Pamela Morales

Pilot study: alterations of intestinal microbiota in obese humans are not associated with colonic inflammation or disturbances of barrier function

Aliment Pharmacol Ther 2010; 32: 1307–1314

Polyphenols Protect the Epithelial Barrier Function of Caco-2 Cells Exposed to Indomethacin through the Modulation of Occludin and Zonula Occludens-1 Expression

The aim of this study was to determine the protective effect of quercetin, epigallocatechingallate, resveratrol, and rutin against the disruption of epithelial integrity induced by indomethacin in Caco-2 cell monolayers. Indomethacin decreased the transepithelial electrical resistance and increased the permeability of the monolayers to fluorescein-dextran. These alterations were abolished by all the tested polyphenols but rutin, with quercetin being the most efficient. The protective effect of quercetin was associated with its capacity to inhibit the redistribution of ZO-1 protein induced in the tight junction by indomethacin or rotenone, a mitochondrial complex-I inhibitor, and to prevent the decrease of ZO-1 and occludin expression induced by indomethacin. The fact that the antioxidant polyphenols assayed in this study differ in their protective capacity against the epithelial damage induced by indomethacin suggests that this damage is due to the ability of this agent to induce not only oxidative stress but also mitochondrial dysfunction.

Phylogenetic and phylogeographic analysis of the genus Orestias (Teleostei: Cyprinodontidae) in the southern Chilean Altiplano: the relevance of ancient and recent divergence processes in speciation

This study presents phylogenetic molecular data of the Chilean species of Orestias to propose an allopatric divergence hypothesis and phylogeographic evidence that suggests the relevance of abiotic factors in promoting population divergence in this complex. The results reveal that diversification is still ongoing, e.g. in the Ascotán salt pan, where populations of Orestias ascotanensis restricted to individual freshwater springs exhibit strong genetic differentiation, reflecting putative independent evolutionary units. Diversification of Orestias in the southern Altiplano may be linked to historical vicariant events and contemporary variation in water level; these processes may have affected the populations from the Plio-Pleistocene until the present.

The gut microbiota of healthy chilean subjects reveals a high abundance of the phylum verrucomicrobia

The gut microbiota is currently recognized as an important factor regulating the homeostasis of the gastrointestinal tract and influencing the energetic metabolism of the host as well as its immune and central nervous systems. Determining the gut microbiota composition of healthy subjects is therefore necessary to establish a baseline allowing the detection of microbiota alterations in pathologic conditions. Accordingly, the aim of this study was to characterize the gut microbiota of healthy Chilean subjects using 16S rRNA gene sequencing. Fecal samples were collected from 41 young, asymptomatic, normal weight volunteers (age: 25 ± 4 years; ♀:48.8%; BMI: 22.5 ± 1.6 kg/m2) with low levels of plasma (IL6 and hsCRP) and colonic (fecal calprotectin) inflammatory markers. The V3-V4 region of the 16S rRNA gene of bacterial DNA was amplified and sequenced using MiSeq Illumina system. 109,180 ± 13,148 sequences/sample were obtained, with an α-diversity of 3.86 ± 0.37. The dominant phyla were Firmicutes (43.6 ± 9.2%) and Bacteroidetes (41.6 ± 13.1%), followed by Verrucomicrobia (8.5 ± 10.4%), Proteobacteria (2.8 ± 4.8%), Actinobacteria (1.8 ± 3.9%) and Euryarchaeota (1.4 ± 2.7%). The core microbiota representing the genera present in all the subjects included Bacteroides, Prevotella, Parabacteroides (phylum Bacteroidetes), Phascolarctobacterium, Faecalibacterium, Ruminococcus, Lachnospira, Oscillospira, Blautia, Dorea, Roseburia, Coprococcus, Clostridium, Streptococcus (phylum Firmicutes), Akkermansia (phylum Verrucomicrobia), and Collinsella (phylum Actinobacteria). Butyrate-producing genera including Faecalibacterium, Roseburia, Coprococcus, and Oscillospira were detected. The family Methanobacteriaceae was reported in 83% of the subjects and Desulfovibrio, the most representative sulfate-reducing genus, in 76%. The microbiota of the Chilean individuals significantly differed from those of Papua New Guinea and the Matses ethnic group and was closer to that of the Argentinians and sub-populations from the United States. Interestingly, the microbiota of the Chilean subjects stands out for its richness in Verrucomicrobia; the mucus-degrading bacterium Akkermansia muciniphila is the only identified member of this phylum. This is an important finding considering that this microorganism has been recently proposed as a hallmark of healthy gut due to its anti-inflammatory and immunostimulant properties and its ability to improve gut barrier function, insulin sensitivity and endotoxinemia. These results constitute an important baseline that will facilitate the characterization of dysbiosis in the main diseases affecting the Chilean population.

