Biomedical subjects

J M Bautista

Publications and source records attributed to J M Bautista.

At least 19 recordsLinked to original sources

PubMed2007

Assessment of dietary nutrients that influence perception of intra-oesophageal acid reflux events in patients with gastro-oesophageal reflux disease.

BACKGROUND: Gastro-oesophageal reflux disease symptoms are most commonly reported postprandially, suggesting that some diet components are likely to induce symptoms more than others. AIMS: To determine which of the various dietary nutrients is a strong predictive factor for symptom generation in association with an acid reflux event. METHODS: Subjects with typical heartburn symptoms were evaluated by the gastro-oesophageal reflux disease Symptom Checklist, demographics questionnaire, upper endoscopy and pH testing. During the pH study, patients completed a detailed 24-h dietary intake record. This included time of meals, description of food components and the amount and type of food preparation. RESULTS: Fifty gastro-oesophageal reflux disease patients completed all stages of the study. A total of 112 (78%) symptoms were considered as sensed reflux event. Body mass index did not correlate with having perceived reflux. Patients who consumed more cholesterol, saturated fatty acids and had more percentage calories from fat were significantly more likely to experience a perceived reflux event. Regression analysis and beta-coefficient were specifically significant for cholesterol. CONCLUSION: Of all dietary nutrients, cholesterol enhances the most the perception of intra-oesophageal acid reflux events in patients with gastro-oesophageal reflux disease.

Adult↗

PubMed2006

Functional analysis of gammaretroviral vector transduction by quantitative PCR.

BACKGROUND: In a clinical setting of gene therapy, quantitative methods are required to determine recombinant viral titres and transgene mRNA expression, avoiding the use of reporter genes. METHODS: We describe procedures based on quantitative polymerase chain reaction (qPCR) designed to assess functional titres of murine leukaemia virus (MLV) vectors, determine proviral copy numbers in transduced cells, and estimate retroviral transgene expression in both target cell lines and mice with transduced chimeric haematopoiesis. RESULTS: Compared to EGFP titration, proviral DNA detection by qPCR was more accurate in assessing the number of infective particles in supernatants, such that average viral titres in terms of proviral copies per cell were two-fold higher. Transgene mRNA expression was directly determined from the vectors used without the need for reporter assays. A new parameter, defined here as the 'transcription index' (TI), served to establish the association between transcribed transgenic mRNA and each proviral insertion. The TI represents the potential expression of every vector or insertion in each cell type, and is thus useful as a control parameter for monitoring preclinical or clinical protocols. CONCLUSIONS: The practical use of qPCR is demonstrated as a valuable alternative to reporter genes for the assessment and surveillance of insertion numbers and transgene expression. In combination with protein expression, this approach should be capable of establishing safer therapeutic gene doses, avoiding the potential side effects of high transduction and expression levels.

3T3 Cells↗

PubMed2005

Dietary protein source affects lipid metabolism in the European seabass (Dicentrarchus labrax).

The study was undertaken to evaluate the effects of dietary protein sources on lipogenesis and fat deposition in a marine teleost, the European seabass (Dicentrarchus labrax). Four isonitrogenous (crude protein (CP, Nx6.25), 44% DM) and isoenergetic (22-23 kJ/g DM) diets were formulated to contain one of the following as the major protein source: fish meal (FM), one of two soy protein concentrates (SPC) and corn gluten meal (CGM). Apparent digestibility coefficients of the diets and raw ingredients, as well as soluble nitrogen (ammonia and urea) and phosphorus excretion were measured. Growth rates of seabass fed plant protein-based diets were significantly lower than those fed fish meal based diet. The protein utilisation was strongly correlated to the dietary essential amino acids index. Measurements of N excretion (ammonia and urea nitrogen) confirmed these data. Daily fat gain at the whole body level ranged between 1.1 to 1.7 g/kg BW, with the highest values being recorded in fish fed the fish meal based diet. Levels of plasma triglycerides and cholesterol were lower in fish fed soy protein diets than in those fed the diet solely based on fish meal. Soy protein rich diets decreased the activities of selected hepatic lipogenic enzymes (glucose 6-phosphate dehydrogenase, malic enzyme, ATP-citrate lysase, acetylcoenzyme A carboxylase and fatty acid synthetase). Highest lipogenic enzyme activities where found in fish fed the fish meal diet, except for fatty acid synthetase which was increased in seabass fed the corn-gluten meal based diets. Overall data suggest that dietary protein sources affects fat deposition and the lipogenic potential in European seabass.

