The EMA assessment of avapritinib in the treatment of gastrointestinal stromal tumours harbouring the PDGFRA D842V mutation.

Avapritinib is a protein kinase inhibitor designed to selectively inhibit oncogenic KIT and platelet-derived growth factor receptor alpha (PDGFRA) mutants by targeting the active conformation of the kinase. On 24 September 2020, a marketing authorisation valid through the European Union was issued for avapritinib as treatment of adult patients with unresectable or metastatic gastrointestinal stromal tumours (GIST) harbouring the PDGFRA D842V mutation. The drug was evaluated in an open-label, phase I, first-in-human, dose-escalation, open-label study to evaluate the safety, tolerability, pharmacokinetics, pharmacodynamics, and efficacy of avapritinib in adults with unresectable or metastatic GIST. The benefit of avapritinib was observed in patients with GIST harbouring the PDGFRA D842V mutation. The overall response rate was 95% (95% confidence interval 82.3%-99.4%), with a median duration of response of 22.1 months (95% confidence interval 14.1-not estimable months). The most common adverse events were nausea, fatigue, anaemia, periorbital and face oedema, hyperbilirubinaemia, diarrhoea, vomiting, increased lacrimation, and decreased appetite. Most of the reported cognitive effects were mild memory impairment. Rarer events were cases of severe encephalopathy and intracranial or gastrointestinal bleeding. The aim of this manuscript is to summarise the scientific review of the application leading to regulatory approval in the European Union.

Novel giardicidal compounds bearing proton pump inhibitor scaffold proceeding through triosephosphate isomerase inactivation.

Giardiasis is a worldwide parasitic disease that affects mainly children and immunosuppressed people. Side effects and the emergence of resistance over current used drugs make imperative looking for new antiparasitics through discovering of new biological targets and designing of novel drugs. Recently, it has determined that gastric proton-pump inhibitors (PPI) have anti-giardiasic activity. The glycolytic enzyme, triosephosphate isomerase (GlTIM), is one of its potential targets. Therefore, we employed the scaffold of PPI to design new compounds aimed to increase their antigiardial capacity by inactivating GlTIM. Here we demonstrated that two novel PPI-derivatives (BHO2 and BHO3), have better anti-giardiasic activity than omeprazole in concentrations around 120-130 µM, without cytotoxic effect on mammal cell cultures. The derivatives inactivated GlTIM through the chemical modification of Cys222 promoting local structural changes in the enzyme. Furthermore, derivatives forms adducts linked to Cys residues through a C-S bond. We demonstrated that PPI can be used as scaffolds to design better antiparasitic molecules; we also are proposing a molecular mechanism of reaction for these novel derivatives.

The A2B adenosine receptor colocalizes with adenosine deaminase in resting parietal cells from gastric mucosa.

The A2B adenosine receptor (A2BR) mediates biological responses to extracellular adenosine in a wide variety of cell types. Adenosine deaminase (ADA) can degrade adenosine and bind extracellularly to adenosine receptors. Adenosine modulates chloride secretion in gastric glands and gastric mucosa parietal cells. A close functional link between surface A2BR and ADA has been found on cells of the immune system, but whether this occurs in the gastrointestinal tract is unknown. The goal of this study was to determine whether A2BR and ADA are coexpressed at the plasma membrane of the acid-secreting gastric mucosa parietal cells. We used isolated gastric parietal cells after purification by centrifugal elutriation. The membrane fraction was obtained by sucrose gradient centrifugation. A2BR mRNA expression was analyzed by RT-PCR. The surface expression of A2BR and ADA proteins was evaluated by Western blotting, flow cytometry and confocal microscopy. Our findings demonstrate that A2BR and ADA are expressed in cell membranes isolated from gastric parietal cells. They show a high degree of colocalization that is particularly evident in the surface of contact between parietal cells. The confocal microscopy data together with flow cytometry analysis suggest a tight association between A2BR and ADA that might be specifically linked to glandular secretory function.

Basolateral expression of GRP94 in parietal cells of gastric mucosa.

