Classical Pathway Research Articles

Liraglutide Reduces Alcohol Consumption, Anxiety, Memory Impairment, and Synapse Loss in Alcohol Dependent Mice.

Glucagon-like peptide 1 (GLP-1) analogues have been commercialized for the management of type 2 diabetes. Recent studies have underscored GLP-1's role as a modulator of alcohol-related behavior. However, the role of the GLP-1 analogue liraglutide on alcohol-withdrawal responses have not been fully elucidated. Liraglutide binds to the G-protein-coupled receptor and activates an adenylyl cyclase and the associated classic growth factor signaling pathway, which acts growth factor-like and neuroprotective properties. The underlying neurobiological mechanisms of liraglutide on alcohol withdrawal remains unknown. This study endeavored to explore the effects of liraglutide on the emotion and memory ability of alcohol-withdrawal mice, and synaptic morphology in the medial prefrontal cortex (mPFC) and the hippocampus (HP), and thus affects the relapse-like drinking of alcohol-withdrawal mice. The alcohol-withdrawal group was reintroduced to a 20% v/v alcohol and water through the two-bottle choice for four consecutive days, a period referred to as alcohol re-drinking. Male C57BL/6J mice were exposed to a regimen of 20% alcohol and water for a duration of 6 weeks. This regimen established the two-bottle choice model of alcohol exposure. Learning capabilities, memory proficiency, and anxiety-like behavior were evaluated using the Morris water maze, open field, and elevated plus maze paradigms. Furthermore, synaptic morphology and the levels of synaptic transport-related proteins were assessed via Golgi staining and Western Blot analysis after a two-week alcohol deprivation period. Alcohol re-drinking of alcohol-withdrawal mice was also evaluated using a two-bottle choice paradigm. Our findings indicate that liraglutide can substantially decrease alcohol consumption and preference (p < 0.05) in the alcohol group and enhance learning and memory performance (p < 0.01), as well as alleviate anxiety-like behavior (p < 0.01) of alcohol-withdrawal mice. Alcohol consumption led to a reduction in dendritic spine density in the mPFC and HP, which was restored to normal levels by liraglutide (p < 0.001). Furthermore, liraglutide was found to augment the levels of synaptic transport-related proteins in mice subjected to alcohol withdrawal (p < 0.01). The study findings corroborate that liraglutide has the potential to mitigate alcohol consumption and ameliorate the memory impairments and anxiety induced by alcohol withdrawal. The therapeutic efficacy of liraglutide might be attributed to its role in counteracting synapse loss in the mPFC and HP regions and thus prevented relapse-like drinking in alcohol-withdrawal mice.

Complement System Deficiencies in Elite Athletes.

Although regular physical activity improves immune competency and reduces the prevalence of inflammatory diseases, strenuous training in elite athletes is associated with an increased susceptibility to infectious complications. Therefore, the objective of our study was to assess the routinely examined parameters of the complement system in elite athletes. The study was carried out in a cohort of elite athletes (n = 134) and healthy control subjects (n = 110). In all subjects, besides a routine laboratory check-up, serum concentrations of the C3 and C4 complement components, mannose-binding lectin (MBL), as well as activation of all three complement pathways were determined. Compared to healthy controls, lower C3 and C4 complement component concentrations were observed in elite athletes (0.96 ± 0.1 vs. 1.08 ± 0.2mg/L, and 0.18 ± 0.1 vs. 0.25 ± 0.1mg/L, respectively, p < 0.05); with much higher frequency rates of C3 and C4 deficiencies in athletes (31.3 vs. 14.5%, and 6 vs. 0%, p < 0.05). Simultaneously, athletes had much higher frequency rates of deficiencies of activation of classical and alternative complement pathways; while, deficiency of activation of the lectin pathway was similar in both cohorts. We confirmed a high frequency of defects in the complement system in elite athletes. Lower concentrations of C3 and C4 complement components, with high frequencies of deficiencies of the classical and alternative complement activation pathways were the most prevalent disorder of the complement system in elite athletes. Further studies are needed to uncover the functional impacts of these observations upon the susceptibility to infectious diseases.

