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1.
PLoS One ; 16(1): e0244855, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33507994

RESUMO

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the causative agent of the pandemic human respiratory illness COVID-19, is a global health emergency. While severe acute disease has been linked to an expansion of antibody-secreting plasmablasts, we sought to identify B cell responses that correlated with positive clinical outcomes in convalescent patients. We characterized the peripheral blood B cell immunophenotype and plasma antibody responses in 40 recovered non-hospitalized COVID-19 subjects that were enrolled as donors in a convalescent plasma treatment study. We observed a significant negative correlation between the frequency of peripheral blood memory B cells and the duration of symptoms for convalescent subjects. Memory B cell subsets in convalescent subjects were composed of classical CD24+ class-switched memory B cells, but also activated CD24-negative and natural unswitched CD27+ IgD+ IgM+ subsets. Memory B cell frequency was significantly correlated with both IgG1 and IgM responses to the SARS-CoV-2 spike protein receptor binding domain (RBD) in most seropositive subjects. IgM+ memory, but not switched memory, directly correlated with virus-specific antibody responses, and remained stable over 3 months. Our findings suggest that the frequency of memory B cells is a critical indicator of disease resolution, and that IgM+ memory B cells may play an important role in SARS-CoV-2 immunity.


Assuntos
Subpopulações de Linfócitos B/imunologia , /imunologia , Adulto , Idoso , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Formação de Anticorpos , Linfócitos B/imunologia , Convalescença , Progressão da Doença , Feminino , Humanos , Imunidade/imunologia , Imunoglobulina G/imunologia , Imunoglobulina M/metabolismo , Imunofenotipagem/métodos , Masculino , Pessoa de Meia-Idade , Recuperação de Função Fisiológica/imunologia , /isolamento & purificação
2.
Biol Direct ; 15(1): 21, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-33138856

RESUMO

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection spreaded rapidly worldwide, as far as it has become a global pandemic. Therefore, the introduction of serological tests for determination of IgM and IgG antibodies has become the main diagnostic tool, useful for tracking the spread of the virus and for consequently allowing its containment. In our study we compared point of care test (POCT) lateral flow immunoassay (FIA) vs automated chemiluminescent immunoassay (CLIA), in order to assess their specificity and sensibility for COVID-19 antibodies detection. RESULTS: We find that different specificities and sensitivities for IgM and IgG tests. Notably IgM POCT FIA method vs CLIA method (gold standard) has a low sensitivity (0.526), while IgG POCT FIA method vs CLIA method (gold standard) test has a much higher sensitivity (0.937); further, with respect of IgG, FIA and CLIA could arguably provide equivalent information. CONCLUSIONS: FIA method could be helpful in assessing in short time, the possible contagiousness of subjects that for work reasons cannot guarantee "social distancing".


Assuntos
Infecções por Coronavirus/sangue , Pneumonia Viral/sangue , Testes Sorológicos , Sequência de Aminoácidos , Feminino , Humanos , Imunoensaio , Imunoglobulina M/metabolismo , Medições Luminescentes , Masculino , Pessoa de Meia-Idade , Pandemias , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo
3.
PLoS Pathog ; 16(8): e1008793, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866189

RESUMO

Transmission to chimpanzees of a precore hepatitis B virus (HBV) mutant implicated in acute liver failure (ALF) in humans did not cause ALF nor the classic form of acute hepatitis B (AHB) seen upon infection with the wild-type HBV strain, but rather a severe AHB with distinct disease features. Here, we investigated the viral and host immunity factors responsible for the unusual severity of AHB associated with the precore HBV mutant in chimpanzees. Archived serial serum and liver specimens from two chimpanzees inoculated with a precore HBV mutant implicated in ALF and two chimpanzees inoculated with wild-type HBV were studied. We used phage-display library and next-generation sequencing (NGS) technologies to characterize the liver antibody response. The results obtained in severe AHB were compared with those in classic AHB and HBV-associated ALF in humans. Severe AHB was characterized by: (i) the highest alanine aminotransferase (ALT) peaks ever seen in HBV transmission studies with a significantly shorter incubation period, compared to classic AHB; (ii) earlier HBsAg clearance and anti-HBs seroconversion with transient or undetectable hepatitis B e antigen (HBeAg); (iii) limited inflammatory reaction relative to hepatocellular damage at the ALT peak with B-cell infiltration, albeit less extensive than in ALF; (iv) detection of intrahepatic germline antibodies against hepatitis B core antigen (HBcAg) by phage-display libraries in the earliest disease phase, as seen in ALF; (v) lack of intrahepatic IgM anti-HBcAg Fab, as seen in classic AHB, but at variance with ALF; and (vi) higher proportion of antibodies in germline configuration detected by NGS in the intrahepatic antibody repertoire compared to classic AHB, but lower than in ALF. This study identifies distinct outcome-specific features associated with severe AHB caused by a precore HBV mutant in chimpanzees, which bear closer resemblance to HBV ALF than to classic AHB. Our data suggest that precore HBV mutants carry an inherently higher pathogenicity that, in addition to specific host factors, may play a critical role in determining the severity of acute HBV disease.


