RESUMEN
Fibrillin microfibrils are polymeric structures present in connective tissues. The importance of fibrillin microfibrils to connective tissue function has been demonstrated by the multiple genetic disorders caused by mutations in fibrillins and in microfibril-associated molecules. However, knowledge of microfibril structure is limited, largely due to their insolubility. Most previous studies have focused on how fibrillin-1 is organized within microfibril polymers. In this study, an immunochemical approach was used to circumvent the insolubility of microfibrils to determine the role of fibrillin-2 in postnatal microfibril structure. Results obtained from studies of wild type and fibrillin-1 null tissues, using monoclonal and polyclonal antibodies with defined epitopes, demonstrated that N-terminal fibrillin-2 epitopes are masked in postnatal microfibrils and can be revealed by enzymatic digestion or by genetic ablation of Fbn1. From these studies, we conclude that fetal fibrillin polymers form an inner core within postnatal microfibrils and that microfibril structure evolves as growth and development proceed into the postnatal period. Furthermore, documentation of a novel cryptic site present in EGF4 in fibrillin-1 underscores the molecular complexity and tissue-specific differences in microfibril structure.
Asunto(s)
Microfibrillas/metabolismo , Proteínas de Microfilamentos/metabolismo , Secuencia de Aminoácidos , Amnios/química , Animales , Anticuerpos Monoclonales/inmunología , Embrión de Pollo , Ectodermo/embriología , Ectodermo/metabolismo , Epítopos/inmunología , Extremidades/embriología , Femenino , Fibrilina-1 , Fibrilina-2 , Fibrilinas , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Lactante , Masculino , Ratones , Ratones Noqueados , Microfibrillas/ultraestructura , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/inmunología , Microscopía Electrónica , Datos de Secuencia Molecular , Músculo Esquelético/química , Homología de Secuencia de Aminoácido , Piel/químicaRESUMEN
A definitive diagnosis of cancer may be rendered by microscopic assessment of only a few cells in an appropriate clinical setting due to the distinctive nuclear structure of most cancer cells in comparison to nuclei of normal human cells. The molecular architecture of non-neoplastic human nuclei--of the nuclear matrix and of matrix-associated proteins and nucleic acids--is being characterized in exquisite molecular detail. What is missing is the application of the findings and tools of molecular biology to understanding the cytological structure of cancer nuclei. This article delves into the basis of nuclear structure at different levels of resolution--light microscopic, electron microscopic, and molecular.
Asunto(s)
Núcleo Celular/patología , Neoplasias/patología , Núcleo Celular/ultraestructura , Forma del Núcleo Celular , Estructuras del Núcleo Celular/patología , Estructuras del Núcleo Celular/ultraestructura , Humanos , Microscopía , Neoplasias/diagnóstico , FenotipoRESUMEN
We used MethyLight assays to analyze DNA methylation status of 27 genes on 49 paired cancerous and noncancerous tissue samples from non-small cell lung cancer (NSCLC) patients who underwent surgical resection. Seven genes (RARB, BVES, CDKN2A, KCNH5, RASSF1, CDH13, and RUNX) were found to be methylated significantly more frequently in tumor tissues than in noncancerous tissues. Only methylation of CCND2 and APC was frequently detected in both cancerous and noncancerous tissues, supporting the hypothesis that the methylation of these two genes is a preneoplastic change and may be associated with tobacco smoking exposure. Methylation of any one of eight genes (RASSF1, DAPK1, BVES, CDH13, MGMT, KCNH5, RARB, or CDH1) was present in 80% of NSCLC tissues but only in 14% of noncancerous tissues. Detection of methylation of these genes in blood might have utility in monitoring and detecting tumor recurrence in early-stage NSCLC after curative surgical resection.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/genética , Metilación de ADN , Neoplasias Pulmonares/genética , Anciano , Antígenos CD , Proteínas Reguladoras de la Apoptosis/genética , Cadherinas/genética , Proteínas Quinasas Dependientes de Calcio-Calmodulina/genética , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico por imagen , Carcinoma de Pulmón de Células no Pequeñas/cirugía , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Proteínas Quinasas Asociadas a Muerte Celular , Canales de Potasio Éter-A-Go-Go/genética , Femenino , Fluorodesoxiglucosa F18 , Humanos , Neoplasias Pulmonares/diagnóstico por imagen , Neoplasias Pulmonares/cirugía , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Radiofármacos , Receptores de Ácido Retinoico/genética , Tomografía Computarizada de Emisión , Proteínas Supresoras de Tumor/genéticaRESUMEN
It remains unknown whether tobacco smoke induces DNA hypermethylation as an early event in carcinogenesis or as a late event, specific to overt cancer tissue. Using MethyLight assays, we analyzed 316 lung tissue samples from 151 cancer-free subjects (121 ever-smokers and 30 never-smokers) for hypermethylation of 19 genes previously observed to be hypermethylated in nonsmall cell lung cancers. Only APC (39%), CCND2 (21%), CDH1 (7%), and RARB (4%) were hypermethylated in >2% of these cancer-free subjects. CCND2 was hypermethylated more frequently in ever-smokers (26%) than in never-smokers (3%). CCND2 hypermethylation was also associated with increased age and upper lobe sample location. APC was frequently hypermethylated in both ever-smokers (41%) and never-smokers (30%). BVES, CDH13, CDKN2A (p16), CDKN2B, DAPK1, IGFBP3, IGSF4, KCNH5, KCNH8, MGMT, OPCML, PCSK6, RASSF1, RUNX, and TMS1 were rarely hypermethylated (<2%) in all subjects. Hypermethylation of CCND2 may reflect a smoking-induced precancerous change in the lung.
