

has many different and sometimes contradictory functions. It suppresses and promotes inflammation as well as innate and adaptive immune responses in a context- and dose-dependent manner.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605819/

 

Studies reveal how plague disables immune system, and how to exploit the process to make a vaccine

July 28, 2005

Two studies by researchers at the University of Chicago show how the bacteria that cause the plague manage to outsmart the immune system and how, by slightly altering one of the microbe's tools, the researchers produced what may be the first safe and effective vaccine.

Both papers--one published online July 28, 2005 in Science Express and one in the August issue of Infection and Immunity--focus on aspects of the type-III pathway, a molecular syringe that Yersinia pestis, the bacterium that may have killed more people throughout history than any other infectious disease, uses to disable its host's immune system.

"Yersinia pestis is the nastiest thing alive," said study author Olaf Schneewind, MD, PhD, professor and chairman of microbiology at the University of Chicago and director of the Great Lakes Regional Center of Excellence in Biodefense and Emerging Infectious Diseases Research (GLRCE). "It's the most virulent bacterial organism known to mankind. But we now know a little more about how it exercises those powers and we think we can use that knowledge to prepare a preemptive strike."

Historically, the terms "plague" or "Black Death" have referred to the bubonic plague, caused by Yersinia pestis and spread by the bites of infected fleas, which acquire the germ from infected rodents. In the mid-14th century, the plague swept through Europe killing nearly one-third of the population. It returned with a slightly reduced death count about once a generation for centuries.

Although far less common now, the plague has not entirely gone away. There are fewer than 2,000 cases a year worldwide, including 10 to 20 each year in the western United States. One out of seven persons infected dies, even with aggressive treatment.

Since 2001, however, many people have worried that terrorists could exploit Y. pestis as a weapon, spreading it widely and rapidly as an aerosol rather than through fleabites and rodents. Contracted this way--infecting the lungs rather than the bloodstream--the disease is known as pneumonic plague. This form of the infection progresses faster, spreads easier from person to person, and is far more deadly, killing 100 percent of those who do not receive the right antibiotics soon after exposure. "There is," the authors note, "an urgent need for vaccine development."

One of this microbe's enduring mysteries has been how it gains a foothold in the host without triggering a protective immune response. In the Science Express paper, Schneewind and colleagues show how Y. pestis annihilates the first line of defense in the host's immune system before it can generate a full response.

The researchers infected mice with Y. pestis. Two to three days later they harvested cells from organs where the bacteria tend to cluster. They used a dye to stain those cells green.

When Y. pestis attacks a cell it uses the type-III pathway--a needle-like projection--to inject various toxins into the cell, killing it. The researchers endowed these bacteria with an additional enzyme, which the microbes also injected in cells. This enzyme can snip the green dye into two pieces. When that happens, those cells, when exposed to fluorescent light, glow blue instead of green.

This technique enabled the team to identify the cell types targeted by the bacteria. Two days after the mice were infected, their spleens were filled with bacteria. Although the overwhelming majority of immune cells in the spleen are B cells or T cells, nearly all of the infected cells were macrophages, neutrophils, or dendritic cells.

These cells make up what immunologists call the "innate" immune system. They are the first to respond to a bacterial invasion. Their role is to rush to the infection site, engulf the bacteria, chew them up into smaller pieces and present those pieces to the T and B cells--the "adaptive" immune system--which enter the fray more slowly but bring powerful and very specific weapons targeted at those individual pieces.

"This is a very clever system for this particular kind of bacteria," said Schneewind. It can take eight to 10 days for the B and T cells to multiply and fully engage. "By that time, with plague," he said, "the host is dead."

The bacteria's Achilles heel, however, may be a protein called LcrV, which Y. pestis transports through the needle and uses to inject its toxins. LcrV plays two roles. It helps the needle to penetrate the membrane surrounding the target cell. It also suppresses the immune response. LcrV causes affected cells to release 40 times the normal levels of interleukin 10 (IL-10), which dampens down the immune response. LcrV also prevents secretion of tumor necrosis factor (TNF), which causes inflammation.

"LcrV is secreted in massive amounts via the type-III pathway during an infection," Schneewind said. "Without it, the bacteria are relatively harmless."

Consequently, researchers have tried to use LcrV alone as a vaccine. Unfortunately, because it suppresses the immune system, immunization with this molecule may be harmful.