Assuntos
Anticorpos Anti-Hepatite B/metabolismo , Vírus da Hepatite B/metabolismo , Hepatite B/metabolismo , Imunoglobulina M/metabolismo , Falência Hepática Aguda/metabolismo , Animais , Modelos Animais de Doenças , Hepatite B/patologia , Antígenos do Núcleo do Vírus da Hepatite B/metabolismo , Humanos , Falência Hepática Aguda/patologia , Pan troglodytes
4.
Int J Nanomedicine ; 15: 4079-4090, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606665

RESUMO

Purpose: The aim of this study is to develop efficient localized therapy of sertaconazole nitrate for the treatment of vaginal candidiasis. Methods: Sertaconazole nitrate-loaded cationic liposomes were prepared by thin-film hydration method and coated with different concentrations of pectin (0.05%, 0.1% and 0.2%) to develop mucoadhesive liposomes. The formulated mucoadhesive vesicles were characterized in terms of morphology, entrapment efficiency, particle size, zeta value, mucoadhesive properties and drug release. The selected formula was incorporated into a gel base and further characterized by an ex vivo permeation study in comparison with conventional sertaconazole gel. Also, the in vivo study was performed to assess the efficacy of sertaconazole mucoadhesive liposomal gel in treating rats with vaginal candidiasis. Results: The mucoadhesive liposomes were spherical. Coating liposomes with pectin results in increased entrapment efficiency and particle size compared with uncoated vesicles. On the contrary, zeta values were reduced upon coating liposomes with pectin indicating efficient coating of liposomes with pectin. Mucoadhesive liposomes showed a more prolonged and sustained drug release compared with uncoated liposomes. Ex vivo study results showed that mucoadhesive liposomal gel increased sertaconazole tissue retention and reduced drug tissue penetration. In the invivo study, the mucoadhesive liposomal gel showed a significant reduction in the microbial count with a subsequent reduction in inflammatory responses with the lowest histopathological change compared with conventional gel. Conclusion: The study confirmed the potentiality of employing mucoadhesive liposomes as a successful carrier for the vaginal delivery of antifungal drugs.


Assuntos
Antifúngicos/uso terapêutico , Candidíase Vulvovaginal/tratamento farmacológico , Imidazóis/uso terapêutico , Muco/química , Tiofenos/uso terapêutico , Adesividade , Animais , Anti-Infecciosos/farmacologia , Biomarcadores/metabolismo , Preparações de Ação Retardada , Liberação Controlada de Fármacos , Feminino , Géis , Humanos , Imidazóis/farmacologia , Imunoglobulina G/metabolismo , Imunoglobulina M/metabolismo , Mediadores da Inflamação/metabolismo , Lipossomos/ultraestrutura , Mucinas/metabolismo , Tamanho da Partícula , Ratos Sprague-Dawley , Ovinos , Eletricidade Estática , Tiofenos/farmacologia , Vagina/patologia , beta-Glucanas/metabolismo
5.
Nat Commun ; 11(1): 3510, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665616

RESUMO

We report Zika virus (ZIKV) vertical transmission in 130 infants born to PCR+ mothers at the time of the Rio de Janeiro epidemic of 2015-2016. Serum and urine collected from birth through the first year of life were tested by quantitative reverse transcriptase polymerase chain reaction (PCR) and/or IgM Zika MAC-ELISA. Four hundred and seven specimens are evaluated; 161 sera tested by PCR and IgM assays, 85 urines by PCR. Sixty-five percent of children (N = 84) are positive in at least one assay. Of 94 children tested within 3 months of age, 70% are positive. Positivity declines to 33% after 3 months. Five children are PCR+ beyond 200 days of life. Concordance between IgM and PCR results is 52%, sensitivity 65%, specificity 40% (positive PCR results as gold standard). IgM and serum PCR are 61% concordant; serum and urine PCR 55%. Most children (65%) are clinically normal. Equal numbers of children with abnormal findings (29 of 45, 64%) and normal findings (55 of 85, 65%) have positive results, p = 0.98. Earlier maternal trimester of infection is associated with positive results (p = 0.04) but not clinical disease (p = 0.98). ZIKV vertical transmission is frequent but laboratory confirmed infection is not necessarily associated with infant abnormalities.


