Complement Component 5a Receptor Research Articles

JR14a: A novel antagonist of C3aR attenuates neuroinflammation in cerebral ischemia-reperfusion injury

Cerebral ischemia-reperfusion injury (CIRI), a prevalent stroke-related complication, can lead to severe brain damage. Inflammation is a crucial factor in CIRI pathogenesis, and the complement component 3a receptor (C3aR) could be a key mediator in the post-CIRI inflammatory cascade. In this study, the role of C3aR in CIRI was investigated utilizing a middle cerebral artery occlusion (MCAO) model in C3aR knockout (KO) mice. Magnetic resonance imaging (MRI) and neurofunctional assessments revealed that C3aR KO mice exhibited significantly diminished cerebral infarction and improved neurological impairments. Consequently, the focus shifted to searching for a small molecule antagonist of C3aR. JR14a, a new potent thiophene antagonist of C3aR, was injected intraperitoneally into mice 1-h post-MCAO model implementation. The mass spectrometry (MS) results indicated the ability of JR14a to penetrate the blood-brain barrier. Subsequent TTC staining and neurofunctional assessments revealed the efficacy of JR14a in reducing cerebral infarct volume and neurological impairment following MCAO. In addition, immunofluorescence (IF) and immunohistochemistry (IHC) demonstrated attenuated microglial activation, neutrophil infiltration, and blood-brain barrier disruption by JR14a in the MCAO model. Furthermore, enzyme-linked immunosorbent assay (ELISA) and Western blotting supported the role of JR14a in downregulating the expression levels of C3aR, tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6), as well as the phosphorylation of p65. In conclusion, the findings suggested that C3aR could be a potential therapeutic target for CIRI, and JR14a emerged as a promising treatment candidate.

Deficiency of Complement C3a and C5a receptors Does Not Prevent Angiotensin II-Induced Hypertension and Hypertensive End-Organ Damage.

Complement may drive the pathology of hypertension through effects on innate and adaptive immune responses. Recently an injurious role for the anaphylatoxin receptors C3aR (complement component 3a receptor) and C5aR1 (complement component 5a receptor) in the development of hypertension was shown through downregulation of Foxp3+ (forkhead box protein 3) regulatory T cells. Here, we deepen our understanding of the therapeutic potential of targeting both receptors in hypertension. Data from the European Renal cDNA Bank, single cell sequencing and immunohistochemistry were examined in hypertensive patients. The effect of C3aR or C3aR/C5aR1 double deficiency was assessed in two models of Ang II (angiotensin II)-induced hypertension in knockout mice. We found increased expression of C3aR, C5aR1 and Foxp3 cells in kidney biopsies of patients with hypertensive nephropathy. Expression of both receptors was mainly found in myeloid cells. No differences in blood pressure, renal injury (albuminuria, glomerular filtration rate, glomerular and tubulointerstitial injury, inflammation) or cardiac injury (cardiac fibrosis, heart weight, gene expression) between control and mutant mice was discerned in C3aR-/- as well as C3aR/C5aR1-/- double knockout mice. The number of renal Tregs was not decreased in Ang II as well as in DOCA salt induced hypertension. Hypertensive nephropathy in mice and men is characterized by an increase of renal regulatory T cells and enhanced expression of anaphylatoxin receptors. Our investigations do not corroborate a role for C3aR/C5aR1 axis in Ang II-induced hypertension hence challenging the concept of anaphylatoxin receptor targeting in the treatment of hypertensive disease.

C5a drives the inflammatory response with bacterial dose effect by binding to C5aR1 in zebrafish infected with Aeromonas hydrophila.