Schneewind and colleagues, however, tested 11 truncated versions of LcrV, snipping out, from different locales, 30 of the protein's 326 amino acids in hopes of eliminating the elements that suppressed the immune response but retaining enough of the normal protein's structure to generate protective antibodies.

Out of 11 altered versions they found one that met both criteria. In mouse and human macrophages, version rV10, missing amino acids 271 through 300, triggered only small amounts of IL-10 and had little effect on TNF secretion. Mice immunized twice over six weeks with rV10 developed antibodies that protected them from many times the lethal dose of the bacteria.

"Our data, the authors conclude, "provide the first evidence of plague vaccines that do not suppress innate immune responses … and that may be useful for plague vaccination in animals, and, perhaps, humans." The next steps include testing in other animal models, said Schneewind.

The two papers combined, Schneewind suggested, are a good example of how, in this era of heightened awareness, "we can use modern tools to learn new things about an ancient scourge, and to prepare for the possible re-emergence of diseases we would like to forget, but better not."

The National Institute of Allergy and Infectious Disease funded this research. Additional authors of the Science Express paper were Melanie Marketon, William DePaolo, Kristin DeBord and Bana Jabri of the University of Chicago. Authors of the Infection and Immunity paper include Katie Overheim, William DePaolo, Kristin DeBord, Elizabeth Morrin, Debra Anderson, Nathaniel Green and Bana Jabri of the University of Chicago, and Robert Brubaker of Michigan State University.

  https://www.uchicagomedicine.org/forefront/news/studies-reveal-howplague-disables-immune-system-and-how-to-exploit-the-process-to-make-a-vaccine

 

 In molecular biology, LcrV is a protein found in Yersinia pestis and several other bacterial species. It forms part of the Yersinia pestis virulence protein factors that also includes all Yops, or Yersinia outer protein, but the name has been kept out of convention. LcrV's main function is not actually known, but it is essential for the production of other Yops.

The type III secretion system of Gram-negative bacteria is used to transport virulence factors from the pathogen directly into the host cell and is only triggered when the bacterium comes into close contact with the host.^[1] Effector proteins secreted by the type III system do not possess a secretion signal, and are considered unique because of this. Yersinia spp. secrete effector proteins called YopB and YopD that facilitate the spread of other translocated proteins through the type III needle and the host cell cytoplasm.^[2] In turn, the transcription of these moieties is thought to be regulated by another gene, lcrV, found on the Yops virulon that encodes the entire type III system.^[3] The product of this gene, LcrV protein, also regulates the secretion of YopD through the type III translocon, and itself acts as a protective "V" antigen for Yersinia pestis, the causative agent of plague.^[4][5]

A homologue of the Y. pestis LcrV protein, PcrV, has been found in Pseudomonas aeruginosa, an opportunistic pathogen. In vivo studies using mice found that immunisation with the protein protected burned animals from infection by P. aeruginosa, and enhanced survival. In addition, it is speculated that PcrV determines the size of the needle pore for type III secreted effectors.^[6]

LcrV is a multifunctional protein that has been shown to act at the level of secretion control by binding the Ysc inner-gate protein LcrG and to modulate the host immune response by altering cytokine production. LcrV is also necessary for full induction of low-calcium response (LCR) stimulon virulence gene transcription.^[7][8]

The polypeptide is encoded on a plasmid and is only present when the surroundings are around 37^o Celsius

https://en.wikipedia.org/wiki/LcrV

 

 Infect Immun. 2005 Aug; 73(8): 5152–5159.

doi: 10.1128/IAI.73.8.5152-5159.2005

PMCID: PMC1201268

PMID: 16041032

LcrV Plague Vaccine with Altered Immunomodulatory Properties

Katie A. Overheim,¹ R. William DePaolo,² Kristin L. DeBord,¹ Elizabeth M. Morrin,¹ Debra M. Anderson,¹ Nathaniel M. Green,² Robert R. Brubaker,³ Bana Jabri,² and Olaf Schneewind^1,*

Author information Article notes Copyright and License information Disclaimer

This article has been cited by other articles in PMC.

