RESUMO
Tannic acid (TA), a naturally occurring polyphenol, is a potent anti-oxidant with anti-proliferative effects on multiple cancers. However, its ability to modulate gene-specific expression of tumour suppressor genes and oncogenes has not been assessed. This work investigates the mechanism of TA to regulate canonical and non-canonical STAT pathways to impose the gene-specific induction of G1-arrest and apoptosis. Regardless of the p53 status and membrane receptors, TA induced G1-arrest and apoptosis in breast cancer cells. Tannic acid distinctly modulated both canonical and non-canonical STAT pathways, each with a specific role in TA-induced anti-cancer effects. Tannic acid enhanced STAT1 ser727 phosphorylation via upstream serine kinase p38. This STAT1 ser727 phosphorylation enhanced the DNA-binding activity of STAT1 and in turn enhanced expression of p21Waf1/Cip1 . However, TA binds to EGF-R and inhibits the tyrosine phosphorylation of both STAT1 and STAT3. This inhibition leads to the inhibition of STAT3/BCL-2 DNA-binding activity. As a result, the expression and mitochondrial localization of BCl-2 are declined. This altered expression and localization of mitochondrial anti-pore factors resulted in the release of cytochrome c and the activation of intrinsic apoptosis cascade involving caspases. Taken together, our results suggest that TA modulates EGF-R/Jak2/STAT1/3 and P38/STAT1/p21Waf1/Cip1 pathways and induce G1-arrest and intrinsic apoptosis in breast carcinomas.
Assuntos
Neoplasias da Mama/metabolismo , Receptores ErbB/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Taninos/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Apoptose/efeitos dos fármacos , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Sinergismo Farmacológico , Feminino , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Gefitinibe , Humanos , Fosforilação/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ligação Proteica/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Quinazolinas/farmacologia , Tamoxifeno/farmacologiaRESUMO
Molecular markers are helpful diagnostic tools, particularly for cytologically indeterminate thyroid nodules. Preoperative RET/PTC1 rearrangement analysis in BRAF and RAS wild-type indeterminate thyroid nodules would permit the formulation of an unambiguous surgical plan. Cycle threshold values according to the cell count for detection of the RET/PTC1 rearrangement by real-time reverse transcription-polymerase chain reaction (RT-PCR) using fresh and routine air-dried TPC1 cells were evaluated. The correlation of RET/PTC1 rearrangement between fine-needle aspiration (FNA) and paired formalin-fixed paraffin-embedded (FFPE) specimens was analyzed. RET/PTC1 rearrangements of 76 resected BRAF and RAS wild-type classical PTCs were also analyzed. Results of RT-PCR and the Nanostring were compared. When 100 fresh and air-dried TPC1 cells were used, expression of RET/PTC1 rearrangement was detectable after 35 and 33 PCR cycles, respectively. The results of RET/PTC1 rearrangement in 10 FNA and paired FFPE papillary thyroid carcinoma (PTC) specimens showed complete correlation. Twenty-nine (38.2%) of 76 BRAF and RAS wild-type classical PTCs had RET/PTC1 rearrangement. Comparison of RET/PTC1 rearrangement analysis between RT-PCR and the Nanostring showed moderate agreement with a κ value of 0.56 (p = 0.002). The RET/PTC1 rearrangement analysis by RT-PCR using routine air-dried FNA specimen was confirmed to be technically applicable. A significant proportion (38.2%) of the BRAF and RAS wild-type PTCs harbored RET/PTC1 rearrangements.
Assuntos
Biópsia por Agulha Fina , Genes ras , Testes Genéticos , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-ret/genética , Nódulo da Glândula Tireoide/diagnóstico , Nódulo da Glândula Tireoide/genética , Perfilação da Expressão Gênica , Rearranjo Gênico , Testes Genéticos/métodos , Humanos , Mutação , Reação em Cadeia da Polimerase , Cuidados Pré-Operatórios , Nódulo da Glândula Tireoide/cirurgiaRESUMO
Mixed carcinoma shows a mixture of glandular and signet ring/poorly cohesive cellular histological components and the prognostic significance of each component is not fully understood. This study aimed to investigate the significance of the poorly cohesive cellular histological component as a risk factor for lymph node metastasis and to examine the diagnostic reliability of endoscopic biopsy. Clinicopathologic characteristics of 202 patients who underwent submucosal invasive gastric carcinoma resection with lymph node dissection in 2005-2012 were reviewed. Mixed carcinoma accounted for 27.2% (56/202) of cases. The overall prevalence of lymph node metastasis was 17.3% (35/202). Lymphatic invasion (P < 0.001), family history of carcinoma (P = 0.025), tumor size (P = 0.004), Lauren classification (P = 0.042), and presence of any poorly cohesive cellular histological component (P = 0.021) positively correlated with the lymph node metastasis rate on univariate analysis. Multivariate analyses revealed lymphatic invasion, family history of any carcinoma, and the presence of any poorly cohesive cellular histological component to be significant and independent factors related to lymph node metastasis. Review of preoperative biopsy slides showed that preoperative biopsy demonstrated a sensitivity of 63.6% and a specificity of 100% in detecting the presence of the poorly cohesive cellular histological component, compared with gastrectomy specimens. The presence of any poorly cohesive cellular histological component was an independent risk factor associated with lymph node metastasis in submucosal invasive gastric carcinoma. Endoscopic biopsy had limited value in predicting the presence and proportion of the poorly cohesive cellular histologic component due to the heterogeneity of mixed carcinoma.
