RESUMO
Complement protein C3dg, a key linkage between innate and adaptive immunity, is capable of stimulating both humoral and cell-mediated immune responses, leading to considerable interest in its use as a molecular adjuvant. However, the potential of C3dg as an adjuvant is limited without ways of controllably assembling multiple copies of it into vaccine platforms. Here, we report a strategy to assemble C3dg into supramolecular nanofibers with excellent compositional control, using ß-tail fusion tags. These assemblies were investigated as therapeutic active immunotherapies, which may offer advantages over existing biologics, particularly toward chronic inflammatory diseases. Supramolecular assemblies based on the Q11 peptide system containing ß-tail-tagged C3dg, B cell epitopes from TNF, and the universal T cell epitope PADRE raised strong antibody responses against both TNF and C3dg, and prophylactic immunization with these materials significantly improved protection in a lethal TNF-mediated inflammation model. Additionally, in a murine model of psoriasis induced by imiquimod, the C3dg-adjuvanted nanofiber vaccine performed as well as anti-TNF monoclonal antibodies. Nanofibers containing only ß-tail-C3dg and lacking the TNF B cell epitope also showed improvements in both models, suggesting that supramolecular C3dg, by itself, played an important therapeutic role. We observed that immunization with ß-tail-C3dg caused the expansion of an autoreactive C3dg-specific T cell population, which may act to dampen the immune response, preventing excessive inflammation. These findings indicate that molecular assemblies displaying C3dg warrant further development as active immunotherapies.
Assuntos
Complemento C3d/imunologia , Nanofibras/química , Psoríase/prevenção & controle , Vacinas/imunologia , Adjuvantes Imunológicos/química , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Linfócitos B/imunologia , Células Cultivadas , Epitopos/química , Epitopos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/imunologia , Vacinas/químicaRESUMO
De novo donor-specific antibodies (dnDSA) are associated with antibody-mediated rejection (ABMR) and allograft loss. We tested Immucor* (IM) Luminex Single-antigen beads (LSAB) assay and C3d-fixing antibodies in the setting of dnDSA and subclinical (s) ABMR. This retrospective multicentric study included 123 patients biopsied because of the presence of subclinical de novo DSA detected by One Lamda* Labscreen (MFI > 1000). In 112 patients, sera of the day of the biopsy were available and tested in a central lab with IM Lifecodes LSAB and C3d fixing antibodies assays. In 16 patients (14.3%), no DSA was detected using Immucor test. In 96 patients, at least one DSA was determined with IM. Systematic biopsies showed active sABMR in 30 patients (31.2%), chronic active sABMR in 17 patients (17.7%) and no lesions of sABMR in 49 KT recipients (51%). Intensitity criteria (BCM, BCR and AD-BCR) of DSA were not statistically different between these 3 histological groups. The proportion of patients with C3d-fixing DSA was not statistically different between the 3 groups and did not offer any prognostic value regarding graft survival. Performing biopsy for dnDSA could not be guided by the intensity criteria of IM LSAB assay. C3d-fixing DSA do not offer added value.
Assuntos
Complemento C3d/imunologia , Rejeição de Enxerto/diagnóstico , Isoanticorpos/sangue , Transplante de Rim/efeitos adversos , Adulto , Biomarcadores/sangue , Feminino , França , Rejeição de Enxerto/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos RetrospectivosRESUMO
BACKGROUND: Donor-specific antibodies (DSA) play a major role in antibody-mediated rejection (AMR) and graft dysfunction. However, the clinical relevance of complement-binding anti-HLA antibodies remains unclear. METHODS: Here, we analyzed DSA detected in the serum (sDSA) using single antigen bead, C1q, and C3d assays combined with the study of intragraft DSA (gDSA) in 86 patients who had DSA and underwent a kidney biopsy for cause (n = 58) or without evidence of kidney dysfunction (n = 28). DSA characteristics were collected and related to the presence of AMR, graft histological features, and allograft survival. RESULTS: Forty-five patients (52%) had C1q DSA, and 42 (51%) had C3d DSA. Allograft biopsies revealed AMR in 63 cases (73%), regardless of kidney function. gDSA were identified in 74% of biopsies. We observed a strong correlation among single antigen bead mean fluorescence intensity and complement assays positivity, presence of gDSA, and AMR occurrence. CONCLUSIONS: Complement-binding DSA per se were not significantly associated with allograft survival in the entire study sample. Finally, gDSA predicted subsequent graft loss in patients who showed a stable renal function at the day of biopsy. Our data suggest that DSA mean fluorescence intensity and presence of gDSA might provide prognostic information during posttransplant monitoring.
