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1.
Anticancer Res ; 40(3): 1691-1695, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32132076

RESUMO

BACKGROUND/AIM: As of 2020, carbohydrate antigen 72.4 (Ca 72.4) has been rarely investigated in the gastric juice (GJ) of patients with gastric cancer (GC). Our aim was to analyze the significance and role of this tumor antigen in the GJ of our GC population. PATIENTS AND METHODS: Between April 2012 and July 2013, 37 patients with operable GC were prospectively investigated to determine the GJ Ca 72.4 levels before surgical manipulation. RESULTS: GJ Ca 72.4 ≥6.49 ng/ml strongly correlated with the traditional categories of aggressive cancer (advanced tumor depth and stage, lymph node invasion and metastatic lymphatic ratio, indication to adjuvant treatment). It also associated with shorter survival (p=0.049) and is, thus, suggested as an independent factor of poor prognosis in GC patients (p=0.047). CONCLUSION: The GJ Ca 72.4 parameter should be considered an indicator of an aggressive tumor phenotype and should be used in the prognostic assessment of GC patients.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Suco Gástrico/metabolismo , Neoplasias Gástricas/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia
2.
Am J Physiol Renal Physiol ; 318(2): F518-F530, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31904283

RESUMO

Mucin-type O-linked glycosylation, a posttranslational modification affecting the stability and biophysical characteristics of proteins, requires C1GalT1 (T synthase) and its obligate, X-linked chaperone Cosmc. Hypomorphic C1GalT1 mutations cause renal failure via not yet established mechanisms. We hypothesize that impaired Cosmc-dependent O-glycosylation in podocytes is sufficient to cause disease. Podocyte-specific Cosmc knockout mice were generated and phenotyped to test this hypothesis. Female heterozygous mice displaying mosaic inactivation of Cosmc in podocytes due to random X-linked inactivation were also examined. Mice with podocyte-specific Cosmc deletion develop profound albuminuria, foot process effacement, glomerular sclerosis, progressive renal failure, and impaired survival. Glomerular transcriptome analysis reveals early changes in cell adhesion, extracellular matrix organization, and chemokine-mediated signaling pathways, coupled with podocyte loss. Expression of the O-glycoprotein podoplanin was lost, while Tn antigen, representing immature O-glycans, was most abundantly found on podocalyxin. In contrast to hemizygous male and homozygous female animals, heterozygous female mosaic animals developed only mild albuminuria, focal foot process effacement, and nonprogressive kidney disease. Ultrastructurally, Cosmc-deficient podocytes formed Tn antigen-positive foot processes interdigitating with those of normal podocytes but not with other Cosmc-deficient cells. This suggests a cell nonautonomous mechanism for mucin-type O-glycoproteins in maintaining podocyte function. In summary, our findings demonstrated an essential and likely cell nonautonomous role for mucin-type O-glycosylation for podocyte function.


Assuntos
Albuminúria/metabolismo , Chaperonas Moleculares/metabolismo , Mucinas/metabolismo , Podócitos/metabolismo , Insuficiência Renal/metabolismo , Albuminúria/genética , Albuminúria/patologia , Albuminúria/fisiopatologia , Animais , Antígenos Glicosídicos Associados a Tumores/metabolismo , Células Cultivadas , Feminino , Predisposição Genética para Doença , Glicosilação , Heterozigoto , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Chaperonas Moleculares/genética , Mosaicismo , Fenótipo , Podócitos/ultraestrutura , Insuficiência Renal/genética , Insuficiência Renal/patologia , Insuficiência Renal/fisiopatologia , Fatores Sexuais , Sialoglicoproteínas/metabolismo
3.
Cancer Immunol Immunother ; 68(8): 1369-1378, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31338558

RESUMO

Intrahepatic cholangiocarcinoma (ICC) is a rare malignancy with poor prognosis. The evaluation of recurrence risk after liver resection is of great importance for ICCs. We aimed to assess the prognostic value of intra- and peritumoral immune infiltrations and to establish a novel histopathology-related immunoscore (HRI) associated with ICC recurrence. A total of 280 ICC patients who received curative resection between February 2005 and July 2011 were enrolled in our study. Patients were randomly assigned to the derivation cohort (n = 176) or the validation cohort (n = 104). Sixteen immune biomarkers in both intra- and peritumoral tissues were examined by immunohistochemistry. The least absolute shrinkage and selection operator (LASSO) Cox model was used to establish the HRI score. Cox regression analysis was used for multivariate analysis. Nine recurrence-related immune features were identified and integrated into the HRI score. The HRI score was used to categorize patients into low-risk and high-risk groups using the X-tile software. Kaplan-Meier analysis presented that the HRI score showed good stratification between low-risk and high-risk groups in both the derivation cohort (P < 0.001) and the validation cohort (P = 0.014), respectively. Multivariate analysis demonstrated that serum γ-glutamyl transpeptidase, carbohydrate antigen 19-9, lymphoid metastasis, tumor numbers, and the HRI score were independent risk factors associated with recurrence-free survival (RFS). The combination of Shen's model and HRI score provided better performance in recurrence prediction compared with traditional staging systems. The HRI score might serve as a promising RFS predictor for ICC with prognostic values.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Biomarcadores Tumorais/metabolismo , Colangiocarcinoma/diagnóstico , Neoplasias Hepáticas/diagnóstico , Fígado/metabolismo , Colangiocarcinoma/mortalidade , Colangiocarcinoma/patologia , Estudos de Coortes , Feminino , Hepatectomia , Humanos , Fígado/patologia , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Recidiva Local de Neoplasia , Prognóstico , Projetos de Pesquisa , Análise de Sobrevida , Carga Tumoral , gama-Glutamiltransferase/metabolismo
4.
Biosci Trends ; 13(3): 216-224, 2019 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-31168022