Two Forms of C-Reactive Protein and Their Implication for Atherogenesis: Focus on Monomeric Form

In this review, we discuss the role of C-reactive protein (CRP), a biomarker of inflammation, in the development of atherosclerosis. CRP exists in two forms: the native pentameric form (nCRP) and the non-native form (mCRP), which is formed during inflammatory processes. The article explores the functions of both forms in atherosclerosis. nCRP binds to molecules with uncoated phosphocholine groups and activates the classical complement pathway. It can bind to enzymatically-modified LDL (E-LDL) and ox-LDL, affecting their pro-inflammatory properties and reducing LDL oxidation. nCRP also interacts with endothelial cells and reduces the absorption of acetylated LDL by these cells. In contrast, mCRP, formed in the presence of pathological conditions, exhibits distinct pro-inflammatory characteristics. While the role of nCRP in atherosclerosis remains unclear in animal models, mCRP has been associated with inflammation and is found in atherosclerotic lesions. The study suggests that nCRP may have atheroprotective effects, while mCRP may contribute to the progression of atherosclerosis. Further research is needed to fully understand the complex mechanisms of CRP in atherosclerosis.

Investigation of Potential Drug Targets Involved in Inflammation Contributing to Alzheimer's Disease Progression.

Alzheimer's disease has become a major public health issue. While extensive research has been conducted in the last few decades, few drugs have been approved by the FDA to treat Alzheimer's disease. There is still an urgent need for understanding the disease pathogenesis, as well as identifying new drug targets for further drug discovery. Alzheimer's disease is known to arise from a build-up of amyloid beta (Aβ) plaques as well as tangles of tau proteins. Along similar lines to Alzheimer's disease, inflammation in the brain is known to stem from the degeneration of tissue and build-up of insoluble materials. A minireview was conducted in this work assessing the genes, proteins, reactions, and pathways that link brain inflammation and Alzheimer's disease. Existing tools in Systems Biology were implemented to build protein interaction networks, mainly for the classical complement pathway and G protein-coupled receptors (GPCRs), to rank the protein targets according to their interactions. The top 10 protein targets were mainly from the classical complement pathway. With the consideration of existing clinical trials and crystal structures, proteins C5AR1 and GARBG1 were identified as the best targets for further drug discovery, through computational approaches like ligand-protein docking techniques.

Bioinformatics analysis identifies GLUD1 as a prognostic indicator for clear cell renal cell carcinoma

BackgroundRenal cell carcinoma (RCC) is a common primary tumor of the kidney and is divided into three major subtypes, of which clear cell renal cell carcinoma (ccRCC) has the highest incidence. Glutamate dehydrogenase 1 (GLUD1) encodes glutamate dehydrogenase 1, which catalyzes the oxidative deamination of glutamate.MethodsWe analyzed TCGA data using R language software and used multiple online databases to explore the relationship of GLUD1 with signaling pathways and drug sensitivity as well as GLUD1 protein expression and methylation.ResultsThe results showed that GLUD1 mRNA expression was reduced in tumor tissues and correlated with the progression of ccRCC. Univariate and multivariate Cox analysis showed that GLUD1 could be used as a prognostic marker for ccRCC. GLUD1 expression in ccRCC was associated with immune cells infiltration and multiple classical signaling pathways. In addition, GLUD1 mRNA expression was related to drug sensitivity.ConclusionsThese findings provide new ideas for finding new prognostic molecular markers and therapeutic targets for ccRCC.

Persistent complement dysregulation with signs of thromboinflammation in active Long Covid.