Assuntos
Carcinoma/patologia , Mucosa Gástrica/patologia , Neoplasias Gástricas/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma/diagnóstico , Feminino , Gastrectomia , Gastroscopia , Humanos , Excisão de Linfonodo , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Razão de Chances , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Neoplasias Gástricas/diagnósticoRESUMO
OBJECTIVE: RAS mutations are the most common mutations in thyroid nodules with indeterminate cytology by fine-needle aspiration cytology (FNAC), and are mutually exclusive with BRAF mutations. However, the diagnostic utility of RAS mutation analysis is uncertain. We evaluated the diagnostic utility of RAS mutation analysis in indeterminate thyroid nodules. DESIGN, PATIENTS, AND MEASUREMENTS: A total of 155 thyroid nodules (90 benign and 65 indeterminate) negative for BRAF(V) (600E) mutations on FNAC were analysed for mutations in RAS codon 61 using pyrosequencing methods. We evaluated diagnostic accuracy of RAS mutation for predicting thyroid malignancy based on the surgical pathologic diagnosis. RESULTS: Among the 65 BRAF(V) (600E) -negative indeterminate thyroid nodules identified by FNAC, 25 (38·5%) exhibited point mutations in RAS 61 consisting of 18 NRAS 61 (72%), and 7 HRAS 61 (28%) mutations. In contrast, only five of 90 (5·6%) nodules with benign cytology had RAS mutations. Only two of 25 (8·0%) RAS 61(+) indeterminate nodules exhibited malignant ultrasonographic features. Of the 15 patients with RAS 61(+) -indeterminate nodules who underwent thyroid surgery, 14 (93·3%) were diagnosed as malignant, including 13 follicular variant of papillary thyroid carcinomas (FVPTC), and one follicular thyroid carcinoma (FTC). The average tumour size was 1·79 ± 0·62 cm. Multifocality was seen in 28·6% of cases, with 7·1% exhibiting extrathyroidal extension; no lymph node or distant metastases were evident. Based on the surgical pathologic diagnosis results, preoperative RAS 61 mutation analysis on FNAC exhibited 93·3% sensitivity, 75·0% specificity, 93·3% positive predictive value, 75·0% negative predictive value and 89·5% diagnostic accuracy for predicting malignancies. CONCLUSION: Our results suggest that RAS mutation analysis holds great promise as a preoperative diagnostic tool for predicting FVPTC in cytologically and sonographically indeterminate nodules negative for BRAF mutations.
Assuntos
Adenocarcinoma Folicular/genética , Carcinoma/genética , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas/genética , Neoplasias da Glândula Tireoide/genética , Nódulo da Glândula Tireoide/patologia , Proteínas ras/genética , Adenocarcinoma Folicular/diagnóstico por imagem , Adulto , Idoso , Biópsia por Agulha Fina , Carcinoma/diagnóstico por imagem , Carcinoma Papilar , Análise Mutacional de DNA , Feminino , Genes ras , Humanos , Masculino , Pessoa de Meia-Idade , Mutação Puntual , Valor Preditivo dos Testes , Proteínas Proto-Oncogênicas B-raf/genética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Câncer Papilífero da Tireoide , Neoplasias da Glândula Tireoide/diagnóstico por imagem , Nódulo da Glândula Tireoide/diagnóstico por imagem , Nódulo da Glândula Tireoide/genética , UltrassonografiaRESUMO
BACKGROUND: Combination therapy, which reduces the dosage intensity of the individual drugs while increasing their efficacy, is not a novel approach for the treatment of cancer. Methylsulfonylmethane (MSM) is an organic sulfur compound shown to act against tumor cells. Tamoxifen is a commercially available therapeutic agent for breast malignancies. METHODS: In the current study, we analyzed the combinatorial effect of MSM and tamoxifen on the suppression of ER-positive breast cancer xenograft growth and metastasis. Additionally, we also validated the molecular targets by which the drug combination regulated tumor growth and metastasis. RESULTS: We observed that the combination of MSM and tamoxifen regulated cell viability and migration in vitro. The intragastric administration of MSM and subcutaneous implantation of tamoxifen tablets led to tumor growth suppression and inhibition of the Janus kinase 2 (Jak2)/signal transducer and activator of transcription 5b (STAT5b) pathway. Our study also assessed the regulation of signaling molecules implicated in the growth, progression, differentiation, and migration of cancer cells, such as Jak2, STAT5b, insulin-like growth factor-1Rß, and their phosphorylation status. CONCLUSIONS: Study results indicated that this combination therapy inhibited tumor growth and metastasis. Therefore, this drug combination may have a synergistic and powerful anticancer effect against breast cancer.