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Brain, Behavior, and Immunity May 2024Complement is dysregulated in the brain in Alzheimer's Disease and in mouse models of Alzheimer's disease. Each of the complement derived effectors, opsonins,...
Complement is dysregulated in the brain in Alzheimer's Disease and in mouse models of Alzheimer's disease. Each of the complement derived effectors, opsonins, anaphylatoxins and membrane attack complex (MAC), have been implicated as drivers of disease but their relative contributions remain unclarified. Here we have focussed on the MAC, a lytic and pro-inflammatory effector, in the App mouse amyloidopathy model. To test the role of MAC, we back-crossed to generate App mice deficient in C7, an essential MAC component. C7 deficiency ablated MAC formation, reduced synapse loss and amyloid load and improved cognition compared to complement-sufficient App mice at 8-10 months age. Adding back C7 caused increased MAC formation in brain and an acute loss of synapses in C7-deficient App mice. To explore whether C7 was a viable therapeutic target, a C7-blocking monoclonal antibody was administered systemically for one month in App mice aged 8-9 months. Treatment reduced brain MAC and amyloid deposition, increased synapse density and improved cognitive performance compared to isotype control-treated App mice. The findings implicate MAC as a driver of pathology and highlight the potential for complement inhibition at the level of MAC as a therapy in Alzheimer's disease.
Topics: Mice; Animals; Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Cognitive Dysfunction; Mice, Transgenic; Plaque, Amyloid; Brain; Cognition; Complement Activation; Disease Models, Animal
PubMed: 38485063
DOI: 10.1016/j.bbi.2024.03.017 -
Cells Mar 2024Disulfiram (DSF), an anti-alcoholism medicine, exerts treatment effects in patients suffering from persistent Borreliosis and also exhibits anti-cancer effects through...
Disulfiram (DSF), an anti-alcoholism medicine, exerts treatment effects in patients suffering from persistent Borreliosis and also exhibits anti-cancer effects through its copper chelating derivatives and induction of oxidative stress in mitochondria. Since chronic/persistent borreliosis is characterized by increased amounts of pro-inflammatory macrophages, this study investigated opsonin-independent phagocytosis, migration, and surface marker expression of in vivo activated and in vitro cultured human monocyte-derived phagocytes (macrophages and dendritic cells) with and without DSF treatment. Phagocytosis of non-opsonized Dynabeads M-450 and migration of macrophages and dendritic cells were monitored using live cell analyzer Juli™ Br for 24 h, imaging every 3.5 min. To simultaneously monitor phagocyte function, results were analyzed by a newly developed software based on the differential phase contrast images of cells before and after ingestion of Dynabeads. DSF decreased the phagocytic capacities exhibited by in vitro enriched and long-lived phagocytes. Although no chemotactic gradient was applied to the test system, vigorous spontaneous migration was observed. We therefore set up an algorithm to monitor and quantify both phagocytosis and migration simultaneously. DSF not only reduced phagocytosis in a majority of these long-lived phagocytes but also impaired their migration. Despite these selective effects by DSF, we found that DSF reduced the expression densities of surface antigens CD45 and CD14 in all of our long-lived phagocytes. In cells with a high metabolic activity and high mitochondrial contents, DSF led to cell death corresponding to mitochondrial oxidative stress, whereas metabolically inactive phagocytes survived our DSF treatment protocol. In conclusion, DSF affects the viability of metabolically active phagocytes by inducing mitochondrial stress and secondly attenuates phagocytosis and migration in some long-lived phagocytes.
Topics: Humans; Disulfiram; Opsonin Proteins; Phagocytosis; Phagocytes; Macrophages
PubMed: 38534379
DOI: 10.3390/cells13060535 -
PeerJ 2024Mammalian models, such as murine, are used widely in pathophysiological studies because they have a high degree of similarity in body temperature, metabolism, and immune...
Mammalian models, such as murine, are used widely in pathophysiological studies because they have a high degree of similarity in body temperature, metabolism, and immune response with humans. However, non-vertebrate animal models have emerged as alternative models to study the host-pathogen interaction with minimal ethical concerns. is an alternative model that has proved useful in studying the interaction of the host with either bacteria or fungi, performing drug testing, and assessing the immunological response to different microorganisms. The immune response includes cellular and humoral components with structural and functional similarities to the immune effectors found in higher vertebrates, such as humans. An important humoral effector stimulated during infections is apolipophorin III (apoLp-III), an opsonin characterized by its lipid and carbohydrate-binding properties that participate in lipid transport, as well as immunomodulatory activity. Despite some parameters, such as the measurement of phenoloxidase activity, melanin production, hemocytes counting, and expression of antimicrobial peptides genes are already used to assess the immune response to pathogens with different virulence degrees, the apoLp-III quantification remains to be a parameter to assess the immune response in this invertebrate. Here, we propose an immunological tool based on an enzyme-linked immunosorbent assay that allows apoLp-III quantification in the hemolymph of larvae challenged with pathogenic agents. We tested the system with hemolymph coming from larvae infected with , , , , and . The results revealed significantly higher concentrations of apoLp-III when each microbial species was inoculated, in comparison with untouched larvae, or inoculated with phosphate-buffered saline. We also demonstrated that the apoLp-III levels correlated with the strains' virulence, which was already reported. To our knowledge, this is one of the first attempts to quantify apoLp-III, using a quick and easy-to-use serological technique.