The complement system is essential to host defense, but its excessive activation caused by severe pathogen invasion is a driving force in adverse inflammatory. The binding of complement component 5a (C5a) and complement component 5a receptor 1 (C5aR1) is the key to trigger complement-mediated inflammatory response in mammals. However, the role of C5a-C5aR1 axis in fish immune response remains obscure. In this study, the role of C5a-C5aR1 axis of zebrafish (Danio rerio) after serious infection with Aeromonas hydrophila was investigated. C5a and C5aR1 of zebrafish were cloned, with CDS sequences of 228 and 1041 bp, respectively, and they were widely expressed in various tissues with the highest expression in the liver and spleen, respectively. The survival of zebrafish was closely correlated to the dose of A. hydrophila. The cytokine storm occurred at high concentrations of A. hydrophila infection. At 24h post infection (hpi), the expression of C5a and C5aR1 in the spleen increased 26.8-fold and 9.9-fold in treatment group 1 (TG1, 3.0×107CFU/mL) (P<0.01), and 4.7-fold and 3.4-fold in treatment group 2 (TG2, 1.0×107CFU/mL) (P<0.05), respectively. Correspondingly, proinflammatory cytokines interleukin-1β (IL-1β), interleukin-8 (IL-8), and interleukin-17 (IL-17) were positively correlated to C5a and C5aR1at mRNA and protein expression levels. The expression of IL-1β was significantly increased in the spleen at 6 hpi, with a 599.2-fold and 203.2-fold upregulation in TG1 and TG2 (P<0.001), respectively. Moreover, after inhibition of C5a-C5aR1 binding treated with C5aR1 antagonist (W-54011), zebrafish showed lower expression of C5a, C5aR1, and cytokines, less intestinal damage, and significantly enhancement of survival (P<0.05) after A. hydrophila challenge. This study revealed that the inflammatory effect of C5a was achieved by binding to C5aR1 in zebrafish, providing novel insights into using C5a-C5aR1 axis as an effective target to reduce bacterial inflammation and disease in fish.

Activation of Type I Interferon and Complement component 3a receptor signaling mediates chronic stress-induced social behavior deficits

Inflammation is involved in the pathophysiology of many neuropsychiatric disorders. Chronic stress increases inflammation and alters protective immune responses. Type I interferons (IFN-I) are key players in the peripheral inflammatory response and are implicated in mood and behavior. In the present study, we investigated the roles of neuroinflammation and synapse loss in IFN-I signaling-mediated social behavior deficits under chronic stress conditions. Our findings show that chronic unpredictable stress (CUS) induces significant increases in serum IFNβ levels. Also, CUS induced microglial activation and synapse loss in the PFC of mice. Peripheral blockade of IFN-I signaling attenuated CUS-induced neuroimmune alterations and social behavioral deficits. Furthermore, complement component 3a receptor 1 (C3ar1) mediates systemic IFNβ-induced neuroinflammation and social behavior deficits. Together, these findings identify a key role of C3ar1 in IFN-I-mediated social behavior deficits under chronic stress conditions.

Stress, immune mechanisms, and mental illness: from etiology to treatment

It is now well-established that a strong relationship exists between psychiatric disorders and disturbances of immune mechanisms at different stages of life. Indeed, a derangement in the expression and function of specific immune elements may predispose and contribute to the development of different psychopathologic conditions and could also represent an important component that must be targeted by therapeutic intervention. The proposed symposium will summarize authoritatively the important recent advances on the immune mechanisms contributing to the etiology of psychiatric disorders, with a specific focus on the role of stress, and it will discuss if and how the effectiveness of therapeutic intervention may depend on the ability to regulate immune mechanisms that are altered under pathologic conditions. The symposium will unfold upon the discussion of preclinical and clinical studies addressing different aspects related to psychiatric disorders. Prof. A. Pillai (University of Texas, U.S.A.) will specifically discuss the roles of neuroinflammation and synapse loss in IFN-I signaling-mediated social deficits under chronic stress conditions. He will discuss how peripheral blockade of IFN-I signaling attenuated stress-induced neuroimmune alterations and social behavioral deficits and also the key role of complement component 3a receptor 1 (C3ar1) in these mechanisms. Along this line, Prof. M.A. Riva (University of Milan, Italy) will show how immune mechanisms and dysregulation of microglial function may contribute to the vulnerability to stress at different life stages, from adolescence to adulthood. Moreover, he will discuss how the normalization of some of these mechanisms following pharmacological intervention may contribute to therapeutic effectiveness in the treatment of different psychiatric disorders characterized, particularly in patients characterized by a higher inflammatory status. At the clinical level, Dr. G. Scaini (University of Texas, U.S.A.) will discuss the interplay between inflammation, stress, and mitochondria in bipolar disorder patients. She will highlight the significant correlation between alterations in mitochondrial proteins and inflammatory markers with a decrease in functional status and an increase in the severity of symptoms. Lastly, Prof. J. Felger (Emory Univ.) will discuss how depressed patients with increased inflammation show altered functional connectivity within the ventral striatum to ventromedial prefrontal cortex reward circuitry in relation to symptoms of anhedonia. These changes can be improved by pharmacological intervention aimed to increase dopamine levels suggesting that functional connectivity in reward circuitry may also serve as a modifiable imaging biomarker for the efficacy of pharmacological and/or behavioral interventions to reverse the impact of inflammation on the brain of depressed subjects.

