RESUMEN
Osteosarcoma is one of the most frequent primary sarcoma of bone among adolescents. Early diagnosis of osteosarcoma is the key factor to achieve high survival rate of patients. Nevertheless, traditional histological biopsy is highly invasive and associated with the risk of arousing tumor spread. Herein, we develop a method integrating microfluidics and surface-enhanced Raman spectroscopy (SERS) to isolate plasma-derived exosomes and profile multiple exosomal biomarkers for the diagnosis of osteosarcoma. The method showed highly efficient isolation of exosomes directly from human plasma and can profile exosomes based on protein biomarkers, with the detection limit down to 2 exosomes per µL. The whole assay can be performed in 5 h and only consumed 50 µL of plasma for one analysis. With the method, we analyzed the level of three protein biomarkers, i.e., CD63, vimentin (VIM) and epithelial cell adhesion molecule (EpCAM), on plasma-derived exosomes from 20 osteosarcoma patients and 20 heathy controls. Significantly higher levels of CD63, VIM and EpCAM were observed on plasma exosomes from the osteosarcoma patients compared to the healthy controls. Based on the level of the exosomal biomarkers, a classification model was built for the rapid diagnosis of osteosarcoma, with the sensitivity, specificity and accuracy of 100%, 90% and 95%, respectively. The proposed method does not require complex operations nor expensive equipment, and has great promise in clinical diagnosis of cancer as a liquid biopsy technique.
Asunto(s)
Técnicas Biosensibles , Neoplasias Óseas , Exosomas , Osteosarcoma , Adolescente , Biomarcadores de Tumor , Neoplasias Óseas/diagnóstico , Neoplasias Óseas/metabolismo , Molécula de Adhesión Celular Epitelial/análisis , Exosomas/química , Humanos , Microfluídica/métodos , Osteosarcoma/diagnóstico , Osteosarcoma/metabolismo , Vimentina/análisis , Vimentina/metabolismoRESUMEN
We developed a strategy using immunomagnetic separation (IMS) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to test seafood allergens. The protocol employed commercial magnetic beads (MBs) functionalized with anti-human IgE antibodies to carry out the IMS of IgEs in blood samples, followed by capture of allergens from seafood protein extracts for allergy analysis. After elution, the captured allergens were identified by MALDI-TOF MS and HPLC-MS/MS. The non-specific adsorption of MBs to biomolecules, the reproducibility and sensitivity of the protocol were investigated. The method shows consistent results with enzyme-linked immunosorbent assay tests. The false positive rate of the present method for the allergy test is 0%. The protocol was applied to detect the allergens in greasy-back shrimp for checking the allergenicity of patients' serum. Cooking fish as soup may effectively decrease the allergenicity. The method can be potentially used to identify unknown allergens of seafood to ensure the safety of allergic patients.
Asunto(s)
Alérgenos , Separación Inmunomagnética , Animales , Humanos , Fenómenos Magnéticos , Reproducibilidad de los Resultados , Alimentos Marinos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Espectrometría de Masas en TándemRESUMEN
Osteosarcoma is the most frequent primary bone cancer, particularly among children and adolescents. The early diagnosis of osteosarcoma is significant for timely clinical treatment to reduce the mortality of patients. Exosomes play a significant role in intercellular communication and serve as promising biomarkers in liquid biopsy for the diagnosis and monitoring of tumors. Herein, we report the utility of surface-enhanced Raman scattering (SERS) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for rapid identification of osteosarcoma. We firstly profiled the intrinsic SERS signals and MALDI-TOF mass fingerprints of different subgroups of extracellular vesicles (EVs) and the corresponding cells, demonstrating that the SERS signals and MALDI-TOF mass spectra of exosomes from different types of cells were more discriminative compared to those of large and medium EVs and the cells themselves. Then, we characterized plasma-derived exosomes of 15 osteosarcoma patients and 15 healthy volunteers using SERS and MALDI-TOF MS, revealing distinctive biochemical differences in the spectra. We further utilized a data fusion approach to combine the two types of spectroscopic techniques, differentiating osteosarcoma patients from healthy controls with higher precision than either technique. The results reveal that the non-invasive liquid biopsy method using SERS and MALDI-TOF MS fingerprinting of exosomes has great potential for rapid diagnosis of osteosarcoma.
