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1.
Med Sci Monit ; 26: e927444, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33320844

RESUMO

BACKGROUND Alpha-fetoprotein (AFP) is widely used to screen for hepatocellular carcinoma (HCC). However, the use of this biomarker has been challenged due to its low sensitivity and high rate of false negatives. In this study, we evaluated the diagnostic capability of cyclin D2 (CCND2) promoter methylation in patients with HCC related to hepatitis B virus (HBV). MATERIAL AND METHODS Using methylation-specific PCR and quantitative real-time PCR, we measured methylation status and mRNA levels of CCND2 in plasma and peripheral blood mononuclear cells (PBMCs) from 275 subjects: 75 patients with chronic hepatitis B (CHB), 47 with liver cirrhosis (LC), 118 with HCC, and 35 healthy controls (HCs). RESULTS The methylation rate of the CCND2 promoter was significantly higher in HCC patients than in patients without HCC (P<0.001). Furthermore, advanced HCC (TNM III/IV) was associated with a significantly higher frequency of CCND2 methylation and lower CCND2 mRNA levels than early-stage disease (TNM I/II; P<0.05). Combined measurement of CCND2 methylation and AFP yielded significantly higher sensitivity and area under the curve (AUC) than AFP alone in distinguishing patients with HCC from subjects with LC and CHB (P<0.001). CONCLUSIONS CCND2 methylation may be useful for predicting HCC progression. In addition, combined measurement of CCND2 methylation and AFP could serve as a non-invasive diagnostic marker for patients with HBV-related HCC.


Assuntos
Carcinoma Hepatocelular , Ciclina D2/genética , Metilação de DNA , Hepatite B Crônica , Cirrose Hepática , Neoplasias Hepáticas , alfa-Fetoproteínas/análise , Área Sob a Curva , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Diagnóstico Diferencial , Progressão da Doença , Feminino , Hepatite B Crônica/sangue , Hepatite B Crônica/diagnóstico , Humanos , Leucócitos Mononucleares/patologia , Cirrose Hepática/sangue , Cirrose Hepática/diagnóstico , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Regiões Promotoras Genéticas , RNA Mensageiro/análise , Sensibilidade e Especificidade
2.
PLoS One ; 15(11): e0233326, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33156824

RESUMO

BACKGROUND: Diabetes is an important risk factor for developing tuberculosis. This association leads to exacerbation of tuberculosis symptoms and delayed treatment of both the diseases. Molecular mechanism and biomarkers/drug targets related to copathogenesis of tuberculosis and diabetes are still poorly understood. In this study, proteomics based 2D-MALDI/MS approach was employed to identify host signature proteins which are altered during copathogenesis of tuberculosis and diabetes. METHODS: Comparative proteome of human peripheral blood mononuclear cells (PBMCs) from healthy controls, tuberculosis and diabetes patients in comparison to comorbid diabetes and tuberculosis patients was analyzed. Gel based proteomics approach followed by in gel trypsin digestion and peptide identification by mass spectrometry was used for signature protein identification. RESULTS: Total of 18 protein spots with differential expression in tuberculosis and diabetes copathogenesis (TBDM) patients in comparison to other groups were identified. These proteins belonged to four functional categories i.e. structural, cell cycle/growth regulation, signaling and intermediary metabolism. These include Vimentin, tubulin beta chain protein, Actin related protein 2/3 complex subunit 2, coffilin 1 (Structural), PDZ LIM domain protein, Rho-GDP dissociation inhibitor, Ras related protein Rab (signaling), superoxide dismutase, dCTPpyrophosphatase 1, Transcription initiation factor TFIID subunit 12, three isoforms of Peptidylprolylcis-trans isomerase A, SH3 domain containing protein (metabolism), three isoforms of Protein S100A9 and S100A8 (cell cycle progression/growth regulation). CONCLUSION: Proteins identified to be differentially expressed in TBDM patient can act as potent biomarkers and as predictors for copathogenesis of tuberculosis and diabetes.


Assuntos
Biomarcadores/metabolismo , Diabetes Mellitus/patologia , Leucócitos Mononucleares/patologia , Proteoma/metabolismo , Tuberculose/patologia , Biomarcadores/análise , Estudos de Casos e Controles , Diabetes Mellitus/metabolismo , Feminino , Humanos , Leucócitos Mononucleares/metabolismo , Pessoa de Meia-Idade , Projetos Piloto , Proteoma/análise , Tuberculose/metabolismo
3.
J Vis Exp ; (163)2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-33044453

RESUMO

Critical limb ischemia (CLI) represents an advanced stage of the peripheral arterial disease. Angioplasty improves the blood flow to the lower limb; however, some patients irreversibly progress to limb amputation. The extent of vascular damage and the mechanisms of vascular repair are factors affecting post-angioplasty outcome. Mononuclear Progenitor Cells (MPCs) are reactive to vascular damage and repair, with the ability to reflect vascular diseases. The present protocol describes quantification of MPCs obtained from blood circulation from vessel close to the angioplasty site, as well as its relationship with endothelial dysfunction and its predictive ability for limb amputation in the next 30 days after angioplasty in patients with CLI.