Sexually‐dimorphic effects of C3aR1 disruption in a mouse model of Alzheimer’s disease

AbstractBackgroundThe complement system can act as part of the innate immune system as well as the adaptive immune system. Complement activation in the brain has been shown to play a role in neurogenesis, synaptic pruning and plasticity. It has also been specifically involved in microglial function, since our group has previously shown that when the complement component 3a receptor 1 (C3aR1) is disrupted in mice, phagocytosis by primary microglia cells is impaired as compared to WT microglia. Therefore, it is hypothesized that complement might have neuroprotective effects early in life but neurotoxic effects later in life or within a disease context, such as during Alzheimer’s disease (AD) impeding to effectively clear hallmarks of the disease.MethodWe performed bulk RNA sequencing (RNAseq) analysis from cortex tissue of 8mo old male and female wild‐type, C3aR1 knock‐out (KO), 5xFAD and 5xFAD‐C3aR1 KO mice. We also performed Barnes Maze on these mice to assess spatial learning and memory, as well as biochemical assays to measure several AD markers.ResultData obtained from RNAseq analysis has shown a sexually‐dimorphic trend in various differentially‐expressed genes (DEGs). For instance, when comparing 5xFAD‐C3aR1 KO with 5xFAD mice, male mice show downregulated DEGs in axon guidance and calcium signaling pathways according to KEGG pathway terms, but this is not seen in females. Similarly, barnes maze has shown sexually‐dimorphic differences, with 5xFAD‐C3aR1 KO male mice showing improve memory and learning when compared to 5xFAD mice, and no differences are seen in female mice.ConclusionInterestingly, other studies in humans have shown increased complement proteins in the cerebrospinal fluid of men, compared to women, showing the importance of studying the complement system in both males and females. Assessing these sexually‐dimorphic differences in this AD mouse model is also important, given that generally not only female mice present more aggressive phenotypes of AD, but also AD in humans is more prevalent in women.

Paclitaxel binds and activates C5aR1: A new potential therapeutic target for the prevention of chemotherapy-induced peripheral neuropathy and hypersensitivity reactions

Chemotherapy-induced peripheral neuropathy (CIPN) and hypersensitivity reactions (HSRs) are among the most frequent and impairing side effects of the antineoplastic agent paclitaxel. Here, we demonstrated that paclitaxel can bind and activate complement component 5a receptor 1 (C5aR1) and that this binding is crucial in the etiology of paclitaxel-induced CIPN and anaphylaxis. Starting from our previous data demonstrating the role of interleukin (IL)-8 in paclitaxel-induced neuronal toxicity, we searched for proteins that activate IL-8 expression and, by using the Exscalate platform for molecular docking simulations, we predicted the high affinity of C5aR1 with paclitaxel. By in vitro studies, we confirmed the specific and competitive nature of the C5aR1-paclitaxel binding and found that it triggers intracellularly the NFkB/P38 pathway and c-Fos. In F11 neuronal cells and rat dorsal root ganglia, C5aR1 inhibition protected from paclitaxel-induced neuropathological effects, while in paclitaxel-treated mice, the absence (knock-out mice) or the inhibition of C5aR1 significantly ameliorated CIPN symptoms—in terms of cold and mechanical allodynia—and reduced the chronic pathological state in the paw. Finally, we found that C5aR1 inhibition can counteract paclitaxel-induced anaphylactic cytokine release in macrophages in vitro, as well as the onset of HSRs in mice. Altogether these data identified C5aR1 as a key mediator and a new potential pharmacological target for the prevention and treatment of CIPN and HSRs induced by paclitaxel.

CD63 and C3AR1: The Potential Molecular Targets in the Progression of Septic Shock.