RESUMO

Protein glycosylation is a diverse form of post-translational modification. Two to three consecutive O-linked N-acetylgalactosamines (Tn-antigens) are recognized by antibodies such as MLS128. MLS128 mAb inhibited cell growth and bound to a 110 kDa glycoprotein (GP) in LS180 and HT29 colon cancer cells. However, purification and identification of the 110 kDa GP was unsuccessful due to its low abundance. The present study used a highly sophisticated and sensitive mass spectrometry method to identify proteins immunoprecipitated with MLS128 and separated by two-dimensional gel electrophoresis. Three desmosome components were identified. Of these, desmocollin and desmoglein shared many similar characteristics, including molecular mass, pI, and potential Tn-antigen sites. Western blotting analyses of LS180 cell lysates revealed a common 110 kDa band recognized by MLS128 and anti-desmocollin, but not by anti-desmoglein. Immunofluorescence microscopy of LS180 cells revealed that desmocollin is membrane-bound, while desmoglein is primarily localized in the cytosol. Confocal microscopy demonstrated colocalization of the desmocollin-specific antibody with the MLS128 antibody on the cell membrane, suggesting that desmocollin may contain Tn-antigens recognized by MLS128. Treatment of LS180 cells with siRNA to knock down desmocollin expression or a desmocollin-specific antibody decreased cell viability, suggesting a critical role for this protein in cell growth and survival. N-glycosidase F digestion of the 110 kDa GP and desmocollin suggested that although both proteins contain N-glycosylation sites, they are not identical. These findings suggest that desmocollin colocalizes with the 110 kDa GP and that growth inhibition induced by the MLS128 antibody may be mediated through a mechanism that involves desmocollin.


Assuntos
Neoplasias do Colo/metabolismo , Desmocolinas/metabolismo , Glicoproteínas/metabolismo , Anticorpos Monoclonais/imunologia , Antígenos Glicosídicos Associados a Tumores/imunologia , Antígenos Glicosídicos Associados a Tumores/metabolismo , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/imunologia , Desmocolinas/imunologia , Glicoproteínas/imunologia , Células HT29 , Humanos , Microscopia Confocal , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/metabolismo , Espectrometria de Massas em Tandem
5.
Proc Natl Acad Sci U S A ; 116(9): 3518-3523, 2019 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808745

RESUMO

The globo-series glycosphingolipids (GSLs) SSEA3, SSEA4, and Globo-H specifically expressed on cancer cells are found to correlate with tumor progression and metastasis, but the functional roles of these GSLs and the key enzyme ß1,3-galactosyltransferase V (ß3GalT5) that converts Gb4 to SSEA3 remain largely unclear. Here we show that the expression of ß3GalT5 significantly correlates with tumor progression and poor survival in patients, and the globo-series GSLs in breast cancer cells form a complex in membrane lipid raft with caveolin-1 (CAV1) and focal adhesion kinase (FAK) which then interact with AKT and receptor-interacting protein kinase (RIP), respectively. Knockdown of ß3GalT5 disrupts the complex and induces apoptosis through dissociation of RIP from the complex to interact with the Fas death domain (FADD) and trigger the Fas-dependent pathway. This finding provides a link between SSEA3/SSEA4/Globo-H and the FAK/CAV1/AKT/RIP complex in tumor progression and apoptosis and suggests a direction for the treatment of breast cancer, as demonstrated by the combined use of antibodies against Globo-H and SSEA4.


