RESUMEN
Clostridium difficile infection is a leading cause of antibiotic-related and healthcare-related diarrhoea. In the past decade, faecal microbiota transplantation or transfer has attracted increasing interest as an effective treatment strategy for severe recurrent C. difficile infection, with a global success rate of >80%. However, experience with this procedure is limited by a lack of randomized trials supporting its efficacy and the lack of standardization of the procedure. This review will address the practical aspects of the protocol.
Asunto(s)
Terapia Biológica/métodos , Clostridioides difficile/aislamiento & purificación , Infecciones por Clostridium/terapia , Infección Hospitalaria/terapia , Diarrea/terapia , Heces , Infecciones por Clostridium/microbiología , Infección Hospitalaria/microbiología , Diarrea/microbiología , Humanos , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
The yoghurt bacteria, Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus, are alleged to have beneficial effects on human health. The objective of this study was to characterise growth, biochemical activity and competitive behaviour of these two bacteria in vitro and in vivo. S. thermophilus LMD-9 and L. bulgaricus ATCC 11842 growth and lactate production were monitored in different media and in the gastrointestinal tract (GIT) of germ-free rats. In vitro, particularly in milk, S. thermophilus had a selective growth advantage over L. bulgaricus. The GIT of germ-free rats not supplemented with lactose was colonised by S. thermophilus but not by L. bulgaricus. Both bacteria were able to colonise the GIT of germ-free rats supplemented with 45 g/l lactose in their drinking water. However, if germ-free rats were inoculated with a mixture of the two bacteria and were supplemented with lactose, S. thermophilus rapidly and extensively colonised the GIT (1010 cfu/g faeces) at the expense of L. bulgaricus, which remained in most cases at levels <102 cfu/g faeces. S. thermophilus specifically produced L-lactate, while L. bulgaricus produced only D-lactate, both in vitro and in vivo. S. thermophilus showed competitive and growth advantage over L. bulgaricus in vitro as well as in vivo in the GIT of germ-free rats and, accordingly, L-lactate was the main lactate isomer produced.
Asunto(s)
Tracto Gastrointestinal/microbiología , Lactobacillus/crecimiento & desarrollo , Streptococcus thermophilus/crecimiento & desarrollo , Animales , Vida Libre de Gérmenes , Ácido Láctico/metabolismo , Masculino , Ratas , Ratas Endogámicas F344 , Yogur/microbiologíaRESUMEN
BACKGROUND: This study aimed to evaluate the implementation of a strategy to prevent postoperative nausea and vomiting (PONV) in patients undergoing general surgery. STUDY DESIGN: Prospective observational study. METHODS: A first period was observational. During a second period, a strategy to prevent PONV was based on five risk factors (RF) identified after the first phase. From two RF, antiemetic treatment was given according to the number of RF. The incidence of PONV was recorded in postoperative anaesthesic care unit (PACU) and at the 24th postoperative hour (24h). RESULTS: We prospectively enrolled 823 patients. Implementation of a prophylactic PONV strategy was associated with a decrease of nausea in PACU from 29.9 to 9.8% (P<0.001) and at 24h from 19 to 10.3% (P<0.001). Vomiting decreased from 12.4 to 2.3% (P<0.001) in PACU and from 5.6 to 3.7% at 24h (non-significant). CONCLUSION: Prophylaxis of PONV by the administration of antiemetic treatment according to a strategy based on a local risk score was efficient and associated with a significant decrease of PONV.
