Strong Innate Immune Response Research Articles

Phase Ib/II study of imprime PGG and pembrolizumab in patients with previously treated advanced non-small cell lung cancer (NSCLC): BTCRC LUN 15-017.

Therapeutic strategies to engage anti-tumor innate immunity are still underdeveloped. Imprime PGG (imprime), a pathogen-associated molecular pattern (PAMP), through pattern recognition receptors, successfully illicit a broad-based innate immune response in preclinical models against various cancers. We aimed to study safety and efficacy of imprime in combination with pembrolizumab in advanced stage non-small cell lung cancer (NSCLC). We conducted an investigator-initiated, multi-institutional, single-arm, phase Ib/II trial in previously treated, advanced stage NSCLC patients. Primary endpoints were maximum-tolerated dose (MTD) of imprime for the phase Ib, and progression-free survival (PFS) for the phase II study (NCT03003468). All 33 eligible patients were included in the safety analysis. No dose-limiting toxicity was observed and the imprime dose of 4 mg/kg was determined as the MTD. Thirty patients treated at the MTD (phase Ib, 6; phase II, 24) were included in the efficacy analysis. Median length of follow-up was 10.8 months. Confirmed objective response rate was 10% [95% confidence interval (CI): 2-27%], with one complete and two partial responses. Median PFS was 2.6 months (95% CI: 1.4-7.0), and 6- and 12-month PFS rates were 37% and 17%, respectively. Median overall survival (OS) was 11.1 months, and 6- and 12-month OS rates were 75% and 46%. Univariate analysis was performed to assess the impact of age, sex, race, disease-stage, programmed death-ligand 1 (PD-L1) expression levels, and prior immunotherapy on PFS and OS. Of these, prior immunotherapy negatively influenced OS [hazard ratio (HR): 2.95, 95% CI: 1.21-7.24]. Overall, the combination was safe and tolerable. The combination of imprime and pembrolizumab is tolerable but did not improve the outcome of advanced stage NSCLC patients who previously progressed on anti-programmed death 1 (PD-1)/PD-L1 immunotherapies. Further investigation is needed to understand the effects of therapeutic PAMPs to mount a strong innate immune response against cancer.

Innate Immune Response Against Batai Virus, Bunyamwera Virus, and Their Reassortants.

Orthobunyaviruses (OBVs) represent a diverse group of RNA viruses, encompassing a progressively increasing number of arboviruses that cause disease in both humans and livestock. Yet, studies investigating these viruses remain scarce despite the critical importance of such knowledge for assessing their zoonotic potential. In this study, we conducted an evaluation of the early immune response against the understudied Batai virus (BATV), as well as the influence of reassortment with the Bunyamwera virus (BUNV) on this response. Using RNA sequencing of infected murine bone marrow-derived dendritic cells, complemented by qPCR assays, we assessed the innate immune response at the transcriptome level. Additionally, we extended the qPCR analysis by including human THP-1-derived dendritic cells and ovine SFT-R cells to identify differences across species. Our results provide the first evidence that BATV elicits a strong innate immune response compared to BUNV, which largely evades early detection. Reassortants exhibited intermediate phenotypes, although unique changes in the early immune response were found as well. These findings provide a starting point for a better understanding of the immune response to BATV. Furthermore, they raise the question of whether reassortment induces changes in the innate immune response that might contribute to the differences in pathogenicity between reassortant OBVs and their parental generations.

Seoul orthohantavirus evades innate immune activation by reservoir endothelial cells.