Assuntos
Neoplasias da Mama/genética , Galactosiltransferases/genética , Glicoesfingolipídeos/genética , Microdomínios da Membrana/genética , Antígenos Glicosídicos Associados a Tumores/genética , Antígenos Glicosídicos Associados a Tumores/metabolismo , Apoptose/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caveolina 1/genética , Caveolina 1/metabolismo , Progressão da Doença , Proteína de Domínio de Morte Associada a Fas/genética , Feminino , Proteína-Tirosina Quinases de Adesão Focal/genética , Regulação Neoplásica da Expressão Gênica/genética , Glicoesfingolipídeos/metabolismo , Humanos , Substâncias Macromoleculares/química , Substâncias Macromoleculares/metabolismo , Microdomínios da Membrana/metabolismo , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-akt/genética , Saporinas/genética , Transdução de Sinais/genética , Antígenos Embrionários Estágio-Específicos/genética , Antígenos Embrionários Estágio-Específicos/metabolismo
6.
EBioMedicine ; 41: 268-275, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30772302

RESUMO

BACKGROUND: Although identification of lymph node (LN) metastasis is a well-recognized strategy for improving outcomes in patients with gastric cancer (GC), currently there is lack of availability of adequate molecular biomarkers that can identify such metastasis. Herein we have developed a robust gene-expression signature for detecting LN metastasis in early stage GC by using a transcriptome-wide biomarker discovery and subsequent validation in multiple clinical cohorts. METHODS: A total of 532 patients with pathological T1 and T2 GC from 4 different cohorts were analyzed. Two independent datasets (n = 96, and n = 188) were used to establish a gene signature for the identification of LN metastasis in GC patients. The diagnostic performance of our gene-expression signature was subsequently assessed in two independent clinical cohorts using qRT-PCR assays (n = 101, and n = 147), and subsequently compared against conventional tumor markers and image-based diagnostics. FINDINGS: We established a 15-gene signature by analyzing multiple high throughput datasets, which robustly distinguished LN status in both training (AUC = 0.765, 95% CI 0.667-0.863) and validation cohorts (AUC = 0.742, 95% CI 0.630-0.852). Notably, the 15-gene signature was significantly superior compared to the conventional tumor markers, CEA (P = .04) and CA19-9 (P = .005), as well as computed tomography-based imaging (P = .04). INTERPRETATION: We have established and validated a 15-gene signature for detecting LN metastasis in GC patients, which offers a robust diagnostic tool for potentially improving treatment outcomes in gastric cancer patients. FUND: NIH: CA72851, CA181572, CA14792, CA202797, CA187956; CPRIT: RP140784: Baylor Sammons Cancer Center polot grants (AG), VPRT: 9610337, CityU 21101115, 11102317, 11103718; JCYJ20170307091256048 (XW).


Assuntos
Neoplasias Gástricas/diagnóstico , Transcriptoma , Idoso , Antígenos Glicosídicos Associados a Tumores/genética , Antígenos Glicosídicos Associados a Tumores/metabolismo , Área Sob a Curva , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Antígeno Carcinoembrionário/metabolismo , Detecção Precoce de Câncer , Feminino , Humanos , Linfonodos/metabolismo , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Curva ROC , Neoplasias Gástricas/patologia , Tomografia Computadorizada por Raios X
7.
Int J Mol Sci ; 20(1)2019 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-30626136

RESUMO

Morniga-G, the Gal-specific black mulberry (Morus nigra) lectin, displays high affinity for T (CD176) and Tn (CD175) antigens, frequently expressed at the cancer cell surface. The effects of Morniga-G were investigated on a Tn-positive leukemic Jurkat cell line. The lectin, used in a concentration range between 5⁻20 µg/mL, induced cell death in leukemic Jurkat cells. Microscopic and cytofluorometric analyses indicated that Jurkat cell death was essentially apoptotic, associated with an increase in the ceramide content and a depolarization of the mitochondrial transmembrane potential. This lectin-mediated cell death was inhibited by the pan caspase-inhibitor zVAD. In addition, cleavage of caspases 8, 9, and 3 was observed in Morniga-G-treated Jurkat cells whereas Jurkat cell lines that are deficient in caspase 8⁻10, caspase 9, or FADD, survived to the lectin-mediated toxicity. Furthermore, in the presence of TRAIL- or DR5-blocking mononoclonal antibodies, Jurkat cells became resistant to Morniga-G, suggesting that the lectin triggers cell death via the TRAIL/DR5 pathway. In silico computer simulations suggest that Morniga-G might facilitate both the DR5 dimerization and the building of TRAIL/DR5 complexes. Finally, upon treatment of Jurkat cells with benzyl-GalNAc, an O-glycosylation inhibitor, a decrease in Tn antigen expression associating with a reduced Morniga-G toxicity, was observed. Taken together, these results suggest that Morniga-G induces the cell death of Tn-positive leukemic cells via concomitant O-glycosylation-, caspase-, and TRAIL/DR5-dependent pathways.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Lectinas/farmacologia , Leucemia/patologia , Morus/química , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Caspases/metabolismo , Morte Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Ceramidas/metabolismo , Glicosilação , Humanos , Células Jurkat , Linfócitos/efeitos dos fármacos , Linfócitos/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Agregados Proteicos
8.
N Biotechnol ; 49: 77-87, 2019 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-30273682

