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1.
Genes (Basel) ; 10(10)2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31635417

RESUMO

PURPOSE: To identify rare genetic variants in early age-related macular degeneration (AMD) utilizing whole-exome sequencing (WES). METHODS: Eight non-related early-AMD families of different Jewish ethnicities were ascertained. Initial mutation screening (phase-1) included common complement factor-H (CFH) p.Y402H; and age related maculopathy susceptibility 2 (ARMS2) p.A69S; and rare variants complement factor-I (CFI) p.V412M; and hemicentin1 (HMCN1) c.4163delC identified previously in our population. Four families, whose initial screening for the aforementioned variants was negative, underwent WES (phase-2). Bioinformatics filtering was based on functionality (from a panel of 234 genes with proven or presumed association to AMD); predicted severity; and frequency (rare variants with minor allele frequency <1%). When applicable, further screening for specific rare variants was carried out on additional cases of similar ethnicities and phenotypes (phase-3). RESULTS: Phase-1 identified three families carrying CFI p.V412M mutation. WES analysis detected probable disease-related variants in three out of the remaining families. These included: a family with a variant in PLEKHA1 gene p.S177N; a family with previously reported variant p.R1210C in CFH gene; and two families with the C3 p.R735W variant. CONCLUSIONS: Rare, high-penetrance variants have a profound contribution to early-AMD pathogenesis. Utilization of WES in genetic research of multifactorial diseases as AMD, allows a thorough comprehensive analysis with the identification of previously unreported rare variants.


Assuntos
Degeneração Macular/genética , Mutação , Idoso , Fator H do Complemento/genética , Fator I do Complemento/genética , Feminino , Heterozigoto , Humanos , Imunoglobulinas/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Judeus/genética , Degeneração Macular/etnologia , Degeneração Macular/patologia , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Linhagem , Penetrância , Proteínas/genética
2.
Dis Markers ; 2019: 9602949, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31583032

RESUMO

Background: To determine the impact of HTRA1 rs1120638, TIMP3 rs9621532, VEGFA rs833068, CFI rs10033900, ERCC6 rs3793784, and KCTD10 rs56209061 genotypes on the development of age-related macular degeneration (AMD) in the Lithuanian population. Methods: A total of 916 subjects were examined: 309 patients with early AMD, 301 patients with exudative AMD, and 306 healthy controls. The genotyping of HTRA1 rs11200638, TIMP3 rs9621532, VEGFA rs833068, CFI rs10033900, ERCC6 rs3793784, and KCTD10 rs56209061 was carried out using the RT-PCR method. Results: Our study showed that single-nucleotide polymorphisms rs3793784 and rs11200638 were associated with increased odds of early and exudative AMD, and the variant in KCTD10 (rs56209061) was found to be associated with decreased odds of early and exudative AMD development after adjustments for age and gender in early AMD analysis and after adjustments only for age in exudative AMD. The haplotype containing two minor alleles C-A and the G-A haplotype in rs3793784-rs11200638 were statistically significantly associated with an increased risk of exudative AMD development after adjustment for age, while the G-G haplotype showed a protective role against early and exudative AMD and the haplotype C-G in rs3793784-rs11200638 was associated with a decreased risk only of exudative AMD development. Conclusions: Our study identified two markers, rs11200638 and rs3793784, as risk factors for early and exudative AMD, and one marker, rs56209061, as a protective factor for early and exudative AMD development. The haplotypes constructed of rs3793784-rs11200638 were found to be associated with AMD development, as well.


Assuntos
DNA Helicases/genética , Enzimas Reparadoras do DNA/genética , Predisposição Genética para Doença , Serina Peptidase 1 de Requerimento de Alta Temperatura A/genética , Degeneração Macular/diagnóstico , Proteínas de Ligação a Poli-ADP-Ribose/genética , Polimorfismo de Nucleotídeo Único , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Idoso , Idoso de 80 Anos ou mais , Alelos , Biomarcadores/sangue , Estudos de Casos e Controles , Fator I do Complemento/genética , Fator I do Complemento/metabolismo , DNA Helicases/sangue , Enzimas Reparadoras do DNA/sangue , Feminino , Expressão Gênica , Frequência do Gene , Haplótipos , Serina Peptidase 1 de Requerimento de Alta Temperatura A/sangue , Humanos , Degeneração Macular/sangue , Degeneração Macular/genética , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Razão de Chances , Proteínas de Ligação a Poli-ADP-Ribose/sangue , Canais de Potássio de Abertura Dependente da Tensão da Membrana/sangue , Risco , Inibidor Tecidual de Metaloproteinase-3/sangue , Inibidor Tecidual de Metaloproteinase-3/genética , Fator A de Crescimento do Endotélio Vascular/sangue , Fator A de Crescimento do Endotélio Vascular/genética
3.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31548314