Assuntos
Doenças Transmissíveis/transmissão , Doenças Transmissíveis/virologia , Infecção por Zika virus/transmissão , Infecção por Zika virus/virologia , Zika virus/patogenicidade , Feminino , Humanos , Imunoglobulina M/metabolismo , Reação em Cadeia da Polimerase , Gravidez , Viroses/virologia
6.
Lancet Infect Dis ; 20(9): e245-e249, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32687805

RESUMO

The collapse of global cooperation and a failure of international solidarity have led to many low-income and middle-income countries being denied access to molecular diagnostics in the COVID-19 pandemic response. Yet the scarcity of knowledge on the dynamics of the immune response to infection has led to hesitation on recommending the use of rapid immunodiagnostic tests, even though rapid serology tests are commercially available and scalable. On the basis of our knowledge and understanding of viral infectivity and host response, we urge countries without the capacity to do molecular testing at scale to research the use of serology tests to triage symptomatic patients in community settings, to test contacts of confirmed cases, and in situational analysis and surveillance. The WHO R&D Blue Print expert group identified eight priorities for research and development, of which the highest is to mobilise research on rapid point-of-care diagnostics for use at the community level. This research should inform control programmes of the required performance and utility of rapid serology tests, which, when applied specifically for appropriate public health measures to then be put in place, can make a huge difference.


Assuntos
Anticorpos Antivirais/metabolismo , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Anticorpos Antivirais/sangue , Betacoronavirus/genética , Betacoronavirus/imunologia , Busca de Comunicante/métodos , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/transmissão , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/metabolismo , Imunoglobulina M/sangue , Imunoglobulina M/metabolismo , Pandemias/prevenção & controle , Pneumonia Viral/imunologia , Pneumonia Viral/prevenção & controle , Pneumonia Viral/transmissão , RNA Viral/análise , Fatores de Tempo , Triagem/métodos , Eliminação de Partículas Virais
7.
APMIS ; 128(8): 487-496, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32562574

RESUMO

Metastatic thyroid cancers are more difficult to treat and have a significantly worse prognosis than localized thyroid cancers. Previous studies have shown that follicular helper T cells (Tfh) may participate in antitumor immune responses. Here, we investigated the characteristics of Tfh cells in patients with differentiated thyroid cancer (DTC) at various severities, including patients with localized disease, cervical metastasis, and distant metastasis. In circulating CD4 T cells, the proportion of CD4+ CXCR5+ Tfh-like cells was significantly higher in patients with distant metastasis than in healthy controls, patients with local disease, and patients with cervical metastasis. Also, the expression of Tfh cell-associated surface molecules, such as PD-1, ICOS, and BTLA, tended to be higher in patients with cervical and distant metastasis than in healthy controls. However, the expression of secreted molecules, such as IL-10, IL-21, and CXCL13, was significantly lower in patients with distant metastasis than in healthy controls and patients with local disease. Additionally, circulating Tfh-like cells from patients with distant metastasis were less capable of supporting B-cell growth and IgM secretion. We also examined the CD4+ CXCR5+ Tfh-like cells in tumor samples. Tumor-infiltrating Tfh-like cells were highly enriched in the pulmonary metastasis compared to the local tumor and the cervical metastasis. However, tumor-infiltrating Tfh-like cells from pulmonary metastasis displayed higher PD-1, TIM-3, and lower IL-21 expression than those from the local tumor. Together, this study identified that the metastasis of DTC patients was associated with an overabundance of defective Tfh cells.