Assuntos
Amputação , Angioplastia , Isquemia/sangue , Leucócitos Mononucleares/patologia , Extremidade Inferior/irrigação sanguínea , Células-Tronco/patologia , Idoso , Amputação/efeitos adversos , Coleta de Amostras Sanguíneas , Endotélio/patologia , Endotélio/fisiopatologia , Feminino , Hemodinâmica , Humanos , Isquemia/etiologia , Isquemia/fisiopatologia , Extremidade Inferior/fisiopatologia , Masculino , Prognóstico , Fatores de Risco , Fatores de Tempo , Resultado do Tratamento
4.
PLoS One ; 15(10): e0240108, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33002096

RESUMO

Dose assessment is an important issue for radiation emergency medicine to determine appropriate clinical treatment. Hematopoietic tissues are extremely vulnerable to radiation exposure. A decrease in blood cell count following radiation exposure is the first quantitative bio-indicator using hematological techniques. We further examined induction of oxidative stress biomarkers in residual lymphocytes to identify new biomarkers for dosimetry. In vivo whole-body radiation to mice exposed to 5 Gy significantly induces DNA double-strand breaks, which were visualized by γ-H2AX in mouse blood cells. Mouse blood smears and peripheral blood mononuclear cells (PBMC) isolated from irradiated mice were used for immunostaining for oxidative biomarkers, parkin or Nrf2. Parkin is the E3 ubiquitin ligase, which is normally localized in the cytoplasm, is relocated to abnormal mitochondria with low membrane potential (ΔΨm), where it promotes clearance via mitophagy. Nrf2 transcription factor controls the major cellular antioxidant responses. Both markers of oxidative stress were more sensitive and persistent over time than nuclear DNA damage. In conclusion, parkin and Nrf2 are potential biomarkers for use in radiation dosimetry. Identification of several biological markers which show different kinetics for radiation response is essential for radiation dosimetry that allows the assessment of radiation injury and efficacy of clinical treatment in emergency radiation incidents. Radiation-induced oxidative damage is useful not only for radiation dose assessment but also for evaluation of radiation risks on humans.


Assuntos
Leucócitos Mononucleares/efeitos da radiação , Fator 2 Relacionado a NF-E2/análise , Estresse Oxidativo/efeitos da radiação , Ubiquitina-Proteína Ligases/análise , Irradiação Corporal Total/efeitos adversos , Animais , Biomarcadores/análise , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Leucócitos Mononucleares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
5.
PLoS One ; 15(10): e0240269, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33007040

RESUMO

OBJECTIVE: It is increasingly common to find patients affected by a combination of type 2 diabetes mellitus (T2DM), dyslipidemia (DLP) and periodontitis (PD), which are chronic inflammatory diseases. More studies able to capture unknown relationships among these diseases will contribute to raise biological and clinical evidence. The aim of this study was to apply association rule mining (ARM) to discover whether there are consistent patterns of clinical features (CFs) and differentially expressed genes (DEGs) relevant to these diseases. We intend to reinforce the evidence of the T2DM-DLP-PD-interplay and demonstrate the ARM ability to provide new insights into multivariate pattern discovery. METHODS: We utilized 29 clinical glycemic, lipid and periodontal parameters from 143 patients divided into five groups based upon diabetic, dyslipidemic and periodontal conditions (including a healthy-control group). At least 5 patients from each group were selected to assess the transcriptome by microarray. ARM was utilized to assess relevant association rules considering: (i) only CFs; and (ii) CFs+DEGs, such that the identified DEGs, specific to each group of patients, were submitted to gene expression validation by quantitative polymerase chain reaction (qPCR). RESULTS: We obtained 78 CF-rules and 161 CF+DEG-rules. Based on their clinical significance, Periodontists and Geneticist experts selected 11 CF-rules, and 5 CF+DEG-rules. From the five DEGs prospected by the rules, four of them were validated by qPCR as significantly different from the control group; and two of them validated the previous microarray findings. CONCLUSIONS: ARM was a powerful data analysis technique to identify multivariate patterns involving clinical and molecular profiles of patients affected by specific pathological panels. ARM proved to be an effective mining approach to analyze gene expression with the advantage of including patient's CFs. A combination of CFs and DEGs might be employed in modeling the patient's chance to develop complex diseases, such as those studied here.


Assuntos
Biologia Computacional/métodos , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patologia , Adulto , Mineração de Dados , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Inflamação/genética , Inflamação/patologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Reação em Cadeia da Polimerase em Tempo Real
6.
Sci Adv ; 6(31)2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32937590