Long Covid is a debilitating condition of unknown etiology. We performed multimodal proteomics analyses of blood serum from COVID-19 patients followed up to 12 months after confirmed severe acute respiratory syndrome coronavirus 2 infection. Analysis of >6500 proteins in 268 longitudinal samples revealed dysregulated activation of the complement system, an innate immune protection and homeostasis mechanism, in individuals experiencing Long Covid. Thus, active Long Covid was characterized by terminal complement system dysregulation and ongoing activation of the alternative and classical complement pathways, the latter associated with increased antibody titers against several herpesviruses possibly stimulating this pathway. Moreover, markers of hemolysis, tissue injury, platelet activation, and monocyte-platelet aggregates were increased in Long Covid. Machine learning confirmed complement and thromboinflammatory proteins as top biomarkers, warranting diagnostic and therapeutic interrogation of these systems.

Soluble terminal complement complex blood levels are elevated in schizophrenia

The role of the complement system in schizophrenia (Sz) is inconclusive due to heterogeneity of the disease and study designs. Here, we assessed the levels of complement activation products and functionality of the classical pathway in acutely ill unmedicated Sz patients at baseline and after 6 weeks of treatment versus matched controls. The study included analyses of the terminal complement complex (sTCC) and C5a in plasma from 96 patients and 96 controls by enzyme-linked immunosorbent assay. Sub-group analysis of serum was conducted for measurement of C4 component and activity of the classical pathway (28 and 24 cases per cohort, respectively). We found no differences in levels of C5a, C4 and classical pathway function in patients versus controls. Plasma sTCC was significantly higher in patients [486 (392–659) ng/mL, n = 96] compared to controls [389 (304–612) ng/mL, n = 96] (p = 0.027, δ = 0.185), but not associated with clinical symptom ratings or treatment. The differences in sTCC between Sz and controls were confirmed using an Aligned Rank Transformation model considering the covariates age and sex (p = 0.040). Additional analysis showed that sTCC was significantly associated with C-reactive protein (CRP; p = 0.006). These findings suggest that sTCC plays a role in Sz as a trait marker of non-specific chronic immune activation, as previously described for CRP. Future longitudinal analyses with more sampling time points from early recognition centres for psychoses may be helpful to better understand the temporal dynamics of innate immune system changes during psychosis development.

Quantitative comparison of nuclear transport inhibition by SARS coronavirus ORF6 reveals the importance of oligomerization

Open Reading Frame 6 (ORF6) proteins, which are unique to severe acute respiratory syndrome-related (SARS) coronavirus, inhibit the classical nuclear import pathway to antagonize host antiviral responses. Several alternative models were proposed to explain the inhibitory function of ORF6 [H. Xia et al., Cell Rep. 33, 108234 (2020); L. Miorin et al., Proc. Natl. Acad. Sci. U.S.A. 117, 28344-28354 (2020); and M. Frieman et al., J. Virol. 81, 9812-9824 (2007)]. To distinguish these models and build quantitative understanding of ORF6 function, we developed a method for scoring both ORF6 concentration and functional effect in single living cells. We combined quantification of untagged ORF6 expression level in single cells with optogenetics-based measurement of nuclear transport kinetics, using methods that could be adapted to measure concentration-dependent effects of any untagged protein. We found that SARS-CoV-2 ORF6 is ~15 times more potent than SARS-CoV-1 ORF6 in inhibiting nuclear import and export, due to differences in the C-terminal region that is required for the NUP98-RAE1 binding. The N-terminal region was required for transport inhibition. This region binds membranes but could be replaced by synthetic constructs which forced oligomerization in solution, suggesting its primary function is oligomerization. We propose that the hydrophobic N-terminal region drives oligomerization of ORF6 to multivalently cross-link the NUP98-RAE1 complexes at the nuclear pore complex, and this multivalent binding inhibits bidirectional transport.

Antibody-Mediated Serum Resistance Protects Pseudomonas aeruginosa During Bloodstream Infections.

