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1.
J Immunol Methods ; 531: 113709, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38862098

RESUMO

The complement system plays a crucial role in orchestrating the activation and regulation of inflammation within the human immune system. Three distinct activation pathways-classical, lectin, and alternative-converge to form the common lytic pathway, culminating in the formation of the membrane-attacking complex that disrupts the structure of pathogens. Dysregulated complement system activity can lead to tissue damage, autoimmune diseases, or immune deficiencies. In this study, the antimicrobial activity of human serum was investigated by using a bioluminescent microbe probe, Escherichia coli (pEGFPluxABCDEamp). This probe has previously been used to determine the antimicrobial activity of complement system and the polymorphonuclear neutrophils. In this study, blocking antibodies against key serum activators and components, including IgG, complement component 1q, factor B, and properdin, were utilized. The influence of body temperature and acute phase proteins, such as C reactive protein (CRP) and serum amyloid alpha (SAA), on the complement system was also examined. The study reveals the critical factors influencing complement system activity and pathway function. Alongside crucial factors like C1q and IgG, alternative pathway components factor B and properdin played pivotal roles. Results indicated that the alternative pathway accounted for approximately one third of the overall serum antimicrobial activity, and blocking this pathway disrupted the entire complement system. Contrary to expectations, elevated body temperature during inflammation did not enhance the antimicrobial activity of human serum. CRP demonstrated complement activation properties, but at higher physiological concentrations, it exhibited antagonistic tendencies, dampening the response. On the other hand, SAA enhanced the serum's activity. Overall, this study sheds a light on the critical factors affecting both complement system activity and pathway functionality, emphasizing the importance of a balanced immune response.


Assuntos
Temperatura Corporal , Proteína C-Reativa , Ativação do Complemento , Complemento C1q , Fator B do Complemento , Properdina , Proteína Amiloide A Sérica , Humanos , Proteína C-Reativa/metabolismo , Proteína C-Reativa/imunologia , Complemento C1q/imunologia , Complemento C1q/metabolismo , Proteína Amiloide A Sérica/metabolismo , Proteína Amiloide A Sérica/imunologia , Properdina/imunologia , Properdina/metabolismo , Fator B do Complemento/metabolismo , Fator B do Complemento/imunologia , Temperatura Corporal/imunologia , Escherichia coli/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia
2.
Pediatr Infect Dis J ; 43(8): e282-e284, 2024 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-38753997

RESUMO

A 13-year-old boy was admitted with severe meningococcal meningitis. Immunologic workup revealed a properdin deficiency, and genetic sequencing of CFP identified a novel, private and predicted pathogenic variant in exon 8. The patient received broad immunizations and penicillin prophylaxis. Children with invasive meningococcal disease should be tested for complement deficiency.


Assuntos
Meningite Meningocócica , Properdina , Humanos , Masculino , Meningite Meningocócica/diagnóstico , Adolescente , Properdina/deficiência , Properdina/genética , Neisseria meningitidis/genética
3.
Perit Dial Int ; 44(1): 37-47, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37794761

RESUMO

BACKGROUND: Despite several advantages compared to haemodialysis (HD), peritoneal dialysis (PD) remains an underused dialysis technique due to its high technique failure rate related to membrane fibrosis and peritonitis events. Previous work has suggested a harmful role for the complement system in these processes, highlighting the need for a more comprehensive examination in PD. METHODS: Plasma levels of C1q, mannose-binding lectin (MBL), Properdin, Factor D, C3d/C3-ratio and soluble membrane attack complex (sC5b-9) were determined in PD patients (n = 55), HD patients (n = 41), non-dialysis chronic kidney disease (CKD) patients (n = 15) and healthy controls (n = 14). Additionally, C1q, MBL, Properdin, Factor D and sC5b-9 levels were assessed in the peritoneal dialysis fluid (PDF). In a subgroup, interleukin-6, matrix metalloproteinase-2 (MMP-2), myeloperoxidase (MPO) and elastase were measured in the PDF. RESULTS: PD patients had significantly higher systemic levels of sC5b-9 compared to healthy controls, CKD and HD patients (p < 0.001). Plasma levels of C1q and C3d/C3-ratios were significantly associated with systemic sC5b-9 levels (p < 0.001). Locally, sC5b-9 was detected in the PDF of all PD patients, and levels were approximately 33% of those in matched plasma, but they did not correlate. In the PDF, only Properdin levels remained significantly associated with PDF sC5b-9 levels in multivariate analysis (p < 0.001). Additionally, PDF levels of sC5b-9 positively correlated with elastase, MPO and MMP-2 levels in the PDF (p < 0.01). CONCLUSIONS: Our data reveal both systemic and local complement activation in PD patients. Furthermore, these two processes seem independent considering the involvement of different pathways and the lack of correlation.