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Dimetil Sulfóxido/administração & dosagem , Janus Quinase 2/genética , Fator de Transcrição STAT5/genética , Sulfonas/administração & dosagem , Tamoxifeno/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Janus Quinase 2/antagonistas & inibidores , Metástase Neoplásica , Receptores de Somatomedina/antagonistas & inibidores , Receptores de Somatomedina/genética , Fator de Transcrição STAT5/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Peroxisome proliferator-activated receptor gamma (PPAR-γ), a ligand-activated transcription factor has been investigated as the target for cancer treatment as well as metabolic disorders. Recent studies have demonstrated that PPAR-γ ligands are anti-tumorigenic in prostate cancer due to anti-proliferative and pro-differentiation effects. The aim of this study was to validate PPAR-γ expression in malignant and benign prostate tissues by immunohistochemistry and quantitative real-time polymerase chain reaction (PCR). A total of 730 prostatic adenocarcinomas (PCAs) including 63 whole sections from radical prostatectomy specimens and tissue microarrays containing 667 PCAs were subject to immunostaining for two PPAR-γ antibodies. Twenty-five benign prostate tissues and PCAs were selected for investigating mRNA expression by quantitative real-time PCR. 10.7% of PCAs (78/730) showed cytoplasmic immunoreactivity of PPAR-γ and no nuclear immunoreactivity was noted in PCAs. Most benign prostatic glands showed negative immunoreactivity of PPAR-γ except for variable weak cytoplasmic staining in some glands. Nuclear immunoreactivity of PPAR-γ was noted some central zone and verumontanum mucosal epithelium. The constitutive PPAR-γ mRNA showed significantly lower level in PCAs compared to that in the benign tissues. There was no difference of PPAR-γ mRNA expression between low (≤7) and high (>7) Gleason score groups. There was no association of PPAR-γ mRNA level or cytoplasmic immunostaining with Gleason grade or pathologic stage. Our study supported the evidence of extra-nuclear localization and nongenomic actions of PPAR-γ. Further studies are needed to assess the functional role of PPAR-γ and to validate its therapeutic implication in prostate cancer.
Assuntos
Adenocarcinoma/patologia , Regulação Neoplásica da Expressão Gênica , PPAR gama/genética , PPAR gama/metabolismo , Neoplasias da Próstata/patologia , Adenocarcinoma/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Próstata/patologia , Prostatectomia , Neoplasias da Próstata/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Análise Serial de TecidosRESUMO
A primary ductal adenocarcinoma (PDA) of the lacrimal gland is a rare distinct subtype of an epithelial tumor arising in the lacrimal gland. PDA is the counterpart of salivary duct carcinoma (SDC) resembling an invasive ductal carcinoma (IDC) of the breast. In our case, PDA revealed histopathological and immunohistochemical results corresponding to SDC. Interestingly, the tumor cells showed intracytoplasmic vacuoles containing dense eosinophilic hyaline globules at light microscopy. Ultrastructurally, the tumor cells exhibited microvilli-lined intracytoplasmic lumen containing homogenous electron-dense secretory products. A previous study demonstrated that numerous intracytoplasmic lumens of tumor cells are favored breast malignant tumor, similar to the histopathology of PDA, rather than benign lesion. This characteristic finding may be meaningful to diagnose high grade epithelial tumors including PDA.
Assuntos
Adenocarcinoma/ultraestrutura , Carcinoma de Células Escamosas/ultraestrutura , Neoplasias de Cabeça e Pescoço/ultraestrutura , Hialina/ultraestrutura , Aparelho Lacrimal/ultraestrutura , Neoplasias Epiteliais e Glandulares/ultraestrutura , Adenocarcinoma/metabolismo , Carcinoma de Células Escamosas/diagnóstico , Neoplasias de Cabeça e Pescoço/diagnóstico , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-Idade , Neoplasias das Glândulas Salivares/diagnóstico , Neoplasias das Glândulas Salivares/metabolismo , Neoplasias das Glândulas Salivares/ultraestrutura , Carcinoma de Células Escamosas de Cabeça e PescoçoRESUMO
We evaluated p53, KRAS, BRAF and CTNNB1 mutation and p53, WT1, p16 and beta-catenin expression in 31 ovarian high-grade serous adenocarcinoma. Twenty-five (80.6%) tumors contained functional mutations of p53; three frameshift, four nonsense and 19 missense mutations. None of the tumors showed KRAS, BRAF or CTNNB1 mutation. In all 18 tumors with missense mutations, ≥60% of tumor cells were strongly positive for p53 immunostaining whereas all tumors with frameshift or nonsense mutations were completely negative. Missense mutation was correlated with diffuse and strong imunoreaction and frameshift/nonsense mutation was correlated with completely negative immunoreaction (P = 0.000). Tumors with wild-type p53 revealed a wide range of immunostaining patterns. In 27 (87.1%) and 18 (58.1%) tumors, ≥50% of tumor cells were moderate to strongly positive for WT1 and p16, respectively. A considerable intratumoral heterogeneity for p16 expression was present. None of the tumors demonstrated nuclear beta-catenin expression. p53 mutations appear to be a powerful molecular marker for ovarian high-grade serous adenocarcinoma. Using p53 with an appropriate interpretation criteria together with WT1, p16 and beta-catenin, most of the high-grade serous adenocarcinoma could be distinguished from other ovarian tumors.