RESUMO

The outbreak of the highly contagious and deadly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), also known as coronavirus disease 2019 (COVID-19), has posed a serious threat to public health across the globe, calling for the development of effective diagnostic markers and therapeutics. Here, we report a highly reliable severity diagnostic biomarker, acetylated 676th lysine transforming growth factor-beta-induced protein (TGFBIp K676Ac). TGFBIp K676Ac was consistently elevated in the blood of patients with SARS-CoV-2 pneumonia (n = 113), especially in patients in the intensive care unit (ICU) compared to non-ICU patients. Patients' blood samples showed increased cytokines and lymphopenia, which are exemplary indicators of SARS-CoV-2 pneumonia. Treatment with TGFBIp neutralizing antibodies suppressed the cytokine storm. The increased level of TGFBIp K676Ac in ICU patients suggests the promise of this protein as a reliable severity diagnostic biomarker for severe SARS-CoV-2 disease.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/diagnóstico , Síndrome da Liberação de Citocina/diagnóstico , Proteínas da Matriz Extracelular/imunologia , Leucócitos Mononucleares/imunologia , Pneumonia Viral/diagnóstico , Processamento de Proteína Pós-Traducional , Insuficiência Respiratória/diagnóstico , Fator de Crescimento Transformador beta/imunologia , Acetilação , Anticorpos Neutralizantes/farmacologia , Betacoronavirus/imunologia , Biomarcadores/sangue , Estudos de Casos e Controles , Infecções por Coronavirus/sangue , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/patologia , Síndrome da Liberação de Citocina/sangue , Síndrome da Liberação de Citocina/imunologia , Síndrome da Liberação de Citocina/patologia , Proteínas da Matriz Extracelular/antagonistas & inibidores , Proteínas da Matriz Extracelular/genética , Expressão Gênica , Humanos , Unidades de Terapia Intensiva , Contagem de Leucócitos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/patologia , Leucócitos Mononucleares/virologia , Pulmão/irrigação sanguínea , Pulmão/efeitos dos fármacos , Pulmão/patologia , Pulmão/virologia , Lisina/metabolismo , NF-kappa B/genética , NF-kappa B/imunologia , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/imunologia , Pneumonia Viral/patologia , Cultura Primária de Células , Prognóstico , Insuficiência Respiratória/sangue , Insuficiência Respiratória/imunologia , Insuficiência Respiratória/patologia , Índice de Gravidade de Doença , Fator de Crescimento Transformador beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/genética
7.
PLoS One ; 15(9): e0237496, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32881912

RESUMO

BACKGROUND: Obesity is often associated with inflammation in adipose tissue (AT) with release of mediators of atherogenesis. We postulated that it would be feasible to collect sufficient abdominal AT to quantify changes in a broad array of adaptive and innate mononuclear white cells in obese non-diabetic adults in response to a dipeptidyl protease inhibitor (DPP4i), known to inhibit activation of immune white cells. METHODS: Adults 18-55 years-of-age were screened for abdominal obesity and insulin resistance or impaired glucose tolerance but without known inflammatory conditions. Twenty-one eligible participants consented for study and were randomized 3:1 to receive sitagliptin (DPP4i) at 100mg or matching placebo daily for 28 days. Abdominal AT collected by percutaneous biopsy and peripheral blood mononuclear cell fractions were evaluated before and after treatment; plasma was stored for batch testing. RESULTS: Highly sensitive C-reactive protein, a global marker of inflammation, was not elevated in the study population. Innate lymphoid cells (ILC) type 3 (ILC-3) in abdominal AT decreased with active treatment compared with placebo (p = 0.04). Other immune white cells in AT and peripheral blood mononuclear cell (PBMC) fractions did not change with treatment compared to placebo (p>0.05); although ILC-2 declined in PBMCs (p = 0.007) in the sitagliptin treatment group. Two circulating biomarkers of atherogenesis, interferon-inducible protein-10 (IP-10) and sCD40L declined in plasma (p = 0.02 and p = 0.07, respectively) in the active treatment group, providing indirect validation of a net reduction in inflammation. CONCLUSIONS: In this pilot study, two cell types of the innate lymphoid system, ILC-3 in AT and ILC-2 PBMCs declined during treatment and as did circulating biomarkers of atherogenesis. Changes in other immune cells were not demonstrable. The study showed that sufficient abdominal AT could be obtained to quantify white cells of both innate and adaptive immunity and to demonstrate changes during therapy with an immune inhibitor. TRIAL REGISTRATION: ClinicalTrials.gov identifier (NCT number): NCT02576.


Assuntos
Gordura Abdominal/patologia , Imunidade Inata , Leucócitos Mononucleares/patologia , Obesidade/imunologia , Adulto , Biomarcadores/sangue , Estudos de Viabilidade , Feminino , Citometria de Fluxo , Humanos , Linfócitos/patologia , Masculino , Pessoa de Meia-Idade , Obesidade/sangue , Fosfato de Sitagliptina/farmacologia , Resultado do Tratamento
8.
Mutat Res ; 856-857: 503233, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32928373

RESUMO

Organophosphate (OP) pesticides are biotransformed into metabolites such as dialkylphosphates (DAPs). We have evaluated the genotoxicity of malathion and its metabolite dimethylthiophosphate (DMTP) in the human hepatic cell lines HepG2 and WRL-68 and in peripheral blood mononuclear cells (PBMC). In the Cytokinesis-Block Micronucleus assay (CBMN), malathion and DMTP increased the frequencies of micronuclei (MN) and nucleoplasmic bridges (NPB). Malathion was primarily clastogenic whereas DMTP was aneuploidogenic. When HepG2 or WRL-68 cells were treated with DMTP in the presence of sulconazole, a non-specific cytochrome P450 inhibitor, MN frequency was reduced, indicating that DMTP genotoxicity requires P450-cataliyzed metabolism.