BackgroundThe molecular mechanism of septic shock is unknown. We studied the pathogenesis of septic shock and provide a novel strategy for treating and improving the prognosis of septic shock.MethodsGluten-Sensitive Enteropathy (GSE) 131761, GSE119217, GSE26378 datasets were downloaded from the Gene Expression Omnibus (GEO) database. The three datasets included 204 septic shock samples and 48 normal samples. The R packages “affy” and “limma” were employed to identify the differently expressed genes (DEGs) between septic shock and normal samples. Weighted gene co-expression network analysis (WGCNA) was performed to search for modules that play an important role in septic shock. Functional annotation of DEGs and construction and analysis of hub genes were used to explore the pathomechanism of septic shock. The receiver operating characteristic (ROC) curves were obtained using MedCalc software. The drug molecules that could regulate hub genes associated with septic shock were searched for in the CMap database. An animal model of septic shock was constructed to analyze the role of these hub genes.ResultsThe merged series contained 321 up-regulated and 255 down-regulated genes. WGCNA showed the brown module had the highest correlation with the status of septic shock. GO and KEGG enrichment analysis results of the brown module genes showed they were mainly enriched in “leukocyte differentiation”, “Ras-proximate-1 (Rap1) signaling pathway”, and “cytokine–cytokine receptor interaction”. Through construction and analysis of a protein–protein interaction (PPI) network, cluster of differentiation 63 (CD63) and complement component 3a receptor 1 (C3AR1) were identified as hub genes of septic shock. The area under curve (AUC) of C3AR1 for the septic shock is 0.772 (P<0.001), and the AUC of CD63 for the septic shock is 0.871 (P<0.001). Small molecule drugs were filtered by the number of instances (n>3) and P-values <0.05, including “monensin”, “verteporfin”, “ikarugamycin”, “tetrahydroalstonine”, “cefamandole”, “etoposide”. In the animal model, the relative expression levels of interleukin-6 (IL-6), Tumor Necrosis Factor-α (TNF-α), and lactic acid were significantly higher in the septic shock group compared with the control group. Results of Real Time Quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA) analysis for CD63 and C3AR1 showed that their relative expression levels were significantly lower in the septic shock group compared with the control group (P<0.05).ConclusionCD63 and C3AR1 are significant hub genes of septic shock and may represent potential molecular targets for future studies of septic shock.

Predictive and Prognostic Biomarkers for Lung Cancer Bone Metastasis and Their Therapeutic Value

Lung cancer is the leading cause of cancer-related death worldwide. Bone metastasis, which usually accompanies severe skeletal-related events, is the most common site for tumor distant dissemination and detected in more than one-third of patients with advanced lung cancer. Biopsy and imaging play critical roles in the diagnosis of bone metastasis; however, these approaches are characterized by evident limitations. Recently, studies regarding potential biomarkers in the serum, urine, and tumor tissue, were performed to predict the bone metastases and prognosis in patients with lung cancer. In this review, we summarize the findings of recent clinical research studies on biomarkers detected in samples obtained from patients with lung cancer bone metastasis. These markers include the following: (1) bone resorption-associated markers, such as N-terminal telopeptide (NTx)/C-terminal telopeptide (CTx), C-terminal telopeptide of type I collagen (CTx-I), tartrate-resistant acid phosphatase isoform 5b (TRACP-5b), pyridinoline (PYD), and parathyroid hormone related peptide (PTHrP); (2) bone formation-associated markers, including total serum alkaline phosphatase (ALP)/bone specific alkaline phosphatase(BAP), osteopontin (OP), osteocalcin (OS), amino-terminal extension propeptide of type I procollagen/carboxy-terminal extension propeptide of type I procollagen (PICP/PINP); (3) signaling markers, including epidermal growth factor receptor/Kirsten rat sarcoma/anaplastic lymphoma kinase (EGFR/KRAS/ALK), receptor activator of nuclear factor κB ligand/receptor activator of nuclear factor κB/osteoprotegerin (RANKL/RANK/OPG), C-X-C motif chemokine ligand 12/C-X-C motif chemokine receptor 4 (CXCL12/CXCR4), complement component 5a receptor (C5AR); and (4) other potential markers, such as calcium sensing receptor (CASR), bone sialoprotein (BSP), bone morphogenetic protein 2 (BMP2), cytokeratin 19 fragment/carcinoembryonic antigen (CYFRA/CEA), tissue factor, cell-free DNA, long non-coding RNA, and microRNA. The prognostic value of these markers is also investigated. Furthermore, we listed some clinical trials targeting hotspot biomarkers in advanced lung cancer referring for their therapeutic effects.

(-)-Oleocanthal Nutraceuticals for Alzheimer's Disease Amyloid Pathology: Novel Oral Formulations, Therapeutic, and Molecular Insights in 5xFAD Transgenic Mice Model.