Assuntos
Biomarcadores Tumorais/genética , Cistadenocarcinoma Seroso/genética , Neoplasias Ovarianas/genética , Proteína Supressora de Tumor p53/genética , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Códon sem Sentido , Inibidor p16 de Quinase Dependente de Ciclina , Cistadenocarcinoma Seroso/metabolismo , Cistadenocarcinoma Seroso/patologia , Primers do DNA/genética , DNA de Neoplasias/química , DNA de Neoplasias/genética , Feminino , Humanos , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas p21(ras) , República da Coreia , Análise de Sequência de DNA , Proteína Supressora de Tumor p53/metabolismo , Proteínas WT1/genética , Proteínas WT1/metabolismo , beta Catenina/genética , beta Catenina/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismoRESUMO
OBJECTIVE: The objective of our study was to evaluate the relationships between BRAF mutation status, sonography findings, and fine-needle aspiration cytology features in patients with papillary thyroid carcinoma (PTC) and to evaluate the diagnostic merits of BRAF mutation status and sonography findings as adjuncts to cytologic diagnoses. MATERIALS AND METHODS: From March 2006 through June 2008, clinicopathologic factors, sonography findings, cytology results, and BRAF mutation status were evaluated in 524 patients (437 women and 87 men) with 553 thyroid nodules; of the 170 malignant nodules, 164 were PTCs. Clinicopathologic factors, sonography findings, and cytology results were correlated with BRAF status. The diagnostic sensitivities and specificities of sonography, cytology, and BRAF analysis and their combinations were compared. RESULTS: The V600E mutation of BRAF (BRAF(V600E)) was detected in 141 of 170 malignant thyroid nodules (82.9%) (140 PTCs and one follicular variant of PTC). Multiple logistic regression revealed that BRAF status was not associated with sonography features with the exception of a negative relation between BRAF(V600E) and an irregular shape (p = 0.004). An indeterminate cytology result was more frequent for BRAF-negative PTC than BRAF-positive PTC (p = 0.035). By adding BRAF status to cytology, diagnostic sensitivity for PTC was significantly increased (94.1%) as compared with cytology alone (81.8%) (p < 0.001). The triple combination-that is, sonography, cytology, and BRAF analysis-showed higher sensitivity than BRAF plus cytology (98.2% vs 94.1%, respectively) (p < 0.05). CONCLUSION: The sonography features of PTC, other than an irregular shape, are not related to BRAF status and the combination of sonography and BRAF testing would increase the diagnostic accuracy of cytologic diagnoses of PTC.
Assuntos
Carcinoma Papilar/diagnóstico por imagem , Carcinoma Papilar/genética , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias da Glândula Tireoide/diagnóstico por imagem , Neoplasias da Glândula Tireoide/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia por Agulha Fina , Carcinoma Papilar/patologia , Distribuição de Qui-Quadrado , Análise Mutacional de DNA , Feminino , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sensibilidade e Especificidade , Neoplasias da Glândula Tireoide/patologia , UltrassonografiaRESUMO
Primary squamous cell carcinoma of the thyroid (SCC-T) is extremely rare. Its clinical presentation is similar to that of anaplastic carcinoma. Metastasis or extension from the head and neck area should be ruled out, as patients with SCC-T have a poorer prognosis than patients who have a thyroid extension from an adjacent tumor. An 87-year-old man presented with a longstanding painless mass in the right thyroid and had experienced 2 months of pain upon swallowing. A right lobectomy was performed with resection of thyroid cartilage, cricoid cartilage, a portion of the first to third tracheal ring and the right neck lymph node. A histological examination revealed pure SCC. The tumor cells showed diffuse immunoreactivity to CK5/6, CK19 and p63. Immunoreactivity to EMA and p53 was focally positive. TTF-1, galectin 3 and thyroglobulin immunoreactivity was restricted to the non-neoplastic thyroid tissue. Both tumor cells and non-neoplastic follicular cells were negative for CD5. The MIB-1 index was 36%. DNA extracted from the tumor identified a BRAF V600E mutation in exon 15 and a BRAF G468A mutation in exon 11, whereas DNA from non-tumorous cells did not contain a mutation. These molecular findings may suggest a direct transformation from papillary carcinoma to SCC-T.
Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Papilar/genética , Carcinoma de Células Escamosas/diagnóstico , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Análise Mutacional de DNA , DNA de Neoplasias/análise , Humanos , Masculino , Mutação/genética , Neoplasias da Glândula Tireoide/diagnósticoRESUMO
Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry-based restriction fragment mass polymorphism (RFMP) assay was adapted to human papillomavirus (HPV) genotyping. The analytical sensitivity and the clinical utility were evaluated by testing defined HPV genome equivalents and a total of 426 specimens composed of normal cytology, atypical squamous cells of undetermined significance, low grade squamous intraepithelial lesion, high grade squamous intraepithelial lesion and invasive squamous cell carcinoma. The RFMP assay was able to detect 38.4-114.6 genomic equivalents of a wide variety of HPV types. The RFMP assay detected 34 different HPV genotypes in cervical samples of which 8% were found to be multiple-type infections. The high-risk HPV positivity rate according to the histological diagnosis was 7.9% (8/101), 31.7% (38/120), 50% (55/110), 86% (37/43), 96.2% (50/52) in normal, atypical squamous cells of undetermined significance, low grade squamous intraepithelial lesion, high grade squamous intraepithelial lesion and squamous cell carcinoma subgroups, respectively. Diagnostic sensitivities/specificities for the cervical lesions of squamous cell carcinoma and high grade squamous intraepithelial lesion or worse histology were found to be 96.2%/92.1% and 91.6%/92.1%, respectively. The sensitivity, accuracy, wide range of genotype identification and high-throughput capacity with cost-effectiveness of the test consumables make the RFMP assay suitable for mass screening and monitoring of HPV-associated cervical cancer.
Assuntos
Tipagem Molecular , Papillomaviridae/genética , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/virologia , Sequência de Bases , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/virologia , Colo do Útero/patologia , Colo do Útero/virologia , DNA Viral , Feminino , Genótipo , Humanos , Programas de Rastreamento , Polimorfismo Genético , Sensibilidade e Especificidade , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/virologia , Displasia do Colo do Útero/patologia , Displasia do Colo do Útero/virologiaRESUMO
CONTEXT: Several ultrasonographic (US) features of thyroid nodules have been reported to predict malignancy. The BRAF(V600E) mutation is a useful diagnostic marker for differentiating papillary thyroid carcinoma from benign thyroid nodules, especially in BRAF(V600E) -prevalent populations such as in Korea. OBJECTIVE: To evaluate the association of BRAF(V600E) mutation with US features of thyroid nodules in predicting the malignancy of thyroid nodules in Korean patients. DESIGN: A total of 991 thyroid nodules from 823 patients in fine-needle aspiration biopsy (FNAB) specimens were investigated. The relationship between US features and the presence of BRAF(V600E) mutation by pyrosequencing method was prospectively analysed. RESULTS: The BRAF(V600E) mutation was associated with the following US features: solid composition [odds ratio (OR) 20·338; 95% confidence interval (CI): 4·952-83·532; P < 0·001], marked hypoechogenicity (OR 30·744; 95% CI: 15·951-59·255; P < 0·001), irregular margin (OR 9·889; 95% CI: 7·005-13·859; P < 0·001), taller-than-wide shape (OR 6·031; 95% CI: 4·343-8·376; P < 0·001) and the presence of microcalcifications (OR 6·664; 95% CI: 4·604-9·648; P < 0·001). The BRAF(V600E) mutation with malignant US features in FNAB enhanced the diagnostic accuracy compared with cytologic diagnosis alone (94·3%vs 69·7%). CONCLUSION: The BRAF(V600E) mutation is significantly associated with malignant US features, such as solid composition, marked hypoechogenicity, irregular margin, taller-than-wide shape and the presence of microcalcifications. The application of BRAF(V600E) mutation analysis in US-guided FNAB can improve the diagnostic accuracy of thyroid nodules.
Assuntos
Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias da Glândula Tireoide/diagnóstico por imagem , Nódulo da Glândula Tireoide/diagnóstico por imagem , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Substituição de Aminoácidos/genética , Biópsia por Agulha Fina , Carcinoma , Carcinoma Papilar , Análise Mutacional de DNA , Diagnóstico Diferencial , Feminino , Predisposição Genética para Doença , Ácido Glutâmico/genética , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/fisiologia , Sensibilidade e Especificidade , Câncer Papilífero da Tireoide , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , Nódulo da Glândula Tireoide/genética , Nódulo da Glândula Tireoide/patologia , Ultrassonografia , Valina/genética , Adulto JovemRESUMO
The cumbersome process required for diagnosis by DNA microarray can be simplified by simple extraction of nucleic acid from cells and by integration of liquid-phase polymerase chain reaction (PCR) and hybridization on the surface of a microarray slide. An unexpected benefit was the large (five- to sixfold) increase in detection signal that also is translated into an increase in sensitivity and the confidence level of diagnosis. The large increase in the detection signal appears to be due to participation of PCR primers as well as to extension of the immobilized capture probes during the hybridization process. The reason for the large increase in signal is not clear in view of only one round of DNA synthesis during the hybridization step. The integrated process correctly identified various genotypes of human papillomavirus (HPV) in the infected clinical human cervical specimens with specificity and efficiency. The process described in this article saves labor, time, and cost and should be applicable for automation of diagnosis by DNA microarray.