Assuntos
Neoplasias Ósseas/patologia , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias Pulmonares/patologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Neoplasias da Glândula Tireoide/patologia , Adulto , Idoso , Antígenos de Superfície/análise , Linfócitos B/imunologia , Neoplasias Ósseas/secundário , Proliferação de Células , Feminino , Neoplasias de Cabeça e Pescoço/secundário , Humanos , Imunoglobulina M/metabolismo , Fatores Imunológicos/metabolismo , Neoplasias Pulmonares/secundário , Masculino , Pessoa de Meia-Idade
8.
Clin Microbiol Infect ; 26(8): 1082-1087, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32473953

RESUMO

OBJECTIVES: To evaluate the diagnostic performance of seven rapid IgG/IgM tests and the Euroimmun IgA/IgG ELISA for antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in COVID-19 patients. METHODS: Specificity was evaluated in 103 samples collected before January 2020. Sensitivity and time to seropositivity was evaluated in 167 samples from 94 patients with COVID-19 confirmed with RT-PCR on nasopharyngeal swab. RESULTS: Specificity (confidence interval) of lateral flow assays (LFAs) was ≥91.3% (84.0-95.5) for IgM, ≥90.3% (82.9-94.8) for IgG, and ≥85.4% (77.2-91.1) for the combination IgM OR IgG. Specificity of the ELISA was 96.1% (90.1-98.8) for IgG and only 73.8% (64.5-81.4) for IgA. Sensitivity 14-25 days after the onset of symptoms was between ≥92.1% (78.5-98.0) and 100% (95.7-100) for IgG LFA compared to 89.5% (75.3-96.4) for IgG ELISA. Positivity of IgM OR IgG for LFA resulted in a decrease in specificity compared to IgG alone without a gain in diagnostic performance, except for VivaDiag. The results for IgM varied significantly between the LFAs with an average overall agreement of only 70% compared to 89% for IgG. The average dynamic trend to seropositivity for IgM was not shorter than for IgG. At the time of hospital admission the sensitivity of LFA was <60%. CONCLUSIONS: Sensitivity for the detection of IgG antibodies 14-25 days after the onset of symptoms was ≥92.1% for all seven LFAs compared to 89.5% for the IgG ELISA. The results for IgM varied significantly, and including IgM antibodies in addition to IgG for the interpretation of LFAs did not improve the diagnostic performance.


Assuntos
Anticorpos Antivirais/análise , Antígenos Virais/imunologia , Betacoronavirus/imunologia , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Infecções por Coronavirus/imunologia , Testes Diagnósticos de Rotina , Feminino , Humanos , Imunoglobulina A/metabolismo , Imunoglobulina G/metabolismo , Imunoglobulina M/metabolismo , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/imunologia , Sensibilidade e Especificidade , Fatores de Tempo , Adulto Jovem
9.
PLoS One ; 15(5): e0232739, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32437393

RESUMO

OBJECTIVES: Systemic sclerosis (SSc) is an auto-immune, multi organ disease marked by severe gastrointestinal (GI) involvement and gut dysbiosis. Here, we aimed to determine the safety and efficacy of fecal microbiota transplantation (FMT) using commercially-available anaerobic cultivated human intestinal microbiota (ACHIM) in SSc. METHODS: Ten patients with SSc were randomized to ACHIM (n = 5) or placebo (n = 5) in a double-blind, placebo-controlled 16-week pilot. All patients had mild to severe upper and lower GI symptoms including diarrhea, distention/bloating and/or fecal incontinence at baseline. Gastroduodenoscopy transfer of ACHIM or placebo was performed at weeks 0 and 2. Primary endpoints were safety and clinical efficacy on GI symptoms assessed at weeks 4 and 16. Secondary endpoints included changes in relative abundance of total, immunoglobulin (Ig) A- and IgM-coated fecal bacteria measured by 16s rRNA sequencing. RESULTS: ACHIM side effects were mild and transient. Two placebo controls experienced procedure-related serious adverse events; one developed laryngospasms at week 0 gastroduodenoscopy necessitating study exclusion whilst one encountered duodenal perforation during gastroduodenoscopy at the last study visit (week 16). Decreased bloating, diarrhea and/or fecal incontinence was observed in four of five patients in the FMT group (week 4 or/and 16) and in two of four in the placebo group (week 4 or 16). Relative abundance, richness and diversity of total and IgA-coated and IgM-coated bacteria fluctuated more after FMT, than after placebo. CONCLUSIONS: FMT of commercially-available ACHIM is associated with gastroduodenoscopy complications but reduces lower GI symptoms by possibly altering the gut microbiota in patients with SSc.