Alzheimer’s disease (AD) is a complex progressive neurodegenerative disorder affecting humans mainly through the deposition of Aβ-amyloid (Aβ) fibrils and accumulation of neurofibrillary tangles in the brain. Currently available AD treatments only exhibit symptomatic relief but do not generally intervene with the amyloid and tau pathologies. The extra-virgin olive oil (EVOO) monophenolic secoiridoid S-(–)-oleocanthal (OC) showed anti-inflammatory activity through COX system inhibition with potency comparable to the standard non-steroidal anti-inflammatory drug (NSAID) like ibuprofen. OC also showed positive in vitro, in vivo, and clinical therapeutic effects against cardiovascular diseases, many malignancies, and AD. Due to its pungent, astringent, and irritant taste, OC should be formulated in acceptable dosage form before its oral use as a potential nutraceutical. The objective of this study is to develop new OC oral formulations, assess whether they maintained OC activity on the attenuation of β-amyloid pathology in a 5xFAD mouse model upon 4-month oral dosing use. Exploration of potential OC formulations underlying molecular mechanism is also within this study scope. OC powder formulation (OC-PF) and OC-solid dispersion formulation with erythritol (OC-SD) were prepared and characterized using FT-IR spectroscopy, powder X-ray diffraction, and scanning electron microscopy (ScEM) analyses. Both formulations showed an improved OC dissolution profile. OC-PF and OC-SD improved memory deficits of 5xFAD mice in behavioral studies. OC-PF and OC-SD exhibited significant attenuation of the accumulation of Aβ plaques and tau phosphorylation in the brain of 5xFAD female mice. Both formulations markedly suppressed C3AR1 (complement component 3a receptor 1) activity by targeting the downstream marker STAT3. Collectively, these results demonstrate the potential for the application of OC-PF as a prospective nutraceutical or dietary supplement to control the progression of amyloid pathogenesis associated with AD.

Structural insights from an in silico molecular docking simulation of complement component 3a receptor 1 with an antagonist

Complement component 3a receptor 1 (C3aR) is an anaphylatoxin receptor that mediates inflammatory processes. Although considerable effort has gone into discovering the antagonists and agonists of C3aR, structural insights are required to search for effective ligands and to elucidate their binding modes and the mechanism of activation and inactivation. No experimental structural data of C3aR have yet been reported. We investigated the binding mode of an antagonist of C3aR using a combination of homology modeling, ligand docking, molecular dynamics simulations, and binding free energy calculations. We produced a plausible binding model consistent with the reported experimental data. We believe that this model is appropriate for the identification of new C3aR antagonists, as it can distinguish between antagonists and decoy compounds.

High Expression of the Component 3a Receptor 1 (C3AR1) Gene in Stomach Adenocarcinomas Infers a Poor Prognosis and High Immune-Infiltration Levels.

BackgroundThis study was designed to explore the incompletely investigated role of the complement component 3a receptor 1 (C3AR1) in the prognosis of stomach adenocarcinomas (STAD).Material/MethodsUsing bioinformatic methods, we systematically determined the expression and prognosis value of C3AR1 in various cancers by using the TIMER (Tumor Immune Estimation Resource) database, UALCAN platform, GEPIA (Gene Expression Profiling Interactive Analysis) server, and the OncoLnc tool. The biological processes influenced by C3AR1 were determined using the GSEA (Gene Set Enrichment Analysis) software (Copyright 2004–2020 Broad Institute, Inc., Massachusetts Institute of Technology, and Regents of the University of California). The correlation between C3AR1 expression and the immune-infiltrating cells as well as the correlation analysis between C3AR1 expression and the corresponding immune-marker sets were conducted using the TIMER and GEPIA databases.ResultsThe expression of C3AR1 was significantly (P<0.001) differentially expressed on several tumor types, while its prognosis value could only be determined on STAD, with a high expression of C3AR1 closely correlated with a poor prognosis. The GSEA analysis revealed that the differential expression of C3AR1 profoundly affected the immune-related biological processes. The expression of C3AR1 was strongly and positively correlated with the infiltration of monocytes, tumor-associated macrophages, M2 macrophages, dendritic cells, and exhausted T cells.ConclusionsOur results have revealed that a high expression of C3AR1 is positively correlated with a poor prognosis and increased tumor-immune infiltration. C3AR1 can promote the polarization of M2 macrophages and T cell exhaustion, leading to the immune escape of STAD. These findings suggest that C3AR1 could be used as a prognostic and immune-infiltration marker in the pathogenesis of STAD.