Assuntos
Análise de Sequência com Séries de Oligonucleotídeos/métodos , Reação em Cadeia da Polimerase/métodos , Antracenos/metabolismo , Sequência de Bases , Extratos Celulares , Linhagem Celular Tumoral , Colo do Útero/patologia , Colo do Útero/virologia , DNA/isolamento & purificação , Sondas de DNA/metabolismo , Feminino , Genoma Humano/genética , Genótipo , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/isolamento & purificação , Humanos , Dados de Sequência Molecular , Mutação/genética , Polimorfismo de Nucleotídeo Único/genética , Sensibilidade e Especificidade , Propriedades de SuperfícieRESUMO
While liver histopathology is heterogeneous in diabetes, the underlying mechanisms remain unclear. We investigated whether glycemic variation resulting from differential diets can induce heterogeneity in diabetic liver and the underlying molecular mechanisms. We generated end-stage non-obese diabetic model rats by subtotal-pancreatectomy in male Sprague- Dawley rats and ad libitum diet for 7 weeks (n = 33). The rats were then divided into three groups, and fed a standard- or a low-protein diet (18 or 6 kcal%, respectively), for another 7 weeks: to maintain hyperglycemia, 11 rats were fed ad libitum (18AL group); to achieve euglycemia, 11 were calorierestricted (18R group), and 11 were both calorie- and proteinrestricted with the low-protein diet (6R group). Overnightfasted liver samples were collected after the differential diets together with sham-control (18S group), and histology and molecular changes were compared. Hyperglycemic-18AL showed glycogenic hepatopathy (GH) without steatosis, with the highest GSK-3ß inactivation because of Akt activation during hyperglycemia; mitochondrial function was not impaired, compared to the 18S group. Euglycemic-18R showed neither GH nor steatosis, with intermediate GSK-3ß activation and mitochondrial dysfunction. However, euglycemic-6R showed both GH and steatosis despite the highest GSK-3ß activity and no molecular evidence of increased lipogenesis or decreased ApoB expression, where mitochondrial dysfunction was highest among the groups. In conclusion, heterogeneous liver histopathology developed in end-stage non-obese diabetic rats as the glycemic levels varied with differential diets, in which protein content in the diets as well as glycemic levels differentially influenced GSK-3ß activity and mitochondrial function in insulin-deficient state. [BMB Reports 2020; 53(2): 100-105].
Assuntos
Diabetes Mellitus Experimental/patologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Hiperglicemia/patologia , Fígado/patologia , Mitocôndrias/metabolismo , Animais , Apolipoproteínas B/genética , Apolipoproteínas B/metabolismo , Glicemia/metabolismo , Restrição Calórica , Diabetes Mellitus Experimental/dietoterapia , Diabetes Mellitus Experimental/metabolismo , Dieta da Carga de Carboidratos , Fígado Gorduroso/dietoterapia , Fígado Gorduroso/enzimologia , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Índice Glicêmico/fisiologia , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta/genética , Hepatócitos/enzimologia , Hepatócitos/metabolismo , Hepatócitos/ultraestrutura , Hiperglicemia/dietoterapia , Hiperglicemia/enzimologia , Hiperglicemia/metabolismo , Insulina/metabolismo , Lipogênese , Fígado/enzimologia , Fígado/metabolismo , Masculino , Mitocôndrias/patologia , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Thymidine phosphorylase (TP) has been reported to stimulate angiogenesis in a variety of human malignancies. We investigated TP expression and its association with angiogenesis in 73 cases of resected gastric cancer. In addition, we compared the expression of the other angiogenesis related factors (VEGF, eNOS and p53) with that of TP with respect to angiogenesis. TP expression was not detected in most of the non-tumoral glandular epithelial cells except for 5 cases. TP expression of the cancer cells and the stroma was assessed separately. The stromal TP expression was not associated with the TP expression of the cancer cells. The mean percent of TP reactive cancer cells was 18.36+/-2.61 (median, 10.00; range, 0-90) and cases showing a percentage higher than the mean were considered as bearing high reactivity. The mean microvessel score assessed was 90.44+/-3.69 (median, 86; range, 31-174). The TP expression of cancer cells was strongly associated with microvessel density (p=0.030), but the stromal TP expression was not. The microvessel density of the tumor showed strong correlation with VEGF expression (p<0.001), but a marginally significant association with eNOS (p=0.055). On the contrary, there was no association with p53 expression and microvessel density of the tumor. No significant correlation was detected between lymph node metastasis and tumoral or stromal TP expression or VEGF/TP coexpression. In gastric cancer, TP expression of the cancer cells, not stromal cells may play an important role in tumor growth by microvessel formation.