Assuntos
Diálise Peritoneal , Insuficiência Renal Crônica , Humanos , Diálise Peritoneal/efeitos adversos , Metaloproteinase 2 da Matriz , Properdina , Fator D do Complemento , Complemento C1q , Ativação do Complemento , Soluções para Diálise , Elastase Pancreática
4.
Kidney Int ; 105(1): 177-188, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37923132

RESUMO

Activation of the alternative pathway (AP) of complement is involved in the pathogenesis of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), although the underlying molecular mechanisms are unclear. To gain insight into the role of the AP, common gene variants in CFH/CFHR1-5, CFB, C3 and MCP, and longitudinal determinations of plasma C3, C4, FH, FHR-1, FHR-2, FHR-5, FB, properdin and sC5b-9 levels were analyzed in a Spanish AAV cohort consisting of 102 patients; 54 with active AAV (active cohort) and 48 in remission not receiving immunosuppressants or dialysis therapy (remission cohort). The validation cohort consisted of 100 patients with ANCA-associated glomerulonephritis. Here, we demonstrated that common genetic variants in complement components of the AP are associated with disease susceptibility (CFB32Q/W) or severity of kidney damage in AAV (CFH-H1, CFH1H2 and ΔCFHR3/1). Plasma levels of complement components were significantly different between active and remission cohorts. In longitudinal observations, a high degree of AP activation at diagnosis was associated with worse disease outcome, while high basal FHR-1 levels and lower FH/FHR-1 ratios determined severe forms of kidney associated AAV. These genetic and plasmatic findings were confirmed in the validation cohort. Additionally, autoantibodies against FH and C3 convertase were identified in one and five active patients, respectively. Thus, our study identified key genetic and plasma components of the AP that determine disease susceptibility, prognosis, and severity in AAV. Our data also suggests that balance between FH and FHR-1 is critical and supports FHR-1 as a novel AP-specific therapeutic target in AAV.


Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos , Anticorpos Anticitoplasma de Neutrófilos , Humanos , Suscetibilidade a Doenças , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/diagnóstico , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/genética , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/complicações , Fatores Imunológicos , Properdina/genética
5.
J Neuroinflammation ; 20(1): 260, 2023 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-37951917

RESUMO

BACKGROUND: Emerging evidence has shown that myeloid cells that infiltrate into the peri-infarct region may influence the progression of ischemic stroke by interacting with microglia. Properdin, which is typically secreted by immune cells such as neutrophils, monocytes, and T cells, has been found to possess damage-associated molecular patterns (DAMPs) properties and can perform functions unrelated to the complement pathway. However, the role of properdin in modulating microglia-mediated post-stroke neuroinflammation remains unclear. METHODS: Global and conditional (myeloid-specific) properdin-knockout mice were subjected to transient middle cerebral artery occlusion (tMCAO). Histopathological and behavioral tests were performed to assess ischemic brain injury in mice. Single-cell RNA sequencing and immunofluorescence staining were applied to explore the source and the expression level of properdin. The transcriptomic profile of properdin-activated primary microglia was depicted by transcriptome sequencing. Lentivirus was used for macrophage-inducible C-type lectin (Mincle) silencing in microglia. Conditioned medium from primary microglia was administered to primary cortex neurons to determine the neurotoxicity of microglia. A series of cellular and molecular biological techniques were used to evaluate the proinflammatory response, neuronal death, protein-protein interactions, and related signaling pathways, etc. RESULTS: The level of properdin was significantly increased, and brain-infiltrating neutrophils and macrophages were the main sources of properdin in the ischemic brain. Global and conditional myeloid knockout of properdin attenuated microglial overactivation and inflammatory responses at the acute stage of tMCAO in mice. Accordingly, treatment with recombinant properdin enhanced the production of proinflammatory cytokines and augmented microglia-potentiated neuronal death in primary culture. Mechanistically, recombinant properdin served as a novel ligand that activated Mincle receptors on microglia and downstream pathways to drive primary microglia-induced inflammatory responses. Intriguingly, properdin can directly bind to the microglial Mincle receptor to exert the above effects, while Mincle knockdown limits properdin-mediated microglial inflammation. CONCLUSION: Properdin is a new medium by which infiltrating peripheral myeloid cells communicate with microglia, further activate microglia, and exacerbate brain injury in the ischemic brain, suggesting that targeted disruption of the interaction between properdin and Mincle on microglia or inhibition of their downstream signaling may improve the prognosis of ischemic stroke.