Pathogenic hantaviruses are maintained world-wide within wild, asymptomatic rodent reservoir hosts, with increasingly frequent human spillover infections resulting in severe hemorrhagic fever or cardio-pulmonary disease. With no approved therapeutics or vaccines, research has, until recently, focused on understanding the drivers of immune-mediated pathogenesis. An emerging body of work is now investigating the mechanisms that allow for asymptomatic, persistent infections of mammalian reservoir hosts with highly pathogenic RNA viruses. Despite limited experimental data, several hypotheses have arisen to explain limited or absent disease pathology in reservoir hosts. In this study, we directly tested two leading hypotheses: 1) that reservoir host cells induce a generally muted response to viral insults, and 2) that these viruses employ host-specific mechanisms of innate antiviral antagonism to limit immune activation in reservoir cells. We demonstrate that, in contrast to human endothelial cells which mount a robust antiviral and inflammatory response to pathogenic hantaviruses, primary Norway rat endothelial cells do not induce antiviral gene expression in response to infection with their endemic hantavirus, Seoul orthohantavirus (SEOV). Reservoir rat cells do, however, induce strong innate immune responses to exogenous stimulatory RNAs, type I interferon, and infection with Hantaan virus, a closely related hantavirus for which the rat is not a natural reservoir. We also find that SEOV-infected rat endothelial cells remain competent for immune activation induced by exogenous stimuli or subsequent viral infection. Importantly, these findings support an alternative model for asymptomatic persistence within hantavirus reservoir hosts: that efficient viral replication within reservoir host cells may prevent the exposure of critical motifs for cellular antiviral recognition and thus limits immune activation that would otherwise result in viral clearance and/or immune-mediated disease. Defining the mechanisms that allow for infection tolerance and persistence within reservoir hosts will reveal novel strategies for viral countermeasures against these highly pathogenic zoonotic threats.

Next generation yellow fever vaccine induces an equivalent immune and transcriptomic profile to the current vaccine: observations from a phase I randomised clinical trial

Yellow fever (YF), a mosquito-borne acute viral haemorrhagic illness, is endemic to many tropical and subtropical areas of Africa and Central and South America. Vaccination remains the most effective prevention strategy; however, as repeated outbreaks have exhausted vaccine stockpiles, there is a need for improved YF vaccines to meet global demand. A live-attenuated YF vaccine candidate (referred to as vYF) cloned from a YF-17D vaccine (YF-VAX®) sub-strain, adapted for growth in Vero cells cultured in serum-free media, is in clinical development. We report the innate and adaptive immune responses and the transcriptome profile of selected genes induced by vYF. Healthy adults aged 18-60 years were randomised at a 1:1:1:1 ratio to receive one dose of vYF at 4, 5 or 6 Log CCID50 or YF-VAX (reference vaccine), administered subcutaneously in the upper arm (ClinicalTrials.gov identifier: NCT04142086). Blood/serum samples were obtained at scheduled time points through 180 days (D180) post-vaccination. The surrogate endpoints assessed were: serum cytokine/chemokine concentrations, measured by bead-based Multiplex assay; peripheral blood vYF-specific IgG and IgM memory B cell frequencies, measured by FluoroSpot assay; and expression of genes involved in the immune response to YF-17D vaccination by RT-qPCR. There was no increase in any of the cytokine/chemokine concentrations assessed through D14 following vaccination with vYF or YF-VAX, except for a slight increase in IP-10 (CXCL10) levels. The gene expression profiles and kinetics following vaccination with vYF and YF-VAX were similar, inclusive of innate (antiviral responses [type-1 interferon, IFN signal transduction; interferon-stimulated genes], activated dendritic cells, viral sensing pattern recognition receptors) and adaptive (cell division in stimulated CD4+ T cells, B cell and antibody) immune signatures, which peaked at D7 and D14, respectively. Increases in vYF-specific IgG and IgM memory B cell frequencies at D28 and D180 were similar across the study groups. vYF-induced strong innate and adaptive immune responses comparable to those induced by YF-VAX, with similar transcriptomic and kinetic profiles observed. Sanofi.

Fitness adaptations of Japanese encephalitis virus in pigs following vector-free serial passaging.