Assuntos
Testes de Fixação de Complemento , Rejeição de Enxerto/diagnóstico , Antígenos HLA/imunologia , Isoanticorpos/sangue , Transplante de Rim/efeitos adversos , Monitorização Imunológica , Adolescente , Adulto , Idoso , Biomarcadores/sangue , Biópsia , Criança , Pré-Escolar , Complemento C1q/imunologia , Complemento C3d/imunologia , Feminino , Rejeição de Enxerto/sangue , Rejeição de Enxerto/imunologia , Sobrevivência de Enxerto , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Estudos Retrospectivos , Fatores de Tempo , Resultado do Tratamento , Adulto JovemRESUMO
BACKGROUND: Porcine circovirus type 2 (PCV2) is an economically important viral pathogen for swine industry worldwide. However, current PCV2 vaccines provide incomplete protection against the PCV2d, which has recently emerged as the predominant pathogenic form of PCV2. METHODS: To develop a novel DNA vaccine with high efficacy against PCV2d virus, we fused the ORF2 of PCV2d to three copies of the minimum-binding domain of the complement C3 cascade terminal component, C3d-P28. Expression of ORF2 alone (pVO) or fused C3d-P28 (pVOC3) were verified by immunofluorescent assay. Vaccine efficacy was tested by measured the DNA copy and T and B cell immune response. RESULTS: Vaccination with pVOC3 reduced the levels of PCV2 genomic DNA after pigs were infected with either PCV2b or PCV2d genotypes, produced potent antibodies against PCV2, and stimulated PCV2-specific interferon-γ secreting cells. CONCLUSION: Results suggested pVOC3 would be a safe and effective DNA vaccine to confer cross-protection against both PCV2b and PCV2d genotypes in pigs.
Assuntos
Proteínas do Capsídeo/imunologia , Infecções por Circoviridae/veterinária , Circovirus/genética , Complemento C3d/imunologia , Doenças dos Suínos/prevenção & controle , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Proteínas do Capsídeo/genética , Infecções por Circoviridae/prevenção & controle , Circovirus/imunologia , Complemento C3d/genética , Proteção Cruzada , DNA Viral/genética , Genoma Viral , Genótipo , Masculino , Suínos , Doenças dos Suínos/virologia , Vacinação , Vacinas de DNA/genética , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologia , Vacinas Virais/genéticaRESUMO
BACKGROUND: One risk factor for antibody-mediated rejection (ABMR) and poor outcome after kidney transplantation is donor-specific antiâhuman leukocyte antigen (anti-HLA) antibodies (DSAs). In this study we sought to determine whether the presence of DSAs that bind complement component C3d could better predict ABMR and graft loss in stable kidney transplant recipients (KTRs). METHODS: We included 220 stable KTRs in this study and screened them for DSAs from July 2013 to July 2016. RESULTS: Of the 220 KTRs, DSAs were detected in 24 (10.9%). The incidence of ABMR was 3.6% (8 of 220) overall, and C3d-DSAâpositive KTRs had a significantly higher incidence than SA-DSAâpositive KTRs (63.3% vs 38.9%, P = .03). Most C3d-binding DSAs were anti-HLA class II antibodies (11 of 13, 84.6%). Class II C3d-binding DSA was also significantly associated with graft failure on multivariate analysis, as were ABMR, chronic ABMR, and high serum creatinine. Class II C3d-binding DSA was also significantly associated with lower graft survival after ABMR. CONCLUSION: C3d-binding DSA, especially class II, was significantly associated with the risk of ABMR and graft loss in stable KTRs. We suggest that monitoring of stable KTRs for C3d-binding DSA, followed by biopsy, could aid in early recognition of ABMR and prevention of graft loss.