Assuntos
Lesões Encefálicas , Isquemia Encefálica , AVC Isquêmico , Camundongos , Animais , Microglia/metabolismo , AVC Isquêmico/metabolismo , Properdina/metabolismo , Properdina/farmacologia , Doenças Neuroinflamatórias , Macrófagos/metabolismo , Infarto da Artéria Cerebral Média/patologia , Lesões Encefálicas/metabolismo , Isquemia Encefálica/metabolismo , Camundongos Endogâmicos C57BL
6.
Front Immunol ; 14: 1174243, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37662914

RESUMO

Hepatic ischemia/reperfusion injury (IRI) often causes serious complications in liver surgeries, including transplantation. Complement activation seems to be involved in hepatic IRI; however, no complement-targeted intervention has been clinically applied. We investigated the therapeutic potential of Properdin-targeted complement regulation in hepatic IRI. Male wild-type mice (B10D2/nSn) were exposed to 90-minute partial hepatic IRI to the left and median lobes with either monoclonal anti-Properdin-antibody (Ab) or control-immunoglobulin (IgG) administration. Since the complement system is closely involved in liver regeneration, the influence of anti-Properdin-Ab on liver regeneration was also evaluated in a mouse model of 70% partial hepatectomy. Anti-Properdin-Ab significantly reduced serum transaminases and histopathological damages at 2 and 6 hours after reperfusion (P <0.001, respectively). These improvements at 2 hours was accompanied by significant reductions in CD41+ platelet aggregation (P =0.010) and ssDNA+ cells (P <0.001), indicating significant amelioration in hepatic microcirculation and apoptosis, respectively. Characteristically, F4/80+ cells representing macrophages, mainly Kupffer cells, were maintained by anti-Properdin-Ab (P <0.001). Western blot showed decreased phosphorylation of only Erk1/2 among MAPKs (P =0.004). After 6 hours of reperfusion, anti-Properdin-Ab significantly attenuated the release of HMGB-1, which provokes the release of proinflammatory cytokines/chemokines (P =0.002). Infiltration of CD11b+ and Ly6-G+ cells, representing infiltrating macrophages and neutrophils, respectively, were significantly alleviated by anti-Properdin-Ab (both P <0.001). Notably, anti-Properdin-Ab did not affect remnant liver weight and BrdU+ cells at 48 hours after 70% partial hepatectomy (P =0.13 and 0.31, respectively). In conclusion, Properdin inhibition significantly ameliorates hepatic IRI without interfering with liver regeneration.


Assuntos
Properdina , Traumatismo por Reperfusão , Masculino , Animais , Camundongos , Regeneração Hepática , Fígado , Traumatismo por Reperfusão/prevenção & controle , Isquemia
7.
Front Immunol ; 14: 1183768, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37207230

RESUMO

Phagocytosis plays vital roles in injury and repair, while its regulation by properdin and innate repair receptor, a heterodimer receptor of erythropoietin receptor (EPOR)/ß common receptor (ßcR), in renal ischaemia-reperfusion (IR) remains unclear. Properdin, a pattern recognition molecule, facilitates phagocytosis by opsonizing damaged cells. Our previous study showed that the phagocytic function of tubular epithelial cells isolated from properdin knockout (PKO) mouse kidneys was compromised, with upregulated EPOR in IR kidneys that was further raised by PKO at repair phase. Here, helix B surface peptide (HBSP), derived from EPO only recognizing EPOR/ßcR, ameliorated IR-induced functional and structural damage in both PKO and wild-type (WT) mice. In particular, HBSP treatment led to less cell apoptosis and F4/80+ macrophage infiltration in the interstitium of PKO IR kidneys compared to the WT control. In addition, the expression of EPOR/ßcR was increased by IR in WT kidneys, and furthered increased in IR PKO kidneys, but greatly reduced by HBSP in the IR kidneys of PKO mice. HBSP also increased PCNA expression in IR kidneys of both genotypes. Moreover, iridium-labelled HBSP (HBSP-Ir) was localized mainly in the tubular epithelia after 17-h renal IR in WT mice. HBSP-Ir also anchored to mouse kidney epithelial (TCMK-1) cells treated by H2O2. Both EPOR and EPOR/ßcR were significantly increased by H2O2 treatment, while further increased EPOR was showed in cells transfected with small interfering RNA (siRNA) targeting properdin, but a lower level of EPOR was seen in EPOR siRNA and HBSP-treated cells. The number of early apoptotic cells was increased by EPOR siRNA in H2O2-treated TCMK-1, but markedly reversed by HBSP. The phagocytic function of TCMK-1 cells assessed by uptake fluorescence-labelled E.coli was enhanced by HBSP dose-dependently. Our data demonstrate for the first time that HBSP improves the phagocytic function of tubular epithelial cells and kidney repair post IR injury, via upregulated EPOR/ßcR triggered by both IR and properdin deficiency.