RESUMO

Leptospirosis is one of the most widespread zoonoses caused by pathogenic Leptospira spp. In this study, we report that the LIC11966/ErpY-like lipoprotein is a surface-exposed outer membrane protein exclusively present in pathogenic species of Leptospira The recombinant ErpY (rErpY)-like protein is recognized by the immunoglobulins of confirmed leptospirosis sera of diverse hosts (human, bovine, and canine), suggesting the expression of the native leptospiral surface protein during infection. Circular dichroism of pure rErpY-like protein showed the secondary structural integrity to be uncompromised during the purification process. Analysis of the rErpY-like protein by native polyacrylamide gel electrophoresis, chemical cross-linking, dynamic light scattering, and field emission transmission electron microscopy demonstrated it undergoes supramolecular assembly. The rErpY-like protein can bind to diverse host extracellular matrices, and it presented a saturable and strong binding affinity (dissociation constant [KD ] of 70.45 ± 4.13 nM) to fibrinogen, a central host plasma component involved in blood clotting. In the presence of the rErpY-like supramolecule, thrombin-catalyzed fibrin clot formation is inhibited up to 7%, implying its role in inhibiting blood coagulation during Leptospira infection. In addition, binding of the rErpY-like supramolecule to complement factors H and I suggests the protein also contributes to Leptospira evading innate host defense during infection by inactivating alternative complement pathways. This study reveals that rErpY-like protein is functionally active in the supramolecular state and performs moonlighting activity under the given in vitro conditions.


Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Coagulação Sanguínea/fisiologia , Fator H do Complemento/metabolismo , Fator I do Complemento/metabolismo , Leptospira/imunologia , Leptospirose/diagnóstico , Animais , Dicroísmo Circular , Via Alternativa do Complemento/imunologia , Feminino , Tempo de Lise do Coágulo de Fibrina , Lipoproteínas/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Ligação Proteica , Estrutura Secundária de Proteína , Trombina/metabolismo
4.
Immunobiology ; 224(4): 511-517, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31109748

RESUMO

Factor I was first discovered in 1966. Its importance became apparent with the description of the original Factor I deficient patient in Boston in 1967. This patient presented with a hyperactive alternative complement pathway resulting in secondary complement deficiency due to continuous complement consumption. On the basis of these findings, the mechanism of the alternative pathway was worked out. In 1975, the surprise finding was made that elevating levels of Factor I in plasma down-regulated the alternative pathway. Attempts to exploit this finding for clinical use had a long and frustrating history and it was not until 2019 that the first patient was treated with the gene therapy vector for age related macular degeneration by Professor Sir Robert MacLaren in Oxford. This review follows the long and contorted course from initial observations to clinical use of complement Factor I.


Assuntos
Fator I do Complemento/fisiologia , Animais , Ativação do Complemento/genética , Ativação do Complemento/imunologia , Fator I do Complemento/química , Fator I do Complemento/uso terapêutico , Via Alternativa do Complemento/genética , Via Alternativa do Complemento/imunologia , Proteínas do Sistema Complemento/genética , Proteínas do Sistema Complemento/imunologia , Proteínas do Sistema Complemento/metabolismo , Suscetibilidade a Doenças , Humanos , Imunoconglutininas/imunologia , Transdução de Sinais , Relação Estrutura-Atividade
5.
Pathol Res Pract ; 215(6): 152369, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30987833

RESUMO

Ovarian carcinoma is one of the most lethal malignancies, but only very few prognostic biomarkers are known. The degradome, comprising proteases, protease non-proteolytic homologues and inhibitors, have been involved in the prognosis of many cancer types, including ovarian carcinoma. The prognostic significance of the whole degradome family has not been specifically studied in high-grade serous ovarian cancer. A targeted DNA microarray known as the CLIP-CHIP microarray was used to identify potential prognostic factors in ten high-grade serous ovarian cancer women who had early recurrence (<1.6 years) or late/no recurrence after first line surgery and chemotherapy. In women with early recurrence, we identified seven upregulated genes (TMPRSS4, MASP1/3, SPC18, PSMB1, IGFBP2, CFI - encoding Complement Factor I - and MMP9) and one down-regulated gene (ADAM-10). Using immunohistochemistry, we evaluated the prognostic effect of these 8 candidate genes in an independent cohort of 112 high-grade serous ovarian cancer women. Outcomes were progression, defined according to CA-125 criteria, and death. Multivariate Cox proportional hazard regression models were done to estimate the associations between each protein and each outcome. High ADAM-10 expression (intensity of 2-3) was associated with a lower risk of progression (adjusted hazard ratio (HR): 0.51; 95% confidence interval (CI): 0.29-0.87). High complement factor I expression (intensity 2-3) was associated with a higher risk of progression (adjusted HR: 2.30, 95% CI: 1.17-4.53) and death (adjusted HR: 3.42; 95% CI: 1.72-6.79). Overall, we identified the prognostic value of two proteases, ADAM-10 and complement factor I, for high-grade serous ovarian cancer which could have clinical significance.