Pseudomonas aeruginosa is a frequent pathogen isolated from bacterial bloodstream infection (BSI) and is associated with high mortality. To survive in the blood, P aeruginosa must resist the bactericidal action of complement (ie, serum killing). Antibodies usually promote serum killing through the classical complement pathway; however, "cloaking antibodies" (cAbs) have been described, which paradoxically protect bacteria from serum killing. The relevance of cAbs in P aeruginosa BSI is unknown. Serum and P aeruginosa were collected from a cohort of 100 patients with BSI. Isolates were tested for sensitivity to healthy control serum (HCS). cAb prevalence was determined in sera. Patient sera were mixed with HCS to determine if killing of the matched isolate was inhibited. Overall, 36 patients had elevated titers of cAbs, and 34 isolates were sensitive to HCS killing. Fifteen patients had cAbs and HCS-sensitive isolates; of these patients, 14 had serum that protected their matched bacteria from HCS killing. Patients with cAbs were less likely to be neutropenic or have comorbidities. cAbs are prevalent in patients with P aeruginosa BSI and allow survival of otherwise serum-sensitive bacteria in the bloodstream. Generation of cAbs may be a risk factor for the development of BSI.

A novel flavonol-polysaccharide from Tamarix chinensis alleviates influenza A virus-induced acute lung injury. Evidences for its mechanism of action

BackgroundTamarix chinensis Lour. is a Chinese medicine used for treating inflammation-related diseases and its crude polysaccharides (MBAP90) exhibited significant anticomplement activities in vitro. PurposeTo obtain anticomplement homogenous polysaccharides from MBAP90 and explore its therapeutic effects and potential mechanism on influenza A virus (IAV)-induced acute lung injury (ALI). MethodsAnticomplement activity-guided fractionation of the water-soluble crude polysaccharides from the leaves and twigs of T. chinensis were performed by diethylaminoethyl-52 (DEAE-52) cellulose and gel permeation columns to yield a homogeneous polysaccharide MBAP-5, which was further characterized using ultra-high-performance liquid chromatography-ion trap tandem mass spectrometry (UPLC-IT-MS) and nuclear magnetic resonance (NMR) analysis. In vitro, the anticomplement activity of MBAP-5 through classical pathway was measured using a hemolytic test. The therapeutic effects of MBAP-5 on ALI were evaluated in H1N1-infected mice. H&E staining, enzyme linked immunosorbent assay (ELISA), immunohistochemistry, and western blot were used to systematically access lung histomorphology, inflammatory cytokines, degree of complement component 3c, 5aR, and 5b-9 (C3c, C5aR, and C5b-9) deposition, and inflammasome signaling pathway protein expressions in lung tissues. ResultsMBAP-5 was a novel flavonol-polysaccharide with the molecular weight (Mw) of 153.6 kDa. Its structure was characterized to process a backbone of →4)-α-D-GlcpA-(1→, →6)-α-D-Glcp-(1→, →3,4)-α-D-Glcp-(1→, →3,4,6)-α-D-Glcp-(1→, and →4,6)-β-D-Glcp-(1→, as well as branches of α-L-Araf-(1→ and β-D-Galp-(1→. Particularly, O-3 of →3,4,6)-α-D-Glcp-(1→ was substituted by quercetin. In vitro assay showed that MBAP-5 had a potent anticomplement activity with a CH50 value of 102 ± 4 µg/ml. Oral administration of MBAP-5 (50 and 100 mg/kg) effectively attenuated the H1N1-induced pulmonary injury in vivo by reducing pulmonary edema, virus replication, and inflammatory responses. Mechanistically, MBAP-5 inhibited the striking deposition and contents of complement activation products (C3c, C5aR, and C5b-9) in the lung. Toll-like receptor 4 (TLR4) /transcription factor nuclear factor κB (NF-κB) signaling pathway was constrained by MBAP-5 treatment. In addition, MBAP-5 could suppress activation of the inflammasome pathways, including Nod‐like receptor pyrin domain 3 (NLRP3), cysteinyl aspartate specific proteinase-1/12 (caspase-1/12), apoptosis‑associated speck‑like protein (ASC), gasdermin D (GSDMD), interleukin (IL)-1β, and IL-18 expressions. ConclusionsA novel flavonol-polysaccharide MBAP-5 isolated from T. chinensis demonstrated a therapeutic effect against ALI induced by IAV attack. The mechanism might be associated with inhibition of complement system and inflammasome pathways activation.