Assuntos
Complemento C3d/imunologia , Rejeição de Enxerto/imunologia , Antígenos HLA/imunologia , Transplante de Rim , Anticorpos/imunologia , Estudos de Coortes , Feminino , Sobrevivência de Enxerto/imunologia , Humanos , Masculino , Estudos Retrospectivos , Fatores de Risco , Doadores de Tecidos , TransplantadosRESUMO
INTRODUCTION: Complement binding activity of donor-specific HLA antibodies (DSA) has been suggested as a new tool to stratify immunologic risk in kidney transplantation (KT). The objective of this study was to evaluate the clinical implication of C1q/C3d binding activity of de novo DSA (dnDSA) in KT recipients. MATERIAL AND METHODS: A total of 161 pretransplant DSA-negative recipients were monitored for dnDSA at the time of biopsy. C1q/C3d binding activities of dnDSA were assessed using C1qScreen assay (One lambda, USA) and Lifecodes C3d detection assay (Immucor, USA), respectively. Clinical outcomes including biopsy-proven antibody mediated rejection (AMR), C4d detection and post-biopsy graft survival were investigated. RESULTS: De-novo DSAs were detected in fifty-four (33.5%) patients (HLA class I only, n = 19; class II only, n = 29; both class I and II, n = 6). Of them, complement binding activities were detected in 26 (48.1%) patients, including 17 C1q+ and 24 C3d+ patients. Both C1q and C3d positivity were associated with increased mean fluorescence intensity values of dnDSA. Complement binding activity of dnDSA enhanced the incidence of AMR (25.0% in C1q-C3d-, 36.4% in C1q+/C3d- or C1q-/C3d+, and 60.0% in C1q+/C3d+ patients) (P <0.001). The incidence of AMR was not different between patients with C1q+ and those with C3d+ dnDSA (64.7%, 11/17 versus 45.8%, 11/24, P = 0.238). In comparison between C1q and C3d assay according to HLA specificity, C1q+ HLA class I ± II dnDSA was the best predictor for AMR (odds ratio: 27.2). C1q+/C3d+ dnDSA was associated with more C4d deposition in allograft tissue and inferior post-biopsy graft survival. Clinical outcomes were not significantly different between C1q+ and C3d+ dnDSA-positive patients. CONCLUSION: Detection of complement binding activity using both C1q and C3d assays can be a further prognostic marker for predicting AMR and allograft outcome in dnDSA+ kidney transplant patients.
Assuntos
Complemento C1q/imunologia , Complemento C3d/imunologia , Antígenos HLA/imunologia , Transplante de Rim/efeitos adversos , Adulto , Anticorpos/genética , Anticorpos/imunologia , Complemento C1q/genética , Complemento C3d/genética , Feminino , Taxa de Filtração Glomerular , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Sobrevivência de Enxerto/genética , Sobrevivência de Enxerto/imunologia , Antígenos HLA/genética , Humanos , Rim/imunologia , Rim/patologia , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Doadores de Tecidos , Transplantados , Transplante Homólogo/efeitos adversosRESUMO
The objective of this study was to evaluate the utility of a complement-dependent C3d assay to risk stratify donor-specific antibodies (DSA) in a multicenter cohort of kidney recipients presenting with new-onset clinical dysfunction. A total of 106 subjects with evidence of DSA at a mean period of 5.3 ± 5.0 years posttransplant underwent testing using C3d reagents. C3d positivity was strongly associated with both the peak and sum IgG DSA MFI, with 98.3% (n = 57/58) of strongly reactive sera (peak MFI > 10 000) eliciting a positive signal. Patients with C3d+ DSA had a higher creatinine (P = .03), more significant graft fibrosis (P = .035), and a faster rate of graft loss posttest compared to those with C3d- DSA (P = .05). Subanalysis of patients with low-moderate level DSA confirmed the inferior outcome associated with C3d positivity. Despite the prognostic value of C3d as a stand-alone test, the assay did not provide independent risk prediction after incorporation of graft fibrosis in a multivariate model (P = .94). Overall, C3d offered limited discriminatory value for strong DSA with peak IgG MFI > 10 000 and in patients where histologic data is available, but its utilization may be considered in those with low-moderate level DSA and where an allograft biopsy is not accessible.
Assuntos
Complemento C3d/imunologia , Rejeição de Enxerto/diagnóstico , Sobrevivência de Enxerto/imunologia , Isoanticorpos/efeitos adversos , Falência Renal Crônica/cirurgia , Transplante de Rim/efeitos adversos , Doadores de Tecidos , Adulto , Bioensaio , Estudos Transversais , Feminino , Seguimentos , Rejeição de Enxerto/etiologia , Antígenos HLA/imunologia , Teste de Histocompatibilidade , Humanos , Masculino , Complicações Pós-Operatórias , Prognóstico , Estudos Prospectivos , Fatores de RiscoRESUMO
Newcastle disease is a serious infectious disease in the poultry industry. The commercial vaccines can only offer limited protection and some of them are expensive and need adjuvants. At present, DNA vaccines are widely used. However, the immune responses induced by DNA vaccines are too slow and low. Here, we constructed the transfer vectors with a different number of C3d as molecular adjuvants (n = 1, 2, 4, or 6), and the vectors were cloned into the optimal eukaryotic expression plasmid (pVAXI-optiF) that expressed the F gene of Newcastle disease virus (NDV), and named pVAXI-F(o)-C3d1, pVAXI -F(o)-C3d2, pVAXI-F(o)-C3d4, and pVAXI-F(o)-C3d6, respectively. Cell transfection test indicated that pVAXI-F(o)-C3d6 showed the highest expression. In vivo immunization showed that the chickens immunized with pVAXI-F(o)-C3d6 intramuscularly induced better immune responses than the chickens immunized with the other plasmids. The protective efficacy of pVAXI-F(o)-C3d6 was 80% after challenge with the highly virulent NDV strain F48E9. The results in this study showed that C3d6 could be used as a molecular adjuvant to quickly induce an effective immune response to control NDV.