Asunto(s)
Antieméticos/uso terapéutico , Náusea y Vómito Posoperatorios/prevención & control , Femenino , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Náusea y Vómito Posoperatorios/epidemiología , Cuidados Preoperatorios , Estudios Prospectivos , Procedimientos Quirúrgicos OperativosRESUMEN
Aspirin consumption has been reported to be able to reduce colorectal cancer risk in humans and in animal models of colon carcinogenesis. Although the mechanism involved in such an effect is not yet clear, both prostaglandin-dependent and -independent effects have been proposed. Using HT-29 Glc(-/+)cells, which originate from a human colon adenocarcinoma, we demonstrated in this study a dose-dependent effect of millimolar concentration of aspirin on cell growth that was concomitant with a rapid accumulation of the cells in the G0/G1 phase, followed by an accumulation in the G2/M phase and by a minor increase in the proportion of cells undergoing nuclear condensation. Cell membrane integrity and cell release into the culture medium were not affected by this treatment. The aspirin effects were apparently unrelated to prostaglandin biosynthesis inhibition, since although these cells were found to express high levels of cyclooxygenase 1 (COX-1) and low levels of COX-2 proteins, they did not produce any measurable net amounts of prostaglandins, based on both utilization of radiolabelled arachidonic acid and the radioimmunoassay of prostaglandins E2 and F2 alpha. In contrast, we identified polyamine biosynthesis as a cellular target of aspirin, since the treatment of HT-29 Glc(-/+) cells with aspirin reduced the flux of L-ornithine through ornithine decarboxylase, an effect that could not be explained by an acute action of the drug on the ornithine decarboxylase catalytic activity. Since polyamine biosynthesis is strictly necessary for HT-29 cell growth, our data suggest that reduced flux through ornithine decarboxylase may participate in the antiproliferative activity of aspirin towards colonic tumoral cells. It is concluded that in HT-29 Glc(-/+) cells that are not functional for prostaglandin production, aspirin can affect cell growth, cell cycle, and polyamine biosynthesis without affecting cell membrane integrity.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Anticarcinógenos/farmacología , Aspirina/farmacología , Prostaglandinas/metabolismo , Putrescina/biosíntesis , Adenocarcinoma , Células CACO-2 , Ciclo Celular/efectos de los fármacos , Membrana Celular/efectos de los fármacos , Neoplasias del Colon , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Células HT29 , Humanos , Isoenzimas/biosíntesis , Proteínas de la Membrana , Ornitina/metabolismo , Poliaminas/metabolismo , Prostaglandina-Endoperóxido Sintasas/biosíntesisRESUMEN
Some colonic luminal molecules resulting from bacterial metabolism of alimentary or endogenous compounds are believed to exert various effects on the colonic epithelial cell physiology. We isolated surface epithelial cells and intact colonic crypts in order to test bacterial metabolites in the pig model, which is often considered relevant for extrapolation to the physiopathology of the human gastrointestinal tract. Using colonocytes isolated with EDTA, we found that the initial cell viability, estimated by the membrane integrity and oxidative capacity measurement, fell rapidly despite several experimental attempts to preserve it such as the use of a medium designed to increase the adherence of epithelial cells and of a coated extracellular matrix, the presence in the culture medium of the oxidative substrate butyrate, and the use of an inhibitor of the caspases involved in cell apoptosis. In contrast, using dispase and collagenase as proteolytic agents, we were able to obtain pig colonic crypts that maintain an excellent membrane integrity after 4 h. Using this preparation, we were able to test the presumably cytotoxic luminal compounds hydrogen sulfide, ammonia, and deoxycholic acid on colonic crypt viability. Of these, only deoxycholic acid was found to significantly alter the cellular membrane integrity. It is concluded that pig colonic crypts can be useful for the in vitro appraisal of the cytotoxic properties of luminal compounds.