Assuntos
Properdina , Traumatismo por Reperfusão , Camundongos , Animais , Properdina/genética , Peróxido de Hidrogênio , Rim , Traumatismo por Reperfusão/genética , Isquemia , Células Epiteliais , Fagocitose/genética , RNA Interferente Pequeno
8.
Pregnancy Hypertens ; 32: 43-49, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37088032

RESUMO

OBJECTIVES: Pre-eclampsia (PE) is a leading cause of obstetric morbidity, with no definitive therapy other than delivery. We aimed to compare complement markers in maternal and fetal circulation, and placental tissue, between women with PE and healthy pregnant controls. STUDY DESIGN: Maternal and umbilical cord blood was tested for iC3b, C3, C4, properdin, Ba and C5b-9, and placental tissue for C3d, C4d, C9 and C1q, from women with PE (n = 34) and healthy pregnant controls (n = 33). Maternal properdin and Ba tests were repeated in a separate validation cohort (PE n = 35; healthy pregnant controls n = 35). MAIN OUTCOME MEASURES: Complement concentrations in maternal and umbilical cord blood, and placental immunohistochemical complement deposition. RESULTS: Women with PE had significantly lower concentrations of properdin (mean: 4828 vs 6877 ng/ml, p < 0.001) and C4 (mean: 0.20 vs 0.31 g/l, p < 0.001), and higher Ba (median: 150 vs 113 ng/ml, p = 0.012), compared to controls. After controlling for gestational age at blood draw, average properdin concentration was 1945 ng/ml lower in PE vs controls (95 % CI: 1487-2402, p < 0.001). Of the cord blood markers assessed, only Ba differed significantly between PE and controls (median: 337 vs 233 ng/ml, p = 0.004). C4d staining of the syncytiotrophoblast membrane was increased in PE vs controls (median immunoreactivity score 3 vs 0, p < 0.001). Maternal properdin and C4 were significantly negatively correlated with placental C4d staining. CONCLUSIONS: Our data confirm excessive placental complement deposition associated with significant concurrent changes in maternal and fetal circulating complement biomarkers in PE. Inhibition of complement activation is a potential therapeutic target.


Assuntos
Placenta , Pré-Eclâmpsia , Gravidez , Feminino , Humanos , Placenta/metabolismo , Properdina/metabolismo , Ativação do Complemento , Trofoblastos/metabolismo
9.
Artigo em Inglês | MEDLINE | ID: mdl-36853837

RESUMO

Complement is a major innate defense system that protects the intravascular space from microbial invasion. Complement activation results in the assembly of C3 convertases, serine proteases that cleave complement protein C3, generating bioactive fragments C3a and C3b. The complement response is rapid and robust, largely due to a positive feedback regulatory loop mediated by alternative pathway (AP) C3 convertase. C3 nephritic factors (C3NEFs) are autoantibodies that stabilize AP convertase, resulting in uncontrolled C3 cleavage, which, in principle, can promote critical tissue injury similar to that seen in certain renal conditions. Investigations of C3NEFs are hampered by a challenging issue: each C3NEF is derived from a different donor source, and there is no method to compare one C3NEF to another. We have identified a widely available mouse anti-C3 mAb that, similar to many C3NEFs, can stabilize functional AP convertase in a form resistant to decay acceleration by multiple complement regulators. The antibody requires the presence of properdin to confer convertase stability, and hampers the activity of Salp20, a tic salivary protein that accelerates convertase dissociation by displacing properdin from the convertase complex. This mAb can serve as an urgently needed standard for the investigation of C3NEFs. This study also provides novel insights into the dynamics of AP convertase.


Assuntos
Anticorpos Monoclonais , Fator Nefrítico do Complemento 3 , Animais , Camundongos , Properdina , Autoanticorpos
10.
Clin Exp Immunol ; 212(1): 52-60, 2023 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-36722378

RESUMO

Inflammation plays a fundamental role in the development of several metabolic diseases, including obesity and type 2 diabetes (T2D); the complement system has been implicated in their development. People of Black African (BA) ethnicity are disproportionately affected by T2D and other metabolic diseases but the impact of ethnicity on the complement system has not been explored. We investigated ethnic differences in complement biomarkers and activation status between men of BA and White European (WE) ethnicity and explored their association with parameters of metabolic health. We measured a panel of 15 complement components, regulators, and activation products in fasting plasma from 89 BA and 96 WE men. Ethnic differences were statistically validated. Association of complement biomarkers with metabolic health indices (BMI, waist circumference, insulin resistance, and HbA1c) were assessed in the groups. Plasma levels of the key complement components C3 and C4, the regulators clusterin and properdin and the activation marker iC3b were significantly higher in BA compared to WE men after age adjustment, while FD levels were significantly lower. C3 and C4 levels positively correlated with some or all markers of metabolic dysfunction in both ethnic groups while FD was inversely associated with HbA1c in both groups, and clusterin and properdin were inversely associated with some markers of metabolic dysfunction only in the WE group. Our findings of increased levels of complement components and activation products in BA compared to WE men suggest differences in complement regulation that may impact susceptibility to poor metabolic health.