Assuntos
Adjuvantes Imunológicos , Complemento C3d/imunologia , Doença de Newcastle/imunologia , Vírus da Doença de Newcastle/imunologia , Proteínas Recombinantes de Fusão/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Proliferação de Células/efeitos dos fármacos , Galinhas/genética , Galinhas/imunologia , Galinhas/virologia , Clonagem Molecular , Complemento C3d/genética , DNA Viral , Modelos Animais de Doenças , Vetores Genéticos , Células HEK293 , Humanos , Imunização , Imunoglobulina A/sangue , Imunoglobulina G/sangue , Linfócitos/imunologia , Doença de Newcastle/genética , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/genética , Plasmídeos , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Proteínas Recombinantes de Fusão/genética , Vacinação , Vacinas de DNA/genética , Vacinas de DNA/farmacologia , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologia , Vacinas Virais/genética , Vacinas Virais/farmacologiaRESUMO
Mycobacterium leprae (M. leprae) infection causes nerve damage and the condition worsens often during and long after treatment. Clearance of bacterial antigens including lipoarabinomannan (LAM) during and after treatment in leprosy patients is slow. We previously demonstrated that M. leprae LAM damages peripheral nerves by in situ generation of the membrane attack complex (MAC). Investigating the role of complement activation in skin lesions of leprosy patients might provide insight into the dynamics of in situ immune reactivity and the destructive pathology of M. leprae. In this study, we analyzed in skin lesions of leprosy patients, whether M. leprae antigen LAM deposition correlates with the deposition of complement activation products MAC and C3d on nerves and cells in the surrounding tissue. Skin biopsies of paucibacillary (n = 7), multibacillary leprosy patients (n = 7), and patients with erythema nodosum leprosum (ENL) (n = 6) or reversal reaction (RR) (n = 4) and controls (n = 5) were analyzed. The percentage of C3d, MAC and LAM deposition was significantly higher in the skin biopsies of multibacillary compared to paucibacillary patients (p = <0.05, p = <0.001 and p = <0.001 respectively), with a significant association between LAM and C3d or MAC in the skin biopsies of leprosy patients (r = 0.9578, p< 0.0001 and r = 0.8585, p<0.0001 respectively). In skin lesions of multibacillary patients, MAC deposition was found on axons and co-localizing with LAM. In skin lesions of paucibacillary patients, we found C3d positive T-cells in and surrounding granulomas, but hardly any MAC deposition. In addition, MAC immunoreactivity was increased in both ENL and RR skin lesions compared to non-reactional leprosy patients (p = <0.01 and p = <0.01 respectively). The present findings demonstrate that complement is deposited in skin lesions of leprosy patients, suggesting that inflammation driven by complement activation might contribute to nerve damage in the lesions of these patients. This should be regarded as an important factor in M. leprae nerve damage pathology.
Assuntos
Ativação do Complemento/imunologia , Hanseníase/imunologia , Hanseníase/patologia , Dermatopatias/imunologia , Dermatopatias/patologia , Linfócitos T/imunologia , Adolescente , Adulto , Carga Bacteriana , Biomarcadores , Biópsia , Criança , Complemento C3d/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Feminino , Granuloma/imunologia , Granuloma/metabolismo , Granuloma/patologia , Humanos , Imuno-Histoquímica , Hanseníase/microbiologia , Lipopolissacarídeos , Masculino , Pessoa de Meia-Idade , Linfócitos T/metabolismo , Adulto JovemRESUMO
Ischemia-reperfusion injury (IRI) is inevitable in solid organ transplantation, due to the transplanted organ being ischemic for prolonged periods prior to transplantation followed by reperfusion. The complement molecule C3 is present in the circulation and is also synthesized by tissue parenchyma in early response to IRI and the final stable fragment of activated C3, C3d, can be detected on injured tissue for several days post-IRI. Complement activation post-IRI was monitored noninvasively by single photon emission computed tomography (SPECT) and CT using (99m) Tc-recombinant complement receptor 2 ((99m) Tc-rCR2) in murine models of cardiac transplantation following the induction of IRI and compared to (99m) Tc-rCR2 in C3(-/-) mice or with the irrelevant protein (99m) Tc-prostate-specific membrane antigen antibody fragment (PSMA). Significant uptake with (99m) Tc-rCR2 was observed as compared to C3(-/-) or (99m) Tc-PSMA. In addition, the transplanted heart to muscle ratio of (99m) Tc-rCR2 was significantly higher than (99m) Tc-PSMA or C3(-/-) . The results were confirmed by histology and autoradiography. (99m) Tc-rCR2 can be used for noninvasive detection of activated complement and in future may be used to quantify the severity of transplant damage due to complement activation postreperfusion.
