The ectoenzymes CD39 and CD73 degrade extracellular ATP to adenosine. ATP is released by stressed or damaged cells and provides pro-inflammatory signals to immune cells through P2 receptors. ...Adenosine, on the other hand, suppresses immune cells by stimulating P1 receptors. Thus, CD39 and CD73 can shape the quality of immune responses. Here we demonstrate that upregulation of CD39 is a consistent feature of activated conventional CD4+ and CD8+ T cells. Following stimulation in vitro, CD4+ and CD8+ T cells from human blood gained surface expression of CD39 but displayed only low levels of CD73. Activated human T cells from inflamed joints largely presented with a CD39+CD73- phenotype. In line, in spleens of mice with acute Listeria monocytogenes, listeria-specific CD4+ and CD8+ T cells acquired a CD39+CD73- phenotype. To test the function of CD39 in control of bacterial infection, CD39-deficient (CD39-/-) mice were infected with L. monocytogenes. CD39-/- mice showed better initial control of L. monocytogenes, which was associated with enhanced production of inflammatory cytokines. In the late stage of infection, CD39-/- mice accumulated more listeria-specific CD8+ T cells in the spleen than wildtype animals suggesting that CD39 attenuates the CD8+ T-cell response to infection. In conclusion, our results demonstrate that CD39 is upregulated on conventional CD4+ and CD8+ T cells at sites of acute infection and inflammation, and that CD39 dampens responses to bacterial infection.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The intracellular development and differentiation of the Plasmodium parasite in the host liver is a prerequisite for the actual onset of malaria disease pathology. Since liver-stage infection is ...clinically silent and can be completely eliminated by sterilizing immune responses, it is a promising target for urgently needed innovative antimalarial drugs and/or vaccines. Discovered more than 65 years ago, these stages remain poorly understood regarding their molecular repertoire and interaction with their host cells in comparison to the pathogenic erythrocytic stages. The differentiating and replicative intrahepatic parasite resides in a membranous compartment called the parasitophorous vacuole, separating it from the host-cell cytoplasm. Here we outline seminal work that contributed to our present understanding of the fundamental dynamic cellular processes of the intrahepatic malarial parasite with both specific host-cell factors and compartments.
The WHO declared the COVID-19 outbreak a public health emergency of international concern. The causative agent of this acute respiratory disease is a newly emerged coronavirus, named SARS-CoV-2, ...which originated in China in late 2019. Exposure to SARS-CoV-2 leads to multifaceted disease outcomes from asymptomatic infection to severe pneumonia, acute respiratory distress and potentially death. Understanding the host immune response is crucial for the development of interventional strategies. Humoral responses play an important role in defending viral infections and are therefore of particular interest. With the aim to resolve SARS-CoV-2-specific humoral immune responses at the epitope level, we screened clinically well-characterized sera from COVID-19 patients with mild and severe disease outcome using high-density peptide microarrays covering the entire proteome of SARS-CoV-2. Moreover, we determined the longevity of epitope-specific antibody responses in a longitudinal approach. Here we present IgG and IgA-specific epitope signatures from COVID-19 patients, which may serve as discriminating prognostic or predictive markers for disease outcome and/or could be relevant for intervention strategies.
Genome editing of malaria parasites is key to the generation of live attenuated parasites used in experimental vaccination approaches. DNA repair in Plasmodium generally occurs only through ...homologous recombination. This has been used to generate transgenic parasites that lack one to three genes, leading to developmental arrest in the liver and allowing the host to launch a protective immune response. While effective in principle, this approach is not safe for use in humans as single surviving parasites can still cause disease. Here we use zinc-finger nucleases to generate attenuated parasite lines lacking an entire chromosome arm, by a timed induction of a double-strand break. Rare surviving parasites also allow the investigation of unconventional DNA repair mechanisms in a rodent malaria parasite.
A single, zinc-finger nuclease-induced DNA double-strand break results in the generation of attenuated parasite lines that show varying degrees of developmental arrest, protection efficacy in an immunisation regime and safety, depending on the timing of zinc-finger nuclease expression within the life cycle. We also identify DNA repair by microhomology-mediated end joining with as little as four base pairs, resulting in surviving parasites and thus breakthrough infections.
Malaria parasites can repair DNA double-strand breaks with surprisingly small mini-homology domains located across the break point. Timely expression of zinc-finger nucleases could be used to generate a new generation of attenuated parasite lines lacking hundreds of genes.
Parasites causing malaria need to migrate in order to penetrate tissue barriers and enter host cells. Here we show that the actin filament-binding protein coronin regulates gliding motility in ...Plasmodium berghei sporozoites, the highly motile forms of a rodent malaria-causing parasite transmitted by mosquitoes. Parasites lacking coronin show motility defects that impair colonization of the mosquito salivary glands but not migration in the skin, yet result in decreased transmission efficiency. In non-motile sporozoites low calcium concentrations mediate actin-independent coronin localization to the periphery. Engagement of extracellular ligands triggers an intracellular calcium release followed by the actin-dependent relocalization of coronin to the rear and initiation of motility. Mutational analysis and imaging suggest that coronin organizes actin filaments for productive motility. Using coronin-mCherry as a marker for the presence of actin filaments we found that protein kinase A contributes to actin filament disassembly. We finally speculate that calcium and cAMP-mediated signaling regulate a switch from rapid parasite motility to host cell invasion by differentially influencing actin dynamics.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Early in the pandemic, we designed a SARS-CoV-2 peptide vaccine containing epitope regions optimized for concurrent B cell, CD4
T cell, and CD8
T cell stimulation. The rationale for this design was ...to drive both humoral and cellular immunity with high specificity while avoiding undesired effects such as antibody-dependent enhancement (ADE).