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Abstract

Yersinia pestis, the causative agent of plague, secretes LcrV (low-calcium-response V or V antigen) during infection. LcrV triggers the release of interleukin 10 (IL-10) by host immune cells and suppresses proinflammatory cytokines such as tumor necrosis factor alpha and gamma interferon as well as innate defense mechanisms required to combat the pathogenesis of plague. Although immunization of animals with LcrV elicits protective immunity, the associated suppression of host defense mechanisms may preclude the use of LcrV as a human vaccine. Here we show that short deletions within LcrV can reduce its immune modulatory properties. An LcrV variant lacking amino acid residues 271 to 300 (rV10) elicited immune responses that protected mice against a lethal challenge with Y. pestis. Compared to full-length LcrV, rV10 displayed a reduced ability to release IL-10 from mouse and human macrophages. Furthermore, the lipopolysaccharide-stimulated release of proinflammatory cytokines by human or mouse macrophages was inhibited by full-length LcrV but not by the rV10 variant. Thus, it appears that LcrV variants with reduced immune modulatory properties could be used as a human vaccine to generate protective immunity against plague.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1201268/

Vaccine. Author manuscript; available in PMC 2013 Jan 8.

Published in final edited form as:

Vaccine. 2011 Sep 2; 29(38): 6572–6583.

Published online 2011 Jul 16. doi: 10.1016/j.vaccine.2011.06.119

PMCID: PMC3538834

NIHMSID: NIHMS313602

PMID: 21763383

Prevention of pneumonic plague in mice, rats, guinea pigs and non-human primates with clinical grade rV10, rV10-2 or F1-V vaccines

Lauriane E. Quenee, Nancy A. Ciletti, Derek Elli, Timothy M. Hermanas, and Olaf Schneewind^*

Abstract

Yersinia pestis causes plague, a disease with high mortality in humans that can be transmitted by fleabite or aerosol. A US Food and Drug Administration (FDA)-licensed plague vaccine is currently not available. Vaccine developers have focused on two subunits of Y. pestis: LcrV, a protein at the tip of type III secretion needles, and F1, the fraction 1 pilus antigen. F1-V, a hybrid generated via translational fusion of both antigens, is being developed for licensure as a plague vaccine. The rV10 vaccine is a non-toxigenic variant of LcrV lacking residues 271–300. Here we developed Current Good Manufacturing Practice (cGMP) protocols for rV10. Comparison of clinical grade rV10 with F1-V did not reveal significant differences in plague protection in mice, guinea pigs or cynomolgus macaques. We also developed cGMP protocols for rV10-2, a variant of rV10 with an altered affinity tag. Immunization with rV10-2 adsorbed to aluminum hydroxide elicited antibodies against LcrV and conferred pneumonic plague protection in mice, rats, guinea pigs, cynomolgus macaques and African Green monkeys. The data support further development of rV10-2 for FDA Investigational New Drug (IND) authorization review and clinical testing.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3538834/

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Complete Protection Against Yersinia pestis in ... - PubMed

https://pubmed.ncbi.nlm.nih.gov › ...

by W Zhang · 2022 — Complete Protection Against Yersinia pestis in BALB/c Mouse Model Elicited by Immunization With Inhalable Formulations of rF1-V10 Fusion Protein ...

                             ORIGINAL RESEARCH article

Front. Immunol., 26 January 2022 | https://doi.org/10.3389/fimmu.2022.793382

Complete Protection Against Yersinia pestis in BALB/c Mouse Model Elicited by Immunization With Inhalable Formulations of rF1-V10 Fusion Protein via Aerosolized Intratracheal Inoculation

Wei Zhang†, Xiaolin Song†, Lina Zhai, Jianshu Guo, Xinying Zheng, Lili Zhang, Meng Lv, Lingfei Hu, Dongsheng Zhou, Xiaolu Xiong* and Wenhui Yang*

• State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, China

Pneumonic plague, caused by Yersinia pestis, is an infectious disease with high mortality rates unless treated early with antibiotics. Currently, no FDA-approved vaccine against plague is available for human use. The capsular antigen F1, the low-calcium-response V antigen (LcrV), and the recombinant fusion protein (rF1-LcrV) of Y. pestis are leading subunit vaccine candidates under intense investigation; however, the inability of recombinant antigens to provide complete protection against pneumonic plague in animal models remains a significant concern. In this study, we compared immunoprotection against pneumonic plague provided by rF1, rV10 (a truncation of LcrV), and rF1-V10, and vaccinations delivered via aerosolized intratracheal (i.t.) inoculation or subcutaneous (s.c.) injection. We further considered three vaccine formulations: conventional liquid, dry powder produced by spray freeze drying, or dry powder reconstituted in PBS. The main findings are: (i) rF1-V10 immunization with any formulation via i.t. or s.c. routes conferred 100% protection against Y. pestis i.t. infection; (ii) rF1 or rV10 immunization using i.t. delivery provided significantly stronger protection than rF1 or rV10 immunization via s.c. delivery; and (iii) powder formulations of subunit vaccines induced immune responses and provided protection equivalent to those elicited by unprocessed liquid formulations of vaccines. Our data indicate that immunization with a powder formulation of rF1-V10 vaccines via an i.t. route may be a promising vaccination strategy for providing protective immunity against pneumonic plague.