Attenuating ischemia/reperfusion injury in rat cardiac transplantation by intracoronary infusion with siRNA cocktail solution.

Tumor necrosis factor-α (TNF-α), caspase-8, and complement component 5a receptor (C5aR) are known to play a crucial role in the myocardial ischemia/reperfusion (I/R) injury in cardiac transplantation. We hypothesized that the intracoronary infusion of TNF-α, caspase-8, and C5aR small interfering RNAs (siRNA) would protect cardiac allograft function and improve graft survival from I/R injury-induced organ failure. I/R injury of cardiac allograft was induced by syngeneic rat cardiac transplantation, in which the transplanted hearts were infused with saline or different amounts of siRNA cocktail solution targeting TNF-α, caspase-8, and C5aR via coronary arteries, and subsequently subjected to 18 h of preservation at 4°C in histidine–tryptophan–ketoglutarate (HTK) solution. The effects of siRNA cocktail solution on prolonged cold I/R injury were determined by assessing graft survival, histopathological changes, myeloperoxidase (MPO) activity, and malondialdehyde (MDA) concentration. The perfused siRNA cocktail solution successfully knocked down the expression of TNF-α, caspase-8, and C5aR in vitro and in vivo. Approximately 91.7% of control hearts that underwent 18 h of cold ischemia ceased their function after transplantation; however, 87.5% of cardiac allografts from the highest dose siRNA cocktail solution-pretreated hearts survived >14 days and exhibited minimal histological changes, with minimal cellular infiltration, interstitial edema, and inflammation and maximal reduced MPO activity and MDA concentration in the cardiac allograft. We demonstrated the feasibility and efficiency of infusion of TNF-α, caspase-8, and C5aR siRNA via the intracoronary route as a promising strategy for gene silencing against I/R injury in cardiac transplantation.

C5a/C5aR1 Pathway Is Critical for the Pathogenesis of Psoriasis.

Psoriasis is one of the most common chronic inflammatory skin diseases, affecting ~2% of the population. The lack of characterization of the pathogenesis of psoriasis has hindered efficient clinical treatment of the disease. In our study, we observed that expression of complement component 5a receptor 1(C5aR1) was significantly increased in skin lesions of both imiquimod (IMQ) and IL23-induced psoriatic mice and patients with psoriasis. C5aR1 deficiency or treatment with C5a receptor 1 antagonist (C5aR1a) in mice significantly attenuated psoriasis-like skin lesions and expression of inflammatory cytokines and chemokines. Moreover, C5aR1 deficiency significantly decreased IMQ-induced infiltration of plasmacytoid dendritic cells (pDCs), monocytes and neutrophils in psoriatic skin lesions and functions of pDCs, evidenced by the remarkable reduction in the IMQ-induced production of interferon-α (IFN-α) and tumor necrosis factor α (TNF-α), and FMS-like tyrosine kinase 3 ligand (FLT3L)-dependent pDCs differentiation. Accordingly, in vitro treatment with recombinant C5a accelerated pDCs migration and the differentiation of bone marrow cells into pDCs. Furthermore, biopsies of psoriatic patients showed a dramatic increase of C5aR1+ pDCs infiltration in psoriatic skin lesions, compared to healthy subjects. Our results provide direct evidence that C5a/C5aR1 signaling plays a critical role in the pathogenesis of psoriasis. Inhibition of C5a/C5aR1 pathway is expected to be beneficial in the treatment of patients with psoriasis.

Sensory Neurons of the Dorsal Root Ganglia Become Hyperexcitable in a T-Cell-Mediated MOG-EAE Model of Multiple Sclerosis