Assuntos
Citocinese/efeitos dos fármacos , Malation/farmacologia , Testes de Mutagenicidade , Mutagênicos/farmacologia , Núcleo Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/patologia , Malation/toxicidade , Mutagênicos/toxicidade , Praguicidas/farmacologia , Praguicidas/toxicidade
9.
Sci Rep ; 10(1): 15805, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978492

RESUMO

Liquid biopsy represents a new frontier of cancer diagnosis and prognosis, which allows the isolation of tumor cells released in the blood stream. The extremely low abundance of these cells needs appropriate methodologies for their identification and enumeration. Herein we present a new protocol based on surface enhanced resonance Raman scattering (SERRS) gold multivalent nanostructures to identify and enumerate tumor cells with epithelial and mesenchimal markers. The validation of the protocol is obtained with spiked samples of peripheral blood mononuclear cells (PBMC). Gold nanostructures are functionalized with SERRS labels and with antibodies to link the tumor cells. Three types of such nanosystems were simultaneously used and the protocol allows obtaining the identification of all individual tumor cells with the help of a Random Forest ensemble learning method.


Assuntos
Células Epiteliais/patologia , Glioblastoma/patologia , Leucócitos Mononucleares/patologia , Células-Tronco Mesenquimais/patologia , Nanoestruturas/química , Neoplasias da Próstata/patologia , Análise Espectral Raman/métodos , Ouro/química , Humanos , Masculino , Células Tumorais Cultivadas
10.
Biosci Rep ; 40(9)2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32808647

RESUMO

Kawasaki disease (KD), also known as mucocutaneous lymph node syndrome, is an acute systemic vasculitis syndrome that mainly occurs in infants under 5 years of age. In the current manuscript, we were aiming to analyze the role of neutrophil extracellular traps (NETs) in the pathogenesis of KD, especially their interplay with peripheral blood mononuclear cells (PBMCs). Neutrophils were exposed to 20 nM phorbol myristate acetate (PMA), we found that neutrophils of KD patients were more likely to form NETs compared with healthy controls (HCs). Furthermore, PBMCs were cultured with NETs for 24 h, and we observed that NETs significantly increased the cell viability, suppressed cell apoptosis, and enhanced the pro-inflammatory cytokines production and NF-κB activation in PBMCs from KD patients. In addition, with the stimulation of NETs, the expression of vascular endothelial growth factor A (VEGF-A) and hypoxia-inducible factor-1α (HIF-1α) were increased, which were related with the pathological mechanism of KD. At last, we examined the activation of phosphoinositide 3 kinase (PI3K)/Akt signaling, and we found NETs treatment obviously enhanced the activation of PI3K and Akt. In conclusion, these findings suggested that the formation of NETs may alter the biologic responses of PBMC and affect the vascular injury in KD.


Assuntos
Armadilhas Extracelulares/fisiologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Síndrome de Linfonodos Mucocutâneos/sangue , Adolescente , Adulto , Apoptose , Estudos de Casos e Controles , Sobrevivência Celular , Citocinas/genética , Citocinas/metabolismo , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Acetato de Tetradecanoilforbol/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adulto Jovem
11.
Sci Rep ; 10(1): 13562, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32782316

RESUMO

NLRP3, an intracellular pattern recognition receptor, recognizes numerous pathogens and/or its own damage-associated molecules, and forms complexes with the adaptor protein ASC. These complexes constitute the NLRP3 inflammasome, a platform for processing interleukin (IL)-1ß and/or IL-18. Several NLRP3 mutations result in constitutive activation of the NLRP3 inflammasome, causing cryopyrin-associated periodic syndrome (CAPS). To the best of our knowledge, small compounds that specifically inhibit inflammasome activation through the pyrin domain (PYD) have not yet been developed. This study describes an attempt to develop small compounds targeting the NLRP3 inflammasome. A core chemical library of 9,600 chemicals was screened against reconstituted NLRP3 inflammasome in a cell-free system with an amplified luminescence proximity homogeneous assay and a cell-based assay by human peripheral blood mononuclear cells (PBMCs). Inflammasome activation was evaluated by ASC-speck formation in human PBMCs, accompanied by IL-1ß secretion and processing, and by using IL-1ß-based dual operating luciferase (IDOL) mice. The activity of these compounds was evaluated clinically using PBMCs from a patient with Muckle-Wells syndrome (MWS), a type of CAPS, with an R260W mutation in NLRP3. Screening identified KN3014, a piperidine-containing compound targeting the interaction between NLRP3 and ASC through the PYD. KN3014 reduced ASC-speck formation in human PBMCs, luminescence from IDOL mice, and auto-secretion of IL-1ß by PBMCs from the patient with MWS. These findings suggest that KN3014 may be an attractive candidate for treatment of MWS, as well as other NLRP3 inflammasomopathies.