RESUMO

Circulating tumour cells (CTCs) originating from a primary tumour, lymph nodes and distant metastases hold great potential for liquid biopsies by providing a molecular fingerprint for disease dissemination and its temporal evolution through the course of disease management. CTC enumeration, classically defined on the basis of surface expression of Epithelial Cell Adhesion Molecule (EpCAM) and absence of the pan-leukocyte marker CD45, has been shown to correlate with clinical outcome. However, existing approaches introduce bias into the subsets of captured CTCs, which may exclude biologically and clinically relevant subpopulations. Here we explore the overexpression of the membrane protein O-glycan sialyl-Tn (STn) antigen in advanced bladder and colorectal tumours, but not in blood cells, to propose a novel CTC isolation technology. Using a size-based microfluidic device, we show that the majority (>90%) of CTCs isolated from the blood of patients with metastatic bladder and colorectal cancers express the STn antigen, supporting a link with metastasis. STn+ CTC counts were significantly higher than EpCAM-based detection in colorectal cancer, providing a more efficient cell-surface biomarker for CTC isolation. Exploring this concept, we constructed a glycan affinity-based microfluidic device for selective isolation of STn+ CTCs and propose an enzyme-based strategy for the recovery of viable cancer cells for downstream investigations. Finally, clinically relevant cancer biomarkers (transcripts and mutations) in bladder and colorectal tumours, were identified in cells isolated by microfluidics, confirming their malignant origin and highlighting the potential of this technology in the context of precision oncology.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Biomarcadores Tumorais/metabolismo , Oncologia/métodos , Microfluídica/métodos , Células Neoplásicas Circulantes/metabolismo , Medicina de Precisão/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Separação Celular , Análise Mutacional de DNA , Molécula de Adesão da Célula Epitelial/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Polissacarídeos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
9.
Acta Histochem ; 121(2): 164-170, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30558911

RESUMO

The latest research suggests cells in the maculae flavae located at both ends of the lamina propria of the human vocal fold mucosa have stemness. This study investigated the differentiation potential of the cells in the maculae flavae of the human vocal fold mucosa. Four normal human adult vocal folds from surgical specimens were used. After extraction of the anterior maculae flavae located at the anterior end of the lamina propria of the human vocal fold mucosa under microscope, the maculae flavae were minced, cultured and proliferated in mesenchymal stem cell growth medium and morphological features were assessed. Cell surface markers were detected using flow cytometry. Cell differentiation into adipogenic, chondrogenic and osteogenic lineages was performed. Cell's differentiation potential was assessed using a human pluripotent stem cell functional identification kit and immunohistochemistry. Subcultured cells formed a colony-forming unit. Subcultured cells expressed CD90, CD105 and CD73 and lacked expression of CD45, CD34, CD11b, CD19 and HLA-DR. They differentiated into adipogenic, chondrogenic and osteogenic lineages. Consequently, the cell features in the maculae flavae meet the minimal criteria defining mesenchymal stromal cells. In addition, subcultured cells differentiated into ectoderm, mesoderm and endoderm and expressed stage-specific embryonic antigen 3 (SSEA-3). The results of this study are consistent with the hypothesis that the cells in the maculae flavae in the lamina propria of the human vocal fold mucosa are putative stem cells.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Diferenciação Celular/fisiologia , Antígenos Embrionários Estágio-Específicos/metabolismo , Células-Tronco/citologia , Prega Vocal/citologia , Condrogênese/fisiologia , Humanos , Células-Tronco Mesenquimais/citologia , Nicho de Células-Tronco/fisiologia
10.
Dis Markers ; 2019: 2721876, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31929839

RESUMO

Objective: We examined serum anti-p53 antibodies (S-p53Ab) in colorectal cancer. Specifically, we retrospectively investigated the use of S-p53Ab as a prognostic marker after surgery for colorectal cancer. Materials and Methods: The levels of S-p53Ab, carcinoembryonic antigen (CEA), and carbohydrate antigen 19-9 (CA19-9) were measured in 160 colorectal cancer patients who underwent surgical treatment. The rate of postoperative change (RPC) in the S-p53Ab titer was calculated as [subsequent antibody titer-lowest titer]/lowest titer. Results: A relationship between recurrence and RPC in the S-p53Ab titer was not observed in patients who tested negative for S-p53Ab preoperatively. In addition, no patients, who tested negative for S-p53Ab preoperatively, tested positive for S-p53Ab at the follow-up after surgery. Of the patients who tested positive for S-p53Ab preoperatively, those recurrences had a significantly higher RPC compared with those who did not (p < 0.001). Conclusions: Although S-p53Ab is not a significant tumor marker in patients who test negative preoperatively, increases in the S-p53Ab titer should be continuously monitored and measured in patients who are positive for this antibody preoperatively, regardless of whether they later test negative.