GRP94 is a member of the heat shock protein family normally confined to the endoplasmic reticulum that sometimes escapes the KDEL-mediated retention system. It is overexpressed in some gastric and other gastrointestinal carcinomas, but little is known about the physiological role of GRP94 in gastric mucosa. We investigated the membrane presence of GRP94 in parietal cells, which secrete acid into the gastric lumen, using subcellular fractionation, selective solubilization of membrane proteins, Western blotting, and radio-ligand binding and provided evidence of functional GRP94 expression at the surface of gastric mucosa parietal cells anchored to the basolateral domain. Our results show that GRP94 is not an integral membrane protein since 50 mM Na2CO3 treatment dissociates part of it from the membrane. However, 100 mM Na2CO3 treatment did not extract all GRP94 from the membrane, which indicates that it is strongly associated with it. The presence of GRP94 in isolated plasma membrane was demonstrated by Western blotting and its functionality by radio-ligand binding experiments. Both the K(D) value obtained in saturation experiments with N-ethylcarboxamido-[3H]adenosine at 4°C, at the nanomolar range, and the inhibition constant of its binding by radicicol, the most specific GRP94 inhibitor, indicate that active receptor regions are exposed at the membrane surface. Western blotting of plasma membrane subfractions showed that GRP94 is mainly expressed in the basolateral membrane of gastric parietal cells, while its presence in the apical domain is negligible, thereby inferring a role for GRP94 in processes operating in this membrane domain.

Matrix-assisted laser desorption ionization imaging mass spectrometry in lipidomics.

The relevant structural, energetics, and regulatory roles of lipids are universally acknowledged. However, the high variability of lipid species and the large differences in concentrations make unraveling the role played by the different species in metabolism a titanic task. A recently developed technique, known as imaging mass spectrometry, may shed some light on the field, as it enables precise information to be obtained on the location of lipids in tissues. A review of the state of the art of the technique is presented in this manuscript, including detailed analysis of sample-preparation steps, data handling, and the identification of the species mapped so far.

Cáncer de mama en mujeres posmenopáusicas / No disponible

Objetivo Revisar nuestra experiencia en el manejo de pacientes posmenopáusicas diagnosticadas y tratadas de cáncer de mama. Material y métodos Se ha realizado estudio descriptivo retrospectivo de 903 pacientes con cáncer de mama entre 1992 y 2008. Se seleccionó a las pacientes posmenopáusicas: 568 (62,90%). Se analizaron factores como edad, paridad, antecedentes familiares y personales, tipo de cáncer, tipo de cirugía, resultados a.p., estadio, tratamientos complementarios y seguimiento. Se ha realizado estudio estadístico mediante SPSS 15.0.ResultadosLa edad media fue 65,26±0,46 (40–95) años. Solo en 117 (20,59%) pacientes existían antecedentes familiares de cáncer de mama. En 312 casos (54,92%) existían factores de riesgo epidemiológico. Eran nuligestas 55 pacientes (9,68%). El motivo de consulta más frecuente fue la palpación de un nódulo 328 (57,7%). Se emplearon diferentes métodos diagnósticos por imagen, siendo la mamografía 420 (73,9%) el más utilizado. En 238 (41,90%) fue necesario estudio mediante biopsia intraoperatoria para confirmación histológica del diagnóstico. El tratamiento neoadyuvante fue empleado en 63 casos (11,09%). El tratamiento quirúrgico como tratamiento inicial fue realizado en 505 casos (88,90%). Pudo realizarse cirugía conservadora en 225 (39,61%) casos. El tipo histológico más frecuente fue el carcinoma ductal infiltrante en 380 (66,9%) pacientes. El número de ganglios fue 12,41±0,26(1–36). La hormonoterapia se pautó en 333 (58,62%), siendo el tamoxifeno el más empleado, en 230 (69,1%).Conclusiones El incremento de la edad conlleva un aumento de la patología oncológica mamaria. El diagnóstico en estas pacientes es mayoritariamente clínico. Los tratamientos en estas pacientes son menos conservadores debido al diagnóstico en estadios más avanzados (AU)