Assuntos
Transplante de Microbiota Fecal , Escleroderma Sistêmico/microbiologia , Escleroderma Sistêmico/terapia , Bactérias , Método Duplo-Cego , Ácidos Graxos/metabolismo , Incontinência Fecal/etiologia , Transplante de Microbiota Fecal/efeitos adversos , Fezes/química , Feminino , Humanos , Imunoglobulina A/metabolismo , Imunoglobulina M/metabolismo , Complexo Antígeno L1 Leucocitário/metabolismo , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Placebos , Resultado do Tratamento
11.
Allergy ; 75(7): 1546-1554, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32329930
12.
Phytomedicine ; 69: 153194, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32146299

RESUMO

BACKGROUND: The frequency of allergic diseases is constantly rising. Dysregulated production of isotype E immunoglobulins is one of the key factors behind allergic reactions and its modulation is therefore an important target for pharmacological intervention. Natural products of the pseurotin family were reported to be inhibitors of IgE production in B-cells. Mechanistic details underlying these effects are however not well understood. PURPOSE: In the present study, we synthesized new analogs of natural pseurotins and extensively investigated their inhibitory effects on activation, proliferation and differentiation of B-cells, as well as on the production of IgE. STUDY DESIGN: Effects of two natural pseurotins (pseurotins A and D) and a collection of fully synthetic pseurotin analogs were studied on mouse B-cells stimulated by the combination of IL-4 and E. coli lipopolysaccharide. The IgE production was determined along with cell viability and cell proliferation. The phosphorylation of selected members of the STAT transcription factor family was subsequently investigated. Finally, the in vivo effect of pseurotin D on the ovalbumin-induced delayed type hypersensitivity response was tested in mice. RESULTS: We discovered that several fully synthetic pseurotin analogs were able to decrease the production of IgE in stimulated B-cells with potency comparable to that of pseurotins A and D. We found that the two natural pseurotins and the active synthetic analogs inhibited the phosphorylation of STAT3, STAT5 and STAT6 proteins in stimulated B-cells, resulting in the inhibition of B-cell proliferation and differentiation into the plasma cells. In vivo, pseurotin D decreased ovalbumin-induced foot pad edema. CONCLUSION: Our results advance the current mechanistic understanding of the pseurotin-induced inhibition of IgE production in B-cells by linking the effect to STAT signaling, and associated modulation of B-cell proliferation and differentiation. Together with our finding that structurally simpler pseurotin analogs were able to reproduce the effects of natural pseurotins, the presented work has implications for the future research on these secondary metabolites in the context of allergic diseases.


Assuntos
Linfócitos B/efeitos dos fármacos , Imunoglobulina E/metabolismo , Plasmócitos/citologia , Pirrolidinonas/química , Pirrolidinonas/farmacologia , Animais , Linfócitos B/citologia , Linfócitos B/fisiologia , Diferenciação Celular/efeitos dos fármacos , Edema/induzido quimicamente , Edema/tratamento farmacológico , Escherichia coli/química , Imunoglobulina E/sangue , Imunoglobulina M/sangue , Imunoglobulina M/metabolismo , Lipopolissacarídeos/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Ovalbumina/toxicidade , Fosforilação/efeitos dos fármacos , Plasmócitos/fisiologia , Fatores de Transcrição STAT/metabolismo
13.
PLoS One ; 15(3): e0229992, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32163462

RESUMO

Recombinant production of IgM antibodies poses a special challenge due to the complex structure of the proteins and their not yet fully elucidated interactions with the immune effector proteins, especially the complement system. In this study, we present transient expression of IgM antibodies (IgM617, IgM012 and IgM012_GL) in HEK cells and compared it to the well-established stable expression system in CHO cells. The presented workflow investigates quality attributes including productivity, polymer distribution, glycosylation, antibody structure and activation of the classical complement pathway. The HEK293E transient expression system is able to generate comparable amounts and polymer distribution as IgM stably produced in CHO. Although the glycan profile generated by HEK293E cells contained a lower degree of sialylation and a higher portion of oligomannose structures, the potency to activate the complement cascade was maintained. Electron microscopy also confirmed the structural integrity of IgM pentamers produced in HEK293E cells, since the conventional star-shaped structure is observed. From our studies, we conclude that the transient expression system provides an attractive alternative for rapid, efficient and high-throughput production of complex IgM antibodies with slightly altered post-translational modifications, but comparable structure and function.