A survey of ficolin-3 activity in Systemic Lupus Erythematosus reveals a link to hematological disease manifestations and autoantibody profile

The complement system plays a central role in the pathogenesis of Systemic Lupus Erythematosus (SLE), but most studies have focused on the classical pathway. Ficolin-3 is the main initiator of the lectin pathway of complement in humans, but its role in systemic autoimmune disease has not been conclusively determined. Here, we combined biochemical and genetic approaches to assess the contribution of ficolin-3 to SLE risk and disease manifestations. Ficolin-3 activity was measured by a functional assay in serum or plasma samples from Swedish SLE patients (n = 786) and controls matched for age and sex (n = 566). Genetic variants in an extended 300 kb genomic region spanning the FCN3 locus were analyzed for their association with ficolin-3 activity and SLE manifestations in a Swedish multicenter cohort (n = 985). Patients with ficolin-3 activity in the highest tertile showed a strong enrichment in an SLE cluster defined by anti-Sm/DNA/nucleosome antibodies (OR 3.0, p < 0.001) and had increased rates of hematological disease (OR 1.4, p = 0.078) and lymphopenia (OR = 1.6, p = 0.039). Genetic variants associated with low ficolin-3 activity mapped to an extended haplotype in high linkage disequilibrium upstream of the FCN3 gene. Patients carrying the lead genetic variant associated with low ficolin-3 activity had a lower frequency of hematological disease (OR 0.67, p = 0.018) and lymphopenia (OR 0.63, p = 0.031) and fewer autoantibodies (p = 0.0019). Loss-of-function variants in the FCN3 gene were not associated with SLE, but four (0.5 %) SLE patients developed acquired ficolin-3 deficiency where ficolin-3 activity in serum was depleted following diagnosis of SLE.Taken together, our results provide genetic and biochemical evidence that implicate the lectin pathway in hematological SLE manifestations. We also identify lectin pathway activation through ficolin-3 as a factor that contributes to the autoantibody response in SLE.

Hedgehog signalling in CD4+ T helper cell polarisation

CD4+ T cells are critical in orchestrating immune responses against various pathogens and cancer but can also be drivers of autoimmune disease, allergy and pro-tumour responses. Naïve CD4+ T cells polarise into specialised T helper cell subsets with unique effector functions. While the guiding transcription factors and effector molecules of the T helper cell lineages are well understood, the signalling pathways orchestrating the intricate T helper cell polarisation programmes remain poorly understood. Here we review an emerging role of Hedgehog signalling – a classical morphogen signalling pathway – in T helper cell polarisation. Importantly, the Hedgehog pathway is pharmacologically highly tractable and existing clinically-approved Hedgehog inhibitors may prove useful therapeutic modulators of T helper cell-driven immune responses.

Trehalosemodulates OVRAS to improve oxidative stress and apoptosis in KGN cells and ovaries of PCOS mice

The etiology of polycystic ovary syndrome (PCOS) is complex and variable, and there is no exact cause or good treatment method. Most of the methods of hormones are used to temporarily meet the needs of patients. Experimental evidence has shown that trehalose has, anti-apoptotic, anti-oxidative, glucose-lowering, and insulin resistance effects. However, whether trehalose has a therapeutic effect on PCOS is unknown. It has been reported that the ovarian renin-angiotensin system (OVRAS) is involved in the development of PCOS, but it has not been fully elucidated. This study aims to explore the effect of trehalose on PCOS and elucidate the related OVRAS mechanism. We first observed that body weight, estrous cycle, ovarian follicles at all levels, glucose tolerance, serum hormones, and insulin resistance were improved by trehalose treatment in the PCOS mouse model. Moreover, trehalose treatment also ameliorated ovarian oxidative stress and apoptosis in PCOS mice, as determined by TUNNEL apoptosis staining, total SOD in ovarian homogenate, and WB assay. OVRAS mainly involves two classic pathways, namely the ACE/AngII/AT1R/AT2R, and ACE2 / Ang1-7/ MASR, Which play different functions. In PCOS mouse ovaries, we found that ACE/AngII/AT1R was up-regulated and ACE2/Ang1-7/MASR and AT2R were down-regulated by PCR and WB experiments, However, trehalose treatment changed its direction. In addition, we also found that trehalose ameliorated DHEA-induced oxidative stress and apoptosis in KGN by PCR and WB experiments, mainly by down-regulating ACE/AngII/AT1R. Our study shows that trehalose improves symptoms of PCOS mainly by down-regulating ACE/AngII/AT1R, revealing a potential therapeutic target for PCOS.