Objective To review our experience in the management and treatment of postmenopausal women with breast cancer. Material and methods A descriptive and retrospective study was performed in 903 women with breast cancer between 1992 and 2008. A total of 568 (62.90%) menopausal women were selected. The factors studied included age, parity, familial and personal history, type of cancer, type of surgery, pathological findings, stage, complementary treatments and follow-up. Statistical analysis was performed using the SPSS statistical package version 15.0.ResultsThe mean age was 65.26±0.46 (40–95) years. Familial breast cancer was found in only 117 patients (20.59%) patients. Epidemiological risk factors were found in 312 (54.92%). Nulliparity was found in only 55 patients (9.68%). The most frequent reason for consultation was palpation of a nodule in 328 (57.7%). Distinct imaging procedures were used, the most frequent being mammography in 420 (73.9%). Intraoperative biopsy was required for histological confirmation of the diagnosis in 238 (41.90%). Neoadjuvant treatment was indicated in 63 patients (11.09%). Surgical treatment was the first step in 505 women (88.90%). Conservative surgery was feasible in 225 (39.61%) patients. The most frequent histological type was infiltrating ductal carcinoma cancer in 380 (66.9%) patients. The mean number of nodes was 12.41±0.26 (1–36). Hormono therapy was used in 333 (58.62%), the most widely used being tamoxifen in 230 (69.1%).Conclusions Increased age is associated with a greater risk of breast cancer. Diagnosis in these patients is mainly clinical. Conservative treatment is infrequent in these patients as tumors are usually diagnosed in the more advanced stages (AU)

Association of SND1 protein to low density lipid droplets in liver steatosis.

Although the human homologue of SND p102, p100 coactivator, was initially described as a nuclear protein, the p100 coactivator protein family members have non-nuclear localization in mammalian cells with active lipid handling, storage, and secretion. However, their role in lipid homeostasis remains unresolved. Here, we investigate the distribution of the rat homologue SND p102 (also called SND1) and its association with newly formed lipid droplets in the liver parenchyma and cultured hepatocytes. Sucrose gradient fractionation showed that SND p102 cofractionated with endoplasmic reticulum and Golgi markers. Such cofractionation was not altered in regenerating steatotic rat liver. However, SND p102 was also detected in lipid droplets from regenerating liver, showing a specific directionalization to the least dense ones. Confocal microscopy of cultured hepatocytes confirmed the findings of gradient fractionation. In addition, p100 coactivator was consistently encountered in microsomes and lipid droplets in control and oleate-treated HepG2 cells. The total amount of SND p102 in hepatocytes was similar in both conditions, suggesting a specific translocation of the protein. Our findings indicate that SND p102 and the human p100 coactivator have a ubiquitous cytoplasmic distribution in hepatocytes and that steatogenic conditions promote the targeting of SND p102 from other cell compartments to specific low density lipid droplets.

Association of SND1 protein to low density lipid droplets in liver steatosis

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Although the human homologue of SND p102, p100 coactivator, was initially described as a nuclear protein, the p100 coactivator protein family members have non-nuclear localization in mammalian cells with active lipid handling, storage, and secretion. However, their role in lipid homeostasis remains unresolved. Here, we investigate the distribution of the rat homologue SND p102 (also called SND1) and its association with newly formed lipid droplets in the liver parenchyma and cultured hepatocytes. Sucrose gradient fractionation showed that SND p102 cofractionated with endoplasmic reticulum and Golgi markers. Such cofractionation was not altered in regenerating steatotic rat liver. However, SND p102 was also detected in lipid droplets from regenerating liver, showing a specific directionalization to the least dense ones. Confocal microscopy of cultured hepatocytes confirmed the findings of gradient fractionation. In addition, p100 coactivator was consistently encountered in microsomes and lipid droplets in control and oleate-treated HepG2 cells. The total amount of SND p102 in hepatocytes was similar in both conditions, suggesting a specific translocation of the protein. Our findings indicate that SND p102 and the human p100 coactivator have a ubiquitous cytoplasmic distribution in hepatocytes and that steatogenic conditions promote the targeting of SND p102 from other cell compartments to specific low density lipid droplets (AU)

Modificación de la acción fisiológica de la FSH por las variantes genéticas de su receptor (FSHr) / No disponible

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[Molecular basis of obesity-related hepatic steatosis]. / Fundamento molecular de la esteatosis hepática asociada a la obesidad.

Non-alcoholic fatty liver disease is a chronic inflammation liver condition that is currently highly relevant because of its strong association with increasingly incident diseases such as obesity and type-2 diabetes mellitus. The primary purpose of this paper is to discuss the best part of current knowledge on the molecular mechanisms involved in hepatic steatosis development, the condition s initial stage, and on progression to steatohepatitis. Special attention has been paid to clinical and experimental obesity-related fatty liver. In the latter, the fa/fa rat is assessed, which constitutes an animal model for obesity with phenotype features similar to human obesity, including insulin resistance and dyslipemia. Hepatic steatosis is a complex, mainly metabolic condition where apparently non-compatible metabolic processes concur, in addition to oxidative stress, endoplasmic reticulum stress, mitochondrial dysfunction, and decreased expression of survival genes. Extrahepatic signals underlie the disorder, such as those arising from peripheral insulin resistance associated with an increase in adipose mass and systemic free fatty acids, together with intrahepatic signals leading to derangement of liver glycostatic and lipidostatic functions, as well as to greater vulnerability to other aggressions.