Assuntos
Anticorpos/sangue , Antígenos Glicosídicos Associados a Tumores/metabolismo , Biomarcadores Tumorais/sangue , Neoplasias Colorretais/cirurgia , Proteína Supressora de Tumor p53/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/imunologia , Neoplasias Colorretais/sangue , Neoplasias Colorretais/patologia , Procedimentos Cirúrgicos do Sistema Digestório , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Estudos Retrospectivos , Resultado do Tratamento
12.
Biochimie ; 151: 14-26, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29802864

RESUMO

Cutaneous melanoma, the most aggressive form of skin cancer, responds poorly to conventional therapy. The appearance of Tn antigen-modified proteins in cancer is correlated with metastasis and poor prognoses. The Tn determinant has been recognized as a powerful diagnostic and therapeutic target, and as an object for the development of anti-tumor vaccine strategies. This study was designed to identify Tn-carrying proteins and reveal their influence on cutaneous melanoma progression. We used a lectin-based strategy to purify Tn antigen-enriched cellular glycoproteome, the LC-MS/MS method to identify isolated glycoproteins, and the DAVID bioinformatics tool to classify the identified proteins. We identified 146 different Tn-bearing glycoproteins, 88% of which are new. The Tn-glycoproteome was generally enriched in proteins involved in the control of ribosome biogenesis, CDR-mediated mRNA stabilization, cell-cell adhesion and extracellular vesicle formation. The differential expression patterns of Tn-modified proteins for cutaneous primary and metastatic melanoma cells supported nonmetastatic and metastatic cell phenotypes, respectively. To our knowledge, this study is the first large-scale proteomic analysis of Tn-bearing proteins in human melanoma cells. The identified Tn-modified proteins are related to the biological and molecular nature of cutaneous melanoma and may be valuable biomarkers and therapeutic targets.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Biomarcadores Tumorais/metabolismo , Glicoproteínas/metabolismo , Melanoma/metabolismo , Proteômica , Neoplasias Cutâneas/metabolismo , Linhagem Celular Tumoral , Cromatografia Líquida , Humanos , Melanoma/patologia , Metástase Neoplásica , Neoplasias Cutâneas/patologia , Espectrometria de Massas em Tandem
13.
Cell Transplant ; 27(2): 285-298, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29637816

RESUMO

Multilineage-differentiating stress-enduring (Muse) cells are endogenous nontumorigenic stem cells collectable as stage-specific embryonic antigen 3 (SSEA-3) + from various organs including the bone marrow and are pluripotent-like. The potential of human bone marrow-derived Muse cells to commit to cardiac lineage cells was evaluated. We found that (1) initial treatment of Muse cells with 5'-azacytidine in suspension culture successfully accelerated demethylation of cardiac marker Nkx2.5 promoter; (2) then transferring the cells onto adherent culture and treatment with early cardiac differentiation factors including wingless-int (Wnt)-3a, bone morphogenetic proteins (BMP)-2/4, and transforming growth factor (TGF) ß1; and (3) further treatment with late cardiac differentiation cytokines including cardiotrophin-1 converted Muse cells into cardiomyocyte-like cells that expressed α-actinin and troponin-I with a striation-like pattern. MLC2a expression in the final step suggested differentiation of the cells into an atrial subtype. MLC2v, a marker for a mature ventricular subtype, was expressed when cells were treated with Dickkopf-related protein 1 (DKK-1) and Noggin, inhibitors of Wnt3a and BMP-4, respectively, between steps (2) and (3). None of the steps included exogenous gene transfection, making induced cells feasible for future clinical application.


Assuntos
Miócitos Cardíacos/metabolismo , Células-Tronco Pluripotentes/metabolismo , Antígenos Glicosídicos Associados a Tumores/metabolismo , Proteína Morfogenética Óssea 2/metabolismo , Proteína Morfogenética Óssea 4/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Proteína Homeobox Nkx-2.5/metabolismo , Humanos , Miócitos Cardíacos/citologia , Células-Tronco Pluripotentes/citologia , Antígenos Embrionários Estágio-Específicos/metabolismo , Fatores de Crescimento Transformadores/metabolismo
14.
Dev Biol ; 436(2): 108-124, 2018 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-29499182