Research Progress on the Mechanism of Milk Fat Synthesis in Cows and the Effect of Conjugated Linoleic Acid on Milk Fat Metabolism and Its Underlying Mechanism: A Review.

Milk fat synthesis in cows mainly includes the synthesis of short- and medium-chain fatty acids, the uptake, transport, and activation of long-chain fatty acids (LCFAs), the synthesis of triglycerides, and the synthesis of the genes, transcription factors, and signaling pathways involved. Although the various stages of milk fat synthesis have been outlined in previous research, only partial processes have been revealed. CLA consists of an aggregation of positional and geometric isomers of linoleic fatty acid, and the accumulated evidence suggests that the two isomers of the active forms of CLA (cis-9, trans-11 conjugated linoleic acid and trans-10, cis-12 conjugated linoleic acid, abbreviated as c9, t11-CLA and t10, c12-CLA) can reduce the fat content in milk by regulating lipogenesis, fatty acid (FA) uptake, oxidation, and fat synthesis. However, the mechanism through which CLA inhibits milk fat synthesis is unique, with most studies focusing only on the effects of CLA on one of the genes, transcription factors, or signaling pathways involved. In this study, we summarized the structure and function of classic genes and pathways (mTOR, SREBP, AMPK, and PPARG) and new genes or pathways (THRSP, METTL3, ELOVL, and LPIN1) involved in each stage of milk fat synthesis and demonstrated the interactions between genes and pathways. We also examined the effects of other substances (melanin, nicotinic acid, SA, etc.). Furthermore, we evaluated the influence of β-sitosterol, sodium butyrate, Met arginine, and Camellia oleifera Abel on milk fat synthesis to improve the mechanism of milk fat synthesis in cows and provide a mechanistic reference for the use of CLA in inhibiting milk fat biosynthesis.

Endogenous complement-activating IgM is not required for primary antibody responses but promotes plasma cell differentiation and secondary antibody responses to a large particulate antigen in mice.

Lack of complement factor C1q of the classical pathway results in severely impaired primary antibody responses. This is a paradox because antibodies, especially IgM, are the most efficient activators of the classical pathway and very little specific IgM will be present at priming. A possible explanation would be that natural IgM, binding with low affinity to the antigen, may suffice to activate complement. In support of this, mice lacking secretory IgM have an impaired antibody response, which can be rescued by transfer of non-immune IgM. Moreover, passive administration of specific IgM together with antigen enhances the antibody response in a complement-dependent fashion. To test the idea, we have used a knock-in mouse strain (Cμ13) carrying a point mutation in the IgM heavy chain, rendering the IgM unable to activate complement. Mutant mice backcrossed to BALB/c or C57BL/6 background were primed and boosted with a low dose of sheep red blood cells. Confirming earlier data, no impairment in early, primary IgM- or IgG-responses were seen in either of the Cμ13 strains. However, in one of the mutant strains, late primary IgG responses were impaired. A more pronounced effect was observed after boost, when the IgG response, the number of germinal center B cells and antibody secreting cells as well as the opsonization of antigen were impaired in mutant mice. We conclude that complement activation by natural IgM cannot explain the role of C1q in primary antibody responses, but that endogenous, specific, wildtype IgM generated after immunization feedback-enhances the response to a booster dose of antigen. Importantly, this mechanism can only partially explain the role of complement in the generation of antibody responses because the IgG response was much lower in C3- or complement receptor 1 and 2-deficient mice than in Cμ13 mice.