1 Introduction

Yersinia pestis, a non-motile, facultative intracellular, gram-negative bacterium, is the causative agent of plague (1–3). Plague, a devastating zoonotic disease prevalent in many parts of the world, is transmitted through infected fleas from rodent reservoirs to humans (4, 5). It is estimated to have claimed over 200 million human lives during the course of three major human pandemics (6). The most recent outbreak in Madagascar (2017–2018) resulted in ~2400 cases and ~200 deaths, raising global concerns (4).The control of human plague outbreaks relies mainly on the rapid confirmation of the diagnosis, isolation and treatment of confirmed and suspected cases (5). For confirmed patients, effective antibiotics (such as tetracycline, streptomycin, chloramphenicol) and preventive therapies must be administered within 24 hours of the onset of symptoms (7). Especially, pneumonic plague transmitting through aerosol droplets is the most dangerous among the three primary clinical forms due to its rapid onset and progression (6, 8). Without the rapid response with appropriate antibiotics, the fatality rate of pneumonic plague approaches 100% (9). Moreover, Y. pestis remains listed as a Tier 1 Select Agent because of its potential use as a biological weapon in an aerosolized form, making it an urgent public health and safety priority (10, 11). Therefore, development of a protective vaccine that provides both rapid and long-lasting immunity in the event of mass exposure to aerosolized Y. pestis is of great interest.

Historically, killed whole-cell vaccines (KWCVs) and live whole-cell vaccines (LWCVs) have been successfully used to protect humans against plague in parts of the world (12). To prepare KWCVs, Y. pestis were inactivated by heating or with chemicals. These vaccines evoked immunity against bubonic plague but were inefficient against pneumonic plague in animal models (13, 14). KWCVs are no longer used due to questionable efficacy and considerable reactogenicity. LWCVs were prepared from fully virulent strains of Y. pestis after multiple passages. The former Soviet Union and other nations still use LWCVs for human vaccination, e.g., the NIIEG line of the pgm-negative strain EV76. LWCVs are able to protect humans against bubonic and pneumonic plague (15–17), however, these vaccines are associated with several adverse effects, and fail to provide long-term immunity (1, 18). In addition, safety concerns have limited enthusiasm for the development of LWCVs and they are only recommended in endemic areas (19).

The subunit vaccine provides the most promise as a plague vaccine. Development efforts for an effective subunit vaccine to pneumonic plague have focused on two primary antigens of Y. pestis, namely the capsular protein (F1) and the low calcium response protein (LcrV). Baker et al. (20) first purified F1 protein and demonstrated that a vaccine with F1 protected mice and rats from bubonic plague. However, the F1 vaccine candidate provided only 65-84% protection against pneumonic plague (21); vaccines based exclusively on F1 were ineffective against F1-negative Y. pestis, which may be as virulent as wild-type (WT) CO92 Y. pestis (22). Burrows (23) discovered that LcrV was an important virulence protein and subsequent studies confirmed it was a critical protective antigen against Y. pestis infection (19, 24, 25). Unfortunately, part of the LcrV protein, acid residues 271-300, partially suppresses host immune response by stimulating interleukin-10 (IL-10), which suppresses Th1 cells (26, 27); this limits its usefulness in vaccines. The combination of recombinant F1 and LcrV antigens (rF1-LcrV) has a good safety profile in various animal models (28, 29), elicits greater protection than either F1 or LcrV alone (30, 31), but rF1-LcrV does not confer complete protection for mice challenged with more than 255× LD₅₀ Y. pestis administered via inhalation (32–34). Most recently, plague vaccines based on the expression of protective antigens of Y. pestis in live vectors (bacterial or viral platform) were developed (35) but had obvious limitations. There is thus a need to continue research on subunit vaccine candidates, which require further modification to minimize shortcomings and elicit more robust immune protection against pneumonic plague.