RESUMEN
Polycystic ovary syndrome (PCOS) is a common endocrinopathy in women of reproductive age. Although genetic linkage analyses have demonstrated a susceptibility locus for PCOS mapping to the fibrillin-3 gene, the presence of fibrillin proteins in normal and polycystic ovaries has not been characterized. This study compared and contrasted fibrillin-1, -2, and -3 localization in normal and polycystic ovaries. Immunohistochemical stainings of ovaries from 21 controls and 9 patients with PCOS were performed. Fibrillin-1 was ubiquitous in ovarian connective tissue. Fibrillin-2 localized around antral follicles and in areas of folliculolysis. Fibrillin-3 was present in a restricted distribution within the specialized perifollicular stroma of follicles in morphological transition from primordial to primary type [transitional follicles (TFs)]. Fibrillin-1 and -2 stainings of PCOS ovaries were similar to those of the controls. However, in eight of the nine PCOS ovaries, there was a decrease in the number of TFs associated with fibrillin-3, including no staining in five PCOS samples; decreased number of fibrillin-3-associated TFs/mm(2) was confirmed by quantitative analysis. Our findings support a role for fibrillin-3 in the pathogenesis of PCOS and suggest fibrillin-3 may function in primordial to primary follicle transition. We propose that loss of fibrillin-3 during folliculogenesis may be an important factor in PCOS pathogenesis.
Asunto(s)
Proteínas de Microfilamentos/metabolismo , Ovario/metabolismo , Síndrome del Ovario Poliquístico/metabolismo , Adolescente , Adulto , Femenino , Fibrilina-1 , Fibrilina-2 , Fibrilinas , Humanos , Persona de Mediana Edad , Folículo Ovárico/metabolismo , Células del Estroma/metabolismo , Adulto JovenRESUMEN
OBJECTIVE: Limited data exist to inform clinicians and patients as to the likelihood of long-term endometrial hyperplasia response to progestin therapy, especially for atypical hyperplasia. We evaluated women with complex and atypical endometrial hyperplasia, comparing those prescribed progestin with those not prescribed progestin. METHODS: This retrospective cohort study was conducted in 1985-2005 among women aged 18-88 years at an integrated health plan in Washington State. Women were ineligible if they achieved an outcome (endometrial carcinoma, hysterectomy, or both) within 8 weeks of hyperplasia diagnosis. Exposure was progestin use for at least 14 days by duration and recency. Outcomes included rate of 1) endometrial carcinoma, 2) hysterectomy, or 3) both. Analyses performed included Kaplan-Meier, incident rate ratios, and Cox proportional hazard ratios. RESULTS: One thousand four hundred forty-three eligible women were identified. One thousand two hundred one had complex (n=164 no progestin) and 242 had atypical (n=62 no progestin) hyperplasia. During follow-up, a median of 5.3 years (range 8 weeks to 20.8 years), 71 women were diagnosed with endometrial carcinoma (35 complex, 36 atypia) and 323 underwent hysterectomy (216 complex, 107 atypia). Among women with complex and atypical hyperplasia, rates of endometrial carcinoma among progestin users were 3.6 and 20.5 per 1,000 woman-years, respectively (compared with women who did not use progestin, 10.8 and 101.4). Among women with complex and atypical hyperplasia, rates of hysterectomy among progestin users were 23.3 and 61.4 per 1,000 woman-years, respectively (compared with women who did not use progestin, 55.1 and 297.3). CONCLUSION: Endometrial carcinoma risk is diminished approximately threefold to fivefold in women diagnosed with complex or atypical endometrial hyperplasia and dispensed progestin; hysterectomy risk is also decreased. LEVEL OF EVIDENCE: II.