Assuntos
Proteína ADAM10/biossíntese , Secretases da Proteína Precursora do Amiloide/biossíntese , Fator I do Complemento/biossíntese , Cistadenocarcinoma Seroso/patologia , Proteínas de Membrana/biossíntese , Neoplasias Ovarianas/patologia , Idoso , Biomarcadores Tumorais/análise , Cistadenocarcinoma Seroso/metabolismo , Cistadenocarcinoma Seroso/mortalidade , Feminino , Humanos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/mortalidade , Prognóstico , Intervalo Livre de Progressão
6.
Nan Fang Yi Ke Da Xue Xue Bao ; 39(1): 13-22, 2019 Jan 30.
Artigo em Chinês | MEDLINE | ID: mdl-30692061

RESUMO

OBJECTIVE: To screen potential plasma protein biomarkers for the progression of cervical precancerous lesions into cervical carcinoma and analyze their functions. METHODS: Plasma samples obtained from healthy control subjects, patients with low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL), cervical cancer (CC), and patients with CC after treatment were enriched for low-abundance proteins for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. The MS data of the samples were analyzed using Discoverer 2.2 software, and the differential proteins (peptide coverage ≥20%, unique peptides≥2) were screened by comparison of LSIL, HSIL and CC groups against the control group followed by verification using target proteomics technology. Protein function enrichment and coexpression analyses were carried out to explore the role of the differentially expressed proteins as potential biomarkers and their pathological mechanisms. RESULTS: Compared with the control group, both LSIL group and HSIL group showed 9 differential proteins; 5 differentially expressed proteins were identified in CC group. The proteins ORM2 and HPR showed obvious differential expressions in LSIL and HSIL groups compared with the control group, and could serve as potential biomarkers for the progression of cervical carcinoma. The expression of F9 increased consistently with the lesion progression from LSIL to HSIL and CC, suggesting its value as a potential biomarker for the progression of cervical cancer. CFI and AFM protein levels were obviously decreased in treated patients with CC compared with the patients before treatment, indicating their predictive value for the therapeutic efficacy. Protein function enrichment analysis showed that all these differentially expressed proteins were associated with the complement system and the coagulation cascades pathway. CONCLUSIONS: We identified 5 new protein biomarkers (F9, CFI, AFM, HPR, and ORM2) for cervical precancerous lesions and for prognostic evaluation of CC, and combined detection of these biomarkers may help in the evaluation of the development and progression of CC and also in improving the diagnostic sensitivity and specificity of cervical lesions.


Assuntos
Biomarcadores Tumorais/sangue , Neoplasia Intraepitelial Cervical/sangue , Cromatografia Líquida , Lesões Pré-Cancerosas/sangue , Espectrometria de Massas em Tandem , Neoplasias do Colo do Útero/sangue , Antígenos de Neoplasias/sangue , Proteínas de Transporte/sangue , Estudos de Casos e Controles , Neoplasia Intraepitelial Cervical/diagnóstico , Fator I do Complemento/análise , Detecção Precoce de Câncer , Feminino , Glicoproteínas/sangue , Haptoglobinas , Humanos , Proteínas de Neoplasias/sangue , Orosomucoide/análise , Lesões Pré-Cancerosas/diagnóstico , Albumina Sérica Humana , Neoplasias do Colo do Útero/diagnóstico
7.
Nephrology (Carlton) ; 24(3): 357-364, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29292855

RESUMO

AIM: The aim of the present study was to characterize the molecular basis of complement factor I deficiency in Tunisian atypical haemolytic and uremic syndrome patients with low factor I levels. METHODS: Six adults and seven children were enrolled in this study. Complement factor I levels were assessed by a homemade sandwich ELISA and ranged between 12.5% and 60%. Genomic DNA was amplified by way of a polymerase chain reaction using intronic primers flanking the 13 coding exons. Sequencing of amplified products was carried out by the dye terminator sequencing method. Molecular study was performed on parental samples for three dead paediatric patients. The control group consisted of 100 healthy Tunisian donors. RESULTS: We identified a total of 13 substitutions and one insertion: seven in introns, four in exons and three in UTR. The new mutations were c.-132G > C, c.71 + 181 T > A in 5'UTR and intron 1, respectively. Three intronic polymorphisms were predicted to have impact on splicing events: c.482 + 6C > T, c.884-42_884-41insTTAAA (rs34422850) and c.1429 + 33 A > G (rs9998151). They were three missense mutations leading to a p.Ile 357Met, p.Ile416Leu and p.GLu548Gln. p.Ile 357Met was found in two patients and one relative. Half of the patients had associated mutation and/or polymorphisms. CONCLUSION: This is the first genetic study in Tunisian and Maghrebin atypical haemolytic and uraemic syndrome patients. The high occurrence of Ile357Met mutation may reflect a founding effect. Functional impact of the two new mutations c.-132G > C and c.71 + 181A > T have to be studied. Association of simultaneous genetic abnormalities may explain the variability of atypical haemolytic and uraemic syndrome, penetrance and disease phenotype.