Assuntos
Clusterina , Resistência à Insulina , Doenças Metabólicas , Properdina , Humanos , Masculino , Biomarcadores , Diabetes Mellitus Tipo 2 , Etnicidade , Hemoglobinas Glicadas , População Branca , População Negra , Doenças Metabólicas/etnologia , Complemento C4 , Complemento C3
11.
Psychol Med ; 53(13): 6102-6112, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-36285542

RESUMO

BACKGROUND: Inflammation plays a crucial role in the pathogenesis of major depressive disorder (MDD) and bipolar disorder (BD). This study aimed to examine whether the dysregulation of complement components contributes to brain structural defects in patients with mood disorders. METHODS: A total of 52 BD patients, 35 MDD patients, and 53 controls were recruited. The human complement immunology assay was used to measure the levels of complement factors. Whole brain-based analysis was performed to investigate differences in gray matter volume (GMV) and cortical thickness (CT) among the BD, MDD, and control groups, and relationships were explored between neuroanatomical differences and levels of complement components. RESULTS: GMV in the medial orbital frontal cortex (mOFC) and middle cingulum was lower in both patient groups than in controls, while the CT of the left precentral gyrus and left superior frontal gyrus were affected differently in the two disorders. Concentrations of C1q, C4, factor B, factor H, and properdin were higher in both patient groups than in controls, while concentrations of C3, C4 and factor H were significantly higher in BD than in MDD. Concentrations of C1q, factor H, and properdin showed a significant negative correlation with GMV in the mOFC at the voxel-wise level. CONCLUSIONS: BD and MDD are associated with shared and different alterations in levels of complement factors and structural impairment in the brain. Structural defects in mOFC may be associated with elevated levels of certain complement factors, providing insight into the shared neuro-inflammatory pathogenesis of mood disorders.


Assuntos
Transtorno Bipolar , Transtorno Depressivo Maior , Córtex Motor , Humanos , Transtorno Depressivo Maior/diagnóstico por imagem , Transtorno Depressivo Maior/patologia , Transtorno Bipolar/diagnóstico por imagem , Transtorno Bipolar/patologia , Fator H do Complemento , Properdina , Complemento C1q , Imageamento por Ressonância Magnética , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Substância Cinzenta/diagnóstico por imagem , Substância Cinzenta/patologia
12.
Immunol Rev ; 313(1): 46-59, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36097870

RESUMO

Structures of alternative pathway proteins have offered a comprehensive structural basis for understanding the molecular mechanisms governing activation and regulation of the amplification pathway of the complement cascade. Although properdin (FP) is required in vivo to sustain a functional alternative pathway, structural studies have been lagging behind due to the extended structure and polydisperse nature of FP. We review recent progress with respect to structure determination of FP and its proconvertase/convertase complexes. These structures identify in detail regions in C3b, factor B and FP involved in their mutual interactions. Structures of FP oligomers obtained by integrative studies have shed light on how FP activity depends on its oligomerization state. The accumulated structural knowledge allows us to rationalize the effect of point mutations causing FP deficiency. The structural basis for FP inhibition by the tick CirpA proteins is reviewed and the potential of alphafold2 predictions for understanding the interaction of FP with other tick proteins and the NKp46 receptor on host immune cells is discussed. The accumulated structural knowledge forms a comprehensive basis for understanding molecular interactions involving FP, pathological conditions arising from low levels of FP, and the molecular strategies used by ticks to suppress the alternative pathway.