Fundamento molecular de la esteatosis hepática asociada a la obesidad / No disponible

La enfermedad del hígado graso no alcohólico es una enfermedadinflamatoria hepática de carácter crónico de gran relevanciaen la actualidad por su fuerte asociación con enfermedades de incidenciacreciente como la obesidad y la diabetes mellitus tipo 2.En este trabajo se recoge buena parte del conocimiento existentesobre los mecanismos moleculares implicados en el establecimientode la esteatosis hepática, el primer estadio de la enfermedad, yen su progreso a esteatohepatitis. Se ha prestado una atención especialal hígado graso asociado a la obesidad, clínica y experimental.En este caso, se valora la rata fa/fa, un modelo animal de obesidadcon rasgos fenotípicos similares a los de la obesidadhumana, incluyendo la resistencia a la insulina y la dislipemia. Laesteatosis hepática se revela como una situación compleja, eminentementemetabólica, en la que se simultanean procesos metabólicosaparentemente contradictorios, así como estrés oxidativo,estrés de retículo endoplasmático, disfunción mitocondrial y descensoen la expresión de genes de supervivencia. En buena medida,en su base se sitúan señales extrahepáticas, como las producidasen una situación de resistencia periférica a la insulina asociadaa un aumento de la masa adiposa y de ácidos grasos libres sistémicos,e internas, causantes de un desajuste de las funciones glucostáticay lipidostática del hígado y de una mayor vulnerabilidad aotras agresiones

Non-alcoholic fatty liver disease is a chronic inflammation livercondition that is currently highly relevant because of its strong associationwith increasingly incident diseases such as obesity andtype-2 diabetes mellitus. The primary purpose of this paper is todiscuss the best part of current knowledge on the molecular mechanismsinvolved in hepatic steatosis development, the condition’sinitial stage, and on progression to steatohepatitis. Special attentionhas been paid to clinical and experimental obesity-related fattyliver. In the latter, the fa/fa rat is assessed, which constitutes ananimal model for obesity with phenotype features similar to humanobesity, including insulin resistance and dyslipemia. Hepaticsteatosis is a complex, mainly metabolic condition where apparentlynon-compatible metabolic processes concur, in addition tooxidative stress, endoplasmic reticulum stress, mitochondrial dysfunction,and decreased expression of survival genes. Extrahepaticsignals underlie the disorder, such as those arising from peripheralinsulin resistance associated with an increase in adipose mass andsystemic free fatty acids, together with intrahepatic signals leadingto derangement of liver glycostatic and lipidostatic functions, aswell as to greater vulnerability to other aggressions

Chitosan-carboxymethylcellulose interpolymer complexes for gastric-specific delivery of clarithromycin.

Chitosan (CS) and carboxymethylcellulose (CMC) sodium interpolymer complexes were formed using the novel method "tablets-in-capsule" for stomach drug delivery. The aim of this study was to investigate the influence of the molecular weight (M.wt.) of CS and the proportion CS/CMC on physical properties and clarithromycin (CAM) release. Swelling was dependent on CS M.wt., on medium pH and on proportion polymer/polymer. The higher M.wt., the major presence of protonated amined moieties that can be ionized and modify the swelling/eroding and dissolution medium uptake capacity. Swelling kinetics at pH 1.2, were close to Fickian diffusion, whereas at pH 4.2 were non-Fickian. Furthermore, dissolution medium uptake capacity can be adjusted to an exponential equation at pH 1.2 (r2> or =0.96), and to a linear equation at pH 4.2 (r2> or =0.99), showing that at low pHs the repulsion among ionized chains is higher. CAM release rates have shown to be dependent on pH and on polymer proportion. At pH 1.2, the fastest profile was obtained when using high M.wt. CS. Drug diffusion was Fickian, so the process is governed by swelling/eroding. Whereas at pH 4.2, CAM release followed zero-order kinetics with no influence on M.wt. So, release is controlled by CAM low solubility.