Assuntos
Síndrome Hemolítico-Urêmica Atípica , Complemento C3/deficiência , Fator I do Complemento , Doenças Genéticas Inatas , Adulto , Síndrome Hemolítico-Urêmica Atípica/diagnóstico , Síndrome Hemolítico-Urêmica Atípica/epidemiologia , Síndrome Hemolítico-Urêmica Atípica/genética , Criança , Pré-Escolar , Estudos de Coortes , Complemento C3/genética , Fator I do Complemento/análise , Fator I do Complemento/genética , Feminino , Doenças Genéticas Inatas/sangue , Doenças Genéticas Inatas/epidemiologia , Doenças Genéticas Inatas/genética , Doenças da Deficiência Hereditária de Complemento , Humanos , Lactente , Masculino , Mutação , Polimorfismo Genético , Tunísia/epidemiologia
8.
Int Ophthalmol ; 39(3): 551-556, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29392637

RESUMO

BACKGROUND: Age-related macular degeneration (AMD) is a complex disease, and recent studies have shown role of complement system genes in its development. Complement factor I regulates the complement pathways, and relationship between CFI polymorphisms and AMD is controversial. We evaluated the possible association of complement factor I rs141853578 (G119R) variation with advanced AMD in Iranian patients. MATERIALS AND METHODS: We included 371 case-control samples consisting of 220 advanced AMD patients and 151 genetically unrelated healthy controls. Extracted DNA samples amplified to obtain fragment including the polymorphic complement factor I rs141853578 (G119R) region. RESULTS: The distribution of the genotypes was significantly different in the AMD patients compared to that of controls (p = 0.035). The TT genotype frequencies for CFI were significantly higher in AMD group (7.7 vs. 2%, OR 4.67, CI 1.33-16.45, p = 0.016). This significant difference was maintained after adjustment for the effects of age and gender (OR 5.09, CI 1.42-18.20, p = 0.012). The minor allele frequency (T allele) was also significantly higher in AMD patients compared to that of controls (29.3 vs. 21.5% OR 1.51, CI 1.07-2.13, p = 0.018). CONCLUSION: Current study showed that CFI rs141853578 (G119R) is a risk factor for developing advanced type AMD. This study also suggests that the frequency of G119R polymorphism in our population is not as rare as reported from other populations.


Assuntos
Fator I do Complemento/genética , DNA/genética , Degeneração Macular/genética , Polimorfismo de Nucleotídeo Único , Idoso , Alelos , Estudos de Casos e Controles , Fator I do Complemento/metabolismo , Feminino , Frequência do Gene , Genótipo , Humanos , Incidência , Irã (Geográfico)/epidemiologia , Degeneração Macular/diagnóstico , Degeneração Macular/epidemiologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Epitélio Pigmentado da Retina/patologia , Tomografia de Coerência Óptica
9.
J Neuroinflammation ; 15(1): 254, 2018 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-30180861

RESUMO

BACKGROUND: The complement system plays an important role in many neurological disorders. Complement modulation, including C3/C3a receptor signaling, shows promising therapeutic effects on cognition and neurodegeneration. Yet, the implications for this pathway in perioperative neurocognitive disorders (PND) are not well established. Here, we evaluated the possible role for C3/C3a receptor signaling after orthopedic surgery using an established mouse model of PND. METHODS: A stabilized tibial fracture surgery was performed in adult male C57BL/6 mice under general anesthesia and analgesia to induce PND-like behavior. Complement activation was assessed in the hippocampus and choroid plexus. Changes in hippocampal neuroinflammation, synapse numbers, choroidal blood-cerebrospinal fluid barrier (BCSFB) integrity, and hippocampal-dependent memory function were evaluated after surgery and treatment with a C3a receptor blocker. RESULTS: C3 levels and C3a receptor expression were specifically increased in hippocampal astrocytes and microglia after surgery. Surgery-induced neuroinflammation and synapse loss in the hippocampus were attenuated by C3a receptor blockade. Choroidal BCSFB dysfunction occurred 1 day after surgery and was attenuated by C3a receptor blockade. Administration of exogenous C3a exacerbated cognitive decline after surgery, whereas C3a receptor blockade improved hippocampal-dependent memory function. CONCLUSIONS: Orthopedic surgery activates complement signaling. C3a receptor blockade may be therapeutically beneficial to attenuate neuroinflammation and PND.


Assuntos
Ativação do Complemento/fisiologia , Proteínas do Sistema Complemento/metabolismo , Hipocampo/metabolismo , Transtornos Neurocognitivos , Complicações Pós-Operatórias/fisiopatologia , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Barreira Hematoencefálica/fisiopatologia , Plexo Corióideo/metabolismo , Ativação do Complemento/efeitos dos fármacos , Fator I do Complemento/farmacologia , Condicionamento Clássico/efeitos dos fármacos , Condicionamento Clássico/fisiologia , Citofagocitose/efeitos dos fármacos , Modelos Animais de Doenças , Medo/psicologia , Hipocampo/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/metabolismo , Transtornos Neurocognitivos/tratamento farmacológico , Transtornos Neurocognitivos/etiologia , Transtornos Neurocognitivos/metabolismo , Transtornos Neurocognitivos/patologia , Complicações Pós-Operatórias/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Sinapses/efeitos dos fármacos , Sinapses/patologia , Fraturas da Tíbia/cirurgia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia
10.
Free Radic Biol Med ; 129: 237-246, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30253188