Assuntos
Neoplasias Gástricas/irrigação sanguínea , Neoplasias Gástricas/enzimologia , Timidina Fosforilase/biossíntese , Feminino , Humanos , Imuno-Histoquímica , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Neovascularização Patológica/enzimologia , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Óxido Nítrico Sintase Tipo III/biossíntese , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Proteína Supressora de Tumor p53/biossíntese , Fator A de Crescimento do Endotélio Vascular/biossínteseRESUMO
Hedgehog protein is essential to gastrointestinal tract development, and disruption of the hedgehog signaling pathway is associated with gastrointestinal tumorigenesis. Here, we analyzed the degree of hedgehog expression in gastric cancer and precancerous tissue. From August 2005 to May 2006, 52 gastric cancers and 16 gastric adenomas were obtained from surgically or endoscopically resected specimens. Immunohistochemical staining using sonic hedgehog (Shh) antibody was performed in cancerous and noncancerous tissue portions. Hedgehog expression was assessed based on the summed scores of the intensity and proportion of Shh staining. According to Lauren's classification, Shh expression was stronger in the intestinal type than in the diffuse type (p<0.001). Although Shh expression was not related to the location, size, metastatic status, or mucin phenotype of the gastric cancer, the expression was stronger in the tubular type of gastric cancer than in the mucinous and signet-ring cell types (p=0.001). Shh expression was stronger in gastric adenoma than in the diffuse-type gastric cancer (p<0.001), but revealed no difference with that of the intestinal-type gastric cancer (p=0.30). Shh expression was strongest in all types of intestinal metaplasia of noncancerous tissues. Shh expression is related to the intestinal type of gastric cancer. The stronger Shh expression in intestinal metaplasia and gastric adenoma indicates that hedgehog protein is involved at an early phase of gastric carcinogenesis.
Assuntos
Adenoma/metabolismo , Proteínas Hedgehog/biossíntese , Neoplasias Gástricas/metabolismo , Adenoma/patologia , Idoso , Feminino , Humanos , Imuno-Histoquímica , Mucosa Intestinal/patologia , Masculino , Metaplasia , Pessoa de Meia-Idade , Lesões Pré-Cancerosas/metabolismo , Lesões Pré-Cancerosas/patologia , Neoplasias Gástricas/patologiaRESUMO
The BRAF V600E mutation test has proven diagnostic value in thyroid fine-needle aspiration. The real-time polymerase chain reaction (RT-PCR) has recently been introduced as a new method for BRAF mutation detection. We performed BRAF V600E detection in 126 cases of thyroid fine-needle aspiration, using RT-PCR and pyrosequencing. BRAF V600E mutation was detected in 78 (61.9%) of 126 cases by RT-PCR and in 74 (57.8%) by pyrosequencing. Of the 98 papillary thyroid carcinoma samples, the BRAF V600E mutation was identified in 72 by RT-PCR and in 70 by pyrosequencing (sensitivities of 71.6% and 71.4%, respectively). Among 28 benign nodules, 6 false-positive cases were detected by RT-PCR, whereas 4 false-positive cases were detected by pyrosequencing (specificities of 78.6% and 85.7%, respectively). When analyzing 104 cases after excluding equivocal samples, pyrosequencing had a marginally higher specificity than RT-PCR (100% vs. 78.3%, P=0.074). After modifying the cut-off criteria, the low RT-PCR specificity improved to a similar or a slightly lower specificity compared with that of pyrosequencing. In the titration assay mixing the mutant DNA with the wild-type DNA in varying proportions, RT-PCR was sensitive enough to detect the mutation in a mixture containing 0.001% mutant DNA, whereas the limit of detection was 10% for pyrosequencing. In conclusion, compared with pyrosequencing, RT-PCR was more sensitive, faster, and more convenient, but less specific, for detecting the BRAF V600E mutation. A readjustment or modification of the interpretation criteria may be necessary to reduce false-positive RT-PCR results and improve the specificity while maintaining the sensitivity.