Assuntos
Imunoglobulina M/metabolismo , Animais , Células CHO , Ativação do Complemento , Complemento C1q/química , Complemento C1q/metabolismo , Cricetinae , Cricetulus , Glicosilação , Células HEK293 , Humanos , Imunoglobulina M/química , Imunoglobulina M/genética , Microscopia Eletrônica de Transmissão , Oligossacarídeos/química , Plasmídeos/genética , Plasmídeos/metabolismo , Transfecção
14.
PLoS One ; 15(3): e0229080, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32196507

RESUMO

Therapeutic monoclonal antibodies have the potential to work as biological therapeutics. OKT3, Herceptin, Keytruda and others have positively impacted healthcare. Antibodies evolved naturally to provide high specificity and high affinity once mature. These characteristics can make them useful as therapeutics. However, we may be missing characteristics that are not obvious. We present a means of measuring antibodies in an unbiased manner that may highlight therapeutic activity. We propose using a microarray of random peptides to assess antibody properties. We tested twenty-four different commercial antibodies to gain some perspective about how much information can be derived from binding antibodies to random peptide libraries. Some monoclonals preferred to bind shorter peptides, some longer, some preferred motifs closer to the C-term, some nearer the N-term. We tested some antibodies with clinical activity but whose function was blinded to us at the time. We were provided with twenty-one different monoclonal antibodies, thirteen mouse and eight human IgM. These antibodies produced a variety of binding patterns on the random peptide arrays. When unblinded, the antibodies with polyspecific binding were the ones with the greatest therapeutic activity. The protein target to these therapeutic monoclonals is still unknown but using common sequence motifs from the peptides we predicted several human and mouse proteins. The same five highest proteins appeared in both mouse and human lists.


Assuntos
Anticorpos Monoclonais/análise , Mapeamento de Epitopos/métodos , Biblioteca de Peptídeos , Análise Serial de Proteínas , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Monoclonais/metabolismo , Especificidade de Anticorpos , Ensaios de Triagem em Larga Escala/métodos , Humanos , Imunoglobulina M/análise , Imunoglobulina M/metabolismo , Camundongos , Peptídeos/química , Peptídeos/imunologia , Peptídeos/metabolismo , Análise Serial de Proteínas/métodos , Ligação Proteica , Proteoma/análise
15.
Mol Immunol ; 121: 99-110, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32199212

RESUMO

The complement cascade consists of cell bound and serum proteins acting together to protect the host from pathogens, remove cancerous cells and effectively links innate and adaptive immune responses. Despite its usefulness in microbial neutralization and clearance of cancerous cells, excessive complement activation causes an immune imbalance and tissue damage in the host. Hence, a series of complement regulatory proteins present at a higher concentration in blood plasma and on cell surfaces tightly regulate the cascade. The complement cascade can be initiated by B-1 B cell production of natural antibodies. Natural antibodies arise spontaneously without any known exogenous antigenic or microbial stimulus and protect against invading pathogens, clear apoptotic cells, provide tissue homeostasis, and modulate adaptive immune functions. Natural IgM antibodies recognize microbial and cancer antigens and serve as an activator of complement mediated lysis. This review will discuss advances in complement activation and regulation in bacterial and viral infections, and cancer. We will also explore the crosstalk of natural antibodies with bacterial populations and cancer.


Assuntos
Infecções Bacterianas/imunologia , Imunidade Humoral , Imunidade Inata , Neoplasias/imunologia , Viroses/imunologia , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/metabolismo , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Antígenos Virais/imunologia , Antígenos Virais/metabolismo , Apoptose/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Ativação do Complemento , Proteínas do Sistema Complemento/imunologia , Proteínas do Sistema Complemento/metabolismo , Humanos , Imunoglobulina M/imunologia , Imunoglobulina M/metabolismo , Receptores de Complemento/imunologia , Receptores de Complemento/metabolismo , Evasão Tumoral
16.
Transfusion ; 60(3): 598-606, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31957888

RESUMO

BACKGROUND: Recent advances in desensitization techniques and immunosuppressive therapy have led to improved outcomes after ABO-incompatible (ABO-i) kidney transplantation (KT). However, questions remain unanswered, particularly regarding which type of ABO isoagglutinin-immunoglobulin M (IgM) or immunoglobulin M (IgG)-is significantly involved in antibody-mediated rejection (AMR). STUDY DESIGN AND METHODS: We retrospectively analyzed data from 120 patients who underwent ABO-i KT between 2012 and 2014. Preoperative plasma exchange was performed until the IgM isoagglutinin titer was 4 or less, regardless of the IgG titer. Clinical data were compared between patient groups with pre-KT IgG isoagglutinin titer 16 or greater (high IgG; titer range, 16-256; n = 39) and 8 or less (low IgG; titer range, -8; n = 81). RESULTS: The median follow-up periods were 59 (high IgG) and 55 (low IgG) months. Patient survival at 5 years (p = 0.314) was 100% (high IgG) and 97.4% (low IgG). Graft survival at 5 years (p = 0.480) was 100% (high IgG) and 98.7% (low IgG). AMR by anti-ABO antibody occurred in only one patient in the low-IgG group. CONCLUSION: Patients with high pre-KT IgG isoagglutinin titers had equally successful outcomes as those with low IgG titers. ABO-i KT can be successfully performed by reducing the pre-KT IgM isoagglutinin titer to 4 or less, as determined by the immediate spin tube method.