RESUMO

Complement Factor H (CFH) is an important inhibitor of the alternate complement pathway in Bruch's membrane (BM), located between the choriocapillaris and the retinal pigment epithelium. Furthermore dysfunction of its activity as occurs with certain polymorphisms is associated with an increased risk of age related macular degeneration (AMD). The retina is a site of high generation of reactive oxygen species (ROS) and dysfunction of redox homeostasis in this milieu also contributes to AMD pathogenesis. In this study we wanted to explore if CFH exists in distinct redox forms and whether these species have unique protective biological functions. CFH can be reduced by the naturally occurring thioredoxin - 1 in CFH domains 1-4, 17-20. We found a duality of function between the oxidised and reduced forms of CFH. The oxidised form was more efficient in binding to C3b and lipid peroxidation by-products that are known to accumulate in the retinae and activate the alternate complement pathway. Oxidised CFH enhances Factor I mediated cleavage of C3 and C3b whereas the reduced form loses this activity. In the setting of oxidative stress (hydrogen peroxide)-mediated death of human retinal pigment epithelial cells as can occur in AMD, the free thiol form of CFH offers a protective function compared to the oxidised form. We found for the first time using a novel ELISA system we have developed for free thiol CFH, that both redox forms of CFH are found in the human plasma. Furthermore there is a distinct ratio of these redox forms in plasma depending if an individual has early or late AMD, with individuals with early AMD having higher levels of the free thiol form compared to late AMD.


Assuntos
Complemento C3b/metabolismo , Fator I do Complemento/metabolismo , Degeneração Macular/genética , Espécies Reativas de Oxigênio/metabolismo , Idoso , Lâmina Basilar da Corioide/imunologia , Lâmina Basilar da Corioide/patologia , Estudos de Casos e Controles , Linhagem Celular , Ativação do Complemento/genética , Complemento C3b/genética , Fator H do Complemento/genética , Fator H do Complemento/metabolismo , Fator I do Complemento/genética , Via Alternativa do Complemento/genética , Células Epiteliais/citologia , Células Epiteliais/imunologia , Feminino , Expressão Gênica , Humanos , Peroxidação de Lipídeos , Degeneração Macular/imunologia , Degeneração Macular/patologia , Masculino , Oxirredução , Ligação Proteica , Proteólise , Espécies Reativas de Oxigênio/imunologia , Epitélio Pigmentado da Retina/imunologia , Epitélio Pigmentado da Retina/patologia , Fatores de Tempo
11.
Molecules ; 23(7)2018 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-30002298

RESUMO

Infusion reactions (IRs) are common immune-mediated side effects in patients treated with a variety of drug products, including, but not limited to, nanotechnology formulations. The mechanism of IRs is not fully understood. One of the best studied mechanisms of IRs to nanomedicines is the complement activation. However, it is largely unknown why some patients develop reactions to nanomedicines while others do not, and why some nanoparticles are more reactogenic than others. One of the theories is that the pre-existing anti-polyethylene glycol (PEG) antibodies initiate the complement activation and IRs in patients. In this study, we investigated this hypothesis in the case of PEGylated liposomal doxorubicin (Doxil), which, when used in a clinical setting, is known to induce IRs; referred to as complement activation-related pseudoallergy (CARPA) in sensitive individuals. We conducted the study in vitro using plasma derived from C57BL/6 mice and twenty human donor volunteers. We used mouse plasma to test a library of well-characterized mouse monoclonal antibodies with different specificity and affinity to PEG as it relates to the complement activation by Doxil. We determined the levels of pre-existing polyclonal antibodies that bind to PEG, methoxy-PEG, and PEGylated liposomes in human plasma, and we also assessed complement activation by Doxil and concentrations of complement inhibitory factors H and I in these human plasma specimens. The affinity, specificity, and other characteristics of the human polyclonal antibodies are not known at this time. Our data demonstrate that under in vitro conditions, some anti-PEG antibodies contribute to the complement activation by Doxil. Such contribution, however, needs to be considered in the context of other factors, including, but not limited to, antibody class, type, clonality, epitope specificity, affinity, and titer. In addition, our data contribute to the knowledge base used to understand and improve nanomedicine safety.