Multiple sclerosis (MS) is an autoimmune, demyelinating disease of the central nervous system. Patients with MS typically present with visual, motor, and sensory deficits. However, an additional complication of MS in large subset of patients is neuropathic pain. To study the underlying immune-mediated pathophysiology of pain in MS we employed the myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalitis (EAE) model in mice. Since sensory neurons are crucial for nociceptive transduction, we investigated the effect of this disease on sensory neurons of the lumbar dorsal root ganglia (DRG). Here, we report the disease was associated with activation of the complement system and the NLRP3 inflammasome in the DRG. We further observe a transient increase in the number of complement component 5a receptor 1-positive (C5aR1+) immune cells, CD4+ T-cells, and Iba1+ macrophages in the DRG. The absence of any significant change in the levels of mRNA for myelin proteins in the DRG and the sciatic nerve suggests that demyelination in the PNS is not a trigger for the immune response in the DRG. However, we did observe an induction of activating transcription factor 3 (ATF3) at disease onset and chronic disruption of cytoskeletal proteins in the DRG demonstrating neuronal injury in the PNS in response to the disease. Electrophysiological analysis revealed the emergence of hyperexcitability in medium-to-large (≥26 µm) diameter neurons, especially at the onset of MOG-EAE signs. These results provide conclusive evidence of immune activation, neuronal injury, and peripheral sensitization in MOG-EAE, a model classically considered to be centrally mediated.

Important roles of C5a and C5aR in tumor development and cancer treatment

The complement system is part of the body's innate defense immune system, which can identify and eliminate invasive pathogenic microorganisms to maintain normal life activities. Complement Component 5a (C5a) is an active anaphylatoxin produced after complement system activation, closely related to tumor formation. C5a is highly expressed in a variety of tumors, and combines with its Complement Component 5a Receptor (C5aR) to increase the proliferation and migration of tumor cells. This review will comprehensively elaborate the important role of C5a/C5aR in the process of tumor genesis and development from the three aspects of signal transduction pathways related to tumor, C5a/C5aR and tumor formation, and C5a/C5aR inhibitors and tumor therapy. Finally, the principle of complement inhibition is used to inhibit tumor metastasis, reduce the rate of tumor diffusion, and control the trend of tumor deterioration.

Distinct neutrophil C5a receptor inflammatory events in cows initiated by chemoattractant C5a and lipopolysaccharide around parturition and in mid lactation

In neutrophils, toll-like receptor and complement component 5a (C5a) signaling are critical pathways regulating innate immunity. In cows, not much is known about the second C5a receptor, complement component 5a receptor 2 (C5AR2). It is an interesting player in sepsis treatment because it is considered to have an anti-inflammatory effect during normal inflammation. Periparturient cows are prone to severe infections, and the objectives of this study were to investigate the expression and functionality of C5AR2 during peripartum. We investigated the effect of 2 major inflammatory stimuli, C5a and lipopolysaccharide (LPS), on the expression of a selected number of genes (C5AR1, C5AR2, TLR4, ITGAM, COX2, and CXCL8) and functions linked to these receptors. Overall, TLR4, ITGAM, and C5AR2, all of which are involved in early inflammation, showed a lower expression in periparturient cows. However, an overall lower expression seems not to be the only explanation for the increased risk of sepsis in periparturient cows. Normally, in response to inflammation and as seen in the mid-lactation group, the expression of these genes increases after stimulation with LPS. However, in periparturient cows, stimulation with LPS led to a decrease in expression of these receptors, indicating a different response of neutrophils in response to LPS during this period. A decrease in ITGAM (coding for CD11b) expression complicates correct neutrophil localization and phagocytosis. Its downregulation upon stimulation might be detrimental for adequate eradication of the pathogen and might increase the risk of an imbalanced inflammation; C5AR2 seems to play a central role in this altered response. In addition, myeloperoxidase (MPO) activity in periparturient cows is lower in response to C5a stimulation. It has been suggested that MPO plays an important role in neutrophil shutdown and, thereby, timely resolution of inflammation. A decreased MPO activity might thus prolong the inflammatory reaction of the neutrophils. This finding was supported by the increased viability of the neutrophils obtained from periparturient cows. Even after stimulation, we found a lower caspase-3 activity in this group, indicating that they might be activated for a longer time compared with the neutrophils from mid-lactation cows. Accordingly, these alterations might contribute to a temporal mismatch in inflammatory responses, as often seen in severe periparturient infections.