Assuntos
Ativação do Complemento/imunologia , Transplante de Coração , Traumatismo por Reperfusão Miocárdica/diagnóstico por imagem , Traumatismo por Reperfusão Miocárdica/imunologia , Receptores de Complemento 3d/imunologia , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Tomografia Computadorizada por Raios X/métodos , Animais , Complemento C3d/imunologia , Feminino , Processamento de Imagem Assistida por Computador/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/imunologia , Tecnécio/administração & dosagemRESUMO
Complement fragment C3d covalently attached to antigens enhances immune responses, particularly for antigens lacking T-cell epitopes. Enhancement has been attributed to receptor cross-linking between complement receptor CR2 (CD21) and polysaccharide antigen to surface IgM on naïve B cells. Paradoxically, C3d has still been shown to increase immune responses in CD21 knockout mice, suggesting that an auxiliary activation pathway exists. In prior studies, we demonstrated the CD21-independent C3d adjuvant effect might be due to T-cell recognition of C3d T-helper epitopes processed and presented by major histocompatibility complex class II on the B-cell surface. C3d peptide sequences containing concentrated clusters of putative human C3 T-cell epitopes were identified using the epitope-mapping algorithm, EpiMatrix. These peptide sequences were synthesized and shown in vitro to bind multiple human leukocyte antigen (HLA)-DR alleles with high affinity, and induce interferon-γ responses in healthy donor peripheral blood mononuclear cells. In the present studies, we establish further correlations between HLA binding and HLA-specific lymphocyte reactions with select epitope clusters. In addition, we show that the T-cell phenotype of C3d-specific reactive T cells is CD4(+)CD45RO(+) memory T cells. Finally, mutation of a single T-cell epitope residing within the P28 peptide segment of C3d resulted in significantly diminished adjuvant activity in BALB/c mice. Collectively, these studies support the hypothesis that the paradoxical enhancement of immune responses by C3d in the absence of CD21 is due to internalization and processing of C3d into peptides that activate autoreactive CD4(+) T-helper cells in the context of HLA class II.
Assuntos
Adjuvantes Imunológicos/metabolismo , Complemento C3d/imunologia , Ativação Linfocitária , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Linfócitos T CD4-Positivos/imunologia , Simulação por Computador , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Feminino , Marcação de Genes , Antígenos de Histocompatibilidade/imunologia , Humanos , Memória Imunológica/imunologia , Interferon gama/metabolismo , Ativação Linfocitária/imunologia , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutação , Peptídeos/metabolismo , Ligação Proteica , Doadores de TecidosRESUMO
UNLABELLED: Age-related macular degeneration (AMD) is the leading cause of blindness in the Western world. AMD is a multifactorial disorder but complement-mediated inflammation at the level of the retina plays a pivotal role. Oral zinc supplementation can reduce the progression of AMD but the precise mechanism of this protective effect is as yet unclear. We investigated whether zinc supplementation directly affects the degree of complement activation in AMD and whether there is a relation between serum complement catabolism during zinc administration and the complement factor H (CFH) gene or the Age-Related Maculopathy susceptibility 2 (ARMS2) genotype. In this open-label clinical study, 72 randomly selected AMD patients in various stages of AMD received a daily supplement of 50 mg zinc sulphate and 1 mg cupric sulphate for three months. Serum complement catabolism-defined as the C3d/C3 ratio-was measured at baseline, throughout the three months of supplementation and after discontinuation of zinc administration. Additionally, downstream inhibition of complement catabolism was evaluated by measurement of anaphylatoxin C5a. Furthermore, we investigated the effect of zinc on complement activation in vitro. AMD patients with high levels of complement catabolism at baseline exhibited a steeper decline in serum complement activation (p<0.001) during the three month zinc supplementation period compared to patients with low complement levels. There was no significant association of change in complement catabolism and CFH and ARMS2 genotype. In vitro zinc sulphate directly inhibits complement catabolism in hemolytic assays and membrane attack complex (MAC) deposition on RPE cells. This study provides evidence that daily administration of 50 mg zinc sulphate can inhibit complement catabolism in AMD patients with increased complement activation. This could explain part of the mechanism by which zinc slows AMD progression. TRIAL REGISTRATION: The Netherlands National Trial Register NTR2605.