Assuntos
Anticorpos Monoclonais , Ativação do Complemento , Inativadores do Complemento , Doxorrubicina/análogos & derivados , Polietilenoglicóis , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Ativação do Complemento/efeitos dos fármacos , Ativação do Complemento/imunologia , Fator H do Complemento/imunologia , Fator I do Complemento/imunologia , Inativadores do Complemento/química , Inativadores do Complemento/imunologia , Doxorrubicina/farmacologia , Hipersensibilidade a Drogas/imunologia , Humanos , Camundongos , Polietilenoglicóis/farmacologia
12.
BMC Nephrol ; 19(1): 148, 2018 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-29940891

RESUMO

BACKGROUND: It has been suggested that C3 glomerulonephritis (C3GN) and atypical hemolytic-uremic syndrome (a stereotypical phenotype of thrombotic microangiopathy), two rare entities caused by complement alternative pathway dysregulation share overlapping genetic origin and can be triggered by infections. CASE PRESENTATION: We report a case of concomitant C3GN and thrombotic microangiopathy (TMA) after pulmonary infection in a young male receiving kidney transplantation. Genetic assessment revealed two missense variations in compound heterozygous form in CFI gene (complement factor I). These two variations are segregated with disease in the core family member of this patient. Plasma CFI levels of the patient and family members were all in normal range. We considered that these two variations only impair CFI function rather than its quantity in the serum. CONCLUSION: Our case supports that C3GN and TMA shared overlapping genetic variations and might be triggered by infection in genetically susceptible patients after kidney transplantation.


Assuntos
Aloenxertos/patologia , Complemento C3/análise , Fator I do Complemento/genética , Glomerulonefrite/diagnóstico , Transplante de Rim/tendências , Microangiopatias Trombóticas/diagnóstico , Adulto , Variação Genética/genética , Glomerulonefrite/etiologia , Glomerulonefrite/genética , Humanos , Rim/patologia , Transplante de Rim/efeitos adversos , Masculino , Microangiopatias Trombóticas/etiologia , Microangiopatias Trombóticas/genética
13.
Clin Genet ; 94(3-4): 330-338, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29888403

RESUMO

Genetic alterations in the complement system have been linked to a variety of diseases, including atypical hemolytic uremic syndrome (aHUS), C3 glomerulopathy (C3G), and age-related macular degeneration (AMD). We performed sequence analysis of the complement genes complement factor H (CFH), complement factor I (CFI), and complement C3 (C3) in 866 aHUS/C3G and 697 AMD patients. In total, we identified 505 low-frequency alleles, representing 121 unique variants, of which 51 are novel. CFH contained the largest number of unique low-frequency variants (n = 64; 53%), followed by C3 (n = 32; 26%) and CFI (n = 25; 21%). A substantial number of variants were found in both patients groups (n = 48; 40%), while 41 (34%) variants were found only in aHUS/C3G and 32 (26%) variants were AMD specific. Genotype-phenotype correlations between the disease groups identified a higher frequency of protein altering alleles in short consensus repeat 20 (SCR20) of factor H (FH), and in the serine protease domain of factor I (FI) in aHUS/C3G patients. In AMD, a higher frequency of protein-altering alleles was observed in SCR3, SCR5, and SCR7 of FH, the SRCR domain of FI, and in the MG3 domain of C3. In conclusion, we observed a substantial overlap of variants between aHUS/C3G and AMD; however, there is a distinct clustering of variants within specific domains.


Assuntos
Síndrome Hemolítico-Urêmica Atípica/genética , Complemento C3/genética , Fator I do Complemento/genética , Genótipo , Glomerulonefrite Membranosa/genética , Degeneração Macular/genética , Fenótipo , Síndrome Hemolítico-Urêmica Atípica/metabolismo , Síndrome Hemolítico-Urêmica Atípica/fisiopatologia , Estudos de Coortes , Complemento C3/metabolismo , Fator H do Complemento/genética , Fator H do Complemento/metabolismo , Fator I do Complemento/metabolismo , Predisposição Genética para Doença , Glomerulonefrite Membranosa/metabolismo , Glomerulonefrite Membranosa/fisiopatologia , Humanos , Degeneração Macular/metabolismo , Degeneração Macular/fisiopatologia
14.
Kidney Int ; 94(2): 408-418, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29907460

RESUMO

Atypical hemolytic uremic syndrome (aHUS) is a rare, genetic, life-threatening disease. The Global aHUS Registry collects real-world data on the natural history of the disease. Here we characterize end-stage renal disease (ESRD)-free survival, the rate of thrombotic microangiopathy, organ involvement and the genetic background of 851 patients in the registry, prior to eculizumab treatment. A sex-specific difference was apparent according to age at initial disease onset as the ratio of males to females was 1.3:1 for childhood presentation and 1:2 for adult presentation. Complement Factor I and Membrane Cofactor Protein mutations were more common in patients with initial presentation as adults and children, respectively. Initial presentation in childhood significantly predicted ESRD risk (adjusted hazard ratio 0.55 [95% confidence interval 0.41-0.73], whereas sex, race, family history of aHUS, and time from initial presentation to diagnosis, did not. Patients with a Complement Factor H mutation had reduced ESRD-free survival, whereas Membrane Cofactor Protein mutation was associated with longer ESRD-free survival. Additionally extrarenal organ manifestations occur in 19%-38% of patients within six months of initial disease presentation (dependent on organ). Thus, our real-world results provide novel insights regarding phenotypic variables and genotypes on the clinical manifestation and progression of aHUS.