Assuntos
Ativação do Complemento , Properdina , Humanos , Properdina/genética , Properdina/metabolismo , Via Alternativa do Complemento
13.
Immunol Rev ; 313(1): 162-180, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36336911

RESUMO

The alternative pathway (AP) is the phylogenetically oldest arm of the complement system and may have evolved to mark pathogens for elimination by phagocytes. Studies using purified AP proteins or AP-specific serum showed that C3b amplification on bacteria commenced following a lag phase of about 5 min and was highly dependent on the concentration of complement. Most pathogens have evolved several elegant mechanisms to evade complement, including expressing proteases that degrade AP proteins and secreting proteins that block function of C3 convertases. In an example of convergent evolution, many microbes recruit the AP inhibitor factor H (FH) using molecular mechanisms that mimic FH interactions with host cells. In most instances, the AP serves to amplify C3b deposited on microbes by the classical pathway (CP). The role of properdin on microbes appears to be restricted to stabilization of C3 convertases; scant evidence exists for its role as an initiator of the AP on pathogens in the context of serum. Therapeutic complement inhibition carries with it an increased risk of infection. Antibody (Ab)-dependent AP activation may be critical for complement activation by vaccine-elicited Ab when the CP is blocked, and its molecular mechanism is discussed.


Assuntos
Infecções Bacterianas , Ativação do Complemento , Via Alternativa do Complemento , Humanos , Ativação do Complemento/fisiologia , Properdina/metabolismo , Infecções Bacterianas/metabolismo , Complemento C3b/metabolismo
14.
Immunol Rev ; 313(1): 194-216, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36203396

RESUMO

This review aimed to capture the key findings that animal models have provided around the role of the alternative pathway and amplification loop (AP/AL) in disease. Animal models, particularly mouse models, have been incredibly useful to define the role of complement and the alternative pathway in health and disease; for instance, the use of cobra venom factor and depletion of C3 provided the initial insight that complement was essential to generate an appropriate adaptive immune response. The development of knockout mice have further underlined the importance of the AP/AL in disease, with the FH knockout mouse paving the way for the first anti-complement drugs. The impact from the development of FB, properdin, and C3 knockout mice closely follows this in terms of mechanistic understanding in disease. Indeed, our current understanding that complement plays a role in most conditions at one level or another is rooted in many of these in vivo studies. That C3, in particular, has roles beyond the obvious in innate and adaptive immunity, normal physiology, and cellular functions, with or without other recognized AP components, we would argue, only extends the reach of this arm of the complement system. Humanized mouse models also continue to play their part. Here, we argue that the animal models developed over the last few decades have truly helped define the role of the AP/AL in disease.


Assuntos
Via Alternativa do Complemento , Properdina , Animais , Camundongos , Humanos , Properdina/metabolismo , Imunidade Adaptativa , Camundongos Knockout , Modelos Animais de Doenças
15.
Front Immunol ; 13: 953674, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36211440

RESUMO

Hidradenitis suppurativa (HS) is a chronic auto-inflammatory skin disease with a complex and multifactorial pathogenesis involving both the innate and adaptive immune system. Despite limited evidence for local complement activation, conflicting results have been published on the role of systemic complement activation in HS. It was hypothesized that complement was consumed in highly inflamed HS skin, trapping complement from the circulation. Therefore, the aim of this study was to evaluate this local complement deposition in HS skin lesions using routine and commonly used complement antibodies.Direct immunofluorescence for C1q, C3c, C4d, C5b-9, and properdin was performed on frozen tissue sections of 19 HS patients and 6 controls. C5a receptor 1 (C5aR1) was visualized using immunohistochemistry. Overall, we found no significant local complement deposition in HS patients versus controls regarding C1q, C3c, C4d, C5b-9, or properdin on either vessels or immune cells. C5aR1 expression was exclusively found on immune cells, predominantly neutrophilic granulocytes, but not significantly different relatively to the total infiltrate in HS lesions compared with controls. In conclusion, despite not being able to confirm local complement depositions of C1q, C3c, C4d, or properdin using highly sensitive and widely accepted techniques, the increased presence of C5aR1 positive immune cells in HS suggests the importance of complement in the pathogenesis of HS and supports emerging therapies targeting this pathway.


Assuntos
Hidradenite Supurativa , Ativação do Complemento , Complemento C1q , Complexo de Ataque à Membrana do Sistema Complemento , Humanos , Inflamação , Properdina , Receptor da Anafilatoxina C5a
16.
Int J Mol Sci ; 23(20)2022 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-36293087