Over the past few decades, pulmonary delivery of vaccines has received increasing attention due to its ability to recruit local immune responses of the bronchopulmonary mucosa in addition to the broader systemic immune response (36–38). In addition, administration of vaccines via the lungs shows better bioavailability and more rapid effectiveness than injection routes because of the lung’s large surface area, abundant blood flow, and highly permeable epithelium (39, 40). Currently, there are two formulations of inhalable vaccines: (i) liquid formulations that require a cold chain transport system to maintain vaccine potency; and (ii) powder formulations that have long-term stability at room temperature for storage and shipping (41, 42). Given its obvious advantages, the latter is attracting more attention for use in aerosolized intratracheal (i.t.) delivery of vaccines. For more than 70 years, the Y. pestis EV NIIEG strain has been used as a human plague vaccine in the former Soviet Union and confers protection against bubonic and pneumonic plague after administration via inhalation (16, 43, 44). However, the protection appears to be short-lived and the vaccine is highly reactogenic, limiting licensing of this vaccine for use in many parts of the world (1, 12, 45). The preparation of live Y. pestis dry powder is rarely reported in the literature, possibly because of bacterial viability being lost during preparation. Subunit vaccine candidates may thus prove a better option for inhalable powder. In this study, we improve the immunoprotection of subunit vaccines against pneumonic plague by preparing the rF1, rV10 (a truncation of LcrV), or rF1-V10 fusion protein using spray freeze drying (SFD) to generate dry powder with the adjuvant CpG for i.t. inoculation. We then explore the immunogenicity and protective efficacy of these three subunit vaccines in different formulations (liquid, powder and reconstituted powder) via i.t. and subcutaneous (s.c.) administration routes in a mouse model of Y. pestis i.t. infection. Our results demonstrate preclinical feasibility of using a powder formulation of rF1-V10 and the potential use of an alternative pulmonary delivery method.

https://www.frontiersin.org/articles/10.3389/fimmu.2022.793382/full

4 Discussion

Due to the sporadic outbreak of pneumonic plague and the threat should it be weaponized, developing effective vaccines against pneumonic plague is important. Compared with F1 or LcrV alone, a subunit vaccine based on a fusion of F1-LcrV proteins is a promising subunit vaccine (30, 31). Previous studies reported that immunization with the rF1-LcrV fusion protein through the s.c. route could fully protect mice only at 10× LD₅₀ virulent Y. pestis CO92 via intranasal (i.n.) challenge (34) and only partially protect mice at 70× LD₅₀ via inhalational challenge (58). The injectable vaccine also failed to adequately protect African green monkeys against aerosolized Y. pestis (19). Other reports have shown that rF1-LcrV immunization through the non-invasive i.n. route conferred 80-90% protection against 70 to 100× LD₅₀ of Y. pestis via inhalational challenge (58, 59). Finally, Jones et al. (32) demonstrated that immunization with Protollin-F1-V through an i.n. route elicits 80% protection against an aerosol challenge at 255× LD₅₀ of Y. pestis and 100% protection against 170× LD₅₀ of Y. pestis. Overall, these studies indicate that the route of inoculation affects vaccine effectiveness against pneumonic plague. Modifying the antigen, we created the rF1-V10 fusion protein and established a mouse model of i.t. delivery to investigate protective immunity against pneumonic plague. Our results demonstrate that i.t.-rF1-V10 immunization induces strong immune responses that lead to 100% protection against an aerosol challenge of high-dose Y. pestis strain 201. Moreover, i.t. inoculation is an improved non-invasive method for pulmonary delivery of vaccines in mice compared to conventional i.n. inoculation that has several disadvantages: a low residence time; mucociliary clearance of vaccines in the nose, throat and upper airways leads to an inefficient uptake of soluble antigens; the route is unsuitable for powder pulmonary delivery; and it is not possible to quantify the given dose (60–62). We propose that the i.t. immunization with inhalable rF1-V10 vaccine may provide an alternative, possibly better, vaccination strategy against pneumonic plague.