Animals↗

PubMed2005

Effect of dietary fish oil substitution with linseed oil on the performance, tissue fatty acid profile, metabolism, and oxidative stability of Atlantic salmon.

The objective of this experiment was to test the effect of total or partial substitution of dietary fish oil (FO) by linseed oil (LO) in Atlantic salmon feeding on performance, liver and muscle fatty acid composition, selected lipogenic and lipolytic enzyme activities, and flesh oxidative stability. For 12 wk, fish (220 +/- 12 g of initial BW) were fed five experimental diets in which the FO was serially replaced by 25, 50, 75, and 100% LO. Total FO replacement by LO did not (P = 0.20) affect fish final weight, biometric indices, or i.m. fat contents. Liver and muscle neutral lipid (NL) composition responded to dietary treatments in different ways. Whereas the sum of n-3 PUFA in muscle followed a linear and quadratic pattern with increasing levels of LO, a linear (P = 0.005) effect was observed in the liver NL fraction. Total n-3 and n-6 PUFA contents in the polar lipid fraction (PL) were unaffected (P = 0.356) by dietary input of LO in muscle. Activity of liver glucose-6-P-dehydrogenase (G6PD) was greater with increasing levels of LO (P = 0.004). A time effect (P < 0.001) was observed in the concentration of lipid peroxidation products, expressed as thiobarbituric acid reactive substances, in fish flesh stored under refrigeration for 9 d; however, the progressive inclusion of LO in the feed did not affect (P = 0.125) flesh oxidation stability. In summary, LO can totally replace FO in Atlantic salmon feed without affecting growth performance and muscle susceptibility to lipid oxidation. Fatty acid metabolism in the liver was affected by LO, promoting G6PD activity and eicosatetraenoic acid accumulation; however, a 100% LO replacement decreased (P < 0.001) concentrations of eicosapentaenoic and docosahexaenoic acids in salmon muscle.

Animal Feed↗

PubMed2004

Adaptation of lipid metabolism, tissue composition and flesh quality in gilthead sea bream (Sparus aurata) to the replacement of dietary fish oil by linseed and soyabean oils.

Linseed (LO) and soyabean (SO) oils were evaluated as fish-oil (FO) substitutes in the diets of marketable-sized gilthead sea bream (Sparus aurata). Practical diets were designed factorially with the lipid added as follows (%): FO 100, LO 60+FO 40, LO 80+FO 20, SO 60+FO 40, SO 80+FO 20. The effects of experimental diets on growth, fatty acids patterns in liver and muscle, flesh quality variables and activities of selected enzymes involved in lipid synthesis and catabolism were determined at the end of a 7-month trial. Fatty acid composition of liver and muscle generally reflected the fatty acid composition of the diets. The n-3 PUFA levels were significantly reduced by the inclusion of vegetable oils. This tendency was more pronounced for EPA than for docosahexaenoic acid. The n-3:n-6 fatty acid ratio reached the lowest values in fish fed the SO diets; this was associated with a higher liver lipid deposition. No differences were found in fillet texture and pH. However, under conditions of forced peroxidation, muscles from fish fed the SO diets had lower peroxidation levels. Vegetable oil substitution decreased lipogenesis in liver and this effect was greatest at the highest substitution level. In contrast, muscle beta-oxidation enzymes had increased activities with vegetable oil substitution. Thus, the lower hepatic lipogenesis was correlated with an increased lipid utilisation in muscle. It is concluded that growth and lipid metabolism were affected by experimental diets.

Animal Feed↗

PubMed2004

The use of fluorescent molecular beacons in real time PCR of IgH gene rearrangements for quantitative evaluation of multiple myeloma.