Assuntos
Anti-Inflamatórios/farmacologia , Síndromes Periódicas Associadas à Criopirina/tratamento farmacológico , Inflamassomos/efeitos dos fármacos , Interleucina-1beta/antagonistas & inibidores , Leucócitos Mononucleares/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Piperidinas/farmacologia , Animais , Anti-Inflamatórios/química , Estudos de Casos e Controles , Síndromes Periódicas Associadas à Criopirina/metabolismo , Síndromes Periódicas Associadas à Criopirina/patologia , Ensaios de Triagem em Larga Escala , Humanos , Inflamassomos/metabolismo , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Masculino , Camundongos , Piperidinas/química
12.
Mol Carcinog ; 59(10): 1129-1139, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32822091

RESUMO

For solid tumors, extravasation of cancer cells and their survival in circulation represents a critical stage of the metastatic process that lacks complete understanding. Gaining insight into interactions between circulating tumor cells (CTCs) and other peripheral blood mononuclear cells (PBMCs) may provide valuable prognostic information. The purpose of this study was to use single-cell RNA-sequencing (scRNA-seq) of liquid biopsies from breast cancer patients to begin defining intravascular interactions. We captured CTCs from the peripheral blood of breast cancer patients using size-exclusion membranes followed by scRNA-seq of enriched CTCs and carry-over PBMCs. Transcriptome analysis identified two populations of CTCs: one enriched for transcripts indicative of estrogen responsiveness and increased proliferation and another enriched for transcripts characteristic of reduced proliferation and epithelial-mesenchymal transition (EMT). We applied interactome and pathway analysis to determine interactions between CTCs and other captured cells. Our analysis predicted for enhanced immune evasion in the CTC population with EMT characteristics. In addition, PD-1/PD-L1 pathway activation and T cell exhaustion were predicted in T cells isolated from breast cancer patients compared with normal T cells. We conclude that scRNA-seq of breast cancer CTCs generally stratifies them into two types based on their proliferative and epithelial state and differential potential to interact with PBMCs. Better understanding of CTC subtypes and their intravascular interactions may help design treatments directed against CTCs with high metastatic and immune-evasive competence.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal , Leucócitos Mononucleares/patologia , Células Neoplásicas Circulantes/patologia , Neoplasias da Mama/sangue , Feminino , Humanos , Prognóstico , Receptor ErbB-2/metabolismo , Receptores Estrogênicos/metabolismo , Receptores de Progesterona/metabolismo , Células Tumorais Cultivadas
13.
PLoS One ; 15(7): e0236375, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32726331

RESUMO

BACKGROUND: Malaria in pregnancy causes maternal, fetal and neonatal morbidity and mortality, and maternal innate immune responses are implicated in pathogenesis of these complications. The effects of malaria exposure and obstetric and demographic factors on the early maternal immune response are poorly understood. METHODS: Peripheral blood mononuclear cell responses to Plasmodium falciparum-infected erythrocytes and phytohemagglutinin were compared between pregnant women from Papua New Guinea (malaria-exposed) with and without current malaria infection and from Australia (unexposed). Elicited levels of inflammatory cytokines at 48 h and 24 h (interferon γ, IFN-γ only) and the cellular sources of IFN-γ were analysed. RESULTS: Among Papua New Guinean women, microscopic malaria at enrolment did not alter peripheral blood mononuclear cell responses. Compared to samples from Australia, cells from Papua New Guinean women secreted more inflammatory cytokines tumor necrosis factor-α, interleukin 1ß, interleukin 6 and IFN-γ; p<0.001 for all assays, and more natural killer cells produced IFN-γ in response to infected erythrocytes and phytohemagglutinin. In both populations, cytokine responses were not affected by gravidity, except that in the Papua New Guinean cohort multigravid women had higher IFN-γ secretion at 24 h (p = 0.029) and an increased proportion of IFN-γ+ Vδ2 γδ T cells (p = 0.003). Cytokine levels elicited by a pregnancy malaria-specific CSA binding parasite line, CS2, were broadly similar to those elicited by CD36-binding line P6A1. CONCLUSIONS: Geographic location and, to some extent, gravidity influence maternal innate immunity to malaria.


Assuntos
Imunidade Inata/genética , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Complicações Parasitárias na Gravidez/imunologia , Adolescente , Adulto , Austrália/epidemiologia , Antígenos CD36/genética , Eritrócitos/imunologia , Eritrócitos/parasitologia , Eritrócitos/patologia , Feminino , Número de Gestações/imunologia , Humanos , Interferon gama/genética , Interferon gama/imunologia , Interleucina-6/genética , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/parasitologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/parasitologia , Leucócitos Mononucleares/patologia , Malária Falciparum/epidemiologia , Malária Falciparum/parasitologia , Pessoa de Meia-Idade , Papua Nova Guiné/epidemiologia , Plasmodium falciparum/patogenicidade , Gravidez , Complicações Parasitárias na Gravidez/parasitologia , Complicações Parasitárias na Gravidez/patologia , Linfócitos T/imunologia , Linfócitos T/parasitologia , Adulto Jovem
14.
Mutat Res ; 854-855: 503200, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32660824