Compared to pulmonary delivery (i.n., i.t. etc.) of liquid vaccine, powder formulation delivered by i.t. route is more advantageous (63). The F1-V SFD powders reconstituted in water delivered by i.n. route provided at most 80% protection against bubonic plague, however, the immunization of powder formulation was not included in this previous study (64). We recently demonstrated the liquid formulation of EV76-B-SHUΔpla delivered by i.t. route represents an excellent live-attenuated vaccine candidate against pneumonic plague (54), but it is likely unsuitable for preparation of powder by SFD, which will affect survivability and overall fitness of the live bacteria. In the current study, we evaluated the protection efficacy of three subunit vaccines administrated with three different formulations via i.t. route. Our results demonstrate that protection against an aerosolized Y. pestis challenge conferred by i.t. delivery of vaccines in dry powder formulations is at least equivalent to that conferred by i.t. delivery of liquid formulations of the same vaccine. Potential advantages of dry powder over liquid vaccines include: (i) the powder formulations could eliminate the cold-chain requirement, thus considerably reducing the costs of storage and shipping (60); (ii) the improved antigen stability may enhance the immunity induced after vaccination (65); (iii) using excipients as bulking agent could increase the total amount of inhaled powders, which makes given dose more accurate (66). These advantages suggest that powder formulation immunization via i.t. can provide a promising improvement over the existing vaccine.

At the mucosal surfaces, the predominant immunoglobulin is secretory IgA (SIgA). SIgA-based protective mucosal immunity can prevent an infectious agent from entering the body and block microbial toxins from binding to, or affecting, epithelial and other target cells (67). Some studies suggest that specific SIgA plays a key role in neutralizing pathogens or toxins (68, 69). One limitation of this study is that we didn’t explain why s.c.-rF1-V10 vaccination conferred complete protection against a high dose Y. pestis challenge without inducing SIgA production. This result indicated that the role of specific SIgA in protecting against respiratory infection of Y. pestis needs to be further investigated, such as evaluating the survival rate of mice challenged with Y. pestis that had been preincubated with SIgA (70), or evaluating the efficacy of subunit vaccines via i.t. delivery in wild-type (WT) and IgA-deficient (IgA (-/-)) mice (71). Another limitation is that we didn’t evaluate the innate immune response in the early stage of i.t. immunization, which is supposed to be more efficient than that of s.c. immunization (72, 73).

Taken together, we have demonstrated that the rF1-V10 fusion protein vaccine can confer complete protection against a high dose aerosolized Y. pestis challenge and that i.t. delivery of vaccines can induce higher protection efficacy in mice compared to that of s.c. immunization. However, i.t. route might be only suitable for animal use. For human pulmonary delivery of dry powder vaccine, the widely accepted inhaled devices named dry powder inhalers (DPIs) are recommended, which are easy to administrate without the assistant of trained medical personnel (65, 74). The use of this alternative method of pulmonary delivery and powder vaccine formulations may directly benefit biodefense vaccination programs and, ultimately, facilitate mass vaccination.

https://www.frontiersin.org/articles/10.3389/fimmu.2022.793382/full

で希望の光が見えてきたので、世界で新型コロナペスト規制緩和を解除中だが・・・

2022年3月21日月曜日

中国東方航空機＠山王パークタワーがFell to EarthでMountain On Fire！

https://tokumei10.blogspot.com/2022/03/fell-to-earthmountain-on-fire.html

・・・ってな感じだったりして！（爆ｗｗｗｗｗｗｗｗｗｗｗｗ

2022年3月21日月曜日

白川伯王家＠源氏の名を語ってた平田篤胤＠平家＠まがいもんの流れを受け継ぐ731部隊由来のバイオ兵器

https://tokumei10.blogspot.com/2022/03/731_21.html

2022年3月21日月曜日

See the mice in their million hordes From Ibiza to the Norfolk Broads Rule Britannia is out of bounds To my mother, my dog, and clowns

https://tokumei10.blogspot.com/2022/03/ok.html

2020年2月12日水曜日

中国製キメラペスト菌流出 と 隠蔽目的のキメラコロナウイルス散布説と７３１部隊

http://tokumei10.blogspot.com/2020/02/blog-post_12.html

2022年3月11日金曜日

日本の731部隊の系譜@ウクライナ生物学研究所

https://tokumei10.blogspot.com/2022/03/731.html

2022年3月13日日曜日

日本人が知らない７３１部隊と満州派と河豚計画の真の黒幕がバレちゃってなみだ目の大阪なおみをベロニカ・クデルメトバ＠ロシアがフルボッコに・・・

http://tokumei10.blogspot.com/2020/02/blog-post_12.html