Topics: Humans; Animals; Mice; Moths; Apolipoproteins; Larva; Host-Pathogen Interactions; Mammals
PubMed: 38500532
DOI: 10.7717/peerj.17117 -
BioRxiv : the Preprint Server For... May 2024Granzymes are a family of serine proteases mainly expressed by CD8 T cells, natural killer cells, and innate-like lymphocytes. Although their major role is thought to be...
Granzymes are a family of serine proteases mainly expressed by CD8 T cells, natural killer cells, and innate-like lymphocytes. Although their major role is thought to be the induction of cell death in virally infected and tumor cells, accumulating evidence suggests some granzymes can regulate inflammation by acting on extracellular substrates. Recently, we found that the majority of tissue CD8 T cells in rheumatoid arthritis (RA) synovium, inflammatory bowel disease and other inflamed organs express granzyme K (GZMK), a tryptase-like protease with poorly defined function. Here, we show that GZMK can activate the complement cascade by cleaving C2 and C4. The nascent C4b and C2a fragments form a C3 convertase that cleaves C3, allowing further assembly of a C5 convertase that cleaves C5. The resulting convertases trigger every major event in the complement cascade, generating the anaphylatoxins C3a and C5a, the opsonins C4b and C3b, and the membrane attack complex. In RA synovium, GZMK is enriched in areas with abundant complement activation, and fibroblasts are the major producers of complement C2, C3, and C4 that serve as targets for GZMK-mediated complement activation. Our findings describe a previously unidentified pathway of complement activation that is entirely driven by lymphocyte-derived GZMK and proceeds independently of the classical, lectin, or alternative pathways. Given the widespread abundance of -expressing T cells in tissues in chronic inflammatory diseases and infection, GZMK-mediated complement activation is likely to be an important contributor to tissue inflammation in multiple disease contexts.
PubMed: 38826230
DOI: 10.1101/2024.05.22.595315 -
Frontiers in Cellular and Infection... 2023The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We...
The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We previously demonstrated that in its protein ligands, Mac-1 binds sequences enriched in basic and hydrophobic residues and strongly disfavors negatively charged sequences. The avoidance by Mac-1 of negatively charged surfaces suggests that the bacterial wall and bacterial capsule possessing net negative electrostatic charge may repel Mac-1 and that the cationic Mac-1 ligands can overcome this evasion by acting as opsonins. Indeed, we previously showed that opsonization of Gram-negative with several cationic peptides, including PF4 (Platelet Factor 4), strongly augmented phagocytosis by macrophages. Here, we investigated the effect of recombinant PF4 (rPF4) on phagocytosis of Gram-positive and examined its impact in a mouse model of peritonitis. Characterization of the interaction of rPF4 with nonencapsulated and encapsulated showed that rPF4 localizes on the bacterial surface, thus making it available for Mac-1. Furthermore, rPF4 did not have direct bactericidal and bacteriostatic activity and was not toxic to host cells. rPF4 enhanced phagocytosis of bioparticles by various primary and cultured Mac-1-expressing leukocytes by several folds. It also increased phagocytosis of live nonencapsulated and encapsulated bacteria. Notably, the augmentation of phagocytosis by rPF4 did not compromise the intracellular killing of by macrophages. Using a murine peritonitis model, we showed that treatment of infected mice with rPF4 caused a significant increase in the clearance of antibiotic-susceptible and its methicillin-resistant (MRSA) variant and markedly improved survival. These findings indicate that rPF4 binding to the bacterial surface circumvents its antiphagocytic properties, improving host defense against antibiotic-susceptible and antibiotic-resistant bacteria.
Topics: Animals; Mice; Methicillin-Resistant Staphylococcus aureus; Anti-Bacterial Agents; Platelet Factor 4; Staphylococcus aureus; Disease Models, Animal; Phagocytosis; Macrophage-1 Antigen; Immunologic Factors; Peritonitis
PubMed: 37868353
DOI: 10.3389/fcimb.2023.1217103 -
PeerJ 2023The complement system is an essential part of innate immunity. It is activated by invading pathogens causing inflammation, opsonization, and lysis complement...