Japanese encephalitis virus (JEV) is a zoonotic mosquito-transmitted Flavivirus circulating in birds and pigs. In humans, JEV can cause severe viral encephalitis with high mortality. Considering that vector-free direct virus transmission was observed in experimentally infected pigs, JEV introduction into an immunologically naïve pig population could result in a series of direct transmissions disrupting the alternating host cycling between vertebrates and mosquitoes. To assess the potential consequences of such a realistic scenario, we passaged JEV ten times in pigs. This resulted in higher in vivo viral replication, increased shedding, and stronger innate immune responses in pigs. Nevertheless, the viral tissue tropism remained similar, and frequency of direct transmission was not enhanced. Next generation sequencing showed single nucleotide deviations in 10% of the genome during passaging. In total, 25 point mutations were selected to reach a frequency of at least 35% in one of the passages. From these, six mutations resulted in amino acid changes located in the precursor of membrane, the envelope, the non-structural 3 and the non-structural 5 proteins. In a competition experiment with two lines of passaging, the mutation M374L in the envelope protein and N275D in the non-structural protein 5 showed a fitness advantage in pigs. Altogether, the interruption of the alternating host cycle of JEV caused a prominent selection of viral quasispecies as well as selection of de novo mutations associated with fitness gains in pigs, albeit without enhancing direct transmission frequency.

Evaluation of a novel intramuscular prime/intranasal boost vaccination strategy against influenza in the pig model.

Live-attenuated influenza vaccines (LAIV) offer advantages over the commonly used inactivated split influenza vaccines. However, finding the optimal balance between sufficient attenuation and immunogenicity has remained a challenge. We recently developed an alternative LAIV based on the 2009 pandemic H1N1 virus with a truncated NS1 protein and lacking PA-X protein expression (NS1(1-126)-ΔPAX). This virus showed a blunted replication and elicited a strong innate immune response. In the present study, we evaluated the efficacy of this vaccine candidate in the porcine animal model as a pertinent in vivo system. Immunization of pigs via the nasal route with the novel NS1(1-126)-ΔPAX LAIV did not cause disease and elicited a strong mucosal immune response that completely blocked replication of the homologous challenge virus in the respiratory tract. However, we observed prolonged shedding of our vaccine candidate from the upper respiratory tract. To improve LAIV safety, we developed a novel prime/boost vaccination strategy combining primary intramuscular immunization with a haemagglutinin-encoding propagation-defective vesicular stomatitis virus (VSV) replicon, followed by a secondary immunization with the NS1(1-126)-ΔPAX LAIV via the nasal route. This two-step immunization procedure significantly reduced LAIV shedding, increased the production of specific serum IgG, neutralizing antibodies, and Th1 memory cells, and resulted in sterilizing immunity against homologous virus challenge. In conclusion, our novel intramuscular prime/intranasal boost regimen interferes with virus shedding and transmission, a feature that will help combat influenza epidemics and pandemics.

Virus-Mimetic Extracellular-Vesicle Vaccine Boosts Systemic and Mucosal Immunity via Immune Recruitment.

Mucosal vaccines can prevent viruses from infecting the respiratory mucosa, rather than only curtailing infection and protecting against the development of disease symptoms. The SARS-CoV-2 spike receptor-binding domain (RBD) is a compelling vaccine target but is undermined by suboptimal mucosal immunogenicity. Here, we report a SARS-CoV-2-mimetic extracellular-vesicle vaccine developed using genetic engineering and dendritic cell membrane budding. After mucosal immunization, the vaccine recruits antigen-presenting cells rapidly initiating a strong innate immune response. Notably, it obviates the need for adjuvants and can induce germinal center formation through both intramuscular and intratracheal vaccination. It not only elicits high levels of RBD-specific antibodies but also stimulates extensive cellular immunity in the respiratory mucosa. A sequential immunization strategy, starting with an intramuscular injection followed by an intratracheal booster, significantly bolsters mucosal immunity with high levels of IgA and tissue-resident memory T cell responses, thereby establishing a formidable defense against pseudovirus infection.

Effects of dam metabolic profile and seasonality (Spring vs. Winter) on their offspring' metabolism, health, and immunity: maternal factors in dairy calves' analytes.