Microglia TREM1-mediated neuroinflammation contributes to central sensitization via the NF-κB pathway in a chronic migraine model

BackgroundNeuroinflammation, mediated by the activation of microglia, contributes to central sensitization, which is associated with the development of chronic migraine (CM). TREM1 receptors amplify the inflammatory response. However, their relationship to CM is unclear. Thus, this study endeavoured to elucidate the exact role of TREM1 in CM.MethodsNitroglycerin (NTG) was repeatedly administered intraperitoneally to establish the CM model. Mechanical and thermal sensitivities were assessed using von Frey filaments and hot plate assays. Using Western blotting, TREM1, NF-κB pathway, NLRP3 inflammasome components, and proinflammatory cytokines were all detected. Immunofluorescence was used to examine the cellular distribution of TREM1 and NLRP3, the number of microglia, immunoreactivity, and morphological changes. We examined the effects of TREM1 antagonists (LR12) and NF-κB inhibitors (PDTC) on pain behaviour, as well as the production of c-fos and CGRP. Additionally, we investigated whether LR12 and PDTC affect the activation of microglia and the NLRP3 inflammasome. We synthesized siRNA and TREM1-overexpressing plasmids to transfect BV2 cells treated with LPS and normal BV2 cells and treated TREM1-overexpressing BV2 cells with PDTC. The NF-κB pathway, NLRP3 inflammasome components, and proinflammatory cytokines were quantified using Western blotting.ResultsFollowing NTG administration, the expression of TREM1 was significantly upregulated and exclusively localized in microglia in the TNC, and was well co-localized with NLRP3. Furthermore, activation of the classical NF-κB pathway was observed. Pre-treatment with LR12 and PDTC effectively attenuated mechanical hypersensitivity, suppressed the expression of c-fos and CGRP, and inhibited NF-κB activity in CM mice. Additionally, inhibition of TREM1 and NF-κB activity mitigated NTG-induced microglia and NLRP3 activation, as well as proinflammatory cytokines production. In vitro, knockdown of TREM1 resulted in attenuated activation of the NF-κB pathway following lipopolysaccharide (LPS) treatment and reduced expression of NLRP3 inflammasome components as well as proinflammatory cytokines. After TREM1 overexpression, the NF-κB pathway was activated, NLRP3 inflammasome components and proinflammatory cytokines were upregulated, and PDTC reversed this phenomenon.ConclusionsOur findings suggest that TREM1 regulates microglia and NLRP3 activation via the NF-κB pathway, thereby contributing to central sensitization and implicating its involvement in chronic migraine pathogenesis.

Deficiency of both classical and alternative end-joining pathways leads to a synergistic defect in double-strand break repair but not to an increase in homology-dependent gene targeting in Arabidopsis.

In eukaryotes, double-strand breaks (DSBs) are either repaired by homologous recombination (HR) or non-homologous end-joining (NHEJ). In somatic plant cells, HR is very inefficient. Therefore, the vast majority of DSBs are repaired by two different pathways of NHEJ. The classical (cNHEJ) pathway depends on the heterodimer KU70/KU80, while polymerase theta (POLQ) is central to the alternative (aNHEJ) pathway. Surprisingly, Arabidopsis plants are viable, even when both pathways are impaired. However, they exhibit severe growth retardation and reduced fertility. Analysis of mitotic anaphases indicates that the double mutant is characterized by a dramatic increase in chromosome fragmentation due to defective DSB repair. In contrast to the single mutants, the double mutant was found to be highly sensitive to the DSB-inducing genotoxin bleomycin. Thus, both pathways can complement for each other efficiently in DSB repair. We speculated that in the absence of both NHEJ pathways, HR might be enhanced. This would be especially attractive for gene targeting (GT) in which predefined changes are introduced using a homologous template. Unexpectedly, the polq single mutant as well as the double mutant showed significantly lower GT frequencies in comparison to wildtype plants. Accordingly, we were able to show that elimination of both NHEJ pathways does not pose an attractive approach for Agrobacterium-mediated GT. However, our results clearly indicate that a loss of cNHEJ leads to an increase in GT frequency, which is especially drastic and attractive for practical applications, in which the in planta GT strategy is used.