Asunto(s)
Amoníaco/toxicidad , Membrana Celular/efectos de los fármacos , Colon/citología , Ácido Desoxicólico/toxicidad , Sulfuro de Hidrógeno/toxicidad , Animales , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Colon/efectos de los fármacos , Colon/ultraestructura , Células Epiteliales/ultraestructura , Mucosa Intestinal/metabolismo , Mucosa Intestinal/ultraestructura , PorcinosRESUMEN
Diallyl disulfide (DADS) is a major organosulphur compound present in garlic with an anti-mitotic potential against colon neoplastic lesions in vivo and colon tumour cell growth in vitro. Using the human colon adenocarcinoma HT-29 Glc(-/+) cell line we identified sub-populations of tumoural cells with markedly different characteristics in terms of metabolic capacities, adhesion properties and distribution in the cell cycle phases. After 1 and 2 days treatment with 100 microM DADS HT-29 cells were largely released into the culture medium. These floating cells accumulated in the G(2)/M phase and were characterized by a 5-fold reduction in cell capacity for de novo protein synthesis. Polyamine metabolism, which is necessary for intestinal epithelial cell attachment and growth, was also severely affected, since 3-fold reductions in polyamine biosynthesis and net accumulation of putrescine were measured after DADS treatment. However, oxidation of L-glutamine, the main precursor of the tricarboxylic acid cycle in these cells, and de novo synthesis of glutathione, a tripeptide involved in tumoural cell chemoresistance, were not affected by DADS treatment. In contrast, the adherent sub-population of HT-29 cells, although partially accumulated in G(2)/M phase, were characterized by unaffected metabolic capacities when compared with control cells except for putrescine accumulation, which was transiently decreased, and L-glutamine oxidation, which was increased 2-fold. DADS-resistant cells selected within 5 days were then able to proliferate at a similar rate to control untreated cells. The DADS-induced changes in HT-29 metabolic capacities, adhesion properties and the cell cycle are discussed from a causal perspective.
Asunto(s)
Adenocarcinoma/metabolismo , Compuestos Alílicos/farmacología , Adhesión Celular/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Neoplasias del Colon/metabolismo , Sulfuros/farmacología , Adenocarcinoma/patología , División Celular/efectos de los fármacos , Membrana Celular , Neoplasias del Colon/patología , Citometría de Flujo , Ajo/química , Células HT29 , Humanos , Plantas MedicinalesRESUMEN
In human colon carcinoma cells (HT-29 cells), L-arginine is the common precursor of L-ornithine which generates polyamines strictly necessary for cellular growth, and nitric oxide which has a strong antiproliferative activity. We show here that proliferative HT-29 cells possess the capacity for de novo synthesis of L-arginine from L-citrulline, but not from L-ornithine. L-Ornithine is apparently not an L-arginine precursor due to the absence of any detectable ornithine carbamoyltransferase activity. In contrast, the newly synthesized L-arginine was competent for urea and thus L-ornithine production in a context of a high putrescine production in the ornithine decarboxylase pathway and a low degradation of this polyamine in the diamine oxidase pathway. However, cells grown in an arginine-free culture medium containing added L-citrulline were unable to reach confluency. Furthermore, the low amount of nitric oxide produced from L-arginine by these cells was apparently not involved in the control of cell growth since inhibition of nitric oxide synthase activity was without effect. On the other hand, the capacity of more differentiated and less proliferative HT-29 cells for de novo L-arginine synthesis from L-citrulline was increased. It is concluded that L-citrulline is a precursor of L-arginine and L-ornithine in proliferative HT-29 cells and that the metabolic fate of L-ornithine in these cells is mainly devoted to polyamine synthesis. The similarity between differentiated HT-29 cells and the enterocytes of newborn animals in terms of L-arginine metabolism is finally discussed.
Asunto(s)
Arginina/biosíntesis , Citrulina/metabolismo , Neoplasias del Colon/metabolismo , Ornitina/biosíntesis , Ornitina/metabolismo , Animales , Animales Recién Nacidos , Diferenciación Celular , División Celular , Neoplasias del Colon/ultraestructura , Inhibidores Enzimáticos/farmacología , Humanos , Mucosa Intestinal/metabolismo , Intestinos/citología , Microscopía Electrónica , Óxido Nítrico Sintasa/antagonistas & inhibidores , Nitroarginina/farmacología , Porcinos , Células Tumorales CultivadasRESUMEN
Ornithine decarboxylase (ODC, EC 4.1.1.17) is the enzyme responsible for the synthesis of polyamines, which are absolutely necessary for cell proliferation. In the present work, we tested the effects of 3 nitric oxide (NO) donors, namely, sodium nitroprusside (SNP), (Z)-1-(N-methyl-N-[6-(N-methylammoniohexyl)amino] diazen-1-ium-1,2-diolate (MAHMA/NO) and 1,1-diethyl-2-hydroxy-2-nitroso-hydrazine sodium (DEA/NO), on ODC activity in human-colon carcinoma cells (HT-29). SNP was the most effective inhibitor of ODC activity with a concentration of 8 micromol/L inducing 50% inhibition of basal activity. The effect of SNP was reversed by haemoglobin (Hb), but not by GSH or L-cysteine (CYS). Very little of the SNP in solution was degraded into nitrite, but the presence of cellular homogenate increased the production of nitrite. MAHMA/NO and DEA/NO were much less effective than SNP as ODC inhibitors, since the concentrations of these agents which induce 50% inhibition of basal activity were 20- to 60-fold higher than that of SNP. The effects of MAHMA/NO and DEA/NO were not reversed by haemoglobin. In solution, these latter 2 agents were totally degraded into nitrites. In conclusion, SNP on the one hand and MAHMA/NO and DEA/NO on the other appeared to release different NOx species with different efficiency on ODC activity.