The complement system is an essential part of innate immunity. It is activated by invading pathogens causing inflammation, opsonization, and lysis complement anaphylatoxins, complement opsonin's and membrane attack complex (MAC), respectively. However, in SARS-CoV-2 infection overactivation of complement system is causing cytokine storm leading to multiple organs damage. In this study, the René Thomas kinetic logic approach was used for the development of biological regulatory network (BRN) to model SARS-CoV-2 mediated complement system signalling pathways. Betweenness centrality analysis in cytoscape was adopted for the selection of the most biologically plausible states in state graph. Among the model results, in strongly connected components (SCCs) pro-inflammatory cytokines (PICyts) oscillatory behaviour between recurrent generation and downregulation was found as the main feature of SARS-CoV-2 infection. Diversion of trajectories from the SCCs leading toward hyper-inflammatory response was found in agreement with studies that overactive innate immunity response caused PICyts storm during SARS-CoV-2 infection. The complex of negative regulators FI, CR1 and DAF in the inhibition of complement peptide (C5a) and PICyts was found desirable to increase immune responses. In modelling role of MAC and PICyts in lowering of SARS-CoV-2 titre was found coherent with experimental studies. Intervention in upregulation of C5a and PICyts by C3 was found helpful in back-and-forth variation of signalling pattern linked with the levels of PICyts. Moreover, intervention in upregulation of PICyts by C5a was found productive in downregulation of all activating factors in the normal SCCs. However, the computational model predictions require experimental studies to be validated by exploring the activation role of C3 and C5a which could change levels of PICyts at various phases of SARS-CoV-2 infection.
Topics: Humans; Cytokines; COVID-19; SARS-CoV-2; Complement System Proteins; Complement Membrane Attack Complex
PubMed: 37744234
DOI: 10.7717/peerj.15794 -
Neural Regeneration Research Jun 2024Spinal cord injury remains a major cause of disability in young adults, and beyond acute decompression and rehabilitation, there are no pharmacological treatments to...
Spinal cord injury remains a major cause of disability in young adults, and beyond acute decompression and rehabilitation, there are no pharmacological treatments to limit the progression of injury and optimize recovery in this population. Following the thorough investigation of the complement system in triggering and propagating cerebral neuroinflammation, a similar role for complement in spinal neuroinflammation is a focus of ongoing research. In this work, we survey the current literature investigating the role of complement in spinal cord injury including the sources of complement proteins, triggers of complement activation, and role of effector functions in the pathology. We study relevant data demonstrating the different triggers of complement activation after spinal cord injury including direct binding to cellular debris, and or activation via antibody binding to damage-associated molecular patterns. Several effector functions of complement have been implicated in spinal cord injury, and we critically evaluate recent studies on the dual role of complement anaphylatoxins in spinal cord injury while emphasizing the lack of pathophysiological understanding of the role of opsonins in spinal cord injury. Following this pathophysiological review, we systematically review the different translational approaches used in preclinical models of spinal cord injury and discuss the challenges for future translation into human subjects. This review emphasizes the need for future studies to dissect the roles of different complement pathways in the pathology of spinal cord injury, to evaluate the phases of involvement of opsonins and anaphylatoxins, and to study the role of complement in white matter degeneration and regeneration using translational strategies to supplement genetic models.
PubMed: 38845224
DOI: 10.4103/NRR.NRR-D-24-00116 -
BioRxiv : the Preprint Server For... Aug 2023The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We...
The complement receptor CR3, also known as integrin Mac-1 (CD11b/CD18), is one of the major phagocytic receptors on the surface of neutrophils and macrophages. We previously demonstrated that in its protein ligands, Mac-1 binds sequences enriched in basic and hydrophobic residues and strongly disfavors negatively charged sequences. The avoidance by Mac-1 of negatively charged surfaces suggests that the bacterial wall and bacterial capsule possessing net negative electrostatic charge may repel Mac-1 and that the cationic Mac-1 ligands can overcome this evasion by acting as opsonins. Indeed, we previously showed that opsonization of Gram-negative with several cationic peptides, including PF4 (Platelet Factor 4), strongly augmented phagocytosis by macrophages. Here, we investigated the effect of recombinant PF4 (rPF4) on phagocytosis of Gram-positive in vitro and examined its impact in a mouse model of peritonitis. Characterization of the interaction of rPF4 with nonencapsulated and encapsulated showed that rPF4 localizes on the bacterial surface, thus making it available for Mac-1. Furthermore, rPF4 did not have direct bactericidal and bacteriostatic activity and was not toxic to host cells. rPF4 enhanced phagocytosis of bioparticles by various primary and cultured Mac-1-expressing leukocytes by several folds. It also increased phagocytosis of live nonencapsulated and encapsulated bacteria. Notably, the augmentation of phagocytosis by rPF4 did not compromise the intracellular killing of by macrophages. Using a murine peritonitis model, we showed that treatment of infected mice with rPF4 caused a significant increase in the clearance of antibiotic-susceptible and its methicillin-resistant (MRSA) variant and markedly improved survival. These findings indicate that rPF4 binding to the bacterial surface circumvents its antiphagocytic properties, improving host defense against antibiotic-susceptible and antibiotic-resistant bacteria.