Maternal status during the transition period can significantly impact the health and performance of Holstein dairy calves, with lasting effects on various variables. However, the relationship between maternal late gestation metabolic status, seasonality, and their impact on offspring remains unclear. This study aimed to assess the influence of maternal variables at calving on the performance, metabolism, and immunity of 28 dairy calves during their first month of life. Blood samples were collected from 28 Holstein cows at calving. Median results for maternal variables including non-esterified fatty acids (NEFA), β-hydroxybutyrate (BHB), glucose, total protein (TP), albumin, triglycerides (TG), total cholesterol (TC), haptoglobin (Hp), body weight (BW), and body condition score (BCS) were determined. These median values served as a basis for categorizing the offspring into two groups based on their dams' high or low degree of each maternal variable. Additionally, calves were categorized by the season of birth (Spring vs. Winter), with 14 in each. Blood samples were collected from the calves at birth and on days 1, 7, 14, and 28 to assess IgG, biochemical parameters, and haptoglobin concentration. Reactive oxygen species (ROS) production by polymorphonuclear cells stimulated by various agents was also evaluated. Clinical assessments were conducted for diarrhea and bovine respiratory disease frequencies. Despite the overall health of the cows, differences were observed in the calves between maternal groups. Heavier cows with high maternal BCS tended to have larger offspring, while high maternal BCS was associated with increased diarrhea prevalence. Low maternal BCS resulted in a stronger innate immune response, indicated by higher ROS production. Calves from cows experiencing metabolic changes during calving displayed elevated Hp concentrations. Spring-born calves were larger but had lower serum IgG concentration and reduced innate immune response compared to winter-born calves. Additionally, spring-born calves exhibited higher Hp and increased diarrhea prevalence on day 28. These findings underscore the importance of the prenatal period in determining neonatal health and suggest further research to elucidate the long-term clinical implications of maternal effects on offspring health and growth. Investigating offspring constituents later in life can provide insight into the persistence of maternal effects over time.

Bacterial cell volume regulation and the importance of cyclic di-AMP.

SUMMARYNucleotide-derived second messengers are present in all domains of life. In prokaryotes, most of their functionality is associated with general lifestyle and metabolic adaptations, often in response to environmental fluctuations of physical parameters. In the last two decades, cyclic di-AMP has emerged as an important signaling nucleotide in many prokaryotic lineages, including Firmicutes, Actinobacteria, and Cyanobacteria. Its importance is highlighted by the fact that both the lack and overproduction of cyclic di-AMP affect viability of prokaryotes that utilize cyclic di-AMP, and that it generates a strong innate immune response in eukaryotes. In bacteria that produce the second messenger, most molecular targets of cyclic di-AMP are associated with cell volume control. Besides, other evidence links the second messenger to cell wall remodeling, DNA damage repair, sporulation, central metabolism, and the regulation of glycogen turnover. In this review, we take a biochemical, quantitative approach to address the main cellular processes that are directly regulated by cyclic di-AMP and show that these processes are very connected and require regulation of a similar set of proteins to which cyclic di-AMP binds. Altogether, we argue that cyclic di-AMP is a master regulator of cell volume and that other cellular processes can be connected with cyclic di-AMP through this core function. We further highlight important directions in which the cyclic di-AMP field has to develop to gain a full understanding of the cyclic di-AMP signaling network and why some processes are regulated, while others are not.

Network-based integration of omics, physiological and environmental data in real-world Elbe estuarine Zander

Coastal and estuarine environments are under endogenic and exogenic pressures jeopardizing survival and diversity of inhabiting biota. Information of possible synergistic effects of multiple (a)biotic stressors and holobiont interaction are largely missing in estuaries like the Elbe but are of importance to estimate unforeseen effects on animals' physiology. Here, we seek to leverage host-transcriptional RNA-seq and gill mucus microbial 16S rRNA metabarcoding data coupled with physiological and abiotic measurements in a network analysis approach to decipher the impact of multiple stressors on the health of juvenile Sander lucioperca along one of the largest European estuaries. We find mesohaline areas characterized by gill tissue specific transcriptional responses matching osmosensing and tissue remodeling. Liver transcriptomes instead emphasized that zander from highly turbid areas were undergoing starvation which was supported by compromised body condition. Potential pathogenic bacteria, including Shewanella, Acinetobacter, Aeromonas and Chryseobacterium, dominated the gill microbiome along the freshwater transition and oxygen minimum zone. Their occurrence coincided with a strong adaptive and innate transcriptional immune response in host gill and enhanced energy demand in liver tissue supporting their potential pathogenicity. Taken together, we show physiological responses of a fish species and its microbiome to abiotic factors whose impact is expected to increase with consequences of climate change. We further present a method for the close-meshed detection of the main stressors and bacterial species with disease potential in a highly productive ecosystem.