RESUMO

T antigen (Galß1-3GalNAcα1-Ser/Thr) is an evolutionary-conserved mucin-type core 1 glycan structure in animals synthesized by core 1 ß1,3-galactosyltransferase 1 (C1GalT1). Previous studies showed that T antigen produced by Drosophila C1GalT1 (dC1GalT1) was expressed in various tissues and dC1GalT1 loss in larvae led to various defects, including decreased number of circulating hemocytes, hyper-differentiation of hematopoietic stem cells in lymph glands, malformation of the central nervous system, mislocalization of neuromuscular junction (NMJ) boutons, and ultrastructural abnormalities in NMJs and muscle cells. Although glucuronylated T antigen (GlcAß1-3Galß1-3GalNAcα1-Ser/Thr) has been identified in Drosophila, the physiological function of this structure has not yet been clarified. In this study, for the first time, we unraveled biological roles of glucuronylated T antigen. Our data show that in Drosophila, glucuronylation of T antigen is predominantly carried out by Drosophila ß1,3-glucuronyltransferase-P (dGlcAT-P). We created dGlcAT-P null mutants and found that mutant larvae showed lower expression of glucuronylated T antigen on the muscles and at NMJs. Furthermore, mislocalization of NMJ boutons and a partial loss of the basement membrane components collagen IV (Col IV) and nidogen (Ndg) at the muscle 6/7 boundary were observed. Those two phenotypes were correlated and identical to previously described phenotypes in dC1GalT1 mutant larvae. In addition, dGlcAT-P null mutants exhibited fewer NMJ branches on muscles 6/7. Moreover, ultrastructural analysis revealed that basement membranes that lacked Col IV and Ndg were significantly deformed. We also found that the loss of dGlcAT-P expression caused ultrastructural defects in NMJ boutons. Finally, we showed a genetic interaction between dGlcAT-P and dC1GalT1. Therefore, these results demonstrate that glucuronylated core 1 glycans synthesized by dGlcAT-P are key modulators of NMJ bouton localization, basement membrane formation, and NMJ arborization on larval muscles.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Membrana Basal/metabolismo , Proteínas de Drosophila/metabolismo , Músculos/metabolismo , Junção Neuromuscular/metabolismo , Animais , Western Blotting , Drosophila/genética , Glucuronosiltransferase/metabolismo , Immunoblotting , Larva/metabolismo , Espectrometria de Massas , Fenótipo , Polissacarídeos/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
15.
Cell Death Dis ; 9(3): 402, 2018 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-29540729

RESUMO

Colorectal cancer (CRC) is one of the most common neoplasms worldwide. However, the mechanisms underlying its development are still poorly understood. Thyroid hormone Receptor Interactor 13 (TRIP13) is a key mitosis regulator, and recent evidence has shown that it is an oncogene. Here, we report that TRIP13, which is overexpressed in CRC, is correlated with the CEA (carcino-embryonic antigen), CA19-9 (carbohydrate antigen 19-9) and pTNM (pathologic primary tumor, lymph nodes, distant metastasis) classification. Multivariate analyses showed that TRIP13 might serve as an independent prognostic marker of CRC. We also found that TRIP13 promoted CRC cell proliferation, invasion and migration in vitro and subcutaneous tumor formation in vivo. Furthermore, the potential mechanism underlying these effects involves the interaction of TRIP13 with a 14-3-3 protein, YWHAZ, which mediates G2-M transition and epithelial-mesenchymal transition (EMT). Together, these findings suggest that TRIP13 may be a potential biomarker and therapeutic target for CRC.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proteínas de Ciclo Celular/metabolismo , Neoplasias Colorretais/metabolismo , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , ATPases Associadas a Diversas Atividades Celulares/genética , Adulto , Idoso , Animais , Antígenos Glicosídicos Associados a Tumores/genética , Antígenos Glicosídicos Associados a Tumores/metabolismo , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/fisiopatologia , Transição Epitelial-Mesenquimal , Feminino , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Prognóstico , Ligação Proteica
16.
Mol Biochem Parasitol ; 221: 56-65, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29581010

RESUMO

Cryptosporidium spp. are the causative agents of diarrheal disease worldwide, but effective treatments are lacking. Cryptosporidium employs mucin-like glycoproteins with O-glycans to attach to and infect host intestinal epithelial cells. The Tn antigen (GalNAcα1-Ser/Thr) is an O-glycan essential for these processes, as Tn-specific lectins and a Tn-specific monoclonal antibody block attachment to and infection of host cells in vitro. The enzymes in Cryptosporidium catalyzing their synthesis, however, have not been studied. Previously, we identified four genes encoding putative UDP N-acetyl-α-d-galactosamine:polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts) in the genomes of three Cryptosporidium spp. Here we report the in silico analysis, cloning, expression, purification, and characterization of one of the four enzymes Cryptosporidium parvum (Cp)-ppGalNAc-T4. This enzyme contains the characteristic domains and motifs conserved in ppGalNAc-Ts and is expressed at multiple time points during in vitro infection. Recombinant soluble Cp-ppGalNAc-T4 was enzymatically active against an unmodified EA2 peptide suggesting that it may function as an "initiating" ppGalNAc-T. Cp-ppGalNAc-T4 also exhibited a strong preference for UDP-GalNAc over other nucleotide sugar donors and was active against unmodified and O-glycosylated versions of the C. parvum gp40-derived peptide, with a preference for the former, suggesting it may play a role in modifying this glycoprotein in vivo. Given the importance of mucin-type O-glycosylation in Cryptosporidium spp., the enzymes that catalyze their synthesis may serve as potential therapeutic targets.