Threatened fishes of the world: Orestias ascotanensis Parenti, 1984 (Cyprinodontidae)

Common name: Killifish; Karachi. Conservation status: Endangered—Chilean Endangered Species List, Boletı́n Museo Nacional de Historia Natural, Chile, 1998. Identification: Orestias ascotanensis Parenti is a small, robust, fusiform, sexually dimorphic species. Females have more complete lateral scale rows than males. Orestias are generally irregularly scaled. Head length 35% of SL. Juvenile color mottled, adults light to dark brown with pale yellow to cream ventrum. Ventrum and basal zone of the pectoral, dorsal and anal fins are unscaled, caudal fin truncate, lower jaw upturned. Outer teeth of jaw are unicuspid, unpigmented and protruding beyond epithelium (Parenti 1984; Martı́nez et al. 1999). Fins are relatively longer than other southern species (D13, A13, P18). Head sensory pattern of clear lyre-shaped neuromasts on the dorsal head surface, continuing laterally along the preopercle and mandible. Adults reach 90 mm SL, females significantly larger as most Orestias species (Figure by Cecilia Fernández). O. ascotanensis differs from the other southern Altiplano species by the combination of: (1) the longest head, (2) a more upturned mouth, (3) 48 chromosomes, (4) 31–32 vertebrae, and (5) 29–34 lateral line scales (Arratia 1982; Parenti 1984; Vila and Pinto 1986; Costa 1997, 2003; Lüssen et al. 2003; Vila 2006). Distribution: Orestias ascotanensis is endemic to the Chilean Altiplano, restricted to springs of Ascotán salt lake, at 3,700 m a.s.l., presently distributed in I. Vila S. Scott P. M. Morales E. Poulin Departamento de Ciencias Ecológicas, Facultad de Ciencias, Universidad de Chile, Las Palmeras 3425, Casilla 653, CP 780-0024 Ñuñoa, Santiago, Chile

Impact of Dietary Lipids on Colonic Function and Microbiota: An Experimental Approach Involving Orlistat-Induced Fat Malabsorption in Human Volunteers

OBJECTIVES:High-fat diets alter gut microbiota and barrier function, inducing metabolic endotoxemia and low-grade inflammation. Whether these effects are due to the high dietary lipid content or to the concomitant decrease of carbohydrate intake is unclear. The aim of this study was to determine whether higher amounts of dietary fat reaching the colon (through orlistat administration) affect the colonic ecosystem in healthy volunteers and the effect of the prebiotic oligofructose (OF) in this model.METHODS:Forty-one healthy young subjects were distributed among four groups: Control (C), Prebiotic (P), Orlistat (O), and Orlistat/Prebiotic (OP). They consumed a fat-standardized diet (60 g/day) during Week-1 (baseline) and after 1 week of washout, Week-3. During Week-3, they also received their respective treatment (Orlistat: 2 × 120 mg/day, OF: 16 g/day, and maltodextrin as placebo). A 72-h stool collection was carried out at the end of Week-1 (T0) and Week-3 (T1). Fecal fat, calprotectin, and short-chain fatty acids (SCFAs) as well as the antioxidant activity of fecal waters (ferric-reducing antioxidant power), fecal microbiota composition (by deep sequencing), and gut permeability (Sucralose/Lactulose/Mannitol test) were determined at these times.RESULTS:Fecal fat excretion was higher in the O (P=0.0050) and OP (P=0.0069) groups. This event was accompanied, in the O group, by an increased calprotectin content (P=0.047) and a decreased fecal antioxidant activity (P=0.047). However, these alterations did not alter gut barrier function and the changes observed in the composition of the fecal microbiota only affected bacterial populations with low relative abundance (<0.01%); in consequences, fecal SCFA remained mainly unchanged. Part of the colonic alterations induced by orlistat were prevented by OF administration.CONCLUSIONS:In the context of an equilibrated diet, the acute exposition of the colonic ecosystem to high amounts of dietary lipids is associated with an incremented excretion of fecal calprotectin and pro-oxidant activity of the colonic content, in the absence of significant changes in the microbiota.