Assuntos
Ativação do Complemento/efeitos dos fármacos , Complemento C3/metabolismo , Complemento C3d/metabolismo , Suplementos Nutricionais , Degeneração Macular/dietoterapia , Sulfato de Zinco/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Células Cultivadas , Complemento C3/imunologia , Complemento C3d/imunologia , Complemento C5a/imunologia , Complemento C5a/metabolismo , Fator B do Complemento/imunologia , Fator B do Complemento/metabolismo , Fator H do Complemento/imunologia , Fator H do Complemento/metabolismo , Sulfato de Cobre/administração & dosagem , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/imunologia , Feminino , Expressão Gênica , Humanos , Degeneração Macular/sangue , Degeneração Macular/imunologia , Degeneração Macular/patologia , Masculino , Mutação , Proteínas/genética , Proteínas/imunologia , Retina/efeitos dos fármacos , Retina/imunologia , Retina/patologia , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/efeitos dos fármacos , Epitélio Pigmentado da Retina/imunologiaRESUMO
Age-related macular degeneration (AMD) is a common condition that leads to severe vision loss and dysregulation of the complement system is thought to be associated with the disease. To investigate associations of polymorphisms in AMD susceptibility genes with systemic complement activation, 2655 individuals were genotyped for 32 single nucleotide polymorphisms (SNPs) in or near 23 AMD associated risk genes. Component 3 (C3) and its catabolic fragment C3d were measured in serum and AMD staging was performed using multimodal imaging. The C3d/C3 ratio was calculated and associations with environmental factors, SNPs and various haplotypes of complement factor H (CFH) genes and complement factor B (CFB) genes were analyzed. Linear models were built to measure the influence of genetic variants on the C3d/C3 ratio. The study cohort included 1387 patients with AMD and 1268 controls. Higher C3d/C3 ratios were found for current smoker (p = 0.002), higher age (p = 1.56 × 10(-7)), AMD phenotype (p = 1.15 × 10(-11)) and the two SNPs in the C3 gene rs6795735 (p = 0.04) and rs2230199 (p = 0.04). Lower C3d/C3 ratios were found for diabetes (p = 2.87 × 10(-6)), higher body mass index (p = 1.00 × 10(-13)), the SNPs rs1410996 (p = 0.0001), rs800292 (p = 0.003), rs12144939 (p = 4.60 × 10(-6)) in CFH, rs4151667 (p = 1.01 × 10(-5)) in CFB and individual haplotypes in CFH and CFB. The linear model revealed a corrected R-square of 0.063 including age, smoking status, gender, and genetic polymorphisms explaining 6.3% of the C3d/C3 ratio. After adding the AMD status the corrected R-square was 0.067. In conclusion, none of the evaluated genetic polymorphisms showed an association with increased systemic complement activation apart from two SNPs in the C3 gene. Major genetic and non-genetic factors for AMD were not associated with systemic complement activation.
Assuntos
Complemento C3d/genética , Fator B do Complemento/genética , Fator H do Complemento/genética , Degeneração Macular/genética , Polimorfismo de Nucleotídeo Único , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Alelos , Ativação do Complemento , Complemento C3d/imunologia , Fator B do Complemento/imunologia , Fator H do Complemento/imunologia , Feminino , Frequência do Gene , Predisposição Genética para Doença , Haplótipos , Humanos , Degeneração Macular/imunologia , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Modelos Genéticos , Fatores de Risco , Fumar/fisiopatologiaRESUMO
Autoimmune haemolytic anaemia (AIHA) is a rare (prevalence 0.2 -1.0 per 100,000 population) but a potentially fatal condition. Diagnosis of AIHA is based mainly on Direct Agglutination Test (DAT/Coomb's test). AIHA is classified into warm (optimal autoantibody reactivity at 37°C) and cold (optimal autoantibody reactivity at 4°C) types. Based on the presence or absence of an underlying cause, it is divided into primary (idiopathic) and secondary (secondary to lympho proliferative disorders, autoimmune disease, drugs and non-haematological malignancies). Warm type accounts for about 70% of cases with AIHA. It can occur at any age, but mostly after 40 years. Haemolysis in warm AIHA is mediated by IgG alone or IgG with complement. Once immunoglobulins are bound, the red cells are taken up by the macrophages of the reticulo-endothelial system which has receptors for the Fc fragments of the immunoglobulins. As the red cells are haemolysed extravascularly this occurs mainly in the spleen. Intravascular haemolysis is unusual in warm AIHA, although it can occur rarely, as warm autoantibodies can fix complements. We report severe warm AIHA in an adult with evidence of intravascular haemolysis, which is an unusual presentation.