Assuntos
Cryptosporidium parvum/enzimologia , N-Acetilgalactosaminiltransferases/genética , N-Acetilgalactosaminiltransferases/metabolismo , Uridina Difosfato N-Acetilgalactosamina/metabolismo , Antígenos Glicosídicos Associados a Tumores/metabolismo , Clonagem Molecular , Cryptosporidium parvum/genética , Células Epiteliais/metabolismo , Expressão Gênica , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Modelos Moleculares , N-Acetilgalactosaminiltransferases/química , N-Acetilgalactosaminiltransferases/isolamento & purificação , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Especificidade por Substrato
17.
Int J Neurosci ; 128(9): 821-827, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29355452

RESUMO

OBJECTIVE: To analyze the clinical features, diagnostic strategies and therapeutic methods associated with paraneoplastic neurological syndromes. METHODS: A retrospective study of paraneoplastic neurological syndromes was performed at a single center in Shandong, East China. The medical records and follow-up data of 28 patients were intensively reviewed between February 2011 and December 2014. RESULTS: Twenty-four (85.7%) patients experienced subacute or chronic onset of disease, and the most common symptoms reported were mild myasthenia and paresthesias. Twenty-five (89.3%) patients presented nervous system lesions prior to occult tumors, and the median time frame between paraneoplastic neurological syndromes onset and the diagnosis of a tumor was 15 weeks. Sensorimotor neuropathy, Lambert-Eaton myasthenic syndrome and limbic encephalitis were the three most common neurological syndromes reported. Elevated serum tumor markers were observed in 44.0% of patients, while 40.7% of patients were positive for onconeural antibodies. Tumors were detected in 21 (75.0%) patients after repeated whole-body screening, and lung carcinomas were the most common primary tumor detected. Seventeen patients received anti-tumor or immunological therapy, and clinical symptoms were relieved in 13 (76.5%) of these patients. CONCLUSIONS: In the majority of paraneoplastic neurological syndromes patients, the onset of disease is subacute or chronic with mild clinical symptoms. Nervous system lesions usually occur prior to occult tumors with complicated and various clinical manifestations. Neither tumor markers nor onconeural antibodies exhibit a high rate of occurrence, while repeated whole-body screening is helpful in identifying occult tumors. Early diagnosis and treatment are crucial to these patients.


Assuntos
Polineuropatia Paraneoplásica/complicações , Polineuropatia Paraneoplásica/epidemiologia , Adulto , Idoso , Antígenos Glicosídicos Associados a Tumores/metabolismo , China/epidemiologia , China/etnologia , Eletroencefalografia , Eletromiografia , Feminino , Seguimentos , Humanos , Queratina-19/metabolismo , Encefalite Límbica/etiologia , Imagem por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Condução Nervosa , Exame Neurológico , Polineuropatia Paraneoplásica/diagnóstico por imagem , Estudos Retrospectivos
18.
Dis Markers ; 2018: 9845123, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30647803

RESUMO

Background: The correlation between tumor markers (TM) and TNM stage of non-small-cell lung cancer (NSCLC) has not been widely reported. Methods: TM levels (CEA, CA125, CA15-3, CA19-9, CA72-4, CYFRA21-1, and SCC-Ag) in 424 cases of lung adenocarcinoma (LAC), 166 cases of lung squamous cell carcinoma (LSCC), and 103 cases of benign chest disease (BCD) were analyzed before treatment. Results: By Kendall's tau-b correlation analysis, CEA, CA125, CA15-3, CA19-9, CA72-4, CYFRA21-1, and SCC-Ag were correlated with T stage of LAC (r = 0.235, p < 0.05; r = 0.298, p < 0.05; r = 0.254, p < 0.05; r = 0.063, p < 0.05; r = 0.080, p < 0.05; r = 0.268, p < 0.05; and r = 0.080, p < 0.05). CEA, CA125, CA15-3, CA19-9, CA72-4, and CYFRA21-1 were correlated with N stage of LAC (r = 0.356, p < 0.05; r = 0.415, p < 0.05; r = 0.340, p < 0.05; r = 0.117, p < 0.05; r = 0.175, p < 0.05; and r = 0.351, p < 0.05). CEA, CA125, CA15-3, CA19-9, CA72-4, and CYFRA21-1 were correlated with M stage of LAC (r = 0.365, p < 0.05; r = 0.353, p < 0.05; r = 0.293, p < 0.05; r = 0.135, p < 0.05; r = 0.169, p < 0.05; and r = 0.312, p < 0.05). CA125, CYFRA21-1, and SCC-Ag were correlated with T stage of LSCC (r = 0.202, p < 0.05; r = 0.233, p < 0.05; and r = 0.099, p < 0.05). CA125 and CYFRA21-1 were correlated with N stage of LSCC (r = 0.178, p < 0.05 and r = 0.284, p < 0.05). CA125, CA15-3, and CYFRA21-1 were correlated with M stage of LSCC (r = 0.214, p < 0.05; r = 0.152, p < 0.05; and r = 0.213, p < 0.05). Combining hazard ratio, AUC, sensitivity, specificity, NPV, and PPV, it was concluded that CEA and CYFRA21-1were the most related TM of LAC. SCC-Ag and CYFRA21-1 were the most related TM of LSCC. Conclusions: CEA combined with CYFRA21-1 contributed to auxiliary diagnosis of LAC. CYFRA21-1 combined with SCC-Ag contributed to auxiliary diagnosis of LSCC. CEA, CA125, CA15-3, CA19-9, CA72-4, and CYFRA21-1 were correlated with primary tissue and metastasis of LAC. CA125 and CYFRA21-1 were correlated with primary tissue and metastasis of LSCC.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Neoplasias Pulmonares/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/metabolismo , Antígenos Glicosídicos Associados a Tumores/metabolismo , Antígeno Ca-125/metabolismo , Antígeno Carcinoembrionário/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Feminino , Humanos , Queratina-19/metabolismo , Neoplasias Pulmonares/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Mucina-1/metabolismo , Estadiamento de Neoplasias , Sensibilidade e Especificidade , Adulto Jovem
19.
Adv Med Sci ; 63(1): 205-211, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29197783