RESUMO

Complement pathway proteins are reported to be increased in polycystic ovary syndrome (PCOS) and may be affected by obesity and insulin resistance. To investigate this, a proteomic analysis of the complement system was undertaken, including inhibitory proteins. In this cohort study, plasma was collected from 234 women (137 with PCOS and 97 controls). SOMALogic proteomic analysis was undertaken for the following complement system proteins: C1q, C1r, C2, C3, C3a, iC3b, C3b, C3d, C3adesArg, C4, C4a, C4b, C5, C5a, C5b-6 complex, C8, properdin, factor B, factor D, factor H, factor I, mannose-binding protein C (MBL), complement decay-accelerating factor (DAF) and complement factor H-related protein 5 (CFHR5). The alternative pathway of the complement system was primarily overexpressed in PCOS, with increased C3 (p < 0.05), properdin and factor B (p < 0.01). In addition, inhibition of this pathway was also seen in PCOS, with an increase in CFHR5, factor H and factor I (p < 0.01). Downstream complement factors iC3b and C3d, associated with an enhanced B cell response, and C5a, associated with an inflammatory cytokine release, were increased (p < 0.01). Hyperandrogenemia correlated positively with properdin and iC3b, whilst insulin resistance (HOMA-IR) correlated with iC3b and factor H (p < 0.05) in PCOS. BMI correlated positively with C3d, factor B, factor D, factor I, CFHR5 and C5a (p < 0.05). This comprehensive evaluation of the complement system in PCOS revealed the upregulation of components of the complement system, which appears to be offset by the concurrent upregulation of its inhibitors, with these changes accounted for in part by BMI, hyperandrogenemia and insulin resistance.


Assuntos
Resistência à Insulina , Lectina de Ligação a Manose , Síndrome do Ovário Policístico , Feminino , Humanos , Properdina/metabolismo , Fator H do Complemento , Fator B do Complemento/metabolismo , Antígenos CD55 , Fator D do Complemento , Estudos de Coortes , Proteômica , Complemento C1q , Complemento C3b , Fibrinogênio , Citocinas
17.
Protein Sci ; 31(10): e4432, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36173177

RESUMO

Structure determination of macromolecular complexes is challenging if subunits can dissociate during crystallization or preparation of electron microscopy grids. We present an approach where a labile complex is stabilized by linking subunits though introduction of a peptide tag in one subunit that is recognized by a nanobody tethered to a second subunit. This allowed crystal structure determination at 3.9 Å resolution of the highly non-globular 320 kDa proconvertase formed by complement components C3b, factor B, and properdin. Whereas the binding mode of properdin to C3b is preserved, an internal rearrangement occurs in the zymogen factor B von Willebrand domain type A domain compared to the proconvertase not bound to properdin. The structure emphasizes the role of two noncanonical loops in thrombospondin repeats 5 and 6 of properdin in augmenting the activity of the C3 convertase. We suggest that linking of subunits through peptide specific tethered nanobodies represents a simple alternative to approaches like affinity maturation and chemical cross-linking for the stabilization of large macromolecular complexes. Besides applications for structural biology, nanobody bridging may become a new tool for biochemical analysis of unstable macromolecular complexes and in vitro selection of highly specific binders for such complexes.


Assuntos
Properdina , Anticorpos de Domínio Único , Convertases de Complemento C3-C5/química , Convertases de Complemento C3-C5/metabolismo , Fator B do Complemento/química , Fator B do Complemento/metabolismo , Precursores Enzimáticos , Substâncias Macromoleculares , Properdina/química , Properdina/metabolismo , Trombospondinas
18.
Front Immunol ; 13: 910993, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36091001

RESUMO

Objectives: The goal of this exploratory study is to determine if urine:serum fractional excretion ratios can outperform the corresponding urinary biomarker proteins in identifying active renal disease in systemic lupus erythematosus (SLE). Methods: Thirty-six adult SLE patients and twelve healthy controls were examined for serum and urine levels of 8 protein markers, namely ALCAM, calpastatin, hemopexin, peroxiredoxin 6 (PRDX6), platelet factor 4 (PF4), properdin, TFPI and VCAM-1, by ELISA. Fractional excretion of analyzed biomarkers was calculated after normalizing both the urine and serum biomarker levels against creatinine. A further validation cohort of fifty SLE patients was included to validate the initial findings. Results: The FE ratios of all 8 proteins interrogated outperformed conventional disease activity markers such as anti-dsDNA, C3 and C4 in identifying renal disease activity. All but VCAM-1FE were superior to the corresponding urine biomarkers levels in differentiating LN activity, exhibiting positive correlation with renal SLEDAI. ALCAMFE, PF4FE and properdinFE ratios exhibited the highest accuracy (AUC>0.9) in distinguishing active LN from inactive SLE. Four of the FE ratios exhibited perfect sensitivity (calpastatin, PRDX6, PF4 and properdin), while ALCAMFE, PF4FE and properdinFE exhibited the highest specificity values for active LN. In addition, several of these novel biomarkers were associated with higher renal pathology activity indices. In the validation cohort ALCAMFE, PF4FE and properdinFE once again exhibited higher accuracy metrics, surpassing corresponding urine and serum biomarkers levels, with ALCAMFE exhibiting 95% accuracy in distinguishing active LN from inactive SLE. Conclusions: With most of the tested proteins, urine:serum fractional excretion ratios outperformed corresponding urine and serum protein measurements in identifying active renal involvement in SLE. Hence, this novel class of biomarkers in SLE ought to be systemically evaluated in larger independent cohorts for their diagnostic utility in LN assessment.