Integrative genomic analysis of the lung tissue microenvironment in SARS-CoV-2 and NL63 patients

The coronavirus disease 2019 (COVID-19) pandemic caused by the SARS-CoV-2 virus has affected over 700 million people, and caused over 7 million deaths throughout the world as of April 2024, and continues to affect people through seasonal waves. While over 675 million people have recovered from this disease globally, the lingering effects of the disease are still under study. Long term effects of SARS-CoV-2 infection, known as 'long COVID,' include a wide range of symptoms including fatigue, chest pain, cellular damage, along with a strong innate immune response characterized by inflammatory cytokine production. Three years after the pandemic, data about long covid studies are finally emerging. More clinical studies and clinical trials are needed to understand and determine the factors that predispose individuals to these long-term side effects.In this methodology paper, our goal was to apply data driven approaches in order to explore the multidimensional landscape of infected lung tissue microenvironment to better understand complex interactions between viral infection, immune response and the lung microbiome of patients with (a) SARS-CoV-2 virus and (b) NL63 coronavirus. The samples were analyzed with several machine learning tools allowing simultaneous detection and quantification of viral RNA amount at genome and gene level; human gene expression and fractions of major types of immune cells, as well as metagenomic analysis of bacterial and viral abundance. To contrast and compare specific viral response to SARS-COV-2, we analyzed deep sequencing data from additional cohort of patients infected with NL63 strain of corona virus.Our correlation analysis of three types of RNA-seq based measurements in patients i.e. fraction of viral RNA (at genome and gene level), Human RNA (transcripts and gene level) and bacterial RNA (metagenomic analysis), showed significant correlation between viral load as well as level of specific viral gene expression with the fractions of immune cells present in lung lavage as well as with abundance of major fractions of lung microbiome in COVID-19 patients.Our methodology-based proof-of-concept study has provided novel insights into complex regulatory signaling interactions and correlative patterns between the viral infection, inhibition of innate and adaptive immune response as well as microbiome landscape of the lung tissue. These initial findings could provide better understanding of the diverse dynamics of immune response and the side effects of the SARS-CoV-2 infection and demonstrates the possibilities of the various types of analyses that could be performed from this type of data.

In Situ Synthesis of an Immune-Checkpoint Blocker from Engineered Bacteria Elicits a Potent Antitumor Response.

Immune-checkpoint blockade (ICB) reinvigorates T cells from exhaustion and potentiates T-cell responses to tumors. However, most patients do not respond to ICB therapy, and only a limited response can be achieved in a "cold" tumor with few infiltrated lymphocytes. Synthetic biology can be used to engineer bacteria as controllable bioreactors to synthesize biotherapeutics in situ. We engineered attenuated Salmonella VNP20009 with synthetic gene circuits to produce PD-1 and Tim-3 scFv to block immunosuppressive receptors on exhausted T cells to reinvigorate their antitumor response. Secreted PD-1 and Tim-3 scFv bound PD-1+ Tim-3+ T cells through their targeting receptors in vitro and potentiated the T-cell secretion of IFN-γ. Engineered bacteria colonized the hypoxic core of the tumor and synthesized PD-1 and Tim-3 scFv in situ, reviving CD4+ T cells and CD8+ T cells to execute an antitumor response. The bacteria also triggered a strong innate immune response, which stimulated the expansion of IFN-γ+ CD4+ T cells within the tumors to induce direct and indirect antitumor immunity.