RESUMO

PURPOSE: Attachment of Helicobacter pylori to the mucous epithelial cells and the mucous layer is said to be a crucial step for infection development. Sugar antigens of gastric mucins (MUC5AC, MUC1) can act as receptors for bacterial adhesins. The aim of the study was to investigate if Lotus tetragonolobus and Maackia amurensis lectins influence the level of MUC1, MUC5AC, Lewis b, H type 1, sialyl Lewis x, phospho-IκBα and interleukin 8 in Helicobacter pylori infected gastric cancer cells. MATERIALS AND METHODS: The study was performed with one clinical H. pylori strain and CRL-1739 gastric cancer cells. To assess the levels of mentioned factors immunosorbent ELISA assays were used. RESULTS: Coculture of cells with bacteria had no clear effect on almost all examined structures. After coculture with H. pylori and lectins, a decrease of the level of both mucins, Lewis b and H type 1 antigens was observed. Lectins addition had no effect on sialyl Lewis x. Maackia amurensis caused slight increase of phospho-IκBα while interleukin 8 level was decreased. CONCLUSIONS: Lotus tetragonolobus and Maackia amurensis lectins can mediate in binding of Helicobacter pylori to gastric epithelium.


Assuntos
Antígenos Glicosídicos Associados a Tumores/metabolismo , Helicobacter pylori/fisiologia , Interleucina-8/metabolismo , Lectinas/uso terapêutico , Lotus/química , Maackia/química , Inibidor de NF-kappaB alfa/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/microbiologia , Linhagem Celular Tumoral , Helicobacter pylori/efeitos dos fármacos , Humanos , Lectinas/farmacologia , Mucinas/metabolismo , Fosforilação/efeitos dos fármacos
20.
Int J Mol Sci ; 18(12)2017 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-29232830

RESUMO

Biomarkers for the early detection of pancreatic cancer are urgently needed. The aim of this pilot study was to evaluate changes in serum N-glycoproteins and their glycosylation status prior to clinical presentation of pancreatic cancer that may be potential biomarkers. Prediagnosis serum samples pooled according to five time-to-diagnosis groups and a non-cancer control pool were digested with trypsin, labelled with mass tags, and subjected to titanium dioxide capture, deglycosylation, and 2D-LC-MS/MS profiling. Unbound peptides were profiled in parallel. Across the sample groups, 703 proteins were quantified and 426 putative sites of N-glycosylation were identified with evidence of several novel sites. Altered proteins with biomarker potential were predominantly abundant inflammatory response, coagulation, and immune-related proteins. Whilst glycopeptide profiles largely paralleled those of their parent proteins, there was evidence of altered N-glycosylation site occupancy or sialic acid content prior to diagnosis for some proteins, most notably of immunoglobulin gamma chains. α-1-Antitrypsin was tested as a biomarker, but found not to complement carbohydrate antigen 19-9 (CA19-9) in early detection of cancer. In conclusion, we provide preliminary evidence of altered glycosylation of several serum proteins prior to pancreatic cancer diagnosis, warranting further investigation of these proteins as early biomarkers. These changes may be largely driven by inflammatory processes that occur in response to tumour formation and progression.


Assuntos
Proteínas Sanguíneas/química , Glicoproteínas/química , Neoplasias Pancreáticas/diagnóstico , Proteômica/métodos , Idoso , Antígenos Glicosídicos Associados a Tumores/metabolismo , Sítios de Ligação , Biomarcadores Tumorais/química , Cromatografia Líquida , Detecção Precoce de Câncer , Feminino , Glicoproteínas/análise , Glicoproteínas/sangue , Glicosilação , Humanos , Pessoa de Meia-Idade , Ácido N-Acetilneuramínico/química , Neoplasias Pancreáticas/sangue , Projetos Piloto , Espectrometria de Massas em Tandem , alfa 1-Antitripsina/sangue , alfa 1-Antitripsina/química , alfa 1-Antitripsina/metabolismo
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