Complement component 3a receptor deficiency attenuates chronic stress-induced monocyte infiltration and depressive-like behavior

Major depressive disorder (MDD) is one of the most common and debilitating neuropsychiatric illnesses. Accumulating evidence suggests a potential role of the immune system in the pathophysiology of MDD. The complement system represents one of the major effector mechanisms of the innate immune system, and plays a critical role in inflammation. However, the role of complement components in MDD is not well understood. Here, we found significant increase in component 3 (C3) expression in the prefrontal cortex (PFC) of depressed suicide subjects. We tested the role of altered C3 expression in mouse model of depression and found that increased C3 expression in PFC as a result of chronic stress causes depressive-like behavior. Conversely, mice lacking C3 were resilient to stress-induced depressive-like behavior. Moreover, selective overexpression of C3 in PFC was sufficient to cause depressive-like behavior in mice. We found that C3a (activated product of C3) receptor, C3aR+ monocytes were infiltrated into PFC following chronic stress. However, C3aR knockout mice displayed significantly reduced monocyte recruitment into PFC and reduced levels of the proinflammatory cytokine IL-1β in PFC after chronic stress. In addition, C3aR knockout mice did not exhibit chronic stress-induced behavior despair. Similarly, chronic stress-induced increases in C3aR+ monocytes and IL-1β in PFC, and depressive-like behavior were attenuated by myeloid cell depletion. These postmortem and preclinical studies identify C3aR signaling as a key factor in MDD pathophysiology.

The Model Structures of the Complement Component 5a Receptor (C5aR) Bound to the Native and Engineered hC5a

The interaction of hC5a with C5aR, previously hypothesized to involve a “two-site” binding, (i) recognition of the bulk of hC5a by the N-terminus (NT) of C5aR (“site1”), and (ii) recognition of C-terminus (CT) of hC5a by the extra cellular surface (ECS) of the C5aR (“site2”). However, the pharmacological landscapes of such recognition sites are yet to be illuminated at atomistic resolution. In the context, unique model complexes of C5aR, harboring pharmacophores of diverse functionality at the “site2” has recently been described. The current study provides a rational illustration of the “two-site” binding paradigm in C5aR, by recruiting the native agonist hC5a and engineered antagonist hC5a(A8). The hC5a-C5aR and hC5a(A8)-C5aR complexes studied over 250 ns of molecular dynamics (MD) each in POPC bilayer illuminate the hallmark of activation mechanism in C5aR. The intermolecular interactions in the model complexes are well supported by the molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) based binding free energy calculation, strongly correlating with the reported mutational studies. Exemplified in two unique and contrasting molecular complexes, the study provides an exceptional understanding of the pharmacological divergence observed in C5aR, which will certainly be useful for search and optimization of new generation “neutraligands” targeting the hC5a-C5aR interaction.

Deficiency of Complement C3a and C5a Receptors Prevents Angiotensin II-Induced Hypertension via Regulatory T Cells.

Inflammation and immunity play crucial roles in the development of hypertension. Complement activation-mediated innate immune response is involved in the regulation of hypertension and target-organ damage. However, whether complement-mediated T-cell functions could regulate blood pressure elevation in hypertension is still unclear. We aim to determine whether C3aR (complement component 3a receptor) and C5aR (complement component 5a receptor) could regulate blood pressure via modulating regulatory T cells (Tregs). We showed that angiotensin II (Ang II)-induced hypertension resulted in an elevated expression of C3aR and C5aR in Foxp3 (forkhead box P3)+ Tregs. By using C3aR and C5aR DKO (double knockout) mice, we showed that C3aR and C5aR deficiency together strikingly decreased both systolic and diastolic blood pressure in response to Ang II compared with WT (wild type), single C3aR-deficient (C3aR-/-), or C5aR-deficient (C5aR-/-) mice. Flow cytometric analysis showed that Ang II-induced Treg reduction in the kidney and blood was also blocked in DKO mice. Histological analysis indicated that renal and vascular structure remodeling and damage after Ang II treatment were attenuated in DKO mice compared with WT mice. In vitro, Ang II was able to stimulate C3aR and C5aR expression in cultured CD4+CD25+ natural Tregs. CD3 and CD28 antibody stimuli downregulated Foxp3 expression in WT but not DKO Tregs. More important, depletion of Tregs with CD25 antibody abolished the protective effects against Ang II-induced hypertension and target-organ damage in DKO mice. Adoptive transfer of DKO Tregs showed much more profound protective effects against Ang II-induced hypertension than WT Treg transfer. Furthermore, we demonstrated that C5aR expression in Foxp3+ Tregs was higher in hypertensive patients compared with normotensive individuals. C3aR and C5aR DKO-mediated Treg function prevents Ang II-induced hypertension and target-organ damage. Targeting C3aR and C5aR in Tregs specifically may be an alternative novel approach for hypertension treatment.