Decoding Toll-like receptors: Recent insights and perspectives in innate immunity.

Toll-like receptors (TLRs) are an evolutionarily conserved family in the innate immune system and are the first line of host defense against microbial pathogens by recognizing pathogen-associated molecular patterns (PAMPs). TLRs, categorized into cell surface and endosomal subfamilies, recognize diverse PAMPs, and structural elucidation of TLRs and PAMP complexes has revealed their intricate mechanisms. TLRs activate common and specific signaling pathways to shape immune responses. Recent studies have shown the importance of post-transcriptional regulation in TLR-mediated inflammatory responses. Despite their protective functions, aberrant responses of TLRs contribute to inflammatory and autoimmune disorders. Understanding the delicate balance between TLR activation and regulatory mechanisms is crucial for deciphering their dual role in immune defense and disease pathogenesis. This review provides an overview of recent insights into the history of TLR discovery, elucidation of TLR ligands and signaling pathways, and their relevance to various diseases.

[Molecular mechanisms underlying the pathogenesis of septic multiple organ failure].

Sepsis is defined as the body's overwhelming and life-threatening response to infection that can lead to tissue damage, organ failure, and death. Since bacterial infection is one of the main causes of sepsis, appropriate antimicrobial therapy remains the cornerstone of sepsis and septic shock management. However, since sepsis is a multifaceted chaos involving inflammation and anti-inflammation disbalance leading to the unregulated widespread release of inflammatory mediators, cytokines, and pathogen-related molecules leading to system-wide organ dysfunction, the whole body control to prevent the progression of organ dysfunction is needed. In sepsis and septic shock, pathogen-associated molecular patterns (PAMPs), such as bacterial exotoxins, cause direct cellular damage and/or trigger an immune response in the host. PAMPs are recognized by pattern recognizing receptors (PRRs) expressed on immune-reactive cells. PRRs are also activated by host nuclear, mitochondrial, and cytosolic proteins, known as damage-associated molecular patterns (DAMPs) that are released from cells during sepsis. Thus, most PRRs respond to PAMPs or DAMPs by triggering activation of transcriptional factors, NF-κB, AP1, and STAT-3. On the other hand, sepsis leads to immune (lymphocytes and macrophages) and nonimmune (endothelial and epithelial cells) cell death. Apoptosis has been the major focus of research on cell death in sepsis, but autophagy, necrosis, necroptosis, pyroptosis, NETosis, and ferroptosis may also play an important role in this critical situation. The recent development in our understanding regarding the cellular pathogenesis of sepsis will help in developing new treatment of sepsis.

Chimeric NOD2 Mincle Agonists as Vaccine Adjuvants.

There is a need for improved vaccine adjuvants to augment vaccine efficacy. One way to address this is by targeting multiple immune cell pathogen recognition receptors (PRRs) using chimeric pathogen-associated molecular patterns (PAMPs). Conjugation of the PAMPs will ensure codelivery of the immunostimulatory molecules to the same cell, enhancing adjuvant activity. The macrophage inducible C-type lectin (Mincle) is a promising PRR for adjuvant development; however, no effective chimeric Mincle adjuvants have been prepared. We addressed this by synthesizing Mincle adjuvant conjugates, MDP-C18Brar and MDP-C18Brar-dilipid, which contain PAMPs recognized by Mincle and the nucleotide-binding oligomerization domain 2 (NOD2). The two PAMPs are joined by a pH-sensitive oxyamine linker which, upon acidification at lysosomal pH, hydrolyzed to release the NOD2 ligands. The conjugates elicited the production of Th1 and Th17 promoting cytokines in vitro, and when using OVA as a model antigen, exhibited enhanced T-cell-mediated immune responses and reduced toxicity in vivo, compared to the coadministration of the adjuvants.

Transcriptomic profiling of immune cells in murine polymicrobial sepsis.