Prevalencia y factores de riesgo de sobrepeso, obesidad y características bioquímicas en adolescentes de los colegios urbanos del cantón Cuenca en el año lectivo 2006-2007: Cuenca, 2006 / Prevalence and risk factors of overweight, obesity and biochemical characteristics in adolescents in urban schools in the canton Cuenca in the academic year 2006-2007: Cuenca, 2006

Estudio de corte transversal para calcular prevalencia de sobrepeso y obesidad e identificar los factores asociados en una muestra aleatoria de 696 estudiantes. la prevalencia de sobrepeso fue del 17,7% y de obesidad del 8,3%. El sobrepeso fue mayor en las mujeres con una razón de prevalencia (RP) de 1,08 (IC95%: 0,7 – 1,4) y la obesidad en los varones, RP de 1,9 (IC95%: 0,1 – 3,5). Hábitos nutricionales no saludables fue el factor asociado más relevante con una RP de 18,4 (IC95%: 12,8 – 26,4) en el grupo con sobrepeso y una RP de 36,1 (IC95%: 21,2 – 61,2) en el grupo con obesidad. La asociación fue altamente significativa. de las pruebas bioquímicas la insulina y la hipertrigliceridemia tuvieron cifras más elevadas en la obesidad que en el sobrepeso (P < 0,05) y la alteración de los niveles de HDL fue mayor del 65% en ambos subgrupos. la prevalencia de sobrepeso y obesidad en nuestra ciudad alcanza porcentajes similares a los reportados en la literatura médica y está asociada a los hábitos nutricionales no saludables con niveles elevados de insulina, triglicéridos y valores alterados de HDL.

Cross-sectional study to calculate prevalence of overweight and obesity and to identify associate factors in a random sample of 696 students. prevalence of overweight was 17,7% and obesity 8,3%. Overweight was bigger in the women with a prevalence rate (RP) 1,08 (IC95%: 0,7 - 1,4) and obesity in the males, RP 1,9 (IC95%: 0,1 - 3,5). Nutritional habits not healthy it was the most outstanding associate factor with a RP 18,4 (IC95%: 12,8 - 26,4) in overweight group and a RP 36,1 (IC95%: 21,2 - 61,2) in obesity group. The association was highly significant. of biochemical tests the insulin and hipertrigliceridemy had higher values in obesity that in overweight (P <0,05) and abnormalities levels of HDL was bigger than 65% in both subgroups. the prevalence of overweight and obesity in our city reach similar percentages to those reported in medical literature and it is associated to the nutritional habits no healthy with high levels of insulin, triglycerides and altered values of HDL.

La infección de hepatocitos de rata en cultivo con adenovirus recombinantes causa un drástico descenso en la secreción de lipoproteínas de muy baja densidad / Recombinant adenoviral infection of rat hepatocyte cultures dramatically decreases very low-density lipoprotein secretion

Introducción. Tras su administración sistémica, los adenovirus recombinantes se concentran en el hígado, lo que les validaría para la transferencia génica a este órgano, pero existen efectos colaterales. En este trabajo se ha analizado el impacto de la infección sobre la cantidad y composición de las lipoproteínas de muy baja densidad (VLDL) secretadas en hepatocitos de rata en cultivo. Métodos. Se generaron 2 vectores adenovirales, con ADNc en dirección sentido o antisentido, ambos con capacidad infectiva pero no de propagación. Se infectaron cultivos de hepatocitos de rata con una dosis no citotóxica y se analizaron las VLDL secretadas durante los 3 períodos de 24 h posteriores a la infección y el contenido en lípidos del citosol. Resultados. La infección causó un moderado descenso en la secreción de apo B48 y de éster de colesterol durante las primeras 24 h, pero posteriormente se redujo drásticamente la secreción de partículas con apo B48 y apo B100 y su contenido en lípidos. Además, modificó sus proporciones, aumentando el porcentaje de colesterol libre a costa del esterificado, que llegó a ser indetectable. La menor secreción de VLDL no conllevó una acumulación de lípido intracelular e, incluso, descendió la masa citosólica de éster de colesterol. Conclusión. La infección con adenovirus reduce la secreción hepatocitaria de VLDL y su porcentaje de colesterol esterificado y, a diferencia de otros virus, estas acciones no parecen ir acompañadas de esteatosis hepatocelular (AU)