Assuntos
Síndrome Hemolítico-Urêmica Atípica/mortalidade , Falência Renal Crônica/epidemiologia , Fenótipo , Adolescente , Adulto , Idade de Início , Síndrome Hemolítico-Urêmica Atípica/genética , Síndrome Hemolítico-Urêmica Atípica/patologia , Criança , Fator H do Complemento/genética , Fator I do Complemento/genética , Progressão da Doença , Feminino , Humanos , Falência Renal Crônica/patologia , Masculino , Proteína Cofatora de Membrana/genética , Estudos Prospectivos , Sistema de Registros/estatística & dados numéricos , Estudos Retrospectivos , Fatores Sexuais , Adulto Jovem
15.
PLoS One ; 13(5): e0195751, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29782502

RESUMO

Activation of the alternative complement cascade has been implicated in the pathogenesis of age related macular degeneration (AMD) and Alzheimer's disease (AD). Amyloid ß (Aß), a component of drusen, may promote complement activation by inhibiting CFI bioactivity. We determined whether Aß reduced CFI bioactivity and whether antibodies against Aß including a monoclonal antibody, GSK933776 could restore CFI bioactivity. We also measured CFI bioactivity in plasma of subjects with AMD and AD. In support of the GSK933776 development program in AMD (geographic atrophy), we developed a quantitative assay to measure CFI bioactivity based on its ability to cleave C3b to iC3b, and repeated it in presence or absence of Aß and anti-Aß antibodies. Using this assay, we measured CFI bioactivity in plasma of 194 subjects with AMD, and in samples from subjects with AD that had been treated with GSK933776 as part of the GSK933776 development program in AD. Aß reduced the CFI bioactivity by 5-fold and pre-incubation with GSK933776 restored CFI bioactivity. In subjects with AMD, plasma CFI levels and bioactivity were not significantly different from non-AMD controls. However, we detected a positive linear trend, suggesting increasing activity with disease severity. In subjects with AD, we observed a 10% and 27% increase in overall CFI bioactivity after treatment with GSK933776 during the second and third dose. Our studies indicate that CFI enzymatic activity can be inhibited by Aß and be altered in proinflammatory diseases such as AMD and AD, in which deposition of Aß and activation of the alternative complement cascade are believed to play a key role in the disease process.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Peptídeos beta-Amiloides/antagonistas & inibidores , Anticorpos Monoclonais/uso terapêutico , Biomarcadores/sangue , Ativação do Complemento/efeitos dos fármacos , Fator I do Complemento/metabolismo , Degeneração Macular/tratamento farmacológico , Idoso , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/imunologia , Estudos de Casos e Controles , Feminino , Humanos , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino
16.
Front Immunol ; 9: 735, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29696024

RESUMO

Cutaneous leukocytoclastic vasculitis arises from immune complex deposition and dysregulated complement activation in small blood vessels. There are many causes, including dysregulated host response to infection, drug reactions, and various autoimmune conditions. It is increasingly recognised that some monogenic autoinflammatory diseases cause vasculitis, although genetic causes of vasculitis are extremely rare. We describe a child of consanguineous parents who presented with chronic cutaneous leukocytoclastic vasculitis, recurrent upper respiratory tract infection, and hypocomplementaemia. A homozygous p.His380Arg mutation in the complement factor I (CFI) gene CFI was identified as the cause, resulting in complete absence of alternative complement pathway activity, decreased classical complement activity, and low levels of serum factor I, C3, and factor H. C4 and C2 levels were normal. The same homozygous mutation and immunological defects were also identified in an asymptomatic sibling. CFI deficiency is thus now added to the growing list of monogenic causes of vasculitis and should always be considered in vasculitis patients found to have persistently low levels of C3 with normal C4.


Assuntos
Complemento C3/deficiência , Fator I do Complemento/genética , Doenças Genéticas Inatas/genética , Vasculite Leucocitoclástica Cutânea/genética , Criança , Complemento C3/genética , Feminino , Doenças da Deficiência Hereditária de Complemento , Humanos , Infecções , Recidiva
17.
J Immunol Methods ; 457: 30-32, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29625075

RESUMO

A sensitive assay for the functional activity of complement Factor I is described. This is based on its third proteolytic clip whereby Factor I cleaves cell-bound iC3b to cell-bound C3dg and soluble C3c, thereby abolishing conglutination of the cells. Factor H is required as a co-factor for Factor I activity. Because of the low affinity of iC3b for Factor H, the assay needs to be performed at low ionic strength. This assay is easier to perform than those based on the conversion of C3b to iC3b (the first two Factor I clips), there being no need for the unstable intermediate EAC142 or for purified C3.