Introduction: Various immune cell types play critical roles in sepsis with numerous distinct subsets exhibiting unique phenotypes even within the same cell population. Single-cell RNA sequencing (scRNA-seq) enables comprehensive transcriptome profiling and unbiased cell classification. In this study, we have unveiled the transcriptomic landscape of immune cells in sepsis through scRNA-seq analysis. Methods: We induced sepsis in mice by cecal ligation and puncture. 20 h after the surgery, the spleen and peritoneal lavage were collected. Single-cell suspensions were processed using a 10× Genomics pipeline and sequenced on an Illumina platform. Count matrices were generated using the Cell Ranger pipeline, which maps reads to the mouse reference transcriptome, GRCm38/mm10. Subsequent scRNA-seq analysis was performed using the R package Seurat. Results: After quality control, we subjected the entire data set to unsupervised classification. Four major clusters were identified as neutrophils, macrophages, B cells, and T cells according to their putative markers. Based on the differentially expressed genes, we identified activated pathways in sepsis for each cell type. In neutrophils, pathways related to inflammatory signaling, such as NF-κB and responses to pathogen-associated molecular patterns (PAMPs), cytokines, and hypoxia were activated. In macrophages, activated pathways were the ones related to cell aging, inflammatory signaling, and responses to PAMPs. In B cells, pathways related to endoplasmic reticulum stress were activated. In T cells, activated pathways were the ones related to inflammatory signaling, responses to PAMPs, and acute lung injury. Next, we further classified each cell type into subsets. Neutrophils consisted of four clusters. Some subsets were activated in inflammatory signaling or cell metabolism, whereas others possessed immunoregulatory or aging properties. Macrophages consisted of four clusters, namely, the ones with enhanced aging, lymphocyte activation, extracellular matrix organization, or cytokine activity. B cells consisted of four clusters, including the ones possessing the phenotype of cell maturation or aging. T cells consisted of six clusters, whose phenotypes include molecular translocation or cell activation. Conclusions: Transcriptomic analysis by scRNA-seq has unveiled a comprehensive spectrum of immune cell responses and distinct subsets in the context of sepsis. These findings are poised to enhance our understanding of sepsis pathophysiology, offering avenues for targeting novel molecules, cells, and pathways to combat infectious diseases.

Different roles of Ca2+ and chitohexose in peanut (Arachis Hypogaea) photosynthetic responses to PAMP-immunity.

Background: During active infections, plants prevent further spread of pathogenic microorganisms by inducing the rapid programmed death of cells around the infection point. This phenomenon is called the hypersensitive response and is a common feature of plant immune responses. Plants recognize conserved structures of pathogenic microorganisms, called pathogen-associated molecular patterns (PAMPs), e.g., flagellin 22 (flg22) and chitohexose, which bind to receptors on plant cells to induce various immune-response pathways. Although abiotic stresses are known to alter photosynthesis, the different effects of flg22 and chitohexose, which are involved into PAMP-induced signaling, on photosynthesis needs further study. Methods: In the present study, we assessed the role of PAMPs in peanut (Arachis hypogaea) photosynthesis, particularly, the interaction between PAMPs and Ca2+ signal transduction pathway. Results: Both flg22 and chitohexose significantly promoted the expression of the pathogenesis-related genes PR-4 and PR-10, as did Ca2+. We found that Ca2+ is involved in downregulating the photosystem II (PSII) reaction center activity induced by the flg22 immune response, but the role of chitohexose is not obvious. Additionally, Ca2+ significantly reduced the non-photochemical energy dissipation in the flg22- and chitohexose-induced immune response. Conclusion: These results indicated that flg22 and chitohexose can trigger peanut immune pathways through the Ca2+ signaling pathway, but they differ in their regulation of the activity of the PSII reaction center.

Spatial IMA1 regulation restricts root iron acquisition on MAMP perception.