Design and evaluation of a multi-epitope DNA vaccine against HPV16

ABSTRACT Cervical cancer, among the deadliest cancers affecting women globally, primarily arises from persistent infection with high-risk human papillomavirus (HPV). To effectively combat persistent infection and prevent the progression of precancerous lesions into malignancy, a therapeutic HPV vaccine is under development. This study utilized an immunoinformatics approach to predict epitopes of cytotoxic T lymphocytes (CTLs) and helper T lymphocytes (HTLs) using the E6 and E7 oncoproteins of the HPV16 strain as target antigens. Subsequently, through meticulous selection of T-cell epitopes and other necessary elements, a multi-epitope vaccine was constructed, exhibiting good immunogenic, physicochemical, and structural characteristics. Furthermore, in silico simulations showed that the vaccine not only interacted well with toll-like receptors (TLR2/TLR3/TLR4), but also induced a strong innate and adaptive immune response characterized by elevated Th1-type cytokines, such as interferon-gamma (IFN-γ) and interleukin-2 (IL2). Additionally, our study investigated the effects of different immunization intervals on immune responses, aiming to optimize a time-efficient immunization program. In animal model experiments, the vaccine exhibited robust immunogenic, therapeutic, and prophylactic effects. Administered thrice, it consistently induced the expansion of specific CD4 and CD8 T cells, resulting in substantial cytokines release and increased proliferation of memory T cell subsets in splenic cells. Overall, our findings support the potential of this multi-epitope vaccine in combating HPV16 infection and signify its candidacy for future HPV vaccine development.

Abstract 2461: Discovery of an orally available and liver-targeted ALPK1 agonist for cancer immunotherapy

Abstract Boosting anti-tumor immune responses can be achieved by activating the innate immune system. Despite the ongoing development of numerous STING and TLR agonists for this purpose, none of them have progressed to late-stage clinical evaluation. Alpha protein kinase-1 (ALPK1) is an intracellular innate immune receptor newly identified for its ability to detect ADP-heptose, a metabolite released during the biosynthesis and degradation of lipopolysaccharides (LPS). Since heptose is not naturally present in mammalian cells, the recognition of heptose metabolites by ALPK1 serves as a reliable mechanism to identify bacterial infections. When ADP-heptose binds to ALPK1, it initiates the phosphorylation of TIFA, thereby triggering a cascade of immune responses. Systemic administration with ADP-heptose induces robust ALPK1-dependent cytokine responses in mice. While high doses of ADP-heptose demonstrate anti-tumor effects, its instability and low in vivo potency hinder its clinical development. Through a SBDD medicinal chemistry effort, a novel, stable, and potent ADP-heptose analog, designated PTT-936, has been discovered and is progressing into clinical evaluation in cancer patients. PTT-936 exhibits remarkable nanomolar potencies for ALPK1 activation in various cellular assays. Notably, in human PBMCs, PTT-936 induces both concentration-dependent release of cytokines and activation of NK and T cells. Consistent with in vitro studies, oral administration of PTT-936 triggered significant cytokine responses in mice at microgram dose levels. However, overt toxicities were observed only at milligram dose levels, indicating a wide therapeutic window for this compound. In syngeneic tumor models, oral administration of PTT-936 induced dose-dependent tumor growth inhibition, which correlated with elevations in several cytokine biomarkers. Mechanistic studies show that PTT-936, working exclusively through the ALPK1 pathway, elicited strong innate and adaptive immune responses, as evidenced by robust cytokine induction and immune cell activation and recruitment. In syngeneic mouse models, the anti-tumor effects of PTT-936 were significantly hindered by either CD8+ T cell depletion or neutralization of select cytokines. Besides monotherapy studies, compelling evidence demonstrates strong synergistic anti-tumor effects when combining PTT-936 with immune checkpoint inhibitors. Intriguingly, PTT-936 exhibits favorable tolerability and a wider therapeutic index compared to certain clinical-stage STING and TLR agonists. Furthermore, PTT-936 does not enhance cytokine elevations induced by checkpoint inhibitors, which have been linked to adverse events (including CRS) observed in clinical combination studies. The dosing frequency of PTT-936 in the upcoming first-in-human clinical trial is carefully designed to prioritize both tolerability and patient compliance. Citation Format: Yupeng Wang, Nana Du, Xiaona Dong, Xuebing Sun, Yuan Hong, Xiaolong Yang, Zhijiang Ji, Qianyu Wang, Yun Lv, Zhichen Shi, Yajuan Gao, Xiaofu Hou, Yaxin Zhang, Juan Wang, Rui Mao, Yinxin Liu, Liang Li, Jingjin Ding, Hexiang Wang, Nan Hu, Counde O’Yang, Zhihong Li, Keith Bley, Feng Shao. Discovery of an orally available and liver-targeted ALPK1 agonist for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2461.