Introduction. After systemic administration, recombinant adenoviruses are rapidly concentrated in the liver, making them good candidates for gene transfer to this organ. However, there are collateral effects. In this study, we analyzed the effect of adenoviral infection on the number and composition of very low-density lipoproteins (VLDL) secreted by cultured rat hepatocytes. Methods. Two recombinant adenoviruses were developed, each containing cDNA in sense or antisense orientation, and with infective but not propagating ability. Cultured rat hepatocytes were infected with a subcytotoxic adenoviral dose. Secretion of VLDL particles during 3 consecutive 24 h-periods after infection and cytosolic lipid content were characterized. Results. During the first 24 h period, apo B48 and cholesteryl ester secretion moderately decreased. However, in longer incubations, infection dramatically reduced both apo B48 and apo B100 secretion, as well as VLDL lipid content. Lipid proportion was also modified: free cholesterol increased while cholesteryl ester decreased to undetectable levels. Decreased VLDL secretion was not associated with intracellular lipid accumulation; indeed, cholesteryl ester mass was even lower than in noninfected cells. Conclusion. Adenovirus infection causes hepatocytes to secrete less VLDL and to reduce their cholesteryl ester percentage. In contrast with the effects of other viruses, these effects are not accompanied by hepatocellular steatosis (AU)

Melanoma maligno nodular acral gigante / Giant nodular malignant melanoma

El Melanoma Maligno Acral es una entidad rara, asociado a un patrón nodular es menos frecuente. El melanoma es un tumor maligno de los melanocitos. La mayoría se localizan en piel y menos frecuentemente en mucosas, retina y meninges. El melanoma cutáneo constituye del 2-3 por ciento del total de neoplasias. El diagnóstico se basa en la sospecha clínica de una lesión cutánea con coloración heterogénea, asimetría, bordes irregulares y diámetro >5mm. Se presenta el caso clínico de paciente femenino de 47 años de edad, quien consultó por presentar tumoración exofítica gigante, de crecimiento rápido y progresivo, en talón derecho. Se realizó exéresis para estudio histopatológico, el cual reportó melanoma maligno nodular ulcerado, con bordes de resección quirúrgica positivos para invasión tumoral.

The fatty acid composition of chylomicron remnants influences their propensity to oxidate.

BACKGROUND AND AIM: Although the replacement of saturated with unsaturated dietary fat has been advocated as a means of reducing the risk of cardiovascular disease, diets high in polyunsaturated fatty acids (PUFAs) may increase lipid peroxidation, thus contributing to the pathogenesis of atherosclerosis. As the susceptibility of individual fatty acids to oxidation directly depends on their degree of unsaturation, and the oxidative modification of lipoproteins may be an important determinant of atherogenesis, the aim of this study was to evaluate the susceptibility to auto-oxidation and copper-mediated oxidation of chylomicron remnants (CMRs) enriched in n-3 or n-6 PUFA. METHODS AND RESULTS: The remnants were prepared in vitro from chylomicrons obtained from rats given an oral dose of fish or corn oil, using rat plasma containing lipoprotein lipase. Their propensity to oxidate and the extent of the oxidation were estimated by measuring the formation of conjugated dienes and the detrimental products of lipid peroxidation. The results showed that: 1) the corn oil CMRs contained a relatively high proportion of n-6 PUFA (mainly linoleic acid), whereas the fish oil CMRs contained more n-3 PUFA, mainly eicosapentanoic and docosahexaenoic acids; 2) n-3-rich CMRs have a significantly lower propensity to oxidate than n-6-rich CMRs despite their 50% lower alpha-tocopherol content and 40% higher unsaturation index. CONCLUSION: Our data indicate that the precise allocation of n-3 PUFA within the lipid core of CMRs may play a pivotal role in lowering the susceptibility to oxidation of fish CMRs by overcoming the effects of unfavourable alpha-tocopherol concentration. Eating n-3 rather than n-6 PUFAs seems to make CMRs more resistant against free radical attack, which may contribute to attenuating their potential atherogenic properties.

Influence of the fatty acid composition of lipids in chylomicron remnants derived from fish or corn oil on the lipid profile of cultured rat hepatocytes.