Iron is critical during host-microorganism interactions1-4. Restriction of available iron by the host during infection is an important defence strategy, described as nutritional immunity5. However, this poses a conundrum for externally facing, absorptive tissues such as the gut epithelium or the plant root epidermis that generate environments that favour iron bioavailability. For example, plant roots acquire iron mostly from the soil and, when iron deficient, increase iron availability through mechanisms that include rhizosphere acidification and secretion of iron chelators6-9. Yet, the elevated iron bioavailability would also be beneficial for the growth of bacteria that threaten plant health. Here we report that microorganism-associated molecular patterns such as flagellin lead to suppression of root iron acquisition through a localized degradation of the systemic iron-deficiency signalling peptide Iron Man 1 (IMA1) in Arabidopsis thaliana. This response is also elicited when bacteria enter root tissues, but not when they dwell on the outer root surface. IMA1 itself has a role in modulating immunity in root and shoot, affecting the levels of root colonization and the resistance to a bacterial foliar pathogen. Our findings reveal an adaptive molecular mechanism of nutritional immunity that affects iron bioavailability and uptake, as well as immune responses.

Retrotransposon-derived transcripts and their functions in immunity and disease.

Retrotransposons, which account for approximately 42% of the human genome, have been increasingly recognized as "non-self" pathogen-associated molecular patterns (PAMPs) due to their virus-like sequences. In abnormal conditions such as cancer and viral infections, retrotransposons that are aberrantly expressed due to impaired epigenetic suppression display PAMPs, leading to their recognition by pattern recognition receptors (PRRs) of the innate immune system and triggering inflammation. This viral mimicry mechanism has been observed in various human diseases, including aging and autoimmune disorders. However, recent evidence suggests that retrotransposons possess highly regulated immune reactivity and play important roles in the development and function of the immune system. In this review, I discuss a wide range of retrotransposon-derived transcripts, their role as targets in immune recognition, and the diseases associated with retrotransposon activity. Furthermore, I explore the implications of chimeric transcripts formed between retrotransposons and known gene mRNAs, which have been previously underestimated, for the increase of immune-related gene isoforms and their influence on immune function. Retrotransposon-derived transcripts have profound and multifaceted effects on immune system function. The aim of this comprehensive review is to provide a better understanding of the complex relationship between retrotransposon transcripts and immune defense.

Evolution of the Major Components of Innate Immunity in Animals.

Innate immunity is present in all animals. In this review, we explore the main conserved mechanisms of recognition and innate immune responses among animals. In this sense, we discuss the receptors, critical for binding to pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs); the downstream signaling proteins; and transcription factors that govern immune responses. We also highlight conserved inflammatory mediators that are induced after the recognition of DAMPs and PAMPs. At last, we discuss the mechanisms that are involved in the regulation and/or generation of reactive oxygen species (ROS), influencing immune responses, like heme-oxygenases (HOs).

Identification, characterization and the inflammatory regulating effect of NOD1/2 in sturgeon.

As an ancient species with both conservation and commercial value, Sturgeon's inflammatory regulation mechanism is a research point. Nucleotide-binding and oligomerization domain-containing proteins 1 and 2 (NOD1/2) are classical intracellular pattern recognition receptors (PRRs) in immunity of anti-bacterial infection. However, the characterization and function of NOD1/2 in Sturgeon are still unclear. In this study, we analyzed the synteny relationship of NOD1/2 genes between Acipenser ruthenus and representative fishes at the genome-level. Results showed that the ArNOD2 collinear genes pair was present in all representative fishes. The duplicated ArNOD1/2 genes were under purifying selection during evolution as indicated by their Ka/Ks values. To explore the function of NOD1/2, we further investigated their expression patterns and the effects of pathogenic infection, PAMPs treatment, and siRNA interference in Acipenser baerii, the sibling species of A. ruthenus. Results showed that both AbNOD1/2 were expressed at early developmental stages and in different tissues. Pathogenic infection in vivo and PAMPs treatment in vitro demonstrated that AbNOD1/2 could respond to pathogen stimulation. siRNA interference with AbNOD1/2 inhibited expression levels of RIPK2 and inflammatory cytokines compared to the control group after iE-DAP or MDP treatment. This study hinted that the AbNOD1/2 could stimulate the inflammatory cytokines response during evolutionary processes.