RESUMO

Germ cell tumour (GCT) patients who fail to respond to chemotherapy or who relapse have a poor prognosis. Timely and accurately stratifying such patients could optimise their therapy. We identified endogenous DNA damage levels as a prognostic marker for progression-free (PFS) and overall (OS) survival in chemotherapy-naïve GCT patients. In the present study, we have extended our previous results and reviewed the prognostic power of DNA damage level in GCTs. Endogenous DNA damage levels were measured with the comet assay. Receiver operator characteristic analysis was applied to determine the optimal cut-off value and to evaluate its prognostic accuracy. PFS and OS were estimated by the Kaplan-Meier method and compared using the log-rank test. Hazard ratio (HR) estimates were calculated by Cox regression analysis. A cut-off value of 6.34 provided the highest sensitivity and specificity, with area under curve values of 0.813 and 0.814 for disease progression and mortality, respectively. A % DNA in tail > 6.34 was significantly associated with shorter PFS (HR = 9.54, 95 % confidence interval [CI]: 3.43-26.55, p < 0.001) and OS (HR = 14.62, 95 % CI: 3.14-67.95, p = 0.001) by univariate analysis. The prognostic value of DNA damage measurement was confirmed by multivariate models (HR = 6.45, 95 % CI: 2.22-18.75, p = 0.001 for PFS and HR = 9.40, 95 % CI: 1.70-52.09, p = 0.010 for OS), when HR was adjusted for relevant clinical categories. The added prognostic value of DNA damage in combination with International Germ Cell Cancer Collaborative Group (IGCCCG) risk groups has been revealed. Endogenous DNA damage is an independent prognosticator for PFS and OS in GCT patients and its clinical use, particularly in combination with IGCCCG risk groups, may help in stratifying these patients.


Assuntos
Células Sanguíneas/patologia , Dano ao DNA/genética , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Embrionárias de Células Germinativas/patologia , Adulto , Células Cultivadas , Ensaio Cometa/métodos , Progressão da Doença , Intervalo Livre de Doença , Feminino , Humanos , Estimativa de Kaplan-Meier , Leucócitos Mononucleares/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Intervalo Livre de Progressão , Modelos de Riscos Proporcionais , Fatores de Risco
15.
Gene ; 754: 144880, 2020 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-32526260

RESUMO

Cancer secretion can change the properties of adjacent cells, including peripheral blood mononuclear cells (PBMCs). We investigated whether such secretion influences messenger RNA expression in PBMCs of patients with head and neck squamous cell carcinoma (HNSCC). In the present study, co-culture model of normal PBMCs and HNSCC cell lines were established. The PBMCs were subsequently subjected to RNA sequencing for transcriptome analysis. Furthermore, expression data from the Gene Expression Omnibus repository, platform GPL4133, series GSE39400, were gathered to analyze, afterward identify zinc finger CysCysHisCys (CCHC)-type domain-containing protein 6 (ZCCHC6) as the main gene involved in HNSCC. This gene was then validated by a quantitative real-time polymerase chain reaction. The results showed that ZCCHC6 was expressed at significantly higher levels in the patients with HNSCC than in the healthy controls, and the sensitivity and specificity of these findings for diagnostic purposes were 100.00% and 70.83%, respectively. In summary, our findings demonstrated that the secretion of HNSCC cells could cause the alterations in messenger RNA expression by PBMCs. The ZCCHC6 expression level may apply in HNSCC screening.


Assuntos
Biomarcadores Tumorais/sangue , Neoplasias de Cabeça e Pescoço/sangue , Leucócitos Mononucleares/metabolismo , RNA Nucleotidiltransferases/sangue , Carcinoma de Células Escamosas de Cabeça e Pescoço/sangue , Transcriptoma , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Feminino , Seguimentos , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Leucócitos Mononucleares/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia
16.
Sci Rep ; 10(1): 7277, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350353

RESUMO

Lung inflammation due to sarcoidosis is characterized by a complex cascade of immunopathologic events, including leukocyte recruitment and granuloma formation. α-melanocyte stimulating hormone (α-MSH) is a melanocortin signaling peptide with anti-inflammatory properties. We aimed to evaluate the effects of α-MSH in a novel in vitro sarcoidosis model. An in vitro sarcoidosis-like granuloma model was developed by challenging peripheral blood mononuclear cells (PBMCs) derived from patients with confirmed treatment-naïve sarcoidosis with microparticles generated from Mycobacterium abscessus cell walls. Unchallenged PBMCsand developed granulomas were treated daily with 10 µM α-MSH or saline as control. Cytokine concentrations in supernatants of culture and in cell extracts were measured using Illumina multiplex Elisa and western blot, respectively. Gene expression was analyzed using RNA-Seq and RT-PCR. Protein secretion and gene expression of IL-7, IL-7R, IFN-γ, MC1R, NF-κB, phosphorylated NF-κB (p-NF-κB), MARCO, and p-CREB were measured with western blot and RNAseq. A significant increase in IL-7, IL-7R, and IFN-γ protein expression was found in developed granulomas comparing to microparticle unchallenged PBMCs. IL-7, IL-7R, and IFN-γ protein expression was significantly reduced in developed granulomas after exposure to α-MSH compared with saline treated granulomas. Compared with microparticle unchallenged PBMCs, total NF-κB and p-NF-κB were significantly increased in developed granulomas, while expression of p-CREB was not changed. Treatment with α-MSH promoted a significantly higher concentration of p-CREB in granulomas. The anti-inflammatory effects of α-MSH were blocked by specific p-CREB inhibition. α-MSH has anti-inflammatory properties in this in vitro granuloma model, which is an effect mediated by induction of phosphorylation of CREB.