UNLABELLED: BACKGROUND AND OBJECTIVES Fluorescent molecular beacons have been employed as hybridization probes in real time quantitative PCR to quantify residual disease in multiple myeloma (MM). DESIGN AND METHODS: After clinical diagnosis of MM, the CDR1, CDR2 and CDR3 regions of the IgH gene were analysed and sequenced to identify its clonal nature. Unique sequences of the clonal IgH rearrangement were used to design specific molecular beacon probes for each MM patient. A molecular beacon probe for the beta-globin gene was used as a reference control to calculate relative amounts of the clonal B-cell population. RESULTS: Optimization of probe design resulted in the use of a competitive sequence at the IgH area target between the loop and part of the stem of the molecular beacon. Cycling conditions and fluorescence temperature acquisition were optimized for a Light Cycler. To validate this method for the follow-up of treated MM patients, we investigated accuracy, as well as interassay and intrassay reproducibility. CONCLUSIONS: Our results indicated that real time PCR with specific molecular beacons provides a feasible, accurate and reproducible method for the determination of minimal residual disease in MM.

DNA Probes↗

PubMed2001

Evolution of the mitochondrial control region in Palaearctic brown trout (Salmo trutta) populations: the biogeographical role of the Iberian Peninsula.

In order to extend present knowledge of brown trout phylogeography in the Palaearctic, we analysed the complete mitochondrial D-loop sequence (1025-1027 bp) of all mitochondrial haplotypes of Salmo trutta found in the Iberian Peninsula and one North African haplotype. Sequence variation in the mitochondrial control region serves to identify four major haplotype groups within the Iberian Peninsula, i.e. Atlantic, Duero, Mediterranean and Andalusian. Including the Iberian haplotypes, the five main European groups previously established were increased to six: (i) an Atlantic group including two different clusters, South European and North Atlantic; (ii) a group representing an endemism restricted to the Duero basin in the Iberian Peninsula; (iii) an Adriatic-Andalusian group found in two vicariant areas including Adriatic-Ionian populations in the Mediterranean and the Andalusian basins of the southern Iberian Peninsula; (iv) a Mediterranean group with a distribution range that extends from the southwestern basins of the Iberian Peninsula to the Ionian basins of the Greek Peninsula; (v) a Danube group of wide distribution in the Black, Aral and Caspian basins; and (vi) a group comprising the S. t. marmoratus subspecies confined to the Adriatic Sea. The Iberian Peninsula appears to have acted as a physical boundary between haplotypes corresponding to Atlantic- and Mediterranean-draining rivers. Owing to its geographical position, this area has played a major role in present Palaearctic species distribution, as illustrated by its haplotype diversity.

Animals↗

PubMed2000

Supercritical carbon dioxide extraction of lipids from Eucalyptus globulus wood.

Various typical lipid components of wood extractives have been isolated from Eucalyptus globulus wood by supercritical carbon dioxide modified with methanol. The influence of various extraction parameters on the yield and qualitative composition of the extracts have been studied. The extracts were analyzed by gas chromatography-mass spectrometry and compared with those obtained by Soxhlet extraction with acetone, the standard method for the determination of wood extractives. The qualitative and quantitative results obtained by both methods were in good agreement. The experimental planning to asses the influence of pressure, temperature and percentage of methanol and their interactions on the extraction efficiency was carried out with a factorial design, followed by multiple linear regression algorithm.

Carbon Dioxide↗

PubMed2000

Structural defects underlying protein dysfunction in human glucose-6-phosphate dehydrogenase A(-) deficiency.

The enzyme variant glucose-6-phosphate dehydrogenase (G6PD) A(-), which gives rise to human glucose-6-phosphate dehydrogenase deficiency, is a protein of markedly reduced structural stability. This variant differs from the normal enzyme, G6PD B, in two amino acid substitutions. A further nondeficient variant, G6PD A, bears only one of these two mutations and is structurally stable. In this study, the synergistic structural defect in recombinant G6PD A(-) was reflected by reduced unfolding enthalpy due to loss of beta-sheet and alpha-helix interactions where both mutations are found. This was accompanied by changes in inner spatial distances between residues in the coenzyme domain and the partial disruption of tertiary structure with no significant loss of secondary structure. However, the secondary structure of G6PD A(-) was qualitatively affected by an increase in beta-sheets substituting beta-turns related to the lower unfolding enthalpy. The structural changes observed did not affect the active site of the mutant proteins, since its spatial position was unmodified. The final result is a loss of folding determinants leading to a protein with decreased intracellular stability. This is suggested as the cause of the enzyme deficiency in the red blood cell, which is unable to perform de novo protein synthesis.