Toll-like receptors and Streptococcus mutans: An updated review article

It has been reported that toll-like receptors (TLRs) are the main innate immune receptors that recognize gram-positive pathogen-associated molecular patterns (PAMPs). The molecules can induce expression of the innate immune-related molecules that are essential against the bacteria. Streptococcus mutans (S. mutans) is a potential caries-associated pathogen, and innate immunity plays a key role in inhibiting its development and the progression of inflammatory responses. Recently, the roles played by TLRs against S. mutans and the induction of inflammatory responses were evaluated by several investigations. This review article discusses updated information regarding the roles played by TLRs and their potential therapeutic effects against S. mutans (AU)

PAMPs and DAMPs in Sepsis: A Review of Their Molecular Features and Potential Clinical Implications.

Sepsis is a serious organ dysfunction caused by a dysregulated immune host reaction to a pathogen. The innate immunity is programmed to react immediately to conserved molecules, released by the pathogens (PAMPs), and the host (DAMPs). We aimed to review the molecular mechanisms of the early phases of sepsis, focusing on PAMPs, DAMPs, and their related pathways, to identify potential biomarkers. We included studies published in English and searched on PubMed® and Cochrane®. After a detailed discussion on the actual knowledge of PAMPs/DAMPs, we analyzed their role in the different organs affected by sepsis, trying to elucidate the molecular basis of some of the most-used prognostic scores for sepsis. Furthermore, we described a chronological trend for the release of PAMPs/DAMPs that may be useful to identify different subsets of septic patients, who may benefit from targeted therapies. These findings are preliminary since these pathways seem to be strongly influenced by the peculiar characteristics of different pathogens and host features. Due to these reasons, while initial findings are promising, additional studies are necessary to clarify the potential involvement of these molecular patterns in the natural evolution of sepsis and to facilitate their transition into the clinical setting.

Toll-like receptors and Streptococcus mutans: An updated review article.

It has been reported that toll-like receptors (TLRs) are the main innate immune receptors that recognize gram-positive pathogen-associated molecular patterns (PAMPs). The molecules can induce expression of the innate immune-related molecules that are essential against the bacteria. Streptococcus mutans (S. mutans) is a potential caries-associated pathogen, and innate immunity plays a key role in inhibiting its development and the progression of inflammatory responses. Recently, the roles played by TLRs against S. mutans and the induction of inflammatory responses were evaluated by several investigations. This review article discusses updated information regarding the roles played by TLRs and their potential therapeutic effects against S. mutans.

A CO2 sensing module modulates ß-1,3-glucan exposure in Candida albicans.

Microbial species capable of co-existing with healthy individuals, such as the commensal fungus Candida albicans, exploit multifarious strategies to evade our immune defenses. These strategies include the masking of immunoinflammatory pathogen-associated molecular patterns (PAMPs) at their cell surface. We reported previously that C. albicans actively reduces the exposure of the proinflammatory PAMP, ß-1,3-glucan, at its cell surface in response to host-related signals such as lactate and hypoxia. Here, we show that clinical isolates of C. albicans display phenotypic variability with respect to their lactate- and hypoxia-induced ß-1,3-glucan masking. We have exploited this variability to identify responsive and non-responsive clinical isolates. We then performed RNA sequencing on these isolates to reveal genes whose expression patterns suggested potential association with lactate- or hypoxia-induced ß-1,3-glucan masking. The deletion of two such genes attenuated masking: PHO84 and NCE103. We examined NCE103-related signaling further because NCE103 has been shown previously to encode carbonic anhydrase, which promotes adenylyl cyclase-protein kinase A (PKA) signaling at low CO2 levels. We show that while CO2 does not trigger ß-1,3-glucan masking in C. albicans, the Sch9-Rca1-Nce103 signaling module strongly influences ß-1,3-glucan exposure in response to hypoxia and lactate. In addition to identifying a new regulatory module that controls PAMP exposure in C. albicans, our data imply that this module is important for PKA signaling in response to environmental inputs other than CO2.IMPORTANCEOur innate immune defenses have evolved to protect us against microbial infection in part via receptor-mediated detection of "pathogen-associated molecular patterns" (PAMPs) expressed by invading microbes, which then triggers their immune clearance. Despite this surveillance, many microbial species are able to colonize healthy, immune-competent individuals, without causing infection. To do so, these microbes must evade immunity. The commensal fungus Candida albicans exploits a variety of strategies to evade immunity, one of which involves reducing the exposure of a proinflammatory PAMP (ß-1,3-glucan) at its cell surface. Most of the ß-1,3-glucan is located in the inner layer of the C. albicans cell wall, hidden by an outer layer of mannan fibrils. Nevertheless, some ß-1,3-glucan can become exposed at the fungal cell surface. However, in response to certain specific host signals, such as lactate or hypoxia, C. albicans activates an anticipatory protective response that decreases ß-1,3-glucan exposure, thereby reducing the susceptibility of the fungus to impending innate immune attack. Here, we exploited the natural phenotypic variability of C. albicans clinical isolates to identify strains that do not display the response to ß-1,3-glucan masking signals observed for the reference isolate, SC5314. Then, using genome-wide transcriptional profiling, we compared these non-responsive isolates with responsive controls to identify genes potentially involved in ß-1,3-glucan masking. Mutational analysis of these genes revealed that a sensing module that was previously associated with CO2 sensing also modulates ß-1,3-glucan exposure in response to hypoxia and lactate in this major fungal pathogen of humans.

