RESUMEN
In situ hybridization analysis provides a means to qualitatively study the heterogeneity of primary tumors and metastases based on the types of genes transcribed. In this study, we have tested some parameters for quantitative analysis of in situ hybridizations with paraffin-embedded human breast tumors and measured mRNA levels for the angiogenic protein, vascular endothelial growth factor (VEGF). VEGF mRNAs were highly tumor specific, with the highest levels near necrotic regions within the tissues (0.1 to 2.7 dpm/mm2). Normal cells within the tissue sections did not have detectable levels of VEGF mRNA. For comparison, tumor levels of c-myc (4 to 46 dpm/mm2) and glyceraldehyde-3-phosphate dehydrogenase mRNAs (48 to 214 dpm/mm2) were measured. The mRNAs for both of these genes were more broadly expressed across the tissue sections. The hybridization pattern for VEGF mRNAs was consistent with hypoxia-induced VEGF mRNA steady-state levels and supports the hypothesis that oxidative stress regulates VEGF expression in breast tumors.
Asunto(s)
Neoplasias de la Mama/química , Factores de Crecimiento Endotelial/análisis , Ganglios Linfáticos/química , Linfocinas/análisis , ARN Mensajero/análisis , Biomarcadores de Tumor , Neoplasias de la Mama/patología , Cartilla de ADN/química , Factores de Crecimiento Endotelial/genética , Femenino , Gliceraldehído-3-Fosfato Deshidrogenasas/análisis , Humanos , Hibridación in Situ , Ganglios Linfáticos/patología , Metástasis Linfática , Linfocinas/genética , Proteínas Proto-Oncogénicas c-myc/análisis , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
In this study, we developed a rabbit polyclonal antibody to a fusion protein containing the envelope (env) transmembrane (TM) region from the human endogenous retroviral family, HERV-E. We used this reagent to document the expression of TM-related protein by Western blot in endothelial, colon and prostate carcinoma and seminoma cell lines and in peripheral blood mononuclear cells from a healthy donor. We detected a 58-kD protein (as compared to murine TM of 15 kD) that had specificity for the env-related antibodies of the polyclonal antiserum.
Asunto(s)
Productos del Gen env/análisis , Proteínas de Neoplasias , Neoplasias/química , Proteínas de los Retroviridae/análisis , Retroviridae , Proteínas del Envoltorio Viral/análisis , Adenocarcinoma/química , Animales , Formación de Anticuerpos , Especificidad de Anticuerpos , Neoplasias del Colon/química , Endotelio/química , Productos del Gen env/inmunología , Humanos , Masculino , Neoplasias de la Próstata/química , Conejos , Proteínas Recombinantes de Fusión/análisis , Proteínas Recombinantes de Fusión/inmunología , Proteínas de los Retroviridae/inmunología , Seminoma/química , Neoplasias Testiculares/química , Células Tumorales Cultivadas/química , Proteínas del Envoltorio Viral/inmunologíaRESUMEN
Tumor-induced immunosuppression by murine retrovirus-induced tumors and nonviral murine and human tumors has been shown to be mediated by the transmembrane (TM) envelope (env) protein p15E. This in vitro activity is inhibitable by anti-(murine)p15E antibodies, implying that a TM-like protein is produced by such tumors. The leading candidate genes that might encode such proteins in human tumors are human endogenous retroviral (HERV) sequences. We have utilized immunohistochemistry to determine what tissues may express HERV env proteins. We subcloned a restriction fragment from the putative TM human env gene of a type C-related HERV (clone-4-1) into a fusion protein gene construct. Using a rabbit polyclonal antiserum against the fusion protein, we observed staining in a variety of human tumor and nontumor tissues.