Assuntos
Sistema ABO de Grupos Sanguíneos/metabolismo , Imunoglobulina M/metabolismo , Adolescente , Adulto , Idoso , Incompatibilidade de Grupos Sanguíneos/metabolismo , Citometria de Fluxo , Humanos , Imunossupressão/métodos , Transplante de Rim , Pessoa de Meia-Idade , Estudos Retrospectivos , Adulto Jovem
17.
Transfusion ; 60(3): 628-636, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31957889

RESUMO

BACKGROUND: Isohemagglutinins (anti-A and anti-B) mediate hemolytic transfusion reactions, antibody-mediated rejection of solid-organ transplants, and delayed engraftment after stem cell transplant. However, quantification of isohemagglutinins is often labor intensive and operator dependent, limiting availability and interfacility comparisons. We evaluated an automated, solid-phase and agglutination-based antibody titer platform versus manual gel testing. STUDY DESIGN AND METHODS: Plasma samples were obtained from 54 randomly selected patients. Titers were determined by our laboratory's standard assay (manual dilution followed by manual gel testing) and were compared to results obtained on a fully automated blood bank analyzer (Galileo NEO, Immucor). The analyzer determined immunoglobulin G (IgG) antibodies using solid-phase and immunoglobulin M (IgM) antibodies by direct hemagglutination. RESULTS: Isohemagglutinin titers obtained by manual gel versus the automated assay generally (>80%) agreed within one doubling dilution, and always (100%) agreed within two dilutions. Among O samples, the gel titer and the highest titer obtained with the automated assay (either IgG or IgM) were similar in paired, nonparametric analysis (p = 0.06 for anti-A; p = 0.13 for anti-B). Gel titers from group A and group B patients were slightly higher than the highest titer obtained using the automated assay (p = 0.04 for group A; p = 0.009 for group B), although these differences were within the accepted error of measurement. CONCLUSION: Manual and automated methodologies yielded similar isohemagglutinin titers. Separate quantification of IgM and IgG isohemagglutinins via automated titration may yield additional insight into hemolysis, graft survival after ABO-incompatible transplantation, and red blood cell engraftment after ABO-incompatible stem cell transplant.


Assuntos
Hemaglutininas/metabolismo , Sistema ABO de Grupos Sanguíneos/imunologia , Sistema ABO de Grupos Sanguíneos/metabolismo , Incompatibilidade de Grupos Sanguíneos/imunologia , Sobrevivência de Enxerto , Hemaglutininas/imunologia , Humanos , Imunoglobulina G/metabolismo , Imunoglobulina M/metabolismo
18.
Infect Immun ; 88(4)2020 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-31988178

RESUMO

Human studies have shown associations between cryptococcal meningitis and reduced IgM memory B cell levels, and studies in IgM- and/or B cell-deficient mice have demonstrated increased Cryptococcus neoformans dissemination from lungs to brain. Since immunoglobulins are part of the immune milieu that C. neoformans confronts in a human host, and its ability to form titan cells is an important virulence mechanism, we determined the effect of human immunoglobulins on C. neoformans titan cell formation in vitro (i) Fluorescence microscopy showed normal human IgG and IgM bind C. neoformans (ii) C. neoformans grown in titan cell-inducing medium with IgM, not IgG, inhibited titan-like cell formation. (iii) Absorption of IgM with laminarin or curdlan (branched and linear 1-3-beta-d-glucans, respectively) decreased this effect. (iv) Transmission electron microscopy revealed that cells grown with IgM had small capsules and unique features not seen with cells grown with IgG. (v) Comparative transcriptional analysis of cell wall, capsule, and stress response genes showed that C. neoformans grown with IgM, not IgG or phosphate-buffered saline (PBS), had decreased expression of chitin synthetase, CHS1, CHS2, and CHS8, and genes encoding cell wall carbohydrate synthetases α-1-3-glucan (AGS1) and ß-1,3-glucan (FKS1). IgM also decreased expression of RIM101 and HOG1, genes encoding central regulators of C. neoformans stress response pathways and cell morphogenesis. Our data show human IgM affects C. neoformans morphology in vitro and suggest that the hypothesis that human immunoglobulins may affect C. neoformans virulence in vivo warrants further investigation.