Amino Acid Sequence↗

PubMed2000

Mitochondrial haplotype variation and phylogeography of Iberian brown trout populations.

The biogeographical distribution of brown trout mitochondrial DNA haplotypes throughout the Iberian Peninsula was established by polymerase chain reaction-restriction fragment polymorphism analysis. The study of 507 specimens from 58 localities representing eight widely separated Atlantic-slope (north and west Iberian coasts) and six Mediterranean drainage systems served to identify five main groups of mitochondrial haplotypes: (i) haplotypes corresponding to non-native, hatchery-reared brown trout that were widely distributed but also found in wild populations of northern Spain (Cantabrian slope); (ii) a widespread Atlantic haplotype group; (iii) a haplotype restricted to the Duero Basin; (iv) a haplotype shown by southern Iberian populations; and (v) a Mediterranean haplotype. The Iberian distribution of these haplotypes reflects both the current fishery management policy of introducing non-native brown trout, and Messinian palaeobiogeography. Our findings complement and extend previous allozyme studies on Iberian brown trout and improve present knowledge of glacial refugia and postglacial movement of brown trout lineages.

Animals↗

PubMed2000

Abdominal fat deposition and fatty acid synthesis are lower and beta-oxidation is higher in broiler chickens fed diets containing unsaturated rather than saturated fat.

We evaluated the effects of dietary fat type on fat metabolism and deposition in broiler chickens. Birds were fed diets containing either 8 g dietary saturated (beef tallow) or polyunsaturated fat (sunflower oil)/100 g for 32 d. The abdominal fat deposition of chickens fed the sunflower oil-enriched diet was significantly lower than that of chickens fed the tallow-enriched diet (2.63 +/- 0.47 versus 3.03 +/- 0.44 g/100 g live wt.; P = 0.033). The specific activities of heart carnitine palmitoyltransferase I and L-3-hydroxyacyl-CoA dehydrogenase were higher (P < or = 0.03) in chickens fed the sunflower oil-enriched diets, indicating a greater rate of beta-oxidation. Liver fatty acid synthetase activity was lower (P = 0.01) in chickens fed the sunflower oil-enriched diet, suggesting reduced hepatic lipogenesis in this group. Postprandial plasma triglyceride levels were significantly lower (P < 0.05) in birds fed the sunflower oil-enriched diet, indicating a higher rate of dietary lipid clearance from the bloodstream to tissues. In conclusion, the lower fat deposition observed in broilers fed sunflower oil-enriched diets appears to be the net result of an increased rate of lipid catabolism and lower rate of fatty acid synthesis despite higher dietary fat absorption.

Adipose Tissue↗

PubMed2000

Short-term modulation of lipogenesis by macronutrients in rainbow trout (Oncorhynchus mykiss) hepatocytes.

Rainbow trout (Oncorhynchus mykiss) hepatocytes were cultured under simulated conditions of varying nutritional status to explore the short-term modulation by dietary substrates of the main lipogenic enzymes: glucose-6-phosphate dehydrogenase (G6PD), malic enzyme (ME), ATP-citrate lyase (ACL), acetyl-CoA carboxylase (ACoAC) and fatty acid synthetase (FAS). Primary cultures were individually exposed to varying amounts of glucose, hydrolysed casein and long-chain polyunsaturated fatty acids (PUFA) for 12 h. A second set of experiments was designed to evaluate the effects of mixing different relative amounts of these macronutrients in the culture medium. Glucose concentrations of up to 20-25 mm showed a stimulatory effect on G6PD, ME, ACL and ACoAC activity while an earlier inhibitory effect on FAS was observed at 10-20 mm glucose The use of hydrolysed casein as a nutritional source of amino acids inhibited the activity of FAS and ME and stimulated G6PD, ACoAC and ACL activity Low levels of linolenic acid exerted a stimulatory effect on all the lipogenic enzymes assayed with the exception of FAS, and increased amounts showed some inhibition of lipogenic activities Eicosapentaenoic acid and docosahexaenoic acid showed a similar effect, although the former strongly inhibited FAS activity while the latter showed greater potential to inhibit ACoAC and G6PD. A complete change in the relative levels of glucose, hydrolysed casein and PUFA in turn led to changes in the enzyme activity patterns observed. The present study shows the feasibility of exploring the direct regulation of lipogenesis in isolated fish cells by varying the relative amounts of main macronutrients, mimicking in vivo dietary conditions. It is felt that such an approach may serve to investigate the macronutrient regulation of other metabolic pathways.