Assuntos
Carcinoma Papilar/diagnóstico , Carcinoma Papilar/genética , Mutação/genética , Proteínas Proto-Oncogênicas B-raf/genética , Reação em Cadeia da Polimerase em Tempo Real/normas , Análise de Sequência de DNA/normas , Neoplasias da Glândula Tireoide/diagnóstico , Neoplasias da Glândula Tireoide/genética , Biópsia por Agulha Fina , Humanos , Câncer Papilífero da TireoideRESUMO
BACKGROUND: The BRAFV600E mutation in papillary thyroid carcinoma (PTC) is particularly prevalent in Korea, and a considerable number of wild-type BRAF PTCs harbor RAS mutations. In addition, subsets of other genetic alterations clearly exist, but their prevalence in the Korean population has not been well studied. Recent increased insight into noninvasive encapsulated follicular variant PTC has prompted endocrine pathologists to reclassify this entity as "noninvasive follicular thyroid neoplasm with papillary-like nuclear features" (NIFTP). This study analyzed the genetic alterations among the histologic variants of PTC, including NIFTP. METHODS: Mutations of the BRAF and RAS genes and rearrangement of the RET/PTC1, NTRK1, and ALK genes using 769 preoperative fine-needle aspiration specimens and resected PTCs were analyzed. RESULTS: Molecular alterations were found in 687 (89.3%) of 769 PTCs. BRAFV600E mutation (80.8%) was the most frequent alteration, followed by RAS mutation and RET/PTC1, NTRK1, and ALK rearrangements (5.6%, 2.1%, 0.4%, and 0%, respectively). The low prevalence of NTRK1 fusions and the absence of an ALK fusion detected in Korea may also be attributed to the higher prevalence of the BRAFV600E mutation. There were significant differences in the frequency of the genetic alterations among the histologic variants of PTC. The prevalence of NIFTP in PTC was 2.7%, and among the NIFTPs, 28.6% and 57.1% harbored BRAF and RAS mutations, respectively. Clinicopathologic factors and mutational profiles between NIFTP and encapsulated follicular variant PTC with capsular invasion group were not significantly different. CONCLUSIONS: Genetic alterations in PTC vary among its different histologic variants and seem to be different in each ethnic group.
Assuntos
Carcinoma Papilar/epidemiologia , Carcinoma Papilar/genética , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias da Glândula Tireoide/epidemiologia , Neoplasias da Glândula Tireoide/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Variação Genética , Genoma Humano , Humanos , Hibridização in Situ Fluorescente , Masculino , Pessoa de Meia-Idade , Neoplasias/genética , Período Pré-Operatório , Prevalência , República da Coreia , Câncer Papilífero da Tireoide , Análise Serial de Tecidos , Adulto Jovem , Proteínas ras/genéticaRESUMO
BACKGROUND: In fine needle aspiration biopsy (FNAB) of thyroid nodules, LBC is adopted in most of the hospitals and clinics in Korea for its convenience. BRAF mutation test has been introduced as an important ancillary test, but its applicability has not been completely proven with LBC samples. METHODS: Five aspirates from thyroidectomy specimens were simultaneously processed into LBC and CS slides, in which BRAF mutation tests were performed using three primer sets with PCR products of 72, 164, and 226 base pairs (bp) at 6, 9, and 12 months after processing. In addition, BRAF mutation tests were performed in nine clinical samples that had been prepared by LBC or CS and stored for 3-5 years after processing. RESULTS: At 9 months after processing, LBC failed to provide DNA of sufficient quality for PCR, whereas CS succeeded with primers for amplifying a 226 bp fragment. Furthermore, CS had successful amplification of DNA despite a delay of more than 1 year. The failure of DNA amplification in LBC was overcome by using primers to amplify shorter PCR products, suggesting that DNA degradation occurred in LBC. However, false positive or negative results were observed in primers for amplifying shorter size. The kind of preservative solutions used in LBC did not affect test results. CONCLUSION: LBC may have disadvantages in long-term DNA preservation because of its accelerated DNA degradation compared with alcohol-fixed CS. Using primers to amplify shorter size fragments might be helpful in mitigating loss of signal due to DNA degradation in LBC.
Assuntos
DNA/química , Testes Genéticos/métodos , Teste de Papanicolaou/métodos , Proteínas Proto-Oncogênicas B-raf/genética , Nódulo da Glândula Tireoide/patologia , DNA/genética , Testes Genéticos/normas , Humanos , Hidrólise , Mutação , Teste de Papanicolaou/efeitos adversos , Sensibilidade e Especificidade , Análise de Sequência de DNA/métodos , Nódulo da Glândula Tireoide/genéticaRESUMO
Spermatogonial stem cells (SSCs), which are unipotent stem cells in the testes that give rise to sperm, can be converted into germline-derived pluripotent stem (gPS) by self-induction. The androgenetic imprinting pattern of SSCs is maintained even after their reprogramming into gPS cells. In this study, we used an in vitro neural differentiation model to investigate whether the imprinting patterns are maintained or altered during differentiation. The androgenetic patterns of H19, Snrpn, and Mest were maintained even after differentiation of gPS cells into NSCs (gPS-NSCs), whereas the fully unmethylated status of Ndn in SSCs was altered to somatic patterns in gPS cells and gPS-NSCs. Thus, our study demonstrates epigenetic alteration of genomic imprinting during the induction of pluripotency in SSCs and neural differentiation, suggesting that gPS-NSCs can be a useful model to study the roles of imprinted genes in brain development and human neurodevelopmental disorders.