RESUMEN
BACKGROUND: BRCA-1/2 germline mutations are responsible for early onset breast cancer and familial association. The underlying causes of the characteristic phenotypic behavior are not completely understood, but mammary stem cells appear to have a key role in this process. MATERIALS AND METHODS: We have investigated the presence of mammary stem / progenitor cells in normal tissues and in tumor tissues obtained from women with and without BRCA1/2 germline mutations by utilizing ALDH-1 immunohistochemistry. RESULTS: Isolated ALDH-1 positive cells were found in 15/28 (54%) of breast cancer samples from women with BRCA 1 or 2 mutations and in 33 /51 (65%) of matched sporadic breast cancer cases (p=0.5949, Chi Square test). While mammary stem cells were also detected in non-malignant breast lesions, only 41% of the tissues contained ALDH-1 positive cells (p=0.0371, Chi Square test). In patients with BRCA germline mutations ALDH-1 positive cells were more common in p53 positive (p=0.0028, Chi Square test) tumors, in high grade (p=0.0796), and in larger tumors (p=0.0604), while no such association was seen in sporadic cancer cases. In our patients, the expression of ALDH-1 positive cells in breast cancer was neither associated with disease-free and overall survival, nor time to metastasis. CONCLUSION: Breast cancers from BRCA mutation carriers do not harbor more ALHD-1 positive cells than sporadic tumors, and their more aggressive phenotype can thus not be explained by an increased stem cell pool. The presence of ALDH-1 in normal breast tissue suggests that additional factors determine the biological behavior of mammary stem cells.
Asunto(s)
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias de la Mama/genética , Mutación de Línea Germinal/genética , Células Madre Neoplásicas/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Familia de Aldehído Deshidrogenasa 1 , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Estudios de Cohortes , Femenino , Humanos , Isoenzimas/biosíntesis , Isoenzimas/genética , Persona de Mediana Edad , Células Madre Neoplásicas/patología , Retinal-Deshidrogenasa/biosíntesis , Retinal-Deshidrogenasa/genéticaRESUMEN
A carboligation was investigated for the first time as an enzymatic gas phase reaction, where benzaldehyde was converted to benzoin using thiamine diphosphate (ThDP)-dependent enzymes, namely benzaldehyde lyase (BAL) and benzoylformate decarboxylase (BFD). The biocatalyst was immobilized per deposition on non-porous support. Some limitations of the gas/solid biocatalysis are discussed based on this carboligation and it is also demonstrated that the solid/gas system is an interesting tool for more volatile products.
Asunto(s)
Benzaldehídos/química , Biodegradación Ambiental , Tiamina Pirofosfato/metabolismo , Aldehído-Liasas/metabolismo , Enzimas/metabolismo , Estudios de Factibilidad , Gases , Espectrometría de FluorescenciaRESUMEN
This paper presents a strategy for preparing an efficient immobilised alcohol dehydrogenase preparation for a gas-phase reaction. The effects of additives such as buffers and sucrose on the immobilisation efficiency (residual activity and protein loading) and on the gas-phase reaction efficiency (initial reaction rate and half-life) of Thermoanaerobacter sp. alcohol dehydrogenase were studied. The reduction of acetophenone to 1-phenylethanol under in situ cofactor regeneration using isopropanol as co-substrate was used as a model reaction at fixed reaction conditions (temperature and thermodynamic activities). A strongly enhanced thermostability of the enzyme in the gas-phase reaction was achieved when the enzyme was immobilised with 50 mM phosphate buffer (pH 7) containing sucrose five times the protein amount (on weight/weight basis). This resulted in a remarkable productivity of 200 g L(-1) day(-1) even at non-optimised reaction conditions. The interaction of additives with the enzyme and water affects the immobilisation and gas-phase efficiencies of the enzyme. However, it was not possible to predict the effect of additives on the gas-phase reaction efficiency even after knowing their effect on the immobilisation efficiency.
Asunto(s)
Alcohol Deshidrogenasa/metabolismo , Biotecnología/métodos , Enzimas Inmovilizadas/metabolismo , Thermoanaerobacter/enzimología , Acetofenonas/metabolismo , Adsorción , Alcohol Deshidrogenasa/química , Alcoholes Bencílicos/metabolismo , Reactores Biológicos , Catálisis , Estabilidad de Enzimas , Enzimas Inmovilizadas/química , Concentración de Iones de Hidrógeno , Cinética , Microesferas , Sacarosa/metabolismoRESUMEN
Confocal laser scanning microscopy was applied to measure the pH value in the carrier of immobilized enzymes during the enzyme-catalyzed synthesis. pH profiles with a high resolution are shown, with the pH increasing in the core of the particles. Significant differences occur for different carrier material, particle size, porosity and surface modification. The increased pH value is identified as one of the reasons leading to reduced enzyme selectivity in the penicillin amidase catalyzed synthesis of cephalosporins and penicillins.