PubMed: 37662328
DOI: 10.1101/2023.08.25.554865 -
3 Biotech Jul 2023The 'enzyme prodrug therapy' represents a promising strategy to overcome limitations of current cancer treatments by the systemic administration of prodrugs, converted...
UNLABELLED
The 'enzyme prodrug therapy' represents a promising strategy to overcome limitations of current cancer treatments by the systemic administration of prodrugs, converted by a foreign enzyme into an active anticancer compound directly in tumor sites. One example is D-amino acid oxidase (DAAO), a dimeric flavoenzyme able to catalyze the oxidative deamination of D-amino acids with production of hydrogen peroxide, a reactive oxygen species (ROS), able to favor cancer cells death. A DAAO variant containing five aminoacidic substitutions (mDAAO) was demonstrated to possess a better therapeutic efficacy under low O concentration than wild-type DAAO (wtDAAO). Recently, aiming to design promising nanocarriers for DAAO, multi-walled carbon nanotubes (MWCNTs) were functionalized with polyethylene glycol (PEG) to reduce their tendency to aggregation and to improve their biocompatibility. Here, wtDAAO and mDAAO were adsorbed on PEGylated MWCNTs and their activity and cytotoxicity were tested. While PEG-MWCNTs-DAAOs have shown a higher activity than pristine MWCNTs-DAAO (independently on the DAAO variant used), PEG-MWCNTs-mDAAO showed a higher cytotoxicity than PEG-MWCNTs-wtDAAO at low O concentration. In order to evaluate the nanocarriers' biocompatibility, PEG-MWCNTs-DAAOs were incubated in human serum and the composition of protein corona was investigated via nLC-MS/MS, aiming to characterize both soft and hard coronas. The mDAAO variant has influenced the bio-corona composition in both number of proteins and presence of opsonins and dysopsonins: notably, the soft corona of PEG-MWCNTs-mDAAO contained less proteins and was more enriched in proteins able to inhibit the immune response than PEG-MWCNTs-wtDAAO. Considering the obtained results, the PEGylated MWCNTs conjugated with the mDAAO variant seems a promising candidate for a selective antitumor oxidative therapy: under anoxic-like conditions, this novel drug delivery system showed a remarkable cytotoxic effect controlled by the substrate addition, against different tumor cell lines, and a bio-corona composition devoted to prolong its blood circulation time, thus improving the drug's biodistribution.
SUPPLEMENTARY INFORMATION
The online version contains supplementary material available at 10.1007/s13205-023-03568-1.
PubMed: 37346390
DOI: 10.1007/s13205-023-03568-1 -
Small (Weinheim An Der Bergstrasse,... Mar 2024Engineering of catalytically active inorganic nanomaterials holds promising prospects for biomedicine. Catalytically active metal oxides show applications in enhancing...
Engineering of catalytically active inorganic nanomaterials holds promising prospects for biomedicine. Catalytically active metal oxides show applications in enhancing wound healing but have also been employed to induce cell death in photodynamic or radiation therapy. Upon introduction into a biological system, nanomaterials are exposed to complex fluids, causing interaction and adsorption of ions and proteins. While protein corona formation on nanomaterials is acknowledged, its modulation of nanomaterial catalytic efficacy is less understood. In this study, proteomic analyses and nano-analytic methodologies quantify and characterize adsorbed proteins, correlating this protein layer with metal oxide catalytic activity in vitro and in vivo. The protein corona comprises up to 280 different proteins, constituting up to 38% by weight. Enhanced complement factors and other opsonins on nanocatalyst surfaces lead to their uptake into macrophages when applied topically, localizing >99% of the nanomaterials in tissue-resident macrophages. Initially, the formation of the protein corona significantly reduces the nanocatalysts' activity, but this activity can be partially recovered in endosomal conditions due to the proteolytic degradation of the corona. Overall, the research reveals the complex relationship between physisorbed proteins and the catalytic characteristics of specific metal oxide nanoparticles, providing design parameters for optimizing nanocatalysts in complex biological environments.
PubMed: 38556634
DOI: 10.1002/smll.202311115