Animals↗

PubMed1999

Molecular basis and enzymatic properties of glucose 6-phosphate dehydrogenase volendam, leading to chronic nonspherocytic anemia, granulocyte dysfunction, and increased susceptibility to infections.

We have investigated the blood cells from a woman with a low degree of chronic nonspherocytic hemolytic anemia and frequent bacterial infections accompanied by icterus and anemia. The activity of glucose 6-phosphate dehydrogenase (G6PD) in her red blood cells (RBCs) was below detection level, and in her leukocytes less than 3% of normal. In cultured skin fibroblasts, G6PD activity was approximately 15% of normal, with 4- to 5-fold increased Michaelis constant (Km) for NADP and for glucose 6-phosphate. Activated neutrophils showed a decreased respiratory burst. Family studies showed normal G6PD activity in the RBCs from all family members, including both parents and the 2 daughters of the patient. Sequencing of polymerase chain reaction (PCR)-amplified genomic DNA showed a novel, heterozygous 514C-->T mutation, predicting a Pro172-->Ser replacement. Analysis of G6PD RNA from the patient's leukocytes and fibroblasts showed only transcripts with the 514C-->T mutation. This was explained by the pattern of X-chromosome inactivation, studied by means of the human androgen receptor (HUMARA) assay, which proved to be skewed in the patient, her mother, and one of the patient's daughters. Thus, the patient has inherited a de novo mutation in G6PD from her father and an X-chromosome inactivation determinant from her mother, causing exclusive expression of the mutated G6PD allele. Purified mutant protein from an Escherichia coli expression system showed strongly decreased specific activity, increased Km for NADP and for glucose 6-phosphate, and increased heat lability, which indicates that the defective phenotype is due to 2 synergistic molecular dysfunctions: decreased catalytic efficiency and protein instability.

Adult↗

PubMed1999

Increased neuronal glucose-6-phosphate dehydrogenase and sulfhydryl levels indicate reductive compensation to oxidative stress in Alzheimer disease.

We analyzed glucose-6-phosphate dehydrogenase, the rate-controlling enzyme of the pentose phosphate pathway and free sulfhydryls, to study redox balance in Alzheimer disease. Glucose-6-phosphate dehydrogenase plays a pivotal role in homeostatic redox control by providing reducing equivalents to glutathione, the major nonenzymatic cellular antioxidant. There is a multitude of evidence that marks oxidative stress proximally in the natural history of Alzheimer disease. Consistent with a role for glutathione in defense against increased reactive oxygen, we found an upregulation of glucose-6-phosphate dehydrogenase together with increased sulfhydryls in Alzheimer disease. These data indicate that reductive compensation may play an important role in combating oxidative stress in Alzheimer disease.

Adolescent↗

PubMed1999

Phylogeography of African fruitbats (Megachiroptera).

Joint sequences from the mitochondrial cytochrome b and 16S rRNA genes of a wide representation of Megachiroptera were employed to evaluate the traditional taxonomic arrangement of African fruitbats and to examine their origins and evolutionary relationships. The resulting phylogenetic hypotheses are inconsistent with the previously established morphology-based subdivisions of Megachiroptera at the suprageneric level. Findings indicate the existence of an African clade, which appears to be formed by two endemic clades: the epomophorines and the myonycterines. According to our topologies, the genus Rousettus is monospecific in mainland Africa. Its traditional subgenera Stenonycteris and Lissonycteris appear closer to the myonycterines than to Rousettus. Topologies also indicate that the African genus Eidolon is not phylogenetically related to any other African fruitbat. It would seem that the arrival of fruitbats in Africa was a complex process involving at least three independent colonization events. One event took place probably in the Miocene via forested corridors that connected the African and Asian rain forest blocks, as for other groups of mammals. The resulting lineage diversified into most of the extant African fruitbats. Related to this clade, the Rousettus species group is thought to have arrived in Africa in more recent times, possibly by progressive displacement from the East through India. Finally, the present topologies suggest an independent colonization of Africa by ancestors of Eidolon.

Africa↗

PubMed1999

Molecular phylogeny and morphological homoplasy in fruitbats.