Additive and Specific Effects of Elicitor Treatments on the Metabolic Profile of Arabidopsis thaliana.

Several elicitors of plant defense have been identified and numerous efforts to use them in the field have been made. Exogenous elicitor treatments mimic the in planta activation of pattern-triggered immunity (PTI), which relies on the perception of pathogen-associated molecular patterns (PAMPs) such as bacterial flg22 or fungal chitins. Early transcriptional responses to distinct PAMPs are mostly overlapping, regardless of the elicitor being used. However, it remains poorly known if the same patterns are observed for metabolites and proteins produced later during PTI. In addition, little is known about the impact of a combination of elicitors on PTI and the level of induced resistance to pathogens. Here, we monitored Arabidopsis thaliana resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pto DC3000) following application of flg22 and chitosan elicitors, used individually or in combination. A slight, but not statistically significant increase in induced resistance was observed when the elicitors were applied together when compared with individual treatments. We investigated the effect of these treatments on the metabolome by using an untargeted analysis. We found that the combination of flg22 and chitosan impacted a higher number of metabolites and deregulated specific metabolic pathways compared with the elicitors individually. These results contribute to a better understanding of plant responses to elicitors, which might help better rationalize their use in the field. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Recombinant expression and immune function analysis of C-reactive protein (CRP) from Hexagrammos otakii.

C-reactive protein (CRP) belongs to the short-chain pentraxin family and functions as a soluble pattern recognition molecule (PRM) aiding in host defense against pathogens. In the present study, a CRP gene, designated HoCRP, was cloned from Hexagrammos otakii for the first time. The full length of the HoCRP cDNA sequence is 821 bp, which contains an open reading frame (ORF) of 675 bp encoding a 224 amino acid protein. The deduced protein is predicted to have a theoretical isoelectric point (pI) of 5.30 and a molecular weight of 25.4 kDa. The recombinant HoCRP protein (rHoCRP) was expressed in E. coli to further characterize the functions of HoCRP. Saccharide binding experiments demonstrated that rHoCRP exhibited a high affinity for various pathogen-associated molecular patterns (PAMPs). Furthermore, bacterial binding and agglutination assays indicated that rHoCRP had the capability to recognize a wide spectrum of microorganisms. These findings suggest that HoCRP functions not only as a PRM for binding PAMPs but also as an immune effector molecule. Considering the role CRP plays in the classical complement pathway, the interaction between rHoCRP and rHoC1q was assessed and proven by a Pull-down and Elisa assay, which implied that rHoCRP may be able to activate complement. In addition, phagocytosis enhancement by rHoCRP in the presence or absence of complement components was analysed by flow cytometry. The results showed that rHoCRP could synergistically enhance the phagocytosis of RAW264.7 cells with complement, providing further evidence of complement activation by rHoCRP through the opsonization of specific complement components. In summary, our findings suggest that rHoCRP may play a crucial role in host antibacterial defense by recognizing pathogens, activating the complement system, and enhancing macrophage function.