Transcriptomic analysis of intestinal organoids, derived from pigs divergent in feed efficiency, and their response to Escherichia coli

BackgroundThere is increasing interest in using intestinal organoids to study complex traits like feed efficiency (FE) and host-microbe interactions. The aim of this study was to investigate differences in the molecular phenotype of organoids derived from pigs divergent for FE as well as their responses to challenge with adherent and invasive Escherichia coli (E. coli).ResultsColon and ileum tissue from low and high FE pigs was used to generate 3D organoids and two dimensional (2D) monolayers of organoid cells for E. coli challenge. Genome-wide gene expression was used to investigate molecular differences between pigs that were phenotypically divergent for FE and to study the difference in gene expression after challenge with E. coli. We showed, (1) minor differences in gene expression of colon organoids from pigs with low and high FE phenotypes, (2) that an E. coli challenge results in a strong innate immune gene response in both colon and ileum organoids, (3) that the immune response seems to be less pronounced in the colon organoids of high FE pigs and (4) a slightly stronger immune response was observed in ileum than in colon organoids.ConclusionsThese findings demonstrate the potential for using organoids to gain insights into complex biological mechanisms such as FE.

Monkeypox virus: insights into pathogenesis and laboratory testing methods.

The monkeypox virus (MPXV) is a zoonotic pathogen that transmits between monkeys and humans, exhibiting clinical similarities with the smallpox virus. Studies on the immunopathogenesis of MPXV revealed that an initial strong innate immune response is elicited on viral infection that subsequently helps in circumventing the host defense. Once the World Health Organization (WHO) declared it a global public health emergency in July 2022, it became essential to clearly demarcate the MPXV-induced symptoms from other viral infections. We have exhaustively searched the various databases involving Google Scholar, PubMed, and Medline to extract the information comprehensively compiled in this review. The primary focus of this review is to describe the diagnostic methods for MPXV such as polymerase chain reaction (PCR), and serological assays, along with developments in viral isolation, imaging techniques, and next-generation sequencing. These innovative technologies have the potential to greatly enhance the accuracy of diagnostic procedures. Significant discoveries involving MPXV immunopathogenesis have also been highlighted. Overall, this will be a knowledge repertoire that will be crucial for the development of efficient monitoring and control strategies in response to the MPXV infection helping clinicians and researchers in formulating healthcare strategies.

Transcriptional Profiling of the Rabbit Liver Infected With Eimeria stiedae Reveals Dynamic Host Cell Responses During the Induction and Resolution of Cholangitis

Eimeria stiedae is one of the few eukaryotic pathogens that exclusively infect the liver and serves as a good model to study the host–pathogen interactions in this vital organ. In this study, we show that rabbits infected with E. stiedae develop severe but self‐healing cholangitis. RNA‐seq analysis of the liver gene expression landscapes over the long course of E. stiedae infection identified 912 differentially expressed genes (DEGs) in the prepatent period (794 up‐ and 118 downregulated genes), 2889 DEGs in the early oocyst shedding period (1870 up‐ and 1019 downregulated genes), 2859 DEGs in the peak oocyst shedding period (1923 up‐ and 936 downregulated genes), and 327 DEGs in the recovery period (164 up‐ and 163 downregulated genes). Combined with pathological observations, we identified dynamic changes in host–parasite interactions involving multiple pathways. They showed that E. stiedae infection induced full‐blown inflammatory, Th1 and Th17 immune responses at all time points. This was associated with the strong innate immune responses during the prepatent period, including increased Toll‐like and NOD‐like receptor signaling. Despite mounting several damage control and repair responses, such as PI3K‐Akt signaling, Ras signaling, and extracellular matrix‐receptor interactions, the liver underwent severe metabolic dysfunction, oxidative damage, and coagulopathy after patency and at peak infection, possibly as a result of suppressed peroxisome activities and downregulated PPAR signaling. These responses largely disappeared during late infection, suggesting that the liver self‐heals after severe cholangitis. These data provide new insights into host–pathogen interactions during Eimeria infection and improve our understanding of the pathogenesis of parasitic cholangitis.