Asunto(s)
Hidrazinas/farmacología , Óxido Nítrico/metabolismo , Nitroprusiato/farmacología , Inhibidores de la Ornitina Descarboxilasa , Relación Dosis-Respuesta a Droga , Células HT29 , Humanos , Nitritos/metabolismo , Óxidos de NitrógenoRESUMEN
Alpha-difluoromethylornithine (DFMO) is commonly used as a specific ornithine decarboxylase (ODC, EC4.1.1.17) irreversible inhibitor. ODC is the enzyme responsible for polyamine biosynthesis, which has been shown to be strictly necessary for cell proliferation. In HT-29 Glc-/+ cells, L-arginine is the major precursor of these molecules through the sequential actions of arginase, which leads to L-ornithine generation and ODC. L-ornithine, a substrate for ODC, retroinhibits arginase. Since DFMO is an ornithine analogue, we searched for a direct effect of this agent upon arginase. The flux of L-arginine through arginase in intact cells was inhibited by 51+/-11% by 10 mM of DFMO whereas 10 mM of L-valine, a known potent arginase inhibitor, inhibited this flux by 73+/-6%. DFMO equilibrated between extracellular and intercellular spaces and, when used at 10-mM concentration, was without effect on L-arginine net uptake. Measurement of arginase activity in HT-29 cell homogenates with increasing concentrations of DFMO and L-arginine led to an inhibition with a calculated Ki (inhibitory constant) equal to 3.9+/-1.0 mM. L-ornithine was less effective than DFMO in inhibiting arginase activity. Bovine liver arginase, used as another source of the enzyme, was also severely inhibited by DFMO. The inhibitory effect of DFMO upon arginase, one step upstream of the ODC reaction in the metabolic conversion of L-arginine to polyamines, is of potential physiological importance, since it could alter the production of ornithine and thus its metabolism in pathways other than the ODC pathway.
Asunto(s)
Arginasa/antagonistas & inhibidores , Eflornitina/farmacología , Inhibidores Enzimáticos/farmacología , Animales , Antineoplásicos/farmacología , Arginina/metabolismo , Bovinos , Células HT29 , Humanos , Hígado/enzimología , Ornitina/metabolismo , Inhibidores de la Ornitina DescarboxilasaRESUMEN
HT-29 cells, originating from a human colon carcinoma, can proliferate in standard culture conditions with an absolute requirement for polyamines. The major precursor provided in the culture medium for polyamine biosynthesis is L-arginine. L-Arginine conversion to L-ornithine by arginase is followed by stepwise conversion of this latter amino acid to putrescine, spermidine and spermine. The aim of the present work was to document the consequences of a total inhibition of L-arginine flux through arginase, resulting in a decreased L-ornithine availability, on HT-29 cell proliferation and polyamine metabolism. L-Valine, a known arginase inhibitor, when used at a high concentration, i.e., 100 mM, inhibits L-arginine flux through arginase almost totally. The addition in the culture medium of 100 mM L-valine or 50 mM NaCl used to mimic the L-valine induced increase in medium osmolality both reduced equally cellular growth. Cell viability, protein synthesis or oxidative metabolism measured in isolated cells were unaffected by the L-valine treatment, suggesting that decreased proliferation was not associated with an acute toxic effect of this aminoacid, but was rather due to the increase in the medium osmolality. L-Valine treated cells displayed an altered polyamine metabolism when compared with control cells grown in the absence of the amino acid. After 4 days of treatment with 100 mM L-valine, L-ornithine flux through ornithine decarboxylase was significantly higher as well as putrescine and spermidine cellular uptakes in treated cells. However, the changes in polyamine metabolism led to similar polyamine cell contents in untreated and L-valine treated cells. In conclusion, we propose that the observed alterations of polyamine metabolism may reflect an adaptative response of HT-29 cells to the presence of L-valine which contribute together with the low amount of L-ornithine present in the culture medium to polyamine homeostasis.