Assuntos
Anemia Hemolítica Autoimune/complicações , Hemólise/imunologia , Doenças Raras/complicações , Adulto , Anemia Hemolítica Autoimune/diagnóstico , Anemia Hemolítica Autoimune/terapia , Complemento C3d/imunologia , Feminino , Hemossiderina/urina , Humanos , Imunoglobulina G/imunologia , Doenças Raras/diagnóstico , Doenças Raras/terapiaAssuntos
Anemia Hemolítica Autoimune/induzido quimicamente , Anti-Inflamatórios não Esteroides/imunologia , Diclofenaco/imunologia , Injúria Renal Aguda/complicações , Idoso , Anemia Hemolítica Autoimune/complicações , Anemia Hemolítica Autoimune/diagnóstico , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/efeitos adversos , Autoanticorpos/sangue , Complemento C3d/imunologia , Cistite/complicações , Diclofenaco/administração & dosagem , Diclofenaco/efeitos adversos , Feminino , Glucocorticoides/uso terapêutico , Hemólise , Humanos , Imunoglobulina G/imunologia , Neutrófilos/patologia , Fagocitose , Prednisolona/uso terapêuticoRESUMO
AIM: To construct recombinant adenovirus Ad/C3d3-sVP1 and investigate the immune effects against coxsackievirus infection in mouse. METHODS: The recombinant adenovirus Ad/sVP1-C3d3 was constructed and packaged. BALB/c mouse were divided into four groups: Ad/sVP1-C3d3 group, Ad/VP1 group, Ad group and PBS group. The mice in each group were immunized by intramuscular injection. The titers of sera IgG and neutralizing antibody were detected by ELISA method and trace neutralization assay, respectively.The specific CTL cytotoxic activity was detected by CCK-8 assay. The mice in each group were challenged with lethal dose of coxsackievirus, the titers of the sera virus were titrated. RESULTS: The recombinant adenovirus Ad/sVP1-C3d3 was successfully constructed. It's observed that the titers of CVB3 VP1 specific antibody and neutralizing antibody were much higher than those of the other three groups(P<0.01). CTL cytotoxicity activities was much higher than PBS and Ad group(P<0.01), but little higher than Ad/VP1 group(P<0.05).The titer of sera virus was lower than Ad and PBS groups after CVB3 challenged(P<0.05). CONCLUSION: Both the celluar and humoral immune responses in mice could been significantly enhanced by Ad/sVP1-C3d3.
Assuntos
Adenoviridae/química , Adenoviridae/imunologia , Complemento C3d/imunologia , Infecções por Coxsackievirus/imunologia , Enterovirus Humano B/imunologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/imunologia , Adenoviridae/genética , Animais , Complemento C3d/genética , DNA Recombinante/química , DNA Recombinante/genética , DNA Recombinante/imunologia , Enterovirus Humano B/genética , Células HEK293 , Células HeLa , Humanos , Imunização/métodos , Imunoglobulina G/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/genética , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/imunologia , Vacinas Virais/química , Vacinas Virais/genética , Vacinas Virais/imunologiaRESUMO
Porcine reproductive and respiratory syndrome virus (PRRSV) has recently caused catastrophic losses in swine industry worldwide. The commercial vaccines only provide a limited protection against PRRSV infection. At present, DNA vaccine is the focus on the new vaccines. The gene fragment (p28) coding for the molecular adjuvants complement protein C3d (mC3d) from BALB/c mouse was cloned and expressed as a fusion protein for its application in the vaccine study of mice. Three potential vaccines construct units were engineered to contain two, four and six copies of mC3d-p28 coding gene linked to the GP5 gene of PRRSV and one vaccine expressing GP5 alone (pcDNA3.1-GP5) was constructed. Subsequently, the vaccines' abilities to elicit the humoral and cellular immune responses were investigated in mice. These results showed that significantly enhanced GP5-specific ELISA antibody, GP5-specific neutralizing antibody, IFN-γ level, and IL-4 level, could be induced in mice immunized with DNA construct units encoding the pcDNA3.1-C3d-p28.n-GP5 than those received DNA vaccine expressing GP5 alone (pcDNA3.1-GP5). Analysis of the immunogenicity of different repeats of mC3d-p28 revealed that mC3d-p28 had an enhancing effect on the immunogenicity of antigens, and that six or more repeats of mC3d-p28 may be necessary for efficient enhancement of antigen specific immune responses. This approach may provide a new strategy for the development of efficient vaccines against the PRRSV for pigs in the future.
