RESUMO
Conservation over three mammalian genera-the mouse, rat, and human-has been found for a subset of the transcripts whose level differs between the adenoma and normal epithelium of the colon. Pde4b is one of the triply conserved transcripts whose level is enhanced both in the colonic adenoma and in the normal colonic epithelium, especially adjacent to adenomas. It encodes the phosphodiesterase PDE4B, specific for cAMP. Loss of PDE4B function in the ApcMin/+ mouse leads to a significant increase in the number of colonic adenomas. Similarly, Pde4b-deficient ApcMin/+ mice are hypersensitive to treatment by the inflammatory agent DSS, becoming moribund soon after treatment. These observations imply that the PDE4B function protects against ApcMin-induced adenomagenesis and inflammatory lethality. The paradoxical enhancement of the Pde4b transcript in the adenoma versus this inferred protective function of PDE4B can be rationalized by a feedback model in which PDE4B is first activated by early oncogenic stress involving cAMP and then, as reported for frank human colon cancer, inactivated by epigenetic silencing.
Assuntos
Adenoma/patologia , Neoplasias Colorretais/patologia , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Doenças Inflamatórias Intestinais/patologia , Adenoma/genética , Adenoma/mortalidade , Proteína da Polipose Adenomatosa do Colo/genética , Animais , Colo/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Feminino , Técnicas de Inativação de Genes , Humanos , Doenças Inflamatórias Intestinais/induzido quimicamente , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Ratos , Análise Serial de TecidosRESUMO
Current screening procedures for colorectal cancer are imperfect and highly invasive and result in increased mortality rates due to low compliance. The goal of the experiments reported herein is to identify potential blood-based biomarkers indicative of early stage intestinal cancers using the ApcMin/+ mouse model of intestinal cancer as an experimental system. Serum proteins from tumor-bearing ApcMin/+ mice were quantitatively compared to tumor-free Apc+/+ wild-type mice via in anima metabolic labeling with 14N/15N-labeled Spirulina algae and an LTQ Orbitrap mass spectrometer. Out of 1116 total serum proteins quantified, this study identified 40 that were differentially expressed and correlated with the increase in intestinal neoplasms. A subset of these differentially expressed proteins underwent a secondary quantitative screen using selected reaction monitoring-mass spectrometry with stable isotope-labeled peptides. Using both quantitative techniques, we identified MGAM and COL1A1 as downregulated and ITIH3 and F5 as upregulated in serum. All but COL1A1 were similarly differentially expressed in the mRNA of neoplastic colonic tissues of ApcMin/+ mice compared to normal wild-type tissue. These differentially expressed proteins identified in the ApcMin/+ mouse model have provided a set of candidate biomarkers for future validation screens in humans.
Assuntos
Biomarcadores Tumorais/sangue , Neoplasias Colorretais/sangue , Animais , Neoplasias Colorretais/diagnóstico , Detecção Precoce de Câncer , Feminino , Genes APC , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Isótopos de Nitrogênio , Mapas de Interação de Proteínas , Proteômica , Coloração e Rotulagem , Regulação para CimaRESUMO
Crop yields are significantly reduced by aluminum (Al) toxicity on acidic soils, which comprise up to 50% of the world's arable land. Al-activated release of ligands (such as organic acids) from the roots is a major Al tolerance mechanism in plants. In maize, Al-activated root citrate exudation plays an important role in tolerance. However, maize Al tolerance is a complex trait involving multiple genes and physiological mechanisms. Recently, transporters from the MATE family have been shown to mediate Al-activated citrate exudation in a number of plant species. Here we describe the cloning and characterization of two MATE family members in maize, ZmMATE1 and ZmMATE2, which co-localize to major Al tolerance QTL. Both genes encode plasma membrane proteins that mediate significant anion efflux when expressed in Xenopus oocytes. ZmMATE1 expression is mostly concentrated in root tissues, is up-regulated by Al and is significantly higher in Al-tolerant maize genotypes. In contrast, ZmMATE2 expression is not specifically localized to any particular tissue and does not respond to Al. [(14)C]-citrate efflux experiments in oocytes demonstrate that ZmMATE1 is a citrate transporter. In addition, ZmMATE1 expression confers a significant increase in Al tolerance in transgenic Arabidopsis. Our data suggests that ZmMATE1 is a functional homolog of the Al tolerance genes recently characterized in sorghum, barley and Arabidopsis, and is likely to underlie the largest maize Al tolerance QTL found on chromosome 6. However, ZmMATE2 most likely does not encode a citrate transporter, and could be involved in a novel Al tolerance mechanism.
Assuntos
Alumínio/toxicidade , Transportadores de Ânions Orgânicos/metabolismo , Proteínas de Plantas/metabolismo , Locos de Características Quantitativas , Zea mays/genética , Sequência de Aminoácidos , Animais , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Mapeamento Cromossômico , Clonagem Molecular , Regulação da Expressão Gênica de Plantas , Dados de Sequência Molecular , Oócitos , Transportadores de Ânions Orgânicos/genética , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , RNA de Plantas/genética , Alinhamento de Sequência , Xenopus , Zea mays/metabolismoRESUMO
Many studies of the response of colonic tumors to therapeutics use tumor multiplicity as the endpoint to determine the effectiveness of the agent. These studies can be greatly enhanced by accurate measurements of tumor volume. Here we present a quantitative method to easily and accurately determine colonic tumor volume. This approach uses a biocompatible alginate to create a negative mold of a tumor-bearing colon; this mold is then used to make positive casts of dental stone that replicate the shape of each original tumor. The weight of the dental stone cast correlates highly with the weight of the dissected tumors. After refinement of the technique, overall error in tumor volume was 16.9% ± 7.9% and includes error from both the alginate and dental stone procedures. Because this technique is limited to molding of tumors in the colon, we utilized the Apc(Pirc/+) rat, which has a propensity for developing colonic tumors that reflect the location of the majority of human intestinal tumors. We have successfully used the described method to determine tumor volumes ranging from 4 to 196 mm³. Alginate molding combined with dental stone casting is a facile method for determining tumor volume in vivo without costly equipment or knowledge of analytic software. This broadly accessible method creates the opportunity to objectively study colonic tumors over time in living animals in conjunction with other experiments and without transferring animals from the facility where they are maintained.