Asunto(s)
División Celular/efectos de los fármacos , Células HT29/efectos de los fármacos , Poliaminas/metabolismo , Valina/farmacología , Arginina/metabolismo , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Glutamina/metabolismo , Humanos , Ornitina/deficiencia , Ornitina/metabolismo , Ornitina Descarboxilasa/metabolismo , Poliaminas/análisis , Poliaminas/farmacocinética , Biosíntesis de Proteínas , Putrescina/farmacocinética , Espermidina/farmacocinéticaRESUMEN
In human colon carcinoma HT-29 Glc(-/+) cells, L-arginine is the common precursor of polyamines which are absolutely necessary for cellular proliferation and nitric oxide (NO) with reported anti-proliferative activity. The aim of the present work was to test the effect of the NO donor sodium nitroprusside (SNP) on polyamine synthesis and cellular growth in HT-29 cells. SNP in the micromolar range inhibits cellular putrescine synthesis and this effect is greatly reversed by haemoglobin, supporting the view that the effect of SNP is related to the generation of NO. This corresponds to the inhibition by SNP of ornithine decarboxylase activity. Furthermore, SNP inhibits cellular proliferation. The effect of SNP is reversed by haemoglobin after 2 days of treatment but not after 4 days. Although no acute toxic effect of SNP was detected after 90 min incubation, it greatly enhanced the cellular death rate after several days in culture as estimated by the LDH leakage test. In conclusion, our data raise the possibility of an inhibitory interrelationship between NO and polyamine metabolic pathways. NO induced inhibition of putrescine synthesis and growth in HT-29 cells is discussed from a causal perspective.
Asunto(s)
Mucosa Intestinal/citología , Nitroprusiato/farmacología , Putrescina/biosíntesis , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Epiteliales , Epitelio/efectos de los fármacos , Epitelio/metabolismo , Células HT29 , Hemoglobinas/farmacología , Humanos , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Putrescina/farmacologíaRESUMEN
We sequenced a 4.8 kb BamHI swine genomic fragment comprising the entire 21-hydroxylase gene (CYP21) and its 5' and 3' flanking segments. The CYP21 coding sequence spanned 3050 bp and as in other species, comprised 10 exons separated by the corresponding introns. The deduced protein corresponded to 492 amino acid residues, 8 of which differed from a previously sequenced swine CYP21 enzyme. The 5' flanking region displayed several putative cis-acting elements which may be involved in either constitutive or cyclic adenosine 3',5'-monophosphate (cAMP) dependent transcriptional expression. We also characterized within the 5' region a 139 bp repetitive element of the short interspersed nucleotide element (SINE) family located on the opposite strand. In addition, we characterized the last five exons of a human-like opposite strand gene (OSG/X) located in the swine at the 3' end of CYP21. The sequenced part of this OSG/X displayed a very strong homology with its human counterpart.