Characterization of immortalized human podocytes infected with lentivirus as an in vitro model of viral infection-associated podocytopathy.

A large number of studies have shown the association of kidney disease with viral infections in the body. Viral infections cause kidney injury in two manners, the systemic inflammation (cytokine storm) and the direct infection of kidney cells. Concerning direct viral infection of podocytes, the mechanism underlying virus-induced podocyte injury remains largely unknown and requires effective experimental models to facilitate its study. Here, we performed molecular characterization of immortalized human podocyte cell line (HPC) infected with lentivirus by RNA-seq. Bioinformatics analysis revealed a strong innate immune response in the cells, including interferon production and signaling. Meanwhile, activations of ferroptosis pathway and TNF-alpha signaling were also found, consistent with an impaired viability of the cells. Lentiviral infection also upregulated expression of APOL1 as observed in patients with HIV associated nephropathy (HIVAN) and diabetic nephropathy (DN). Interestingly, when the lentiviral infected cells were treated with Adriamycin (ADR), the ADR-associated signaling pathways were not interfered and remained activated as that in the cells treated with ADR only, suggesting that the virus and ADR have distinct mechanisms in damaging podocytes. Thus, the lentivirus-infected HPC cells represent a useful in vitro model of viral infection-associated podocytopathy.

Secretome Analysis of High- and Low-Virulent Bovine Pasteurella multocida Cultured in Different Media

Bovine Pasteurella multocida (P. multocida) serotype A is one of the major causes of bovine respiratory disease (BRD). We used data-independent acquisition (DIA) LC-MS/MS combined with bioinformatics analysis to identify proteins secreted by P. multocida. A total of 154 proteins were obtained from the supernatants of two isolates of bovine P. multocida serotype A (high virulent PmCQ2 and low virulent PmCQ6) cultured in Martin or BHI media, of which 50 were identified as putative secreted proteins. Further studies showed that Tuf, an elongation factor Tu, was highly expressed in P. multocida and secreted into infected tissues. Tuf stimulated strong innate immune responses of macrophages and had protective efficacy against P. multocida infection in a mouse model. The results provide insight into the secreted proteins of P. multocida and suggest new targets for vaccine development against P. multocida.

Role of Glutathione in Neutrophil Chemotaxis in Periodontitis

Periodontitis is a common non-communicable inflammatory disease that leads to the destruction of periodontal tissues and tooth loss. Initiated by the plaque biofilm, there is a strong innate immune response with an abundance of neutrophils in the periodontium of affected individuals. Previous reports have shown that the intracellular concentration of glutathione in peripheral blood neutrophils from periodontitis patients and the chemotactic ability of these cells are compromised. Furthermore, other studies have described that in oxidative stress conditions neutrophil chemotaxis is aberrant and causes the glutathionylation of F-actin, a key player in chemotaxis. In this study, the effects of glutathione-modulating compounds were assessed in neutrophils isolated from healthy donors, showing that the perturbation of glutathione homeostasis decreases the chemotaxis of neutrophils. Following this, the intracellular glutathione status and chemotactic ability of neutrophils isolated from periodontitis patients was compared to that of age- and sex-matched controls. A decrease in glutathione and chemotactic ability were confirmed. Finally, the proteome of these neutrophils was explored, demonstrating a change in the abundance of proteins involved in glutathione homeostasis. Together these data suggest that peripheral blood neutrophils from periodontitis patients are compromised in their ability to cope with oxidative stress and move.