The aim of this work was to characterise the lipid and fatty acid composition of chylomicron remnants enriched in n-3 or n-6 polyunsaturated fatty acids (PUFA) and to investigate their influence on the fatty acid profiles of the lipids of rat hepatocytes cultured in monolayers. Chylomicrons were prepared from the lymph collected from the thoracic duct of rats given an oral dose of fish or corn oil (high in n-3 and n-6 PUFA, respectively), and remnants were prepared in vitro from such chylomicrons using rat plasma containing lipoprotein lipase. The fatty acids predominating in the oils abounded also in their respective chylomicrons and remnants, especially in triacylglycerols. Chylomicrons as well as remnants contained small amounts of phospholipids and long-chain PUFA that were minor in, or absent from, the dietary oils, evidently provided by the intestinal epithelium. The incubation of hepatocytes for 6 h, with either n-3 or n-6 PUFA-rich remnants (0.25-0.75 mM triacylglycerol) resulted in a dose-dependent increase in the amount of triacylglycerols and phospholipids in the cells, which was not affected further by increasing the incubation time to 19 h. Whereas hepatocyte triacylglycerols mostly incorporated the PUFA predominating in each remnant type, the fatty acid profile of cell phospholipids was virtually unchanged. In addition, irrespective of whether they were enriched in n-3 or n-6 PUFA, remnants promoted a relative decrease in the amount of cholesteryl esters, a minor hepatocyte lipid class poor in PUFA. The results demonstrate that the hepatocyte fatty acid profile is modulated in a lipid-class specific way by the amount and type of dietary PUFA delivered to cells in chylomicron remnants.

Lipid and fatty acid composition of canine lipoproteins.

Lipid classes and their fatty acids were studied in the major lipoprotein fractions from canine, in comparison with human, plasma. In dogs, high-density-lipoprotein (HDL), the main carrier of plasma phospholipid (PL), cholesterol ester (CE) and free cholesterol, was the most abundant lipoprotein, followed by low and very-low density lipoproteins (LDL and VLDL). Notably, LDL and VLDL contributed similarly to the total dog plasma triacylglycerol (TG). The PL composition was similar in all three lipoproteins, dominated by phosphatidylcholine (PC). Even though the content and composition of lipids within and among lipoproteins differed markedly between dog and man, the total amount of circulating lipid was similar. All canine lipoproteins were relatively richer than those from humans in long-chain (C20-C22) n-6 and n-3 polyunsaturated fatty acids (PUFA) but had comparable proportions of total saturated and monoenoic fatty acids, with 18:2n-6 being the main PUFA in both mammals. The fatty acid profile of canine and human lipoproteins differed because they had distinct proportions of their major lipids. There were more n-3 and n-6 long-chain PUFA in canine than in human plasma, because dogs had more HDL, their HDL had more PC and CE, and both these lipids were richer in such PUFA.

Immunolocalization of a novel cholesteryl ester hydrolase in the endoplasmic reticulum of murine and human hepatocytes.

We have recently purified a cholesteryl ester hydrolase (CEH) from rat liver microsomes. Antibodies raised against the purified protein specifically reacted with a 106-kd protein and neutralized 90% of the CEH activity of rat liver microsomes (J Lipid Res 1999;40:715-725). In this work we have used the anti-CEH antibody to study both the subcellular distribution of the protein in hepatocytes as well as its tissue-specific expression in rat. Western blotting of subcellular fractions obtained from isolated rat hepatocytes revealed that the immunoreactive 106-kd CEH was exclusively localized in microsomes. The antibody also recognized a 106-kd protein in microsomes from mouse and human liver but not from rat nonparenchymal liver cells. Confocal microscopy of HepG2 cells revealed that CEH immunoreactive material colocalized with calnexin, a marker of the endoplasmic reticulum. Furthermore, high-resolution immunoelectron microscopy of rat liver thin sections exclusively localized the CEH immunoreactivity to the endoplasmic reticulum of the hepatocyte. No CEH immunoreactivity was observed in microsomes derived from adrenal glands, ovaries, testis, pancreas, intestine, white adipose tissue, mammary gland, lung, spleen, brain, aorta, and macrophages. We report a CEH localized to the endoplasmic reticulum, erCEH, in the mammalian hepatocyte. The subcellular localization and tissue-restricted pattern of expression of erCEH suggests that it might have unique functions in liver cholesterol metabolism.

OPGLl/RANKL y remodelado óseo / OPGL/RANKL and bone remodeling

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