Reduced Bioactive Microbial Products (Pathogen-Associated Molecular Patterns) Contribute to Dysregulated Immune Responses and Impaired Healing in Infected Wounds in Mice with Diabetes.

Diabetic chronic ulcers are plagued with persistent nonresolving inflammation. However, diabetic wound environment early after injury suffers from inadequate inflammatory responses due to reductions in proinflammatory cytokines levels. Diabetic neutrophils have known impairments in bactericidal functions. We hypothesized that reduced bacterial killing by diabetic neutrophils, due to their bactericidal functional impairments, results in reduced bioactive bacterial products, known as pathogen-associated molecular patterns, which in turn contribute to reduced signaling through toll-like receptors, leading to inadequate production of proinflammatory cytokines in infected diabetic wound early after injury. We tested our hypothesis in db/db type 2 obese diabetic mouse wound infection model with Pseudomonas aeruginosa. Our data indicate that despite substantially higher levels of infection, toll-like receptor 4-mediated signaling is reduced in diabetic wounds early after injury owing to reduced bioactive levels of lipopolysaccharide. We further demonstrate that topical treatment with lipopolysaccharide enhances toll-like receptor 4 signaling, increases proinflammatory cytokine production, restores leukocyte trafficking, reduces infection burden, and stimulates healing in diabetic wounds. We posit that lipopolysaccharide may be a viable therapeutic option for the treatment of diabetic foot ulcers if it is applied topically after the surgical debridement process, which is intended to reset chronic ulcers into acute fresh wounds.

Bacteria- and fungus-derived PAMPs induce innate immune memory via similar functional, metabolic, and transcriptional adaptations.

Exposure to pathogen-associated molecular patterns (PAMPs) induces an augmented, broad-spectrum antimicrobial response to subsequent infection, a phenomenon termed innate immune memory. This study examined the effects of treatment with ß-glucan, a fungus-derived dectin-1 ligand, or monophosphoryl lipid A (MPLA), a bacteria-derived Toll-like receptor 4 ligand, on innate immune memory with a focus on identifying common cellular and molecular pathways activated by these diverse PAMPs. Treatment with either PAMP prepared the innate immune system to respond more robustly to Pseudomonas aeruginosa infection in vivo by facilitating mobilization of innate leukocytes into blood, recruitment of leukocytes to the site of infection, augmentation of microbial clearance, and attenuation of cytokine production. Examination of macrophages ex vivo showed amplification of metabolism, phagocytosis, and respiratory burst after treatment with either agent, although MPLA more robustly augmented these activities and more effectively facilitated killing of bacteria. Both agents activated gene expression pathways in macrophages that control inflammation, antimicrobial functions, and protein synthesis and suppressed pathways regulating cell division. ß-glucan treatment minimally altered macrophage differential gene expression in response to lipopolysaccharide (LPS) challenge, whereas MPLA attenuated the magnitude of the LPS-induced transcriptional response, especially cytokine gene expression. These results show that ß-glucan and MPLA similarly augment the innate response to infection in vivo. Yet, MPLA more potently induces alterations in macrophage metabolism, antimicrobial functions, gene transcription and the response to LPS.

Impact of Bifidobacterium longum1714® on maternal cytokine response in peripheral blood mononuclear cells.