Asunto(s)
Genes Reguladores , Esteroide 21-Hidroxilasa/genética , TATA Box , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Codón , ADN/genética , ADN/aislamiento & purificación , Humanos , Datos de Secuencia Molecular , ARN Mensajero/genética , Secuencias Repetitivas de Ácidos Nucleicos , Mapeo Restrictivo , Homología de Secuencia de Ácido Nucleico , PorcinosRESUMEN
1. The efficiency and time course of dietary fatty acid incorporation into lipids of egg yolk and abdominal adipose tissue was compared in "White Leghorn", normal (Dw) and dwarf (dw) laying hens at 56 weeks of age, using 14C labelled linoleic acid. 2. The sex-linked dwarfing gene, dw, was shown to reduce not only body weight and abdominal fat pad deposition, but also yolk production and the average clutch size. 3. Higher peak incorporation and total recovery of the linoleic acid radioactivity into yolk lipids, but lower label recovery into adipose tissue triglycerides were found in dwarf hens. 4. The higher esterification of the dietary linoleic acid in its native form into dwarf yolk triglycerides indicates that dwarf hens use more dietary lipids to synthesise yolk lipids but these results also suggest that the dw allele might reduce the lipogenic capacities of the liver and adipose tissue in laying hens.
Asunto(s)
Pollos/genética , Grasas de la Dieta/metabolismo , Enanismo/veterinaria , Lípidos/biosíntesis , Enfermedades de las Aves de Corral/genética , Tejido Adiposo/crecimiento & desarrollo , Tejido Adiposo/metabolismo , Animales , Peso Corporal/genética , Enanismo/genética , Yema de Huevo , Ácidos Grasos/metabolismo , Femenino , Ligamiento Genético , Genotipo , Ácidos Linoleicos/metabolismo , Hígado/crecimiento & desarrollo , Hígado/metabolismo , Tamaño de los Órganos , Triglicéridos/biosíntesis , Cromosoma XRESUMEN
The levels and fatty acid composition of lipids were determined in very low density lipoproteins (VLDL, d less than 1.006), yolk and abdominal adipose tissue of normal (Dw) and sex-linked dwarf (dw) White Leghorn laying hens. Effects of adding 4% tallow to the diet were also examined. In 40-wk-old hens, neither plasma lipids (triglycerides, phospholipids and cholesterol), VLDL levels, nor the chemical composition of VLDL was altered by the dw gene or dietary fat. Dwarfism reduced egg and yolk weights. Though the yolk lipid content was similar in normal and dwarf hens, yolk from dwarfs had slightly more phospholipids and less triglycerides than yolk from normal hens. Higher linoleic acid [18:2(n-6)] and lower oleic acid [18:1(n-9)] levels were observed in triglycerides of VLDL, yolk and adipose tissue from dwarf hens. In addition, the dietary fatty acid pattern had a greater influence on the fatty acid composition of the yolk lipid major precursors (VLDL triglycerides) in dwarf laying hens than in normal hens. These results suggest that the dwarfing gene might reduce the hepatic de novo fatty acid synthesis and/or dwarf hens might incorporate more dietary lipids into yolk than do normal hens.
Asunto(s)
Tejido Adiposo/análisis , Pollos/genética , Grasas de la Dieta/farmacología , Enanismo/genética , Yema de Huevo/análisis , Regulación de la Expresión Génica , Genes Recesivos/fisiología , Lípidos/genética , Cromosomas Sexuales/fisiología , Animales , Colesterol/análisis , Colesterol/genética , Enanismo/metabolismo , Ácidos Grasos/análisis , Ácidos Grasos/biosíntesis , Ácidos Grasos/genética , Femenino , Lípidos/sangre , Lipoproteínas VLDL/análisis , Lipoproteínas VLDL/genética , Hígado/metabolismo , Ovulación/genética , Fosfolípidos/análisis , Fosfolípidos/genética , Triglicéridos/análisis , Triglicéridos/genéticaRESUMEN
1. In vitro activities of glucose oxidation, de novo lipogenesis and lipolysis were compared in normal (Dw) and dwarf (dw) laying hens. 2. Dwarfism reduced the hepatic glucose oxidation while de novo lipogenesis was not altered. As liver weight was depressed, total liver lipogenesis capacity was probably reduced by dwarfism. 3. As compared to normal hens, de novo lipogenesis and basal or stimulated lipolysis were lower in dwarf adipose tissue while its lipid content was enhanced in dwarfs. 4. Results suggest that in laying hens dwarfism reduces the adipose tissue lipid mobilization but probably also the liver de novo lipogenesis.