Assuntos
Complemento C3d/imunologia , Vacinas de DNA/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Complemento C3d/genética , Ensaio de Imunoadsorção Enzimática , Feminino , Interferon gama/metabolismo , Interleucina-4/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Vacinas de DNA/genética , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Proteínas do Envelope Viral/genéticaAssuntos
Complexo Antígeno-Anticorpo/sangue , Artrite Reumatoide/diagnóstico , Complemento C3d/imunologia , Testes Imunológicos/métodos , Lúpus Eritematoso Sistêmico/diagnóstico , Anticorpos Monoclonais , Biomarcadores/sangue , Humanos , Neoplasias/diagnóstico , Kit de Reagentes para Diagnóstico , Valores de Referência , Manejo de Espécimes/métodosRESUMO
Although the critical role of complement component C3d as a molecular adjuvant in preventing virus infection is well established, its role in cancer therapies is unclear. In this study, we have engineered a DNA vaccine that expresses extracellular region of murine VEGFR-2 (FLK1(265-2493)) and 3 copies of C3d (C3d3), a component of complement as a molecular adjuvant, designed to increase antitumor immunity. VEGFR-2 has a more restricted expression on endothelial cells and is upregulated once these cells proliferate during angiogenesis in the tumor vasculature. Immunization of mice with vector encoding FLK1(265-2493) alone generated only background levels of anti-VEGFR-2 antibodies and slight inhibitory effect on tumor growth. However, the addition of C3d3 to the vaccine construct significantly augmented the anti-VEGFR-2 humoral immune response and inhibited the tumor growth. The antitumor activity induced by vaccination with vector encoding FLK1(265-2493)-C3d3 fusion protein was also demonstrated via growth inhibition of established tumors following passive transfer of immune serum from vaccinated mice. Our results suggest that vaccination with vector encoding FLK1(265-2493) with C3d3 as a molecular adjuvant induces adaptive humoral activity, which is directed against the murine VEGFR-2 and can significantly inhibit tumor growth, and that administration of C3d as a molecular adjuvant to increase antibodies levels to VEGFR-2 may provide an alternative treatment modality for cancer therapies.
Assuntos
Adjuvantes Imunológicos/farmacologia , Vacinas Anticâncer/imunologia , Complemento C3d/administração & dosagem , Melanoma Experimental/terapia , Vacinas de DNA/imunologia , Animais , Complemento C3d/imunologia , Células Endoteliais/imunologia , Feminino , Células HeLa , Humanos , Imunidade Humoral , Imunoterapia Adotiva , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/imunologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/administração & dosagem , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/imunologiaRESUMO
BACKGROUND: Rift Valley fever (RVF) is an arthropod-borne viral zoonosis. Rift Valley fever virus (RVFV) is an important biological threat with the potential to spread to new susceptible areas. In addition, it is a potential biowarfare agent. METHODOLOGY/PRINCIPAL FINDINGS: We developed two potential vaccines, DNA plasmids and alphavirus replicons, expressing the Gn glycoprotein of RVFV alone or fused to three copies of complement protein, C3d. Each vaccine was administered to mice in an all DNA, all replicon, or a DNA prime/replicon boost strategy and both the humoral and cellular responses were assessed. DNA plasmids expressing Gn-C3d and alphavirus replicons expressing Gn elicited high titer neutralizing antibodies that were similar to titers elicited by the live-attenuated MP12 virus. Mice vaccinated with an inactivated form of MP12 did elicit high titer antibodies, but these antibodies were unable to neutralize RVFV infection. However, only vaccine strategies incorporating alphavirus replicons elicited cellular responses to Gn. Both vaccines strategies completely prevented weight loss and morbidity and protected against lethal RVFV challenge. Passive transfer of antisera from vaccinated mice into naïve mice showed that both DNA plasmids expressing Gn-C3d and alphavirus replicons expressing Gn elicited antibodies that protected mice as well as sera from mice immunized with MP12. CONCLUSION/SIGNIFICANCE: These results show that both DNA plasmids expressing Gn-C3d and alphavirus replicons expressing Gn administered alone or in a DNA prime/replicon boost strategy are effective RVFV vaccines. These vaccine strategies provide safer alternatives to using live-attenuated RVFV vaccines for human use.