Assuntos
Cryptococcus neoformans/efeitos dos fármacos , Cryptococcus neoformans/crescimento & desenvolvimento , Interações Hospedeiro-Patógeno , Imunoglobulina M/metabolismo , Fatores Imunológicos/metabolismo , Cryptococcus neoformans/citologia , Humanos , Imunoglobulina G/metabolismo , Virulência/efeitos dos fármacos
19.
Transfusion ; 60(3): 488-497, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31951028

RESUMO

BACKGROUND: Single antigen bead (SAB) assays are used to identify human leukocyte antigen (HLA) antibodies in patients with platelet refractoriness due to HLA Class I alloimmunization. Some laboratories use serum pretreatment regimens to eliminate interference from immunoglobulin M antibodies and complement. These modifications may contribute to interlaboratory variability, which is a recognized problem with the SAB assay. STUDY DESIGN AND METHODS: Five patients' sera were overnight shipped to 12 laboratories in the United States and internationally. Recipients used their lab's SAB procedure to identify HLA Class I antibodies. The resultant mean fluorescence intensity (MFI) data were compared by instrumentation, bead lot, and pretreatment regimens. Laboratory-specific cutoffs for positive antibodies were applied to the results. RESULTS: Interlaboratory variability for MFI values appears to be associated with different pretreatment regimens. The coefficient of variation (CV) of MFI from samples pretreated with ethylenediaminetetraacetic acid, dithiothreitol, or heat inactivation (EDHI) were similar, ranging from 14% to 56% (mean, 22%). For samples with no pretreatment, the CVs were significantly higher than EDHI-treated samples, ranging from 25% to 74% (mean, 39%; 95% confidence interval, 12.10-21.90; p < 0.0001). An intralaboratory comparison of pretreatment regimens confirmed these findings. Some positive antibody specificities present in EDHI-treated samples were negative in corresponding samples with no pretreatment when laboratory-specific cutoffs for positive antibodies were applied. CONCLUSION: Our results show that greater interlaboratory precision can be achieved when samples are pretreated with EDHI as opposed to no pretreatment, likely because these pretreatments eliminate interference from inhibitors. Inhibitors may mask antibodies, leading to missed (or uncalled) specificities when no pretreatment is used.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Isoanticorpos/imunologia , Especificidade de Anticorpos , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Ditiotreitol/farmacologia , Ácido Edético/farmacologia , Feminino , Antígenos HLA/metabolismo , Teste de Histocompatibilidade , Humanos , Imunoglobulina M/metabolismo , Masculino
20.
Muscle Nerve ; 61(4): 512-515, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31650561

RESUMO

INTRODUCTION: Causes of small-fiber peripheral neuropathies (SFN) are often undefined. In this study we investigated associations of serum autoantibodies, immunoglobulin G (IgG) vs fibroblast growth factor receptor-3 (FGFR-3), and immunoglobulin M (IgM) vs trisulfated heparan disaccharide (TS-HDS) in cryptogenic SFN. METHODS: One hundred fifty-five patients with biopsy-proven SFN and no identified cause for their neuropathy were blindly tested for serum IgM vs TS-HDS and IgG vs FGFR-3. RESULTS: Forty-eight percent of SFN patients had serum antibodies, 37% with IgM vs TS-HDS and 15% with IgG vs FGFR-3. TS-HDS antibodies were more frequent in SFN patients than in controls (P = .0012). Both antibodies were more common in females, and with non-length-dependent nerve pathology. Nintey-two percent of patients with acute-onset SFN had serum IgM vs TS-HDS. DISCUSSION: Autoantibodies directed against TS-HDS and FGFR-3 suggest an immune disorder in otherwise idiopathic SFN. Serum IgM vs TS-HDS may be a marker for SFN with an acute onset.


Assuntos
Autoanticorpos/imunologia , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/metabolismo , Neuropatia de Pequenas Fibras/imunologia , Feminino , Humanos , Imunoglobulina G/metabolismo , Imunoglobulina M/metabolismo , Masculino , Neuropatia de Pequenas Fibras/metabolismo
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