Asunto(s)
Exosomas , Osteosarcoma , Adolescente , Biomarcadores , Detección Precoz del Cáncer , Humanos , Osteosarcoma/diagnóstico , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Osteosarcoma is the most common primary sarcoma of bone among adolescents, often characterized by early lung metastasis resulting in high mortality. Recently, exosomes have been used in liquid biopsy to monitor tumors. Herein, we used matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to profile human plasma exosomes for the evaluation of osteosarcoma lung metastasis. Forty patients with osteosarcoma with (n = 20) or without (n = 20) lung metastasis as well as 12 heathy controls were recruited. Exosomes were isolated from human plasma for MALDI-TOF MS analysis. Multivariate statistical analyses were performed based on the MALDI-TOF mass spectra. The strategy can efficiently differentiate osteosarcomas from healthy controls and further discriminate osteosarcoma lung metastasis from non-lung metastasis. We identified seven exosomal proteins as potential biomarkers of osteosarcoma lung metastasis. The proposed method holds great promise to clinically diagnose osteosarcoma and monitor osteosarcoma lung metastasis.
RESUMEN
Aim: To dynamically analyze the differential m6A methylation during the progression and reversal of hepatic fibrosis. Materials & methods: We induced hepatic fibrosis in C57/BL6 mice by intraperitoneal injection of CCl4. The reversal model of hepatic fibrosis was established by stopping drug after continuous injection of CCl4. Dynamic m6A methylation was evaluated using MeRIP-Seq in the progression and reversal of hepatic fibrosis at different stages. Result: During the hepatic fibrosis, differential m6A methylation was mainly enriched in processes associated with oxidative stress and cytochrome metabolism, while differential m6A methylation was mainly enriched in processes associated with immune response and apoptosis in the hepatic fibrosis reversal. Conclusion: m6A methylation plays an important role in the progression and reversal of hepatic fibrosis.
Asunto(s)
Adenosina/análogos & derivados , Cirrosis Hepática/genética , ARN/metabolismo , Adenosina/metabolismo , Animales , Progresión de la Enfermedad , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/patología , Masculino , Metilación , Ratones Endogámicos C57BL , ARN/genética , Análisis Espectral , TranscriptomaRESUMEN
Phosphoribosylaminoimidazole carboxylase, phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS), an essential enzyme involved in de novo purine biosynthesis, is connected with formation of various tumors. However, the specific biological roles and related mechanisms of PAICS in gastric cancer (GC) remain unclear. In the present study, we identified for the first time that PAICS was significantly upregulated in GC and high expression of PAICS was correlated with poor prognosis of patients with GC. In addition, knockdown of PAICS significantly induced cell apoptosis, and inhibited GC cell growth both in vitro and in vivo. Mechanistic studies first found that PAICS was engaged in DNA damage response, and knockdown of PAICS in GC cell lines induced DNA damage and impaired DNA damage repair efficiency. Further explorations revealed that PAICS interacted with histone deacetylase HDAC1 and HDAC2, and PAICS deficiency decreased the expression of DAD51 and inhibited its recruitment to DNA damage sites by impairing HDAC1/2 deacetylase activity, eventually preventing DNA damage repair. Consistently, PAICS deficiency enhanced the sensitivity of GC cells to DNA damage agent, cisplatin (CDDP), both in vitro and in vivo. Altogether, our findings demonstrate that PAICS plays an oncogenic role in GC, which act as a novel diagnosis and prognostic biomarker for patients with GC.