Asunto(s)
Hiperplasia Endometrial/patología , Neoplasias Endometriales/epidemiología , Progestinas/farmacología , Adulto , Anciano , Estudios de Cohortes , Hiperplasia Endometrial/inducido químicamente , Neoplasias Endometriales/cirugía , Endometrio/efectos de los fármacos , Femenino , Humanos , Histerectomía/estadística & datos numéricos , Incidencia , Persona de Mediana Edad , Progestinas/efectos adversos , Estudios Retrospectivos , Factores de RiesgoRESUMEN
INTRODUCTION: We compared primary non-small cell lung cancer (NSCLC) F-fluorodeoxyglucose (FDG) uptake at positron emission tomography (PET) to tumor histologic features and Ki-67 proliferation index. This large, prospectively-recruited patient cohort has previously been analyzed based on differences in FDG uptake across stage groups; the current analysis adds further dimensions to this characterization. MATERIALS AND METHODS: One hundred seventy-eight patients with potentially-resectable NSCLC were scanned with FDG PET before therapy. A partial volume correction algorithm was used to correct FDG uptake values for their dependence on tumor size. Primary tumor resection specimens, core biopsies, and biopsies of metastatic lymph nodes were used to assess each tumor's NSCLC histologic subtype, degree of differentiation, and Ki-67 proliferation index. RESULTS: Bronchioalveolar carcinomas were found to have lower FDG uptake at PET and lower Ki-67 scores than any other histologic subtype. Non-bronchioalveolar adenocarcinomas had lower FDG uptake and Ki-67 scores than squamous cell carcinomas or large cell undifferentiated carcinomas. Better differentiated NSCLCs had lower FDG uptake and Ki-67 scores than more poorly differentiated NSCLCs. There was a significant positive correlation between FDG uptake and Ki-67 scores. Partial volume correction increased the strength of this correlation, while also diminishing the strong positive correlation between FDG uptake and tumor size. CONCLUSIONS: There are significant differences in NSCLC FDG uptake across histologic subtypes and differentiation groups. These differences parallel nearly identical differences in Ki-67 scores, implying that differences in NSCLC tumor cell proliferation may give rise to commensurate differences in tumor glucose metabolism.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Fluorodesoxiglucosa F18/farmacocinética , Antígeno Ki-67/metabolismo , Neoplasias Pulmonares/metabolismo , Tomografía de Emisión de Positrones , Radiofármacos/farmacocinética , Adenocarcinoma/diagnóstico por imagen , Adenocarcinoma/metabolismo , Adenocarcinoma/secundario , Adenocarcinoma Bronquioloalveolar/diagnóstico por imagen , Adenocarcinoma Bronquioloalveolar/metabolismo , Adenocarcinoma Bronquioloalveolar/secundario , Biomarcadores de Tumor/metabolismo , Carcinoma de Células Grandes/diagnóstico por imagen , Carcinoma de Células Grandes/metabolismo , Carcinoma de Células Grandes/secundario , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico por imagen , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Células Escamosas/diagnóstico por imagen , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/secundario , Estudios de Cohortes , Femenino , Humanos , Neoplasias Pulmonares/diagnóstico por imagen , Neoplasias Pulmonares/secundario , Masculino , Estadificación de Neoplasias , Pronóstico , Estudios Prospectivos , Radiografía Torácica , Tomografía Computarizada por Rayos XRESUMEN
OBJECTIVE: To validate the DNA microarray results on a subset of genes that could potentially serve as biomarkers of oral squamous cell carcinoma (OSCC) by examining their expression with an alternate quantitative method and by assessing their protein levels. DESIGN: Based on DNA microarray data from our laboratory and data reported in the literature, we identified 6 potential biomarkers of OSCC to investigate further. We used quantitative real-time polymerase chain reaction to examine expression changes of CDH11, MMP3, SPARC, POSTN, TNC, and TGM3 in OSCC and histologically normal control tissues. We further examined validated markers at the protein level by immunohistochemical analysis of OSCC tissue microarray sections. RESULTS: Quantitative real-time polymerase chain reaction analysis revealed upregulation of CDH11, SPARC, POSTN, and TNC gene expression and decreased TGM3 expression in OSCC tissue compared with control tissue; MMP3 was not found to be differentially expressed. In tissue microarray immunohistochemical analyses, SPARC (secreted protein, acidic, rich in cysteine), periostin, and tenascin C exhibited increased protein expression in tumor tissue compared with control tissue, and their expression was primarily localized within tumor-associated stroma rather than tumor epithelium. Conversely, transglutaminase 3 protein expression was found only within keratinocytes in control tissue and was significantly downregulated in cancer cells. CONCLUSIONS: Of 6 potential gene markers of OSCC, initially identified by DNA microarray analyses, differential expression of CDH11, SPARC, POSTN, TNC, and TGM3 were validated by quantitative real-time polymerase chain reaction. Differential expression and localization of proteins encoded by SPARC, POSTN, TNC, and TGM3 were clearly shown by tissue microarray immunohistochemical analysis.