The present study evaluates the evolutionary framework of the Old World fruitbats based on the cytochrome b and 16S rRNA mitochondrial gene sequences from a wide range of taxa. Phylogenetic analyses indicated that morphology-based subfamilies and most suprageneric groups are nonnatural assemblages. They also support the existence of an endemic African clade of fruitbats. The discrepancy between the evolutionary relationships yielded by molecular and morphological data sets may be, at least in part, explained by the recurrent retention of primitive morphology (Rousettus-like) across different lineages. The maintenance of primitive characters in different groups of flying foxes, as well as morphological convergence in nectar-feeding bats and possibly also in short-muzzle bats, may have led to high levels of homoplasy, resulting in misleading taxonomic arrangements. This may be particularly so with respect to high taxonomic levels based on morphological characters.

Africa↗

PubMed1999

The partial substitution of digestible protein with gelatinized starch as an energy source reduces susceptibility to lipid oxidation in rainbow trout (Oncorhynchus mykiss) and sea bass (Dicentrarchus labrax) muscle.

We evaluated the influence of dietary gelatinized starch and protein on the fatty acid composition of muscle in rainbow trout and European sea bass and on the susceptibility of flesh to lipid peroxidation. The possibility that flesh peroxidation could be accounted for by lipogenesis and the deposition of fat was also explored. The inclusion of gelatinized starch in the diet of rainbow trout improved growth with respect to that observed in fish fed crude starch (P<.001). This was especially noticeable at the lowest concentration of dietary protein tested (P = .037); suggesting that gelatinized starch may partially replace protein in the production of energy without inducing a negative effect on growth. However, in European sea bass, the gelatinization of starch and dietary protein concentration showed no significant effect on final body weight. The intramuscular neutral lipid concentration of the sea bass was reduced by the gelatinization of dietary starch (P = .034). The highest dietary protein concentration increased the proportion of saturated fatty acids in the neutral (P = .0742) and polar (P = .0033) lipid fractions. The dietary inclusion of high levels of protein in rainbow trout led to a lower concentration of total (n-3) (P = .0457) and (n-6) (P = .0522) fatty acids and a higher concentration of total monounsaturated fatty acids (P = .0006). The inclusion of gelatinized starch led to a lower concentration of (n-3) fatty acids (P = .0034) and a higher concentration of saturated fatty acids (P = .0007). The polar fraction was hardly affected by the same treatment. A significantly lower susceptibility of the dorsal muscle to oxidation was observed in groups of European sea bass fed gelatinized starch (P<.01). A similar trend was observed in rainbow trout, although differences were not significant. The findings suggest that the digestible protein concentration of nutrient-dense diets for rainbow trout and European sea bass can be reduced with a beneficial effect on tissue lipid oxidation and no negative effects on growth and muscle composition.

Animals↗

PubMed1998

Amino acid substitutions at the dimer interface of human glucose-6-phosphate dehydrogenase that increase thermostability and reduce the stabilising effect of NADP.

Over 100 mutations of the G6PD gene have been documented. With the construction of the molecular model of glucose-6-phosphate dehydrogenase, based on the structure of the bacterial Leuconostoc mesenteroides glucose-6-phosphate dehydrogenase, it has been possible to superimpose these amino acid changes on to the structure of the glucose-6-phosphate dehydrogenase molecule. There are a large number of severe disease causing mutations at the dimer interface which usually cause decreased thermostability. We have used this knowledge to predict amino acid changes which would effect an increase in the stability of the dimer. The aspartic acid at residue 421 was chosen as it is a negatively charged residue at the centre of the dimer interface in an area rich in negatively charged residues. This residue was changed to a neutrally charged alanine or asparagine, or a positively charged lysine or arginine. The thermostability of the enzyme was increased when residue 421 was neutral (A or N) and increased further when positive (K or R). NADP is known to exert a concentration dependent stabilising effect on the glucose-6-phosphate dehydrogenase dimer. However the concentration-dependent stabilising effect of NADP was reduced in the residue-421 substitutions in a manner which was inversely proportional to charge change. These results suggest that changes at the dimer interface can also affect the distant (> 20 A) NADP-binding site, and vice versa; an attempt has been made to explain these interactions based on the molecular model of human glucose-6-phosphate dehydrogenase.

Dimerization↗