Asunto(s)
Carcinogénesis/patología , Daño del ADN , Histona Desacetilasa 1/metabolismo , Histona Desacetilasa 2/metabolismo , Péptido Sintasas/metabolismo , Neoplasias Gástricas/enzimología , Neoplasias Gástricas/patología , Animales , Apoptosis/genética , Carcinogénesis/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Cisplatino/farmacología , Reparación del ADN , Regulación Enzimológica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Modelos Biológicos , Péptido Sintasas/deficiencia , Pronóstico , Unión Proteica , Regulación hacia Arriba/genéticaRESUMEN
Pathogenesis of colorectal cancer (CRC) is associated with alterations in gut microbiome. Previous studies have focused on the changes of taxonomic abundances by metagenomics. Variations of the function of intestinal bacteria in CRC patients compared to healthy crowds remain largely unknown. Here we collected fecal samples from CRC patients and healthy volunteers and characterized their microbiome using quantitative metaproteomic method. We have identified and quantified 91,902 peptides, 30,062 gut microbial protein groups, and 195 genera of microbes. Among the proteins, 341 were found significantly different in abundance between the CRC patients and the healthy volunteers. Microbial proteins related to iron intake/transport; oxidative stress; and DNA replication, recombination, and repair were significantly alternated in abundance as a result of high local concentration of iron and high oxidative stress in the large intestine of CRC patients. Our study shows that metaproteomics can provide functional information on intestinal microflora that is of great value for pathogenesis research, and can help guide clinical diagnosis in the future.
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Bacterias/clasificación , Proteínas Bacterianas/análisis , Neoplasias Colorrectales/microbiología , Proteómica/métodos , Bacterias/aislamiento & purificación , Bacterias/metabolismo , Estudios de Casos y Controles , Cromatografía Liquida , Replicación del ADN , Heces/microbiología , Femenino , Microbioma Gastrointestinal , Humanos , Hierro/metabolismo , Masculino , Estrés Oxidativo , Filogenia , Espectrometría de Masas en TándemRESUMEN
Bence-Jones protein is a biomarker in urine for multiple myeloma. Traditional methods for urine Bence-Jones protein detection are either less-sensitive or laborious. Herein, we describe a new method for the detection of urine Bence-Jones protein using nanoporous materials and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Macroporous ordered silica foams (MOSF) were used to enrich proteins in urine, and then the materials-proteins composites were analyzed by MALDI-TOF MS. Based on the presence of specific mass spectrometric signals, Bence-Jones protein can be detected for the diagnosis of multiple myeloma. Twenty-one clinical positive and twenty-seven clinical negative urine samples were analyzed by the method. High sensitivity (95.24%, 20/21) and specificity (100%, 27/27) for the diagnosis of multiple myeloma were achieved. Compared to other methods for multiple myeloma diagnosis, e.g. immunofixation electrophoresis and immunonephelometry, our approach is more rapid, economical and convenient, which can be a new choice for the clinical diagnosis of Bence-Jones protein related diseases.
Asunto(s)
Proteína de Bence Jones/orina , Mieloma Múltiple/diagnóstico , Mieloma Múltiple/orina , Nanopartículas/química , Dióxido de Silicio/química , Humanos , Tamaño de la Partícula , Porosidad , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Propiedades de SuperficieRESUMEN
OBJECTIVES: The National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) was used to determine the chronic prostatitis syndrome in young men (from 20 to 48 years old) of clinical validity. METHODS: 227 patients with chronic prostatitis syndrome (CPS)/chronic pelvic pain syndrome and 32 patients with BPH were randomized to study using NIH-CPSI. RESULTS: 1. The main manifestations of CPS patients were pain or uncomfort. Those were more common in CPS than BPH. 2. 79.30% CPS patients had a sensation of not emptying bladder completely after finished urinating, 44.93% patients had to urinate again less than two hours after finished urinating. 3. 51.51% patients with CPS have more effect on work, 90.31% patients on free life, and 68.72% patients on quality of life than BPH. CONCLUSIONS: According NIH-CPSI, the main manifestations of CPS are pain or uncomfort. CPS patients have more effect on work and quality of life.