Assuntos
Lúpus Eritematoso Sistêmico , Nefrite Lúpica , Molécula de Adesão de Leucócito Ativado , Adulto , Biomarcadores , Humanos , Lúpus Eritematoso Sistêmico/diagnóstico , Nefrite Lúpica/diagnóstico , Fator Plaquetário 4 , Properdina , Molécula 1 de Adesão de Célula Vascular
19.
Front Endocrinol (Lausanne) ; 13: 918320, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35909516

RESUMO

Introduction: Complement factors mediate the recruitment and activation of immune cells and are associated with metabolic changes during pregnancy. The aim of this study was to determine whether complement factors in the maternal serum and follicular fluid (FF) are associated with in vitro fertilization (IVF) outcomes in overweight/obese women. Methods: Forty overweight/obese (BMI = 30.8 ± 5.2 kg/m2) female patients, 33.6 ± 6.3 years old, undergoing IVF treatment for unexplained infertility were recruited. Baseline demographic information, including biochemical hormonal, metabolic, and inflammatory markers, and pregnancy outcome, was collected. Levels of 14 complement markers (C2, C4b, C5, C5a, C9, adipsin, mannose-binding lectin, C1q, C3, C3b/iC3b, C4, factor B, factor H, and properdin) were assessed in the serum and FF and compared to IVF outcome, inflammatory, and metabolic markers using multivariate and univariate models. Results: Out of 40 IVF cycles, 14 (35%) resulted in pregnancy. Compared to women with failed pregnancies, women with successful pregnancies had higher levels of adipsin in the serum and FF (p = 0.01) but lower C5a levels (p = 0.05). Serum adipsin levels were positively correlated with circulating levels of vitamin D (R = 0.5, p = 0.02), glucagon (R = 0.4, p = 0.03), leptin (R = 0.4, p = 0.01), resistin (R = 0.4, p = 0.02), and visfatin (R = 0.4, p = 0.02), but negatively correlated with total protein (R = -0.5, p = 0.03). Higher numbers of top-quality embryos were associated with increased levels of C3, properdin, C1q, factors H and B, C4, and adipsin, but with reduced C2 and C5a levels (p ≤ 0.01). Conclusions: Higher adipsin and lower C5a levels in the maternal serum during implantation are potential markers of successful outcome in obese women undergoing IVF-assisted pregnancies.


Assuntos
Complemento C5a , Fator D do Complemento , Líquido Folicular , Adulto , Biomarcadores/metabolismo , Complemento C5a/metabolismo , Fator D do Complemento/metabolismo , Feminino , Líquido Folicular/metabolismo , Humanos , Obesidade/metabolismo , Sobrepeso/metabolismo , Gravidez , Resultado da Gravidez , Properdina/metabolismo
20.
Immunobiology ; 227(4): 152246, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35843030

RESUMO

The complement system does not only play an important role in the defence against microorganism and pathogens, but also contributes to the regulation of innate and adaptive immunity. Especially activation fragments C3a and C5a and complement activation at the interface of antigen presenting cell (APC) and T cell, were shown to have a role in T cell activation and proliferation. Whereas most complement factors are produced by the liver, properdin, a positive regulator of the C3 convertase, is mainly produced by myeloid cells. Here we show that properdin can be detected in myeloid cell infiltrate during human renal allograft rejection. In vitro, properdin is produced and secreted by human immature dendritic cells (iDCs), which is further increased by CD40-L-matured DCs (mDCs). Transfection with a specific properdin siRNA reduced properdin secretion by iDCs and mDCs, without affecting the expression of co-stimulatory markers CD80 and CD86. Co-culture of properdin siRNA-transfected iDCs and mDCs with human allogeneic T cells resulted in reduced T cell proliferation, especially under lower DC-T cell ratio's (1:30 and 1:90 ratio). In addition, T cell cytokines were altered, including a reduced TNF-α and IL-17 secretion by T cells co-cultured with properdin siRNA-transfected iDCs. Taken together, these results indicate a local role for properdin during the interaction of DCs and allogeneic T cells, contributing to the shaping of T cell proliferation and activation.


Assuntos
Transplante de Rim , Properdina , Células Cultivadas , Células Dendríticas , Humanos , Properdina/genética , Properdina/metabolismo , RNA Interferente Pequeno , Linfócitos T
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