PURPOSE: The maternal immune system is implicated in adverse pregnancy outcomes. Manipulation of maternal immune response by probiotics holds potential to reduce pregnancy complications. The MicrobeMom2 study investigates the impact of probiotic supplementation on maternal immune responses to pathogen associated molecular patterns (PAMPs) in peripheral blood mononuclear cells (PBMCs) during pregnancy. METHODS: This double-blinded randomised-controlled trial involved oral supplementation of Bifidobacterium longum subsp. longum 1714® (B. longum 1714; daily ingestion of a minimum of 1x109 colony forming units) or placebo from 16 to 20-weeks' gestation until delivery in healthy pregnant women. The primary outcome was a change in IL-10 production, after stimulation with Lipopolysaccharide (LPS) or anti-CD3/28/2, in PBMCs isolated from blood samples taken at baseline (11-15 weeks' gestation) and late pregnancy (28-32 weeks' gestation) after 48 h incubation. 68 subjects were needed (34ineachgroup) for 80 % power at an alpha significance of 0.05 to detect differences in IL10. RESULTS: 72 women (mean ± SD age 33.17 ± 4.53 years and median (25th, 75th centile) body mass index 24.93 (21.93, 27.57 kg/m2)) were recruited with primary outcome data. Using LPS, late pregnancy fold change in IL-10 in PBMCs after 48 h incubation was median (25th, 75th centile) 88.45 (4.88, 488.78) in the intervention, 24.18 (6.36, 141.17) in the control group, p = 0.183. Using anti-CD3/28/2, values were 189.69 (425.96, 866.57),148.74 (31.67, 887.03) in intervention and control groups, respectively, p = 0.506. No significant differences were observed between the two groups. CONCLUSION: Maternal antenatal supplementation with B. longum 1714 did not alter cytokine production by maternal PBMCs in response to PAMPs or anti-CD3/28/2. TRIAL REGISTRATION NUMBER: ISRCTN registry ISRCTN43013285.

Exploring the heterogeneous transcriptional response of the CNS to systemic LPS and Poly(I:C).

Peripheral contact to pathogen-associated molecular patterns (PAMPs) evokes a systemic innate immune response which is rapidly relayed to the central nervous system (CNS). The remarkable cellular heterogeneity of the CNS poses a significant challenge to the study of cell type and stimulus dependent responses of neural cells during acute inflammation. Here we utilized single nuclei RNA sequencing (snRNAseq), serum proteome profiling and primary cell culture methods to systematically compare the acute response of the mammalian brain to the bacterial PAMP lipopolysaccharide (LPS) and the viral PAMP polyinosinic:polycytidylic acid (Poly(I:C)), at single cell resolution. Our study unveiled convergent transcriptional cytokine and cellular stress responses in brain vascular and ependymal cells and a downregulation of several key mediators of directed blood brain barrier (BBB) transport. In contrast the neuronal response to PAMPs was limited in acute neuroinflammation. Moreover, our study highlighted the dominant role of IFN signalling upon Poly(I:C) challenge, particularly in cells of the oligodendrocyte lineage. Collectively our study unveils heterogeneous, shared and distinct cell type and stimulus dependent acute responses of the CNS to bacterial and viral PAMP challenges. Our findings highlight inflammation induced dysregulations of BBB-transporter gene expression, suggesting potential translational implications on drug pharmacokinetics variability during acute neuroinflammation. The pronounced dependency of oligodendrocytes on IFN stimulation during viral PAMP challenges, emphasizes their limited molecular viral response repertoire.

Transforming Intratumor Bacteria into Immunopotentiators to Reverse Cold Tumors for Enhanced Immuno-chemodynamic Therapy of Triple-Negative Breast Cancer.

Immunotherapy of triple-negative breast cancer (TNBC) has an unsatisfactory therapeutic outcome due to an immunologically "cold" microenvironment. Fusobacterium nucleatum (F. nucleatum) was found to be colonized in triple-negative breast tumors and was responsible for the immunosuppressive tumor microenvironment and tumor metastasis. Herein, we constructed a bacteria-derived outer membrane vesicle (OMV)-coated nanoplatform that precisely targeted tumor tissues for dual killing of F. nucleatum and cancer cells, thus transforming intratumor bacteria into immunopotentiators in immunotherapy of TNBC. The as-prepared nanoparticles efficiently induced immunogenic cell death through a Fenton-like reaction, resulting in enhanced immunogenicity. Meanwhile, intratumoral F. nucleatum was killed by metronidazole, resulting in the release of pathogen-associated molecular patterns (PAMPs). PAMPs cooperated with OMVs further facilitated the maturation of dendritic cells and subsequent T-cell infiltration. As a result, the "kill two birds with one stone" strategy warmed up the cold tumor environment, maximized the antitumor immune response, and achieved efficient therapy of TNBC as well as metastasis prevention. Overall, this strategy based on a microecology distinction in tumor and normal tissue as well as microbiome-induced reversal of cold tumors provides new insight into the precise and efficient immune therapy of TNBC.