Assuntos
Complemento C3b/metabolismo , Fator I do Complemento/metabolismo , Imunoensaio/métodos , Colectinas/análise , Colectinas/metabolismo , Fator H do Complemento/metabolismo , Fator I do Complemento/análise , Humanos , Fragmentos de Peptídeos/metabolismo , Peptídeo Hidrolases/metabolismo , Proteólise , Sensibilidade e Especificidade , Soroglobulinas/análise , Soroglobulinas/metabolismo
18.
Mol Vis ; 24: 75-82, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29410599

RESUMO

Purpose: A recent genome-wide association study by the International Age-related Macular Degeneration Genomics Consortium (IAMDGC) identified seven rare variants that are individually associated with age-related macular degeneration (AMD), the most common cause of vision loss in the elderly. In literature, several of these rare variants have been reported with different frequencies and odds ratios across populations of Europe and North America. Here, we aim to describe the representation of these seven AMD-associated rare variants in different geographic regions based on 24 AMD studies. Methods: We explored the occurrence of seven rare variants independently associated with AMD (CFH rs121913059 (p.Arg1210Cys), CFI rs141853578 (p.Gly119Arg), C3 rs147859257 (p.Lys155Gln), and C9 rs34882957 (p.Pro167Ser)) and three non-coding variants in or near the CFH gene (rs148553336, rs35292876, and rs191281603) in 24 AMD case-control studies. We studied the difference in distribution, interaction, and effect size for each of the rare variants based on the minor allele frequency within the different geographic regions. Results: We demonstrate that two rare AMD-associated variants in the CFH gene (rs121913059 [p.Arg1210Cys] and rs35292876) deviate in frequency among different geographic regions (p=0.004 and p=0.001, respectively). The risk estimates of each of the seven rare variants were comparable across the geographic regions. Conclusions: The results emphasize the importance of identifying population-specific rare variants, for example, by performing sequencing studies in case-control studies of various populations, because their identification may have implications for diagnostic screening and personalized treatment.


Assuntos
Fator H do Complemento/genética , Predisposição Genética para Doença , Degeneração Macular/genética , Polimorfismo de Nucleotídeo Único , Idoso , Alelos , Estudos de Casos e Controles , Complemento C3/genética , Complemento C3/imunologia , Complemento C9/genética , Complemento C9/imunologia , Fator H do Complemento/imunologia , Fator I do Complemento/genética , Fator I do Complemento/imunologia , Europa (Continente) , Feminino , Expressão Gênica , Frequência do Gene , Estudo de Associação Genômica Ampla , Geografia , Humanos , Degeneração Macular/diagnóstico , Degeneração Macular/imunologia , Degeneração Macular/patologia , Masculino
19.
Dev Comp Immunol ; 82: 66-71, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29329952

RESUMO

Complement factor I (CFI) is a novel regulatory serine protease that plays an important role in resistance to pathogen infection. In this study, the CFI gene of Pelteobagrus vachellii (Pv-CFI) was sequenced and characterized. The full-length cDNA of 2320 bp includes a 155 bp 5'-untranslated region (UTR), a 164 bp 3'-UTR, and a 2001 bp open reading frame (ORF) encoding a 667 amino acids. Multiple sequence alignment revealed five highly conserved domains with a typical modular architecture and identical active sites in vertebrates, indicating a conserved function. Pv-CFI mRNA was constitutively expressed in all examined tissues and most abundant in liver. During infection with Aeromonas hydrophila, Pv-CFI mRNA expression was significantly up-regulated in liver at 3-24 h, spleen at 3-48 h and head kidney at 3-48 h. The results suggest Pv-CFI plays an important role in resistance to pathogenic bacteria in P. vachellii.


Assuntos
Aeromonas hydrophila/fisiologia , Peixes-Gato/genética , Fator I do Complemento/genética , Proteínas de Peixes/genética , Infecções por Bactérias Gram-Negativas/imunologia , Fígado/fisiologia , Animais , Peixes-Gato/imunologia , Clonagem Molecular , Fator I do Complemento/metabolismo , DNA Complementar/genética , Proteínas de Peixes/metabolismo , Infecções por Bactérias Gram-Negativas/genética , Imunidade Inata , Filogenia , Alinhamento de Sequência
20.
Ocul Immunol Inflamm ; 26(1): 51-56, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-27380463

RESUMO

PURPOSE: To investigate the roles of CFI, genotype-phenotype associations were identified in AAU. METHODS: A case-control study was conducted in a total of 575 subjects consisting of 279 AAU patients and 296 healthy controls. Genotypic analyses were performed using Sequenom MassARRAY technology. Analyses were stratified to a series of clinical ophthalmic confounding factors. RESULTS: A lower frequency of the CFI-rs13104777 C allele was found in the AAU cohort compared with the controls, and, thus, was significantly associated with AAU pathogenesis (p = 0.041, OR = 0.712, 95% CI: 0.513-0.987). Stratified analysis also demonstrated the associations may differ depending on the HLA-B27 status and laterality status. CONCLUSIONS: This study has revealed a significant genetic role for CFI-rs13104777 in AAU. This influence may be dependent on human leukocyte antigen (HLA)-B27 and disease laterality. Overall, the results provide evidence for a pathogenic role for CFI in AAU and expand our knowledge on the genetic basis of AAU.


Assuntos
Fator I do Complemento/genética , Marcadores Genéticos/genética , Polimorfismo de Nucleotídeo Único , Uveíte Anterior/genética , Doença Aguda , Adulto , Estudos de Casos e Controles , Feminino , Frequência do Gene , Estudos de Associação Genética , Técnicas de Genotipagem , Antígeno HLA-B27/genética , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase
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