La microbiota intestinal: Un nuevo actor en el desarrollo de la obesidad

Intestinal microbiota (IM) plays a role in the development of obesity and its associated low-grade inflammation. Bacterial colonization of the gastrointestinal tract of germ-free mice (without microbiota) increases by 60% their fat mass, alters their fasting glucose and insulin levels, triples their hepatic triglycerides and induces adipocyte hypertrophy. IM favors fat storage in adipocytes through the inhibition of Fiaf (Fasting-Induced Adipocyte Factor), an inhibitor of lipoprotein lipase. Compa-red with normal weight subjects, the IM from obese exhibits a higher proportion of Firmicutes/Bacteroidetes and is more efficient in extracting energy from foodstuffs. The loss of bodyweight by a hypocaloric diet reverts the proportion of bacteria to that of lean subjects. The intake of a high fat diet also alters the IM, affecting intes-tinal barrier function and favoring endotoxinemia. These events increase oxidative and pro-inflammatory processes in plasma and peripheral tissues and increment the risk of insulin resistance. Such events are reverted by the administration of pre-biotics which stimulate the growth of Bifidobacterium and Lactobacillus species in the colon, reestablishing the gut homeostasis. Interestingly, products resulting from the fermentation of prebiotics stimulate the differentiation of enteroendocrine cells and the release of glucagon-like peptide 1 and peptide YY, that have insulin-like and anorexigenic activities, thus contributing to body weight equilibrium. (Rev Med Chile 2010; 138: 1020-1027).

Taxonomic identity of the forms of Orestias agassii (Teleostei: Cyprinodontidae) from Chile: A morphological comparison with the syntypes of the species of Peru and Bolivia

Se realizo una comparacion morfologica de dos poblaciones asignadas a Orestias agassii del Altiplano chileno con los sintipos de Peru y Bolivia. Se sugiere que las poblaciones de Chile no corresponden a las de Peru (Lago Titicaca), sin embargo, la poblacion de Isluga estaria relacionada con O. agassii de Bolivia.

Pattern of genetic differentiation of an incipient speciation process: The case of the high Andean killifish Orestias

During the Pleistocene and Holocene, the southwest Andean Altiplano (17°-22°S) was affected by repeated fluctuations in water levels, high volcanic activity and major tectonic movements. In the early Holocene the humid Tauca phase shifted to the arid conditions that have lasted until the present, producing endorheic rivers, lakes, lagoons and wetlands. The endemic fish Orestias (Cyprinodontidae) represents a good model to observe the genetic differentiation that characterizes an incipient speciation process in allopatry since the morphospecies described inhabit a restricted geographic area, with present habitat fragmentation. The genetic diversity and population structure of four endemic morphospecies of Orestias (Cyprinodontidae) found in the Lauca National Park (LNP) analyzed with mitochondrial markers (Control Region) and eight microsatellites, revealed the existence of genetic groups that matches the fragmentation of these systems. High values of genetic and phylogeographic differentiation indices were observed between Chungará Lake and Piacota lagoon. The group composed of the Lauca River, Copapujo and Chuviri wetlands sampling sites showed a clear signal of expansion, with a star-like haplotype network. Levels of genetic differentiation were lower than in Chungará and Piacota, suggesting that these localities would have differentiated after the bottlenecks linked to the collapse of Parinacota volcano. The Parinacota sample showed a population signal that differed from the other localities revealing greater genetic diversity and a disperse network, presenting haplotypes shared with other LNP localities. A mixing pattern of the different genetic groups was evident using the microsatellite markers. The chronology of the vicariance events in LNP may indicate that the partition process of the Orestias populations was gradual. Considering this, and in view of the genetic results, we may conclude that the morphospecies from LNP are populations in ongoing differentiation process.