We explored the set of computationally predicted SARS-CoV-2 HLA-I and HLA-II ligands, examining protein source, concurrent human/murine coverage, and population coverage. Beyond MHC affinity, T cell vaccine candidates were further refined by predicted immunogenicity, sequence conservation, source protein abundance, and coverage of high frequency HLA alleles. B cell epitope regions were chosen from linear epitope mapping studies of convalescent patient serum, followed by filtering for surface accessibility, sequence conservation, spatial localization near functional domains of the spike glycoprotein, and avoidance of glycosylation sites.
From 58 initial candidates, three B cell epitope regions were identified. From 3730 (MHC-I) and 5045 (MHC-II) candidate ligands, 292 CD8
and 284 CD4
T cell epitopes were identified. By combining these B cell and T cell analyses, as well as a manufacturability heuristic, we proposed a set of 22 SARS-CoV-2 vaccine peptides for use in subsequent murine studies. We curated a dataset of ~ 1000 observed T cell epitopes from convalescent COVID-19 patients across eight studies, showing 8/15 recurrent epitope regions to overlap with at least one of our candidate peptides. Of the 22 candidate vaccine peptides, 16 (n = 10 T cell epitope optimized; n = 6 B cell epitope optimized) were manually selected to decrease their degree of sequence overlap and then synthesized. The immunogenicity of the synthesized vaccine peptides was validated using ELISpot and ELISA following murine vaccination. Strong T cell responses were observed in 7/10 T cell epitope optimized peptides following vaccination. Humoral responses were deficient, likely due to the unrestricted conformational space inhabited by linear vaccine peptides.
Overall, we find our selection process and vaccine formulation to be appropriate for identifying T cell epitopes and eliciting T cell responses against those epitopes. Further studies are needed to optimize prediction and induction of B cell responses, as well as study the protective capacity of predicted T and B cell epitopes.
Cerebral malaria is a life-threatening complication of Plasmodia infection and a major cause of child mortality in Sub-Saharan Africa. We report that protection from experimental cerebral malaria in ...the rodent model is obtained by a single intravenous or subcutaneous whole-parasite immunization. Whole-parasite immunization with radiation-attenuated sporozoites was equally protective as immunization with non-attenuated sporozoites under chemoprophylaxis. Both immunization regimens delayed the development of blood-stage parasites, but differences in cellular and humoral immune mechanisms were observed. Single-dose whole-parasite vaccination might serve as a relatively simple and feasible immunization approach to prevent life-threatening cerebral malaria.
Whole sporozoite vaccines represent one of the most promising strategies to induce protection against malaria. However, the development of efficient vaccination protocols still remains a major ...challenge. To understand how the generation of immunity is affected by variations in vaccination dosage and frequency, we systematically analyzed intrasplenic and intrahepatic CD8
T cell responses following varied immunizations of mice with radiation-attenuated sporozoites. By combining experimental data and mathematical modeling, our analysis indicates a reversing role of spleen and liver in the generation of protective liver-resident CD8
T cells during priming and booster injections: While the spleen acts as a critical source compartment during priming, the increase in vaccine-induced hepatic T cell levels is likely due to local reactivation in the liver in response to subsequent booster injections. Higher dosing accelerates the efficient generation of liver-resident CD8
T cells by especially affecting their local reactivation. In addition, we determine the differentiation and migration pathway from splenic precursors toward hepatic memory cells thereby presenting a mechanistic framework for the impact of various vaccination protocols on these dynamics. Thus, our work provides important insights into organ-specific CD8
T cell dynamics and their role and interplay in the formation of protective immunity against malaria.
During the clinically silent liver stage of a Plasmodium infection the parasite replicates from a single sporozoite into thousands of merozoites. Infection of humans and rodents with large numbers of ...sporozoites that arrest their development within the liver can cause sterile protection from subsequent infections. Disruption of genes essential for liver stage development of rodent malaria parasites has yielded a number of attenuated parasite strains. A key question to this end is how increased attenuation relates to vaccine efficacy. Here, we generated rodent malaria parasite lines that arrest during liver stage development and probed the impact of multiple gene deletions on attenuation and protective efficacy. In contrast to P. berghei strain ANKA LISP2(-) or uis3(-) single knockout parasites, which occasionally caused breakthrough infections, the double mutant lacking both genes was completely attenuated even when high numbers of sporozoites were administered. However, different vaccination protocols showed that LISP2(-) parasites protected better than uis3(-) and double mutants. Hence, deletion of several genes can yield increased safety but might come at the cost of protective efficacy.
Sterile protection against malaria infection can be achieved by the inoculation of intact sporozoites while treating concomitantly with the 4-aminoquinoline chloroquine. We present an analysis of ...protective immunity elicited by successive immunization with Plasmodium berghei sporozoites under chemoprophylaxis. Immunization resulted in a protective, stage-specific immune response. Protection appeared to be mediated by CD8(+) T cells and was abrogated upon their specific depletion. Adoptive transfer of splenocytes rendered recipient animals resistant to sporozoite infection, but not to blood-stage challenge. Immunization with sporozoites under chemoprophylaxis results in robust immunity, and the presence of blood-stage infection at sporozoite immunization had no downregulating effect on the protective immune response.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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