Assuntos
Anti-Inflamatórios/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Granuloma , Leucócitos Mononucleares , Modelos Biológicos , Infecções por Mycobacterium não Tuberculosas/metabolismo , Mycobacterium abscessus/metabolismo , Sarcoidose , alfa-MSH/farmacologia , Criança , Granuloma/metabolismo , Granuloma/microbiologia , Granuloma/patologia , Humanos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/microbiologia , Leucócitos Mononucleares/patologia , Masculino , Infecções por Mycobacterium não Tuberculosas/patologia , Sarcoidose/metabolismo , Sarcoidose/microbiologia , Sarcoidose/patologia
17.
Indian J Gastroenterol ; 39(2): 186-195, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32436176

RESUMO

BACKGROUND: Though a few studies in animal models suggest that intestinal helminths (IH) favorably affect evolution of gastritis associated with Helicobacter pylori (H. pylori) the studies supporting this concept in humans are only a few and are based on serological data. METHODS: To evaluate the possible influence of IH on the human gastric mucosa, three groups of Venezuelan adults with gastropathy (endoscopically diagnosed) were studied: H. pylori-/IH- (n = 17), H. pylori+/IH- (n = 18), and H. pylori+/IH+ (n = 11). Histological analysis (hematoxylin-eosin) and immunohistochemical staining (peroxidase) for cytokines interleukin-1beta (IL-1ß), tumor necrosis factor alpha (TNF-α), gamma interferon (IFN-γ), and interleukin 4 (IL-4) were undertaken in gastric antral biopsies. RESULTS: Expression of the four cytokines was detected in all individuals in varying degrees, but proinflammatory cytokines were expressed in a higher degree in the H. pylori+/IH- group, mainly IL-1ß (Th1-dominant immune response), associated with a higher degree of both histological inflammation and gastric cancer risk index (GCRI), as compared to the H. pylori-/IH- group. In contrast, an increased expression of IL-4 and a reduced expression of proinflammatory cytokines (Th2-dominant response), plus the tendency to a lower degree of mononuclear infiltration, mucosal atrophy in gastric corpus, and GCRI, were evidenced in the coinfected group. CONCLUSIONS: The findings of the present study is perhaps the first histological evidence of a possible modulatory effect of IH on the gastric mucosal inflammatory response due to H. pylori infection in humans.


Assuntos
Coinfecção/metabolismo , Coinfecção/patologia , Citocinas/metabolismo , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Gastrite/microbiologia , Gastrite/patologia , Infecções por Helicobacter , Helicobacter pylori , Mediadores da Inflamação/metabolismo , Enteropatias Parasitárias/metabolismo , Enteropatias Parasitárias/patologia , Adolescente , Adulto , Atrofia , Coinfecção/imunologia , Feminino , Mucosa Gástrica/imunologia , Gastrite/imunologia , Gastrite/metabolismo , Humanos , Imuno-Histoquímica , Enteropatias Parasitárias/imunologia , Leucócitos Mononucleares/patologia , Masculino , Pessoa de Meia-Idade , Adulto Jovem
18.
Cancer Immunol Res ; 8(6): 819-828, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32238380

RESUMO

Myeloid derived suppressor cells (MDSC) are a heterogeneous group of immature cells that accumulate in the peripheral blood and tumor microenvironment and are barriers to cancer therapy. MDSCs serve as prognostic biomarkers and are targets for therapy. On the basis of surface markers, three subsets of MDSCs have been defined in humans: granulocytic, monocytic, and early stage (e-MDSC). The markers attributed to e-MDSCs overlap with those of basophils, which are rare circulating myeloid cells with unrecognized roles in cancer. Thus, we asked whether e-MDSCs in circulation and the tumor microenvironment include basophils. On average, 58% of cells with e-MDSC surface markers in blood and 36% in ascites from patients with ovarian cancer were basophils based on CD123high expression and cytology, whereas cells with immature features were rare. Circulating and ascites basophils did not suppress proliferation of stimulated T cells, a key feature of MDSCs. Increased accumulation of basophils and basogranulin, a marker of basophil degranulation, were observed in ascites compared to serum in patients with newly diagnosed ovarian cancer. Basophils recruited to the tumor microenvironment may exacerbate fluid accumulation by their release of proinflammatory granular constituents that promote vascular leakage. No significant correlation was observed between peripheral basophil counts and survival in patients with ovarian cancer. Our results suggest that studies in which e-MDSCs were defined solely by surface markers should be reevaluated to exclude basophils. Both immaturity and suppression are criteria to define e-MDSCs in future studies.