Microbial evasion of the complement system: a continuous and evolving story.

The complement system is a fundamental part of the innate immune system that plays a key role in the battle of the human body against invading pathogens. Through its three pathways, represented by the classical, alternative, and lectin pathways, the complement system forms a tightly regulated network of soluble proteins, membrane-expressed receptors, and regulators with versatile protective and killing mechanisms. However, ingenious pathogens have developed strategies over the years to protect themselves from this complex part of the immune system. This review briefly discusses the sequence of the complement activation pathways. Then, we present a comprehensive updated overview of how the major four pathogenic groups, namely, bacteria, viruses, fungi, and parasites, control, modulate, and block the complement attacks at different steps of the complement cascade. We shed more light on the ability of those pathogens to deploy more than one mechanism to tackle the complement system in their path to establish infection within the human host.

Enhancement of salt tolerance of alfalfa: Physiological and molecular responses of transgenic alfalfa plants expressing Syntrichia caninervis-derived ScABI3

Alfalfa (Medicago sativa L.), a perennial forage plant, is a rich source of nutrients such as vitamins, minerals, and proteins. Salt stress, however, impedes its growth. The plant-specific transcription factor abscisic acid insensitive 3 (ABI3) has a critical contribution to the control of abscisic acid (ABA) signaling pathway and abiotic stress response. The gene ScABI3 from Syntrichia caninervis, a moss species tolerant to desiccation, could be considered a potential candidate gene to modify alfalfa's nutritional and growth aspects. However, it remains unclear how ScABI3 affects the salt stress response of transgenic alfalfa. Therefore, we elucidated the role and molecular mechanism of ScABI3 from S. caninervis as an ABA signaling factor in transgenic alfalfa. Our findings demonstrate that ScABI3 overexpression in transgenic alfalfa improves salt tolerance by promoting relative water content, antioxidant enzyme activity, and photosynthetic parameters. Furthermore, the key genes of plant hormone signaling and the classical salt tolerance pathway were activated in ScABI3 transgenic lines under salt stress. Based on these results, ScABI3 could be considered a potentially critical candidate gene to alleviate salt stress in alfalfa. The present study provides valuable insights for developing transgenic crop breeding strategies for saline-alkaline soils.

Thrombotic microangiopathy following systemic AAV administration is dependent on anti-capsid antibodies.

BACKGROUNDSystemic administration of adeno-associated virus (AAV) can trigger life-threatening inflammatory responses, including thrombotic microangiopathy (TMA), acute kidney injury due to atypical hemolytic uremic syndrome-like complement activation, immune-mediated myocardial inflammation, and hepatic toxicity.METHODSWe describe the kinetics of immune activation following systemic AAV serotype 9 (AAV9) administration in 38 individuals following 2 distinct prophylactic immunomodulation regimens. Group 1 received corticosteroids and Group 2 received rituximab plus sirolimus in addition to steroids to prevent anti-AAV antibody formation.RESULTSGroup 1 participants had a rapid increase in immunoglobulin M (IgM) and IgG. Increase in D-dimer, decline in platelet count, and complement activation are indicative of TMA. All Group 1 participants demonstrated activation of both classical and alternative complement pathways, as indicated by depleted C4 and elevated soluble C5b-9, Ba, and Bb antigens. Group 2 patients did not have a significant change in IgM or IgG and had minimal complement activation.CONCLUSIONSThis study demonstrates that TMA in the setting of AAV gene therapy is antibody dependent (classical pathway) and amplified by the alternative complement pathway. Critical time points and interventions are identified to allow for management of immune-mediated events that impact the safety and efficacy of systemic gene therapy.