Assuntos
Ascite/patologia , Basófilos/patologia , Biomarcadores Tumorais/sangue , Leucócitos Mononucleares/patologia , Células Supressoras Mieloides/patologia , Neoplasias Ovarianas/patologia , Microambiente Tumoral , Estudos de Casos e Controles , Feminino , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/sangue , Neoplasias Ovarianas/cirurgia , Prognóstico , Estudos Prospectivos , Estudos Retrospectivos , Taxa de Sobrevida , Células Tumorais Cultivadas
19.
Toxicol Lett ; 328: 1-6, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32315709

RESUMO

The genotoxicity of cationic lipopeptide nanoparticles (cLPNPs) was evaluated in vivo and in vitro comet assay and the in vivo chromosome aberrations test. In vitro comet assay, human blood cells were exposed to cLPNPs at the concentration of 2.5, 5, 10, 20, 40 and 100 µg/mL. Significant DNA damage was observed after 1 h exposure, but no effects were detected after 3 h. In vivo, cLPNPs were administered in single or five daily injection doses at 8, 20 and 40 mg/kg of body weight by subcutaneous injection to male mice. The cLPNPs caused DNA damage in the liver, lung and kidney, but not in the spleen. The kidney was more prone to genotoxic effects that persisted from 24 h to 14d after a single injection of cLPNPs. No statistically significant increase in the percentage of cells with chromosomal aberrations above the vehicle control was observed in mice bone marrow after a single or repeated injection of cLPNPs. In summary, cLPNPs shown to be genotoxic both in vivo and in vitro. The results suggest the importance of the use of highly sensitive methods, such as the comet assay, in order to determine the full genotoxic potential of nanoparticles.


Assuntos
Aberrações Cromossômicas/induzido quimicamente , Dano ao DNA , Lipopeptídeos/toxicidade , Nanopartículas/toxicidade , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Relação Dose-Resposta a Droga , Humanos , Injeções Subcutâneas , Rim/efeitos dos fármacos , Rim/patologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/patologia , Lipopeptídeos/química , Fígado/efeitos dos fármacos , Fígado/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Nanopartículas/química
20.
Exp Oncol ; 42(1): 16-24, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32231196

RESUMO

Response of chronic lymphocytic leukemia (CLL) patients to classical chemoimmunotherapy that remains the main strategy in treatment of this disease is strikingly variable. This issue requires the finding of biomarkers which could predict efficiency of drug administration and choose the best treatment option for each patient individually. The aim of this study was to find out association between cell surface receptors expression levels and CLL B cells sensitivity to chemotherapeutic drugs ex vivo. MATERIALS AND METHODS: The study was performed on malignant B cells isolated from peripheral blood of primary CLL patients. Flow cytometry, qPCR, ex vivo drug sensitivity assay, and cell viability assay were used in this study. RESULTS: The high CD5 expression level was linked to better bendamustine (BEN) and cyclophosphamide (CP) CLL B cells response in contrast to B cells with low CD5 expression. Sensitivity of CLL B cells to CP also could be predicted by high level of CD20 expression. Expression of CD38 and high levels of CD37 and CD40 showed CLL B cells resistance to BEN ex vivo. CLL B cells sensitivity to analyzed chemotherapeutic drugs was not dependent on CD22 expression status. The CD180 expression was detected in CLL B cells which were more susceptible to fludarabine and cyclophosphamide (FC) combinatory action. CLL B cells that coexpressed CD150 and CD180 on the cell surface were characterized by significantly decreased cell viability under fludarabine (FLU) exposure alone or FC in comparison with CD150-CD180- B cells. Cell surface expression level of CD150 was not associated with CLL B cells chemosensitivity. However, high mRNA expression level of mCD150 isoform in CLL B cells was linked to their FLU sensitivity and CP resistance, while high nCD150 mRNA expression level showed resistance to FLU. Simultaneous CD150 and CD180 ligation increased FLU resistance, but BEN susceptibility of CLL B cells. CD150 and CD180 alone or in combination are involved in upregulation of CD20 cell surface expression. CONCLUSION: Expression status of the CD5, CD20, CD37, CD38, CD40, CD150, and CD180 cell surface receptors could be used in prediction CLL B cells sensitivity to FLU, CP, BEN and FC ex vivo. Moreover, CD150 and CD180 receptors are involved in regulation of CLL B cells susceptibility to FLU and BEN. The CD150 and CD180 are positive regulators of CD20 expression that could make CD150+CD180+ CLL B cells more responsive to CD20-based immunotherapy.


Assuntos
Antígenos CD/metabolismo , Antineoplásicos/farmacologia , Linfócitos B/efeitos dos fármacos , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucócitos Mononucleares/efeitos dos fármacos , Receptores de Superfície Celular/metabolismo , Linfócitos B/metabolismo , Linfócitos B/patologia , Cloridrato de Bendamustina/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ciclofosfamida/farmacologia , Humanos , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Vidarabina/análogos & derivados , Vidarabina/farmacologia
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